Sample records for amniotic epithelial cells
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Directory of Open Access Journals (Sweden)
Ting-gang Wang
2016-01-01
Full Text Available Treatment and functional reconstruction after central nervous system injury is a major medical and social challenge. An increasing number of researchers are attempting to use neural stem cells combined with artificial scaffold materials, such as fibroin, for nerve repair. However, such approaches are challenged by ethical and practical issues. Amniotic tissue, a clinical waste product, is abundant, and amniotic epithelial cells are pluripotent, have low immunogenicity, and are not the subject of ethical debate. We hypothesized that amniotic epithelial cells combined with silk fibroin scaffolds would be conducive to the repair of spinal cord injury. To test this, we isolated and cultured amniotic epithelial cells, and constructed complexes of these cells and silk fibroin scaffolds. Implantation of the cell-scaffold complex into a rat model of spinal cord injury resulted in a smaller glial scar in the damaged cord tissue than in model rats that received a blank scaffold, or amniotic epithelial cells alone. In addition to a milder local immunological reaction, the rats showed less inflammatory cell infiltration at the transplant site, milder host-versus-graft reaction, and a marked improvement in motor function. These findings confirm that the transplantation of amniotic epithelial cells combined with silk fibroin scaffold can promote the repair of spinal cord injury. Silk fibroin scaffold can provide a good nerve regeneration microenvironment for amniotic epithelial cells.
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Directory of Open Access Journals (Sweden)
Hua Jin
2015-01-01
Full Text Available The transplantation of embryonic stem cells can effectively improve the creeping strength of nerves near an injury site in animals. Amniotic epithelial cells have similar biological properties as embryonic stem cells; therefore, we hypothesized that transplantation of amniotic epithelial cells can repair peripheral nerve injury and recover the creeping strength of the brachial plexus nerve. In the present study, a brachial plexus injury model was established in rabbits using the C 6 root avulsion method. A suspension of human amniotic epithelial cells was repeatedly injected over an area 4.0 mm lateral to the cephal and caudal ends of the C 6 brachial plexus injury site (1 × 10 6 cells/mL, 3 μL/injection, 25 injections immediately after the injury. The results showed that the decrease in stress and increase in strain at 7,200 seconds in the injured rabbit C 6 brachial plexus nerve were mitigated by the cell transplantation, restoring the viscoelastic stress relaxation and creep properties of the brachial plexus nerve. The forepaw functions were also significantly improved at 26 weeks after injury. These data indicate that transplantation of human amniotic epithelial cells can effectively restore the mechanical properties of the brachial plexus nerve after injury in rabbits and that viscoelasticity may be an important index for the evaluation of brachial plexus injury in animals.
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Institute of Scientific and Technical Information of China (English)
Qi Yang; Min Luo; Peng Li; Hai Jin
2014-01-01
A brachial plexus injury model was established in rabbits by stretching the C6 nerve root. Imme-diately after the stretching, a suspension of human amniotic epithelial cells was injected into the injured brachial plexus. The results of tensile mechanical testing of the brachial plexus showed that the tensile elastic limit strain, elastic limit stress, maximum stress, and maximum strain of the injured brachial plexuses were signiifcantly increased at 24 weeks after the injection. The treat-ment clearly improved the pathological morphology of the injured brachial plexus nerve, as seen by hematoxylin eosin staining, and the functions of the rabbit forepaw were restored. These data indicate that the injection of human amniotic epithelial cells contributed to the repair of brachial plexus injury, and that this technique may transform into current clinical treatment strategies.
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Directory of Open Access Journals (Sweden)
Sangwan Virender
2004-01-01
Full Text Available Purpose: To assess the efficacy of amniotic membrane for treatment of partial limbal stem cell deficiency (LSCD. Methods: Medical records of four patients with partial LSCD who underwent pannus resection and amniotic membrane transplantation (AMT were reviewed for ocular surface stability and improvement in visual acuity. Clinico-histopathological correlation was done with the resected pannus tissue. Results: All the eyes exhibited stable corneal epithelial surface by an average of 7 weeks postoperatively with improvement in subjective symptoms. Best corrected visual acuity improved from preoperative (range: 6/9p-6/120 to postoperative (range: 6/6p-6/15 by an average of 4.5 lines on Snellen visual acuity charts. Histopathological examination of excised tissue showed features of conjunctivalisation. Conclusion: Amniotic membrane transplantation appears to be an effective means of reconstructing the corneal epithelial surface and for visual rehabilitation of patients with partial limbal stem cell deficiency. It may be considered as an alternative primary procedure to limbal transplantation in these cases.
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Directory of Open Access Journals (Sweden)
Lai JY
2013-10-01
Full Text Available Jui-Yang Lai,1–3 David Hui-Kang Ma4,5 1Institute of Biochemical and Biomedical Engineering, 2Biomedical Engineering Research Center, 3Molecular Medicine Research Center, Chang Gung University, Taoyuan, Taiwan; 4Limbal Stem Cell Laboratory, Department of Ophthalmology, Chang Gung Memorial Hospital, Taoyuan, Taiwan; 5Department of Chinese Medicine, Chang Gung University, Taoyuan, Taiwan Abstract: Given that the cells can sense nanometer dimensions, the chemical cross-linking-mediated alteration in fibrillar structure of collagenous tissue scaffolds is critical to determining their cell culture performances. This article explores, for the first time, the effect of nanofibrous structure of glutaraldehyde (GTA cross-linked amniotic membrane (AM on limbal epithelial cell (LEC cultivation. Results of ninhydrin assays demonstrated that the amount of new cross-links formed between the collagen chains is significantly increased with increasing the cross-linking time from 1 to 24 hours. By transmission electron microscopy, the AM treated with GTA for a longer duration exhibited a greater extent of molecular aggregation, thereby leading to a considerable increase in nanofiber diameter and resistance against collagenase degradation. In vitro biocompatibility studies showed that the samples cross-linked with GTA for 24 hours are not well-tolerated by the human corneal epithelial cell cultures. When the treatment duration is less than 6 hours, the biological tissues cross-linked with GTA for a longer time may cause slight reductions in 3-(4,5-dimethylthiazol-2-yl-5-(3-carboxymethoxyphenyl-2-(4-sulfophenyl-2H-tetrazolium, inner salt, and anti-inflammatory activities. Nevertheless, significant collagen molecular aggregation also enhances the stemness gene expression, indicating a high ability of these AM matrices to preserve the progenitors of LECs in vitro. It is concluded that GTA cross-linking of collagenous tissue materials may affect their nanofibrous
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Zhao, B; Wu, G F; Zhang, Y J; Zhang, W; Yang, F F; Xiao, D; Zeng, K X; Shi, J H; Su, L L; Hu, D H
2017-01-20
Objective: To investigate the effects of human amniotic epithelial stem cells-derived exosomes on healing of wound with full-thickness skin defect in rats. Methods: (1) Human amniotic epithelial stem cells were isolated from the amnion tissue of 5 full-term pregnant women in Department of Obstetrics of our hospital by the method of trypsin digestion, and their morphology was observed. The third passage of cells were stained with rhodamine-phalloidin for cytoskeleton observation. The third passage of cells were identified with flow cytometry through the detection of expressions of cell surface markers CD29, CD31, CD34, CD90, CD105, SSEA3, SSEA4 and immunity-related marker human leukocyte antigen-D related site (HLA-DR). The third passage of cells were also assessed the ability of adipogenic and osteogenic differentiation. (2) The third passage of human amniotic epithelial stem cells were cultured in DMEM medium supplemented with 10% exosome-free fetal bovine serum. Exosomes were isolated from culture supernatant by the method of ultracentrifugation and represented with scanning electron microscope for morphologic observation. (3) Six adult SD rats were anesthetized, and four 1 cm×1 cm sized wounds with full-thickness skin defect were made on the back of each rat. The wounds on the back of each rat were divided into control group, 25 μg/mL exosomes group, 50 μg/mL exosomes group, and 100 μg/mL exosomes group according to the random number table (with 6 wounds in each group), and a total volume of 100 μL phosphate buffered saline, 25 μg/mL exosomes, 50 μg/mL exosomes, and 100 μg/mL exosomes were evenly injected around the wound through multiple subcutaneous sites, respectively. The wound healing rate was calculated based on measurement on post injury day (PID) 7, 14, and 21. On PID 21, the healed wound tissue of each group was collected and stained with HE to observe and count skin accessories, and the arrangement of collagen fibers was observed with Masson
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Kim, Tae-Hyun; Park, Young-Woo; Ahn, Jae-Sang; Ahn, Jeong-Taek; Kim, Se-Eun; Jeong, Man-Bok; Seo, Min-Su; Kang, Kyung-Sun
2013-01-01
This study was performed to evaluate the effects of conditioned media (CM) from human amniotic epithelial cells (HAECs) on the corneal wound healing process. Eighteen rabbits (36 eyes) were used and randomly assigned to three groups according treatment: CM from HAECs (group 1), vehicle alone (group 2), and saline (group 3). Corneal alkali injuries were induced with 1 N sodium hydroxide. Each reagent used for treatment evaluation was injected into the dorsal bulbar subconjunctiva and the area of the corneal epithelial defect was measured every other day. Two animals from each group were euthanized at a time on days 3, 7, and 15, and the cornea was removed for histological examination. The sum of the epithelial defect areas measured on day 0 to day 6 as well as day 0 to day 14 in group 1 was significantly smaller than those of other groups. Histological examination revealed that the group 1 corneas had less inflammatory cell infiltration and showed more intact epithelial features compared to the other groups. These results suggest that CM from HAECs promote corneal wound healing in rabbits. PMID:23388445
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Alternative Sources of Adult Stem Cells: Human Amniotic Membrane
Wolbank, Susanne; van Griensven, Martijn; Grillari-Voglauer, Regina; Peterbauer-Scherb, Anja
Human amniotic membrane is a highly promising cell source for tissue engineering. The cells thereof, human amniotic epithelial cells (hAEC) and human amniotic mesenchymal stromal cells (hAMSC), may be immunoprivileged, they represent an early developmental status, and their application is ethically uncontroversial. Cell banking strategies may use freshly isolated cells or involve in vitro expansion to increase cell numbers. Therefore, we have thoroughly characterized the effect of in vitro cultivation on both phenotype and differentiation potential of hAEC. Moreover, we present different strategies to improve expansion including replacement of animal-derived supplements by human platelet products or the introduction of the catalytic subunit of human telomerase to extend the in vitro lifespan of amniotic cells. Characterization of the resulting cultures includes phenotype, growth characteristics, and differentiation potential, as well as immunogenic and immunomodulatory properties.
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Directory of Open Access Journals (Sweden)
Lai JY
2014-11-01
Full Text Available Jui-Yang Lai,1–3 Pei-Ran Wang,1 Li-Jyuan Luo,1 Si-Tan Chen1 1Institute of Biochemical and Biomedical Engineering, 2Biomedical Engineering Research Center, 3Molecular Medicine Research Center, Chang Gung University, Taoyuan, Taiwan, Republic of ChinaAbstract: To overcome the drawbacks associated with limited cross-linking efficiency of carbodiimide modified amniotic membrane, this study investigated the use of L-lysine as an additional amino acid bridge to enhance the stability of a nanofibrous tissue matrix for a limbal epithelial cell culture platform. Results of ninhydrin assays and zeta potential measurements showed that the amount of positively charged amino acid residues incorporated into the tissue collagen chains is highly correlated with the L-lysine -pretreated concentration. The cross-linked structure and hydrophilicity of amniotic membrane scaffolding materials affected by the lysine molecular bridging effects were determined. With an increase in the L-lysine-pretreated concentration from 1 to 30 mM, the cross-linking density was significantly increased and water content was markedly decreased. The variations in resistance to thermal denaturation and enzymatic degradation were in accordance with the number of cross-links per unit mass of amniotic membrane, indicating L-lysine-modulated stabilization of collagen molecules. It was also noteworthy that the carbodiimide cross-linked tissue samples prepared using a relatively high L-lysine-pretreated concentration (ie, 30 mM appeared to have decreased light transmittance and biocompatibility, probably due to the influence of a large nanofiber size and a high charge density. The rise in stemness gene and protein expression levels was dependent on improved cross-link formation, suggesting the crucial role of amino acid bridges in constructing suitable scaffolds to preserve limbal progenitor cells. It is concluded that mild to moderate pretreatment conditions (ie, 3–10 mM L-lysine can
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Ghaderi, Shima; Soheili, Zahra-Soheila; Ahmadieh, Hamid; Davari, Maliheh; Jahromi, Fatemeh Sanie; Samie, Shahram; Rezaie-Kanavi, Mozhgan; Pakravesh, Jalil; Deezagi, Abdolkhalegh
2011-09-01
To evaluate the effect of human amniotic fluid (HAF) on retinal pigmented epithelial cells growth and trans-differentiation into retinal neurons, retinal pigmented epithelium (RPE) cells were isolated from neonatal human cadaver eye globes and cultured in Dulbecco's modified Eagle's medium-F12 supplemented with 10% fetal bovine serum (FBS). Confluent monolayer cultures were trypsinized and passaged using FBS-containing or HAF-containing media. Amniotic fluid samples were received from pregnant women in the first trimester of gestation. Cell proliferation and death enzyme-linked immunosorbent assays were performed to assess the effect of HAF on RPE cell growth. Trans-differentiation into rod photoreceptors and retinal ganglion cells was also studied using immunocytochemistry and real-time polymerase chain reaction techniques. Primary cultures of RPE cells were successfully established under FBS-containing or HAF-containing media leading to rapid cell growth and proliferation. When RPE cells were moved to in vitro culture system, they began to lose their differentiation markers such as pigmentation and RPE65 marker and trans-differentiated neural-like cells followed by spheroid colonies pertaining to stem/progenitor cells were morphologically detected. Immunocytochemistry (ICC) analysis of HAF-treated cultures showed a considerable expression of Rhodopsin gene (30% Rhodopsin-positive cells) indicating trans-differentiation of RPE cells to rod photoreceptors. Real-time polymerase chain reaction revealed an HAF-dose-dependant expression of Thy-1 gene (RGC marker) and significant promoting effect of HAF on RGCs generation. The data presented here suggest that HAF possesses invaluable stimulatory effect on RPE cells growth and trans-differentiation into retinal neurons. It can be regarded as a newly introduced enriched supplement in serum-free kinds of media used in neuro-retinal regeneration studies.
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Kang, N-H; Hwang, K-A; Kim, S U; Kim, Y-B; Hyun, S-H; Jeung, E-B; Choi, K-C
2012-08-01
As stem cells are capable of self-renewal and can generate differentiated progenies for organ development, they are considered as potential source for regenerative medicine and tissue replacement after injury or disease. Along with this capacity, stem cells have the therapeutic potential for treating human diseases including cancers. According to the origins, stem cells are broadly classified into two types: embryonic stem cells (ESCs) and adult stem cells. In terms of differentiation potential, ESCs are pluripotent and adult stem cells are multipotent. Amnion, which is a membranous sac that contains the fetus and amniotic fluid and functions in protecting the developing embryo during gestation, is another stem cell source. Amnion-derived stem cells are classified as human amniotic membrane-derived epithelial stem cells, human amniotic membrane-derived mesenchymal stem cells and human amniotic fluid-derived stem cells. They are in an intermediate stage between pluripotent ESCs and lineage-restricted adult stem cells, non-tumorigenic, and contribute to low immunogenicity and anti-inflammation. Furthermore, they are easily available and do not cause any controversial issues in their recovery and applications. Not only are amnion-derived stem cells applicable in regenerative medicine, they have anticancer capacity. In non-engineered stem cells transplantation strategies, amnion-derived stem cells effectively target the tumor and suppressed the tumor growth by expressing cytotoxic cytokines. Additionally, they also have a potential as novel delivery vehicles transferring therapeutic genes to the cancer formation sites in gene-directed enzyme/prodrug combination therapy. Owing to their own advantageous properties, amnion-derived stem cells are emerging as a new candidate in anticancer therapy.
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The Immunomodulatory Properties of Amniotic Cells: The Two Sides of the Coin.
Magatti, Marta; Vertua, Elsa; Cargnoni, Anna; Silini, Antonietta; Parolini, Ornella
2018-01-01
Among the many cell types useful in developing therapeutic treatments, human amniotic cells from placenta have been proposed as valid candidates. Both human amniotic epithelial and mesenchymal stromal cells, and the conditioned medium generated from their culture, exert multiple immunosuppressive activities. Indeed, they inhibit T and B cell proliferation, suppress inflammatory properties of monocytes, macrophages, dendritic cells, neutrophils, and natural killer cells, while promoting induction of cells with regulatory functions such as regulatory T cells and anti-inflammatory M2 macrophages. These properties have laid the foundation for their use for the treatment of inflammatory-based diseases, and encouraging results have been obtained in different preclinical disease models where exacerbated inflammation is present. Moreover, an immune-privileged status of amniotic cells has been often highlighted. However, even if long-term engraftment of amniotic cells has been reported into immunocompetent animals, only few cells survive after infusion. Furthermore, amniotic cells have been shown to be able to induce immune responses in vivo and, under specific culture conditions, they can stimulate T cell proliferation in vitro. Although immunosuppressive properties are a widely recognized characteristic of amniotic cells, immunogenic and stimulatory activities appear to be less reported, sporadic events. In order to improve therapeutic outcome, the mechanisms responsible for the suppressive versus stimulatory activity need to be carefully addressed. In this review, both the immunosuppressive and immunostimulatory activity of amniotic cells will be discussed.
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Lai, Jui-Yang; Ma, David Hui-Kang
2013-01-01
Given that the cells can sense nanometer dimensions, the chemical cross-linking-mediated alteration in fibrillar structure of collagenous tissue scaffolds is critical to determining their cell culture performances. This article explores, for the first time, the effect of nanofibrous structure of glutaraldehyde (GTA) cross-linked amniotic membrane (AM) on limbal epithelial cell (LEC) cultivation. Results of ninhydrin assays demonstrated that the amount of new cross-links formed between the collagen chains is significantly increased with increasing the cross-linking time from 1 to 24 hours. By transmission electron microscopy, the AM treated with GTA for a longer duration exhibited a greater extent of molecular aggregation, thereby leading to a considerable increase in nanofiber diameter and resistance against collagenase degradation. In vitro biocompatibility studies showed that the samples cross-linked with GTA for 24 hours are not well-tolerated by the human corneal epithelial cell cultures. When the treatment duration is less than 6 hours, the biological tissues cross-linked with GTA for a longer time may cause slight reductions in 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium, inner salt, and anti-inflammatory activities. Nevertheless, significant collagen molecular aggregation also enhances the stemness gene expression, indicating a high ability of these AM matrices to preserve the progenitors of LECs in vitro. It is concluded that GTA cross-linking of collagenous tissue materials may affect their nanofibrous structures and corneal epithelial stem cell culture characteristics. The AM treated with GTA for 6 hours holds promise for use as a niche for the expansion and transplantation of limbal epithelial progenitor cells.
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Liu, Te; Cheng, Weiwei; Liu, Tianjin; Guo, Lihe; Huang, Qin; Jiang, Lizhen; Du, Xiling; Xu, Fuhui; Liu, Zhixue; Lai, Dongmei
2010-02-01
Mouse embryonic stem cells (ESCs) are typically cultured on a feeder layer of mouse embryonic fibroblasts (MEFs), with leukemia inhibitory factor (LIF) added to maintain them in an undifferentiated state. We have previously shown that human amniotic epithelial cells (hAECs) can be used as feeder cells to maintain mouse ESC pluripotency, but the mechanism for this is unknown. In the present study, we found that CpG islands 5' of the c-Myc gene remain hypomethylated in mouse ESCs cultured on hAECs. In addition, levels of acetylation of histone H3 and trimethylation of histone H3K4 in the c-Myc gene promoter were higher in ES cells cultured on hAECs than those in ES cells cultured on MEFs. These data suggested that hAECs can alter mouse ESC gene expression via epigenetic modification of c-Myc, providing a possible mechanism for the hAEC-induced maintenance of ESCs in an undifferentiated state.
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Amniotic membrane-derived stem cells: immunomodulatory properties and potential clinical application
Directory of Open Access Journals (Sweden)
Insausti CL
2014-03-01
Full Text Available Carmen L Insausti,1 Miguel Blanquer,1 Ana M García-Hernández,1 Gregorio Castellanos,2 José M Moraleda11Unidad de Trasplante Hematopoyético y Terapia Celular, 2Servicio de Cirugía, Hospital Clínico Universitario Virgen de la Arrixaca, IMIB, Campus Mare Nostrum, Universidad de Murcia, El Palmar, Murcia, SpainAbstract: Epithelial and mesenchymal cells isolated from the amniotic membrane (AM possess stem cell characteristics, differentiation potential toward lineages of different germ layers, and immunomodulatory properties. While their expansion and differentiation potential have been well studied and characterized, knowledge about their immunomodulatory properties and the mechanisms involved is still incomplete. These mechanisms have been evaluated on various target cells of the innate and the adaptive system and in animal models of different inflammatory diseases. Some results have evidenced that the immunomodulatory effect of AM-derived cells is dependent on cell-cell contact, but many of them have demonstrated that these properties are mediated through the secretion of suppressive molecules. In this review, we present an update on the described immunomodulatory properties of the derived amniotic cells and some of the proposed involved mechanisms. Furthermore, we describe some assays in animal models of different inflammatory diseases which reveal the potential use of these cells to treat such diseases.Keywords: epithelial cells, mesenchymal cells, cell therapy, immunomodulation
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Bone morphogenetic protein-7 promotes chondrogenesis in human amniotic epithelial cells.
Zhou, Junjie; Yu, Guangrong; Cao, Chengfu; Pang, Jinhui; Chen, Xianqi
2011-06-01
Bone morphogenetic proteins (BMPs) play important roles at multiple stages of chondrogenesis. This study was undertaken to investigate the potential role of bone morphogenetic protein-7 (BMP-7) in the differentiation of chondrocytes using tissue engineering techniques. The impact of BMP-7 on human amniotic epithelial cells (hAECs) was tested. The hAECs were treated either with recombinant human BMP-7 cDNA or with transforming growth factor beta 1 (TGF-β1) as a positive control for three weeks in vitro. Cartilaginous differentiation and proliferation were assayed by quantitative RT-PCR, histology, and in situ hybridization. Our results were such that hAECs treated with either BMP-7 or TGF-β1 expressed cartilage markers (aggrecan, Sox9, CEP-68, and type II and X collagens) within three weeks. Compared with a control vector, BMP-7 induced a decrease in type I collagen expression, while the transcription of the cartilage-specific type II collagen remained stable. In induction experiments, BMP-7 transgenic hAECs exhibited the largest amount of matrix synthesis. In conclusion, these data indicate that BMP-7 plays an important role in inducing the production of cartilage by hAECs in vitro. Cartilage differentiation and matrix maturation can be promoted by BMPs in a cartilage engineering paradigm. These properties make BMPs promising tools in the engineering of cartilaginous joint bio-prostheses and as candidate biological agents or genes for cartilage stabilisation.
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Triantafilou, Martha; De Glanville, Benjamin; Aboklaish, Ali F; Spiller, O Brad; Kotecha, Sailesh; Triantafilou, Kathy
2013-01-01
Ureaplasma species are the most frequently isolated microorganisms inside the amniotic cavity and have been associated with spontaneous abortion, chorioamnionitis, premature rupture of the membranes (PROM), preterm labour (PL) pneumonia in neonates and bronchopulmonary dysplasia in neonates. The mechanisms by which Ureaplasmas cause such diseases remain unclear, but it is believed that inappropriate induction of inflammatory responses is involved, triggered by the innate immune system. As part of its mechanism of activation, the innate immune system employs germ-lined encoded receptors, called pattern recognition receptors (PRRs) in order to "sense" pathogens. One such family of PRRs are the Toll like receptor family (TLR). In the current study we aimed to elucidate the role of TLRs in Ureaplasma-induced inflammation in human amniotic epithelial cells. Using silencing, as well as human embryonic kidney (HEK) transfected cell lines, we demonstrate that TLR2, TLR6 and TLR9 are involved in the inflammatory responses against Ureaplasma parvum and urealyticum serovars. Ureaplasma lipoproteins, such as Multiple Banded antigen (MBA), trigger responses via TLR2/TLR6, whereas the whole bacterium is required for TLR9 activation. No major differences were observed between the different serovars. Cell activation by Ureaplasma parvum and urealyticum seem to require lipid raft function and formation of heterotypic receptor complexes comprising of TLR2 and TLR6 on the cell surface and TLR9 intracellularly.
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Directory of Open Access Journals (Sweden)
Martha Triantafilou
Full Text Available Ureaplasma species are the most frequently isolated microorganisms inside the amniotic cavity and have been associated with spontaneous abortion, chorioamnionitis, premature rupture of the membranes (PROM, preterm labour (PL pneumonia in neonates and bronchopulmonary dysplasia in neonates. The mechanisms by which Ureaplasmas cause such diseases remain unclear, but it is believed that inappropriate induction of inflammatory responses is involved, triggered by the innate immune system. As part of its mechanism of activation, the innate immune system employs germ-lined encoded receptors, called pattern recognition receptors (PRRs in order to "sense" pathogens. One such family of PRRs are the Toll like receptor family (TLR. In the current study we aimed to elucidate the role of TLRs in Ureaplasma-induced inflammation in human amniotic epithelial cells. Using silencing, as well as human embryonic kidney (HEK transfected cell lines, we demonstrate that TLR2, TLR6 and TLR9 are involved in the inflammatory responses against Ureaplasma parvum and urealyticum serovars. Ureaplasma lipoproteins, such as Multiple Banded antigen (MBA, trigger responses via TLR2/TLR6, whereas the whole bacterium is required for TLR9 activation. No major differences were observed between the different serovars. Cell activation by Ureaplasma parvum and urealyticum seem to require lipid raft function and formation of heterotypic receptor complexes comprising of TLR2 and TLR6 on the cell surface and TLR9 intracellularly.
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Akrami, Hassan; Soheili, Zahra-Soheila; Sadeghizadeh, Majid; Khalooghi, Keynoush; Ahmadieh, Hamid; Kanavi, Mojgan Rezaie; Samiei, Shahram; Pakravesh, Jalil
2011-06-01
The retinal pigment epithelium (RPE) plays a key role in the maintenance of the normal functions of the retina. Tissue engineering using amniotic membrane as a substrate to culture RPE cells may provide a promising new strategy to replace damaged RPE. We established a method of culturing RPE cells over the amniotic membrane as a support for their growth and transplantation. The transcription of specific genes involved in cellular function of native RPE, including RPE65, CRALBP, VEGF, CD68, and tyrosinase, were then measured using quantitative real-time PCR. Data showed a considerable increase in transcription of RPE65, CD68, and VEGF in RPE cells cultured on amniotic membrane. The amounts of CRALBP and tyrosinase transcripts were not affected. This may simply indicate that amniotic membrane restricted dedifferentiation of RPE cells in culture. The results suggest that amniotic membrane may be considered as an elective biological substrate for RPE cell culture.
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Bioprinted Amniotic Fluid-Derived Stem Cells Accelerate Healing of Large Skin Wounds
Skardal, Aleksander; Mack, David; Kapetanovic, Edi; Atala, Anthony; Jackson, John D.; Yoo, James
2012-01-01
Stem cells obtained from amniotic fluid show high proliferative capacity in culture and multilineage differentiation potential. Because of the lack of significant immunogenicity and the ability of the amniotic fluid-derived stem (AFS) cells to modulate the inflammatory response, we investigated whether they could augment wound healing in a mouse model of skin regeneration. We used bioprinting technology to treat full-thickness skin wounds in nu/nu mice. AFS cells and bone marrow-derived mesenchymal stem cells (MSCs) were resuspended in fibrin-collagen gel and “printed” over the wound site. At days 0, 7, and 14, AFS cell- and MSC-driven wound closure and re-epithelialization were significantly greater than closure and re-epithelialization in wounds treated by fibrin-collagen gel only. Histological examination showed increased microvessel density and capillary diameters in the AFS cell-treated wounds compared with the MSC-treated wounds, whereas the skin treated only with gel showed the lowest amount of microvessels. However, tracking of fluorescently labeled AFS cells and MSCs revealed that the cells remained transiently and did not permanently integrate in the tissue. These observations suggest that the increased wound closure rates and angiogenesis may be due to delivery of secreted trophic factors, rather than direct cell-cell interactions. Accordingly, we performed proteomic analysis, which showed that AFS cells secreted a number of growth factors at concentrations higher than those of MSCs. In parallel, we showed that AFS cell-conditioned media induced endothelial cell migration in vitro. Taken together, our results indicate that bioprinting AFS cells could be an effective treatment for large-scale wounds and burns. PMID:23197691
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Vaghjiani, Vijesh; Cain, Jason E; Lee, William; Vaithilingam, Vijayaganapathy; Tuch, Bernard E; St John, Justin C
2017-10-15
Mitochondrial deoxyribonucleic acid (mtDNA) copy number is tightly regulated during pluripotency and differentiation. There is increased demand of cellular adenosine triphosphate (ATP) during differentiation for energy-intensive cell types such as hepatocytes and neurons to meet the cell's functional requirements. During hepatocyte differentiation, mtDNA copy number should be synchronously increased to generate sufficient ATP through oxidative phosphorylation. Unlike bone marrow mesenchymal cells, mtDNA copy number failed to increase by 28 days of differentiation of human amniotic epithelial cells (hAEC) into hepatocyte-like cells (HLC) despite their expression of some end-stage hepatic markers. This was due to higher levels of DNA methylation at exon 2 of POLGA, the mtDNA-specific replication factor. Treatment with a DNA demethylation agent, 5-azacytidine, resulted in increased mtDNA copy number, reduced DNA methylation at exon 2 of POLGA, and reduced hepatic gene expression. Depletion of mtDNA followed by subsequent differentiation did not increase mtDNA copy number, but reduced DNA methylation at exon 2 of POLGA and increased expression of hepatic and pluripotency genes. We encapsulated hAEC in barium alginate microcapsules and subsequently differentiated them into HLC. Encapsulation resulted in no net increase of mtDNA copy number but a significant reduction in DNA methylation of POLGA. RNAseq analysis showed that differentiated HLC express hepatocyte-specific genes but also increased expression of inflammatory interferon genes. Differentiation in encapsulated cells showed suppression of inflammatory genes as well as increased expression of genes associated with hepatocyte function pathways and networks. This study demonstrates that an increase in classical hepatic gene expression can be achieved in HLC through encapsulation, although they fail to effectively regulate mtDNA copy number.
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* Human Amniotic Mesenchymal Stromal Cells as Favorable Source for Cartilage Repair.
Muiños-López, Emma; Hermida-Gómez, Tamara; Fuentes-Boquete, Isaac; de Toro-Santos, Javier; Blanco, Francisco Javier; Díaz-Prado, Silvia María
2017-09-01
Localized trauma-derived breakdown of the hyaline articular cartilage may progress toward osteoarthritis, a degenerative condition characterized by total loss of articular cartilage and joint function. Tissue engineering technologies encompass several promising approaches with high therapeutic potential for the treatment of these focal defects. However, most of the research in tissue engineering is focused on potential materials and structural cues, while little attention is directed to the most appropriate source of cells endowing these materials. In this study, using human amniotic membrane (HAM) as scaffold, we defined a novel static in vitro model for cartilage repair. In combination with HAM, four different cell types, human chondrocytes, human bone marrow-derived mesenchymal stromal cells (hBMSCs), human amniotic epithelial cells, and human amniotic mesenchymal stromal cells (hAMSCs) were assessed determining their therapeutic potential. A chondral lesion was drilled in human cartilage biopsies simulating a focal defect. A pellet of different cell types was implanted inside the lesion and covered with HAM. The biopsies were maintained for 8 weeks in culture. Chondrogenic differentiation in the defect was analyzed by histology and immunohistochemistry. HAM scaffold showed good integration and adhesion to the native cartilage in all groups. Although all cell types showed the capacity of filling the focal defect, hBMSCs and hAMSCs demonstrated higher levels of new matrix synthesis. However, only the hAMSCs-containing group presented a significant cytoplasmic content of type II collagen when compared with chondrocytes. More collagen type I was identified in the new synthesized tissue of hBMSCs. In accordance, hBMSCs and hAMSCs showed better International Cartilage Research Society scoring although without statistical significance. HAM is a useful material for articular cartilage repair in vitro when used as scaffold. In combination with hAMSCs, HAM showed better
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Directory of Open Access Journals (Sweden)
Jia-Xin Xie
Full Text Available Effective therapy for visual loss caused by optic nerve injury or diseases has not been achieved even though the optic nerve has the regeneration potential after injury. This study was designed to modify amniotic epithelial cells (AECs with basic fibroblast growth factor (bFGF gene, preliminarily investigating its effect on transected optic nerve.A human bFGF gene segment was delivered into rat AECs (AECs/hbFGF by lentiviral vector, and the gene expression was examined by RT-PCR and ELISA. The AECs/hbFGF and untransfected rat AECs were transplanted into the transected site of the rat optic nerve. At 28 days post transplantation, the survival and migration of the transplanted cells was observed by tracking labeled cells; meanwhile retinal ganglion cells (RGCs were observed and counted by employing biotin dextran amine (BDA and Nissl staining. Furthermore, the expression of growth associated protein 43 (GAP-43 within the injury site was examined with immunohistochemical staining.The AECs/hbFGF was proven to express bFGF gene and secrete bFGF peptide. Both AECs/hbFGF and AECs could survive and migrate after transplantation. RGCs counting implicated that RGCs numbers of the cell transplantation groups were significantly higher than that of the control group, and the AECs/hbFGF group was significantly higher than that of the AECs group. Moreover GAP-43 integral optical density value in the control group was significantly lower than that of the cell transplantation groups, and the value in the AECs/hbFGF group was significantly higher than that of the AECs group.AECs modified with bFGF could reduce RGCs loss and promote expression of GAP-43 in the rat optic nerve transected model, facilitating the process of neural restoration following injury.
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Amniotic Fluid Cells Proliferation in Normal and Down Syndrome Subjects
Directory of Open Access Journals (Sweden)
Honcea Adina
2016-02-01
Full Text Available Down Syndrome/Trisomy 21 is the most common chromosomal anomaly, and it represents the most common congenital cause of infants’ intellectual disability. Subjects with this syndrome are affected by degenerative processes caused by accelerated aging or unknown ethyologies. In recent years, accumulating evidence revealed increased potential of amniotic fluid-derived stem cells to be used in regenerative therapy. Our aim was to assess differences in immunophenotype, cell morphology and proliferation of amniotic fluid cells from normal and Down Syndrome pregnancies using a quantitative cytometry approach. Results revealed the emergence of a population of small sized cells in Down Syndrome derived amniotic fluid cells that are readily visible upon microscopic inspection. Hence, the fluorescence–based quantitative image cytometry determinations showed a tendency of decrease in both cell and nuclei size in trisomy, with no significant modification in nuclei circularity, as measured following actin cytoskeleton and nuclei labeling. The propensity of Ki67 positive cells was found to be increased in Down Syndrome derived cells (48.92% as compared to normal specimens (28.68%. However, cells in S and G2/M cell cycle phases decreased from 32.91% to 4.49% in diseased cells. Further studies are devoted to understanding the molecular basis of the observed differences in the proliferation ability of Down Syndrome amniotic cells, in order to evaluate the potential therapeutic effect of amniotic fluid stem cells for tissue regeneration in subjects with trisomy and to find correlations between amniotic cells phenotype and patient prognosis.
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Russo, Valentina [Faculty of Veterinary Medicine, University of Teramo, Campus Universitario Coste S. Agostino Via R. Balzarini 1, 64100 Teramo (Italy); StemTeCh Group (Italy); Tammaro, Loredana [Department of Industrial Engineering, University of Salerno, Via Giovanni Paolo II 132, 84084 Fisciano, SA (Italy); Di Marcantonio, Lisa, E-mail: ldimarcantonio@unite.it [Faculty of Veterinary Medicine, University of Teramo, Campus Universitario Coste S. Agostino Via R. Balzarini 1, 64100 Teramo (Italy); Sorrentino, Andrea [Institute for Polymers, Composite and Biomaterials (IPCB), CNR, P.le Enrico Fermi 1, I-80055 Portici, Napoli (Italy); Ancora, Massimo [Istituto Zooprofilattico Sperimentale dell' Abruzzo e del Molise ‘G. Caporale’, Teramo (Italy); Valbonetti, Luca [Faculty of Veterinary Medicine, University of Teramo, Campus Universitario Coste S. Agostino Via R. Balzarini 1, 64100 Teramo (Italy); StemTeCh Group (Italy); Turriani, Maura; Martelli, Alessandra [Faculty of Veterinary Medicine, University of Teramo, Campus Universitario Coste S. Agostino Via R. Balzarini 1, 64100 Teramo (Italy); Cammà, Cesare [Istituto Zooprofilattico Sperimentale dell' Abruzzo e del Molise ‘G. Caporale’, Teramo (Italy); Barboni, Barbara [Faculty of Veterinary Medicine, University of Teramo, Campus Universitario Coste S. Agostino Via R. Balzarini 1, 64100 Teramo (Italy); StemTeCh Group (Italy)
2016-12-01
Three biodegradable thermoplastic polymers, poly(ε-caprolactone) (PCL), poly(L-lactide-co-D,L-lactide) (PLA) and poly(L-lactide-co-glycolide) (PLGA), have been used to produce nonwovens scaffolds with uniform micrometer fibres. Scaffolds' physical and morphological characterization was performed by X-ray diffraction, Scanning Electron Microscopy and Contact-Angle test. Morphological investigations revealed that all produced fibres were randomly orientated with interconnected pores ranging between 5 and 12 μm in diameter. An average fibre diameter of 1.5, 0.75 and 1.2 μm was found for PCL, PLA and PLGA, respectively. Moreover, experiments were designed to verify whether the fabricated electrospun substrates were biocompatible for ovine amniotic epithelial stem cells (oAECs) under in vitro conditions. Cell adhesion, survival, spatial organization on fibres, proliferation index, and DNA quantification after 48 h culture, showed an enhanced adhesion and proliferation, especially for PLGA scaffolds. The favourable interaction between oAECs and the fibrous scaffolds was attributed to the greatly improved porosity and pore size distribution of the electrospun scaffolds. In addition, AECs can be considered ideal for tissue engineering especially when using biocompatible and opportunely produced scaffolds. - Highlights: • Scaffolds have random oriented, beadless fibres and similar wettability. • Porosity and pore size distribution are determinant on boosting cell activity. • oAECs activities are influenced by scaffold chemical and physical structure. • In PLGA oAECs showed higher spatial distribution efficiency. • PLGA seeded cells present a rise in cell proliferation activity and in DNA amount.
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García-Castro, Irma Lydia; García-López, Guadalupe; Ávila-González, Daniela; Flores-Herrera, Héctor; Molina-Hernández, Anayansi; Portillo, Wendy; Ramón-Gallegos, Eva; Díaz, Néstor Fabián
2015-01-01
Human pluripotent stem cells (hPSC) have promise for regenerative medicine due to their auto-renovation and differentiation capacities. Nevertheless, there are several ethical and methodological issues about these cells that have not been resolved. Human amniotic epithelial cells (hAEC) have been proposed as source of pluripotent stem cells. Several groups have studied hAEC but have reported inconsistencies about their pluripotency properties. The aim of the present study was the in vitro characterization of hAEC collected from a Mexican population in order to identify transcription factors involved in the pluripotency circuitry and to determine their epigenetic state. Finally, we evaluated if these cells differentiate to cortical progenitors. We analyzed qualitatively and quantitatively the expression of the transcription factors of pluripotency (OCT4, SOX2, NANOG, KLF4 and REX1) by RT-PCR and RT-qPCR in hAEC. Also, we determined the presence of OCT4, SOX2, NANOG, SSEA3, SSEA4, TRA-1-60, E-cadherin, KLF4, TFE3 as well as the proliferation and epigenetic state by immunocytochemistry of the cells. Finally, hAEC were differentiated towards cortical progenitors using a protocol of two stages. Here we show that hAEC, obtained from a Mexican population and cultured in vitro (P0-P3), maintained the expression of several markers strongly involved in pluripotency maintenance (OCT4, SOX2, NANOG, TFE3, KLF4, SSEA3, SSEA4, TRA-1-60 and E-cadherin). Finally, when hAEC were treated with growth factors and small molecules, they expressed markers characteristic of cortical progenitors (TBR2, OTX2, NeuN and β-III-tubulin). Our results demonstrated that hAEC express naïve pluripotent markers (KLF4, REX1 and TFE3) as well as the cortical neuron phenotype after differentiation. This highlights the need for further investigation of hAEC as a possible source of hPSC. PMID:26720151
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Effects of maternal subclinical hypothyroidism on amniotic fluid cells oxidative status.
Novakovic, Tanja R; Dolicanin, Zana C; Djordjevic, Natasa Z
2018-06-01
In this study, we researched the effects of maternal subclinical hypothyroidism on the amniotic fluid cells oxidative metabolism during the first trimester of pregnancy. Oxidative stress and damage biomarkers were assayed in the amniotic fluid cells of healthy and pregnant women with subclinical hypothyroidism. Obtained results show that amniotic fluid cells of pregnant women with subclinical hypothyroidism have significantly higher concentrations of oxidative stress biomarkers (superoxide anion, nitric oxide, peroxynitrite) and oxidative damage (lipid peroxide and micronuclei frequency), but lower concentrations of hydrogen peroxide and oxidized glutathione in comparison to healthy pregnant women. We also showed that oxidative stress biomarkers were positively correlated with micronuclei frequency and lipid peroxide concentration in amniotic fluid cells of pregnant women with subclinical hypothyroidism. The present study provides the first evidence for prooxidative effects of maternal subclinical hypothyroidism on the fetus obtained by the estimating oxidative metabolism in the amniotic fluid cells. Copyright © 2018 Elsevier Inc. All rights reserved.
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Amniotic fluid as a source of engraftable stem cells
Directory of Open Access Journals (Sweden)
Cesar V Borlongan
2017-01-01
Full Text Available The ability of stem cells to differentiate into various lineages has made them powerful tools of regenerative medicine and applicable to multiple human diseases. Of particular interest, amniotic fluid-derived stem cells (AFSC have been characterized to express both adult and embryonic cell markers, indicating them as cells within an intermediate stage between embryonic and adult phenotype. AFSC can differentiate into cells of all three germ layers, including hepatic, myogenic, osteogenic, and neurogenic cell types. Furthermore, AFSC have minimal replicative senescence, retaining the ability to divide effectively for over 250 doublings. These facts indicate that amniotic fluid may exist as a promising donor source of stem cells for the treatment of multiple clinically relevant conditions. Of particular interest is the convenience of harvesting stem cells from the amniotic fluid stem for the treatment of newborns, as well as for banking or cryopreserving purposes to be used at a later date. Importantly, the promise of amniotic fluid as a source of stem cells merits ongoing research into their potential therapeutic applications. This paper is a review article. Referred literature in this paper has been listed in the references section. The datasets supporting the conclusions of this article are available online by searching various databases, including PubMed. Some original points in this article come from the laboratory practice in our research center and the authors' experiences.
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Amniotic membrane allografts: development and clinical utility in ophthalmology
Directory of Open Access Journals (Sweden)
Rizzuti A
2014-12-01
Full Text Available Allison Rizzuti,1,2 Adam Goldenberg,1 Douglas R Lazzaro1,2 1SUNY Downstate Medical Center, 2Kings County Hospital Center, Brooklyn, NY, USA Abstract: Amniotic membrane, the innermost layer of the placenta, is a tissue that promotes epithelialization, while decreasing inflammation, neovascularization, and scarring. It is used in the surgical management of a wide variety of ophthalmic conditions where it functions as a graft or patch in ocular surface reconstruction. The development of new preservation techniques, as well as a sutureless amniotic membrane, has allowed for easier, in-office placement, without the disadvantages of an operating room procedure. The purpose of this review is to describe the historical development of amniotic membrane in ophthalmology and to describe its current clinical applications, particularly focusing on recent advances. Keywords: ocular surface, cornea, stem cells, prokera, allograft, patch, transplantation
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Amniotic Fluid Cells Show Higher Pluripotency-Related Gene Expression Than Allantoic Fluid Cells.
Kehl, Debora; Generali, Melanie; Görtz, Sabrina; Geering, Diego; Slamecka, Jaroslav; Hoerstrup, Simon P; Bleul, Ulrich; Weber, Benedikt
2017-10-01
Amniotic fluid represents an abundant source of multipotent stem cells, referred as broadly multipotent given their differentiation potential and expression of pluripotency-related genes. However, the origin of this broadly multipotent cellular fraction is not fully understood. Several sources have been proposed so far, including embryonic and extraembryonic tissues. In this regard, the ovine developmental model uniquely allows for direct comparison of fetal fluid-derived cells from two separate fetal fluid cavities, the allantois and the amnion, over the entire duration of gestation. As allantoic fluid mainly collects fetal urine, cells originating from the efferent urinary tract can directly be compared with cells deriving from the extraembryonic amniotic tissues and the fetus. This study shows isolation of cells from the amniotic [ovine amniotic fluid cells (oAFCs)] and allantoic fluid [ovine allantoic fluid cells (oALCs)] in a strictly paired fashion with oAFCs and oALCs derived from the same fetus. Both cell types showed cellular phenotypes comparable to standard mesenchymal stem cells (MSCs), with trilineage differentiation potential, and expression of common ovine MSC markers. However, the expression of MSC markers per single cell was higher in oAFCs as measured by flow cytometry. oAFCs exhibited higher proliferative capacities and showed significantly higher expression of pluripotency-related genes OCT4, STAT3, NANOG, and REX1 by quantitative real-time polymerase chain reaction compared with paired oALCs. No significant decrease of pluripotency-related gene expression was noted over gestation, implying that cells with high differentiation potential may be isolated at the end of pregnancy. In conclusion, this study suggests that cells with highest stem cell characteristics may originate from the fetus itself or the amniotic fetal adnexa rather than from the efferent urinary tract or the allantoic fetal adnexa.
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Development of a novel method for amniotic fluid stem cell storage.
Zavatti, Manuela; Beretti, Francesca; Casciaro, Francesca; Comitini, Giuseppina; Franchi, Fabrizia; Barbieri, Veronica; Bertoni, Laura; De Pol, Anto; La Sala, Giovanni B; Maraldi, Tullia
2017-08-01
Current procedures for collection of human amniotic fluid stem cells (hAFSCs) indicate that cells cultured in a flask for 2 weeks can then be used for research. However, hAFSCs can be retrieved directly from a small amount of amniotic fluid that can be obtained at the time of diagnostic amniocentesis. The aim of this study was to determine whether direct freezing of amniotic fluid cells is able to maintain or improve the potential of a sub-population of stem cells. We compared the potential of the hAFSCs regarding timing of freezing, cells obtained directly from amniotic fluid aspiration (D samples) and cells cultured in a flask before freezing (C samples). Colony-forming-unit ability, proliferation, morphology, stemness-related marker expression, senescence, apoptosis and differentiation potential of C and D samples were compared. hAFSCs isolated from D samples expressed mesenchymal stem cells markers until later passages, had a good proliferation rate and exhibited differentiation capacity similar to hAFSCs of C samples. Interestingly, direct freezing induced a higher concentration of cells positive for pluripotency stem cell markers, without teratoma formation in vivo. This study suggests that minimal processing may be adequate for the banking of amniotic fluid cells, avoiding in vitro passages before the storage and exposure to high oxygen concentration, which affect stem cell properties. This technique might be a cost-effective and reasonable approach to the process of Good Manufacturing Process accreditation for stem-cell banks. Copyright © 2017 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.
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[Characterization of epithelial primary culture from human conjunctiva].
Rivas, L; Blázquez, A; Muñoz-Negrete, F J; López, S; Rebolleda, G; Domínguez, F; Pérez-Esteban, A
2014-01-01
To evaluate primary cultures from human conjunctiva supplemented with fetal bovine serum, autologous serum, and platelet-rich autologous serum, over human amniotic membrane and lens anterior capsules. One-hundred and forty-eight human conjunctiva explants were cultured in CnT50(®) supplemented with 1, 2.5, 5 and 10% fetal bovine serum, autologous serum and platelet-rich autologous serum. Conjunctival samples were incubated at 37°C, 5% CO2 and 95% HR, for 3 weeks. The typical phenotype corresponding to conjunctival epithelial cells was present in all primary cultures. Conjunctival cultures had MUC5AC-positive secretory cells, K19-positive conjunctival cells, and MUC4-positive non-secretory conjunctival cells, but were not corneal phenotype (cytokeratin K3-negative) and fibroblasts (CD90-negative). Conjunctiva epithelial progenitor cells were preserved in all cultures; thus, a cell culture in CnT50(®) supplemented with 1 to 5% autologous serum over human amniotic membrane can provide better information of epithelial cell differentiation for the conjunctival surface reconstruction. Copyright © 2013 Sociedad Española de Oftalmología. Published by Elsevier Espana. All rights reserved.
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Biological Activity Alterations of Human Amniotic Membrane Pre and Post Irradiation Tissue Banking.
Nemr, Waleed; Bashandy, A S; Araby, Eman; Khamiss, O
Innate immunity of Human Amniotic Membrane (HAM) and its highly active secretome that rich with various types of growth factors and anti-inflammatory substances proposed it as a promising material for many medical studies and applications. This study evaluate the biological activity of cultivated HAM pre and post tissue banking process in which freeze-dried HAM was sterilized by 25 KGray (kGy) dose of γ radiation. The HAM's antimicrobial activity, viability, growth of isolated human amniotic epithelial cells (HAECs), hematopoietic stimulation of co-cultivated murine bone marrow cells (mammalian model), scaffold efficiency for fish brain building up (non-mammalian model) and self re-epithelialization after trypsin denuding treatment were examined as supposed biological activity features. Native HAM revealed viability indications and was active to kill all tested microorganisms; 6 bacterial species (3 Gram-positive and 3 Gram-negative) and Candida albicans as a pathogenic fungus. Also, HAM activity promoted colony formation of murine hematopoietic cells, Tilapia nilotica brain fragment building-up and self re-epithelialization after trypsin treatment. In contrary, radiation-based tissue banking of HAM caused HAM cellular death and consequently lacked almost all of examined biological activity features. Viable HAM was featured with biological activity than fixed HAM prepared by irradiation tissue banking.
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Zhang, Kehua; Cai, Zhe; Li, Yang; Shu, Jun; Pan, Lin; Wan, Fang; Li, Hong; Huang, Xiaojie; He, Chun; Liu, Yanqiu; Cui, Xiaohui; Xu, Yang; Gao, Yan; Wu, Liqun; Cao, Shanxia; Li, Lingsong
2011-08-01
Human embryonic stem (ES) cells are usually maintained in the undifferentiated state by culturing on feeder cells layers of mouse embryonic fibroblasts (MEFs). However, MEFs are not suitable to support human ES cells used for clinical purpose because of risk of zoonosis from animal cells. Therefore, human tissue-based feeder layers need to be developed for human ES cells for clinical purpose. Hereof we report that human amniotic mesenchymal cells (hAMCs) could act as feeder cells for human ES cells, because they are easily obtained and relatively exempt from ethical problem. Like MEFs, hAMCs could act as feeder cells for human ES cells to grow well on. The self-renewal rate of human ES cells cultured on hAMCs feeders was higher than that on MEFs and human amniotic epithelial cells determined by measurement of colonial diameters and growth curve as well as cell cycle analysis. Both immunofluorescence staining and immunoblotting showed that human ES cells cultured on hAMCs expressed stem cell markers such as Oct-3/4, Sox2, and NANOG. Verified by embryoid body formation in vitro and teratoma formation in vivo, we found out that after 20 passages of culture, human ES cells grown on hAMCs feeders could still retain the potency of differentiating into three germ layers. Taken together, our data suggested hAMCs may be safe feeder cells to sustain the propagation of human ES cells in undifferentiated state for future therapeutic use.
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Developmental mechanisms of the tympanic membrane in mammals and non-mammalian amniotes.
Takechi, Masaki; Kitazawa, Taro; Hirasawa, Tatsuya; Hirai, Tamami; Iseki, Sachiko; Kurihara, Hiroki; Kuratani, Shigeru
2016-01-01
The tympanic membrane is a thin layer that originates from the ectoderm, endoderm, and mesenchyme. Molecular-genetic investigations have revealed that interaction between epithelial and mesenchymal cells in the pharyngeal arches is essential for development of the tympanic membrane. We have recently reported that developmental mechanisms underlying the tympanic membrane seem to be different between mouse and chicken, suggesting that the tympanic membrane evolved independently in mammals and non-mammalian amniotes. In this review, we summarize previous studies of tympanic membrane formation in the mouse. We also discuss its formation in amniotes from an evolutionary point of view. © 2015 Japanese Teratology Society.
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Sanie-Jahromi, Fatemeh; Ahmadieh, Hamid; Soheili, Zahra-Soheila; Davari, Maliheh; Ghaderi, Shima; Kanavi, Mozhgan Rezaei; Samiei, Shahram; Deezagi, Abdolkhalegh; Pakravesh, Jalil; Bagheri, Abouzar
2012-04-10
Retinal progenitor cells are a convenient source of cell replacement therapy in retinal degenerative disorders. The purpose of this study was to evaluate the expression patterns of the homeobox genes PAX6 and CHX10 (retinal progenitor markers) during treatment of human retinal pigment epithelium (RPE) cells with amniotic fluid (AF), RPE cells harvested from neonatal cadaver globes were cultured in a mixture of DMEM and Ham's F12 supplemented with 10% FBS. At different passages, cells were trypsinized and co-cultured with 30% AF obtained from normal fetuses of 1416 weeks gestational age. Compared to FBS-treated controls, AF-treated cultures exhibited special morphological changes in culture, including appearance of spheroid colonies, improved initial cell adhesion and ordered cell alignment. Cell proliferation assays indicated a remarkable increase in the proliferation rate of RPE cells cultivated in 30% AF-supplemented medium, compared with those grown in the absence of AF. Immunocytochemical analyses exhibited nuclear localization of retinal progenitor markers at a ratio of 33% and 27% for CHX10 and PAX6, respectively. This indicated a 3-fold increase in retinal progenitor markers in AF-treated cultures compared to FBS-treated controls. Real-time PCR data of retinal progenitor genes (PAX6, CHX10 and VSX-1) confirmed these results and demonstrated AF's capacity for promoting retinal progenitor cell generation. Taken together, the results suggest that AF significantly promotes the rate of retinal progenitor cell generation, indicating that AF can be used as an enriched supplement for serum-free media used for the in vitro propagation of human progenitor cells.
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Moraghebi, Roksana; Kirkeby, Agnete; Chaves, Patricia; Rönn, Roger E; Sitnicka, Ewa; Parmar, Malin; Larsson, Marcus; Herbst, Andreas; Woods, Niels-Bjarne
2017-08-25
Mesenchymal stromal cells (MSCs) are currently being evaluated in numerous pre-clinical and clinical cell-based therapy studies. Furthermore, there is an increasing interest in exploring alternative uses of these cells in disease modelling, pharmaceutical screening, and regenerative medicine by applying reprogramming technologies. However, the limited availability of MSCs from various sources restricts their use. Term amniotic fluid has been proposed as an alternative source of MSCs. Previously, only low volumes of term fluid and its cellular constituents have been collected, and current knowledge of the MSCs derived from this fluid is limited. In this study, we collected amniotic fluid at term using a novel collection system and evaluated amniotic fluid MSC content and their characteristics, including their feasibility to undergo cellular reprogramming. Amniotic fluid was collected at term caesarean section deliveries using a closed catheter-based system. Following fluid processing, amniotic fluid was assessed for cellularity, MSC frequency, in-vitro proliferation, surface phenotype, differentiation, and gene expression characteristics. Cells were also reprogrammed to the pluripotent stem cell state and differentiated towards neural and haematopoietic lineages. The average volume of term amniotic fluid collected was approximately 0.4 litres per donor, containing an average of 7 million viable mononuclear cells per litre, and a CFU-F content of 15 per 100,000 MNCs. Expanded CFU-F cultures showed similar surface phenotype, differentiation potential, and gene expression characteristics to MSCs isolated from traditional sources, and showed extensive expansion potential and rapid doubling times. Given the high proliferation rates of these neonatal source cells, we assessed them in a reprogramming application, where the derived induced pluripotent stem cells showed multigerm layer lineage differentiation potential. The potentially large donor base from caesarean section
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International Nuclear Information System (INIS)
Liu, Te; Cheng, Weiwei; Huang, Yongyi; Huang, Qin; Jiang, Lizhen; Guo, Lihe
2012-01-01
Currently, human induced pluripotent stem (iPS) cells were generated from patient or disease-specific sources and share the same key properties as embryonic stem cells. This makes them attractive for personalized medicine, drug screens or cellular therapy. Long-term cultivation and maintenance of normal iPS cells in an undifferentiated self-renewing state are a major challenge. Our previous studies have shown that human amniotic epithelial cells (HuAECs) could provide a good source of feeder cells for mouse and human embryonic stem cells, or spermatogonial stem cells, but the mechanism for this is unknown. Here, we examined the effect of endogenous microRNA-145 regulation on Sox2 expression in human iPS cells by HuAECs feeder cells regulation, and in turn on human iPS cells pluripotency. We found that human IPS cells transfected with a microRNA-145 mutant expressed Sox2 at high levels, allowing iPS to maintain a high level of AP activity in long-term culture and form teratomas in SCID mice. Expression of stem cell markers was increased in iPS transfected with the microRNA-145 mutant, compared with iPS was transfected with microRNA-145. Besides, the expression of Drosha proteins of the microRNA-processor complex, required for the generation of precursor pre-miRNA, was significantly increased in human iPS cells cultured on MEF but not on HuAECs. Taken together, these results suggest that endogenous Sox2 expression may be regulated by microRNA-145 in human iPS cells with HuAECs feeder cells, and Sox2 is a crucial component required for maintenance of them in an undifferentiated, proliferative state capable of self-renewal. Highlights: ► microRNA-145 inhibits Sox2 expression in human iPS cells. ► microRNA-145 suppresses the self-renewal and pluripotency of human iPS cells. ► HuAECs regulate expression of microRNA-145 and Sox2 in human iPS cells. ► HuAECs feeder layers maintain human iPS cells pluripotency. ► HuAECs negatively regulates the synthesis of
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Energy Technology Data Exchange (ETDEWEB)
Liu, Te, E-mail: liute79@yahoo.com [School of Environmental Science and Engineering, Donghua University, Shanghai 201620 (China); Shanghai Geriatric Institute of Chinese Medicine, Shanghai 200031 (China); Cheng, Weiwei [International Peace Maternity and Child Health Hospital, Shanghai Jiaotong University, Shanghai 200030 (China); Huang, Yongyi [Laboratoire PROTEE, Batiment R, Universite du Sud Toulon-Var, 83957 LA GARDE Cedex (France); Huang, Qin; Jiang, Lizhen [Institute of Biochemistry and Cell Biology, Shanghai Institute for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031 (China); Guo, Lihe, E-mail: liute79@yahoo.com [Institute of Biochemistry and Cell Biology, Shanghai Institute for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031 (China)
2012-02-15
Currently, human induced pluripotent stem (iPS) cells were generated from patient or disease-specific sources and share the same key properties as embryonic stem cells. This makes them attractive for personalized medicine, drug screens or cellular therapy. Long-term cultivation and maintenance of normal iPS cells in an undifferentiated self-renewing state are a major challenge. Our previous studies have shown that human amniotic epithelial cells (HuAECs) could provide a good source of feeder cells for mouse and human embryonic stem cells, or spermatogonial stem cells, but the mechanism for this is unknown. Here, we examined the effect of endogenous microRNA-145 regulation on Sox2 expression in human iPS cells by HuAECs feeder cells regulation, and in turn on human iPS cells pluripotency. We found that human IPS cells transfected with a microRNA-145 mutant expressed Sox2 at high levels, allowing iPS to maintain a high level of AP activity in long-term culture and form teratomas in SCID mice. Expression of stem cell markers was increased in iPS transfected with the microRNA-145 mutant, compared with iPS was transfected with microRNA-145. Besides, the expression of Drosha proteins of the microRNA-processor complex, required for the generation of precursor pre-miRNA, was significantly increased in human iPS cells cultured on MEF but not on HuAECs. Taken together, these results suggest that endogenous Sox2 expression may be regulated by microRNA-145 in human iPS cells with HuAECs feeder cells, and Sox2 is a crucial component required for maintenance of them in an undifferentiated, proliferative state capable of self-renewal. Highlights: Black-Right-Pointing-Pointer microRNA-145 inhibits Sox2 expression in human iPS cells. Black-Right-Pointing-Pointer microRNA-145 suppresses the self-renewal and pluripotency of human iPS cells. Black-Right-Pointing-Pointer HuAECs regulate expression of microRNA-145 and Sox2 in human iPS cells. Black-Right-Pointing-Pointer HuAECs feeder
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Zeng, Wei; Li, Yanwei; Zeng, Guangwei; Yang, Bo; Zhu, Yu
2014-01-01
Amniotic membranes (AM) have been used in a wide range of clinical applications. We successfully extracted mesenchymal stem cells (MSCs) from human AM, but little is known about the use and efficacy of human amniotic membrane-derived mesenchymal stem cells (hAM-dMSCs) for the treatment of alkali burns. We utilized hAM-dMSCs transplantation, AM grafting, and their combined use in the treatment of alkali burns. An experimental model in rabbits was devised to analyze the use of these techniques with immunocytochemistry and ELISA. The survival and migration of hAM-dMSCs labeled by SPION in the host were assessed with Prussian blue staining. Compared with the control group, the treated groups demonstrated faster reconstruction of the corneal epithelium, and lower levels of corneal opacification and neovascularization within corneal alkali burns. Furthermore, dark blue-stained particles were detected in the limbus corneae at day 28. These results demonstrated the ability of hAM-dMSCs to enhance epithelial healing and reduce corneal opacification and neovascularization in corneal alkali wounds.
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Directory of Open Access Journals (Sweden)
Sanie-Jahromi Fatemeh
2012-04-01
Full Text Available Abstract Background Retinal progenitor cells are a convenient source of cell replacement therapy in retinal degenerative disorders. The purpose of this study was to evaluate the expression patterns of the homeobox genes PAX6 and CHX10 (retinal progenitor markers during treatment of human retinal pigment epithelium (RPE cells with amniotic fluid (AF, RPE cells harvested from neonatal cadaver globes were cultured in a mixture of DMEM and Ham's F12 supplemented with 10% FBS. At different passages, cells were trypsinized and co-cultured with 30% AF obtained from normal fetuses of 1416 weeks gestational age. Results Compared to FBS-treated controls, AF-treated cultures exhibited special morphological changes in culture, including appearance of spheroid colonies, improved initial cell adhesion and ordered cell alignment. Cell proliferation assays indicated a remarkable increase in the proliferation rate of RPE cells cultivated in 30% AF-supplemented medium, compared with those grown in the absence of AF. Immunocytochemical analyses exhibited nuclear localization of retinal progenitor markers at a ratio of 33% and 27% for CHX10 and PAX6, respectively. This indicated a 3-fold increase in retinal progenitor markers in AF-treated cultures compared to FBS-treated controls. Real-time PCR data of retinal progenitor genes (PAX6, CHX10 and VSX-1 confirmed these results and demonstrated AF's capacity for promoting retinal progenitor cell generation. Conclusion Taken together, the results suggest that AF significantly promotes the rate of retinal progenitor cell generation, indicating that AF can be used as an enriched supplement for serum-free media used for the in vitro propagation of human progenitor cells.
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Directory of Open Access Journals (Sweden)
Zahra Vojdani
2016-01-01
Full Text Available Objective(s: Umbilical cord blood is a good source of the mesenchymal stem cells that can be banked, expanded and used in regenerative medicine. The objective of this study was to test whether amniotic membrane extract, as a rich source of growth factors such as basic-fibroblast growth factor, can promote the proliferation potential of the umbilical cord mesenchymal stem cells. Materials and Methods: The study design was interventional. Umbilical cord mesenchymal stem cells were isolated from voluntary healthy infants from hospitals in Shiraz, Iran, cultured in the presence of basic-fibroblast growth factor and amniotic membrane extracts (from pooled - samples, and compared with control cultures. Proliferation assay was performed and duplication number and time were calculated. The expression of stem cell’s specific markers and the differentiation capacity toward osteogenic and adipogenic lineages were evaluated. Results: Amniotic membrane extract led to a significant increase in the proliferation rate and duplication number and a decrease in the duplication time without any change in the cell morphology. Both amniotic membrane extract and basic-fibroblast growth factor altered the expressing of CD44 and CD105 in cell population. Treating basic-fibroblast growth factor but not the amniotic membrane extract favored the differentiation potential of the stem cells toward osteogenic lineage. Conclusion: The amniotic membrane extract administration accelerated cell proliferation and modified the CD marker characteristics which may be due to the induction of differentiation toward a specific lineage. Amniotic membrane extract may enhance the proliferation rate and duplication number of the stem cell through changing the duplication time.
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Directory of Open Access Journals (Sweden)
Shiva Gholizadeh-Ghaleh Aziz
Full Text Available Human stem cells and progenitor cells can be used to treat cancer and replace dysfunctional cells within a tissue or organ. The objective of this study was to identify the appropriate cells type in regenerative medicine and targeted therapy. As an alternative to embryonic and bone marrow stem cells, we examined human amniotic fluid stem cells (hAFSCs, one of the potential source of multipotent stem cells isolated from both cell pellet (using single-stage method, and supernatant of human amniotic fluid. Source of isolation and unique property of the cells emphasize that these cells are one of the promising new tools in therapeutic field. Double sources for isolation and availability of the left over samples in diagnostic laboratory at the same time have less legal and ethical concerns compared with embryonic stem cell studies. Cells were isolated, cultured for 18th passage for 6 months and characterized using qPCR and flow cytometry. Cells showed good proliferative ability in culture condition. The cells successfully differentiated into the adipogenic and osteogenic lineages. Based on these findings, amniotic fluid can be considered as an appropriate and convenient source of human amniotic fluid stem cells. These cells provide potential tools for therapeutic applications in the field of regenerative medicine. To get a better understanding of crosstalk between Oct4/NANOG with osteogenesis and adipogenesis, we used network analysis based on Common Targets algorithm and Common Regulators algorithm as well as subnetwork discovery based on gene set enrichment. Network analysis highlighted the possible role of MIR 302A and MIR let-7g. We demonstrated the high expression of MIR 302A and low expression of MIR let7g in hAFSCs by qPCR.
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Topoluk, Natasha; Hawkins, Richard; Tokish, John; Mercuri, Jeremy
2017-09-01
Therapeutic efficacy of various mesenchymal stromal cell (MSC) types for orthopaedic applications is currently being investigated. While the concept of MSC therapy is well grounded in the basic science of healing and regeneration, little is known about individual MSC populations in terms of their propensity to promote the repair and/or regeneration of specific musculoskeletal tissues. Two promising MSC sources, adipose and amnion, have each demonstrated differentiation and extracellular matrix (ECM) production in the setting of musculoskeletal tissue regeneration. However, no study to date has directly compared the differentiation potential of these 2 MSC populations. To compare the ability of human adipose- and amnion-derived MSCs to undergo osteogenic and chondrogenic differentiation. Controlled laboratory study. MSC populations from the human term amnion were quantified and characterized via cell counting, histologic assessment, and flow cytometry. Differentiation of these cells in comparison to commercially purchased human adipose-derived mesenchymal stromal cells (hADSCs) in the presence and absence of differentiation media was evaluated via reverse transcription polymerase chain reaction (PCR) for bone and cartilage gene transcript markers and histology/immunohistochemistry to examine ECM production. Analysis of variance and paired t tests were performed to compare results across all cell groups investigated. The authors confirmed that the human term amnion contains 2 primary cell types demonstrating MSC characteristics-(1) human amniotic epithelial cells (hAECs) and (2) human amniotic mesenchymal stromal cells (hAMSCs)-and each exhibited more than 90% staining for MSC surface markers (CD90, CD105, CD73). Average viable hAEC and hAMSC yields at harvest were 2.3 × 10 6 ± 3.7 × 10 5 and 1.6 × 10 6 ± 4.7 × 10 5 per milliliter of amnion, respectively. As well, hAECs and hAMSCs demonstrated significantly greater osteocalcin ( P = .025), aggrecan ( P
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Multilineage Potential Research of Bovine Amniotic Fluid Mesenchymal Stem Cells
Directory of Open Access Journals (Sweden)
Yuhua Gao
2014-02-01
Full Text Available The use of amnion and amniotic fluid (AF are abundant sources of mesenchymal stem cells (MSCs that can be harvested at low cost and do not pose ethical conflicts. In human and veterinary research, stem cells derived from these tissues are promising candidates for disease treatment, specifically for their plasticity, their reduced immunogenicity, and high anti-inflammatory potential. This work aimed to obtain and characterize bovine amniotic fluid mesenchymal stem cells (AFMSC. The bovine AF from the amniotic cavity of pregnant gilts in the early stages of gestation (3- and 4-m-old bovine embryos was collected. AFMSCs exhibit a fibroblastic-like morphology only starting from the fourth passage, being heterogeneous during the primary culture. Immunofluorescence results showed that AFMSCs were positive for β-integrin, CD44, CD73 and CD166, but negative for CD34, CD45. Meanwhile, AFMSCs expressed ES cell markers, such as Oct4, and when appropriately induced, are capable of differentiating into ectodermal and mesodermal lineages. This study reinforces the emerging importance of these cells as ideal tools in veterinary medicine; future studies aimed at a deeper evaluation of their immunological properties will allow a better understanding of their role in cellular therapy.
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Davari, Maliheh; Soheili, Zahra-Soheila; Ahmadieh, Hamid; Sanie-Jahromi, Fateme; Ghaderi, Shima; Kanavi, Mozhgan Rezaei; Samiei, Shahram; Akrami, Hassan; Haghighi, Massoud; Javidi-Azad, Fahimeh
2013-01-01
Retinal pigment epithelial (RPE) cells are capable of differentiating into retinal neurons when induced by the appropriate growth factors. Amniotic fluid contains a variety of growth factors that are crucial for the development of a fetus. In this study, the effects of human amniotic fluid (HAF) on primary RPE cell cultures were evaluated. RPE cells were isolated from the globes of postnatal human cadavers. The isolated cells were plated and grown in DMEM/F12 with 10% fetal bovine serum. To confirm the RPE identity of the cultured cells, they were immunocytochemically examined for the presence of the RPE cell-specific marker RPE65. RPE cultures obtained from passages 2-7 were treated with HAF and examined morphologically for 1 month. To determine whether retinal neurons or progenitors developed in the treated cultures, specific markers for bipolar (protein kinase C isomer α, PKCα), amacrine (cellular retinoic acid-binding protein I, CRABPI), and neural progenitor (NESTIN) cells were sought, and the amount of mRNA was quantified using real-time PCR. Treating RPE cells with HAF led to a significant decrease in the number of RPE65-positive cells, while PKCα- and CRABPI-positive cells were detected in the cultures. Compared with the fetal bovine serum-treated cultures, the levels of mRNAs quantitatively increased by 2-, 20- and 22-fold for NESTIN, PKCα, and CRABPI, respectively. The RPE cultures treated with HAF established spheres containing both pigmented and nonpigmented cells, which expressed neural progenitor markers such as NESTIN. This study showed that HAF can induce RPE cells to transdifferentiate into retinal neurons and progenitor cells, and that it provides a potential source for cell-based therapies to treat retinal diseases.
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Lal, Ikeda; Panchal, Bhavik Uttam; Basu, Sayan; Sangwan, Virender S
2013-01-01
A 20-year-old man from Bangladesh suffered accidental alkali injury to his right eye in May 2010 leading to total limbal stem cell deficiency. An amniotic membrane graft was performed 5 days after the accident and the patient presented to our institute 6 months later. On ocular examination, his best corrected visual acuity (BCVA) was 20/50 with a 360° pannus at the periphery and central area was spared but had stromal scarring. He underwent simple limbal epithelial transplantation (SLET) taki...
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Tidu, Aurélien; Ghoubay-Benallaoua, Djida; Teulon, Claire; Asnacios, Sophie; Grieve, Kate; Portier, François; Schanne-Klein, Marie-Claire; Borderie, Vincent; Mosser, Gervaise
2018-05-29
This study aimed at controlling both the organization and the transparency of dense collagen scaffolds making use of the lyotropic mesogen properties of collagen. Cholesteric or plywood-like liquid crystal phases were achieved using mixtures of acetic and hydrochloric acids as solvents. The critical pH at which the switch between the two phases occurred was around pH = 3. The use of the two acids led to fibrillated collagen I scaffolds, whose visual aspect ranged from opaque to transparent. Rheological investigations showed that viscoelastic properties of the plywood-like solutions were optimized for molding due to faster recovery. They also confirmed the correlation between the elastic modulus and the diameter of collagen fibrils obtained after fibrillogenesis under ammonia vapor. Human corneal epithelial cells, grown from donor limbal explants, were cultured both on transparent plywood-like matrices and on human amniotic membranes for 14 days. The development of corneal epithelium and the preservation of epithelial stem cells were checked by optical microscopy, colony formation assay, immuno-fluorescence and quantitative polymerase chain reaction. A higher level of amplification of limbal stem cells was obtained with collagen matrices compared with amniotic membranes, showing the high biocompatibility of our scaffolds. We therefore suggest that collagen solutions presenting both plywood-like organization and transparency might be of interest for biomedical applications in ophthalmology.
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Loukogeorgakis, Stavros P; De Coppi, Paolo
2017-07-01
The amniotic fluid has been identified as an untapped source of cells with broad potential, which possess immunomodulatory properties and do not have the ethical and legal limitations of embryonic stem cells. CD117(c-Kit)+ cells selected from amniotic fluid have been shown to differentiate into cell lineages representing all three embryonic germ layers without generating tumors, making them ideal candidates for regenerative medicine applications. Moreover, their ability to engraft in injured organs and modulate immune and repair responses of host tissues, suggest that transplantation of such cells may be useful for the treatment of various degenerative and inflammatory diseases. Although significant questions remain regarding the origin, heterogeneous phenotype, and expansion potential of amniotic fluid stem cells, evidence to date supports their potential role as a valuable stem cell source for the field of regenerative medicine. Stem Cells 2017;35:1663-1673. © 2016 AlphaMed Press.
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Liu, Junfeng; Hua, Rong; Gong, Zhangbin; Shang, Bin; Huang, Yongyi; Guo, Lihe; Liu, Te; Xue, Jun
2017-01-01
In the pathogenesis of acute kidney injury (AKI), the release of multiple interleukins can lead to increased kidney damage. Human amniotic epithelial cells (HuAECs) can inhibit immune cell activation in vivo and in vitro. We hypothesized that HuAECs could weaken patient-derived peripheral blood CD4+ T-cell activation and decreasing the ability of these cells to express and release IL-2. -Cell proliferation assay revealed that under the same culture conditions, activated AKI patient-derived CD4+ T cells had a significantly reduced proliferation rate when were co-cultured with HuAECs. And the level of IL-2 released was also significantly reduced. Western blot and qRT-PCR assays showed that the expression of c-Rel in the CD4+ T cells was also significantly reduced. However, the expression level of endogenous miR-101 in the CD4+ T cells co-cultured with HuAECs was significantly increased. Luciferase reporter assay results suggested that miR-101 could bind to a specific site in the c-Rel 3' UTR and induce the post-transcriptional silencing of c-Rel. Subsequently, we over-expressed miR-101 in AKI patient-derived CD4+ T cells. The qRT-PCR and western blot assay results revealed that the expression of endogenous c-Rel was significantly reduced, while the ELISA results indicated that the level of IL-2 released was also significantly decreased. Finally, ChIP-PCR assay results showed that the miR-101-overexpressing CD4+ T-cell group and the HuAEC co-culture CD4+ T-cell group exhibited significantly decreased binding capacities between the 'c-Rel-NFκB' complex and the IL-2 gene promoter, and the transcriptional activity of IL-2 was also significantly decreased. Therefore, we confirmed that HuAECs can stimulate miR-101 expression in AKI patient-derived peripheral blood CD4+ T cells, thus inhibiting the expression of the miR-101 target gene c-Rel and leading to a reduction in IL-2 expression and release. Copyright © 2016 Elsevier Ltd. All rights reserved.
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Applications of Amniotic Membrane and Fluid in Stem Cell Biology and Regenerative Medicine
Directory of Open Access Journals (Sweden)
Kerry Rennie
2012-01-01
Full Text Available The amniotic membrane (AM and amniotic fluid (AF have a long history of use in surgical and prenatal diagnostic applications, respectively. In addition, the discovery of cell populations in AM and AF which are widely accessible, nontumorigenic and capable of differentiating into a variety of cell types has stimulated a flurry of research aimed at characterizing the cells and evaluating their potential utility in regenerative medicine. While a major focus of research has been the use of amniotic membrane and fluid in tissue engineering and cell replacement, AM- and AF-derived cells may also have capabilities in protecting and stimulating the repair of injured tissues via paracrine actions, and acting as vectors for biodelivery of exogenous factors to treat injury and diseases. Much progress has been made since the discovery of AM and AF cells with stem cell characteristics nearly a decade ago, but there remain a number of problematic issues stemming from the inherent heterogeneity of these cells as well as inconsistencies in isolation and culturing methods which must be addressed to advance the field towards the development of cell-based therapies. Here, we provide an overview of the recent progress and future perspectives in the use of AM- and AF-derived cells for therapeutic applications.
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Ginsberg, Michael; Schachterle, William; Shido, Koji; Rafii, Shahin
2016-01-01
Endothelial cells (ECs) have essential roles in organ development and regeneration, and therefore they could be used for regenerative therapies. However, generation of abundant functional endothelium from pluripotent stem cells has been difficult because ECs generated by many existing strategies have limited proliferative potential and display vascular instability. The latter difficulty is of particular importance because cells that lose their identity over time could be unsuitable for therapeutic use. Here, we describe a 3-week platform for directly converting human mid-gestation lineage-committed amniotic fluid–derived cells (ACs) into a stable and expandable population of vascular ECs (rAC-VECs) without using pluripotency factors. By transient expression of the ETS transcription factor ETV2 for 2 weeks and constitutive expression the ETS transcription factors FLI1 and ERG1, concomitant with TGF-β inhibition for 3 weeks, epithelial and mesenchymal ACs are converted, with high efficiency, into functional rAC-VECs. These rAC-VECs maintain their vascular repertoire and morphology over numerous passages in vitro, and they form functional vessels when implanted in vivo. rAC-VECs can be detected in recipient mice months after implantation. Thus, rAC-VECs can be used to establish a cellular platform to uncover the molecular determinants of vascular development and heterogeneity and potentially represent ideal ECs for the treatment of regenerative disorders. PMID:26540589
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International Nuclear Information System (INIS)
Tsuno, Hiroaki; Yoshida, Toshiko; Nogami, Makiko; Koike, Chika; Okabe, Motonori; Noto, Zenko; Arai, Naoya; Noguchi, Makoto; Nikaido, Toshio
2012-01-01
Autogenous mesenchymal stem cells (MSCs) have therapeutic applications in bone regenerative therapy due to their pluripotency. However, the ability of MSCs to proliferate and differentiate varies between donors. Furthermore, alternative sources of MSCs are required for patients with contraindications to autogenous cell therapy. The aim of this study was to evaluate the potential of mesenchymal cells from the human amniotic membrane (HAM) as a source of cells for allogeneic transplantation in bone regenerative therapy. Cells that retained a proliferative capacity of more than 50 population doubling level were distinguished from other HAM cells as HAMα cells and induced to osteogenic status—their in vivo osteogenesis was subsequently investigated in rats. It was found that HAMα cells were spindle shaped and were positive for MSC markers and negative for hematopoietic stem cell markers. Alkaline phosphatase activity and calcium deposition increased with osteogenic status of HAMα cells. The expression of osteocalcin mRNA was increased in HAMα cells cultured on calcium phosphate scaffolds. Moreover, xenografted HAMα cells remained viable and produced extracellular matrix for several weeks. Thus, this study suggests that human amniotic mesenchymal cells possess osteogenic differentiation potential and could be applied to allogeneic transplantation in bone regenerative therapy. - Highlights: ► Human amniotic mesenchymal cells include cells (HAMα cells) that have the properties of MSCs. ► HAMα cells have excellent osteogenic differentiation potential. ► Osteogenic differentiation ability of HAMα was amplified by calcium phosphate scaffolds. ► HAMα cells can be applicable to allogeneic cell transplantation in bone regenerative therapy.
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Directory of Open Access Journals (Sweden)
Annalucia Carbone
2018-01-01
Full Text Available We previously found that human amniotic mesenchymal stem cells (hAMSCs in coculture with CF immortalised airway epithelial cells (CFBE41o- line, CFBE on Transwell® filters acquired an epithelial phenotype and led to the expression of a mature and functional CFTR protein. In order to explore the role of gap junction- (GJ- mediated intercellular communication (GJIC in this rescue, cocultures (hAMSC : CFBE, 1 : 5 ratio were studied for the formation of GJIC, before and after silencing connexin 43 (Cx43, a major component of GJs. Functional GJs in cocultures were inhibited when the expression of the Cx43 protein was downregulated. Transfection of cocultures with siRNA against Cx43 resulted in the absence of specific CFTR signal on the apical membrane and reduction in the mature form of CFTR (band C, and in parallel, the CFTR-dependent chloride channel activity was significantly decreased. Cx43 downregulation determined also a decrease in transepithelial resistance and an increase in paracellular permeability as compared with control cocultures, implying that GJIC may regulate CFTR expression and function that in turn modulate airway epithelium tightness. These results indicate that GJIC is involved in the correction of CFTR chloride channel activity upon the acquisition of an epithelial phenotype by hAMSCs in coculture with CF cells.
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Carbone, Annalucia; Zefferino, Roberto; Beccia, Elisa; Casavola, Valeria; Castellani, Stefano; Di Gioia, Sante; Giannone, Valentina; Seia, Manuela; Angiolillo, Antonella; Colombo, Carla; Favia, Maria; Conese, Massimo
2018-01-01
We previously found that human amniotic mesenchymal stem cells (hAMSCs) in coculture with CF immortalised airway epithelial cells (CFBE41o- line, CFBE) on Transwell® filters acquired an epithelial phenotype and led to the expression of a mature and functional CFTR protein. In order to explore the role of gap junction- (GJ-) mediated intercellular communication (GJIC) in this rescue, cocultures (hAMSC : CFBE, 1 : 5 ratio) were studied for the formation of GJIC, before and after silencing connexin 43 (Cx43), a major component of GJs. Functional GJs in cocultures were inhibited when the expression of the Cx43 protein was downregulated. Transfection of cocultures with siRNA against Cx43 resulted in the absence of specific CFTR signal on the apical membrane and reduction in the mature form of CFTR (band C), and in parallel, the CFTR-dependent chloride channel activity was significantly decreased. Cx43 downregulation determined also a decrease in transepithelial resistance and an increase in paracellular permeability as compared with control cocultures, implying that GJIC may regulate CFTR expression and function that in turn modulate airway epithelium tightness. These results indicate that GJIC is involved in the correction of CFTR chloride channel activity upon the acquisition of an epithelial phenotype by hAMSCs in coculture with CF cells.
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Gholizadeh-Ghaleh Aziz, Shiva; Fathi, Ezzatollah; Rahmati-Yamchi, Mohammad; Akbarzadeh, Abolfazl; Fardyazar, Zahra; Pashaiasl, Maryam
2017-06-01
Recent studies have elucidated that cell-based therapies are promising for cancer treatments. The human amniotic fluid stem (AFS) cells are advantageous cells for such therapeutic schemes that can be innately changed to express therapeutic proteins. HAFSCs display a natural tropism to cancer cells in vivo. They can be useful in cancer cells targeting. Moreover, they are easily available from surplus diagnostic samples during pregnancy and less ethical and legal concern are associated with the collection and application than other putative cells are subjected. This review will designate representatives of amniotic fluid and stem cell derived from amniotic fluid. For this propose, we collect state of human AFS cells data applicable in cancer therapy by dividing this approach into two main classes (nonengineered and engineered based approaches). Our study shows the advantage of AFS cells over other putative cells types in terms differentiation ability to a wide range of cells by potential and effective use in preclinical studies for a variety of diseases. This study has shown the elasticity of human AFS cells and their favorable potential as a multipotent cell source for regenerative stem cell therapy and capable of giving rise to multiple lineages including such as osteoblasts and adipocyte.
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Directory of Open Access Journals (Sweden)
Hsiu-Huei Peng
2012-09-01
Conclusion: In conclusion, human amniotic fluid mesenchymal stem cells contain a unique epigenetic signature during in vitro cell culture. H19 and KCNQ1OT1 possessed a substantial degree of hypermethylation status, and variable DNA methylation patterns of SNRPN was observed during in vitro cell culture of human amniotic fluid mesenchymal stem cells. Our results urge further understanding of epigenetic status of human amniotic fluid mesenchymal stem cells before it is applied in cell replacement therapy.
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In Vitro Cardiomyogenic Potential of Human Amniotic Fluid Stem Cells
Guan, Xuan; Delo, Dawn M.; Atala, Anthony; Soker, Shay
2011-01-01
Stem cell therapy for damaged cardiac tissue is currently limited by a number of factors, including the inability to obtain sufficient cell numbers, the potential tumorigenicity of certain types of stem cells, and the possible link between stem cell therapy and the development of malignant arrhythmias. In this study, we investigated whether human amniotic fluid-derived stem (hAFS) cells could be a potential source of cells for cardiac cell therapy by testing the in vitro differentiation capab...
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Energy Technology Data Exchange (ETDEWEB)
Tsuno, Hiroaki [Department of Regenerative Medicine, Graduate School of Medicine and Pharmaceutical Sciences for Research, University of Toyama, 2630 Sugitani Toyama, Toyama 930-0194 (Japan); Department of Oral and Maxillofacial Surgery, Graduate School of Medicine and Pharmaceutical Sciences for Research, University of Toyama, 2630 Sugitani Toyama, Toyama 930-0194 (Japan); Yoshida, Toshiko [Department of Regenerative Medicine, Graduate School of Medicine and Pharmaceutical Sciences for Research, University of Toyama, 2630 Sugitani Toyama, Toyama 930-0194 (Japan); Nogami, Makiko [Department of Regenerative Medicine, Graduate School of Medicine and Pharmaceutical Sciences for Research, University of Toyama, 2630 Sugitani Toyama, Toyama 930-0194 (Japan); Department of Orthopedic Surgery, Graduate School of Medicine and Pharmaceutical Sciences for Research, University of Toyama, 2630 Sugitani Toyama, Toyama 930-0194 (Japan); Koike, Chika; Okabe, Motonori [Department of Regenerative Medicine, Graduate School of Medicine and Pharmaceutical Sciences for Research, University of Toyama, 2630 Sugitani Toyama, Toyama 930-0194 (Japan); Noto, Zenko [Department of Regenerative Medicine, Graduate School of Medicine and Pharmaceutical Sciences for Research, University of Toyama, 2630 Sugitani Toyama, Toyama 930-0194 (Japan); Department of Oral and Maxillofacial Surgery, Graduate School of Medicine and Pharmaceutical Sciences for Research, University of Toyama, 2630 Sugitani Toyama, Toyama 930-0194 (Japan); Arai, Naoya; Noguchi, Makoto [Department of Oral and Maxillofacial Surgery, Graduate School of Medicine and Pharmaceutical Sciences for Research, University of Toyama, 2630 Sugitani Toyama, Toyama 930-0194 (Japan); Nikaido, Toshio, E-mail: tnikaido@med.u-toyama.ac.jp [Department of Regenerative Medicine, Graduate School of Medicine and Pharmaceutical Sciences for Research, University of Toyama, 2630 Sugitani Toyama, Toyama 930-0194 (Japan)
2012-12-01
Autogenous mesenchymal stem cells (MSCs) have therapeutic applications in bone regenerative therapy due to their pluripotency. However, the ability of MSCs to proliferate and differentiate varies between donors. Furthermore, alternative sources of MSCs are required for patients with contraindications to autogenous cell therapy. The aim of this study was to evaluate the potential of mesenchymal cells from the human amniotic membrane (HAM) as a source of cells for allogeneic transplantation in bone regenerative therapy. Cells that retained a proliferative capacity of more than 50 population doubling level were distinguished from other HAM cells as HAM{alpha} cells and induced to osteogenic status-their in vivo osteogenesis was subsequently investigated in rats. It was found that HAM{alpha} cells were spindle shaped and were positive for MSC markers and negative for hematopoietic stem cell markers. Alkaline phosphatase activity and calcium deposition increased with osteogenic status of HAM{alpha} cells. The expression of osteocalcin mRNA was increased in HAM{alpha} cells cultured on calcium phosphate scaffolds. Moreover, xenografted HAM{alpha} cells remained viable and produced extracellular matrix for several weeks. Thus, this study suggests that human amniotic mesenchymal cells possess osteogenic differentiation potential and could be applied to allogeneic transplantation in bone regenerative therapy. - Highlights: Black-Right-Pointing-Pointer Human amniotic mesenchymal cells include cells (HAM{alpha} cells) that have the properties of MSCs. Black-Right-Pointing-Pointer HAM{alpha} cells have excellent osteogenic differentiation potential. Black-Right-Pointing-Pointer Osteogenic differentiation ability of HAM{alpha} was amplified by calcium phosphate scaffolds. Black-Right-Pointing-Pointer HAM{alpha} cells can be applicable to allogeneic cell transplantation in bone regenerative therapy.
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He, Xiao-Ying; Zheng, Yue-Mao; Qiu, Shuang; Qi, Ying-Pei; Zhang, Yong
2011-08-01
The aims of this study were to determine whether stem cells could be isolated from amniotic fluid of goat fetus at terminal gestational age and to determine if these stem cells could differentiate into adipogenic cells and be transfected with a reporter gene, EGFP (enhanced green fluorescent protein). The stem cells were isolated from amniotic fluid of goat fetus at terminal gestational age, induced to differentiate into adipogenic cells in vitro and transfected with the EGFP gene using lipofection. Markers associated with undifferentiated AFS (amniotic fluid-derived stem) cells were tested by RT (reverse transcription)-PCR. The results demonstrated that AFS cells could be isolated from amniotic fluid of goat fetus at terminal gestational age and could differentiate into adipogenic cells. The EGFP gene was transfected into AFS cells successfully. EGFP gene transfection efficiency of the three groups of transgenic AFS cells were 26.0, 29.9 and 30.5%, respectively. Both transgenic and wild-type AFS cells could express Hes1 (hairy and enhancer of split 1), Oct4 (octamer-binding protein 4) and Nanog.
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Directory of Open Access Journals (Sweden)
Elly Munadziroh
2008-09-01
Full Text Available Background: Secretory leukocyte protease inhibitor (SLPI has been found to facilitate epithelialization, maintain a normal epithelial phenotype, reduce inflammation, secrete growth factors such as IL-4, IL-6, IL-10, EGF, FGF, TGF, HGFand 2-microbulin. SLPI is serine protease inhibitor, which found in secretions such as whole saliva, seminal fluid, cervical mucus, synovial fluid, breast milk, tears, amniotic fluid and amniotic membrane. Impaired healing states are characterized by excessive proteolysis and oftenbacterial infection, leading to the hypothesis that SLPI may have a role in the healing process in oral inflammation and contributes to tissue repair in oral mucosa. The oral wound healing response is impaired in the SLPI sufficient mice since matrix synthesis and collagen deposition delayed. The objective of this research is to isolate and identify the amniotic membrane of Java Race SLPI Gene. Methods: SLPI RNA was isolated from Java Race amniotic membrane and the cDNA was amplified by polymerase chain reaction (PCR. Result: Through sequence analyses, SLPI cDNA was 530 nucleotide in length with a predicted molecular mass about 12 kDa. The nucleotide sequence showed that human SLPI from sample was 98% identical with human SLPI from gene bank. PCR analysis revealed that the mRNA of SLPI was highly expressed in the amniotic membrane from Java Race sample. Conclusion: it is demonstrated that human SLPI are highly conserved in sequence content as compared to the human SLPI from gene.
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Some biological properties of the human amniotic membrane interferon
Directory of Open Access Journals (Sweden)
P. C. P. Ferreira
1992-03-01
Full Text Available Human amniotic interferon was investigated to define the species specificity of its antiviral action and compare its anti-cellular and NK cell stimulating activities with those of other human interferons. The antiviral effect was titrated in bovine (RV-IAL and monkey (VERO cells. Amniotic interferon exhibited, in bovine cells, 5% of the activity seen in monkey cells, while alpha interferon displayed 200%. No effect was detected with either beta or gamma interferon in bovine cells. Daudi cells were exposed to different concentrations of various interferons and the cell numbers were determined. The anticellular effect of the amniotic interferon reached its peak on the third day of incubation. Results suggested a higher activity for alpha and gamma interferons and a lower activity for beta when compared to amniotic interferon. Using total mononuclear cells as effector cells and K 562 as target cell in a 51Cr release assay, it was demonstrated that low concentrations of amniotic interferon consistently stimulated NK cell activity in cells derived from several donors, the results indicating a higher level of activity with this interferon than with alpha and beta interferons.
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Directory of Open Access Journals (Sweden)
Liru Li
Full Text Available The mesenchymal stem cells (MSCs derived from amniotic fluid (AF have become an attractive stem cells source for cell-based therapy because they can be harvested at low cost and avoid ethical disputes. In human research, stem cells derived from AF gradually became a hot research direction for disease treatment, specifically for their plasticity, their reduced immunogenicity and their tumor tropism regardless of the tumor size, location and source. Our work aimed to obtain and characterize human amniotic fluid mesenchymal stem cells (AFMSCs and detect their ovarian cancer tropsim in nude mice model. Ten milliliters of twenty independent amniotic fluid samples were collected from 16-20 week pregnant women who underwent amniocentesis for fetal genetic determination in routine prenatal diagnosis in the first affiliated hospital of Harbin medical university. We successfully isolated the AFMSCs from thirteen of twenty amniotic fluid samples. AFMSCs presented a fibroblastic-like morphology during the culture. Flow cytometry analyses showed that the cells were positive for specific stem cell markers CD73,CD90, CD105, CD166 and HLA-ABC (MHC class I, but negative for CD 45,CD40, CD34, CD14 and HLA-DR (MHC class II. RT-PCR results showed that the AFMSCs expressed stem cell marker OCT4. AFMSCs could differentiate into bone cells, fat cells and chondrocytes under certain conditions. AFMSCs had the high motility to migrate to ovarian cancer site but didn't have the tumorigenicity. This study enhances the possibility of AFMSCs as drug carrier in human cell-based therapy. Meanwhile, the research emphasis in the future can also put in targeting therapy of ovarian cancer.
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Myogel supports the ex-vivo amplification of corneal epithelial cells.
Francis, D; Abberton, K; Thompson, E; Daniell, M
2009-03-01
Limbal stem cell deficiency leads to conjunctivalisation of the cornea and subsequent loss of vision. The recent development of transplantation of ex-vivo amplified corneal epithelium, derived from limbal stem cells, has shown promise in treating this challenging condition. The purpose of this research was to compare a variety of cell sheet carriers for their suitability in creating a confluent corneal epithelium from amplified limbal stem cells. Cadaveric donor limbal cells were cultured using an explant technique, free of 3T3 feeder cells, on a variety of cell sheet carriers, including denuded amniotic membrane, Matrigel, Myogel and stromal extract. Comparisons in rate of growth and degree of differentiation were made, using immunocytochemistry (CK3, CK19 and ABCG2). The most rapid growth was observed on Myogel and denuded amniotic membrane, these two cell carriers also provided the most reliable substrata for achieving confluence. The putative limbal stem cell marker, ABCG2, stained positively on cells grown over Myogel and Matrigel but not for those propagated on denuded amniotic membrane. In the clinical setting amniotic membrane has been demonstrated to provide a suitable carrier for limbal stem cells and the resultant epithelium has been shown to be successful in treating limbal stem cell deficiency. Myogel may provide an alternative cell carrier with a further reduction in risk as it is has the potential to be derived from an autologous muscle biopsy in the clinical setting.
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Effect of Lead on Human Middle Ear Epithelial Cells
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Shin Hye Kim
2018-01-01
Full Text Available Lead is a ubiquitous metal in the environment, but no studies have examined lead toxicity on the middle ear. Here, we investigated lead toxicity and its mechanism in human middle ear epithelial cells (HMEECs. Moreover, we investigated the protective effects of amniotic membrane extract (AME and chorionic membrane extract (CME against lead toxicity in HMEECs. Cell viability was analyzed using the cell counting kit, and reactive oxygen species (ROS activity was measured using a cellular ROS detection kit. After lead(II acetate trihydrate treatment, mRNA levels of various genes were assessed by semiquantitative real-time polymerase chain reaction. Following treatment with AME or CME after lead exposure, the changes in cell viability, ROS activity, and gene expression were analyzed. Exposure to >100 μg/mL of lead(II acetate trihydrate caused a significant decrease in cell viability and increased ROS production in HMEECs. Lead exposure significantly increased the mRNA expression of genes encoding inflammatory cytokines and mucins. Administration of AME or CME restored cell viability, reduced ROS activity, and ameliorated mRNA levels. Our findings suggest that environmental lead exposure is related to the development of otitis media, and AME and CME may have antioxidative and anti-inflammatory effects against lead toxicity.
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Wu, Huei-Wen; Lin, Xi-Zhang; Hwang, Shiaw-Min; Lee, Gwo-Bin
2009-12-01
Human mesenchymal stem cells can differentiate into multiple lineages for cell therapy and, therefore, have attracted considerable research interest recently. This study presents a new microfluidic device for bead and cell separation utilizing a combination of T-junction focusing and tilted louver-like structures. For the first time, a microfluidic device is used for continuous separation of amniotic stem cells from amniotic fluids. An experimental separation efficiency as high as 82.8% for amniotic fluid mesenchymal stem cells is achieved. Furthermore, a two-step separation process is performed to improve the separation efficiency to 97.1%. These results are based on characterization experiments that show that this microfluidic chip is capable of separating beads with diameters of 5, 10, 20, and 40 microm by adjusting the volume-flow-rate ratio between the flows in the main and side channels of the T-junction focusing structure. An optimal volume-flow-rate ratio of 0.5 can lead to high separation efficiencies of 87.8% and 85.7% for 5-microm and 10-microm beads, respectively, in a one-step separation process. The development of this microfluidic chip may be promising for future research into stem cells and for cell therapy.
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Amniotic Membrane Transplantation
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Alireza Baradaran-Rafii
2008-12-01
Full Text Available
The past decade has witnessed the revival of amniotic membrane transplantation (AMT in ophthalmology. The importance of amniotic membrane lies in its ability to reduce inflammation and scarring, enhance epithelialization and wound healing, and in its antimicrobial properties. Amniotic membrane has recently been used as a substrate for culturing limbal stem cells for transplantation. It has also been used extensively in corneal conditions such as neurotrophic ulcers, persistent epithelial defects, shield ulcers, microbial keratitis, band keratopathy, bullous keratopathy, and following photorefractive keratectomy and chemical injuries. Other indications for AMT include ocular surface reconstruction surgery for conjunctival pathologies such as squamous neoplasia, pterygium, and symblepharon. In this review we describe the basic structure and properties of amniotic membrane, its preparation process and its applications in ophthalmology.
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Lal, Ikeda; Panchal, Bhavik Uttam; Basu, Sayan; Sangwan, Virender S
2013-01-01
A 20-year-old man from Bangladesh suffered accidental alkali injury to his right eye in May 2010 leading to total limbal stem cell deficiency. An amniotic membrane graft was performed 5 days after the accident and the patient presented to our institute 6 months later. On ocular examination, his best corrected visual acuity (BCVA) was 20/50 with a 360° pannus at the periphery and central area was spared but had stromal scarring. He underwent simple limbal epithelial transplantation (SLET) taking a limbal biopsy from his left eye and was prescribed steroid and antibiotic eye drops postoperatively as per the standard regimen. At 2 year follow-up, the patient's ocular surface is stable with improvement in BCVA to 20/25 post-SLET. PMID:23704435
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Gan, Lu; Duan, Hua; Xu, Qian; Tang, Yi-Qun; Li, Jin-Jiao; Sun, Fu-Qing; Wang, Sha
2017-05-01
Intrauterine adhesion (IUA) is a common uterine cavity disease characterized by the unsatisfactory regeneration of damaged endometria. Recently, stem cell transplantation has been proposed to promote the recovery process. Here we investigated whether human amniotic mesenchymal stromal cells (hAMSCs), a valuable resource for transplantation therapy, could improve endometrial regeneration in rodent IUA models. Forty female Sprague-Dawley rats were randomly assigned to five groups: normal, sham-operated, mechanical injury, hAMSC transplantation, and negative control group. One week after intervention and transplantation, histological analyses were performed, and immunofluorescent and immunohistochemical expression of cell-specific markers and messenger RNA expression of cytokines were measured. Thicker endometria, increased gland numbers and fewer fibrotic areas were found in the hAMSC transplantation group compared with the mechanical injury group. Engraftment of hAMSCs was detected by the presence of anti-human nuclear antigen-positive cells in the endometrial glands of the transplantation uteri. Transplantation of hAMSCs significantly decreased messenger RNA levels of pro-inflammatory cytokines (tumor necrosis factor-α and interleukin-1β), and increased those of anti-inflammatory cytokines (basic fibroblast growth factor, and interleukin-6) compared with the injured uterine horns. Immunohistochemical expression of endometrial epithelial cells was revealed in specimens after hAMSC transplantation, whereas it was absent in the mechanically injured uteri. hAMSC transplantation promotes endometrial regeneration after injury in IUA rat models, possibly due to immunomodulatory properties. These cells provide a more easily accessible source of stem cells for future research into the impact of cell transplantation on damaged endometria. Copyright © 2017 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.
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Directory of Open Access Journals (Sweden)
Vidane AS
2014-08-01
Full Text Available Atanasio S Vidane,1 Aline F Souza,1 Rafael V Sampaio,1 Fabiana F Bressan,2 Naira C Pieri,1 Daniele S Martins,2 Flavio V Meirelles,2 Maria A Miglino,1 Carlos E Ambrósio2 1Department of Surgery, Faculty of Veterinary Medicine and Animal Science, University of São Paulo, São Paulo, Brazil; 2Department of Veterinary Medicine, Faculty of Animal Sciences and Food Engineering, University of São Paulo, São Paulo, Brazil Abstract: Amnion-derived mesenchymal stem cells (AMSCs are multipotent cells with an enhanced ability to differentiate into multiple lineages. AMSCs can be acquired through noninvasive methods, and therefore are exempt from the typical ethical issues surrounding stem cell use. The objective of this study was to isolate and characterize AMSCs from a cat amniotic membrane for future application in regenerative medicine. The cat AMSCs were harvested after mechanical and enzymatic digestion of amnion. In culture medium, the cat AMSCs adhered to a plastic culture dish and displayed a fibroblast-like morphology. Immunophenotyping assays were positive for the mesenchymal stem cell-specific markers CD73 and CD90 but not the hematopoietic markers CD34, CD45, and CD79. Under appropriate conditions, the cat AMSCs differentiated into osteogenic, chondrogenic, and adipogenic cell lineages. One advantage of cat AMSCs was nonteratogenicity, assessed 4 weeks post injection of undifferentiated AMSCs into immunodeficient mice. These findings suggest that cat amniotic membranes may be an important and useful source of mesenchymal stem cells for clinical applications, especially for cell or tissue replacement in chronic and degenerative diseases. Keywords: amnion, cats, cell differentiation, fetal membranes, mesenchymal cells
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Directory of Open Access Journals (Sweden)
Araki Hiromasa
2007-04-01
Full Text Available Abstract Background Proteinase-activated receptors (PARs; PAR1–4 that can be activated by serine proteinases such as thrombin and neutrophil catepsin G are known to contribute to the pathogenesis of various pulmonary diseases including fibrosis. Among these PARs, especially PAR4, a newly identified subtype, is highly expressed in the lung. Here, we examined whether PAR4 stimulation plays a role in the formation of fibrotic response in the lung, through alveolar epithelial-mesenchymal transition (EMT which contributes to the increase in myofibroblast population. Methods EMT was assessed by measuring the changes in each specific cell markers, E-cadherin for epithelial cell, α-smooth muscle actin (α-SMA for myofibroblast, using primary cultured mouse alveolar epithelial cells and human lung carcinoma-derived alveolar epithelial cell line (A549 cells. Results Stimulation of PAR with thrombin (1 U/ml or a synthetic PAR4 agonist peptide (AYPGKF-NH2, 100 μM for 72 h induced morphological changes from cobblestone-like structure to elongated shape in primary cultured alveolar epithelial cells and A549 cells. In immunocytochemical analyses of these cells, such PAR4 stimulation decreased E-cadherin-like immunoreactivity and increased α-SMA-like immunoreactivity, as observed with a typical EMT-inducer, tumor growth factor-β (TGF-β. Western blot analyses of PAR4-stimulated A549 cells also showed similar changes in expression of these EMT-related marker proteins. Such PAR4-mediated changes were attenuated by inhibitors of epidermal growth factor receptor (EGFR kinase and Src. PAR4-mediated morphological changes in primary cultured alveolar epithelial cells were reduced in the presence of these inhibitors. PAR4 stimulation increased tyrosine phosphorylated EGFR or tyrosine phosphorylated Src level in A549 cells, and the former response being inhibited by Src inhibitor. Conclusion PAR4 stimulation of alveolar epithelial cells induced epithelial
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2017-09-01
repair in the upper extremity using processed nerve allograft. J Hand Surg Am 2012 Nov;37(11):2340-9. (9) Joo S, Ko IK, Atala A, Yoo JJ , Lee SJ. Amniotic...nerve grafts implanted with autologous mesenchymal stem cells.Exp Neurol. 2007 Apr;204(2):658-66. (18) Kim BS, Chun SY, Atala A, Soker S, Yoo JJ , Kwon TG...wounds. Stem Cells Transl Med. 2012 ;1(11):792-802 4. Joo S, Ko IK, Atala A, Yoo JJ , Lee SJ. Amniotic fluid-derived stem cells in regenerative
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Human Mammary Luminal Epithelial Cells Contain Progenitors to Myoepithelial Cells
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Pechoux, Christine; Gudjonsson, Thorarinn; Ronnov-Jessen, Lone; Bissell, Mina J; Petersen, Ole
1999-02-01
The origin of the epithelial and myoepithelial cells in the human breast has not been delineated. In this study we have addressed whether luminal epithelial cells and myoepithelial cells are vertically connected, i.e., whether one is the precursor for the other. We used a primary culture assay allowing preservation of basic phenotypic traits of luminal epithelial and myoepithelial cells in culture. The two cell types were then separated immunomagnetically using antibodies directed against lineage-specific cell surface antigens into at best 100% purity. The cellular identity was ascertained by cytochemistry, immunoblotting, and 2-D gel electrophoresis. Luminal epithelial cells were identified by strong expression of cytokeratins 18 and 19 while myoepithelial cells were recognized by expression of vimentin and {alpha}-smooth muscle actin. We used a previously devised culture medium (CDM4) that allows vigorous expansion of proliferative myoepithelial cells and also devised a medium (CDM6) that allowed sufficient expansion of differentiated luminal epithelial cells based on addition of hepatocyte growth factor/scatter factor. The two different culture media supported each lineage for at least five passages without signs of interconversion. We used parallel cultures where we switched culture media, thus testing the ability of each lineage to convert to the other. Whereas the myoepithelial lineage showed no signs of interconversion, a subset of luminal epithelial cells, gradually, but distinctly, converted to myoepithelial cells. We propose that in the mature human breast, it is the luminal epithelial cell compartment that gives rise to myoepithelial cells rather than the other way around.
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Man, Rohaina Che; Yong, Then Kong; Hwei, Ng Min; Halim, Wan Haslina Wan Abdul; Zahidin, Aida Zairani Mohd; Ramli, Roszalina; Saim, Aminuddin Bin; Idrus, Ruszymah Binti Hj
2017-01-01
Various clinical disorders and injuries, such as chemical, thermal, or mechanical injuries, may lead to corneal loss that results in blindness. PURPOSE : The aims of this study were to differentiate human buccal mucosa (BMuc) into corneal epithelial-like cells, to fabricate engineered corneal tissue using buccal mucosal epithelial cells, and to reconstruct a damaged corneal epithelium in a nude rat model. BMuc were subjected to 10 d of induction factors to investigate the potential of cells to differentiate into corneal lineages. Corneal stem cell markers β1-integrin, C/EBPδ, ABCG2, p63, and CK3 were upregulated in the gene expression analysis in induced BMuc, whereas CK3 and p63 showed significant protein expression in induced BMuc compared to the uninduced cells. BMuc were then left to reach 80% confluency after differential trypsinization. The cells were harvested and cultivated on a commercially available untreated air-dried amniotic membrane (AM) in a Transwell system in induction medium. The corneal constructs were fabricated and then implanted into damaged rat corneas for up to 8 weeks. A significant improvement was detected in the treatment group at 8 weeks post-implantation, as revealed by slit lamp biomicroscopy analysis. The structure and thickness of the corneal layer were also analyzed using histological staining and time-domain optical coherence tomography scans and were found to resemble a native corneal layer. The protein expression for CK3 and p63 were continuously detected throughout the corneal epithelial layer in the corneal construct. In conclusion, human BMuc can be induced to express a corneal epithelial-like phenotype. The addition of BMuc improves corneal clarity, prevents vascularization, increases corneal thickness and stromal alignment, and appears to have no adverse effect on the host after implantation.
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Epithelial cell-cell junctions and plasma membrane domains
Giepmans, Ben N. G.; van Ijzendoorn, Sven C. D.
Epithelial cells form a barrier against the environment, but are also required for the regulated exchange of molecules between an organism and its surroundings. Epithelial cells are characterised by a remarkable polarization of their plasma membrane, evidenced by the appearance of structurally,
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Engraftment of mouse amniotic fluid-derived progenitor cells after in utero transplantation in mice.
Lin, Kun-Yi; Peng, Shao-Yu; Chou, Chih-Jen; Wu, Chia-Chun; Wu, Shinn-Chih
2015-11-01
Amniotic fluid-derived progenitor cells (AFPCs) are oligopotent and shed from the fetus into the amniotic fluid. It was reported that AFPCs express stem cell-like markers and are capable of differentiating into specific cell type in in vitro experiments. However, no study has fully investigated the potentiality and destiny of these cells in in vivo experiments. Ds-red transgenic mice (on Day 13.5 of pregnancy) were transplanted in utero with enhanced green fluorescent protein-labeled mouse AFPC (EGFP-mAFPCs). After birth, baby mice were euthanized at 3-week intervals beginning 3 weeks postnatally, and the specimens were examined by polymerase chain reaction, histology, and flow cytometry. Our results demonstrate the transplantability of mAFPCs into all three germ layers and the potential of mAFPCs in the study of progenitor cell homing, differentiation, and function. Engraftment of EGFP-mAFPCs was detected in the intestine, kidney, muscle, skin, bladder, heart, stomach, etc., at 3 weeks after delivery. This model using EGFP-mAFPCs injected in utero may provide an ideal method for determining the fate of transplanted cells in recipients and these findings may justify a clinical trial of in utero transplantation during gestation for patients who have inherited genetic disorders. Copyright © 2014. Published by Elsevier B.V.
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Clinical analysis of amniotic membrane patches and grafts for acute ocular surface burn
Directory of Open Access Journals (Sweden)
Lin Li
2015-01-01
Full Text Available AIM: To investigate the effect and value of amniotic membrane patches and grafts for acute ocular surface burn at different degrees.METHODS: A retrospective analysis of 28 cases(28 eyesaffected by ocular chemical or thermal burn with different degree were included in our hospital from March 2007 to March 2012. Amniotic membrane patched was undergone in 13 eyes with fresh amnion that the patients corneal burns degree Ⅱ or Ⅲ with partial limbal buns at degree Ⅳ. Amniotic membrane grafts was performed in 15 eyes with fresh amnion that the patients all corneal burns at degree Ⅲ with the whole limbal necrosis without severe eyelid defect. The follow-up time ranged 6～24mo. The postoperative visual acuity, the condition of amniotic membrane transplant, renovation of cornea and complications were observed. RESULTS: Postoperative corrected visual acuity was improved in 20 eyes(71%, it was not changed in 5 eyes(18%, the visual acuity declined in 3 eyes(11%. The amniotic membrane survived in 23 eyes and the survival rate was up to 82%. The cornea of 4 eyes recovered to transparent, nebula emceed in 8 eyes eventually, corneal macula emerged in 10 eyes, 4 eyes ended up with leukoma, 2 eyes developed corneal melting after therapy, then received lamellar keratoplasty. Corneal surface become epithelization after amnion patches or grafts, but any of them have recurrent epithelial erosion, and become stable epithalization after repeat operation.CONCLUSION: Amniotic membrane patches and grafts is an effective method to deal with acute ocular surface burn.
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Directory of Open Access Journals (Sweden)
Rifat Rasier
2014-12-01
Full Text Available AIM: To avoid the side effects of the suture usage by welding amniotic membrane (AM to contact lens (CL with laser.METHODS:AM was taken from pregnant women and cleaned from blood clots with sterile phosphate-buffered physiological saline solution which included antibiotics. Stromal side of the AM was spread inside of the CL and it was welded to CL by 1470 nm diode laser. 600 μm diameter fiber tip of the laser was contacted with the epithelial side of the AM from 4 separate points. After welding excess amniotic membrane around the CL was cut with a scalpel.RESULTS:Stromal side of the AM was spread inside of the CL and then with laser fiber, different power levels and exposure times were applied on the epithelium of AM and 340 mW for seven seconds was found optimal. CL and AM attached with the spot welding effect in 4 points by touching fiber tip. CL-AM welded complex did not separated from each other while holding AM that extend beyond the CL with the help of two forceps.CONCLUSION:As a conclusion, it was aimed in this study to achieve the success of the conventional amniotic membrane transplantation (AMTwith the easiness of applying a CL and to avoid risks and side effects of corneal or conjunctival suturing. The results showed that the application of the CL–AM complex will be as easy as the application of a CL and lasts shortly.
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Directory of Open Access Journals (Sweden)
Takashi Kawaguchi
1999-01-01
Full Text Available This study was designed to examine the effects of adrenomedullin (AM on airway epithelial cells. Primary cultures of guinea-pig tracheal epithelial cells and the human bronchiolar epithelial cell line NCI-H441 were used. Intracellular cyclic adenosine monophosphate (cAMP, cyclic guanosine monophosphate (cGMP, prostaglandin E2 (PGE2, and stable end-products of nitric oxide were assayed. Adrenomedullin (10−6 mol/L stimulated cAMP production in guinea-pig epithelial cells. Indomethacin (10−5 mol/L significantly decreased the basal level of intracellular cAMP in guinea-pig epithelial cells, but not in NCI-H441 cells. However, AM did not stimulate production of PGE2, a major product that can increase cAMP formation. In the case of NCI-H441 cells, AM (10−8 – 10−6 mol/L did not significantly affect intracellular cGMP levels or nitrite content in conditioned medium. Adrenomedullin and calcitonin gene-related peptide (CGRP each stimulated cAMP production in NCI-H441 cells, but AM-stimulated cAMP production was antagonized by the CGRP fragment CGRP8–37. These findings suggest that AM stimulates cAMP production and functionally competes with CGRP for binding sites in airway epithelial cells, at least in human epithelial cells, but that it does not stimulate the release of PGE2 and nitric oxide. Though cyclooxygenase products contribute to some extent to cAMP formation in guinea-pigs, AM independently stimulates intracellular cAMP formation in airway epithelial cells.
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DNA repair in human bronchial epithelial cells
International Nuclear Information System (INIS)
Fornace, A.J. Jr.; Lechner, J.F.; Grafstrom, R.C.; Harris, C.C.
1982-01-01
The purpose of this investigation was to compare the response of human cell types (bronchial epithelial cells and fibroblasts and skin fibroblasts) to various DNA damaging agents. Repair of DNA single strand breaks (SSB) induced by 5 krads of X-ray was similar for all cell types; approximately 90% of the DNA SSB were rejoined within one hour. During excision repair of DNA damage from u.v.-radiation, the frequencies of DNA SSB as estimated by the alkaline elution technique, were similar in all cell types. Repair replication as measured by BND cellulose chromatography was also similar in epithelial and fibroblastic cells after u.v.-irradiation. Similar levels of SSB were also observed in epithelial and fibroblastic cells after exposure to chemical carcinogens: 7,12-dimethylbenz[a]anthracene; benzo[a]pyrene diol epoxide (BPDE); or N-methyl-N-nitro-N-nitrosoguanidine. Significant repair replication of BPDE-induced DNA damage was detected in both bronchial epithelial and fibroblastic cells, although the level in fibroblasts was approximately 40% of that in epithelial cells. The pulmonary carcinogen asbestos did not damage DNA. DNA-protein crosslinks induced by formaldehyde were rapidly removed in bronchial cells. Further, epithelial and fibroblastic cells, which were incubated with formaldehyde and the polymerase inhibitor combination of cytosine arabinoside and hydroxyurea, accumulated DNA SSB at approximately equal frequencies. These results should provide a useful background for further investigations of the response of human bronchial cells to various DNA damaging agents
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Energy Technology Data Exchange (ETDEWEB)
Nelson, Celeste M.; Khauv, Davitte; Bissell, Mina J.; Radisky, Derek C.
2008-06-26
Cell morphology dictates response to a wide variety of stimuli, controlling cell metabolism, differentiation, proliferation, and death. Epithelial-mesenchymal transition (EMT) is a developmental process in which epithelial cells acquire migratory characteristics, and in the process convert from a 'cuboidal' epithelial structure into an elongated mesenchymal shape. We had shown previously that matrix metalloproteinase-3 (MMP3) can stimulate EMT of cultured mouse mammary epithelial cells through a process that involves increased expression of Rac1b, a protein that stimulates alterations in cytoskeletal structure. We show here that cells treated with MMP-3 or induced to express Rac1b spread to cover a larger surface, and that this induction of cell spreading is a requirement of MMP-3/Rac1b-induced EMT. We find that limiting cell spreading, either by increasing cell density or by culturing cells on precisely defined micropatterned substrata, blocks expression of characteristic markers of EMT in cells treated with MMP-3. These effects are not caused by general disruptions in cell signaling pathways, as TGF-{beta}-induced EMT is not affected by similar limitations on cell spreading. Our data reveal a previously unanticipated cell shape-dependent mechanism that controls this key phenotypic alteration and provide insight into the distinct mechanisms activated by different EMT-inducing agents.
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Qi, Fangfang; Yoshida, Toshiko; Koike, Takeshi; Aizawa, Hitoshi; Shimane, Tetsu; Li, Yinghui; Yamada, Shinichi; Okabe, Motonori; Nikaido, Toshio; Kurita, Hiroshi
2016-05-01
Human amniotic membrane(HAM) as a graft material has been used in various fields. Hyper-dry amniotic membrane (HD-AM) is a novel dried amniotic membrane that is easy to handle and can be preserved at room temperature without time limitation. The purpose of this study was to investigate the useful properties of HD-AM in reconstruction of the oral mucosa. Human oral keratinocytes were isolated and seeded on HD-AM in serum-free culture system. Oral mucosa equivalent (OME) was developed and transplanted onto full-thickness wound on athymic mice. The wound healing was analyzed and the OME both before and after transplantation was analyzed with hematoxylin-eosin staining and immunohistochemical staining for Cytokines 10 (CK10), Cytokines 16 (CK16), and Ivolucrin (IVL). Oral keratinocytes spread and proliferated well on HD-AM. Two weeks after air-lifting, OME had formed with good differentiation and morphology. We confirmed immunohistochemically that the expression of CK10 was positive in all suprabasal layers, as was CK16 in the upper layers, while IVL was present in all cell layers. Three weeks after transplantation to athymic mice, the newly generated tissue had survived well with the smallest contraction. The epithelial cells of newly generated tissue expressed CK10 throughout in all suprabasal layers, IVL was mainly in the granular layer, and CK16 positive cells were observed in all spinous layer and granular layer but were not expressed in the mouse skin, all of which were similar to native gingival mucosa. The OME with HD-AM as a matrix revealed a good morphology and stable wound healing. This study demonstrates that HD-AM is a useful and feasible biomaterial for oral mucosa reconstruction. Copyright © 2016 Elsevier Ltd. All rights reserved.
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Brown, Karl David; Low, Suet; Mariappan, Indumathi; Abberton, Keren Maree; Short, Robert; Zhang, Hong; Maddileti, Savitri; Sangwan, Virender; Steele, David; Daniell, Mark
2014-02-01
Extensive damage to the limbal region of the cornea leads to a severe form of corneal blindness termed as limbal stem cell deficiency (LSCD). Whereas most cases of corneal opacity can be treated with full thickness corneal transplants, LSCD requires stem cell transplantation for successful ocular surface reconstruction. Current treatments for LSCD using limbal stem cell transplantation involve the use of murine NIH 3T3 cells and human amniotic membranes as culture substrates, which pose the threat of transmission of animal-derived pathogens and donor tissue-derived cryptic infections. In this study, we aimed to produce surface modified therapeutic contact lenses for the culture and delivery of corneal epithelial cells for the treatment of LSCD. This approach avoids the possibility of suture-related complications and is completely synthetic. We used plasma polymerization to deposit acid functional groups onto the lenses at various concentrations. Each surface was tested for its suitability to promote corneal epithelial cell adhesion, proliferation, retention of stem cells, and differentiation and found that acid-based chemistries promoted better cell adhesion and proliferation. We also found that the lenses coated with a higher percentage of acid functional groups resulted in a higher number of cells transferred onto the corneal wound bed in rabbit models of LSCD. Immunohistochemistry of the recipient cornea confirmed the presence of autologous, transplanted 5-bromo-2'-deoxyuridine (BrdU)-labeled cells. Hematoxylin staining has also revealed the presence of a stratified epithelium at 26 days post-transplantation. This study provides the first evidence for in vivo transfer and survival of cells transplanted from a contact lens to the wounded corneal surface. It also proposes the possibility of using plasma polymer-coated contact lenses with high acid functional groups as substrates for the culture and transfer of limbal cells in the treatment of LSCD.
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Early fish myoseptal cells: insights from the trout and relationships with amniote axial tenocytes.
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Yoann Bricard
Full Text Available The trunk muscle in fish is organized as longitudinal series of myomeres which are separated by sheets of connective tissue called myoseptum to which myofibers attach. In this study we show in the trout that the myoseptum separating two somites is initially acellular and composed of matricial components such as fibronectin, laminin and collagen I. However, myoseptal cells forming a continuum with skeletogenic cells surrounding axial structures are observed between adjacent myotomes after the completion of somitogenesis. The myoseptal cells do not express myogenic markers such as Pax3, Pax7 and myogenin but express several tendon-associated collagens including col1a1, col5a2 and col12a1 and angiopoietin-like 7, which is a secreted molecule involved in matrix remodelling. Using col1a1 as a marker gene, we observed in developing trout embryo an initial labelling in disseminating cells ventral to the myotome. Later, labelled cells were found more dorsally encircling the notochord or invading the intermyotomal space. This opens the possibility that the sclerotome gives rise not only to skeletogenic mesenchymal cells, as previously reported, but also to myoseptal cells. We furthermore show that myoseptal cells differ from skeletogenic cells found around the notochord by the specific expression of Scleraxis, a distinctive marker of tendon cells in amniotes. In conclusion, the location, the molecular signature and the possible sclerotomal origin of the myoseptal cells suggest that the fish myoseptal cells are homologous to the axial tenocytes in amniotes.
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Endothelial Induced EMT in Breast Epithelial Cells with Stem Cell Properties
Sigurdsson, Valgardur; Hilmarsdottir, Bylgja; Sigmundsdottir, Hekla; Fridriksdottir, Agla J. R.; Ringnér, Markus; Villadsen, Rene; Borg, Ake; Agnarsson, Bjarni A.; Petersen, Ole William; Magnusson, Magnus K.; Gudjonsson, Thorarinn
2011-01-01
Epithelial to mesenchymal transition (EMT) is a critical event in cancer progression and is closely linked to the breast epithelial cancer stem cell phenotype. Given the close interaction between the vascular endothelium and cancer cells, especially at the invasive front, we asked whether endothelial cells might play a role in EMT. Using a 3D culture model we demonstrate that endothelial cells are potent inducers of EMT in D492 an immortalized breast epithelial cell line with stem cell proper...
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Wei, Zhengxi; Shan, Zhongguo; Shaikh, Zahir A
2018-04-01
Epidemiological and experimental studies have implicated cadmium (Cd) with breast cancer. In breast epithelial MCF10A and MDA-MB-231 cells, Cd has been shown to promote cell growth. The present study examined whether Cd also promotes epithelial-mesenchymal transition (EMT), a hallmark of cancer progression. Human breast epithelial cells consisting of non-cancerous MCF10A, non-metastatic HCC 1937 and HCC 38, and metastatic MDA-MB-231 were treated with 1 or 3 μM Cd for 4 weeks. The MCF10A epithelial cells switched to a more mesenchymal-like morphology, which was accompanied by a decrease in the epithelial marker E-cadherin and an increase in the mesenchymal markers N-cadherin and vimentin. In both non-metastatic HCC 1937 and HCC 38 cells, treatment with Cd decreased the epithelial marker claudin-1. In addition, E-cadherin also decreased in the HCC 1937 cells. Even the mesenchymal-like MDA-MB-231 cells exhibited an increase in the mesenchymal marker vimentin. These changes indicated that prolonged treatment with Cd resulted in EMT in both normal and cancer-derived breast epithelial cells. Furthermore, both the MCF10A and MDA-MB-231 cells labeled with Zcad, a dual sensor for tracking EMT, demonstrated a decrease in the epithelial marker E-cadherin and an increase in the mesenchymal marker ZEB-1. Treatment of cells with Cd significantly increased the level of Snail, a transcription factor involved in the regulation of EMT. However, the Cd-induced Snail expression was completely abolished by actinomycin D. Luciferase reporter assay indicated that the expression of Snail was regulated by Cd at the promotor level. Snail was essential for Cd-induced promotion of EMT in the MDA-MB-231 cells, as knockdown of Snail expression blocked Cd-induced cell migration. Together, these results indicate that Cd promotes EMT in breast epithelial cells and does so by modulating the transcription of Snail. Copyright © 2018 Elsevier Inc. All rights reserved.
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Energy Technology Data Exchange (ETDEWEB)
Parrinello, Simona; Coppe, Jean-Philippe; Krtolica, Ana; Campisi, Judith
2004-07-14
Cellular senescence suppresses cancer by arresting cells at risk for malignant tumorigenesis. However, senescent cells also secrete molecules that can stimulate premalignant cells to proliferate and form tumors, suggesting the senescence response is antagonistically pleiotropic. We show that premalignant mammary epithelial cells exposed to senescent human fibroblasts in mice irreversibly lose differentiated properties, become invasive and undergo full malignant transformation. Moreover, using cultured mouse or human fibroblasts and non-malignant breast epithelial cells, we show that senescent fibroblasts disrupt epithelial alveolar morphogenesis, functional differentiation, and branching morphogenesis. Further, we identify MMP-3 as the major factor responsible for the effects of senescent fibroblasts on branching morphogenesis. Our findings support the idea that senescent cells contribute to age-related pathology, including cancer, and describe a new property of senescent fibroblasts--the ability to alter epithelial differentiation--that might also explain the loss of tissue function and organization that is a hallmark of aging.
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Rat glomerular epithelial cells in culture. Parietal or visceral epithelial origin
International Nuclear Information System (INIS)
Norgaard, J.O.
1987-01-01
Isolated glomeruli from rats were explanted under standard culture conditions and outgrowths were studied by light and electron microscopy in order to identify the cells. Rat glomerular samples contained 20 to 30% structurally well-preserved encapsulated glomeruli which had a large rate of attachment to the substrate and very constantly gave rise to cellular outgrowth. In order to label cells from which outgrowth originated the glomerular incorporation of [ 3 H]thymidine was studied in the preattachment phase. By light and electron microscope autoradiograph it was demonstrated that label was located only over visceral and parietal epithelial cells during the first 3 days of culture. Incorporation of [ 3 H]thymidine was seen in mesangial cells after 5 days, i.e., after the glomeruli had attached to the culture vessels and the initial outgrowth had appeared. Consequently the first cells to grow out were of epithelial origin. Glomeruli were then incubated with [ 3 H]thymidine for the first 2 1/2 days of culture in order to label the epithelial cells, then were allowed to attach to the substrate and induce cell outgrowth. By light microscope autoradiography performed with the outgrowths in situ two types of cells with labeled nuclei were seen: (a) a small, polyhedral ciliated cell which grew in colonies where the cells were joined by junctional complexes (type I), and (b) a second very large, often multinucleated cell (type II). Based on the structural resemblance with their counterparts in situ and on comparisons with positively identified visceral epithelial cells in outgrowths from other species it is suggested that type I cells are derived from the parietal epithelium of Bowman's capsule and type II cells from the visceral epithelium
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Kim, Sun Min; Romero, Roberto; Lee, JoonHo; Chaemsaithong, Piya; Docheva, Nikolina; Yoon, Bo Hyun
2016-01-01
Early neonatal sepsis is often due to intra-amniotic infection. The stomach of the neonate contains fluid swallowed before and during delivery. The presence of bacteria as well as neutrophils detected by culture or Gram stain of the gastric fluid during the first day of life is suggestive of exposure to bacteria or inflammation. We undertook this study to determine the relationship between gastric fluid analysis and amniotic fluid obtained by transabdominal amniocentesis in the detection of Ureaplasma species, the most frequent microorganisms responsible for intra-amniotic infection. The study population consisted of 100 singleton pregnant women who delivered preterm neonates (Ureaplasma species was performed. Intra-amniotic inflammation was defined as an elevated amniotic fluid matrix metalloproteinase-8 concentration (>23 ng/mL). (1) Ureaplasma species were detected by culture or PCR in 18% (18/100) of amniotic fluid samples and in 5% (5/100) of gastric fluid samples; (2) among the amniotic fluid cases positive for Ureaplasma species, these microorganisms were identified in 27.8% (5/18) of gastric fluid samples; (3) none of the cases negative for Ureaplasma species in the amniotic fluid were found to be positive for these microorganisms in the gastric fluid; (4) patients with amniotic fluid positive for Ureaplasma species but with gastric fluid negative for these microorganisms had a significantly higher rate of intra-amniotic inflammation, acute histologic chorioamnionitis, and neonatal death than those with both amniotic fluid and gastric fluid negative for Ureaplasma species; and (5) no significant differences were observed in the rate of intra-amniotic inflammation, acute histologic chorioamnionitis, and neonatal death between patients with amniotic fluid positive for Ureaplasma species but with gastric fluid negative for these microorganisms and those with both amniotic fluid and gastric fluid positive for Ureaplasma species. Gastric fluid analysis has 100
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SOX9 as a Predictor for Neurogenesis Potentiality of Amniotic Fluid Stem Cells
Wei, Pei-Cih; Chao, Angel; Peng, Hsiu-Huei; Chao, An-Shine; Chang, Yao-Lung; Chang, Shuenn-Dyh; Wang, Hsin-Shih; Chang, Yu-Jen; Tsai, Ming-Song; Sieber, Martin; Chen, Hua-Chien; Chen, Shu-Jen; Lee, Yun-Shien
2014-01-01
Preclinical studies of amniotic fluid-derived cell therapy have been successful in the research of neurodegenerative diseases, peripheral nerve injury, spinal cord injury, and brain ischemia. Transplantation of human amniotic fluid stem cells (AFSCs) into rat brain ventricles has shown improvement in symptoms of Parkinson's disease and also highlighted the minimal immune rejection risk of AFSCs, even between species. Although AFSCs appeared to be a promising resource for cell-based regenerative therapy, AFSCs contain a heterogeneous pool of distinct cell types, rendering each preparation of AFSCs unique. Identification of predictive markers for neuron-prone AFSCs is necessary before such stem cell-based therapeutics can become a reality. In an attempt to identify markers of AFSCs to predict their ability for neurogenesis, we performed a two-phase study. In the discovery phase of 23 AFSCs, we tested ZNF521/Zfp521, OCT6, SOX1, SOX2, SOX3, and SOX9 as predictive markers of AFSCs for neural differentiation. In the validation phase, the efficacy of these predictive markers was tested in independent sets of 18 AFSCs and 14 dental pulp stem cells (DPSCs). We found that high expression of SOX9 in AFSCs is associated with good neurogenetic ability, and these positive correlations were confirmed in independent sets of AFSCs and DPSCs. Furthermore, knockdown of SOX9 in AFSCs inhibited their neuronal differentiation. In conclusion, the discovery of SOX9 as a predictive marker for neuron-prone AFSCs could expedite the selection of useful clones for regenerative medicine, in particular, in neurological diseases and injuries. PMID:25154783
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Directory of Open Access Journals (Sweden)
Satoru Yoshida
Full Text Available BACKGROUND: Application of induced pluripotent stem (iPS cells in regenerative medicine will bypass ethical issues associated with use of embryonic stem cells. In addition, patient-specific IPS cells can be useful to elucidate the pathophysiology of genetic disorders, drug screening, and tailor-made medicine. However, in order to apply iPS cells to mitotic tissue, induction of tissue stem cells that give rise to progeny of the target organ is required. METHODOLOGY/PRINCIPAL FINDINGS: We induced stratified epithelial cells from mouse iPS cells by co-culture with PA6 feeder cells (SDIA-method with use of BMP4. Clusters of cells positive for the differentiation markers KRT1 or KRT12 were observed in KRT14-positive colonies. We successfully cloned KRT14 and p63 double-positive stratified epithelial progenitor cells from iPS-derived epithelial cells, which formed stratified epithelial sheets consisting of five- to six-polarized epithelial cells in vitro. When these clonal cells were cultured on denuded mouse corneas, a robust stratified epithelial layer was observed with physiological cell polarity including high levels of E-cadherin, p63 and K15 expression in the basal layer and ZO-1 in the superficial layer, recapitulating the apico-basal polarity of the epithelium in vivo. CONCLUSIONS/SIGNIFICANCE: These results suggest that KRT14 and p63 double-positive epithelial progenitor cells can be cloned from iPS cells in order to produce polarized multilayer epithelial cell sheets.
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Epithelial Cells in Urine: MedlinePlus Lab Test Information
... page: https://medlineplus.gov/labtests/epithelialcellsinurine.html Epithelial Cells in Urine To use the sharing features on ... page, please enable JavaScript. What is an Epithelial Cells in Urine Test? Epithelial cells are a type ...
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Amniotic fluid stem cells: a promising therapeutic resource for cell-based regenerative therapy.
Antonucci, Ivana; Pantalone, Andrea; Tete, Stefano; Salini, Vincenzo; Borlongan, Cesar V; Hess, David; Stuppia, Liborio
2012-01-01
Stem cells have been proposed as a powerful tool in the treatment of several human diseases, both for their ability to represent a source of new cells to replace those lost due to tissue injuries or degenerative diseases, and for the ability of produce trophic molecules able to minimize damage and promote recovery in the injured tissue. Different cell types, such as embryonic, fetal or adult stem cells, human fetal tissues and genetically engineered cell lines, have been tested for their ability to replace damaged cells and to restore the tissue function after transplantation. Amniotic fluid -derived Stem cells (AFS) are considered a novel resource for cell transplantation therapy, due to their high renewal capacity, the "in vitro" expression of embryonic cell lineage markers, and the ability to differentiate in tissues derived from all the three embryonic layers. Moreover, AFS do not produce teratomas when transplanted into animals and are characterized by a low antigenicity, which could represent an advantage for cell transplantation or cell replacement therapy. The present review focuses on the biological features of AFS, and on their potential use in the treatment of pathological conditions such as ischemic brain injury and bone damages.
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Silk Film Topography Directs Collective Epithelial Cell Migration
Rosenblatt, Mark I.
2012-01-01
The following study provides new insight into how surface topography dictates directed collective epithelial cell sheet growth through the guidance of individual cell movement. Collective cell behavior of migrating human corneal limbal-epithelial cell sheets were studied on highly biocompatible flat and micro-patterned silk film surfaces. The silk film edge topography guided the migratory direction of individual cells making up the collective epithelial sheet, which resulted in a 75% increase in total culture elongation. This was due to a 3-fold decrease in cell sheet migration rate efficiency for movement perpendicular to the topography edge. Individual cell migration direction is preferred in the parallel approach to the edge topography where localization of cytoskeletal proteins to the topography’s edge region is reduced, which results in the directed growth of the collective epithelial sheet. Findings indicate customized biomaterial surfaces may be created to direct both the migration rate and direction of tissue epithelialization. PMID:23185573
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Basolateral BMP signaling in polarized epithelial cells.
Directory of Open Access Journals (Sweden)
Masao Saitoh
Full Text Available Bone morphogenetic proteins (BMPs regulate various biological processes, mostly mediated by cells of mesenchymal origin. However, the roles of BMPs in epithelial cells are poorly understood. Here, we demonstrate that, in polarized epithelial cells, BMP signals are transmitted from BMP receptor complexes exclusively localized at the basolateral surface of the cell membrane. In addition, basolateral stimulation with BMP increased expression of components of tight junctions and enhanced the transepithelial resistance (TER, counteracting reduction of TER by treatment with TGF-β or an anti-tumor drug. We conclude that BMPs maintain epithelial polarity via intracellular signaling from basolaterally localized BMP receptors.
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Epithelial cells as alternative human biomatrices for comet assay.
Rojas, Emilio; Lorenzo, Yolanda; Haug, Kristiane; Nicolaissen, Bjørn; Valverde, Mahara
2014-01-01
The comet assay is a valuable experimental tool aimed at mapping DNA damage in human cells in vivo for environmental and occupational monitoring, as well as for therapeutic purposes, such as storage prior to transplant, during tissue engineering, and in experimental ex vivo assays. Furthermore, due to its great versatility, the comet assay allows to explore the use of alternative cell types to assess DNA damage, such as epithelial cells. Epithelial cells, as specialized components of many organs, have the potential to serve as biomatrices that can be used to evaluate genotoxicity and may also serve as early effect biomarkers. Furthermore, 80% of solid cancers are of epithelial origin, which points to the importance of studying DNA damage in these tissues. Indeed, studies including comet assay in epithelial cells have either clear clinical applications (lens and corneal epithelial cells) or examine genotoxicity within human biomonitoring and in vitro studies. We here review improvements in determining DNA damage using the comet assay by employing lens, corneal, tear duct, buccal, and nasal epithelial cells. For some of these tissues invasive sampling procedures are needed. Desquamated epithelial cells must be obtained and dissociated prior to examination using the comet assay, and such procedures may induce varying amounts of DNA damage. Buccal epithelial cells require lysis enriched with proteinase K to obtain free nucleosomes. Over a 30 year period, the comet assay in epithelial cells has been little employed, however its use indicates that it could be an extraordinary tool not only for risk assessment, but also for diagnosis, prognosis of treatments and diseases.
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Molecular Characterization of Gastric Epithelial Cells Using Flow Cytometry
Directory of Open Access Journals (Sweden)
Kevin A. Bockerstett
2018-04-01
Full Text Available The ability to analyze individual epithelial cells in the gastric mucosa would provide important insight into gastric disease, including chronic gastritis and progression to gastric cancer. However, the successful isolation of viable gastric epithelial cells (parietal cells, neck cells, chief cells, and foveolar cells from gastric glands has been limited due to difficulties in tissue processing. Furthermore, analysis and interpretation of gastric epithelial cell flow cytometry data has been difficult due to the varying sizes and light scatter properties of the different epithelial cells, high levels of autofluorescence, and poor cell viability. These studies were designed to develop a reliable method for isolating viable single cells from the corpus of stomachs and to optimize analyses examining epithelial cells from healthy and diseased stomach tissue by flow cytometry. We performed a two stage enzymatic digestion in which collagenase released individual gastric glands from the stromal tissue of the corpus, followed by a Dispase II digestion that dispersed these glands into greater than 1 × 106 viable single cells per gastric corpus. Single cell suspensions were comprised of all major cell lineages found in the normal gastric glands. A method describing light scatter, size exclusion, doublet discrimination, viability staining, and fluorescently-conjugated antibodies and lectins was used to analyze individual epithelial cells and immune cells. This technique was capable of identifying parietal cells and revealed that gastric epithelial cells in the chronically inflamed mucosa significantly upregulated major histocompatibility complexes (MHC I and II but not CD80 or CD86, which are costimulatory molecules involved in T cell activation. These studies describe a method for isolating viable single cells and a detailed description of flow cytometric analysis of cells from healthy and diseased stomachs. These studies begin to identify effects of
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Chromosome aberration induction in human diploid fibroblast and epithelial cells
International Nuclear Information System (INIS)
Scott, D.
1986-01-01
The relative sensitivity of cultured human fibroblasts and epithelial cells to radiation-induced chromosomal aberrations was investigated. Lung fibroblast and kidney epithelial cells from the same fetus were compared, as were skin fibroblasts and epithelial keratinocytes from the same foreskin sample. After exposure of proliferating fetal cells to 1.5 Gy X-rays there was a very similar aberration yield in the fibroblasts and epithelial cells. Observations of either little or no difference in chromosomal sensitivity between human fibroblasts and epithelial cells give added confidence that quantitative cytogenetic data obtained from cultured fibroblasts are relevant to the question of sensitivity of epithelial cells which are the predominant cell type in human cancers. (author)
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Directory of Open Access Journals (Sweden)
Kimura M
2012-01-01
Full Text Available Introduction. Amniotic membrane contains a multipotential stem cell population and is expected to possess the machinery to regulate immunological reactions. We investigated the safety and efficacy of allogeneic amniotic membrane-derived mesenchymal stromal cell (AMSC transplantation in a porcine model of chronic myocardial ischemia as a preclinical trial. Methods. Porcine AMSCs were isolated from amniotic membranes obtained by cesarean section just before delivery and were cultured to increase their numbers before transplantation. Chronic myocardial ischemia was induced by implantation of an ameroid constrictor around the left circumflex coronary artery. Four weeks after ischemia induction, nine swine were assigned to undergo either allogeneic AMSC transplantation or normal saline injection. Functional analysis was performed by echocardiography, and histological examinations were carried out by immunohistochemistry 4 weeks after AMSC transplantation. Results. Echocardiography demonstrated that left ventricular ejection fraction was significantly improved and left ventricular dilatation was well attenuated 4 weeks after AMSC transplantation. Histological assessment showed a significant reduction in percentage of fibrosis in the AMSC transplantation group. Injected allogeneic green fluorescent protein (GFP-expressing AMSCs were identified in the immunocompetent host heart without the use of any immunosuppressants 4 weeks after transplantation. Immunohistochemistry revealed that GFP colocalized with cardiac troponin T and cardiac troponin I. Conclusions. We have demonstrated that allogeneic AMSC transplantation produced histological and functional improvement in the impaired myocardium in a porcine model of chronic myocardial ischemia. The transplanted allogeneic AMSCs survived without the use of any immunosuppressants and gained cardiac phenotype through either their transdifferentiation or cell fusion.
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Management of herpes simplex virus epithelial keratitis.
Roozbahani, Mehdi; Hammersmith, Kristin M
2018-04-24
To review recent advancements in the management of herpes simplex virus (HSV) epithelial keratitis. Trifluridine eye drop, acyclovir (ACV) ointment, ganciclovir gel, and oral ACV are still the main therapeutic agents. Cryopreserved amniotic membrane has been recently used as an adjuvant treatment. Resistance to ACV has become a concerning issue. The animal models of HSV vaccine are able to reduce HSV keratitis. New antivirals are under development. Current cases of HSV epithelial keratitis are manageable with available medications, but new advancements are required to decrease disease burden in the future. HSV vaccine can be revolutionary.
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Gomez-Lopez, Nardhy; Romero, Roberto; Xu, Yi; Miller, Derek; Unkel, Ronald; Shaman, Majid; Jacques, Suzanne M; Panaitescu, Bogdan; Garcia-Flores, Valeria; Hassan, Sonia S
2017-08-01
Neutrophil extracellular traps (NETs) control microbial infections through their antimicrobial activities attributed to DNA, histones, granules, and cytoplasmic proteins (eg, elastase). Intra-amniotic infection is characterized by the influx of neutrophils into the amniotic cavity; therefore, the aim of this study was to determine whether amniotic fluid neutrophils form NETs in this inflammatory process. Amniotic fluid samples from women with intra-amniotic infection (n = 15) were stained for bacteria detection using fluorescent dyes. Amniotic fluid neutrophils were purified by filtration. As controls, neutrophils from maternal blood samples (n = 3) were isolated by density gradients. Isolated neutrophils were plated onto glass cover slips for culture with and without 100 nM of phorbol-12-myristate-13-acetate (PMA). NET formation was assessed by 4',6-diamidino-2-phenylindole (DAPI) staining and scanning electron microscopy. Different stages of NET formation were visualized using antibodies against elastase and histone H3, in combination with DAPI staining, by confocal microscopy. Finally, maternal or neonatal neutrophils were added to amniotic fluid samples from women without intra-amniotic infection (n = 4), and NET formation was evaluated by DAPI staining. (1) NETs were present in the amniotic fluid of women with intra-amniotic infection; (2) all of the amniotic fluid samples had detectable live and dead bacteria associated with the presence of NETs; (3) in contrast to neutrophils from the maternal circulation, amniotic fluid neutrophils did not require PMA stimulation to form NETs; (4) different stages of NET formation were observed by co-localizing elastase, histone H3, and DNA in amniotic fluid neutrophils; and (5) neither maternal nor neonatal neutrophils form NETs in the amniotic fluid of women without intra-amniotic infection. NETs are detectable in the amniotic fluid of women with intra-amniotic infection.
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2014-01-01
Introduction Stromal-epithelial interactions play a fundamental role in tissue homeostasis, controlling cell proliferation and differentiation. Not surprisingly, aberrant stromal-epithelial interactions contribute to malignancies. Studies of the cellular and molecular mechanisms underlying these interactions require ex vivo experimental model systems that recapitulate the complexity of human tissue without compromising the differentiation and proliferation potentials of human primary cells. Methods We isolated and characterized human breast epithelial and mesenchymal precursors from reduction mammoplasty tissue and tagged them with lentiviral vectors. We assembled heterotypic co-cultures and compared mesenchymal and epithelial cells to cells in corresponding monocultures by analyzing growth, differentiation potentials, and gene expression profiles. Results We show that heterotypic culture of non-immortalized human primary breast epithelial and mesenchymal precursors maintains their proliferation and differentiation potentials and constrains their growth. We further describe the gene expression profiles of stromal and epithelial cells in co-cultures and monocultures and show increased expression of the tumor growth factor beta (TGFβ) family member inhibin beta A (INHBA) in mesenchymal cells grown as co-cultures compared with monocultures. Notably, overexpression of INHBA in mesenchymal cells increases colony formation potential of epithelial cells, suggesting that it contributes to the dynamic reciprocity between breast mesenchymal and epithelial cells. Conclusions The described heterotypic co-culture system will prove useful for further characterization of the molecular mechanisms mediating interactions between human normal or neoplastic breast epithelial cells and the stroma, and will provide a framework to test the relevance of the ever-increasing number of oncogenomic alterations identified in human breast cancer. PMID:24916766
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CUX1/Wnt signaling regulates Epithelial Mesenchymal Transition in EBV infected epithelial cells
International Nuclear Information System (INIS)
Malizia, Andrea P.; Lacey, Noreen; Walls, Dermot; Egan, Jim J.; Doran, Peter P.
2009-01-01
Idiopathic pulmonary fibrosis (IPF) is a refractory and lethal interstitial lung disease characterized by alveolar epithelial cells apoptosis, fibroblast proliferation and extra-cellular matrix protein deposition. EBV, localised to alveolar epithelial cells of pulmonary fibrosis patients is associated with a poor prognosis. A strategy based on microarray-differential gene expression analysis to identify molecular drivers of EBV-associated lung fibrosis was utilized. Alveolar epithelial cells were infected with EBV to identify genes whose expression was altered following TGFβ1-mediated lytic phase. EBV lytic reactivation by TGFβ1 drives a selective alteration in CUX1 variant (a) (NCBI accession number NM 1 81552) expression, inducing activation of non-canonical Wnt pathway mediators, implicating it in Epithelial Mesenchymal Transition (EMT), the molecular event underpinning scar production in tissue fibrosis. The role of EBV in EMT can be attenuated by antiviral strategies and inhibition of Wnt signaling by using All-Trans Retinoic Acids (ATRA). Activation of non-canonical Wnt signaling pathway by EBV in epithelial cells suggests a novel mechanism of EMT via CUX1 signaling. These data present a framework for further description of the link between infectious agents and fibrosis, a significant disease burden.
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CUX1/Wnt signaling regulates Epithelial Mesenchymal Transition in EBV infected epithelial cells
Energy Technology Data Exchange (ETDEWEB)
Malizia, Andrea P.; Lacey, Noreen [Clinical Research Centre, School of Medicine and Medical Science, University College Dublin. 21, Nelson Street. Dublin, 7. Ireland (Ireland); Walls, Dermot [School of Biotechnology, Dublin City University. Dublin, 9. Ireland (Ireland); Egan, Jim J. [Advanced Lung Disease and Lung Transplant Program, Mater Misericordiae University Hospital. 44, Eccles Street. Dublin, 7. Ireland (Ireland); Doran, Peter P., E-mail: peter.doran@ucd.ie [Clinical Research Centre, School of Medicine and Medical Science, University College Dublin. 21, Nelson Street. Dublin, 7. Ireland (Ireland)
2009-07-01
Idiopathic pulmonary fibrosis (IPF) is a refractory and lethal interstitial lung disease characterized by alveolar epithelial cells apoptosis, fibroblast proliferation and extra-cellular matrix protein deposition. EBV, localised to alveolar epithelial cells of pulmonary fibrosis patients is associated with a poor prognosis. A strategy based on microarray-differential gene expression analysis to identify molecular drivers of EBV-associated lung fibrosis was utilized. Alveolar epithelial cells were infected with EBV to identify genes whose expression was altered following TGF{beta}1-mediated lytic phase. EBV lytic reactivation by TGF{beta}1 drives a selective alteration in CUX1 variant (a) (NCBI accession number NM{sub 1}81552) expression, inducing activation of non-canonical Wnt pathway mediators, implicating it in Epithelial Mesenchymal Transition (EMT), the molecular event underpinning scar production in tissue fibrosis. The role of EBV in EMT can be attenuated by antiviral strategies and inhibition of Wnt signaling by using All-Trans Retinoic Acids (ATRA). Activation of non-canonical Wnt signaling pathway by EBV in epithelial cells suggests a novel mechanism of EMT via CUX1 signaling. These data present a framework for further description of the link between infectious agents and fibrosis, a significant disease burden.
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Jijon, H B; Suarez-Lopez, L; Diaz, O E; Das, S; De Calisto, J; Yaffe, M B; Pittet, M J; Mora, J R; Belkaid, Y; Xavier, R J; Villablanca, E J
2018-05-01
Retinoic acid (RA), a dietary vitamin A metabolite, is crucial in maintaining intestinal homeostasis. RA acts on intestinal leukocytes to modulate their lineage commitment and function. Although the role of RA has been characterized in immune cells, whether intestinal epithelial cells (IECs) rely on RA signaling to exert their immune-regulatory function has not been examined. Here we demonstrate that lack of RA receptor α (RARα) signaling in IECs results in deregulated epithelial lineage specification, leading to increased numbers of goblet cells and Paneth cells. Mechanistically, lack of RARα resulted in increased KLF4 + goblet cell precursors in the distal bowel, whereas RA treatment inhibited klf4 expression and goblet cell differentiation in zebrafish. These changes in secretory cells are associated with increased Reg3g, reduced luminal bacterial detection, and an underdeveloped intestinal immune system, as evidenced by an almost complete absence of lymphoid follicles and gut resident mononuclear phagocytes. This underdeveloped intestinal immune system shows a decreased ability to clear infection with Citrobacter rodentium. Collectively, our findings indicate that epithelial cell-intrinsic RARα signaling is critical to the global development of the intestinal immune system.
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Directory of Open Access Journals (Sweden)
Imran S. Chaudhry
2011-01-01
Full Text Available The biological effects of only a finite number of tobacco toxins have been studied. Here, we describe exposure of cultures of human bronchial epithelial cells to low concentrations of tobacco carcinogens: nickel sulphate, benzo(bfluoranthene, N-nitrosodiethylamine, and 4-(methylnitrosamino-1-(3-pyridyl-1-butanone (NNK. After a 24-hour exposure, EGFR was expressed in cell membrane and cytoplasm, BCL-2 was expressed only in the irregular nuclei of large atypical cells, MKI67 was expressed in nuclei with no staining in larger cells, cytoplasmic BIRC5 with stronger nuclear staining was seen in large atypical cells, and nuclear TP53 was strongly expressed in all cells. After only a 24-hour exposure, cells exhibited atypical nuclear and cytoplasmic features. After a 48-hour exposure, EGFR staining was localized to the nucleus, BCL-2 was slightly decreased in intensity, BIRC5 was localized to the cytoplasm, and TP53 staining was increased in small and large cells. BCL2L1 was expressed in both the cytoplasm and nuclei of cells at 24- and 48-hour exposures. We illustrate that short-termexposure of a bronchial epithelial cell line to smoking-equivalent concentrations of tobacco carcinogens alters the expression of key proliferation regulatory genes, EGFR, BCL-2, BCL2L1, BIRC5, TP53, and MKI67, similar to that reported in biopsy specimens of pulmonary epithelium described to be preneoplastic lesions.
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Globoside accelerates the differentiation of dental epithelial cells into ameloblasts
Institute of Scientific and Technical Information of China (English)
Takashi Nakamura; Yuta Chiba; Masahiro Naruse; Kan Saito; Hidemitsu Harada; Satoshi Fukumoto
2016-01-01
Tooth crown morphogenesis is tightly regulated by the proliferation and differentiation of dental epithelial cells. Globoside (Gb4), a globo-series glycosphingolipid, is highly expressed during embryogenesis as well as organogenesis, including tooth development. We previously reported that Gb4 is dominantly expressed in the neutral lipid fraction of dental epithelial cells. However, because its functional role in tooth development remains unknown, we investigated the involvement of Gb4 in dental epithelial cell differentiation. The expression of Gb4 was detected in ameloblasts of postnatal mouse molars and incisors. A cell culture analysis using HAT-7 cells, a rat-derived dental epithelial cell line, revealed that Gb4 did not promote dental epithelial cell proliferation. Interestingly, exogenous administration of Gb4 enhanced the gene expression of enamel extracellular matrix proteins such as ameloblastin, amelogenin, and enamelin in dental epithelial cells as well as in developing tooth germs. Gb4 also induced the expression of TrkB, one of the key receptors required for ameloblast induction in dental epithelial cells. In contrast, Gb4 downregulated the expression of p75, a receptor for neurotrophins (including neurotrophin-4) and a marker of undifferentiated dental epithelial cells. In addition, we found that exogenous administration of Gb4 to dental epithelial cells stimulated the extracellular signal-regulated kinase and p38 mitogen-activated protein kinase signalling pathways. Furthermore, Gb4 induced the expression of epiprofin and Runx2, the positive regulators for ameloblastin gene transcription. Thus, our results suggest that Gb4 contributes to promoting the differentiation of dental epithelial cells into ameloblasts.
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Heterogeneity of limbal basal epithelial progenitor cells.
Hayashida, Yasutaka; Li, Wei; Chen, Ying-Ting; He, Hua; Chen, Szu-yu; Kheirkah, Ahmad; Zhu, Ying-Tien; Matsumoto, Yukihiro; Tseng, Scheffer C G
2010-11-01
Although corneal epithelial stem cells (SCs) are located at the limbus between the cornea and the conjunctiva, not all limbal basal epithelial cells are SCs. Using 2 dispase digestions to remove different amounts of limbal basal epithelial cells for cross-sections, flat mounts, and cytospin preparations, double immunostaining to pancytokeratins (PCK) and vimentin (Vim) identified 3 p63+ epithelial progenitors such as PCK-/Vim+, PCK/Vim, and PCK-/Vim+ and 1 p63+ mesenchymal cell, PCK-/Vim+. PCK-/Vim- progenitors had the smallest cell size were 10-20 times more enriched on collagen I-coated dishes in the 5-minute rapid adherent fraction that contained the highest percentage of p63+ cells but the lowest percentage of cytokeratin12+ cells, and gave rise to high Ki67 labeling and vivid clonal growth. In contrast, PCK+/Vim+ and PCK+/Vim- progenitors were found more in the slow-adherent fraction and yielded poor clonal growth. PCK/Vim progenitors and clusters of PCK-/Vim+ mesenchymal cells, which were neither melanocytes nor Langerhans cells, were located in the limbal basal region. Therefore, differential expression of PCK and Vim helps identify small PCK-/Vim- cells as the most likely candidate for SCs among a hierarchy of heterogeneous limbal basal progenitors, and their close association with PCK-/Vim+ presumed "niche" cells.
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Interaction with Epithelial Cells Modifies Airway Macrophage Response to Ozone
The initial innate immune response to ozone (03) in the lung is orchestrated by structural cells, such as epithelial cells, and resident immune cells, such as airway macrophages (Macs). We developed an epithelial cell-Mac coculture model to investigate how epithelial cell-derived...
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Rabbit uterine epithelial cells: Co-culture with spermatozoa
International Nuclear Information System (INIS)
Boice, M.L.
1988-01-01
A primary culture of rabbit uterine epithelial cells was established and their effects on sperm function were examined in vitro. Epithelial cells were isolated from uteri of estrous rabbits and cultured on floating collagen gels in phenol red-free medium supplemented with 5% fetal bovine serum. Light microscopy and keratin staining showed that the epithelial cell population established in culture had morphological characteristics similar to that seen in the intact endometrium. Cells were cultured with 3 H-leucine and uptake of label by cells and its incorporation into cellular and secretory proteins determined. When compared to cells cultured for 24-48 h, incorporation of label into cellular protein was lower at 72-96 h, but secretion increased. Estradiol 17-β did not affect label uptake or incorporation, but did enhance proliferation of cells as judged by total DNA content of the cell population. Analysis of proteins in media by sodium dodecyl sulfate polyacrylamide gel electrophoresis and fluorography suggested that epithelial and stromal cells synthesis proteins that may be secretory in nature during 72-96 h culture. Twenty-nine to thirty-one h after initiation of epithelial cultures, 1-2 x 10 6 sperm were co-incubated with cells and sperm viability, motility, loss of acrosome and fertilizing ability determined
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Cell volume regulation in epithelial physiology and cancer
DEFF Research Database (Denmark)
Pedersen, Stine Helene Falsig; Hoffmann, Else Kay; Novak, Ivana
2013-01-01
expression of ion transporters and channels is now recognized as one of the hallmarks of cancer, it is timely to consider this especially for epithelia. Epithelial cells are highly proliferative and epithelial cancers, carcinomas, account for about 90% of all cancers. In this review we will focus on ion...... such as cancer, transepithelial and cell volume regulatory ion transport are dys-regulated. Furthermore, epithelial architecture and coordinated ion transport function are lost, cell survival/death balance is altered, and new interactions with the stroma arise, all contributing to drug resistance. Since altered...... transporters and channels with key physiological functions in epithelia and known roles in the development of cancer in these tissues. Their roles in cell survival, cell cycle progression, and development of drug resistance in epithelial cancers will be discussed....
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No junctional communication between epithelial cells in hydra
DEFF Research Database (Denmark)
de Laat, S W; Tertoolen, L G; Grimmelikhuijzen, C J
1980-01-01
junctions between epithelial cells of hydra. However, until now, there has been no report published on whether these junctions enable the epithelial cells to exchange molecules of small molecular weight, as has been described in other organisms. Therefore we decided to investigate the communicative...... properties of the junctional membranes by electrophysiological methods and by intracellular-dye iontophoresis. We report here that no electrotonic coupling is detectable between epithelial cells of Hydra attenuata in: (1) intact animals, (2) head-regenerating animals, (3) cell re-aggregates, and (4) hydra...
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Human retinal pigment epithelial cell-induced apoptosis in activated T cells
DEFF Research Database (Denmark)
Jørgensen, A; Wiencke, A K; la Cour, M
1998-01-01
human retinal pigment epithelial (RPE) cells can induce apoptosis in activated T cells. METHODS: Fas ligand (FasL) expression was detected by flow cytometry and immunohistochemistry. Cultured RPE cells were cocultured with T-cell lines and peripheral blood lymphocytes for 6 hours to 2 days. Induction...... of apoptosis was detected by 7-amino-actinomycin D and annexin V staining. RESULTS: Retinal pigment epithelial cells expressed FasL and induced apoptosis in activated Fas+ T cells. Blocking of Fas-FasL interaction with antibody strongly inhibited RPE-mediated T-cell apoptosis. Retinal pigment epithelial cells...... induced apoptosis in several activated T-cell populations and T-cell lines, including T-cell antigen receptor (TCR)-CD3-negative T-cell lines. In contrast, RPE cells induced little or no apoptosis in resting peripheral T cells. Major histocompatibility complex (MHC) class II monoclonal antibodies, which...
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Ding, Chenyue; Li, Hong; Wang, Yun; Wang, Fuxin; Wu, Huihua; Chen, Rulei; Lv, Jinghuan; Wang, Wei; Huang, Boxian
2017-07-27
Many reports have shown that various kinds of stem cells have the ability to recover premature ovarian aging (POA) function. Transplantation of human amniotic epithelial cells (hAECs) improves ovarian function damaged by chemotherapy in a mice model. Understanding of how to evaluate the distinct effects of adult stem cells in curing POA and how to choose stem cells in clinical application is lacking. To build a different degrees of POA model, mice were administered different doses of cyclophosphamide: light dose (70 mg/kg, 2 weeks), medium dose (70 mg/kg, 1 week; 120 mg/kg, 1 week), and high dose (120 mg/kg, 2 weeks). Enzyme-linked immunosorbent assay detected serum levels of sex hormones, and hematoxylin and eosin staining allowed follicle counting and showed the ovarian tissue structure. DiIC 18 (5)-DS was employed to label human amniotic mesenchymal stem cells (hAMSCs) and hAECs for detecting the cellular retention time in ovaries by a live imaging system. Proliferation of human ovarian granule cells (ki67, AMH, FSHR, FOXL2, and CYP19A1) and immunological rejection of human peripheral blood mononuclear cells (CD4, CD11b, CD19, and CD56) were measured by flow cytometry (fluorescence-activated cell sorting (FACS)). Distinction of cellular biological characteristics between hAECs and hAMSCs was evaluated, such as collagen secretory level (collagen I, II, III, IV, and VI), telomerase activity, pluripotent markers tested by western blot, expression level of immune molecules (HLA-ABC and HLA-DR) analyzed by FACS, and cytokines (growth factors, chemotactic factors, apoptosis factors, and inflammatory factors) measured by a protein antibody array methodology. After hAMSCs and hAECs were transplanted into a different degrees of POA model, hAMSCs exerted better therapeutic activity on mouse ovarian function in the high-dose administration group, promoting the proliferation rate of ovarian granular cells from premature ovarian failure patients, but also provoking immune
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Peng, Shao-Yu; Chen, Yu-Hsu; Chou, Chih-Jen; Wang, Yao-Horng; Lee, Hung-Maan; Cheng, Winston Teng-Kui; Shaw, S W Steven; Wu, Shinn-Chih
2014-05-01
Amniotic fluid stem cells (AFSCs) are derived from the amniotic fluid of the developing fetus and can give rise to diverse differentiated cells of ectoderm, mesoderm, and endoderm lineages. Intrauterine transplantation is an approach used to cure inherited genetic fetal defects during the gestation period of pregnant dams. Certain disease such as osteogenesis imperfecta was successfully treated in affected fetal mice using this method. However, the donor cell destiny remains uncertain. The purpose of this study was to investigate the biodistribution and cell fate of Ds-red-harboring porcine AFSCs (Ds-red pAFSCs) after intrauterine transplantation into enhanced green fluorescent protein-harboring fetuses of pregnant mice. Pregnant mice (12.5 days) underwent open laparotomy with intrauterine pAFSC transplantation (5 × 10(4) cells per pup) into fetal peritoneal cavity. Three weeks after birth, the mice were sacrificed. Several samples from different organs were obtained for histological examination and flow cytometric analysis. Ds-red pAFSCs migrated most frequently into the intestines. Furthermore, enhanced green fluorescent protein and red fluorescent protein signals were co-expressed in the intestine and liver cells via immunohistochemistry studies. In utero xenotransplantation of pAFSCs fused with recipient intestinal cells instead of differentiating or maintaining the undifferentiated status in the tissue. © 2014 John Wiley & Sons, Ltd.
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Jiang, Chunmiao; Zhang, Qunzhou; Shanti, Rabie M; Shi, Shihong; Chang, Ting-Han; Carrasco, Lee; Alawi, Faizan; Le, Anh D
2017-09-01
Epithelial-mesenchymal transition (EMT), a biological process associated with cancer stem-like or cancer-initiating cell formation, contributes to the invasiveness, metastasis, drug resistance, and recurrence of the malignant tumors; it remains to be determined whether similar processes contribute to the pathogenesis and progression of ameloblastoma (AM), a benign but locally invasive odontogenic neoplasm. Here, we demonstrated that EMT- and stem cell-related genes were expressed in the epithelial islands of the most common histologic variant subtype, the follicular AM. Our results revealed elevated interleukin (IL)-6 signals that were differentially expressed in the stromal compartment of the follicular AM. To explore the stromal effect on tumor pathogenesis, we isolated and characterized both mesenchymal stromal cells (AM-MSCs) and epithelial cells (AM-EpiCs) from follicular AM and demonstrated that, in in vitro culture, AM-MSCs secreted a significantly higher level of IL-6 as compared to the counterpart AM-EpiCs. Furthermore, both in vitro and in vivo studies revealed that exogenous and AM-MSC-derived IL-6 induced the expression of EMT- and stem cell-related genes in AM-EpiCs, whereas such effects were significantly abrogated either by a specific inhibitor of STAT3 or ERK1/2, or by knockdown of Slug gene expression. These findings suggest that AM-MSC-derived IL-6 promotes tumor-stem like cell formation by inducing EMT process in AM-EpiCs through STAT3 and ERK1/2-mediated signaling pathways, implying a role in the etiology and progression of the benign but locally invasive neoplasm. Stem Cells 2017;35:2083-2094. © 2017 AlphaMed Press.
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Replication of cultured lung epithelial cells
International Nuclear Information System (INIS)
Guzowski, D.; Bienkowski, R.
1986-01-01
The authors have investigated the conditions necessary to support replication of lung type 2 epithelial cells in culture. Cells were isolated from mature fetal rabbit lungs (29d gestation) and cultured on feeder layers of mitotically inactivated 3T3 fibroblasts. The epithelial nature of the cells was demonstrated by indirect immunofluorescent staining for keratin and by polyacid dichrome stain. Ultrastructural examination during the first week showed that the cells contained myofilaments, microvilli and lamellar bodies (markers for type 2 cells). The following changes were observed after the first week: increase in cell size; loss of lamellar bodies and appearance of multivesicular bodies; increase in rough endoplasmic reticulum and golgi; increase in tonafilaments and well-defined junctions. General cell morphology was good for up to 10 wk. Cells cultured on plastic surface degenerated after 1 wk. Cell replication was assayed by autoradiography of cultures exposed to ( 3 H)-thymidine and by direct cell counts. The cells did not replicate during the first week; however, between 2-10 wk the cells incorporated the label and went through approximately 6 population doublings. They have demonstrated that lung alveolar epithelial cells can replicate in culture if they are maintained on an appropriate substrate. The coincidence of ability to replicate and loss of markers for differentiation may reflect the dichotomy between growth and differentiation commonly observed in developing systems
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Human retinal pigment epithelial cell-induced apoptosis in activated T cells
DEFF Research Database (Denmark)
Jørgensen, A; Wiencke, A K; la Cour, M
1998-01-01
PURPOSE: The immune privilege of the eye has been thought to be dependent on physical barriers and absence of lymphatic vessels. However, the immune privilege may also involve active immunologic processes, as recent studies have indicated. The purpose of the present study was to investigate whether...... human retinal pigment epithelial (RPE) cells can induce apoptosis in activated T cells. METHODS: Fas ligand (FasL) expression was detected by flow cytometry and immunohistochemistry. Cultured RPE cells were cocultured with T-cell lines and peripheral blood lymphocytes for 6 hours to 2 days. Induction...... of apoptosis was detected by 7-amino-actinomycin D and annexin V staining. RESULTS: Retinal pigment epithelial cells expressed FasL and induced apoptosis in activated Fas+ T cells. Blocking of Fas-FasL interaction with antibody strongly inhibited RPE-mediated T-cell apoptosis. Retinal pigment epithelial cells...
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Directory of Open Access Journals (Sweden)
Raquel Rodrigues-Diez
2014-01-01
Full Text Available Gremlin is a developmental gene upregulated in human chronic kidney disease and in renal cells in response to transforming growth factor-β (TGF-β. Epithelial mesenchymal transition (EMT is one process involved in renal fibrosis. In tubular epithelial cells we have recently described that Gremlin induces EMT and acts as a downstream TGF-β mediator. Our aim was to investigate whether Gremlin participates in EMT by the regulation of the Smad pathway. Stimulation of human tubular epithelial cells (HK2 with Gremlin caused an early activation of the Smad signaling pathway (Smad 2/3 phosphorylation, nuclear translocation, and Smad-dependent gene transcription. The blockade of TGF-β, by a neutralizing antibody against active TGF-β, did not modify Gremlin-induced early Smad activation. These data show that Gremlin directly, by a TGF-β independent process, activates the Smad pathway. In tubular epithelial cells long-term incubation with Gremlin increased TGF-β production and caused a sustained Smad activation and a phenotype conversion into myofibroblasts-like cells. Smad 7 overexpression, which blocks Smad 2/3 activation, diminished EMT changes observed in Gremlin-transfected tubuloepithelial cells. TGF-β neutralization also diminished Gremlin-induced EMT changes. In conclusion, we propose that Gremlin could participate in renal fibrosis by inducing EMT in tubular epithelial cells through activation of Smad pathway and induction of TGF-β.
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Nakajima, Ryota; Takeda, Shizu
2014-01-01
The use of murine 3T3 feeder cells needs to be avoided when fabricating corneal epithelial cell sheets for use in treating ocular surface diseases. However, the expression level of the epithelial stem/progenitor cell marker, p63, is down-regulated in feeder-free culture systems. In this study, in order to fabricate corneal epithelial cell sheets that maintain colony-forming cells without using any feeder cells, we investigated the use of an oxygen-controlled method that was developed previously to fabricate cell sheets efficiently. Rabbit limbal epithelial cells were cultured under hypoxia (1-10% O2) and under normoxia during stratification after reaching confluence. Multilayered corneal epithelial cell sheets were fabricated using an oxygen-controlled method, and immunofluorescence analysis showed that cytokeratin 3 and p63 was expressed in appropriate localization in the cell sheets. The colony-forming efficiency of the cell sheets fabricated by the oxygen-controlled method without feeder cells was significantly higher than that of cell sheets fabricated under 20% O2 without feeder cells. These results indicate that the oxygen-controlled method has the potential to achieve a feeder-free culture system for fabricating corneal epithelial cell sheets for corneal regeneration. Copyright © 2013 Elsevier Ltd. All rights reserved.
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Lingual Epithelial Stem Cells and Organoid Culture of Them
Directory of Open Access Journals (Sweden)
Hiroko Hisha
2016-01-01
Full Text Available As tongue cancer is one of the major malignant cancers in the world, understanding the mechanism of maintenance of lingual epithelial tissue, which is known to be the origin of tongue cancer, is unquestionably important. However, the actual stem cells that are responsible for the long-term maintenance of the lingual epithelium have not been identified. Moreover, a simple and convenient culture method for lingual epithelial stem cells has not yet been established. Recently, we have shown that Bmi1-positive cells, residing at the second or third layer of the epithelial cell layer at the base of the interpapillary pit (IPP, were slow-cycling and could supply keratinized epithelial cells for over one year, indicating that Bmi1-positive cells are long-term lingual epithelial stem cells. In addition, we have developed a novel lingual epithelium organoid culture system using a three-dimensional matrix and growth factors. Here, we discuss current progress in the identification of lingual stem cells and future applications of the lingual culture system for studying the regulatory mechanisms of the lingual epithelium and for regenerative medicine.
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Role of Corneal Stromal Cells on Epithelial Cell Function during Wound Healing
Directory of Open Access Journals (Sweden)
Bhavani S. Kowtharapu
2018-02-01
Full Text Available Following injury, corneal stromal keratocytes transform into repair-phenotype of activated stromal fibroblasts (SFs and participate in wound repair. Simultaneously, ongoing bi-directional communications between corneal stromal-epithelial cells also play a vital role in mediating the process of wound healing. Factors produced by stromal cells are known to induce proliferation, differentiation, and motility of corneal epithelial cells, which are also subsequently the main processes that occur during wound healing. In this context, the present study aims to investigate the effect of SFs conditioned medium (SFCM on corneal epithelial cell function along with substance P (SP. Antibody microarrays were employed to profile differentially expressed cell surface markers and cytokines in the presence of SFCM and SP. Antibody microarray data revealed enhanced expression of the ITGB1 in corneal epithelial cells following stimulation with SP whereas SFCM induced abundant expression of IL-8, ITGB1, PD1L1, PECA1, IL-15, BDNF, ICAM1, CD8A, CD44 and NTF4. All these proteins have either direct or indirect roles in epithelial cell growth, movement and adhesion related signaling cascades during tissue regeneration. We also observed activation of MAPK signaling pathway along with increased expression of focal adhesion kinase (FAK, paxillin, vimentin, β-catenin and vasodilator-stimulated phosphoprotein (VASP phosphorylation. Additionally, epithelial-to-mesenchymal transition (EMT regulating transcription factors Slug and ZEB1 expression were enhanced in the presence of SFCM. SP enriched the expression of integrin subunits α4, α5, αV, β1 and β3 whereas SFCM increased α4, α5, αV, β1 and β5 integrin subunits. We also observed increased expression of Serpin E1 following SP and SFCM treatment. Wound healing scratch assay revealed enhanced migration of epithelial cells following the addition of SFCM. Taken together, we conclude that SFCM-mediated sustained
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CD8+ T cells induce thyroid epithelial cell hyperplasia and fibrosis.
Yu, Shiguang; Fang, Yujiang; Sharav, Tumenjargal; Sharp, Gordon C; Braley-Mullen, Helen
2011-02-15
CD8(+) T cells can be important effector cells in autoimmune inflammation, generally because they can damage target cells by cytotoxicity. This study shows that activated CD8(+) T cells induce thyroid epithelial cell hyperplasia and proliferation and fibrosis in IFN-γ(-/-) NOD.H-2h4 SCID mice in the absence of CD4(+) T cells. Because CD8(+) T cells induce proliferation rather than cytotoxicity of target cells, these results describe a novel function for CD8(+) T cells in autoimmune disease. In contrast to the ability of purified CD8(+) T cells to induce thyrocyte proliferation, CD4(+) T cells or CD8 T cell-depleted splenocytes induced only mild thyroid lesions in SCID recipients. T cells in both spleens and thyroids highly produce TNF-α. TNF-α promotes proliferation of thyrocytes in vitro, and anti-TNF-α inhibits development of thyroid epithelial cell hyperplasia and proliferation in SCID recipients of IFN-γ(-/-) splenocytes. This suggests that targeting CD8(+) T cells and/or TNF-α may be effective for treating epithelial cell hyperplasia and fibrosis.
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International Nuclear Information System (INIS)
Hubbs, A.F.; Hahn, F.F.; Kelly, G.; Thomassen, D.G.
1988-01-01
Transforming growth factor beta (TGFβ) markedly inhibited growth of canine tracheal epithelial (CTE) cells. Reduced responsiveness to TGFβ-induced growth inhibition accompanied neoplastic progression of these cells from primary to transformed to neoplastic. This was similar to the relationship between neoplastic progression and increased resistance to TGFβ-induced growth inhibition seen for rat tracheal epithelial (RTE) cells. The canine cells were more sensitive than rat cells to TGFβ-induced growth inhibition at all stages in the neoplastic process. (author)
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Membrane lipidome of an epithelial cell line
DEFF Research Database (Denmark)
Sampaio, Julio L; Gerl, Mathias J; Klose, Christian
2011-01-01
Tissue differentiation is an important process that involves major cellular membrane remodeling. We used Madin-Darby canine kidney cells as a model for epithelium formation and investigated the remodeling of the total cell membrane lipidome during the transition from a nonpolarized morphology...... to an epithelial morphology and vice versa. To achieve this, we developed a shotgun-based lipidomics workflow that enabled the absolute quantification of mammalian membrane lipidomes with minimal sample processing from low sample amounts. Epithelial morphogenesis was accompanied by a major shift from sphingomyelin...... to generate an apical membrane domain that serves as a protective barrier for the epithelial sheet....
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Implantation of Induced Pluripotent Stem Cell-Derived Tracheal Epithelial Cells.
Ikeda, Masakazu; Imaizumi, Mitsuyoshi; Yoshie, Susumu; Nakamura, Ryosuke; Otsuki, Koshi; Murono, Shigeyuki; Omori, Koichi
2017-07-01
Compared with using autologous tissue, the use of artificial materials in the regeneration of tracheal defects is minimally invasive. However, this technique requires early epithelialization on the inner side of the artificial trachea. After differentiation from induced pluripotent stem cells (iPSCs), tracheal epithelial tissues may be used to produce artificial tracheas. Herein, we aimed to demonstrate that after differentiation from fluorescent protein-labeled iPSCs, tracheal epithelial tissues survived in nude rats with tracheal defects. Red fluorescent tdTomato protein was electroporated into mouse iPSCs to produce tdTomato-labeled iPSCs. Embryoid bodies derived from these iPSCs were then cultured in differentiation medium supplemented with growth factors, followed by culture on air-liquid interfaces for further differentiation into tracheal epithelium. The cells were implanted with artificial tracheas into nude rats with tracheal defects on day 26 of cultivation. On day 7 after implantation, the tracheas were exposed and examined histologically. Tracheal epithelial tissue derived from tdTomato-labeled iPSCs survived in the tracheal defects. Moreover, immunochemical analyses showed that differentiated tissues had epithelial structures similar to those of proximal tracheal tissues. After differentiation from iPSCs, tracheal epithelial tissues survived in rat bodies, warranting the use of iPSCs for epithelial regeneration in tracheal defects.
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Oral epithelial cells are susceptible to cell-free and cell-associated HIV-1 infection in vitro
International Nuclear Information System (INIS)
Moore, Jennifer S.; Rahemtulla, Firoz; Kent, Leigh W.; Hall, Stacy D.; Ikizler, Mine R.; Wright, Peter F.; Nguyen, Huan H.; Jackson, Susan
2003-01-01
Epithelial cells lining the oral cavity are exposed to HIV-1 through breast-feeding and oral-genital contact. Genital secretions and breast milk of HIV-1-infected subjects contain both cell-free and cell-associated virus. To determine if oral epithelial cells can be infected with HIV-1 we exposed gingival keratinocytes and adenoid epithelial cells to cell-free virus and HIV-1-infected peripheral blood mononuclear cells and monocytes. Using primary isolates we determined that gingival keratinocytes are susceptible to HIV-1 infection via cell-free CD4-independent infection only. R5 but not X4 viral strains were capable of infecting the keratinocytes. Further, infected cells were able to release infectious virus. In addition, primary epithelial cells isolated from adenoids were also susceptible to infection; both cell-free and cell-associated virus infected these cells. These data have potential implications in the transmission of HIV-1 in the oral cavity
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Yamada, Mitsuhiro; Kubo, Hiroshi; Ota, Chiharu; Takahashi, Toru; Tando, Yukiko; Suzuki, Takaya; Fujino, Naoya; Makiguchi, Tomonori; Takagi, Kiyoshi; Suzuki, Takashi; Ichinose, Masakazu
2013-09-24
The excess and persistent accumulation of fibroblasts due to aberrant tissue repair results in fibrotic diseases such as idiopathic pulmonary fibrosis. Recent reports have revealed significant changes in microRNAs during idiopathic pulmonary fibrosis and evidence in support of a role for microRNAs in myofibroblast differentiation and the epithelial-mesenchymal transition in the context of fibrosis. It has been reported that microRNA-21 is up-regulated in myofibroblasts during fibrosis and promotes transforming growth factor-beta signaling by inhibiting Smad7. However, expression changes in microRNA-21 and the role of microRNA-21 in epithelial-mesenchymal transition during lung fibrosis have not yet been defined. Lungs from saline- or bleomycin-treated C57BL/6 J mice and lung specimens from patients with idiopathic pulmonary fibrosis were analyzed. Enzymatic digestions were performed to isolate single lung cells. Lung epithelial cells were isolated by flow cytometric cell sorting. The expression of microRNA-21 was analyzed using both quantitative PCR and in situ hybridization. To induce epithelial-mesenchymal transition in culture, isolated mouse lung alveolar type II cells were cultured on fibronectin-coated chamber slides in the presence of transforming growth factor-β, thus generating conditions that enhance epithelial-mesenchymal transition. To investigate the role of microRNA-21 in epithelial-mesenchymal transition, we transfected cells with a microRNA-21 inhibitor. Total RNA was isolated from the freshly isolated and cultured cells. MicroRNA-21, as well as mRNAs of genes that are markers of alveolar epithelial or mesenchymal cell differentiation, were quantified using quantitative PCR. The lung epithelial cells isolated from the bleomycin-induced lung fibrosis model system had decreased expression of epithelial marker genes, whereas the expression of mesenchymal marker genes was increased. MicroRNA-21 was significantly upregulated in isolated lung epithelial
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Schallenberg, Maurice; Westekemper, Henrike; Steuhl, Klaus-Peter; Meller, Daniel
2013-12-17
Mooren's ulcer is a severe ulcerative inflammation of the cornea. The exact pathogenesis remains unclear. Therefore many therapies of Mooren's ulcer are recommended in literature. To shed more light on the ongoing question of optimal treatment of severe progressive Mooren's ulcer, we here report on a retrospective case series of patients treated with systemic immunosuppressive therapy and additional amniotic membrane transplantation. Medical records from seven patients (eleven eyes), 4 male and 3 female, with severe progressive Mooren's ulcer were analysed retrospectively. The mean follow up was 88.4 ± 80.8 months (range 12-232 month). A HLA-typing was performed in all patients. A systemic immunosuppressive therapy was administered in all patients. The amniotic membrane was transplanted after the base of the ulcer was resected. Multiple amniotic membrane transplantations were necessary in six patients. The visual outcome of all patients was poor. No patient achieved a visual acuity better than 20/630 Snellen chart. Five patients were positive for HLA-DQ2 and four patients were positive for HLA-DR17(3). The aggressive and highly inflammatory form of Mooren's ulcer is difficult to treat and the progression of the disease is hard to influence positively even under systemic immunosuppressive therapy. Therefore, the main intention of therapy is to achieve a stable epithelialized corneal surface without the risk of perforation. Amniotic membrane transplantation is not able to cure severe forms of Mooren's ulcer. However it supports the immunosuppressive therapy in acute situations as in critical corneal thinning.
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Probiotics promote endocytic allergen degradation in gut epithelial cells
International Nuclear Information System (INIS)
Song, Chun-Hua; Liu, Zhi-Qiang; Huang, Shelly; Zheng, Peng-Yuan; Yang, Ping-Chang
2012-01-01
Highlights: ► Knockdown of A20 compromised the epithelial barrier function. ► The fusion of endosome/lysosome was disturbed in the A20-deficient HT-29 cells. ► Antigens transported across A20-deficient HT-29 monolayers conserved antigenicity. ► Probiotic proteins increased the expression of A20 in HT-29 cells. -- Abstract: Background and aims: Epithelial barrier dysfunction plays a critical role in the pathogenesis of allergic diseases; the mechanism is to be further understood. The ubiquitin E3 ligase A20 (A20) plays a role in the endocytic protein degradation in the cells. This study aims to elucidate the role of A20 in the maintenance of gut epithelial barrier function. Methods: Gut epithelial cell line, HT-29 cell, was cultured into monolayers to evaluate the barrier function in transwells. RNA interference was employed to knock down the A20 gene in HT-29 cells to test the role of A20 in the maintenance of epithelial barrier function. Probiotic derived proteins were extracted from the culture supernatants using to enhance the expression of A20 in HT-29 cells. Results: The results showed that the knockdown of A20 compromised the epithelial barrier function in HT-29 monolayers, mainly increased the intracellular permeability. The fusion of endosome/lysosome was disturbed in the A20-deficient HT-29 cells. Allergens collected from the transwell basal chambers of A20-deficient HT-29 monolayers still conserved functional antigenicity. Treating with probiotic derived proteins increased the expression of A20 in HT-29 cells and promote the barrier function. Conclusion: A20 plays an important role in the maintenance of epithelial barrier function as shown by HT-29 monolayer. Probiotic derived protein increases the expression of A20 and promote the HT-29 monolayer barrier function.
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Probiotics promote endocytic allergen degradation in gut epithelial cells
Energy Technology Data Exchange (ETDEWEB)
Song, Chun-Hua [Department of Epidemiology and Biostatistics, College of Public Health, Zhengzhou University, Zhengzhou (China); Liu, Zhi-Qiang [Department of Gastroenterology, The Second Hospital, Zhengzhou University, Zhengzhou (China); Department of Pathology and Molecular Medicine, McMaster University, Hamilton, ON (Canada); Huang, Shelly [Department of Pathology and Molecular Medicine, McMaster University, Hamilton, ON (Canada); Zheng, Peng-Yuan, E-mail: medp7123@126.com [Department of Gastroenterology, The Second Hospital, Zhengzhou University, Zhengzhou (China); Yang, Ping-Chang, E-mail: yangp@mcmaster.ca [Department of Pathology and Molecular Medicine, McMaster University, Hamilton, ON (Canada)
2012-09-14
Highlights: Black-Right-Pointing-Pointer Knockdown of A20 compromised the epithelial barrier function. Black-Right-Pointing-Pointer The fusion of endosome/lysosome was disturbed in the A20-deficient HT-29 cells. Black-Right-Pointing-Pointer Antigens transported across A20-deficient HT-29 monolayers conserved antigenicity. Black-Right-Pointing-Pointer Probiotic proteins increased the expression of A20 in HT-29 cells. -- Abstract: Background and aims: Epithelial barrier dysfunction plays a critical role in the pathogenesis of allergic diseases; the mechanism is to be further understood. The ubiquitin E3 ligase A20 (A20) plays a role in the endocytic protein degradation in the cells. This study aims to elucidate the role of A20 in the maintenance of gut epithelial barrier function. Methods: Gut epithelial cell line, HT-29 cell, was cultured into monolayers to evaluate the barrier function in transwells. RNA interference was employed to knock down the A20 gene in HT-29 cells to test the role of A20 in the maintenance of epithelial barrier function. Probiotic derived proteins were extracted from the culture supernatants using to enhance the expression of A20 in HT-29 cells. Results: The results showed that the knockdown of A20 compromised the epithelial barrier function in HT-29 monolayers, mainly increased the intracellular permeability. The fusion of endosome/lysosome was disturbed in the A20-deficient HT-29 cells. Allergens collected from the transwell basal chambers of A20-deficient HT-29 monolayers still conserved functional antigenicity. Treating with probiotic derived proteins increased the expression of A20 in HT-29 cells and promote the barrier function. Conclusion: A20 plays an important role in the maintenance of epithelial barrier function as shown by HT-29 monolayer. Probiotic derived protein increases the expression of A20 and promote the HT-29 monolayer barrier function.
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DEFF Research Database (Denmark)
Toft, Gunnar; Jönsson, Bo A; Bonde, Jens P
2016-01-01
BACKGROUND: Exposure to perfluorooctane sulfonate (PFOS) may potentially disturb fetal Leydig cell hormone production and male genital development. OBJECTIVES: We aimed to study the associations between levels of amniotic fluid PFOS, fetal steroid hormone, and insulin-like factor 3 (INSL3...... associations with fetal hormone levels may have long-term implications for reproductive health. CITATION: Toft G, Jönsson BA, Bonde JP, Nørgaard-Pedersen B, Hougaard DM, Cohen A, Lindh CH, Ivell R, Anand-Ivell R, Lindhard MS. 2016. Perfluorooctane sulfonate concentrations in amniotic fluid, biomarkers of fetal...
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Chitosan Cross-linked Reconstituted Amniotic Collagen Membrane ...
Indian Academy of Sciences (India)
First page Back Continue Last page Overview Graphics. Chitosan Cross-linked Reconstituted Amniotic Collagen Membrane – An Excellent Cell Substratum. The KERATINOCYTE proliferation and Differentiation into multiple layers is due to the presence of type - IV collagen in the amnion. Cultured FIBROBLASTS had good ...
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Sequestration of human cytomegalovirus by human renal and mammary epithelial cells
Energy Technology Data Exchange (ETDEWEB)
Twite, Nicolas [Institute for Medical Immunology, Université Libre de Bruxelles, Rue A. Bolland 8, B-6041 Charleroi (Belgium); Andrei, Graciela [Laboratory of Virology and Chemotherapy, Department of Microbiology and Immunology, Rega Institute for Medical Research, KU Leuven (Belgium); Kummert, Caroline [ImmuneHealth, Rue A. Bolland 8, B-6041 Charleroi (Belgium); Donner, Catherine [Department of Obstetrics and Gynecology, Erasme Hospital, Route de Lennik 808, 1070 Brussels (Belgium); Perez-Morga, David [Laboratory of Molecular Parasitology, Institut de Biologie et Médecine Moléculaires, Université Libre de Bruxelles, Gosselies (Belgium); De Vos, Rita [Pathology Department, U.Z. Leuven, Minderbroedersstraat 12, Leuven (Belgium); Snoeck, Robert, E-mail: Robert.Snoeck@Rega.kuleuven.be [Laboratory of Virology and Chemotherapy, Department of Microbiology and Immunology, Rega Institute for Medical Research, KU Leuven (Belgium); Marchant, Arnaud, E-mail: arnaud.marchant@ulb.ac.be [Institute for Medical Immunology, Université Libre de Bruxelles, Rue A. Bolland 8, B-6041 Charleroi (Belgium); ImmuneHealth, Rue A. Bolland 8, B-6041 Charleroi (Belgium)
2014-07-15
Urine and breast milk represent the main routes of human cytomegalovirus (HCMV) transmission but the contribution of renal and mammary epithelial cells to viral excretion remains unclear. We observed that kidney and mammary epithelial cells were permissive to HCMV infection and expressed immediate early, early and late antigens within 72 h of infection. During the first 24 h after infection, high titers of infectious virus were measured associated to the cells and in culture supernatants, independently of de novo synthesis of virus progeny. This phenomenon was not observed in HCMV-infected fibroblasts and suggested the sequestration and the release of HCMV by epithelial cells. This hypothesis was supported by confocal and electron microscopy analyses. The sequestration and progressive release of HCMV by kidney and mammary epithelial cells may play an important role in the excretion of the virus in urine and breast milk and may thereby contribute to HCMV transmission. - Highlights: • Primary renal and mammary epithelial cells are permissive to HCMV infection. • HCMV is sequestered by epithelial cells and this phenomenon does not require viral replication. • HCMV sequestration by epithelial cells is reduced by antibodies and IFN-γ.
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Energy Technology Data Exchange (ETDEWEB)
Lasalvia, Maria [Department of Clinical and Experimental Medicine, University of Foggia, Foggia (Italy); Istituto Nazionale di Fisica Nucleare, Sezione di Bari, Bari (Italy); Castellani, Stefano [Department of Medical and Surgical Sciences, University of Foggia, Foggia (Italy); D’Antonio, Palma [Department of Clinical and Experimental Medicine, University of Foggia, Foggia (Italy); Perna, Giuseppe [Department of Clinical and Experimental Medicine, University of Foggia, Foggia (Italy); Istituto Nazionale di Fisica Nucleare, Sezione di Bari, Bari (Italy); Carbone, Annalucia [Department of Medical and Surgical Sciences, University of Foggia, Foggia (Italy); Colia, Anna Laura; Maffione, Angela Bruna [Department of Clinical and Experimental Medicine, University of Foggia, Foggia (Italy); Capozzi, Vito [Department of Clinical and Experimental Medicine, University of Foggia, Foggia (Italy); Istituto Nazionale di Fisica Nucleare, Sezione di Bari, Bari (Italy); Conese, Massimo, E-mail: massimo.conese@unifg.it [Department of Medical and Surgical Sciences, University of Foggia, Foggia (Italy)
2016-10-15
The pathophysiology of cystic fibrosis (CF) airway disease stems from mutations in the CF Transmembrane Conductance Regulator (CFTR) gene, leading to a chronic respiratory disease. Actin cytoskeleton is disorganized in CF airway epithelial cells, likely contributing to the CF-associated basic defects, i.e. defective chloride secretion and sodium/fluid hypersorption. In this work, we aimed to find whether this alteration could be pointed out by means of Atomic Force Microscopy (AFM) investigation, as roughness and Young's elastic module. Moreover, we also sought to determine whether disorganization of actin cytoskeleton is linked to hypersoption of apical fluid. Not only CFBE41o- (CFBE) cells, immortalized airway epithelial cells homozygous for the F508del CFTR allele, showed a different morphology in comparison with 16HBE14o- (16HBE) epithelial cells, wild-type for CFTR, but also they displayed a lack of stress fibers, suggestive of a disorganized actin cytoskeleton. AFM measurements showed that CFBE cells presented a higher membrane roughness and decreased rigidity as compared with 16HBE cells. CFBE overexpressing wtCFTR became more elongated than the parental CFBE cell line and presented actin stress fibers. CFBE cells absorbed more fluid from the apical compartment. Study of fluid absorption with the F-actin-depolymerizing agent Latrunculin B demonstrated that actin cytoskeletal disorganization increased fluid absorption, an effect observed at higher magnitude in 16HBE than in CFBE cells. For the first time, we demonstrate that actin cytoskeleton disorganization is reflected by AFM parameters in CF airway epithelial cells. Our data also strongly suggest that the lack of stress fibers is involved in at least one of the early step in CF pathophysiology at the levels of the airways, i.e. fluid hypersorption. - Highlights: • CF bronchial epithelial (CFBE) cells show a disorganized actin cytoskeleton. • CFBE cells present high roughness and low rigidity in
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International Nuclear Information System (INIS)
Lasalvia, Maria; Castellani, Stefano; D’Antonio, Palma; Perna, Giuseppe; Carbone, Annalucia; Colia, Anna Laura; Maffione, Angela Bruna; Capozzi, Vito; Conese, Massimo
2016-01-01
The pathophysiology of cystic fibrosis (CF) airway disease stems from mutations in the CF Transmembrane Conductance Regulator (CFTR) gene, leading to a chronic respiratory disease. Actin cytoskeleton is disorganized in CF airway epithelial cells, likely contributing to the CF-associated basic defects, i.e. defective chloride secretion and sodium/fluid hypersorption. In this work, we aimed to find whether this alteration could be pointed out by means of Atomic Force Microscopy (AFM) investigation, as roughness and Young's elastic module. Moreover, we also sought to determine whether disorganization of actin cytoskeleton is linked to hypersoption of apical fluid. Not only CFBE41o- (CFBE) cells, immortalized airway epithelial cells homozygous for the F508del CFTR allele, showed a different morphology in comparison with 16HBE14o- (16HBE) epithelial cells, wild-type for CFTR, but also they displayed a lack of stress fibers, suggestive of a disorganized actin cytoskeleton. AFM measurements showed that CFBE cells presented a higher membrane roughness and decreased rigidity as compared with 16HBE cells. CFBE overexpressing wtCFTR became more elongated than the parental CFBE cell line and presented actin stress fibers. CFBE cells absorbed more fluid from the apical compartment. Study of fluid absorption with the F-actin-depolymerizing agent Latrunculin B demonstrated that actin cytoskeletal disorganization increased fluid absorption, an effect observed at higher magnitude in 16HBE than in CFBE cells. For the first time, we demonstrate that actin cytoskeleton disorganization is reflected by AFM parameters in CF airway epithelial cells. Our data also strongly suggest that the lack of stress fibers is involved in at least one of the early step in CF pathophysiology at the levels of the airways, i.e. fluid hypersorption. - Highlights: • CF bronchial epithelial (CFBE) cells show a disorganized actin cytoskeleton. • CFBE cells present high roughness and low rigidity in the
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Ramasamy, Thamil Selvee; Velaithan, Vithya; Yeow, Yelena; Sarkar, Fazlul H
2018-01-01
Regenerative medicine aims to provide therapeutic treatment for disease or injury, and cell-based therapy is a newer therapeutic approach different from conventional medicine. Ethical issues that rose by the utilisation of human embryonic stem cells (hESC) and the limited capacity of adult stem cells, however, hinder the application of these stem cells in regenerative medicine. Recently, isolation and characterisation of c-kit positive cells from human amniotic fluid, which possess intermediate characteristics between hESCs and adult stem cells, provided a new approach towards realising their promise for fetal and adult regenerative medicine. Despite the number of studies that have been initiated to characterize their molecular signature, research on developing approaches to maintain and enhance their regenerative potential is urgently needed and must be developed. Thus, this review is focused on understanding their potential uses and factors influencing their pluripotent status in vitro. In short, this cell source could be an ideal cellular resource for pluripotent cells for potential applications in allogeneic cellular replacement therapies, fetal tissue engineering, pharmaceutical screening, and in disease modelling. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.
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Hakai reduces cell-substratum adhesion and increases epithelial cell invasion
International Nuclear Information System (INIS)
Rodríguez-Rigueiro, Teresa; Valladares-Ayerbes, Manuel; Haz-Conde, Mar; Aparicio, Luis A; Figueroa, Angélica
2011-01-01
The dynamic regulation of cell-cell adhesions is crucial for developmental processes, including tissue formation, differentiation and motility. Adherens junctions are important components of the junctional complex between cells and are necessary for maintaining cell homeostasis and normal tissue architecture. E-cadherin is the prototype and best-characterized protein member of adherens junctions in mammalian epithelial cells. Regarded as a tumour suppressor, E-cadherin loss is associated with poor prognosis in carcinoma. The E3 ubiquitin-ligase Hakai was the first reported posttranslational regulator of the E-cadherin complex. Hakai specifically targetted E-cadherin for internalization and degradation and thereby lowered epithelial cell-cell contact. Hakai was also implicated in controlling proliferation, and promoted cancer-related gene expression by increasing the binding of RNA-binding protein PSF to RNAs encoding oncogenic proteins. We sought to investigate the possible implication of Hakai in cell-substratum adhesions and invasion in epithelial cells. Parental MDCK cells and MDCK cells stably overexpressing Hakai were used to analyse cell-substratum adhesion and invasion capabilities. Western blot and immunofluoresecence analyses were performed to assess the roles of Paxillin, FAK and Vinculin in cell-substratum adhesion. The role of the proteasome in controlling cell-substratum adhesion was studied using two proteasome inhibitors, lactacystin and MG132. To study the molecular mechanisms controlling Paxillin expression, MDCK cells expressing E-cadherin shRNA in a tetracycline-inducible manner was employed. Here, we present evidence that implicate Hakai in reducing cell-substratum adhesion and increasing epithelial cell invasion, two hallmark features of cancer progression and metastasis. Paxillin, an important protein component of the cell-matrix adhesion, was completely absent from focal adhesions and focal contacts in Hakai-overexpressing MDCK cells. The
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Kayastha, Forum; Johar, Kaid; Gajjar, Devarshi; Arora, Anshul; Madhu, Hardik; Ganatra, Darshini; Vasavada, Abhay
2015-06-01
Epithelial mesenchymal transition (EMT) of lens epithelial cells (LECs) may contribute to the development of posterior capsular opacification (PCO), which leads to visual impairment. Andrographolide has been shown to have therapeutic potential against various cancers. However, its effect on human LECs is still unknown. The purpose of this study is to evaluate the effect of andrographolide on EMT induced by growth factors in the fetal human lens epithelial cell line (FHL 124). Initially the LECs were treated with growth factors (TGF-beta 2 and bFGF) to induce EMT. Subsequently these EMT-induced cells were treated with andrographolide at 100 and 500 nM concentrations for 24 h. Our results showed that FHL 124 cells treated with growth factors had a significant decrease in protein and m-RNA levels of epithelial markers pax6 and E-Cadherin. After administering andrographolide, these levels significantly increased. It was noticed that EMT markers alpha-SMA, fibronectin and collagen IV significantly decreased after treatment with andrographolide when compared to the other group. Treatment with andrographolide significantly inhibited phosphorylation of ERK and JNK. Cell cycle analysis showed that andrographolide did not arrest cells at G0/G1 or G2/M at tested concentrations. Our findings suggest that andrographolide helps sustain epithelial characteristics by modulating EMT markers and inhibiting the mitogen-activated protein kinase (MAPK) signalling pathway in LECs. Hence it can prove to be useful in curbing EMT-mediated PCO.
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Directory of Open Access Journals (Sweden)
İsmet Durak
2012-05-01
Full Text Available Pur po se: To evaluate the 1-year follow-up results of cultivated limbal epithelial cell (CLEC transplantation in unilateral limbal stem cell deficiency (LSCD. Ma te ri al and Met hod: One-year follow-up results of five unilateral LSCD patients who had undergone CLEC transplantation were evaluated. Parameters for this evaluation were: fluorescein staining of ocular surface, corneal vascularization and status of epithelium with slit lamp, and visual acuity. 1.5-mm limbal biopsy was performed from the superior limbus of the healthy eyes, broke into two equal pieces, expanded on human amniotic membrane (hAM and inserts for 14 days until getting 20 mm in size. CLECs on hAMs were used directly, and cells on inserts were usedafter detachment procedure. The symblepharon and pannus tissues were removed, superficial keratectomy was performed. CLEC on hAMs were transplanted with the epithelial side up onto the bare corneal stroma, sutured to the conjunctiva with 10-0 nylon sutures. Free CLEC layer from insert was placed on hAM as a second layer, additional hAM was used as a protective layer all over other tissues. Re sults: Median age was 44.4 years (14-71. The etiology was chemical burn in all patients. Median duration of symptoms was 10 years (2-18, median follow-up period was 12.6 (12-12.5 months. Preoperative best corrected visual acuities (BCVA were light perception in three patients, counting fingers at 50 cm in one patient and 3/10 in one patient. Visions were improved in all patients. Postoperative BCVA 12 months after the surgery were between counting fingers at 3 meters to 6/10. There was a temporary hemorrhage between the two layers of hAMs in one patient at the early postoperative period. Peripheral corneal vascularization has occurred in three patients, in patient corneal vascularization has reached to the paracentral area. Dis cus si on: CLEC transplantation is an efficient treatment option for unilateral LSCD in mid-long term. (Turk J
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Cigarette smoke suppresses Bik to cause epithelial cell hyperplasia and mucous cell metaplasia.
Mebratu, Yohannes A; Schwalm, Kurt; Smith, Kevin R; Schuyler, Mark; Tesfaigzi, Yohannes
2011-06-01
Aberrant regulation of airway epithelial cell numbers in airways leads to increased mucous secretions in chronic lung diseases such as chronic bronchitis. Because the Bcl-2 family of proteins is crucial for airway epithelial homeostasis, identifying the players that reduce cigarette smoke (CS)-induced mucous cell metaplasia can help to develop effective therapies. To identify the Bcl-2 family of proteins that play a role in reducing CS-induced mucous cell metaplasia. We screened for dysregulated expression of the Bcl-2 family members. We identified Bik to be significantly reduced in bronchial brushings of patients with chronic epithelial cell hyperplasia compared with nondiseased control subjects. Reduced Bik but increased MUC5AC mRNA levels were also detected when normal human airway epithelial cells (HAECs) were exposed to CS or when autopsy tissues from former smokers with and without chronic bronchitis were compared. Similarly, exposure of C57Bl/6 mice to CS resulted in increased numbers of epithelial and mucous cells per millimeter of basal lamina, along with reduced Bik but increased Muc5ac expression, and this change was sustained even when mice were allowed to recover in filtered air for 8 weeks. Restoring Bik expression significantly suppressed CS-induced mucous cell metaplasia in differentiated primary HAEC cultures and in airways of mice in vivo. Bik blocked nuclear translocation of phospho-ERK1/2 to induce apoptosis of HAECs. The conserved Leu61 within Bik and ERK1/2 activation were essential to induce cell death in hyperplastic mucous cells. These studies show that CS suppresses Bik expression to block airway epithelia cell death and thereby increases epithelial cell hyperplasia in chronic bronchitis.
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Epimorphin Functions as a Key Morphoregulator for Mammary Epithelial Cells
Energy Technology Data Exchange (ETDEWEB)
Hirai, H.; Lochter, A.; Galosy, S.; Koshida, S.; Niwa, S.; Bissell, M.J.
1997-10-13
Hepatocyte growth factor (HGF) and EGF have been reported to promote branching morphogenesis of mammary epithelial cells. We now show that it is epimorphin that is primarily responsible for this phenomenon. In vivo, epimorphin was detected in the stromal compartment but not in lumenal epithelial cells of the mammary gland; in culture, however, a subpopulation of mammary epithelial cells produced significant amounts of epimorphin. When epimorphin-expressing epithelial cell clones were cultured in collagen gels they displayed branching morphogenesis in the presence of HGF, EGF, keratinocyte growth factor, or fibroblast growth factor, a process that was inhibited by anti-epimorphin but not anti-HGF antibodies. The branch length, however, was roughly proportional to the ability of the factors to induce growth. Accordingly, epimorphin-negative epithelial cells simply grew in a cluster in response to the growth factors and failed to branch. When recombinant epimorphin was added to these collagen gels, epimorphin-negative cells underwent branching morphogenesis. The mode of action of epimorphin on morphogenesis of the gland, however, was dependent on how it was presented to the mammary cells. If epimorphin was overexpressed in epimorphin-negative epithelial cells under regulation of an inducible promoter or was allowed to coat the surface of each epithelial cell in a nonpolar fashion, the cells formed globular, alveoli-like structures with a large central lumen instead of branching ducts. This process was enhanced also by addition of HGF, EGF, or other growth factors and was inhibited by epimorphin antibodies. These results suggest that epimorphin is the primary morphogen in the mammary gland but that growth factors are necessary to achieve the appropriate cell numbers for the resulting morphogenesis to be visualized.
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Yoshioka, Masahiro; Ohashi, Shinya; Ida, Tomomi; Nakai, Yukie; Kikuchi, Osamu; Amanuma, Yusuke; Matsubara, Junichi; Yamada, Atsushi; Miyamoto, Shin'ichi; Natsuizaka, Mitsuteru; Nakagawa, Hiroshi; Chiba, Tsutomu; Seno, Hiroshi; Muto, Manabu
2017-08-01
Epidermal growth factor receptor (EGFR) plays a pivotal role in the pathophysiology of esophageal squamous cell carcinoma (ESCC). However, the clinical effects of EGFR inhibitors on ESCC are controversial. This study sought to identify the factors determining the therapeutic efficacy of EGFR inhibitors in ESCC cells. Immortalized-human esophageal epithelial cells (EPC2-hTERT), transformed-human esophageal epithelial cells (T-Epi and T-Mes), and ESCC cells (TE-1, TE-5, TE-8, TE-11, TE-11R, and HCE4) were treated with the EGFR inhibitors erlotinib or cetuximab. Inhibitory effects on cell growth were assessed by cell counting or cell-cycle analysis. The expression levels of genes and proteins such as involucrin and cytokeratin13 (a squamous differentiation marker), E-cadherin, and vimentin were evaluated by real-time polymerase chain reaction or western blotting. To examine whether mesenchymal phenotype influenced the effects of EGFR inhibitors, we treated T-Epi cells with TGF-β1 to establish a mesenchymal phenotype (mesenchymal T-Epi cells). We then compared the effects of EGFR inhibitors on parental T-Epi cells and mesenchymal T-Epi cells. TE-8 (mesenchymal-like ESCC cells)- or TE-11R (epithelial-like ESCC cells)-derived xenograft tumors in mice were treated with cetuximab, and the antitumor effects of EGFR inhibitors were evaluated. Cells were classified as epithelial-like or mesenchymal-like phenotypes, determined by the expression levels of E-cadherin and vimentin. Both erlotinib and cetuximab reduced cell growth and the ratio of cells in cell-cycle S phase in epithelial-like but not mesenchymal-like cells. Additionally, EGFR inhibitors induced squamous cell differentiation (defined as increased expression of involucrin and cytokeratin13) in epithelial-like but not mesenchymal-like cells. We found that EGFR inhibitors did not suppress the phosphorylation of EGFR in mesenchymal-like cells, while EGFR dephosphorylation was observed after treatment with EGFR
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Comparison of para-aminophenol cytotoxicity in rat renal epithelial cells and hepatocytes.
Li, Ying; Bentzley, Catherine M; Tarloff, Joan B
2005-04-01
Several chemicals, including para-aminophenol (PAP), produce kidney damage in the absence of hepatic damage. Selective nephrotoxicity may be related to the ability of the kidney to reabsorb filtered water, thereby raising the intraluminal concentration of toxicants and exposing tubular epithelial cells to higher concentrations than would be present in other tissues. The present experiments tested the hypothesis that hepatocytes and renal epithelial cells exposed to equivalent concentrations of PAP would be equally susceptible to toxicity. Hepatocytes and renal epithelial cells were prepared by collagenase digestion of tissues obtained from female Sprague-Dawley rats. Toxicity was monitored using trypan blue exclusion, oxygen consumption and ATP content. We measured the rate of PAP clearance and formation of PAP-glutathione conjugate by HPLC. We found that renal epithelial cells accumulated trypan blue and showed declines in oxygen consumption and ATP content at significantly lower concentrations of PAP and at earlier time points than hepatocytes. The half-life of PAP in hepatocyte incubations was significantly shorter (0.71+/-0.07 h) than in renal epithelial cell incubations (1.33+/-0.23 h), suggesting that renal epithelial cells were exposed to PAP for longer time periods than hepatocytes. Renal epithelial cells formed significantly less glutathione conjugates of PAP (PAP-SG) than did hepatocytes, consistent with less efficient detoxification of reactive PAP intermediates by renal epithelial cells. Finally, hepatocytes contained significant more reduced glutathione (NPSH) than did renal epithelial cells, possibly explaining the enhanced formation of PAP-SG by this cell population. In conclusion, our data indicates that renal epithelial cells are intrinsically more susceptible to PAP cytotoxicity than are hepatocytes. This enhanced cytotoxicity may be due to longer exposure to PAP and/or reduced detoxification of reactive intermediates due to lower concentrations
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Brzeszczynska, J; Samuel, K; Greenhough, S; Ramaesh, K; Dhillon, B; Hay, D C; Ross, J A
2014-06-01
It has been suggested that the isolation of scalable populations of limbal stem cells may lead to radical changes in ocular therapy. In particular, the derivation and transplantation of corneal stem cells from these populations may result in therapies providing clinical normality of the diseased or damaged cornea. Although feasible in theory, the lack of donor material in sufficient quantity and quality currently limits such a strategy. A potential scalable source of corneal cells could be derived from pluripotent stem cells (PSCs). We developed an in vitro and serum-free corneal differentiation model which displays significant promise. Our stepwise differentiation model was designed with reference to development and gave rise to cells which displayed similarities to epithelial progenitor cells which can be specified to cells displaying a corneal epithelial phenotype. We believe our approach is novel, provides a robust model of human development and in the future, may facilitate the generation of corneal epithelial cells that are suitable for clinical use. Additionally, we demonstrate that following continued cell culture, stem cell-derived corneal epithelial cells undergo transdifferentiation and exhibit squamous metaplasia and therefore, also offer an in vitro model of disease.
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Isolation, separation, and characterization of epithelial and connective cells from rat palate
Energy Technology Data Exchange (ETDEWEB)
Terranova, Victor Paul [Univ. of Rochester, NY (United States)
1979-01-01
Epithelial and connective tissue cells were isolated from rat palate by sequential collagenase, hyaluronidase and trypsin digestion of the extracellular matrix. Differences between the two populations were noted with respect to total cell protein, total cell water, proline uptake and incorporation, percent collagen synthesized, effects of parathyroid hormone, metabolism of D-valine and cell density. Basal epithelial cells were subsequently separated from the heterogeneous epithelial cell population on shallow linear density gradients by velocity centrifugation. The type of collagen synthesized by the basal epithelial cells was compared to the type of collagen synthesized by the connective tissue cells by means of labeled amino acid incorporation ratios. Cells isolated from the epithelial and connective tissue were compared. From these studies it can be concluded that epithelial and connective tissue cells can be isolated from rat palate as viable and distinct populations with respect to the biochemical parameters examined. Furthermore, subpopulations can be separated and biochemically characterized.
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Viktorinová, Ivana; Henry, Ian; Tomancak, Pavel
2017-11-01
Symmetry breaking is involved in many developmental processes that form bodies and organs. One of them is the epithelial rotation of developing tubular and acinar organs. However, how epithelial cells move, how they break symmetry to define their common direction, and what function rotational epithelial motions have remains elusive. Here, we identify a dynamic actomyosin network that breaks symmetry at the basal surface of the Drosophila follicle epithelium of acinar-like primitive organs, called egg chambers, and may represent a candidate force-generation mechanism that underlies the unidirectional motion of this epithelial tissue. We provide evidence that the atypical cadherin Fat2, a key planar cell polarity regulator in Drosophila oogenesis, directs and orchestrates transmission of the intracellular actomyosin asymmetry cue onto a tissue plane in order to break planar actomyosin symmetry, facilitate epithelial rotation in the opposite direction, and direct the elongation of follicle cells. In contrast, loss of this rotational motion results in anisotropic non-muscle Myosin II pulses that are disorganized in plane and causes cell deformations in the epithelial tissue of Drosophila eggs. Our work demonstrates that atypical cadherins play an important role in the control of symmetry breaking of cellular mechanics in order to facilitate tissue motion and model epithelial tissue. We propose that their functions may be evolutionarily conserved in tubular/acinar vertebrate organs.
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Amniotic membrane for burn trauma
International Nuclear Information System (INIS)
Jamaluddin Zainol; Hasim Mohammad
1999-01-01
Amniotic membranes are derived from human placentae at birth. They have two layers mainly the amniotic and the chorionic surfaces which are separated by a thin layer of connective tissues. The two layers are separated during procurement, the placenta and the chorionic side are discarded and the amnion membranes are then further processed. Amnion membranes are normally procured from placentae which are normally free of infections, i.e; the mothers are antenatally screened for sexually transmitted diseases or AlDs related diseases. Intrapartum the mother should not be having chorioamnionitis or jaundice. Sometimes the amniotic membranes are acquired from fresh elective caeserian sections. After processing, the amniotic membranes are packed in two layers of polypropylene and radiated with cobalt 60 at a dose of about 25 kGy. The amniotic membranes are clinically used to cover burn surfaces especially effective for superficial or partial thickness burns. The thin membranes adhered well to the trauma areas and peeled off automatically by the second week. No change of dressing were necessary during these times because of the close adherence, there were less chance of external contamination or infections of these wounds. Due to their flexibility they are very useful to cover difference contours of the human body for example the face, body, elbows or knees. However our experience revealed that amniotic membranes are not useful for third degree bums because the membranes dissolves by the enzymes present in the wounds
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Ortega, Fabian E.; Rengarajan, Michelle; Chavez, Natalie; Radhakrishnan, Prathima; Gloerich, Martijn; Bianchini, Julie; Siemers, Kathleen; Luckett, William S.; Lauer, Peter; Nelson, W. James; Theriot, Julie A.
2017-01-01
The intestinal epithelium is the first physiological barrier breached by the Gram-positive facultative pathogen Listeria monocytogenes during an in vivo infection. Listeria monocytogenes binds to the epithelial host cell receptor E-cadherin, which mediates a physical link between the bacterium and filamentous actin (F-actin). However, the importance of anchoring the bacterium to F-actin through E-cadherin for bacterial invasion has not been tested directly in epithelial cells. Here we demonstrate that depleting αE-catenin, which indirectly links E-cadherin to F-actin, did not decrease L. monocytogenes invasion of epithelial cells in tissue culture. Instead, invasion increased due to increased bacterial adhesion to epithelial monolayers with compromised cell–cell junctions. Furthermore, expression of a mutant E-cadherin lacking the intracellular domain was sufficient for efficient L. monocytogenes invasion of epithelial cells. Importantly, direct biotin-mediated binding of bacteria to surface lipids in the plasma membrane of host epithelial cells was sufficient for uptake. Our results indicate that the only requirement for L. monocytogenes invasion of epithelial cells is adhesion to the host cell surface, and that E-cadherin–mediated coupling of the bacterium to F-actin is not required. PMID:28877987
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Ma, Xiaorong; Zhang, Shengli; Zhou, Junmei; Chen, Baisong; Shang, Yafeng; Gao, Tongbing; Wang, Xue; Xie, Hua; Chen, Fang
2012-08-01
Stem cell-based therapy may be the most promising method to cure skeletal muscle degenerative diseases such as Duchenne muscular dystrophy (DMD) and trauma in the future. Human amniotic fluid is enriched with early-stage stem cells from developing fetuses and these cells have cardiomyogenic potential both in vitro and in vivo. In the present study, we investigated the characteristics of human amniotic fluid-derived AF-type stem (HAF-AFS) cells by flow cytometry, immunofluorescence staining, reverse-transcription polymerase chain reaction, and osteogenic and adipogenic differentiation analysis. After confirming the stemness of HAF-AFS cells, we tested whether HAF-AFS cells could differentiate into skeletal myogenic cells in vitro and incorporate into regenerating skeletal muscle in vivo. By temporary exposure to the DNA demethylation agent 5-aza-2'-deoxycytidine (5-Aza dC) or co-cultured with C2C12 myoblasts, HAF-AFS cells differentiated into skeletal myogenic cells, expressing skeletal myogenic cell-specific markers such as Desmin, Troponin I (Tn I) and α-Actinin. Four weeks after transplantation into cardiotoxin-injured and X-ray-irradiated tibialis anterior (TA) muscles of NOD/SCID mice, HAF-AFS cells survived, differentiated into myogenic precursor cells and fused with host myofibres. The findings that HAF-AFS cells differentiate into myogenic cells in vitro and incorporate in skeletal muscle regeneration in vivo hold the promise of HAF-AFS cell-based therapy for skeletal muscle degenerative diseases. Copyright © 2012 John Wiley & Sons, Ltd.
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Type 3 innate lymphoid cells maintain intestinal epithelial stem cells after tissue damage
P. Aparicio-Domingo (Patricia); M. Romera Hernández (Mónica); J.J. Karrich (Julien J.); F.H.J. Cornelissen (Ferry); N. Papazian (Natalie); D.J. Lindenbergh-Kortleve (Dicky); J.A. Butler (James A.); L. Boon (Louis); M. Coles (Mark); J.N. Samsom (Janneke); T. Cupedo (Tom)
2015-01-01
textabstractDisruption of the intestinal epithelial barrier allows bacterial translocation and predisposes to destructive inflammation. To ensure proper barrier composition, crypt-residing stem cells continuously proliferate and replenish all intestinal epithelial cells within days. As a consequence
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Endothelial induced EMT in breast epithelial cells with stem cell properties.
Directory of Open Access Journals (Sweden)
Valgardur Sigurdsson
Full Text Available Epithelial to mesenchymal transition (EMT is a critical event in cancer progression and is closely linked to the breast epithelial cancer stem cell phenotype. Given the close interaction between the vascular endothelium and cancer cells, especially at the invasive front, we asked whether endothelial cells might play a role in EMT. Using a 3D culture model we demonstrate that endothelial cells are potent inducers of EMT in D492 an immortalized breast epithelial cell line with stem cell properties. Endothelial induced mesenchymal-like cells (D492M derived from D492, show reduced expression of keratins, a switch from E-Cadherin (E-Cad to N-Cadherin (N-Cad and enhanced migration. Acquisition of cancer stem cell associated characteristics like increased CD44(high/CD24(low ratio, resistance to apoptosis and anchorage independent growth was also seen in D492M cells. Endothelial induced EMT in D492 was partially blocked by inhibition of HGF signaling. Basal-like breast cancer, a vascular rich cancer with stem cell properties and adverse prognosis has been linked with EMT. We immunostained several basal-like breast cancer samples for endothelial and EMT markers. Cancer cells close to the vascular rich areas show no or decreased expression of E-Cad and increased N-Cad expression suggesting EMT. Collectively, we have shown in a 3D culture model that endothelial cells are potent inducers of EMT in breast epithelial cells with stem cell properties. Furthermore, we demonstrate that basal-like breast cancer contains cells with an EMT phenotype, most prominently close to vascular rich areas of these tumors. We conclude that endothelial cells are potent inducers of EMT and may play a role in progression of basal-like breast cancer.
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Endothelial induced EMT in breast epithelial cells with stem cell properties.
Sigurdsson, Valgardur; Hilmarsdottir, Bylgja; Sigmundsdottir, Hekla; Fridriksdottir, Agla J R; Ringnér, Markus; Villadsen, Rene; Borg, Ake; Agnarsson, Bjarni A; Petersen, Ole William; Magnusson, Magnus K; Gudjonsson, Thorarinn
2011-01-01
Epithelial to mesenchymal transition (EMT) is a critical event in cancer progression and is closely linked to the breast epithelial cancer stem cell phenotype. Given the close interaction between the vascular endothelium and cancer cells, especially at the invasive front, we asked whether endothelial cells might play a role in EMT. Using a 3D culture model we demonstrate that endothelial cells are potent inducers of EMT in D492 an immortalized breast epithelial cell line with stem cell properties. Endothelial induced mesenchymal-like cells (D492M) derived from D492, show reduced expression of keratins, a switch from E-Cadherin (E-Cad) to N-Cadherin (N-Cad) and enhanced migration. Acquisition of cancer stem cell associated characteristics like increased CD44(high)/CD24(low) ratio, resistance to apoptosis and anchorage independent growth was also seen in D492M cells. Endothelial induced EMT in D492 was partially blocked by inhibition of HGF signaling. Basal-like breast cancer, a vascular rich cancer with stem cell properties and adverse prognosis has been linked with EMT. We immunostained several basal-like breast cancer samples for endothelial and EMT markers. Cancer cells close to the vascular rich areas show no or decreased expression of E-Cad and increased N-Cad expression suggesting EMT. Collectively, we have shown in a 3D culture model that endothelial cells are potent inducers of EMT in breast epithelial cells with stem cell properties. Furthermore, we demonstrate that basal-like breast cancer contains cells with an EMT phenotype, most prominently close to vascular rich areas of these tumors. We conclude that endothelial cells are potent inducers of EMT and may play a role in progression of basal-like breast cancer.
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Intestinal Epithelial Cells Modulate Antigen-Presenting Cell Responses to Bacterial DNA
Campeau, J. L.; Salim, S. Y.; Albert, E. J.; Hotte, N.
2012-01-01
Intestinal epithelial cells and antigen-presenting cells orchestrate mucosal innate immunity. This study investigated the role of bacterial DNA in modulating epithelial and bone marrow-derived antigen-presenting cells (BM-APCs) and subsequent T-lymphocyte responses. Murine MODE-K epithelial cells and BM-APCs were treated with DNA from either Bifidobacterium breve or Salmonella enterica serovar Dublin directly and under coculture conditions with CD4+ T cells. Apical stimulation of MODE-K cells with S. Dublin DNA enhanced secretion of cytokines from underlying BM-APCs and induced interleukin-17 (IL-17) and gamma interferon (IFN-γ) secretion from CD4+ T cells. Bacterial DNA isolated from either strain induced maturation and increased cytokine secretion from BM-APCs. Conditioned medium from S. Dublin-treated MODE-K cells elicited an increase in cytokine secretion similar to that seen for S. Dublin DNA. Treatment of conditioned medium from MODE-K cells with RNase and protease prevented the S. Dublin-induced increased cytokine secretion. Oral feeding of mice with B. breve DNA resulted in enhanced levels of colonic IL-10 and transforming growth factor β (TGFβ) compared with what was seen for mice treated with S. Dublin DNA. In contrast, feeding mice with S. Dublin DNA increased levels of colonic IL-17 and IL-12p70. T cells from S. Dublin DNA-treated mice secreted high levels of IL-12 and IFN-γ compared to controls and B. breve DNA-treated mice. These results demonstrate that intestinal epithelial cells are able to modulate subsequent antigen-presenting and T-cell responses to bacterial DNA with pathogenic but not commensal bacterial DNA inducing effector CD4+ T lymphocytes. PMID:22615241
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RESEARCH ON REDUCING PREMATURITY RUPTURE OF MEMBRANE
Maria URSACHI (BOLOTA); Emil ANTON; Sorana Caterina ANTON
2016-01-01
The membranes surrounding the amniotic cavity are composed from amnion and chorion, tightly adherent layers which are composed of several cell types, including epithelial cells, trophoblasts cells and mesenchyme cells, embedded in a collagenous matrix. They retain amniotic fluid, secret substances into the amniotic fluid, as well as to the uterus and protect the fetus against upward infections from urogenital tract. Normally, the membranes it breaks during labor. Premature rupture of the amn...
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Mas-Oliva, Jaime; Navarro-Vidal, Enrique; Tapia-Vieyra, Juana Virginia
2014-01-01
Neoplastic epithelial cells generate the most aggressive types of cancers such as those located in the lung, breast, colon, prostate and ovary. During advanced stages of prostate cancer, epithelial cells are associated to the appearance of androgen-independent tumors, an apoptotic-resistant phenotype that ultimately overgrows and promotes metastatic events. We have previously identified and electrophysiologically characterized a novel Ca(2+)-permeable channel activated during apoptosis in the androgen-independent prostate epithelial cancer cell line, LNCaP. In addition, we reported for the first time the cloning and characterization of this channel-like molecule named apoptosis regulated protein 2 (ARP2) associated to a lethal influx of Ca(2+) in Xenopus oocytes. In the present study, LNCaP cells and Chinese hamster ovary cells (CHO cell line) transfected with arp2-cDNA are induced to undergo apoptosis showing an important impact on cell viability and activation of caspases 3 and 7 when compared to serum deprived grown cells and ionomycin treated cells. The subcellular localization of ARP2 in CHO cells undergoing apoptosis was studied using confocal microscopy. While apoptosis progresses, ARP2 initially localized in the peri-nuclear region of cells migrates with time towards the plasma membrane region. Based on the present results and those of our previous studies, the fact that ARP2 constitutes a novel cation channel is supported. Therefore, ARP2 becomes a valuable target to modulate the influx and concentration of calcium in the cytoplasm of epithelial cancer cells showing an apoptotic-resistant phenotype during the onset of an apoptotic event.
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induced acute cytotoxicity in human cervical epithelial carcinoma cells
African Journals Online (AJOL)
Molecular basis of arsenite (As +3 )-induced acute cytotoxicity in human cervical epithelial carcinoma cells. ... Libyan Journal of Medicine ... Methods: After performing cytotoxic assays on a human epithelial carcinoma cell line, expression analysis was done by quantitative polymerase chain reaction, western blotting, and ...
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Directory of Open Access Journals (Sweden)
Shuxian Jiang
2010-03-01
Full Text Available Mammary stem cells are maintained within specific microenvironments and recruited throughout lifetime to reconstitute de novo the mammary gland. Mammary stem cells have been isolated through the identification of specific cell surface markers and in vivo transplantation into cleared mammary fat pads. Accumulating evidence showed that during the reformation of mammary stem cell niches by dispersed epithelial cells in the context of the intact epithelium-free mammary stroma, non-mammary epithelial cells may be sequestered and reprogrammed to perform mammary epithelial cell functions and to adopt mammary epithelial characteristics during reconstruction of mammary epithelium in regenerating mammary tissue in vivo.To examine whether other types of progenitor cells are able to contribute to mammary branching morphogenesis, we examined the potential of murine embryonic stem (mES cells, undergoing hematopoietic differentiation, to support mammary reconstitution in vivo. We observed that cells from day 14 embryoid bodies (EBs under hematopoietic differentiation condition, but not supernatants derived from these cells, when transplanted into denuded mammary fat pads, were able to contribute to both the luminal and myoepithelial lineages in branching ductal structures resembling the ductal-alveolar architecture of the mammary tree. No teratomas were observed when these cells were transplanted in vivo.Our data provide evidence for the dominance of the tissue-specific mammary stem cell niche and its role in directing mES cells, undergoing hematopoietic differentiation, to reprogram into mammary epithelial cells and to promote mammary epithelial morphogenesis. These studies should also provide insights into regeneration of damaged mammary gland and the role of the mammary microenvironment in reprogramming cell fate.
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Coronavirus infection of polarized epithelial cells
Rossen, J W; Horzinek, M C; Rottier, P J
1995-01-01
Epithelial cells are the first host cells to be infected by incoming c oronaviruses. Recent observations in vitro show that coronaviruses are released from a specific side of these polarized cells, and this polarized release might be important for the spread of the infection in vivo. Mechanisms for
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Uranium induces oxidative stress in lung epithelial cells
International Nuclear Information System (INIS)
Periyakaruppan, Adaikkappan; Kumar, Felix; Sarkar, Shubhashish; Sharma, Chidananda S.; Ramesh, Govindarajan T.
2007-01-01
Uranium compounds are widely used in the nuclear fuel cycle, antitank weapons, tank armor, and also as a pigment to color ceramics and glass. Effective management of waste uranium compounds is necessary to prevent exposure to avoid adverse health effects on the population. Health risks associated with uranium exposure includes kidney disease and respiratory disorders. In addition, several published results have shown uranium or depleted uranium causes DNA damage, mutagenicity, cancer and neurological defects. In the current study, uranium toxicity was evaluated in rat lung epithelial cells. The study shows uranium induces significant oxidative stress in rat lung epithelial cells followed by concomitant decrease in the antioxidant potential of the cells. Treatment with uranium to rat lung epithelial cells also decreased cell proliferation after 72 h in culture. The decrease in cell proliferation was attributed to loss of total glutathione and superoxide dismutase in the presence of uranium. Thus the results indicate the ineffectiveness of antioxidant system's response to the oxidative stress induced by uranium in the cells. (orig.)
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O'Beirne, Sarah L; Walsh, Sinead M; Fabre, Aurélie; Reviriego, Carlota; Worrell, Julie C; Counihan, Ian P; Lumsden, Robert V; Cramton-Barnes, Jennifer; Belperio, John A; Donnelly, Seamas C; Boylan, Denise; Marchal-Sommé, Joëlle; Kane, Rosemary; Keane, Michael P
2015-09-15
Epithelial to mesenchymal cell transition (EMT), whereby fully differentiated epithelial cells transition to a mesenchymal phenotype, has been implicated in the pathogenesis of idiopathic pulmonary fibrosis (IPF). CXCR3 and its ligands are recognized to play a protective role in pulmonary fibrosis. In this study, we investigated the presence and extent of EMT and CXCR3 expression in human IPF surgical lung biopsies and assessed whether CXCR3 and its ligand CXCL9 modulate EMT in alveolar epithelial cells. Coexpression of the epithelial marker thyroid transcription factor-1 and the mesenchymal marker α-smooth muscle actin and CXCR3 expression was examined by immunohistochemical staining of IPF surgical lung biopsies. Epithelial and mesenchymal marker expression was examined by quantitative real-time PCR, Western blotting, and immunofluorescence in human alveolar epithelial (A549) cells treated with TGF-β1 and CXCL9, with Smad2, Smad3, and Smad7 expression and cellular localization examined by Western blotting. We found that significantly more cells were undergoing EMT in fibrotic versus normal areas of lung in IPF surgical lung biopsy samples. CXCR3 was expressed by type II pneumocytes and fibroblasts in fibrotic areas in close proximity to cells undergoing EMT. In vitro, CXCL9 abrogated TGF-β1-induced EMT. A decrease in TGF-β1-induced phosphorylation of Smad2 and Smad3 occurred with CXCL9 treatment. This was associated with increased shuttling of Smad7 from the nucleus to the cytoplasm where it inhibits Smad phosphorylation. This suggests a role for EMT in the pathogenesis of IPF and provides a novel mechanism for the inhibitory effects of CXCL9 on TGF-β1-induced EMT. Copyright © 2015 by The American Association of Immunologists, Inc.
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Contraction and elongation: Mechanics underlying cell boundary deformations in epithelial tissue.
Hara, Yusuke
2017-06-01
The cell-cell boundaries of epithelial cells form cellular frameworks at the apical side of tissues. Deformations in these boundaries, for example, boundary contraction and elongation, and the associated forces form the mechanical basis of epithelial tissue morphogenesis. In this review, using data from recent Drosophila studies on cell boundary contraction and elongation, I provide an overview of the mechanism underlying the bi-directional deformations in the epithelial cell boundary, that are sustained by biased accumulations of junctional and apico-medial non-muscle myosin II. Moreover, how the junctional tensions exist on cell boundaries in different boundary dynamics and morphologies are discussed. Finally, some future perspectives on how recent knowledge about single cell boundary-level mechanics will contribute to our understanding of epithelial tissue morphogenesis are discussed. © 2017 Japanese Society of Developmental Biologists.
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Quantification of epithelial cells in coculture with fibroblasts by fluorescence image analysis.
Krtolica, Ana; Ortiz de Solorzano, Carlos; Lockett, Stephen; Campisi, Judith
2002-10-01
To demonstrate that senescent fibroblasts stimulate the proliferation and neoplastic transformation of premalignant epithelial cells (Krtolica et al.: Proc Natl Acad Sci USA 98:12072-12077, 2001), we developed methods to quantify the proliferation of epithelial cells cocultured with fibroblasts. We stained epithelial-fibroblast cocultures with the fluorescent DNA-intercalating dye 4,6-diamidino-2-phenylindole (DAPI), or expressed green fluorescent protein (GFP) in the epithelial cells, and then cultured them with fibroblasts. The cocultures were photographed under an inverted microscope with appropriate filters, and the fluorescent images were captured with a digital camera. We modified an image analysis program to selectively recognize the smaller, more intensely fluorescent epithelial cell nuclei in DAPI-stained cultures and used the program to quantify areas with DAPI fluorescence generated by epithelial nuclei or GFP fluorescence generated by epithelial cells in each field. Analysis of the image areas with DAPI and GFP fluorescences produced nearly identical quantification of epithelial cells in coculture with fibroblasts. We confirmed these results by manual counting. In addition, GFP labeling permitted kinetic studies of the same coculture over multiple time points. The image analysis-based quantification method we describe here is an easy and reliable way to monitor cells in coculture and should be useful for a variety of cell biological studies. Copyright 2002 Wiley-Liss, Inc.
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DA-6034 Induces [Ca(2+)]i Increase in Epithelial Cells.
Yang, Yu-Mi; Park, Soonhong; Ji, Hyewon; Kim, Tae-Im; Kim, Eung Kweon; Kang, Kyung Koo; Shin, Dong Min
2014-04-01
DA-6034, a eupatilin derivative of flavonoid, has shown potent effects on the protection of gastric mucosa and induced the increases in fluid and glycoprotein secretion in human and rat corneal and conjunctival cells, suggesting that it might be considered as a drug for the treatment of dry eye. However, whether DA-6034 induces Ca(2+) signaling and its underlying mechanism in epithelial cells are not known. In the present study, we investigated the mechanism for actions of DA-6034 in Ca(2+) signaling pathways of the epithelial cells (conjunctival and corneal cells) from human donor eyes and mouse salivary gland epithelial cells. DA-6034 activated Ca(2+)-activated Cl(-) channels (CaCCs) and increased intracellular calcium concentrations ([Ca(2+)]i) in primary cultured human conjunctival cells. DA-6034 also increased [Ca(2+)]i in mouse salivary gland cells and human corneal epithelial cells. [Ca(2+)]i increase of DA-6034 was dependent on the Ca(2+) entry from extracellular and Ca(2+) release from internal Ca(2+) stores. Interestingly, these effects of DA-6034 were related to ryanodine receptors (RyRs) but not phospholipase C/inositol 1,4,5-triphosphate (IP3) pathway and lysosomal Ca(2+) stores. These results suggest that DA-6034 induces Ca(2+) signaling via extracellular Ca(2+) entry and RyRs-sensitive Ca(2+) release from internal Ca(2+) stores in epithelial cells.
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Directory of Open Access Journals (Sweden)
Alexandra Mikhailova
2014-02-01
Full Text Available Human induced pluripotent stem cells (hiPSCs offer unique opportunities for developing novel cell-based therapies and disease modeling. In this study, we developed a directed differentiation method for hiPSCs toward corneal epithelial progenitor cells capable of terminal differentiation toward mature corneal epithelial-like cells. In order to improve the efficiency and reproducibility of our method, we replicated signaling cues active during ocular surface ectoderm development with the help of two small-molecule inhibitors in combination with basic fibroblast growth factor (bFGF in serum-free and feeder-free conditions. First, small-molecule induction downregulated the expression of pluripotency markers while upregulating several transcription factors essential for normal eye development. Second, protein expression of the corneal epithelial progenitor marker p63 was greatly enhanced, with up to 95% of cells being p63 positive after 5 weeks of differentiation. Third, corneal epithelial-like cells were obtained upon further maturation.
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Pancreatic stellate cells promote epithelial-mesenchymal transition in pancreatic cancer cells
International Nuclear Information System (INIS)
Kikuta, Kazuhiro; Masamune, Atsushi; Watanabe, Takashi; Ariga, Hiroyuki; Itoh, Hiromichi; Hamada, Shin; Satoh, Kennichi; Egawa, Shinichi; Unno, Michiaki; Shimosegawa, Tooru
2010-01-01
Research highlights: → Recent studies have shown that pancreatic stellate cells (PSCs) promote the progression of pancreatic cancer. → Pancreatic cancer cells co-cultured with PSCs showed loose cell contacts and scattered, fibroblast-like appearance. → PSCs decreased the expression of epithelial markers but increased that of mesenchymal markers, along with increased migration. → This study suggests epithelial-mesenchymal transition as a novel mechanism by which PSCs contribute to the aggressive behavior of pancreatic cancer cells. -- Abstract: The interaction between pancreatic cancer cells and pancreatic stellate cells (PSCs), a major profibrogenic cell type in the pancreas, is receiving increasing attention. There is accumulating evidence that PSCs promote the progression of pancreatic cancer by increasing cancer cell proliferation and invasion as well as by protecting them from radiation- and gemcitabine-induced apoptosis. Because epithelial-mesenchymal transition (EMT) plays a critical role in the progression of pancreatic cancer, we hypothesized that PSCs promote EMT in pancreatic cancer cells. Panc-1 and SUIT-2 pancreatic cancer cells were indirectly co-cultured with human PSCs isolated from patients undergoing operation for pancreatic cancer. The expression of epithelial and mesenchymal markers was examined by real-time PCR and immunofluorescent staining. The migration of pancreatic cancer cells was examined by scratch and two-chamber assays. Pancreatic cancer cells co-cultured with PSCs showed loose cell contacts and a scattered, fibroblast-like appearance. The expression of E-cadherin, cytokeratin 19, and membrane-associated β-catenin was decreased, whereas vimentin and Snail (Snai-1) expression was increased more in cancer cells co-cultured with PSCs than in mono-cultured cells. The migration of pancreatic cancer cells was increased by co-culture with PSCs. The PSC-induced decrease of E-cadherin expression was not altered by treatment with anti
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Point mutations in EBV gH that abrogate or differentially affect B cell and epithelial cell fusion
International Nuclear Information System (INIS)
Wu Liguo; Hutt-Fletcher, Lindsey M.
2007-01-01
Cell fusion mediated by Epstein-Barr virus requires three conserved glycoproteins, gB and gHgL, but activation is cell type specific. B cell fusion requires interaction between MHC class II and a fourth virus glycoprotein, gp42, which complexes non-covalently with gHgL. Epithelial cell fusion requires interaction between gHgL and a novel epithelial cell coreceptor and is blocked by excess gp42. We show here that gp42 interacts directly with gH and that point mutations in the region of gH recognized by an antibody that differentially inhibits epithelial and B cell fusion significantly impact both the core fusion machinery and cell-specific events. Substitution of alanine for glycine at residue 594 completely abrogates fusion with either B cells or epithelial cells. Substitution of alanine for glutamic acid at residue 595 reduces fusion with epithelial cells, greatly enhances fusion with B cells and allows low levels of B cell fusion even in the absence of gL
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Amniotic fluid inflammatory cytokines
DEFF Research Database (Denmark)
Abdallah, Morsi; Larsen, Nanna; Grove, Jakob
2013-01-01
The aim of the study was to analyze cytokine profiles in amniotic fluid (AF) samples of children developing autism spectrum disorders (ASD) and controls, adjusting for maternal autoimmune disorders and maternal infections during pregnancy.......The aim of the study was to analyze cytokine profiles in amniotic fluid (AF) samples of children developing autism spectrum disorders (ASD) and controls, adjusting for maternal autoimmune disorders and maternal infections during pregnancy....
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Nicotine transport in lung and non-lung epithelial cells.
Takano, Mikihisa; Kamei, Hidetaka; Nagahiro, Machi; Kawami, Masashi; Yumoto, Ryoko
2017-11-01
Nicotine is rapidly absorbed from the lung alveoli into systemic circulation during cigarette smoking. However, mechanism underlying nicotine transport in alveolar epithelial cells is not well understood to date. In the present study, we characterized nicotine uptake in lung epithelial cell lines A549 and NCI-H441 and in non-lung epithelial cell lines HepG2 and MCF-7. Characteristics of [ 3 H]nicotine uptake was studied using these cell lines. Nicotine uptake in A549 cells occurred in a time- and temperature-dependent manner and showed saturation kinetics, with a Km value of 0.31mM. Treatment with some organic cations such as diphenhydramine and pyrilamine inhibited nicotine uptake, whereas treatment with organic cations such as carnitine and tetraethylammonium did not affect nicotine uptake. Extracellular pH markedly affected nicotine uptake, with high nicotine uptake being observed at high pH up to 11.0. Modulation of intracellular pH with ammonium chloride also affected nicotine uptake. Treatment with valinomycin, a potassium ionophore, did not significantly affect nicotine uptake, indicating that nicotine uptake is an electroneutral process. For comparison, we assessed the characteristics of nicotine uptake in another lung epithelial cell line NCI-H441 and in non-lung epithelial cell lines HepG2 and MCF-7. Interestingly, these cell lines showed similar characteristics of nicotine uptake with respect to pH dependency and inhibition by various organic cations. The present findings suggest that a similar or the same pH-dependent transport system is involved in nicotine uptake in these cell lines. A novel molecular mechanism of nicotine transport is proposed. Copyright © 2017 Elsevier Inc. All rights reserved.
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Melanin dependent survival of Apergillus fumigatus conidia in lung epithelial cells.
Amin, Shayista; Thywissen, Andreas; Heinekamp, Thorsten; Saluz, Hans Peter; Brakhage, Axel A
2014-07-01
Aspergillus fumigatus is the most important air-borne pathogenic fungus of humans. Upon inhalation of conidia, the fungus makes close contact with lung epithelial cells, which only possess low phagocytic activity. These cells are in particular interesting to address the question whether there is some form of persistence of conidia of A. fumigatus in the human host. Therefore, by also using uracil-auxotrophic mutant strains, we were able to investigate the interaction of A549 lung epithelial cells and A. fumigatus conidia in detail for long periods. Interestingly, unlike professional phagocytes, our study showed that the presence of conidial dihydroxynaphthalene (DHN) melanin enhanced the uptake of A. fumigatus conidia by epithelial cells when compared with non-pigmented pksP mutant conidia. Furthermore, conidia of A. fumigatus were able to survive within epithelial cells. This was due to the presence of DHN melanin in the cell wall of conidia, because melanised wild-type conidia showed a higher survival rate inside epithelial cells and led to inhibition of acidification of phagolysosomes. Both effects were not observed for white (non-melanised) conidia of the pksP mutant strain. Moreover, in contrast to pksP mutant conidia, melanised wild-type conidia were able to inhibit the extrinsic apoptotic pathway in A549 lung epithelial cells even for longer periods. The anti-apoptotic effect was not restricted to conidia, because both conidia-derived melanin ghosts (cell-free DHN melanin) and a different type of melanin, dihydroxyphenylalanine (DOPA) melanin, acted anti-apoptotically. Taken together, these data indicate the possibility of melanin-dependent persistence of conidia in lung epithelial cells. Copyright © 2014 Elsevier GmbH. All rights reserved.
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Interaction of the pathogenic mold Aspergillus fumigatus with lung epithelial cells
Directory of Open Access Journals (Sweden)
Nir eOsherov
2012-09-01
Full Text Available Aspergillus fumigatus is an opportunistic environmental mold that can cause severe allergic responses in atopic individuals and poses a life-threatening risk for severely immunocompromised patients. Infection is caused by inhalation of fungal spores (conidia into the lungs. The initial point of contact between the fungus and the host is a monolayer of lung epithelial cells. Understanding how these cells react to fungal contact is crucial to elucidating the pathobiology of Aspergillus-related disease states. The experimental systems, both in vitro and in vivo, used to study these interactions, are described. Distinction is made between bronchial and alveolar epithelial cells. The experimental findings suggest that lung epithelial cells are more than just innocent bystanders or a purely physical barrier against infection. They can be better described as an active extension of our innate immune system, operating as a surveillance mechanism that can specifically identify fungal spores and activate an offensive response to block infection. This response includes the internalization of adherent conidia and the release of cytokines, antimicrobial peptides and reactive oxygen species. In the case of allergy, lung epithelial cells can dampen an over-reactive immune response by releasing anti-inflammatory compounds such as kinurenine. This review summarizes our current knowledge regarding the interaction of A. fumigatus with lung epithelial cells. A better understanding of the interactions between A. fumigatus and lung epithelial cells has therapeutic implications, as stimulation or inhibition of the epithelial response may alter disease outcome.
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Application of Amniotic Membrane in Ocular Surface Diseases: Clinical Features and Treatment Outcome
Directory of Open Access Journals (Sweden)
Derya Cindarik
2012-05-01
Full Text Available Pur po se: To investigate the effectiveness of amniotic membrane transplantation in cases with corneal thinning, desmatocele and refractive corneal ulcer. Ma te ri al and Met hod: Fifty-four eyes of 54 patients who were applied amniotic membrane transplantation for various ocular surface disease between January 2004 and February 2009 in Çukurova University Ophthalmology Department were included in the study. A complete ophthalmologic examination was performed. Corneal culture and corneal cytology samples were collected from the patients with the diagnosis of corneal ulcers. The patients were informed about the surgical procedure and the possible complications and informed consent was obtained. The amniotic membranes that were prepared under optimal conditions and protected in frozen forms were used in the operations. Follow-up examinations were done at postoperative 1st day, 1st week, 1st month, 3rd month, 6th month and then once in a year. Re sults: Of 54 patients, 26 (48.1% were men and 28 (51.8% were women. The mean age of patients was 52.53±19.75 (2-87 years. The cases were separated into 2 groups according to the etiology: group 1 - eyes with corneal ulcer (n:26 and group 2 - eyes with corneal stromal thinning, persistent epithelial defects and desmatocel (n:28. The transplantations were performed using cover technique in 17 eyes (31.4%, graft technique in 37 eyes (68.5% and graft technique with corneal patch in 2 eyes (3.7%. Partial penetrating keratoplasty was required in 38 of 54 eyes (70.3%. One eye was enucleated. Dis cus si on: The amniotic membrane transplantation has advantages like: it can be prepared easily and is cost-effective. It is a safe and effective procedure in ocular surface disease. (Turk J Ophthalmol 2012; 42: 177-82
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Amniotic membrane extract ameliorates benzalkonium chloride-induced dry eye in a murine model.
Xiao, Xinye; Luo, Pingping; Zhao, Hui; Chen, Jingyao; He, Hui; Xu, Yuxue; Lin, Zhirong; Zhou, Yueping; Xu, Jianjiang; Liu, Zuguo
2013-10-01
Human amniotic membrane (AM) is avascular but contains various beneficial bioactive factors, its extract (AE) is also effective in treating many ocular surface disorders. In this study, we for the first time evaluated the therapeutic effects of AE on dry eye induced by benzalkonium chloride in a BALB/c mouse model. Topical application of AE (1.5 and 3 μg/eye/day) resulted in significantly longer tear break-up time on Day 3 and 6, lower fluorescein staining scores on Day 3, and lower inflammatory index on Day 6. AE reduced corneal epithelial K10 expression, inflammatory infiltration, and levels of TNF-α, IL-1β and IL-6 in BAC treated mice than that in the control mice. Moreover, decreased TUNEL positive cells in cornea and increased goblet cells in conjunctiva were also observed in AE treated corneas. Finally, AE induced more Ki-67 positive cells in corneal epithelium of dry eye mouse. Taken together, our data provide further support for BAC induced dry eye model as a valuable for dry eye study and suggest a great potential for AE as a therapeutic agent in the clinical treatment of dry eye. Copyright © 2013 Elsevier Ltd. All rights reserved.
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Sweat JMDunigan, D D; Wright, S D
2001-06-01
The West-Indian manatee, Trichechus manatus latirostris, is a herbivorous marine mammal found in the coastal waters of Florida. Because of their endangered status, animal experimentation is not allowed. Therefore, a cell line was developed and characterized from tissue collected during necropsies of the manatees. A primary cell culture was established by isolating single cells from kidney tissue using both enzymatic and mechanical techniques. Primary manatee kidney (MK) cells were subcultured for characterization. These cells were morphologically similar to the cell lines of epithelial origin. An immunocytochemistry assay was used to localize the cytokeratin filaments common to cells of epithelial origin. At second passage, epithelial-like cells had an average population-doubling time of 48 h, had an optimum seeding density of 5 x 10(3) cells/cm2, and readily attached to plastic culture plates with a high level of seeding efficiency. Although the epithelial-like cells had a rapid growth rate during the first three passages, the cloning potential was low. These cells did not form colonies in agar medium, were serum dependent, had a limited life span of approximately nine passages, and possessed cell-contact inhibition. These data suggest that the cells were finite (noncontinuous growth), did not possess transformed properties, and were of epithelial origin. These cells are now referred to as MK epithelial cells.
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Interaction of chitin/chitosan with salivary and other epithelial cells-An overview.
Patil, Sharvari Vijaykumar; Nanduri, Lalitha S Y
2017-11-01
Chitin and its deacetylated form, chitosan, have been widely used for tissue engineering of both epithelial and mesenchymal tissues. Epithelial cells characterised by their sheet-like tight cellular arrangement and polarised nature, constitute a major component in various organs and play a variety of roles including protection, secretion and maintenance of tissue homeostasis. Regeneration of damaged epithelial tissues has been studied using biomaterials such as chitin, chitosan, hyaluronan, gelatin and alginate. Chitin and chitosan are known to promote proliferation of various embryonic and adult epithelial cells. However it is not clearly understood how this activity is achieved or what are the mechanisms involved in the chitin/chitosan driven proliferation of epithelial cells. Mechanistic understanding of influence of chitin/chitosan on epithelial cells will guide us to develop more targeted regenerative scaffold/hydrogel systems. Therefore, current review attempts to elicit a mechanistic insight into how chitin and chitosan interact with salivary, mammary, skin, nasal, lung, intestinal and bladder epithelial cells. Copyright © 2017 Elsevier B.V. All rights reserved.
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Roles of Wnt/β-catenin signaling in epithelial differentiation of mesenchymal stem cells
International Nuclear Information System (INIS)
Wang, Yajing; Sun, Zhaorui; Qiu, Xuefeng; Li, Yan; Qin, Jizheng; Han, Xiaodong
2009-01-01
Bone marrow-derived mesenchymal stem cells (MSCs) have been demonstrated to be able to differentiate into epithelial lineage, but the precise mechanisms controlling this process are unclear. Our aim is to explore the roles of Wnt/β-catenin in the epithelial differentiation of MSCs. Using indirect co-culture of rat MSCs with rat airway epithelial cells (RTE), MSCs expressed several airway epithelial markers (cytokeratin 18, tight junction protein occudin, cystic fibrosis transmembrance regulator). The protein levels of some important members in Wnt/β-catenin signaling were determined, suggested down-regulation of Wnt/β-catenin with epithelial differentiation of MSCs. Furthermore, Wnt3α can inhibit the epithelial differentiation of MSCs. A loss of β-catenin induced by Dickkopf-1 can enhance MSCs differentiation into epithelial cells. Lithium chloride transiently activated β-catenin expression and subsequently decreased β-catenin level and at last inhibited MSCs to differentiate into airway epithelium. Taken together, our study indicated that RTE cells can trigger epithelial differentiation of MSCs. Blocking Wnt/β-catenin signaling may promote MSCs to differentiate towards airway epithelial cells.
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Stem cell factor expression after renal ischemia promotes tubular epithelial survival.
Directory of Open Access Journals (Sweden)
Geurt Stokman
Full Text Available BACKGROUND: Renal ischemia leads to apoptosis of tubular epithelial cells and results in decreased renal function. Tissue repair involves re-epithelialization of the tubular basement membrane. Survival of the tubular epithelium following ischemia is therefore important in the successful regeneration of renal tissue. The cytokine stem cell factor (SCF has been shown to protect the tubular epithelium against apoptosis. METHODOLOGY/PRINCIPAL FINDINGS: In a mouse model for renal ischemia/reperfusion injury, we studied how expression of c-KIT on tubular epithelium and its ligand SCF protect cells against apoptosis. Administration of SCF specific antisense oligonucleotides significantly decreased specific staining of SCF following ischemia. Reduced SCF expression resulted in impaired renal function, increased tubular damage and increased tubular epithelial apoptosis, independent of inflammation. In an in vitro hypoxia model, stimulation of tubular epithelial cells with SCF activated survival signaling and decreased apoptosis. CONCLUSIONS/SIGNIFICANCE: Our data indicate an important role for c-KIT and SCF in mediating tubular epithelial cell survival via an autocrine pathway.
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Fibrosis of Two: Epithelial Cell-Fibroblast Interactions in Pulmonary Fibrosis
Sakai, Norihiko; Tager, Andrew M.
2013-01-01
Idiopathic pulmonary fibrosis (IPF) is characterized by the progressive and ultimately fatal accumulation of fibroblasts and extracellular matrix in the lung that distorts its architecture and compromises its function. IPF is now thought to result from wound-healing processes that, although initiated to protect the host from injurious environmental stimuli, lead to pathological fibrosis due to these processes becoming aberrant or over-exuberant. Although the environmental stimuli that trigger IPF remain to be identified, recent evidence suggests that they initially injure the alveolar epithelium. Repetitive cycles of epithelial injury and resultant alveolar epithelial cell death provoke the migration, proliferation, activation and myofibroblast differentiation of fibroblasts, causing the accumulation of these cells and the extracellular matrix that they synthesize. In turn, these activated fibroblasts induce further alveolar epithelial cell injury and death, thereby creating a vicious cycle of pro-fibrotic epithelial cell-fibroblast interactions. Though other cell types certainly make important contributions, we focus here on the “pas de deux” (steps of two), or perhaps more appropriate to IPF pathogenesis, the “folie à deux” (madness of two) of epithelial cells and fibroblasts that drives the progression of pulmonary fibrosis. We describe the signaling molecules that mediate the interactions of these cell types in their “fibrosis of two”, including transforming growth factor-β, connective tissue growth factor, sonic hedgehog, prostaglandin E2, angiotensin II and reactive oxygen species. PMID:23499992
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Breast fibroblasts modulate epithelial cell proliferation in three-dimensional in vitro co-culture
International Nuclear Information System (INIS)
Sadlonova, Andrea; Novak, Zdenek; Johnson, Martin R; Bowe, Damon B; Gault, Sandra R; Page, Grier P; Thottassery, Jaideep V; Welch, Danny R; Frost, Andra R
2005-01-01
Stromal fibroblasts associated with in situ and invasive breast carcinoma differ phenotypically from fibroblasts associated with normal breast epithelium, and these alterations in carcinoma-associated fibroblasts (CAF) may promote breast carcinogenesis and cancer progression. A better understanding of the changes that occur in fibroblasts during carcinogenesis and their influence on epithelial cell growth and behavior could lead to novel strategies for the prevention and treatment of breast cancer. To this end, the effect of CAF and normal breast-associated fibroblasts (NAF) on the growth of epithelial cells representative of pre-neoplastic breast disease was assessed. NAF and CAF were grown with the nontumorigenic MCF10A epithelial cells and their more transformed, tumorigenic derivative, MCF10AT cells, in direct three-dimensional co-cultures on basement membrane material. The proliferation and apoptosis of MCF10A cells and MCF10AT cells were assessed by 5-bromo-2'-deoxyuridine labeling and TUNEL assay, respectively. Additionally, NAF and CAF were compared for expression of insulin-like growth factor II as a potential mediator of their effects on epithelial cell growth, by ELISA and by quantitative, real-time PCR. In relatively low numbers, both NAF and CAF suppressed proliferation of MCF10A cells. However, only NAF and not CAF significantly inhibited proliferation of the more transformed MCF10AT cells. The degree of growth inhibition varied among NAF or CAF from different individuals. In greater numbers, NAF and CAF have less inhibitory effect on epithelial cell growth. The rate of epithelial cell apoptosis was not affected by NAF or CAF. Mean insulin-like growth factor II levels were not significantly different in NAF versus CAF and did not correlate with the fibroblast effect on epithelial cell proliferation. Both NAF and CAF have the ability to inhibit the growth of pre-cancerous breast epithelial cells. NAF have greater inhibitory capacity than CAF
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Evaluation of a dental pulp-derived cell sheet cultured on amniotic membrane substrate.
Honjo, Ken-ichi; Yamamoto, Toshiro; Adachi, Tetsuya; Amemiya, Takeshi; Mazda, Osam; Kanamura, Narisato; Kita, Masakazu
2015-01-01
Mesenchymal stem cells (MSC) are transplanted for periodontal tissue regeneration, and the periodontal ligament (PDL) is regenerated using a cultured cell sheet. This cultured cell sheet is prepared using PDL-derived cells, growth factors, and amniotic membrane (AM). Dental pulp (DP)-derived cells can be easily obtained from extracted wisdom teeth, proliferate rapidly, and are less susceptible to bacterial infection than PDL-derived cells. Thus, to prepare a novel cell sheet, DP-derived cells were cultured on AM as a culture substrate for immunohistochemical examination. Wisdom teeth extracted from three adults were cut along the cement-enamel border. DP tissue was collected, minced, and primarily cultured. After three or four passage cultures, DP-derived cells were cultured on AM, followed by hematoxylin-eosin (H-E) and immunofluorescence staining. DP-derived cells cultured on AM formed a layered structure. Cells positive for vimentin, Ki-67, ZO-1, desmoplakin, CD29, 44, 105 or 146, STRO-1, collagen IV or VII or laminin 5 or α5 chain were localized. DP-derived cells proliferated on AM, while retaining the properties of DP, which allowed the cultured cell sheet to be prepared. In addition, the cultured cell sheet contained MSC, which suggests its potential application in periodontal tissue regeneration.
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Cellular Plasticity of Epithelial Cells-Cause of Metastasis
National Research Council Canada - National Science Library
Sukumar, Saraswati
2005-01-01
.... We present a novel concept that cancer epithelial cells, possibly of stem cell origin, have inherent cellular plasticity and can differentiate into endothelial cells and form microvessels that serve...
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Directory of Open Access Journals (Sweden)
Ryuhei Hayashi
Full Text Available Induced pluripotent stem (iPS cells can be established from somatic cells. However, there is currently no established strategy to generate corneal epithelial cells from iPS cells. In this study, we investigated whether corneal epithelial cells could be differentiated from iPS cells. We tested 2 distinct sources: human adult dermal fibroblast (HDF-derived iPS cells (253G1 and human adult corneal limbal epithelial cells (HLEC-derived iPS cells (L1B41. We first established iPS cells from HLEC by introducing the Yamanaka 4 factors. Corneal epithelial cells were successfully induced from the iPS cells by the stromal cell-derived inducing activity (SDIA differentiation method, as Pax6(+/K12(+ corneal epithelial colonies were observed after prolonged differentiation culture (12 weeks or later in both the L1B41 and 253G1 iPS cells following retinal pigment epithelial and lens cell induction. Interestingly, the corneal epithelial differentiation efficiency was higher in L1B41 than in 253G1. DNA methylation analysis revealed that a small proportion of differentially methylated regions still existed between L1B41 and 253G1 iPS cells even though no significant difference in methylation status was detected in the specific corneal epithelium-related genes such as K12, K3, and Pax6. The present study is the first to demonstrate a strategy for corneal epithelial cell differentiation from human iPS cells, and further suggests that the epigenomic status is associated with the propensity of iPS cells to differentiate into corneal epithelial cells.
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Dijkman, H.B.P.M.; Weening, J.J.; Smeets, B.; Verrijp, K.; Kuppevelt, A.H.M.S.M. van; Assmann, K.K.; Steenbergen, E.; Wetzels, J.F.M.
2006-01-01
Collapsing focal segmental glomerulosclerosis (cFSGS) is characterized by hyperplasia of glomerular epithelial cells. In a mouse model of FSGS and in a patient with recurrent idiopathic FSGS, we identified the proliferating cells as parietal epithelial cells (PECs). In the present study, we have
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Dijkman, H. B. P. M.; Weening, J. J.; Smeets, B.; Verrijp, K. C. N.; van Kuppevelt, T. H.; Assmann, K. K. J. M.; Steenbergen, E. J.; Wetzels, J. F. M.
2006-01-01
Collapsing focal segmental glomerulosclerosis (cFSGS) is characterized by hyperplasia of glomerular epithelial cells. In a mouse model of FSGS and in a patient with recurrent idiopathic FSGS, we identified the proliferating cells as parietal epithelial cells (PECs). In the present study, we have
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Epithelial cells as active player in fibrosis: findings from an in vitro model.
Directory of Open Access Journals (Sweden)
Solange Moll
Full Text Available Kidney fibrosis, a scarring of the tubulo-interstitial space, is due to activation of interstitial myofibroblasts recruited locally or systemically with consecutive extracellular matrix deposition. Newly published clinical studies correlating acute kidney injury (AKI to chronic kidney disease (CKD challenge this pathological concept putting tubular epithelial cells into the spotlight. In this work we investigated the role of epithelial cells in fibrosis using a simple controlled in vitro system. An epithelial/mesenchymal 3D cell culture model composed of human proximal renal tubular cells and fibroblasts was challenged with toxic doses of Cisplatin, thus injuring epithelial cells. RT-PCR for classical fibrotic markers was performed on fibroblasts to assess their modulation toward an activated myofibroblast phenotype in presence or absence of that stimulus. Epithelial cell lesion triggered a phenotypical modulation of fibroblasts toward activated myofibroblasts as assessed by main fibrotic marker analysis. Uninjured 3D cell culture as well as fibroblasts alone treated with toxic stimulus in the absence of epithelial cells were used as control. Our results, with the caveats due to the limited, but highly controllable and reproducible in vitro approach, suggest that epithelial cells can control and regulate fibroblast phenotype. Therefore they emerge as relevant target cells for the development of new preventive anti-fibrotic therapeutic approaches.
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Traction forces exerted by epithelial cell sheets
International Nuclear Information System (INIS)
Saez, A; Anon, E; Ghibaudo, M; Di Meglio, J-M; Hersen, P; Ladoux, B; Du Roure, O; Silberzan, P; Buguin, A
2010-01-01
Whereas the adhesion and migration of individual cells have been well described in terms of physical forces, the mechanics of multicellular assemblies is still poorly understood. Here, we study the behavior of epithelial cells cultured on microfabricated substrates designed to measure cell-to-substrate interactions. These substrates are covered by a dense array of flexible micropillars whose deflection enables us to measure traction forces. They are obtained by lithography and soft replica molding. The pillar deflection is measured by video microscopy and images are analyzed with home-made multiple particle tracking software. First, we have characterized the temporal and spatial distributions of traction forces of cellular assemblies of various sizes. The mechanical force balance within epithelial cell sheets shows that the forces exerted by neighboring cells strongly depend on their relative position in the monolayer: the largest deformations are always localized at the edge of the islands of cells in the active areas of cell protrusions. The average traction stress rapidly decreases from its maximum value at the edge but remains much larger than the inherent noise due to the force resolution of our pillar tracking software, indicating an important mechanical activity inside epithelial cell islands. Moreover, these traction forces vary linearly with the rigidity of the substrate over about two decades, suggesting that cells exert a given amount of deformation rather than a force. Finally, we engineer micropatterned substrates supporting pillars with anisotropic stiffness. On such substrates cellular growth is aligned with respect to the stiffest direction in correlation with the magnitude of the applied traction forces.
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Ginsberg, Michael; James, Daylon; Ding, Bi-Sen; Nolan, Daniel; Geng, Fuqiang; Butler, Jason M; Schachterle, William; Pulijaal, Venkat R; Mathew, Susan; Chasen, Stephen T; Xiang, Jenny; Rosenwaks, Zev; Shido, Koji; Elemento, Olivier; Rabbany, Sina Y; Rafii, Shahin
2012-01-01
ETS transcription factors ETV2, FLI1 and ERG1 specify pluripotent stem cells into endothelial cells (ECs). However, these ECs are unstable and drift towards non-vascular cell fates. We show that human mid-gestation c-Kit− lineage-committed amniotic cells (ACs) can be readily reprogrammed into induced vascular endothelial cells (iVECs). Transient ETV2 expression in ACs generated proliferative but immature iVECs, while co-expression with FLI1/ERG1 endowed iVECs with a vascular repertoire and morphology matching mature stable ECs. Brief TGFβ-inhibition functionalized VEGFR2 signaling, augmenting specification of ACs to iVECs. Genome-wide transcriptional analyses showed that iVECs are similar to adult ECs in which vascular-specific genes are turned on and non-vascular genes are silenced. Functionally, iVECs form long-lasting patent vasculature in Matrigel plugs and regenerating livers. Thus, short-term ETV2 expression and TGFβ-inhibition along with constitutive ERG1/FLI1 co-expression reprogram mature ACs into durable and functional iVECs with clinical-scale expansion potential. Public banking of HLA-typed iVECs would establish a vascular inventory for treatment of genetically diverse disorders. PMID:23084400
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Uterine epithelial cell proliferation and endometrial hyperplasia: evidence from a mouse model.
Gao, Yang; Li, Shu; Li, Qinglei
2014-08-01
In the uterus, epithelial cell proliferation changes during the estrous cycle and pregnancy. Uncontrolled epithelial cell proliferation results in implantation failure and/or cancer development. Transforming growth factor-β (TGF-β) signaling is a fundamental regulator of diverse biological processes and is indispensable for multiple reproductive functions. However, the in vivo role of TGF-β signaling in uterine epithelial cells remains poorly defined. We have shown that in the uterus, conditional deletion of the Type 1 receptor for TGF-β (Tgfbr1) using anti-Müllerian hormone receptor type 2 (Amhr2) Cre leads to myometrial defects. Here, we describe enhanced epithelial cell proliferation by immunostaining of Ki67 in the uteri of these mice. The aberration culminated in endometrial hyperplasia in aged females. To exclude the potential influence of ovarian steroid hormones, the proliferative status of uterine epithelial cells was assessed following ovariectomy. Increased uterine epithelial cell proliferation was also revealed in ovariectomized Tgfbr1 Amhr2-Cre conditional knockout mice. We further demonstrated that transcript levels for fibroblast growth factor 10 (Fgf10) were markedly up-regulated in Tgfbr1 Amhr2-Cre conditional knockout uteri. Consistently, treatment of primary uterine stromal cells with TGF-β1 significantly reduced Fgf10 mRNA expression. Thus, our findings suggest a potential involvement of TGFBR1-mediated signaling in the regulation of uterine epithelial cell proliferation, and provide genetic evidence supporting the role of uterine epithelial cell proliferation in the pathogenesis of endometrial hyperplasia. © The Author 2014. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology. All rights reserved. For Permissions, please email: journals.permissions@oup.com.
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Proteomic profiling of the amniotic fluid to detect inflammation, infection, and neonatal sepsis.
Directory of Open Access Journals (Sweden)
Catalin S Buhimschi
2007-01-01
Full Text Available Proteomic analysis of amniotic fluid shows the presence of biomarkers characteristic of intrauterine inflammation. We sought to validate prospectively the clinical utility of one such proteomic profile, the Mass Restricted (MR score.We enrolled 169 consecutive women with singleton pregnancies admitted with preterm labor or preterm premature rupture of membranes. All women had a clinically indicated amniocentesis to rule out intra-amniotic infection. A proteomic fingerprint (MR score was generated from fresh samples of amniotic fluid using surface-enhanced laser desorption ionization (SELDI mass spectrometry. Presence or absence of the biomarkers of the MR score was interpreted in relationship to the amniocentesis-to-delivery interval, placental inflammation, and early-onset neonatal sepsis for all neonates admitted to the Newborn Special Care Unit (n = 104. Women with "severe" amniotic fluid inflammation (MR score of 3 or 4 had shorter amniocentesis-to-delivery intervals than women with "no" (MR score of 0 inflammation or even "minimal" (MR score of 1 or 2 inflammation (median [range] MR 3-4: 0.4 d [0.0-49.6 d] versus MR 1-2: 3.8 d [0.0-151.2 d] versus MR 0: 17.0 d [0.1-94.3 d], p 100 cells/mm3, whereas the combination of Gram stain and MR score was best for rapid prediction of intra-amniotic infection (positive amniotic fluid culture.High MR scores are associated with preterm delivery, histological chorioamnionitis, and early-onset neonatal sepsis. In this study, proteomic analysis of amniotic fluid was shown to be the most accurate test for diagnosis of intra-amniotic inflammation, whereas addition of the MR score to the Gram stain provides the best combination of tests to rapidly predict infection.
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Feng, Christina; D Graham, Christopher; Connors, John Patrick; Brazzo, Joseph; Zurakowski, David; Fauza, Dario O
2016-06-01
We compared placental-derived and amniotic fluid-derived mesenchymal stem cells (pMSCs and afMSCs, respectively) in transamniotic stem cell therapy (TRASCET) for experimental spina bifida. Pregnant dams (n=29) exposed to retinoic acid for the induction of fetal spina bifida were divided into four groups. Three groups received volume-matched intraamniotic injections of either saline (n=38 fetuses) or a suspension of 2×10(6) cells/mL of syngeneic, labeled afMSCs (n=73) or pMSCs (n=115) on gestational day 17 (term=21-22days). Untreated fetuses served as controls. Animals were killed before term. Statistical comparisons were by Fisher's exact test (pcell source for TRASCET as a potential alternative in the prenatal management of spina bifida. Copyright © 2016 Elsevier Inc. All rights reserved.
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Van Itallie, Christina M; Tietgens, Amber Jean; Aponte, Angel; Gucek, Marjan; Cartagena-Rivera, Alexander X; Chadwick, Richard S; Anderson, James M
2018-02-02
Treatment of epithelial cells with interferon-γ and TNF-α (IFN/TNF) results in increased paracellular permeability. To identify relevant proteins mediating barrier disruption, we performed proximity-dependent biotinylation (BioID) of occludin and found that tagging of MARCKS-related protein (MRP; also known as MARCKSL1) increased ∼20-fold following IFN/TNF administration. GFP-MRP was focused at the lateral cell membrane and its overexpression potentiated the physiological response of the tight junction barrier to cytokines. However, deletion of MRP did not abrogate the cytokine responses, suggesting that MRP is not required in the occludin-dependent IFN/TNF response. Instead, our results reveal a key role for MRP in epithelial cells in control of multiple actin-based structures, likely by regulation of integrin signaling. Changes in focal adhesion organization and basal actin stress fibers in MRP-knockout (KO) cells were reminiscent of those seen in FAK-KO cells. In addition, we found alterations in cell-cell interactions in MRP-KO cells associated with increased junctional tension, suggesting that MRP may play a role in focal adhesion-adherens junction cross talk. Together, our results are consistent with a key role for MRP in cytoskeletal organization of cell contacts in epithelial cells. © 2018. Published by The Company of Biologists Ltd.
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A Preliminary Study of Human Amniotic Membrane as a Potential Chondrocyte Carrier
Directory of Open Access Journals (Sweden)
L Boo
2009-11-01
Full Text Available PURPOSE: To investigate the feasibility of using processed human amniotic membrane (HAM to support the attachment and proliferation of chondrocytes in vitro which in turn can be utilised as a cell delivery vehicle in tissue engineering applications. METHODS: Fresh HAM obtained from patients undergoing routine elective caesarean sections was harvested, processed and dried using either freeze drying (FD or air drying (AD methods prior to sterilisation by gamma irradiation. Isolated, processed and characterised rabbit autologous chondrocytes were seeded on processed HAM and cultured for up to three weeks. Cell attachment and proliferation were examined qualitatively using inverted brightfield microscopy. RESULTS: Processed HAM appeared to allow cell attachment when implanted with chondrocytes. Although cells seeded on AD and FD HAM did not appear to attach as strongly as those seeded on glycerol preserved intact human amniotic membrane, these cells to be proliferated in cell culture conditions. CONCLUSION: Preliminary results show that processed HAM promotes chondrocyte attachment and proliferation.
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Sasaki, Ayana; Nagatake, Takahiro; Egami, Riku; Gu, Guoqiang; Takigawa, Ichigaku; Ikeda, Wataru; Nakatani, Tomoya; Kunisawa, Jun; Fujita, Yasuyuki
2018-04-24
Recent studies have revealed that newly emerging transformed cells are often eliminated from epithelial tissues via cell competition with the surrounding normal epithelial cells. This cancer preventive phenomenon is termed epithelial defense against cancer (EDAC). However, it remains largely unknown whether and how EDAC is diminished during carcinogenesis. In this study, using a cell competition mouse model, we show that high-fat diet (HFD) feeding substantially attenuates the frequency of apical elimination of RasV12-transformed cells from intestinal and pancreatic epithelia. This process involves both lipid metabolism and chronic inflammation. Furthermore, aspirin treatment significantly facilitates eradication of transformed cells from the epithelial tissues in HFD-fed mice. Thus, our work demonstrates that obesity can profoundly influence competitive interaction between normal and transformed cells, providing insights into cell competition and cancer preventive medicine. Copyright © 2018 The Author(s). Published by Elsevier Inc. All rights reserved.
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Endothelial cells stimulate growth of normal and cancerous breast epithelial cells in 3D culture
Directory of Open Access Journals (Sweden)
Magnusson Magnus K
2010-07-01
Full Text Available Abstract Background Epithelial-stromal interaction provides regulatory signals that maintain correct histoarchitecture and homeostasis in the normal breast and facilitates tumor progression in breast cancer. However, research on the regulatory role of the endothelial component in the normal and malignant breast gland has largely been neglected. The aim of the study was to investigate the effects of endothelial cells on growth and differentiation of human breast epithelial cells in a three-dimensional (3D co-culture assay. Methods Breast luminal and myoepithelial cells and endothelial cells were isolated from reduction mammoplasties. Primary cells and established normal and malignant breast cell lines were embedded in reconstituted basement membrane in direct co-culture with endothelial cells and by separation of Transwell filters. Morphogenic and phenotypic profiles of co-cultures was evaluated by phase contrast microscopy, immunostaining and confocal microscopy. Results In co-culture, endothelial cells stimulate proliferation of both luminal- and myoepithelial cells. Furthermore, endothelial cells induce a subpopulation of luminal epithelial cells to form large acini/ducts with a large and clear lumen. Endothelial cells also stimulate growth and cloning efficiency of normal and malignant breast epithelial cell lines. Transwell and gradient co-culture studies show that endothelial derived effects are mediated - at least partially - by soluble factors. Conclusion Breast endothelial cells - beside their role in transporting nutrients and oxygen to tissues - are vital component of the epithelial microenvironment in the breast and provide proliferative signals to the normal and malignant breast epithelium. These growth promoting effects of endothelial cells should be taken into consideration in breast cancer biology.
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International Nuclear Information System (INIS)
Gao, Rundi; Chen, Ruilin; Cao, Yu; Wang, Yuan; Song, Kang; Zhang, Ya; Yang, Junchao
2017-01-01
Pulmonary fibrosis is characterized by the destruction of lung tissue architecture and the formation of fibrous foci, currently has no satisfactory treatment. Emodin is a component of Chinese herb that has been reported to be medicament on pancreatic fibrosis and liver fibrosis. However, its role in pulmonary fibrosis has not been established yet. In the present study, we investigated the hypothesis that Emodin plays an inhibitory role in TGF-β1 induced epithelial-mesenchymal transition (EMT) of alveolar epithelial cell, and Emodin exerts its effect through the Notch signaling pathway. Emodin inhibits the proliferation of Rat alveolar type II epithelial cells RLE-6TN in a concentration-dependent manner; reduces the expression of Collagen I, α-SMA and Vimentin, promotes the expression of E-cadherin. Moreover, Emodin could regulate the expression patterns of the Notch signaling pathway-related factors and reduce the Notch-1 nucleus translocation. Knockdown of Notch-1 enhances the inhibitory effect of Emodin on TGF-β1-induced EMT in RLE-6TN cells. In conclusion, the data of the present study suggests that Emodin suppresses TGF-β1-induced EMT in alveolar epithelial cells through Notch signaling pathway and shows the potential to be effective in the treatment of pulmonary fibrosis. - Highlights: • Emodin inhibits TGF-β1-induced EMT in alveolar epithelial cells. • Emodin regulates the expression patterns of the Notch signaling pathway-related factors. • Emodin inhibits TGF-β1-induced Notch-1 nucleus translocation and activation.
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Energy Technology Data Exchange (ETDEWEB)
Gao, Rundi; Chen, Ruilin; Cao, Yu [Department of Respiration, The First Affiliated Hospital of Zhejiang Chinese Medicine University, NO. 56, Youdian Road, Shangcheng District, Hangzhou, Zhejiang Province 310006 (China); Wang, Yuan [Department of Pulmonary Function, The First Affiliated Hospital of Zhejiang Chinese Medicine University, NO. 56, Youdian Road, Shangcheng District, Hangzhou, Zhejiang Province 310006 (China); Song, Kang [Department of Respiration, The First Affiliated Hospital of Zhejiang Chinese Medicine University, NO. 56, Youdian Road, Shangcheng District, Hangzhou, Zhejiang Province 310006 (China); Zhang, Ya [Zhejiang Chinese Medicine University, No. 548, Binwen Road, Binjiang District, Hangzhou, Zhejiang Province 310006 (China); Yang, Junchao, E-mail: yangjunchaozj@zcmu.edu.cn [Department of Respiration, The First Affiliated Hospital of Zhejiang Chinese Medicine University, NO. 56, Youdian Road, Shangcheng District, Hangzhou, Zhejiang Province 310006 (China)
2017-03-01
Pulmonary fibrosis is characterized by the destruction of lung tissue architecture and the formation of fibrous foci, currently has no satisfactory treatment. Emodin is a component of Chinese herb that has been reported to be medicament on pancreatic fibrosis and liver fibrosis. However, its role in pulmonary fibrosis has not been established yet. In the present study, we investigated the hypothesis that Emodin plays an inhibitory role in TGF-β1 induced epithelial-mesenchymal transition (EMT) of alveolar epithelial cell, and Emodin exerts its effect through the Notch signaling pathway. Emodin inhibits the proliferation of Rat alveolar type II epithelial cells RLE-6TN in a concentration-dependent manner; reduces the expression of Collagen I, α-SMA and Vimentin, promotes the expression of E-cadherin. Moreover, Emodin could regulate the expression patterns of the Notch signaling pathway-related factors and reduce the Notch-1 nucleus translocation. Knockdown of Notch-1 enhances the inhibitory effect of Emodin on TGF-β1-induced EMT in RLE-6TN cells. In conclusion, the data of the present study suggests that Emodin suppresses TGF-β1-induced EMT in alveolar epithelial cells through Notch signaling pathway and shows the potential to be effective in the treatment of pulmonary fibrosis. - Highlights: • Emodin inhibits TGF-β1-induced EMT in alveolar epithelial cells. • Emodin regulates the expression patterns of the Notch signaling pathway-related factors. • Emodin inhibits TGF-β1-induced Notch-1 nucleus translocation and activation.
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Moderate plasma activated media suppresses proliferation and migration of MDCK epithelial cells
International Nuclear Information System (INIS)
Mohades, Soheila; Laroussi, Mounir; Maruthamuthu, Venkat
2017-01-01
Low-temperature plasma has been shown to have diverse biomedical uses, including its applications in cancer and wound healing. One recent approach in treating mammalian cells with plasma is through the use of plasma activated media (PAM), which is produced by exposing cell culture media to plasma. While the adverse effects of PAM treatment on cancerous epithelial cell lines have been recently studied, much less is known about the interaction of PAM with normal epithelial cells. In this paper, non-cancerous canine kidney MDCK (Madin-Darby Canine Kidney) epithelial cells were treated by PAM and time-lapse microscopy was used to directly monitor their proliferation and random migration upon treatment. While longer durations of PAM treatment led to cell death, we found that moderate levels of PAM treatment inhibited proliferation in these epithelial cells. We also found that PAM treatment reduced random cell migration within epithelial islands. Immunofluorescence staining showed that while there were no major changes in the actin/adhesion apparatus, there was a significant change in the nuclear localization of proliferation marker Ki-67, consistent with our time-lapse results. (paper)
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Bio-synthesis of gold nanoparticles by human epithelial cells, in vivo
International Nuclear Information System (INIS)
Larios-Rodriguez, E; Rangel-Ayon, C; Herrera-Urbina, R; Castillo, S J; Zavala, G
2011-01-01
Healthy epithelial cells, in vivo, have the ability to synthesize gold nanoparticles when aqueous tetrachloroauric acid is made to react with human skin. Neither a reducing agent nor a protecting chemical is needed for this bio-synthesis method. The first indication of gold nanoparticle formation is the staining of the skin, which turns deep purple. Stereoscopic optical micrographs of human skin tissue in contact with aqueous tetrachloroauric acid clearly show the staining of the epithelial cells. The UV-Vis spectrum of these epithelial cells shows an absorption band with a maximum at 553 nm. This absorption peak is within the wavelength region where the surface plasmon resonance (SPR) band of aqueous colloidal gold exhibits a maximum. Transmission electron micrographs show that gold nanoparticles synthesized by epithelial cells have sizes between 1 and 100 nm. The electron diffraction pattern of these nanoparticles reveals a crystalline structure whose interplanar distances correspond to fcc metallic gold. Transmission electron micrographs of ultra-thin (70 nm thick) slices of epithelial cells clearly and undoubtedly demonstrate that gold nanoparticles are inside the cell. According to high resolution transmission electron micrographs of intracellular single gold nanoparticles, they have the shape of a polyhedron.
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Bio-synthesis of gold nanoparticles by human epithelial cells, in vivo
Energy Technology Data Exchange (ETDEWEB)
Larios-Rodriguez, E; Rangel-Ayon, C; Herrera-Urbina, R [Departamento de Ingenieria Quimica y Metalurgia, Universidad de Sonora, Rosales y Luis Encinas S/N, Hermosillo, Sonora, C.P. 83000 (Mexico); Castillo, S J [Departamento de Investigacion en Fisica, Universidad de Sonora, Rosales y Luis Encinas S/N, Hermosillo, Sonora, C.P. 83000 (Mexico); Zavala, G, E-mail: elarios@polimeros.uson.mx [Instituto de Biotecnologia, Universidad Nacional Autonoma de Mexico, Cuernavaca, Morelos (Mexico)
2011-09-02
Healthy epithelial cells, in vivo, have the ability to synthesize gold nanoparticles when aqueous tetrachloroauric acid is made to react with human skin. Neither a reducing agent nor a protecting chemical is needed for this bio-synthesis method. The first indication of gold nanoparticle formation is the staining of the skin, which turns deep purple. Stereoscopic optical micrographs of human skin tissue in contact with aqueous tetrachloroauric acid clearly show the staining of the epithelial cells. The UV-Vis spectrum of these epithelial cells shows an absorption band with a maximum at 553 nm. This absorption peak is within the wavelength region where the surface plasmon resonance (SPR) band of aqueous colloidal gold exhibits a maximum. Transmission electron micrographs show that gold nanoparticles synthesized by epithelial cells have sizes between 1 and 100 nm. The electron diffraction pattern of these nanoparticles reveals a crystalline structure whose interplanar distances correspond to fcc metallic gold. Transmission electron micrographs of ultra-thin (70 nm thick) slices of epithelial cells clearly and undoubtedly demonstrate that gold nanoparticles are inside the cell. According to high resolution transmission electron micrographs of intracellular single gold nanoparticles, they have the shape of a polyhedron.
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Directory of Open Access Journals (Sweden)
Evelyne Beerling
2016-03-01
Full Text Available Forced overexpression and/or downregulation of proteins regulating epithelial-to-mesenchymal transition (EMT has been reported to alter metastasis by changing migration and stem cell capacity of tumor cells. However, these manipulations artificially keep cells in fixed states, while in vivo cells may adapt transient and reversible states. Here, we have tested the existence and role of epithelial-mesenchymal plasticity in metastasis of mammary tumors without artificially modifying EMT regulators. In these tumors, we found by intravital microscopy that the motile tumor cells have undergone EMT, while their epithelial counterparts were not migratory. Moreover, we found that epithelial-mesenchymal plasticity renders any EMT-induced stemness differences, as reported previously, irrelevant for metastatic outgrowth, because mesenchymal cells that arrive at secondary sites convert to the epithelial state within one or two divisions, thereby obtaining the same stem cell potential as their arrived epithelial counterparts. We conclude that epithelial-mesenchymal plasticity supports migration but additionally eliminates stemness-enhanced metastatic outgrowth differences.
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PKC activation induces inflammatory response and cell death in human bronchial epithelial cells.
Directory of Open Access Journals (Sweden)
Hyunhee Kim
Full Text Available A variety of airborne pathogens can induce inflammatory responses in airway epithelial cells, which is a crucial component of host defence. However, excessive inflammatory responses and chronic inflammation also contribute to different diseases of the respiratory system. We hypothesized that the activation of protein kinase C (PKC is one of the essential mechanisms of inflammatory response in airway epithelial cells. In the present study, we stimulated human bronchial lung epithelial (BEAS-2B cells with the phorbol ester Phorbol 12, 13-dibutyrate (PDBu, and examined gene expression profile using microarrays. Microarray analysis suggests that PKC activation induced dramatic changes in gene expression related to multiple cellular functions. The top two interaction networks generated from these changes were centered on NFκB and TNF-α, which are two commonly known pathways for cell death and inflammation. Subsequent tests confirmed the decrease in cell viability and an increase in the production of various cytokines. Interestingly, each of the increased cytokines was differentially regulated at mRNA and/or protein levels by different sub-classes of PKC isozymes. We conclude that pathological cell death and cytokine production in airway epithelial cells in various situations may be mediated through PKC related signaling pathways. These findings suggest that PKCs can be new targets for treatment of lung diseases.
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Retinoic Acid Signaling in Thymic Epithelial Cells Regulates Thymopoiesis
DEFF Research Database (Denmark)
Wendland, Kerstin; Niss, Kristoffer; Kotarsky, Knut
2018-01-01
Despite the essential role of thymic epithelial cells (TEC) in T cell development, the signals regulating TEC differentiation and homeostasis remain incompletely understood. In this study, we show a key in vivo role for the vitamin A metabolite, retinoic acid (RA), in TEC homeostasis. In the abse......Despite the essential role of thymic epithelial cells (TEC) in T cell development, the signals regulating TEC differentiation and homeostasis remain incompletely understood. In this study, we show a key in vivo role for the vitamin A metabolite, retinoic acid (RA), in TEC homeostasis...
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Parietal epithelial cells: their role in health and disease.
Romagnani, Paola
2011-01-01
Parietal epithelial cells of Bowman's capsules were first described by Sir William Bowman in 1842 in his paper On the Structure and Use of the Malpighian Bodies of the Kidney [London, Taylor, 1842], but since then their functions have remained poorly understood. A large body of evidence has recently suggested that parietal epithelial cells represent a reservoir of renal progenitors in adult human kidney which generate novel podocytes during childhood and adolescence, and can regenerate injured podocytes. The discovery that parietal epithelial cells represent a potential source for podocyte regeneration suggests that podocyte injury can be repaired. However, recent results also suggest that an abnormal proliferative response of renal progenitors to podocyte injury can generate hyperplastic glomerular lesions that are observed in crescentic glomerulonephritis and other types of glomerular disorders. Taken together, these results establish an entirely novel view that changes the way of thinking about renal physiology and pathophysiology, and suggest that understanding how self-renewal and fate decision of parietal epithelial cells in response to podocyte injury may be perturbed or modulated will be crucial for obtaining novel tools for prevention and treatment of glomerulosclerosis. Copyright © 2011 S. Karger AG, Basel.
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Metabolic cooperativity between epithelial cells and adipocytes of mice
International Nuclear Information System (INIS)
Bartley, J.C.; Emerman, J.T.; Bissell, M.J.
1981-01-01
We have demonstrated that glycogen and lipid synthesis in adipocytes is modulated by the lactational state and that this modulation in mammary adipocytes requires the presence of the adjacent epithelial cells. Glycogen and lipid synthesis from [ 14 C]glucose was measured in mammary fat pads cleared of epithelium, in abdominal fat pads, and in adipocytes from both sources and from intact mammary gland of mature virgin, pregnant, and lactating mice. Accumulation of glycogen, the activity of glycogen synthase, and the lipogenic rate in abdominal and mammary adipocytes remained high during pregnancy but decreased to insignificant levels by early lactation. The depressant effects of lactation were observed solely in those mammary adipocytes isolated from intact glands. The presence of mammary epithelial cells was also required to effect the stimulated lipogenesis in mammary adipocytes during pregnancy. We conclude that the metabolic activity of adipocytes is modulated both during pregnancy and lactation to channel nutrients to the mammary epithelial cell. The fact that the changes occur in mammary adipocytes only when epithelial cells are present indicates that local as well as systemic factors are operating in these modulations
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Alterations in Helicobacter pylori triggered by contact with gastric epithelial cells
Directory of Open Access Journals (Sweden)
Elizabeth M. Johnson
2012-02-01
Full Text Available Helicobacter pylori lives within the mucus layer of the human stomach, in close proximity to gastric epithelial cells. While a great deal is known about the effects of H. pylori on human cells and the specific bacterial products that mediate these effects, relatively little work has been done to investigate alterations in H. pylori that may be triggered by bacterial contact with human cells. In this review, we discuss the spectrum of changes in bacterial physiology and morphology that occur when H. pylori is in contact with gastric epithelial cells. Several studies have reported that cell contact causes alterations in H. pylori gene transcription. In addition, H. pylori contact with gastric epithelial cells promotes the formation of pilus-like structures at the bacteria-host cell interface. The formation of these structures requires multiple genes in the cag pathogenicity island, and these structures are proposed to have an important role in the type IV secretion system-dependent process through which CagA enters host cells. Finally, H. pylori contact with epithelial cells can promote bacterial replication and the formation of microcolonies, phenomena that are facilitated by the acquisition of iron and other nutrients from infected cells. In summary, the gastric epithelial cell surface represents an important niche for H. pylori, and upon entry into this niche, the bacteria alter their behavior in a manner that optimizes bacterial proliferation and persistent colonization of the host.
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Response of cultured normal human mammary epithelial cells to X rays
International Nuclear Information System (INIS)
Yang, T.C.; Stampfer, M.R.; Smith, H.S.
1983-01-01
The effect of X rays on the reproductive death of cultured normal human mammary epithelial cells was examined. Techniques were developed for isolating and culturing normal human mammary epithelial cells which provide sufficient cells at second passage for radiation studies, and an efficient clonogenic assay suitable for measuring radiation survival curves. It was found that the survival curves for epithelial cells from normal breast tissue were exponential and had D 0 values of about 109-148 rad for 225 kVp X rays. No consistent change in cell radiosensitivity with the age of donor was observed, and no sublethal damage repair in these cells could be detected with the split-dose technique
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Taurine Protects Lens Epithelial Cells Against Ultraviolet B-Induced Apoptosis.
Dayang, Wu; Dongbo, Pang
2017-10-01
The massive uptake of compatible osmolytes is a self-protective response shared by lens exposed to hypertonic stress and ultraviolet stress. This study aimed to investigate the protective effects of taurine against ultraviolet B-induced cytotoxicity in the lens epithelial cells. Real-time PCR was used to measure osmolytes transport. Radioimmunoassay was used to measure osmolytes uptake. Cell counting kit-8 assays were used to measure cellular viability. Flow cytometry analysis was used to measure apoptosis level. Compared with normotonic stress, hypertonic stress-induced osmolytes uptake into the lens epithelial cells such as betaine, myoinositol and taurine. UVB exposure increased osmolytes transporter mRNA expression together with osmolytes uptake. Moreover, taurine suppressed UVB-induced cell apoptosis in the lens epithelial cells significantly. The effect of compatible osmolyte taurine on cell survival rate may play an important role in cell resistance and adaption to UVB exposure.
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Kamata, Hirofumi; Yamamoto, Kazuko; Wasserman, Gregory A; Zabinski, Mary C; Yuen, Constance K; Lung, Wing Yi; Gower, Adam C; Belkina, Anna C; Ramirez, Maria I; Deng, Jane C; Quinton, Lee J; Jones, Matthew R; Mizgerd, Joseph P
2016-09-01
Airway epithelial cell responses are critical to the outcome of lung infection. In this study, we aimed to identify unique contributions of epithelial cells during lung infection. To differentiate genes induced selectively in epithelial cells during pneumonia, we compared genome-wide expression profiles from three sorted cell populations: epithelial cells from uninfected mouse lungs, epithelial cells from mouse lungs with pneumococcal pneumonia, and nonepithelial cells from those same infected lungs. Of 1,166 transcripts that were more abundant in epithelial cells from infected lungs compared with nonepithelial cells from the same lungs or from epithelial cells of uninfected lungs, 32 genes were identified as highly expressed secreted products. Especially strong signals included two related secreted and transmembrane (Sectm) 1 genes, Sectm1a and Sectm1b. Refinement of sorting strategies suggested that both Sectm1 products were induced predominantly in conducting airway epithelial cells. Sectm1 was induced during the early stages of pneumococcal pneumonia, and mutation of NF-κB RelA in epithelial cells did not diminish its expression. Instead, type I IFN signaling was necessary and sufficient for Sectm1 induction in lung epithelial cells, mediated by signal transducer and activator of transcription 1. For target cells, Sectm1a bound to myeloid cells preferentially, in particular Ly6G(bright)CD11b(bright) neutrophils in the infected lung. In contrast, Sectm1a did not bind to neutrophils from uninfected lungs. Sectm1a increased expression of the neutrophil-attracting chemokine CXCL2 by neutrophils from the infected lung. We propose that Sectm1a is an epithelial product that sustains a positive feedback loop amplifying neutrophilic inflammation during pneumococcal pneumonia.
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Urokinase and the intestinal mucosa: evidence for a role in epithelial cell turnover
Gibson, P; Birchall, I; Rosella, O; Albert, V; Finch, C; Barkla, D; Young, G
1998-01-01
Background—The functions of urokinase in intestinal epithelia are unknown. Aims—To determine the relation of urokinase expressed by intestinal epithelial cells to their position in the crypt-villus/surface axis and of mucosal urokinase activity to epithelial proliferative kinetics in the distal colon. Methods—Urokinase expression was examined immunohistochemically in human intestinal mucosa. Urokinase activity was measured colorimetrically in epithelial cells isolated sequentially from the crypt-villus axis of the rat small intestine. In separate experiments, urokinase activity and epithelial kinetics (measured stathmokinetically) were measured in homogenates of distal colonic mucosa of 14 groups of eight rats fed diets known to alter epithelial turnover. Results—From the crypt base, an ascending gradient of expression and activity of urokinase was associated with the epithelial cells. Median mucosal urokinase activities in each of the dietary groups of rats correlated positively with autologous median number of metaphase arrests per crypt (r=0.68; p<0.005) and per 100 crypt cells (r=0.75; p<0.001), but not with crypt column height. Conclusions—Localisation of an enzyme capable of leading to digestion of cell substratum in the region where cells are loosely attached to their basement membrane, and the association of its activity with indexes of cell turnover, suggest a role for urokinase in facilitating epithelial cell loss in the intestine. Keywords: urokinase; intestinal epithelium; colon; epithelial proliferation PMID:9824347
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Invasion of Human Oral Epithelial Cells by Prevotella intermedia
Dorn, Brian R.; Leung, K.-P.; Progulske-Fox, Ann
1998-01-01
Invasion of oral epithelial cells by pathogenic oral bacteria may represent an important virulence factor in the progression of periodontal disease. Here we report that a clinical isolate of Prevotella intermedia, strain 17, was found to invade a human oral epithelial cell line (KB), whereas P. intermedia 27, another clinical isolate, and P. intermedia 25611, the type strain, were not found to invade the cell line. Invasion was quantified by the recovery of viable bacteria following a standard antibiotic protection assay and observed by electron microscopy. Cytochalasin D, cycloheximide, monodansylcadaverine, and low temperature (4°C) inhibited the internalization of P. intermedia 17. Antibodies raised against P. intermedia type C fimbriae and against whole cells inhibited invasion, but the anti-type-C-fimbria antibody inhibited invasion to a greater extent than the anti-whole-cell antibody. This work provides evidence that at least one strain of P. intermedia can invade an oral epithelial cell line and that the type C fimbriae and a cytoskeletal rearrangement are required for this invasion. PMID:9826397
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Acrolein stimulates eicosanoid release from bovine airway epithelial cells
International Nuclear Information System (INIS)
Doupnik, C.A.; Leikauf, G.D.
1990-01-01
Injury to the airway mucosa after exposure to environmental irritants is associated with pulmonary inflammation and bronchial hyperresponsiveness. To better understand the relationships between mediator release and airway epithelial cell injury during irritant exposures, we studied the effects of acrolein, a low-molecular-weight aldehyde found in cigarette smoke, on arachidonic acid metabolism in cultured bovine tracheal epithelial cells. Confluent airway epithelial cell monolayers, prelabeled with [3H]arachidonic acid, released significant levels of 3H activity when exposed (20 min) to 100 microM acrolein. [3H]arachidonic acid products were resolved using reverse-phase high-performance liquid chromatography. Under control conditions the released 3H activity coeluted predominantly with the cyclooxygenase product, prostaglandin (PG) E2. After exposure to acrolein, significant peaks in 3H activity coeluted with the lipoxygenase products 12-hydroxyeicosatetraenoic acid (HETE) and 15-HETE, as well as with PGE2, PGF2 alpha, and 6-keto-PGF1 alpha. Dose-response relationships for acrolein-induced release of immunoreactive PGF2 alpha and PGE2 from unlabeled epithelial monolayers demonstrated 30 microM acrolein as the threshold dose, with 100 microM acrolein inducing nearly a fivefold increase in both PGF2 alpha and PGE2. Cellular viability after exposure to 100 microM acrolein, determined by released lactate dehydrogenase activity, was not affected until exposure periods were greater than or equal to 2 h. These results implicate the airway epithelial cell as a possible source of eicosanoids after exposure to acrolein
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Gene transfer to primary corneal epithelial cells with an integrating lentiviral vector
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Lauro Augusto de Oliveira
2010-10-01
Full Text Available PURPOSE: To evaluate the transfer of heterologous genes carrying a Green Fluorescent Protein (GFP reporter cassette to primary corneal epithelial cells ex vivo. METHODS: Freshly enucleated rabbit corneoscleral tissue was used to obtain corneal epithelial cell suspension via enzymatic digestion. Cells were plated at a density of 5×10³ cells/cm² and allowed to grow for 5 days (to 70-80% confluency prior to transduction. Gene transfer was monitored using fluorescence microscopy and fluorescence activated cell sorter (FACS. We evaluated the transduction efficiency (TE over time and the dose-response effect of different lentiviral particles. One set of cells were dual sorted by fluorescence activated cell sorter for green fluorescent protein expression as well as Hoechst dye exclusion to evaluate the transduction of potentially corneal epithelial stem cells (side-population phenotypic cells. RESULTS: Green fluorescent protein expressing lentiviral vectors were able to effectively transduce rabbit primary epithelial cells cultured ex vivo. Live cell imaging post-transduction demonstrated GFP-positive cells with normal epithelial cell morphology and growth. The transduction efficiency over time was higher at the 5th post-transduction day (14.1% and tended to stabilize after the 8th day. The number of transduced cells was dose-dependent, and at the highest lentivirus concentrations approached 7%. When double sorted by fluorescence activated cell sorter to isolate both green fluorescent protein positive and side population cells, transduced side population cells were identified. CONCLUSIONS: Lentiviral vectors can effectively transfer heterologous genes to primary corneal epithelial cells expanded ex vivo. Genes were stably expressed over time, transferred in a dose-dependence fashion, and could be transferred to mature corneal cells as well as presumable putative stem cells.
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International Nuclear Information System (INIS)
Miura, Yuka; Hagiwara, Natsumi; Radisky, Derek C.; Hirai, Yohei
2014-01-01
Activation of the epithelial-mesenchymal transition (EMT) program promotes cell invasion and metastasis, and is reversed through mesenchymal-epithelial transition (MET) after formation of distant metastases. Here, we show that an imbalance of gene products encoded by the transcriptional factor C/EBPβ, LAP (liver-enriched activating protein) and LIP (liver-enriched inhibitory protein), can regulate both EMT- and MET-like phenotypic changes in mouse mammary epithelial cells. By using tetracycline repressive LIP expression constructs, we found that SCp2 cells, a clonal epithelial line of COMMA1-D cells, expressed EMT markers, lost the ability to undergo alveolar-like morphogenesis in 3D Matrigel, and acquired properties of benign adenoma cells. Conversely, we found that inducible expression of LAP in SCg6 cells, a clonal fibroblastic line of COMMA1-D cells, began to express epithelial keratins with suppression of proliferation. The overexpression of the C/EBPβ gene products in these COMMA1-D derivatives was suppressed by long-term cultivation on tissue culture plastic, but gene expression was maintained in cells grown on Matrigel or exposed to proteasome inhibitors. Thus, imbalances of C/EBPβ gene products in mouse mammary epithelial cells, which are affected by contact with basement membrane, are defined as a potential regulator of metastatic potential. - Highlights: • We created a temporal imbalance of C/EBPβ gene products in the mammary model cells. • The temporal up-regulation of LIP protein induced EMT-like cell behaviors. • The temporal up-regulation of LAP protein induced MET-like cell behaviors. • Excess amount of C/EBPβ gene products were eliminated by proteasomal-degradation. • Basement membrane components attenuated proteasome-triggered protein elimination
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Directory of Open Access Journals (Sweden)
Whittington Kate
2009-07-01
Full Text Available Abstract Background Epithelial cells in first catch urine (FCU specimens from 87 men with and without urethritis were quantified. Epithelial cells were broadly categorised into transitional and squamous populations using morphological characteristics and immunostaining with anti-pan leukocyte and anti-cytokeratin monoclonal antibodies. Findings The majority (77/87 = 89% of samples contained both transitional (76/87 = 87%; range 1 × 104 – 6 × 105, median 6 × 104 and squamous (57/87 = 66%; range 1 × 104 – 8 × 105, median 2 × 104 epithelial cells. The number of transitional cells correlated with the number of squamous cells (Spearman's rho = 0.697 p Conclusion Further studies are required to explore the complexity of epithelial cell populations in urine. These would provide novel opportunities for studying cellular interactions of C. trachomatis in male urethral infections, about which little is currently known.
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Romo1 expression contributes to oxidative stress-induced death of lung epithelial cells
International Nuclear Information System (INIS)
Shin, Jung Ar; Chung, Jin Sil; Cho, Sang-Ho; Kim, Hyung Jung; Yoo, Young Do
2013-01-01
Highlights: •Romo1 mediates oxidative stress-induced mitochondrial ROS production. •Romo1 induction by oxidative stress plays an important role in oxidative stress-induced apoptosis. •Romo1 overexpression correlates with epithelial cell death in patients with IPF. -- Abstract: Oxidant-mediated death of lung epithelial cells due to cigarette smoking plays an important role in pathogenesis in lung diseases such as idiopathic pulmonary fibrosis (IPF). However, the exact mechanism by which oxidants induce epithelial cell death is not fully understood. Reactive oxygen species (ROS) modulator 1 (Romo1) is localized in the mitochondria and mediates mitochondrial ROS production through complex III of the mitochondrial electron transport chain. Here, we show that Romo1 mediates mitochondrial ROS production and apoptosis induced by oxidative stress in lung epithelial cells. Hydrogen peroxide (H 2 O 2 ) treatment increased Romo1 expression, and Romo1 knockdown suppressed the cellular ROS levels and cell death triggered by H 2 O 2 treatment. In immunohistochemical staining of lung tissues from patients with IPF, Romo1 was mainly localized in hyperplastic alveolar and bronchial epithelial cells. Romo1 overexpression was detected in 14 of 18 patients with IPF. TUNEL-positive alveolar epithelial cells were also detected in most patients with IPF but not in normal controls. These findings suggest that Romo1 mediates apoptosis induced by oxidative stress in lung epithelial cells
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Romo1 expression contributes to oxidative stress-induced death of lung epithelial cells
Energy Technology Data Exchange (ETDEWEB)
Shin, Jung Ar [Department of Internal Medicine, Yonsei University College of Medicine, Yonsei University Health System, Seoul 135-270 (Korea, Republic of); Chung, Jin Sil [Laboratory of Molecular Cell Biology, College of Life Sciences and Biotechnology, Korea University, Seoul 136-713 (Korea, Republic of); Cho, Sang-Ho [Department of Pathology, Pochon CHA University, College of Medicine, Gyeonggi-do (Korea, Republic of); Kim, Hyung Jung, E-mail: khj57@yuhs.ac.kr [Department of Internal Medicine, Yonsei University College of Medicine, Yonsei University Health System, Seoul 135-270 (Korea, Republic of); Yoo, Young Do, E-mail: ydy1130@korea.ac.kr [Laboratory of Molecular Cell Biology, College of Life Sciences and Biotechnology, Korea University, Seoul 136-713 (Korea, Republic of)
2013-09-20
Highlights: •Romo1 mediates oxidative stress-induced mitochondrial ROS production. •Romo1 induction by oxidative stress plays an important role in oxidative stress-induced apoptosis. •Romo1 overexpression correlates with epithelial cell death in patients with IPF. -- Abstract: Oxidant-mediated death of lung epithelial cells due to cigarette smoking plays an important role in pathogenesis in lung diseases such as idiopathic pulmonary fibrosis (IPF). However, the exact mechanism by which oxidants induce epithelial cell death is not fully understood. Reactive oxygen species (ROS) modulator 1 (Romo1) is localized in the mitochondria and mediates mitochondrial ROS production through complex III of the mitochondrial electron transport chain. Here, we show that Romo1 mediates mitochondrial ROS production and apoptosis induced by oxidative stress in lung epithelial cells. Hydrogen peroxide (H{sub 2}O{sub 2}) treatment increased Romo1 expression, and Romo1 knockdown suppressed the cellular ROS levels and cell death triggered by H{sub 2}O{sub 2} treatment. In immunohistochemical staining of lung tissues from patients with IPF, Romo1 was mainly localized in hyperplastic alveolar and bronchial epithelial cells. Romo1 overexpression was detected in 14 of 18 patients with IPF. TUNEL-positive alveolar epithelial cells were also detected in most patients with IPF but not in normal controls. These findings suggest that Romo1 mediates apoptosis induced by oxidative stress in lung epithelial cells.
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Crystalline structure of pulverized dental calculus induces cell death in oral epithelial cells.
Ziauddin, S M; Yoshimura, A; Montenegro Raudales, J L; Ozaki, Y; Higuchi, K; Ukai, T; Kaneko, T; Miyazaki, T; Latz, E; Hara, Y
2018-06-01
Dental calculus is a mineralized deposit attached to the tooth surface. We have shown that cellular uptake of dental calculus triggers nucleotide-binding oligomerization domain-like receptor family pyrin domain-containing 3 (NLRP3) inflammasome activation, leading to the processing of the interleukin-1β precursor into its mature form in mouse and human phagocytes. The activation of the NLRP3 inflammasome also induced a lytic form of programmed cell death, pyroptosis, in these cells. However, the effects of dental calculus on other cell types in periodontal tissue have not been investigated. The aim of this study was to determine whether dental calculus can induce cell death in oral epithelial cells. HSC-2 human oral squamous carcinoma cells, HOMK107 human primary oral epithelial cells and immortalized mouse macrophages were exposed to dental calculus or 1 of its components, hydroxyapatite crystals. For inhibition assays, the cells were exposed to dental calculus in the presence or absence of cytochalasin D (endocytosis inhibitor), z-YVAD-fmk (caspase-1 inhibitor) or glyburide (NLRP3 inflammasome inhibitor). Cytotoxicity was determined by measuring lactate dehydrogenase (LDH) release and staining with propidium iodide. Tumor necrosis factor-α production was quantified by enzyme-linked immunosorbent assay. Oral epithelial barrier function was examined by permeability assay. Dental calculus induced cell death in HSC-2 cells, as judged by LDH release and propidium iodide staining. Dental calculus also induced LDH release from HOMK107 cells. Following heat treatment, dental calculus lost its capacity to induce tumor necrosis factor-α in mouse macrophages, but could induce LDH release in HSC-2 cells, indicating a major role of inorganic components in cell death. Hydroxyapatite crystals also induced cell death in both HSC-2 and HOMK107 cells, as judged by LDH release, indicating the capacity of crystal particles to induce cell death. Cell death induced by dental
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Epithelial WNT Ligands Are Essential Drivers of Intestinal Stem Cell Activation
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Winnie Y. Zou
2018-01-01
Full Text Available Intestinal stem cells (ISCs maintain and repair the intestinal epithelium. While regeneration after ISC-targeted damage is increasingly understood, injury-repair mechanisms that direct regeneration following injuries to differentiated cells remain uncharacterized. The enteric pathogen, rotavirus, infects and damages differentiated cells while sparing all ISC populations, thus allowing the unique examination of the response of intact ISC compartments during injury-repair. Upon rotavirus infection in mice, ISC compartments robustly expand and proliferating cells rapidly migrate. Infection results specifically in stimulation of the active crypt-based columnar ISCs, but not alternative reserve ISC populations, as is observed after ISC-targeted damage. Conditional ablation of epithelial WNT secretion diminishes crypt expansion and ISC activation, demonstrating a previously unknown function of epithelial-secreted WNT during injury-repair. These findings indicate a hierarchical preference of crypt-based columnar cells (CBCs over other potential ISC populations during epithelial restitution and the importance of epithelial-derived signals in regulating ISC behavior.
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Inhibition of PTP1B disrupts cell-cell adhesion and induces anoikis in breast epithelial cells.
Hilmarsdottir, Bylgja; Briem, Eirikur; Halldorsson, Skarphedinn; Kricker, Jennifer; Ingthorsson, Sævar; Gustafsdottir, Sigrun; Mælandsmo, Gunhild M; Magnusson, Magnus K; Gudjonsson, Thorarinn
2017-05-11
Protein tyrosine phosphatase 1B (PTP1B) is a well-known inhibitor of insulin signaling pathways and inhibitors against PTP1B are being developed as promising drug candidates for treatment of obesity. PTP1B has also been linked to breast cancer both as a tumor suppressor and as an oncogene. Furthermore, PTP1B has been shown to be a regulator of cell adhesion and migration in normal and cancer cells. In this study, we analyzed the PTP1B expression in normal breast tissue, primary breast cells and the breast epithelial cell line D492. In normal breast tissue and primary breast cells, PTP1B is widely expressed in both epithelial and stromal cells, with highest expression in myoepithelial cells and fibroblasts. PTP1B is widely expressed in branching structures generated by D492 when cultured in 3D reconstituted basement membrane (3D rBM). Inhibition of PTP1B in D492 and another mammary epithelial cell line HMLE resulted in reduced cell proliferation and induction of anoikis. These changes were seen when cells were cultured both in monolayer and in 3D rBM. PTP1B inhibition affected cell attachment, expression of cell adhesion proteins and actin polymerization. Moreover, epithelial to mesenchymal transition (EMT) sensitized cells to PTP1B inhibition. A mesenchymal sublines of D492 and HMLE (D492M and HMLEmes) were more sensitive to PTP1B inhibition than D492 and HMLE. Reversion of D492M to an epithelial state using miR-200c-141 restored resistance to detachment induced by PTP1B inhibition. In conclusion, we have shown that PTP1B is widely expressed in the human breast gland with highest expression in myoepithelial cells and fibroblasts. Inhibition of PTP1B in D492 and HMLE affects cell-cell adhesion and induces anoikis-like effects. Finally, cells with an EMT phenotype are more sensitive to PTP1B inhibitors making PTP1B a potential candidate for further studies as a target for drug development in cancer involving the EMT phenotype.
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Lowe Syndrome protein OCRL1 supports maturation of polarized epithelial cells.
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Adam G Grieve
Full Text Available Mutations in the inositol polyphosphate 5-phosphatase OCRL1 cause Lowe Syndrome, leading to cataracts, mental retardation and renal failure. We noted that cell types affected in Lowe Syndrome are highly polarized, and therefore we studied OCRL1 in epithelial cells as they mature from isolated individual cells into polarized sheets and cysts with extensive communication between neighbouring cells. We show that a proportion of OCRL1 targets intercellular junctions at the early stages of their formation, co-localizing both with adherens junctional components and with tight junctional components. Correlating with this distribution, OCRL1 forms complexes with junctional components α-catenin and zonula occludens (ZO-1/2/3. Depletion of OCRL1 in epithelial cells growing as a sheet inhibits maturation; cells remain flat, fail to polarize apical markers and also show reduced proliferation. The effect on shape is reverted by re-expressed OCRL1 and requires the 5'-phosphatase domain, indicating that down-regulation of 5-phosphorylated inositides is necessary for epithelial development. The effect of OCRL1 in epithelial maturation is seen more strongly in 3-dimensional cultures, where epithelial cells lacking OCRL1 not only fail to form a central lumen, but also do not have the correct intracellular distribution of ZO-1, suggesting that OCRL1 functions early in the maturation of intercellular junctions when cells grow as cysts. A role of OCRL1 in junctions of polarized cells may explain the pattern of organs affected in Lowe Syndrome.
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Effect of the Human Amniotic Membrane on Liver Regeneration in Rats
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Mesut Sipahi
2015-01-01
Full Text Available Introduction. Operations are performed for broader liver surgery indications for a better understanding of hepatic anatomy/physiology and developments in operation technology. Surgery can cure some patients with liver metastasis of some tumors. Nevertheless, postoperative liver failure is the most feared complication causing mortality in patients who have undergone excision of a large liver mass. The human amniotic membrane has regenerative effects. Thus, we investigated the effects of the human amniotic membrane on regeneration of the resected liver. Methods. Twenty female Wistar albino rats were divided into control and experimental groups and underwent a 70% hepatectomy. The human amniotic membrane was placed over the residual liver in the experimental group. Relative liver weight, histopathological features, and biochemical parameters were assessed on postoperative day 3. Results. Total protein and albumin levels were significantly lower in the experimental group than in the control group. No difference in relative liver weight was observed between the groups. Hepatocyte mitotic count was significantly higher in the experimental group than in the control group. Hepatic steatosis was detected in the experimental group. Conclusion. Applying the amniotic membrane to residual liver adversely affected liver regeneration. However, mesenchymal stem cell research has the potential to accelerate liver regeneration investigations.
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MUC-1-ESA+ progenitor cells in normal benign and malignant human breast epithelial cells
Lu, Xinquan; Li, Huixiang; Xu, Kejia; Nesland, Jahn M.; Suo, Zhenhe
2009-01-01
The existence of mammary epithelial stem/progenitor cells has been demonstrated in MUC-1-/ ESA+ subpopulations of breast epithelial cells. However, knowledge about the expression and localization in benign and malignant breast lesions is unknown. Using a double-staining immunohistochemistry method, we investigated MUC-1-/ESA+ cells in 10 normal breast tissues, 49 cases with fibrocystic disease, 40 fibroadenomas, 36 invasive ductal carcinomas and the breast cancer ce...
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International Nuclear Information System (INIS)
Li Hongzhen; Zhou Jianjun; Miki, Jun; Furusato, Bungo; Gu Yongpeng; Srivastava, Shiv; McLeod, David G.; Vogel, Jonathan C.; Rhim, Johng S.
2008-01-01
Understanding prostate stem cells may provide insight into the origin of prostate cancer. Primary cells have been cultured from human prostate tissue but they usually survive only 15-20 population doublings before undergoing senescence. We report here that RC-170N/h/clone 7 cells, a clonal cell line from hTERT-immortalized primary non-malignant tissue-derived human prostate epithelial cell line (RC170N/h), retain multipotent stem cell properties. The RC-170N/h/clone 7 cells expressed a human embryonic stem cell marker, Oct-4, and potential prostate epithelial stem cell markers, CD133, integrin α2β1 hi and CD44. The RC-170N/h/clone 7 cells proliferated in KGM and Dulbecco's Modified Eagle Medium with 10% fetal bovine serum and 5 μg/ml insulin (DMEM + 10% FBS + Ins.) medium, and differentiated into epithelial stem cells that expressed epithelial cell markers, including CK5/14, CD44, p63 and cytokeratin 18 (CK18); as well as the mesenchymal cell markers, vimentin, desmin; the neuron and neuroendocrine cell marker, chromogranin A. Furthermore the RC170 N/h/clone 7 cells differentiated into multi tissues when transplanted into the sub-renal capsule and subcutaneously of NOD-SCID mice. The results indicate that RC170N/h/clone 7 cells retain the properties of multipotent stem cells and will be useful as a novel cell model for studying the mechanisms of human prostate stem cell differentiation and transformation
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Y-27632, a ROCK Inhibitor, Promoted Limbal Epithelial Cell Proliferation and Corneal Wound Healing.
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Chi-Chin Sun
Full Text Available Transplantation of ex vivo cultured limbal epithelial cells is proven effective in restoring limbal stem cell deficiency. The present study aimed to investigate the promoting effect of Y-27632 on limbal epithelial cell proliferation. Limbal explants isolated from human donor eyes were expanded three weeks on culture dishes and outgrowth of epithelial cells was subsequently subcultured for in vitro experiments. In the presence of Y-27632, the ex vivo limbal outgrowth was accelerated, particularly the cells with epithelial cell-like morphology. Y-27632 dose-dependently promoted the proliferation of in vitro cultured human limbal epithelial cells as examined by phase contrast microscopy and luminescent cell-viability assay 30 hours after the treatment. The colony forming efficacy determined 7 days after the treatment was enhanced by Y-27632 also in a dose-dependent manner. The number of p63- or Ki67-positive cells was dose-dependently increased in Y-27632-treated cultures as detected by immunofluorescent staining and western blotanalysis. Cell cycle analysis by flow cytometric method revealed an increase in S-phase proliferating cells. The epithelial woundclosure rate was shown to be faster in experimental group received topical treatment withY-27632 than the sham control using a rat corneal wounding model. These resultsdemonstrate that Y-27632 can promote both the ex vivo and in vitro proliferation oflimbal epithelial cell proliferation. The in vivo enhanced epithelial wound healingfurther implies that the Y-27632 may act as a new strategy for treating limbal stem cell deficiency.
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International Nuclear Information System (INIS)
Yang, Min; Ma, Bo; Shao, Hanshuang; Clark, Amanda M.; Wells, Alan
2016-01-01
Metastatic progression of breast cancer involves phenotypic plasticity of the carcinoma cells moving between epithelial and mesenchymal behaviors. During metastatic seeding and dormancy, even highly aggressive carcinoma cells take on an E-cadherin-positive epithelial phenotype that is absent from the emergent, lethal metastatic outgrowths. These phenotypes are linked to the metastatic microenvironment, though the specific cells and induction signals are still to be deciphered. Recent evidence suggests that macrophages impact tumor progression, and may alter the balance between cancer cell EMT and MErT in the metastatic microenvironment. Here we explore the role of M1/M2 macrophages in epithelial-mesenchymal plasticity of breast cancer cells by coculturing epithelial and mesenchymal cells lines with macrophages. We found that after polarizing the THP-1 human monocyte cell line, the M1 and M2-types were stable and maintained when co-cultured with breast cancer cells. Surprisingly, M2 macrophages may conferred a growth advantage to the epithelial MCF-7 cells, with these cells being driven to a partial mesenchymal phenotypic as indicated by spindle morphology. Notably, E-cadherin protein expression is significantly decreased in MCF-7 cells co-cultured with M2 macrophages. M0 and M1 macrophages had no effect on the MCF-7 epithelial phenotype. However, the M1 macrophages impacted the highly aggressive mesenchymal-like MDA-MB-231 breast cancer cells to take on a quiescent, epithelial phenotype with re-expression of E-cadherin. The M2 macrophages if anything exacerbated the mesenchymal phenotype of the MDA-MB-231 cells. Our findings demonstrate M2 macrophages might impart outgrowth and M1 macrophages may contribute to dormancy behaviors in metastatic breast cancer cells. Thus EMT and MErT are regulated by selected macrophage phenotype in the liver metastatic microenvironment. These results indicate macrophage could be a potential therapeutic target for limiting death due
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Apoptosis resistance in epithelial tumors is mediated by tumor-cell-derived interleukin-4.
Todaro, M; Lombardo, Y; Francipane, M G; Alea, M Perez; Cammareri, P; Iovino, F; Di Stefano, A B; Di Bernardo, C; Agrusa, A; Condorelli, G; Walczak, H; Stassi, G
2008-04-01
We investigated the mechanisms involved in the resistance to cell death observed in epithelial cancers. Here, we identify that primary epithelial cancer cells from colon, breast and lung carcinomas express high levels of the antiapoptotic proteins PED, cFLIP, Bcl-xL and Bcl-2. These cancer cells produced interleukin-4 (IL-4), which amplified the expression levels of these antiapoptotic proteins and prevented cell death induced upon exposure to TRAIL or other drug agents. IL-4 blockade resulted in a significant decrease in the growth rate of epithelial cancer cells and sensitized them, both in vitro and in vivo, to apoptosis induction by TRAIL and chemotherapy via downregulation of the antiapoptotic factors PED, cFLIP, Bcl-xL and Bcl-2. Furthermore, we provide evidence that exogenous IL-4 was able to upregulate the expression levels of these antiapoptotic proteins and potently stabilized the growth of normal epithelial cells rendering them apoptosis resistant. In conclusion, IL-4 acts as an autocrine survival factor in epithelial cells. Our results indicate that inhibition of IL-4/IL-4R signaling may serve as a novel treatment for epithelial cancers.
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The amniotic band syndrome: antenatal sonographic diagnosis and potential pitfalls.
Mahony, B S; Filly, R A; Callen, P W; Golbus, M S
1985-05-01
Amniotic band syndrome causes a variety of fetal malformations involving the limbs, craniofacial region, and trunk. Six prenatally diagnosed cases of amniotic band syndrome are discussed. The diagnosis was based on sonographic visualization of either amniotic sheets or bands associated with fetal deformation or deformities in nonembryologic distributions known to characterize the amniotic band syndrome. Seven additional cases are considered in which an aberrant sheet of tissue with a free edge was visualized within the amniotic cavity but no restriction of fetal motion or subsequent deformity was demonstrated.
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γδ T cells in homeostasis and host defence of epithelial barrier tissues
DEFF Research Database (Denmark)
Nielsen, Morten M.; Witherden, Deborah A.; Havran, Wendy L.
2017-01-01
Epithelial surfaces line the body and provide a crucial interface between the body and the external environment. Tissue-resident epithelial γδ T cells represent a major T cell population in the epithelial tissues and are ideally positioned to carry out barrier surveillance and aid in tissue...... homeostasis and repair. In this Review, we focus on the intraepithelial γδ T cell compartment of the two largest epithelial tissues in the body — namely, the epidermis and the intestine — and provide a comprehensive overview of the crucial contributions of intraepithelial γδ T cells to tissue integrity...
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Cytotoxic effects of air freshener biocides in lung epithelial cells.
Kwon, Jung-Taek; Lee, Mimi; Seo, Gun-Baek; Kim, Hyun-Mi; Shim, Ilseob; Lee, Doo-Hee; Kim, Taksoo; Seo, Jung Kwan; Kim, Pilje; Choi, Kyunghee
2013-09-01
This study evaluated the cytotoxicity of mixtures of citral (CTR) and either benzisothiazolinone (BIT, Mix-CTR-BIT) or triclosan (TCS, Mix-CTR-TCS) in human A549 lung epithelial cells. We investigated the effects of various mix ratios of these common air freshener ingredients on cell viability, cell proliferation, reactive oxygen species (ROS) generation, and DNA damage. Mix-CTR-BIT and Mix-CTR-TCS significantly decreased the viability of lung epithelial cells and inhibited cell growth in a dose-dependent manner. In addition, both mixtures increased ROS generation, compared to that observed in control cells. In particular, cell viability, growth, and morphology were affected upon increase in the proportion of BIT or TCS in the mixture. However, comet analysis showed that treatment of cells with Mix-CTR-BIT or Mix-CTR-TCS did not increase DNA damage. Taken together, these data suggested that increasing the content of biocides in air fresheners might induce cytotoxicity, and that screening these compounds using lung epithelial cells may contribute to hazard assessment.
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The status of intercellular junctions in established lens epithelial cell lines.
Dave, Alpana; Craig, Jamie E; Sharma, Shiwani
2012-01-01
Cataract is the major cause of vision-related disability worldwide. Mutations in the crystallin genes are the most common known cause of inherited congenital cataract. Mutations in the genes associated with intercellular contacts, such as Nance-Horan Syndrome (NHS) and Ephrin type A receptor-2 (EPHA2), are other recognized causes of congenital cataract. The EPHA2 gene has been also associated with age-related cataract, suggesting that intercellular junctions are important in not only lens development, but also in maintaining lens transparency. The purpose of this study was to analyze the expression and localization of the key cell junction and cytoskeletal proteins, and of NHS and EPHA2, in established lens epithelial cell lines to determine their suitability as model epithelial systems for the functional investigation of genes involved in intercellular contacts and implicated in cataract. The expression and subcellular localization of occludin and zona occludens protein-1 (ZO-1), which are associated with tight junctions; E-cadherin, which is associated with adherence junctions; and the cytoskeletal actin were analyzed in monolayers of a human lens epithelial cell line (SRA 01/04) and a mouse lens epithelial cell line (αTN4). In addition, the expression and subcellular localization of the NHS and EPHA2 proteins were analyzed in these cell lines. Protein or mRNA expression was respectively determined by western blotting or reverse transcription-polymerase chain reaction (RT-PCR), and localization was determined by immunofluorescence labeling. Human SRA 01/04 and mouse αTN4 lens epithelial cells expressed either the proteins of interest or their encoding mRNA. Occludin, ZO-1, and NHS proteins localized to the cellular periphery, whereas E-cadherin, actin, and EPHA2 localized in the cytoplasm in these cell lines. The human SRA 01/04 and mouse αTN4 lens epithelial cells express the key junctional proteins. The localization patterns of these proteins suggest that
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Human thymic epithelial cells express functional HLA-DP molecules
DEFF Research Database (Denmark)
Jørgensen, A; Röpke, C; Nielsen, M
1996-01-01
T lymphocytes, we examined whether human thymic epithelial cells (TEC) expressed HLA-DP molecules. We present evidence that TEC obtained from short time culture express low but significant levels of HLA-DP molecules. The expression of HLA-DP molecules was comparable to or higher than the expression...... of HLA-DP allospecific primed lymphocyte typing (PLT) CD4 T cell lines. IFN-gamma treatment strongly upregulated the HLA-DP allospecific PLT responses whereas other PLT responses remained largely unchanged. In conclusion, these data indicate that human thymus epithelial cells express significant levels...
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Serratia marcescens is injurious to intestinal epithelial cells.
Ochieng, John B; Boisen, Nadia; Lindsay, Brianna; Santiago, Araceli; Ouma, Collins; Ombok, Maurice; Fields, Barry; Stine, O Colin; Nataro, James P
2014-01-01
Diarrhea causes substantial morbidity and mortality in children in low-income countries. Although numerous pathogens cause diarrhea, the etiology of many episodes remains unknown. Serratia marcescens is incriminated in hospital-associated infections, and HIV/AIDS associated diarrhea. We have recently found that Serratia spp. may be found more commonly in the stools of patients with diarrhea than in asymptomatic control children. We therefore investigated the possible enteric pathogenicity of S. marcescens in vitro employing a polarized human colonic epithelial cell (T84) monolayer. Infected monolayers were assayed for bacterial invasion, transepithelial electrical resistance (TEER), cytotoxicity, interleukin-8 (IL-8) release and morphological changes by scanning electron microscopy. We observed significantly greater epithelial cell invasion by S. marcescens compared to Escherichia coli strain HS (p = 0.0038 respectively). Cell invasion was accompanied by reduction in TEER and secretion of IL-8. Lactate dehydrogenase (LDH) extracellular concentration rapidly increased within a few hours of exposure of the monolayer to S. marcescens. Scanning electron microscopy of S. marcescens-infected monolayers demonstrated destruction of microvilli and vacuolization. Our results suggest that S. marcescens interacts with intestinal epithelial cells in culture and induces dramatic alterations similar to those produced by known enteric pathogens.
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Cytological analysis of the epithelial cells in patients with oral candidiasis.
Loss, Rafael; Sandrin, Rodrigo; França, Beatriz Helena Sottile; de Azevedo-Alanis, Luciana Reis; Grégio, Ana Maria Trindade; Machado, Maria Ângela Naval; de Lima, Antonio Adilson Soares
2011-07-01
The aim of this study was to evaluate oral epithelial cells of the oral mucosa infected by Candida albicans using exfoliative cytology. Oral smears were collected from clinically normal-appearing mucosa by liquid-based exfoliative cytology of 60 individuals (30 patients with oral candidiasis and 30 healthy controls matched for age and gender) and analysed for morphologic and cytomorphometric technique. Morphologically, candida-infected epithelial cells exhibited nuclear enlargement, perinuclear rings, discrete orangeophilia, and cytoplasmic vacuoles. The cytomorphometric analysis demonstrated that the cytoplasmic area (CA) of the epithelial cells was diminished in patients undergoing candidiasis as compared to the non-infected controls. In addition, there was an augmentation in nuclear area (NA) and NA/CA area ratio. This study revealed that oral mucosa of patients undergoing candidal infection exhibited significant changes in the size and shape of the oral epithelial cells. © 2010 Blackwell Verlag GmbH.
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Intrinsic pro-angiogenic status of cystic fibrosis airway epithelial cells
International Nuclear Information System (INIS)
Verhaeghe, Catherine; Tabruyn, Sebastien P.; Oury, Cecile; Bours, Vincent; Griffioen, Arjan W.
2007-01-01
Cystic fibrosis is a common genetic disorder characterized by a severe lung inflammation and fibrosis leading to the patient's death. Enhanced angiogenesis in cystic fibrosis (CF) tissue has been suggested, probably caused by the process of inflammation, as similarly described in asthma and chronic bronchitis. The present study demonstrates an intrinsic pro-angiogenic status of cystic fibrosis airway epithelial cells. Microarray experiments showed that CF airway epithelial cells expressed several angiogenic factors such as VEGF-A, VEGF-C, bFGF, and PLGF at higher levels than control cells. These data were confirmed by real-time quantitative PCR and, at the protein level, by ELISA. Conditioned media of these cystic fibrosis cells were able to induce proliferation, migration and sprouting of cultured primary endothelial cells. This report describes for the first time that cystic fibrosis epithelial cells have an intrinsic angiogenic activity. Since excess of angiogenesis is correlated with more severe pulmonary disease, our results could lead to the development of new therapeutic applications
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Loerke, D.; le Duc, Q.; Blonk, I.; Kerstens, A.; Spanjaard, E.; Machacek, M.; Danuser, G.; de Rooij, J.
2012-01-01
The scattering of cultured epithelial cells in response to hepatocyte growth factor (HGF) is a model system that recapitulates key features of metastatic cell behavior in vitro, including disruption of cell-cell adhesions and induction of cell migration. We have developed image analysis tools that
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CCL20, (gamma)(delta) T cells, and IL-22 in corneal epithelial healing
After corneal epithelial abrasion, leukocytes and platelets rapidly enter the corneal stroma, and CCR6 (+) IL-17(+) gamma delta T cells migrate into the epithelium. Gamma delta T-cell-deficient (TCRd(-/-)) mice have significantly reduced inflammation and epithelial wound healing. Epithelial CCL20 mR...
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Millaku, Agron; Drobne, Damjana; Torkar, Matjaz; Novak, Sara; Remškar, Maja; Pipan-Tkalec, Živa
2013-09-15
We provide data obtained by scanning electron microscopy (SEM) and energy dispersive X-ray spectroscopy (EDS) on the interaction of ingested tungsten nanofibers with epithelial cells of the digestive tubes of a test organism Porcellio scaber. Conventional toxicity endpoints including feeding behaviour, weight loss and mortality were also measured in each investigated animal. No toxicity was detected in any of exposed animals after 14 days of feeding on tungsten nanofiber dosed food, but when nanofibers enter the digestive system they can react with epithelial cells of the digestive tubes, becoming physically inserted into the cells. In this way, nanofibers can injure the epithelial cells of digestive gland tubes when they are ingested with food. Our SEM data suggest that peristaltic forces may have an important role, not predicted by in vitro experiments, in the interactions of nanomaterials with digestive intestinal cells. Copyright © 2013 Elsevier B.V. All rights reserved.
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Prenatal diagnosis of Bartter syndrome: amniotic fluid aldosterone.
Rachid, Myriam; Dreux, Sophie; Pean de Ponfilly, Gauthier; Vargas-Poussou, Rosa; Czerkiewicz, Isabelle; Chevenne, Didier; Oury, Jean-François; Deschênes, Georges; Muller, Françoise
2017-04-01
Bartter syndrome is a severe inherited tubulopathy characterized at birth by salt wasting, severe polyuria, dehydration, growth retardation and secondary hyperaldosteronism. Prenatally, the disease is usually discovered following onset of severe polyhydramnios. We studied amniotic fluid aldosterone concentration in cases of Bartter syndrome and in control groups. Amniotic fluid aldosterone was assayed by radioimmunoassay. We undertook a retrospective case-control study based on 36 cases of postnatally diagnosed Bartter syndrome and 144 controls matched for gestational age. Two controls groups were defined: controls with polyhydramnios (n=72) and control without polyhydramnios (n=72). Amniotic fluid aldosterone was compared between the three groups. The median amniotic fluid aldosterone concentration in the Bartter syndrome group (90 pg/mL) did not differ significantly from that in the controls with polyhydramnios (90 pg/mL, p=0.33) or the controls without polyhydramnios (87 pg/mL, p=0.41). In conclusion, amniotic fluid aldosterone assay cannot be used for prenatal diagnosis of Bartter syndrome.
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Directory of Open Access Journals (Sweden)
Guillaume Golovkine
2016-01-01
Full Text Available To achieve systemic infection, bacterial pathogens must overcome the critical and challenging step of transmigration across epithelial barriers. This is particularly true for opportunistic pathogens such as Pseudomonas aeruginosa, an agent which causes nosocomial infections. Despite extensive study, details on the mechanisms used by this bacterium to transmigrate across epithelial tissues, as well as the entry sites it uses, remain speculative. Here, using real-time microscopy and a model epithelial barrier, we show that P. aeruginosa employs a paracellular transmigration route, taking advantage of altered cell-cell junctions at sites of cell division or when senescent cells are expelled from the cell layer. Once a bacterium transmigrates, it is followed by a cohort of bacteria using the same entry point. The basal compartment is then invaded radially from the initial penetration site. Effective transmigration and propagation require type 4 pili, the type 3 secretion system (T3SS and a flagellum, although flagellum-deficient bacteria can occasionally invade the basal compartment from wounded areas. In the basal compartment, the bacteria inject the T3SS toxins into host cells, disrupting the cytoskeleton and focal contacts to allow their progression under the cells. Thus, P. aeruginosa exploits intrinsic host cell processes to breach the epithelium and invade the subcellular compartment.
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Tissue Banking in Malaysia-amniotic membrane
International Nuclear Information System (INIS)
Hashim bin Mohamad; Norimah binti Yusof
1991-01-01
Burn treatment using amniotic membranes in some of our patients initiate our own tissue bank starting with a pilot project on procurement, processing and clinical application of irradiated amniotic membrane. The irradiation of amniotic membrane was made possible by the availability of cobalt source at the Nuclear Energy Agency (UTN). With the technical help from the Inter-national Atomic Energy Agency (IAEA) we soon should be able to embark on bone bank to supply local surgeons. Thus the establishment of tissue bank at our institution will further enhance our programme which will include keratinocytes culture for burn, osteocytes culture for bone replacement as well as the use of animal skin for temporary coverage of open wounds
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A novel method for isolation of epithelial cells from ovine esophagus for tissue engineering.
Macheiner, Tanja; Kuess, Anna; Dye, Julian; Saxena, Amulya K
2014-01-01
The yield of a critical number of basal epithelial cells with high mitotic rates from native tissue is a challenge in the field of tissue engineering. There are many protocols that use enzymatic methods for isolation of epithelial cells with unsatisfactory results for tissue engineering. This study aimed to develop a protocol for isolating a sufficient number of epithelial cells with a high Proliferating Index from ovine esophagus for tissue engineering applications. Esophageal mucosa was pretreated with dispase-collagenase solution and plated on collagen-coated culture dishes. Distinction of the various types of epithelial cells and developmental stages was done with specific primary antibodies to Cytokeratins and to Proliferating Cell Nuclear Antigen (PCNA). Up to approximately 8100 epithelial cells/mm2 of mucosa tissue were found after one week of migration. Cytokeratin 14 (CK 14) was positive identified in cells even after 83 days. At the same time the Proliferating Index was 71%. Our protocol for isolation of basal epithelial cells was successful to yield sufficient numbers of cells predominantly with proliferative character and without noteworthy negative enzymatic affection. The results at this study offer the possibility of generation critical cell numbers for tissue engineering applications.
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Expression of a fms-related oncogene in carcinogen-induced neoplastic epithelial cells
International Nuclear Information System (INIS)
Walker, C.; Nettesheim, P.; Barrett, J.C.; Gilmer, T.M.
1987-01-01
Following carcinogen exposure in vitro, normal rat tracheal epithelial cells are transformed in a multistage process in which the cultured cells become immortal and ultimately, neoplastic. Five cell lines derived from tumors produced by neoplastically transformed rat tracheal epithelial cells were examined for the expression of 11 cellular oncogenes previously implicated in pulmonary or epithelial carcinogenesis. RNA homologous to fms was expressed at a level 5-19 times higher than normal tracheal epithelial cells in three of five of the tumor-derived lines. All three lines expressing high levels of fms-related RNA gave rise to invasive tumors of epithelial origin when injected into nude mice. Increased expression of the fms-related mRNA was not due to gene amplification, and no gene rearrangement was detected by Southern analyses. RNA blot analysis using a 3' v-fms probe detected a 9.5-kilobase message in the three tumor-derived lines, whereas both normal rat aveolar macrophages and the human choriocarcinoma line BeWo expressed a fms transcript of ≅ 4 kilobases. The authors conclude from these data that the gene expressed as a 9.5-kilobase transcript in these neoplastic epithelial cells is a member of a fms-related gene family but may be distinct from the gene that encodes the macrophage colony-stimulating factor (CSF-1) receptor
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Jones, B A; Gores, G J
1997-12-01
Cell death of gastrointestinal epithelial cells occurs by a process referred to as apoptosis. In this review, we succinctly define apoptosis and summarize the role of apoptosis in the physiology and pathophysiology of epithelial cells in the liver, pancreas, and small and large intestine. The physiological mediators regulating apoptosis in gastrointestinal epithelial cells, when known, are discussed. Selected pathophysiological consequences of excessive apoptosis and inhibition of apoptosis are used to illustrate the significance of apoptosis in disease processes. These examples demonstrate that excessive apoptosis may result in epithelial cell atrophy, injury, and dysfunction, whereas inhibition of apoptosis results in hyperplasia and promotes malignant transformation. The specific cellular mechanisms responsible for dysregulation of epithelial cell apoptosis during pathophysiological disturbances are emphasized. Potential future areas of physiological research regarding apoptosis in gastrointestinal epithelia are highlighted when appropriate.
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Oral epithelial cell reaction after exposure to Invisalign plastic material.
Premaraj, Thyagaseely; Simet, Samantha; Beatty, Mark; Premaraj, Sundaralingam
2014-01-01
Invisalign plastic aligners (Align Technology, Santa Clara, Calif) are used to correct malocclusions. The aligners wrap around the teeth and are in contact with gingival epithelium during treatment. The purpose of this study was to evaluate the cellular responses of oral epithelium exposed to Invisalign plastic in vitro. Oral epithelial cells were exposed to eluate obtained by soaking Invisalign plastic in either saline solution or artificial saliva for 2, 4, and 8 weeks. Cells grown in media containing saline solution or saliva served as controls. Morphologic changes were assessed by light microscopy. The 3-[4, 5-dimethythiazol- 2-yl]-2, 5-diphenyl tetrazolium bromide assay and flow cytometry were used to determine cell viability and membrane integrity, respectively. Cellular adhesion and micromotion of epithelial cells were measured in real time by electrical cell-substrate impedance sensing. Cells exposed to saline-solution eluate appeared rounded, were lifted from the culture plates, and demonstrated significantly increased metabolic inactivity or cell death (P <0.05). Saliva eluates did not induce significant changes in cell viability compared with untreated cells. Flow cytometry and electric cell-substrate impedance sensing showed that cells treated with saline-solution eluate exhibited compromised membrane integrity, and reduced cell-to-cell contact and mobility when compared with saliva-eluate treatment. Exposure to Invisalign plastic caused changes in viability, membrane permeability, and adhesion of epithelial cells in a saline-solution environment. Microleakage and hapten formation secondary to compromised epithelial integrity might lead to isocyanate allergy, which could be systemic or localized to gingiva. However, these results suggest that saliva might offer protection. Copyright © 2014 American Association of Orthodontists. Published by Mosby, Inc. All rights reserved.
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Loss of γ-cytoplasmic actin triggers myofibroblast transition of human epithelial cells.
Lechuga, Susana; Baranwal, Somesh; Li, Chao; Naydenov, Nayden G; Kuemmerle, John F; Dugina, Vera; Chaponnier, Christine; Ivanov, Andrei I
2014-10-15
Transdifferentiation of epithelial cells into mesenchymal cells and myofibroblasts plays an important role in tumor progression and tissue fibrosis. Such epithelial plasticity is accompanied by dramatic reorganizations of the actin cytoskeleton, although mechanisms underlying cytoskeletal effects on epithelial transdifferentiation remain poorly understood. In the present study, we observed that selective siRNA-mediated knockdown of γ-cytoplasmic actin (γ-CYA), but not β-cytoplasmic actin, induced epithelial-to-myofibroblast transition (EMyT) of different epithelial cells. The EMyT manifested by increased expression of α-smooth muscle actin and other contractile proteins, along with inhibition of genes responsible for cell proliferation. Induction of EMyT in γ-CYA-depleted cells depended on activation of serum response factor and its cofactors, myocardial-related transcriptional factors A and B. Loss of γ-CYA stimulated formin-mediated actin polymerization and activation of Rho GTPase, which appear to be essential for EMyT induction. Our findings demonstrate a previously unanticipated, unique role of γ-CYA in regulating epithelial phenotype and suppression of EMyT that may be essential for cell differentiation and tissue fibrosis. © 2014 Lechuga, Baranwal, et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).
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Intrauterine Defensive Mechanism of Amniotic Fluid and Fetal Membranes
金山, 尚裕
1994-01-01
To determine the intrauterine defensive role of urinary trypsin inhibitor (UTI), we studied the effects of UTI in amniotic fluid, fetal membranes and myometrium. The level of UTI was 94±34U/ml in neonatal urine (compared to adult urine 8.0±6.0U/ml) and 88±37U/ml in amniotic fluid. This may indicate that the main source of UTI in the amniotic fluid is the fetal urine. UTI was found to be concentrated in vernix, fetal intestine, amniotic membranes and uterine myometrium. Immunostaining of term ...
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Production of arachidonic and linoleic acid metabolites by guinea pig tracheal epithelial cells
International Nuclear Information System (INIS)
Oosthuizen, M.J.; Engels, F.; Van Esch, B.; Henricks, P.A.; Nijkamp, F.P.
1990-01-01
Pulmonary epithelial cells may be responsible for regulating airway smooth muscle function, in part by release of fatty acid-derived mediators. Incubation of isolated guinea pig tracheal epithelial cells with radiolabeled arachidonic acid (AA) leads to the production of 5- and 15-hydroxyeicosatetraenoic acid (5- and 15-HETE) and smaller amounts of leukotriene (LT) B4 and C4 and 12-hydroxyheptadecatrienoic acid (HHT). Epithelial cells also are able to release linoleic acid (LA) metabolites. Incubation with radiolabeled linoleic acid leads to the formation of 9- and 13-hydroxyoctadecadienoic acid (9- and 13-HODE). The biological significance of these mediators produced by epithelial cells is discussed
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Filipovski, Vanja; Kubelka-Sabit, Katerina; Jasar, Dzengis; Janevska, Vesna
2017-08-15
Prostatic carcinoma (PCa) derives from prostatic epithelial cells. However stromal microenvironment, associated with malignant epithelium, also plays a role in prostatic carcinogenesis. Alterations in prostatic stromal cells contribute to the loss of growth control in epithelial cells that lead to progression of PCa. To analyse the differences between Androgen Receptor (AR) expression in both epithelial and stromal cells in PCa and the surrounding benign prostatic hyperplasia (BPH) and to compare the results with tumour grade. Samples from 70 cases of radical prostatectomy specimens were used. The expression and intensity of the signal for AR was analysed in the epithelial and stromal cells of PCa and BPH, and the data was quantified using histological score (H-score). AR showed significantly lower expression in both epithelial and stromal cells of PCa compared to BPH. In PCa a significant positive correlation of AR expression was found between stromal and epithelial cells of PCa. AR expression showed a correlation between the stromal cells of PCa and tumour grade. AR expression is reduced in epithelial and stromal cells of PCa. Expression of AR in stromal cells of PCa significantly correlates with tumour grade.
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Lee, Sarah Y.; Buhimschi, Irina A.; Dulay, Antonette T.; Ali, Unzila A.; Zhao, Guomao; Abdel-Razeq, Sonya S.; Bahtiyar, Mert O.; Thung, Stephen F.; Funai, Edmund F.; Buhimschi, Catalin S.
2013-01-01
Classic IL-6 signaling is conditioned by the transmembrane receptor (IL-6R) and homodimerization of gp130. During trans-signaling, IL-6 binds to soluble IL-6R (sIL-6R) enabling activation of cells expressing solely gp130. Soluble gp130 (sgp130) selectively inhibits IL-6 trans-signaling. To characterize amniotic fluid IL-6 trans-signaling molecules (IL-6, sIL-6R, sgp130) in normal gestations and pregnancies complicated by intra-amniotic inflammation (IAI) we studied 301 women during second trimester (n=39), third trimester (n=40) and preterm labor with intact (n=131, 85 IAI negative & 46 IAI positive) or preterm premature rupture of membranes (PPROM: n=91, 61 IAI negative & 30 IAI positive). ELISA, Western blotting and RT-PCR were used to investigate amniotic fluid, placenta and amniochorion for protein and mRNA expression of sIL-6R, sgp130, IL-6R and gp130. Tissues were immunostained for IL-6R, gp130, CD15+ (polymorphonuclear) and CD3+ (T-cell) inflammatory cells. The ability of sIL-6R and sgp130 to modulate basal and LPS-stimulated release of amniochorion matrix-metalloprotease-9 (MMP-9) was tested ex-vivo. We showed that in physiologic gestations amniotic fluid sgp130 decreases toward term. Amniotic fluid IL-6 and sIL-6R were elevated in IAI whereas sgp130 was decreased in PPROM. Our results suggested that fetal membranes are the probable source of amniotic fluid sIL-6R and sgp130. Immunohistochemistry and RT-PCR revealed increased IL-6R and decreased gp130 expression in amniochorion of women with IAI. Ex-vivo, sIL-6R and LPS augmented amniochorion MMP-9 release whereas sgp130 opposed this effect. We conclude that IL-6 trans-signaling molecules are physiologic constituents of the amniotic fluid regulated by gestational age and inflammation. PPROM likely involves functional loss of sgp130. PMID:21282511
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Directory of Open Access Journals (Sweden)
Jennifer S. Liu
2012-11-01
Full Text Available Variability in signaling pathway activation between neighboring epithelial cells can arise from local differences in the microenvironment, noisy gene expression, or acquired genetic changes. To investigate the consequences of this cell-to-cell variability in signaling pathway activation on coordinated multicellular processes such as morphogenesis, we use DNA-programmed assembly to construct three-dimensional MCF10A microtissues that are mosaic for low-level expression of activated H-Ras. We find two emergent behaviors in mosaic microtissues: cells with activated H-Ras are basally extruded or lead motile multicellular protrusions that direct the collective motility of their wild-type neighbors. Remarkably, these behaviors are not observed in homogeneous microtissues in which all cells express the activated Ras protein, indicating that heterogeneity in Ras activity, rather than the total amount of Ras activity, is critical for these processes. Our results directly demonstrate that cell-to-cell variability in pathway activation within local populations of epithelial cells can drive emergent behaviors during epithelial morphogenesis.
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The fate of epithelial cells in the human large intestine.
Barkla, D H; Gibson, P R
1999-08-01
One hundred and forty biopsies of the colon and rectum, collected during routine colonoscopies of 51 patients aged 19 to 74 years, were examined using light microscopy and transmission and scanning electron microscopy. The results indicated that surface epithelial cells undergo apoptosis, passing through fenestrations in the basement membrane to where they enter the lamina propria and are taken up by macrophages; and it is hypothesized that apoptotic cells are carried through the fenestrations on a current of fluid. The study also found that epithelial cells positioned over the crypts are better attached and more robust than those more distant from the crypt opening; and it is further hypothesized that, after reaching the top of the crypts, some goblet cells cease secreting mucus and pass onto the surface compartment of absorptive cells. An unexpected finding was that the lower regions of the crypts commonly contain isolated necrotic colonocytes. Apoptotic cells were rarely observed in the crypt epithelium. The findings of this study support the "recycling" model of epithelial cell death in the surface compartment of the human colon.
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Xie, Chunfeng; Jin, Jianliang; Lv, Xianhui; Tao, Jianguo; Wang, Rong; Miao, Dengshun
2015-01-01
To determine whether transplanted amniotic membrane mesenchymal stem cells (AMSCs) ameliorated the premature senescent phenotype of Bmi-1-deficient mice, postnatal 2-day-old Bmi-1−/− mice were injected intraperitoneally with the second-passage AMSCs from amniotic membranes of β-galactosidase (β-gal) transgenic mice or wild-type (WT) mice labeled with DiI. Three reinjections were given, once every seven days. Phenotypes of 5-week-old β-gal+ AMSC-transplanted or 6-week-old DiI+ AMSC-transplanted Bmi-1−/− mice were compared with vehicle-transplanted Bmi-1−/− and WT mice. Vehicle-transplanted Bmi-1−/− mice displayed growth retardation and premature aging with decreased cell proliferation and increased cell apoptosis; a decreased ratio and dysmaturity of lymphocytic series; premature osteoporosis with reduced osteogenesis and increased adipogenesis; redox imbalance and DNA damage in multiple organs. Transplanted AMSCs carried Bmi-1 migrated into multiple organs, proliferated and differentiated into multiple tissue cells, promoted growth and delayed senescence in Bmi-1−/− transplant recipients. The dysmaturity of lymphocytic series were ameliorated, premature osteoporosis were rescued by promoting osteogenesis and inhibiting adipogenesis, the oxidative stress and DNA damage in multiple organs were inhibited by the AMSC transplantation in Bmi-1−/− mice. These findings indicate that AMSC transplantation ameliorated the premature senescent phenotype of Bmi-1-deficient mice and could be a novel therapy to delay aging and prevent aging-associated degenerative diseases. PMID:26370922
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International Nuclear Information System (INIS)
Baltzell, Kimberly A; Moghadassi, Michelle; Rice, Terri; Sison, Jennette D; Wrensch, Margaret
2008-01-01
Past studies have shown that women with abnormal cytology or epithelial cells in nipple aspirate fluid (NAF) have an increased relative risk (RR) of breast cancer when compared to women from whom NAF was attempted but not obtained (non-yielders). This study analyzed NAF results from a group of women seen in a breast clinic between 1970–1991 (N = 2480). Our analysis presented here is an aggregate of two sub-groups: women with questionnaire data (n = 712) and those with NAF visits beginning in 1988 (n = 238), the year in which cancer case information was uniformly collected in California. Cytological classification was determined for a group of 946 women using the most abnormal epithelial cytology observed in fluid specimens. Breast cancer incidence and mortality status was determined through June 2006 using data from the California Cancer Registry, California Vital Statistics and self-report. We estimated odd ratios (ORs) for breast cancer using logistic regression analysis, adjusting for age. We analyzed breast cancer risk related to abnormality of NAF cytology using non-yielders as the referent group and breast cancer risk related to the presence or absence of epithelial cells in NAF, using non-yielders/fluid without epithelial cells as the referent group. Overall, 10% (93) of the 946 women developed breast cancer during the follow-up period. Age-adjusted ORs and 95% confidence intervals (C.I.) compared to non-yielders were 1.4 (0.3 to 6.4), 1.7 (0.9 to 3.5), and 2.0 (1.1 to 3.6) for women with fluid without epithelial cells, normal epithelial cells and hyperplasia/atypia, respectively. Comparing the presence or absence of epithelial cells in NAF, women with epithelial cells present in NAF were more likely to develop breast cancer than non-yielders or women with fluid without epithelial cells (RR = 1.9, 1.2 to 3.1). These results support previous findings that 1) women with abnormal epithelial cells in NAF have an increased risk of breast cancer when compared to
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Allergen recognition by innate immune cells: critical role of dendritic and epithelial cells
Directory of Open Access Journals (Sweden)
Fabian eSalazar
2013-11-01
Full Text Available Allergy is an exacerbated response of the immune system against non-self-proteins called allergens and is typically characterized by biased type-2 T helper cell and deleterious IgE mediated immune responses. The allergic cascade starts with the recognition of allergens by antigen presenting cells, mainly dendritic cells, culminating in mast cell sensitization and triggering. Dendritic cells have been demonstrated to play a crucial role in orchestrating allergic diseases. Using different C-type lectin receptors dendritic cells are able to recognize and internalize a number of allergens from diverse sources leading to sensitization. Furthermore, there is increasing evidence highlighting the role of epithelial cells in triggering and modulating immune responses to allergens. As well as providing a physical barrier, epithelial cells can interact with allergens and influence dendritic cells behaviour through the release of a number of Th2 promoting cytokines. In this review we will summarise current understanding of how allergens are recognised by dendritic cells and epithelial cells and what are the consequences of such interaction in the context of allergic sensitisation and downstream events leading to allergic inflammation. Better understanding of the molecular mechanisms of allergen recognition and associated signalling pathways could enable developing more effective therapeutic strategies that target the initial steps of allergic sensitisation hence hindering development or progression of allergic diseases.
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Energy Technology Data Exchange (ETDEWEB)
Millaku, Agron, E-mail: agron.mi@hotmail.com [Limnos-Company for Applied Ecology Ltd, Podlimbarskega 31, 1000 Ljubljana (Slovenia); Drobne, Damjana [University of Ljubljana, Biotechnical Faculty, Department of Biology, Večna pot 111, 1000 Ljubljana (Slovenia); Centre of Excellence, Advanced Materials and Technologies for the Future (CO NAMASTE), Jamova cesta 39, 1000 Ljubljana (Slovenia); Centre of Excellence, Nanoscience and Nanotechnology (Nanocentre), Jamova cesta 39, 1000 Ljubljana (Slovenia); Torkar, Matjaz [Institute of Metals and Technology IMT, Lepi pot 11, 1000 Ljubljana (Slovenia); Jožef Stefan Institute, Condensed Matter Physics Department, Jamova cesta 39, 1000 Ljubljana (Slovenia); Novak, Sara [University of Ljubljana, Biotechnical Faculty, Department of Biology, Večna pot 111, 1000 Ljubljana (Slovenia); Remškar, Maja [Jožef Stefan Institute, Condensed Matter Physics Department, Jamova cesta 39, 1000 Ljubljana (Slovenia); Pipan-Tkalec, Živa [University of Ljubljana, Biotechnical Faculty, Department of Biology, Večna pot 111, 1000 Ljubljana (Slovenia)
2013-09-15
Graphical abstract: Scanning electron microscopy is particularly well suited to the observation of nanofibre/cell interaction in the endothelial cells lining the hepatopancreas. (a) Tungsten oxide nanofibres, (b) test organism Porcellio scaber and schematic appearance of digestive tubes, (c) digestive tube (hepatopancreas) prepared for SEM investigation, (d) digestive gland cells (C) with nanofibres (NF) embedded in the cell membrane and (e) nanofibres inserted deeply in the cells and damaged nanofibres due to peristalsis. -- Highlights: • Tungsten oxide nanofibres react physically with digestive gland epithelial cells in Porcellio scaber. • Physical peristaltic forces of lead to insertion of nanofibres into the cells. • No toxic responses as measured by conventional toxicity biomarkers were detected. • Physical interactions were observed in a majority of the investigated animals. -- Abstract: We provide data obtained by scanning electron microscopy (SEM) and energy dispersive X-ray spectroscopy (EDS) on the interaction of ingested tungsten nanofibers with epithelial cells of the digestive tubes of a test organism Porcellio scaber. Conventional toxicity endpoints including feeding behaviour, weight loss and mortality were also measured in each investigated animal. No toxicity was detected in any of exposed animals after 14 days of feeding on tungsten nanofiber dosed food, but when nanofibers enter the digestive system they can react with epithelial cells of the digestive tubes, becoming physically inserted into the cells. In this way, nanofibers can injure the epithelial cells of digestive gland tubes when they are ingested with food. Our SEM data suggest that peristaltic forces may have an important role, not predicted by in vitro experiments, in the interactions of nanomaterials with digestive intestinal cells.
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International Nuclear Information System (INIS)
Millaku, Agron; Drobne, Damjana; Torkar, Matjaz; Novak, Sara; Remškar, Maja; Pipan-Tkalec, Živa
2013-01-01
Graphical abstract: Scanning electron microscopy is particularly well suited to the observation of nanofibre/cell interaction in the endothelial cells lining the hepatopancreas. (a) Tungsten oxide nanofibres, (b) test organism Porcellio scaber and schematic appearance of digestive tubes, (c) digestive tube (hepatopancreas) prepared for SEM investigation, (d) digestive gland cells (C) with nanofibres (NF) embedded in the cell membrane and (e) nanofibres inserted deeply in the cells and damaged nanofibres due to peristalsis. -- Highlights: • Tungsten oxide nanofibres react physically with digestive gland epithelial cells in Porcellio scaber. • Physical peristaltic forces of lead to insertion of nanofibres into the cells. • No toxic responses as measured by conventional toxicity biomarkers were detected. • Physical interactions were observed in a majority of the investigated animals. -- Abstract: We provide data obtained by scanning electron microscopy (SEM) and energy dispersive X-ray spectroscopy (EDS) on the interaction of ingested tungsten nanofibers with epithelial cells of the digestive tubes of a test organism Porcellio scaber. Conventional toxicity endpoints including feeding behaviour, weight loss and mortality were also measured in each investigated animal. No toxicity was detected in any of exposed animals after 14 days of feeding on tungsten nanofiber dosed food, but when nanofibers enter the digestive system they can react with epithelial cells of the digestive tubes, becoming physically inserted into the cells. In this way, nanofibers can injure the epithelial cells of digestive gland tubes when they are ingested with food. Our SEM data suggest that peristaltic forces may have an important role, not predicted by in vitro experiments, in the interactions of nanomaterials with digestive intestinal cells
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Endogenous Sheet-Averaged Tension Within a Large Epithelial Cell Colony.
Dumbali, Sandeep P; Mei, Lanju; Qian, Shizhi; Maruthamuthu, Venkat
2017-10-01
Epithelial cells form quasi-two-dimensional sheets that function as contractile media to effect tissue shape changes during development and homeostasis. Endogenously generated intrasheet tension is a driver of such changes, but has predominantly been measured in the presence of directional migration. The nature of epithelial cell-generated forces transmitted over supracellular distances, in the absence of directional migration, is thus largely unclear. In this report, we consider large epithelial cell colonies which are archetypical multicell collectives with extensive cell-cell contacts but with a symmetric (circular) boundary. Using the traction force imbalance method (TFIM) (traction force microscopy combined with physical force balance), we first show that one can determine the colony-level endogenous sheet forces exerted at the midline by one half of the colony on the other half with no prior assumptions on the uniformity of the mechanical properties of the cell sheet. Importantly, we find that this colony-level sheet force exhibits large variations with orientation-the difference between the maximum and minimum sheet force is comparable to the average sheet force itself. Furthermore, the sheet force at the colony midline is largely tensile but the shear component exhibits significantly more variation with orientation. We thus show that even an unperturbed epithelial colony with a symmetric boundary shows significant directional variation in the endogenous sheet tension and shear forces that subsist at the colony level.
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Pesavento, Patricia; Liu, Hongwei; Ossiboff, Robert J; Stucker, Karla M; Heymer, Anna; Millon, Lee; Wood, Jason; van der List, Deborah; Parker, John S L
2009-04-01
Mucosal epithelial cells are the primary targets for many common viral pathogens of cats. Viral infection of epithelia can damage or disrupt the epithelial barrier that protects underlying tissues. In vitro cell culture systems are an effective means to study how viruses infect and disrupt epithelial barriers, however no true continuous or immortalized feline epithelial cell culture lines are available. A continuous cell culture of feline mammary epithelial cells (FMEC UCD-04-2) that forms tight junctions with high transepithelial electrical resistance (>2000Omegacm(-1)) 3-4 days after reaching confluence was characterized. In addition, it was shown that FMECs are susceptible to infection with feline calicivirus (FCV), feline herpesvirus (FHV-1), feline coronavirus (FeCoV), and feline panleukopenia virus (FPV). These cells will be useful for studies of feline viral disease and for in vitro studies of feline epithelia.
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Regulation of osteogenesis of human amniotic mesenchymal stem cells by sodium butyrate.
Fan, Xiaoting; Li, Lei; Ye, Zhaoyang; Zhou, Yan; Tan, Wen-Song
2018-04-01
Human amniotic membrane-derived mesenchymal stem cells (hAMSCs) draw great interests for regenerative medicine due to convenient availability and low immunogenicity. However, suboptimal culture conditions limit their application. In recent years, small molecules have proven powerful in regulating stem cell fates and can be applied to stimulate their function. In the present study, the impacts of sodium butyrate (NaBu), a histone deacetylase inhibitor (HDACi), on hAMSCs were investigated. It was shown that NaBu at a low concentration inhibited cell proliferation by arresting cell cycle at G0/G1 rather than inducing apoptosis. When NaBu was supplemented at a concentration of generated and the expression of osteogenesis-related genes (ALP, Runx2, Opn, and Ocn) and proteins (Col1a1, OPN, OCN, Runx2, and TAZ) were both significantly enhanced. However, a higher concentration (1.0 mM) and longer exposure time (14 days) of NaBu showed no such effects, which may be partially attributed to both the increased expression of histone deacetylase 8 (HDAC8) and reduced level of H3K9-Ace, thus leading to the transcriptional inhibition during osteogenesis. Further, it was indicated that ERK might be involved in the stimulatory effects of NaBu. These findings may be helpful to develop an efficient culture process for hAMSCs towards bone regeneration. © 2018 International Federation for Cell Biology.
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Sunpaweravong, S; Sunpaweravong, P; Sathitruangsak, C; Mai, S
2016-05-01
Telomeres are repetitive nucleotide sequences (TTAGGG)n located at the ends of chromosomes that function to preserve chromosomal integrity and prevent terminal end-to-end fusions. Telomere loss or dysfunction results in breakage-bridge-fusion cycles, aneuploidy, gene amplification and chromosomal rearrangements, which can lead to genomic instability and promote carcinogenesis. Evaluating the hypothesis that changes in telomeres contribute to the development of esophageal squamous cell carcinoma (ESCC) and to determine whether there are differences between young and old patients, we compared the three-dimensional (3D) nuclear telomere architecture in ESCC tumor cells with that of normal epithelial cells obtained from the same patient. Patients were equally divided by age into two groups, one comprising those less than 45 years of age and the other consisting of those over 80 years of age. Tumor and normal epithelial cells located at least 10 cm from the border of the tumor were biopsied in ESCC patients. Hematoxylin and eosin staining was performed for each sample to confirm and identify the cancer and normal epithelial cells. This study was based on quantitative 3D fluorescence in situ hybridization (Q-FISH), 3D imaging and 3D analysis of paraffin-embedded slides. The 3D telomere architecture data were computer analyzed using 100 nuclei per slide. The following were the main parameters compared: the number of signals (number of telomeres), signal intensity (telomere length), number of telomere aggregates, and nuclear volume. Tumor and normal epithelial samples from 16 patients were compared. The normal epithelial cells had more telomere signals and higher intensities than the tumor cells, with P-values of P architecture and found no statistically significant differences in any parameter tested between the young and old patients in either the tumor or epithelial cells. The 3D nuclear telomeric signature was able to detect differences in telomere architecture
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Prenatal diagnosis of amniotic band syndrome
Directory of Open Access Journals (Sweden)
Laxmi Devi Padmanabhan
2016-01-01
Full Text Available Amniotic band can cause a broad spectrum of anomalies ranging from simple band constrictions to major craniofacial and visceral defects. It can cause significant neonatal morbidity. Accurate diagnosis will help in the management of the present pregnancy and in counseling with regard to future pregnancies. Here we report three cases of amniotic band syndrome detected in the prenatal period.
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UV-induced changes in cell cycle and gene expression within rabbit lens epithelial cells
Energy Technology Data Exchange (ETDEWEB)
Sidjanin, D. [Northern Illinois Univ., De Kalb, IL (United States). Dept. of Biological Sciences; Grdina, D. [Argonne National Lab., IL (United States); Woloschak, G.E. [Northern Illinois Univ., De Kalb, IL (United States). Dept. of Biological Sciences
1994-11-01
Damage to lens epithelial cells is a probable initiation process in cataract formation induced by ultraviolet radiation. These experiments investigated the ability of 254 nm radiation on cell cycle progression and gene expression in rabbit lens epithelial cell line N/N1003A. No changes in expression of c-fos, c-jun, alpha- tubulin, or vimentin was observed following UV exposure. Using flow cytometry, an accumulation of cells in G1/S phase of the cell cycle 1 hr following exposure. The observed changes in gene expression, especially the decreased histone transcripts reported here may play a role in UV induced inhibition of cell cycle progression.
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Differential sensitivity of epithelial cells to extracellular matrix in polarity establishment.
Directory of Open Access Journals (Sweden)
Shigenobu Yonemura
Full Text Available Establishment of apical-basal polarity is crucial for epithelial sheets that form a compartment in the body, which function to maintain the environment in the compartment. Effects of impaired polarization are easily observed in three-dimensional (3-D culture systems rather than in two-dimensional (2-D culture systems. Although the mechanisms for establishing the polarity are not completely understood, signals from the extracellular matrix (ECM are considered to be essential for determining the basal side and eventually generating polarity in the epithelial cells. To elucidate the common features and differences in polarity establishment among various epithelial cells, we analyzed the formation of epithelial apical-basal polarity using three cell lines of different origin: MDCK II cells (dog renal tubules, EpH4 cells (mouse mammary gland, and R2/7 cells (human colon expressing wild-type α-catenin (R2/7 α-Cate cells. These cells showed clear apical-basal polarity in 2-D cultures. In 3-D cultures, however, each cell line displayed different responses to the same ECM. In MDCK II cells, spheroids with a single lumen formed in both Matrigel and collagen gel. In R2/7 α-Cate cells, spheroids showed similar apical-basal polarity as that seen in MDCK II cells, but had multiple lumens. In EpH4 cells, the spheroids displayed an apical-basal polarity that was opposite to that seen in the other two cell types in both ECM gels, at least during the culture period. On the other hand, the three cell lines showed the same apical-basal polarity both in 2-D cultures and in 3-D cultures using the hanging drop method. The three lines also had similar cellular responses to ECM secreted by the cells themselves. Therefore, appropriate culture conditions should be carefully determined in advance when using various epithelial cells to analyze cell polarity or 3-D morphogenesis.
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Active Vertex Model for cell-resolution description of epithelial tissue mechanics.
Barton, Daniel L; Henkes, Silke; Weijer, Cornelis J; Sknepnek, Rastko
2017-06-01
We introduce an Active Vertex Model (AVM) for cell-resolution studies of the mechanics of confluent epithelial tissues consisting of tens of thousands of cells, with a level of detail inaccessible to similar methods. The AVM combines the Vertex Model for confluent epithelial tissues with active matter dynamics. This introduces a natural description of the cell motion and accounts for motion patterns observed on multiple scales. Furthermore, cell contacts are generated dynamically from positions of cell centres. This not only enables efficient numerical implementation, but provides a natural description of the T1 transition events responsible for local tissue rearrangements. The AVM also includes cell alignment, cell-specific mechanical properties, cell growth, division and apoptosis. In addition, the AVM introduces a flexible, dynamically changing boundary of the epithelial sheet allowing for studies of phenomena such as the fingering instability or wound healing. We illustrate these capabilities with a number of case studies.
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Lens epithelial cell apoptosis is an early event in the development of UVB-induced cataract.
Li, W C; Spector, A
1996-01-01
Epidemiological and experimental studies have revealed that exposure to UV can induce cataractogenesis. To investigate the mechanism of this induction, viability of the lens epithelial cells from UVB-treated rat lenses were examined. Irradiation of the cultured rat lenses with 8 J/s/m2 UVB for 60 min triggers lens epithelial cell apoptosis as determined by terminal deoxyribonucleotide transferase (TdT) labeling and DNA fragmentation assays. The apoptotic lens epithelial cells were initially found in the equatorial region and then quickly appeared in both equatorial and central regions. The percentage of apoptotic cells continuously increased during the postirradiation incubation. After a 5-h post-UVB incubation, more than 50% of the lens epithelial cells were apoptotic. By 24 h, all of the lens epithelial cells in the irradiated lenses were dead through apoptosis. Associated with this apoptotic process is a large upregulation of the proto-oncogene, c-fos. Opacification appears to follow the death of lens epithelial cells occurring first in the equatorial region and then in the central area. This is also true of classical cataract parameters such as non-protein thiol and wet weight, which are significantly modified only after appreciable epithelial cell apoptosis. Together, these results suggest that the rapid apoptotic death of the lens epithelial cells induced by UVB initiates cataract development.
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Polarity, cell division, and out-of-equilibrium dynamics control the growth of epithelial structures
Cerruti, Benedetta; Puliafito, Alberto; Shewan, Annette M.; Yu, Wei; Combes, Alexander N.; Little, Melissa H.; Chianale, Federica; Primo, Luca; Serini, Guido; Mostov, Keith E.; Celani, Antonio
2013-01-01
The growth of a well-formed epithelial structure is governed by mechanical constraints, cellular apico-basal polarity, and spatially controlled cell division. Here we compared the predictions of a mathematical model of epithelial growth with the morphological analysis of 3D epithelial structures. In both in vitro cyst models and in developing epithelial structures in vivo, epithelial growth could take place close to or far from mechanical equilibrium, and was determined by the hierarchy of time-scales of cell division, cell–cell rearrangements, and lumen dynamics. Equilibrium properties could be inferred by the analysis of cell–cell contact topologies, and the nonequilibrium phenotype was altered by inhibiting ROCK activity. The occurrence of an aberrant multilumen phenotype was linked to fast nonequilibrium growth, even when geometric control of cell division was correctly enforced. We predicted and verified experimentally that slowing down cell division partially rescued a multilumen phenotype induced by altered polarity. These results improve our understanding of the development of epithelial organs and, ultimately, of carcinogenesis. PMID:24145168
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Yang, Zhengtao; Fu, Yunhe; Gong, Pengtao; Zheng, Jingtong; Liu, Li; Yu, Yuqiang; Li, Jianhua; Li, He; Yang, Ju; Zhang, Xichen
2015-08-01
Cryptosporidium parvum (C. parvum) is an intestinal parasite that causes diarrhea in neonatal calves. It results in significant morbidity of neonatal calves and economic losses for producers worldwide. Innate resistance against C. parvum is thought to depend on engagement of pattern recognition receptors. However, the role of innate responses to C. parvum has not been elucidated in bovine. The aim of this study was to evaluate the role of TLRs in host-cell responses during C. parvum infection of cultured bovine intestinal epithelial cells. The expressions of TLRs in bovine intestinal epithelial cells were detected by qRT-PCR. To determine which, if any, TLRs may play a role in the response of bovine intestinal epithelial cells to C. parvum, the cells were stimulated with C. parvum and the expression of TLRs were tested by qRT-PCR. The expression of NF-κB was detected by western blotting. Further analyses were carried out in bovine TLRs transfected HEK293 cells and by TLRs-DN transfected bovine intestinal epithelial cells. The results showed that bovine intestinal epithelial cells expressed all known TLRs. The expression of TLR2 and TLR4 were up-regulated when bovine intestinal epithelial cells were treated with C. parvum. Meanwhile, C. parvum induced IL-8 production in TLR2 or TLR4/MD-2 transfected HEK293 cells. Moreover, C. parvum induced NF-κB activation and cytokine expression in bovine intestinal epithelial cells. The induction of NF-κB activation and cytokine expression by C. parvum were reduced in TLR2-DN and TLR4-DN transfected cells. The results showed that bovine intestinal epithelial cells expressed all known TLRs, and bovine intestinal epithelial cells recognized and responded to C. parvum via TLR2 and TLR4. Copyright © 2015 Elsevier Ltd. All rights reserved.
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Mechanisms of Cell Polarity-Controlled Epithelial Homeostasis and Immunity in the Intestine.
Klunder, Leon J; Faber, Klaas Nico; Dijkstra, Gerard; van IJzendoorn, Sven C D
2017-07-05
Intestinal epithelial cell polarity is instrumental to maintain epithelial homeostasis and balance communications between the gut lumen and bodily tissue, thereby controlling the defense against gastrointestinal pathogens and maintenance of immune tolerance to commensal bacteria. In this review, we highlight recent advances with regard to the molecular mechanisms of cell polarity-controlled epithelial homeostasis and immunity in the human intestine. Copyright © 2017 Cold Spring Harbor Laboratory Press; all rights reserved.
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Amniotic fluid index predicts the relief of variable decelerations after amnioinfusion bolus.
Spong, C Y; McKindsey, F; Ross, M G
1996-10-01
Our purpose was to determine whether intrapartum amniotic fluid index before amnioinfusion can be used to predict response to therapeutic amnioinfusion. Intrapartum patients (n = 85) with repetitive variable decelerations in fetal heart rate that necessitated amnioinfusion (10 ml/min for 60 minutes) underwent determination of amniotic fluid index before and after bolus amnioinfusion. The fetal heart tracing was scored (scorer blinded to amniotic fluid index values) for number and characteristics of variable decelerations before and 1 hour after initiation of amnioinfusion. The amnioinfusion was considered successful if it resulted in a decrease of > or = 50% in total number of variable decelerations or a decrease of > or = 50% in the rate of atypical or severe variable decelerations after administration of the bolus. Spontaneous vaginal births before completion of administration of the bolus (n = 18) were excluded from analysis. The probability of success of amnioinfusion in relation to amniotic fluid index was analyzed with the chi(2) test for progressive sequence. The mean amniotic fluid index before amnioinfusion was 6.2 +/- 3.3 cm. An amniotic fluid index of amnioinfusion decreased with increasing amniotic fluid index before amnioinfusion (76% [16/21] when initial amniotic fluid index was 0 to 4 cm, 63% [17/27] when initial amniotic fluid index was 4 to 8 cm, 44% [7/16] when initial amniotic fluid index was 8 to 12 cm, and 33% [1/3] when initial amniotic fluid index was > 12 cm, p = 0.03). The incidence of nuchal cords or true umbilical cord knots increased in relation to amniotic fluid index before amnioinfusion. Amniotic fluid index before amnioinfusion can be used to predict the success of amnioinfusion for relief of variable decelerations in fetal heart rate. Failure of amnioinfusion at a high amniotic fluid index before amnioinfusion may be explained by the increased prevalence of nuchal cords or true knots in the umbilical cord.
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Establishment of three-dimensional cultures of human pancreatic duct epithelial cells
International Nuclear Information System (INIS)
Gutierrez-Barrera, Angelica M.; Menter, David G.; Abbruzzese, James L.; Reddy, Shrikanth A.G.
2007-01-01
Three-dimensional (3D) cultures of epithelial cells offer singular advantages for studies of morphogenesis or the role of cancer genes in oncogenesis. In this study, as part of establishing a 3D culture system of pancreatic duct epithelial cells, we compared human pancreatic duct epithelial cells (HPDE-E6E7) with pancreatic cancer cell lines. Our results show, that in contrast to cancer cells, HPDE-E6E7 organized into spheroids with what appeared to be apical and basal membranes and a luminal space. Immunostaining experiments indicated that protein kinase Akt was phosphorylated (Ser473) and CTMP, a negative Akt regulator, was expressed in both HPDE-E6E7 and cancer cells. However, a nuclear pool of CTMP was detectable in HPDE-E6E7 cells that showed a dynamic concentrated expression pattern, a feature that further distinguished HPDE-E637 cells from cancer cells. Collectively, these data suggest that 3D cultures of HPDE-E6E7 cells are useful for investigating signaling and morphological abnormalities in pancreatic cancer cells
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Modeling Alveolar Epithelial Cell Behavior In Spatially Designed Hydrogel Microenvironments
Lewis, Katherine Jean Reeder
The alveolar epithelium consists of two cell phenotypes, elongated alveolar type I cells (AT1) and rounded alveolar type II cells (ATII), and exists in a complex three-dimensional environment as a polarized cell layer attached to a thin basement membrane and enclosing a roughly spherical lumen. Closely surrounding the alveolar cysts are capillary endothelial cells as well as interstitial pulmonary fibroblasts. Many factors are thought to influence alveolar epithelial cell differentiation during lung development and wound repair, including physical and biochemical signals from the extracellular matrix (ECM), and paracrine signals from the surrounding mesenchyme. In particular, disrupted signaling between the alveolar epithelium and local fibroblasts has been implicated in the progression of several pulmonary diseases. However, given the complexity of alveolar tissue architecture and the multitude of signaling pathways involved, designing appropriate experimental platforms for this biological system has been difficult. In order to isolate key factors regulating cellular behavior, the researcher ideally should have control over biophysical properties of the ECM, as well as the ability to organize multiple cell types within the scaffold. This thesis aimed to develop a 3D synthetic hydrogel platform to control alveolar epithelial cyst formation, which could then be used to explore how extracellular cues influence cell behavior in a tissue-relevant cellular arrangement. To accomplish this, a poly(ethylene glycol) (PEG) hydrogel network containing enzymatically-degradable crosslinks and bioadhesive pendant peptides was employed as a base material for encapsulating primary alveolar epithelial cells. First, an array of microwells of various cross-sectional shapes was photopatterned into a PEG gel containing photo-labile crosslinks, and primary ATII cells were seeded into the wells to examine the role of geometric confinement on differentiation and multicellular arrangement
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Amniotic fluid embolism and isolated coagulopathy: atypical presentation of amniotic fluid embolism.
LENUS (Irish Health Repository)
Awad, I T
2012-02-03
A 41-year-old multigravida presented at 32 weeks of gestation with polyhydramnios and an anencephalic fetus. Abnormal bleeding as a result of disseminated intravascular coagulation complicated an emergency Caesarean section for severe abdominal pain thought to be due to uterine rupture. Massive transfusion with blood products was necessary and the abdomen packed to control bleeding. The patient was transferred to the intensive care unit where she made a slow but complete recovery. Amniotic fluid embolism with atypical presentation of isolated coagulopathy is the likely diagnosis in this case. The case serves to demonstrate that amniotic fluid embolism may present with symptoms and signs other than the classical pattern of dyspnoea, cyanosis and hypotension.
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The genetic and molecular basis of bacterial invasion of epithelial cells
African Journals Online (AJOL)
Invasion of epithelial cells was demonstrated to be triggered by invasion plasmid antigens B, C, and D ( IpaB, IpaC and IpaD ) which is accomplished by intracellular spread gene icsA. The invasion of epithelial cells by some individual species of bacteria were also reviewed.Yersinia enterocolitica invasiveness was shown ...
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File list: Unc.Utr.50.AllAg.Fallopian_tube_secretory_epithelial_cell [Chip-atlas[Archive
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File list: Unc.Utr.20.AllAg.Fallopian_tube_secretory_epithelial_cell [Chip-atlas[Archive
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File list: Unc.Utr.05.AllAg.Fallopian_tube_secretory_epithelial_cell [Chip-atlas[Archive
Lifescience Database Archive (English)
Full Text Available Unc.Utr.05.AllAg.Fallopian_tube_secretory_epithelial_cell hg19 Unclassified Uterus Fallopian tube secret...ory epithelial cell http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/Unc.Utr.05.AllAg.Fallopian_tube_secretory_epithelial_cell.bed ...
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File list: Unc.Utr.10.AllAg.Fallopian_tube_secretory_epithelial_cell [Chip-atlas[Archive
Lifescience Database Archive (English)
Full Text Available Unc.Utr.10.AllAg.Fallopian_tube_secretory_epithelial_cell hg19 Unclassified Uterus Fallopian tube secret...ory epithelial cell http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/Unc.Utr.10.AllAg.Fallopian_tube_secretory_epithelial_cell.bed ...
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Robust cell tracking in epithelial tissues through identification of maximum common subgraphs.
Kursawe, Jochen; Bardenet, Rémi; Zartman, Jeremiah J; Baker, Ruth E; Fletcher, Alexander G
2016-11-01
Tracking of cells in live-imaging microscopy videos of epithelial sheets is a powerful tool for investigating fundamental processes in embryonic development. Characterizing cell growth, proliferation, intercalation and apoptosis in epithelia helps us to understand how morphogenetic processes such as tissue invagination and extension are locally regulated and controlled. Accurate cell tracking requires correctly resolving cells entering or leaving the field of view between frames, cell neighbour exchanges, cell removals and cell divisions. However, current tracking methods for epithelial sheets are not robust to large morphogenetic deformations and require significant manual interventions. Here, we present a novel algorithm for epithelial cell tracking, exploiting the graph-theoretic concept of a 'maximum common subgraph' to track cells between frames of a video. Our algorithm does not require the adjustment of tissue-specific parameters, and scales in sub-quadratic time with tissue size. It does not rely on precise positional information, permitting large cell movements between frames and enabling tracking in datasets acquired at low temporal resolution due to experimental constraints such as phototoxicity. To demonstrate the method, we perform tracking on the Drosophila embryonic epidermis and compare cell-cell rearrangements to previous studies in other tissues. Our implementation is open source and generally applicable to epithelial tissues. © 2016 The Authors.
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A method for isolating identifying and culturing of rat trachea-bronchia epithelial cells
International Nuclear Information System (INIS)
Cui Fengmei; Su Shibiao; Nie Jihua; Li Bingyan; Tong Jian
2005-01-01
Objective: To explore a method for isolating identifying and culturing the rat trachea-bronchia epithelial cells. Methods: The rat trachea-bronchia epithelial cells were isolated by digestion with pronase and brushing with cell brush, identified using confocul and cultured in entire F12 media with no serum. Results: With this method, cells in high purity and high viability could be obtained, and about 10 6 cells per rat. The cells grow well in entire F12 media with no serum. Conclusion: The method is useful for isolating rate trachea-bronchia epithelial cells and the entire F12 media with no serum is effective for culturing. (authors)
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Directory of Open Access Journals (Sweden)
Han Hu
Full Text Available BACKGROUND: The objective of this study was to establish a culture system and elucidate the unique characteristics of a bovine mammary epithelial cell line in vitro. METHODOLOGY: Mammary tissue from a three year old lactating dairy cow (ca. 100 d relative to parturition was used as a source of the epithelial cell line, which was cultured in collagen-coated tissue culture dishes. Fibroblasts and epithelial cells successively grew and extended from the culturing mammary tissue at the third day. Pure epithelial cells were obtained by passages culture. PRINCIPAL FINDINGS: The strong positive immunostaining to cytokeratin 18 suggested that the resulting cell line exhibited the specific character of epithelial cells. Epithelial cells cultured in the presence of 10% FBS, supraphysiologic concentrations of insulin, and hydrocortisone maintained a normal diploid chromosome modal number of 2n=60. Furthermore, they were capable of synthesizing beta-casein (CSN2, acetyl-CoA carboxylase-alpha (ACACA and butyrophilin (BTN1A1. An important finding was that frozen preservation in a mixture of 90% FBS and 10% DMSO did not influence the growth characteristics, chromosome number, or protein secretion of the isolated epithelial cell line. CONCLUSIONS: The obtained mammary epithelial cell line had normal morphology, growth characteristics, cytogenetic and secretory characteristics, thus, it might represent an useful tool for studying the function of Chinese Holstein dairy cows mammary epithelial cell (CMECs.
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Directory of Open Access Journals (Sweden)
Z. Liu
2010-05-01
Full Text Available Epithelium, a highly dynamic system, plays a key role in the homeostasis of the intestine. However, thus far a human intestinal epithelial cell line has not been established in many countries. Fetal tissue was selected to generate viable cell cultures for its sterile condition, effective generation, and differentiated character. The purpose of the present study was to culture human intestinal epithelial cells by a relatively simple method. Thermolysin was added to improve the yield of epithelial cells, while endothelin-3 was added to stimulate their growth. By adding endothelin-3, the achievement ratio (viable cell cultures/total cultures was enhanced to 60% of a total of 10 cultures (initiated from 8 distinct fetal small intestines, allowing the generation of viable epithelial cell cultures. Western blot, real-time PCR and immunofluorescent staining showed that cytokeratins 8, 18 and mouse intestinal mucosa-1/39 had high expression levels in human intestinal epithelial cells. Differentiated markers such as sucrase-isomaltase, aminopeptidase N and dipeptidylpeptidase IV also showed high expression levels in human intestinal epithelial cells. Differentiated human intestinal epithelial cells, with the expression of surface markers (cytokeratins 8, 18 and mouse intestinal mucosa-1/39 and secretion of cytokines (sucrase-isomaltase, aminopeptidase N and dipeptidylpeptidase IV, may be cultured by the thermolysin and endothelin-3 method and maintained for at least 20 passages. This is relatively simple, requiring no sophisticated techniques or instruments, and may have a number of varied applications.
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Three-dimensional epithelial tissues generated from human embryonic stem cells.
Hewitt, Kyle J; Shamis, Yulia; Carlson, Mark W; Aberdam, Edith; Aberdam, Daniel; Garlick, Jonathan A
2009-11-01
The use of pluripotent human embryonic stem (hES) cells for tissue engineering may provide advantages over traditional sources of progenitor cells because of their ability to give rise to multiple cell types and their unlimited expansion potential. We derived cell populations with properties of ectodermal and mesenchymal cells in two-dimensional culture and incorporated these divergent cell populations into three-dimensional (3D) epithelial tissues. When grown in specific media and substrate conditions, two-dimensional cultures were enriched in cells (EDK1) with mesenchymal morphology and surface markers. Cells with a distinct epithelial morphology (HDE1) that expressed cytokeratin 12 and beta-catenin at cell junctions became the predominant cell type when EDK1 were grown on surfaces enriched in keratinocyte-derived extracellular matrix proteins. When these cells were incorporated into the stromal and epithelial tissue compartments of 3D tissues, they generated multilayer epithelia similar to those generated with foreskin-derived epithelium and fibroblasts. Three-dimensional tissues demonstrated stromal cells with morphologic features of mature fibroblasts, type IV collagen deposition in the basement membrane, and a stratified epithelium that expressed cytokeratin 12. By deriving two distinct cell lineages from a common hES cell source to fabricate complex tissues, it is possible to explore environmental cues that will direct hES-derived cells toward optimal tissue form and function.
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File list: Pol.Utr.50.AllAg.Fallopian_tube_secretory_epithelial_cell [Chip-atlas[Archive
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File list: Oth.Utr.50.AllAg.Fallopian_tube_secretory_epithelial_cell [Chip-atlas[Archive
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File list: Oth.Utr.05.AllAg.Fallopian_tube_secretory_epithelial_cell [Chip-atlas[Archive
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File list: DNS.Utr.05.AllAg.Fallopian_tube_secretory_epithelial_cell [Chip-atlas[Archive
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File list: Pol.Utr.20.AllAg.Fallopian_tube_secretory_epithelial_cell [Chip-atlas[Archive
Lifescience Database Archive (English)
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File list: DNS.Utr.50.AllAg.Fallopian_tube_secretory_epithelial_cell [Chip-atlas[Archive
Lifescience Database Archive (English)
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File list: DNS.Utr.20.AllAg.Fallopian_tube_secretory_epithelial_cell [Chip-atlas[Archive
Lifescience Database Archive (English)
Full Text Available DNS.Utr.20.AllAg.Fallopian_tube_secretory_epithelial_cell hg19 DNase-seq Uterus Fallopian tube secret...ory epithelial cell http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/DNS.Utr.20.AllAg.Fallopian_tube_secretory_epithelial_cell.bed ...
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File list: Pol.Utr.10.AllAg.Fallopian_tube_secretory_epithelial_cell [Chip-atlas[Archive
Lifescience Database Archive (English)
Full Text Available Pol.Utr.10.AllAg.Fallopian_tube_secretory_epithelial_cell hg19 RNA polymerase Uterus Fallopian tube secret...ory epithelial cell http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/Pol.Utr.10.AllAg.Fallopian_tube_secretory_epithelial_cell.bed ...
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File list: DNS.Utr.10.AllAg.Fallopian_tube_secretory_epithelial_cell [Chip-atlas[Archive
Lifescience Database Archive (English)
Full Text Available DNS.Utr.10.AllAg.Fallopian_tube_secretory_epithelial_cell hg19 DNase-seq Uterus Fallopian tube secret...ory epithelial cell http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/DNS.Utr.10.AllAg.Fallopian_tube_secretory_epithelial_cell.bed ...
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File list: Oth.Utr.20.AllAg.Fallopian_tube_secretory_epithelial_cell [Chip-atlas[Archive
Lifescience Database Archive (English)
Full Text Available Oth.Utr.20.AllAg.Fallopian_tube_secretory_epithelial_cell hg19 TFs and others Uterus Fallopian tube secret...ory epithelial cell http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/Oth.Utr.20.AllAg.Fallopian_tube_secretory_epithelial_cell.bed ...
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From cells to tissue: A continuum model of epithelial mechanics
Ishihara, Shuji; Marcq, Philippe; Sugimura, Kaoru
2017-08-01
A two-dimensional continuum model of epithelial tissue mechanics was formulated using cellular-level mechanical ingredients and cell morphogenetic processes, including cellular shape changes and cellular rearrangements. This model incorporates stress and deformation tensors, which can be compared with experimental data. Focusing on the interplay between cell shape changes and cell rearrangements, we elucidated dynamical behavior underlying passive relaxation, active contraction-elongation, and tissue shear flow, including a mechanism for contraction-elongation, whereby tissue flows perpendicularly to the axis of cell elongation. This study provides an integrated scheme for the understanding of the orchestration of morphogenetic processes in individual cells to achieve epithelial tissue morphogenesis.
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Anxiolytic-Like Actions of Fatty Acids Identified in Human Amniotic Fluid
Directory of Open Access Journals (Sweden)
Rosa Isela García-Ríos
2013-01-01
Full Text Available Eight fatty acids (C12–C18 were previously identified in human amniotic fluid, colostrum, and milk in similar proportions but different amounts. Amniotic fluid is well known to be the natural environment for development in mammals. Interestingly, amniotic fluid and an artificial mixture of fatty acids contained in amniotic fluid produce similar anxiolytic-like actions in Wistar rats. We explored whether the lowest amount of fatty acids contained in amniotic fluid with respect to colostrum and milk produces such anxiolytic-like effects. Although a trend toward a dose-response effect was observed, only an amount of fatty acids that was similar to amniotic fluid fully mimicked the effect of diazepam (2 mg/kg, i.p. in the defensive burying test, an action devoid of effects on locomotor activity and motor coordination. Our results confirm that the amount of fatty acids contained in amniotic fluid is sufficient to produce anxiolytic-like effects, suggesting similar actions during intrauterine development.
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Renal epithelial cells can release ATP by vesicular fusion
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Randi G Bjaelde
2013-09-01
Full Text Available Renal epithelial cells have the ability to release nucleotides as paracrine factors. In the intercalated cells of the collecting duct, ATP is released by connexin30 (cx30, which is selectively expressed in this cell type. However, ATP is released by virtually all renal epithelia and the aim of the present study was to identify possible alternative nucleotide release pathways in a renal epithelial cell model. We used MDCK (type1 cells to screen for various potential ATP release pathways. In these cells, inhibition of the vesicular H+-ATPases (bafilomycin reduced both the spontaneous and hypotonically (80%-induced nucleotide release. Interference with vesicular fusion using N-ethylamide markedly reduced the spontaneous nucleotide release, as did interference with trafficking from the endoplasmic reticulum to the Golgi apparatus (brefeldin A1 and vesicular transport (nocodazole. These findings were substantiated using a siRNA directed against SNAP-23, which significantly reduced spontaneous ATP release. Inhibition of pannexin and connexins did not affect the spontaneous ATP release in this cell type, which consists of ∼90% principal cells. TIRF-microscopy of either fluorescently-labeled ATP (MANT-ATP or quinacrine-loaded vesicles, revealed that spontaneous release of single vesicles could be promoted by either hypoosmolality (50% or ionomycin. This vesicular release decreased the overall cellular fluorescence by 5.8% and 7.6% respectively. In summary, this study supports the notion that spontaneous and induced ATP release can occur via exocytosis in renal epithelial cells.
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Amniocar as a proliferative medium for mesenchymal cells
Directory of Open Access Journals (Sweden)
V. V. Chestkov
2014-01-01
Full Text Available Objectives. To develop the Amniocar nutrient medium that contains fetal calf serum (FCS and growth factors cocktail for mass cultivation of human fibroblasts. To study proliferative activity of the medium on cultures of HUVEC cells of mesenchymal origin and mesenchymal stromal cells, as well as on cell culture of human amniotic fluid.Materials and methods. Determination of the rate of accumulation of the cellular mass and cell morphology in the course of cultivation of cells of various histogenesis in the Amniocar medium and nutrient medium that contains 10 % of FCS.Results. It has been demonstrated that the Amniocar medium is prevalent as compared to the standard DMEM medium with 10 % of FCS by 2 to 5 times for cultivation of skin fibroblasts, HUVEC, and mesenchymal stem cells. The Amniocar medium increased the quantity of endothelial cells that enter mitosis and maintained the culture of HUVEC cells with prolonged passaging in vitro. Clonal cultivation of human amniotic fluid cells in the Amniocar medium secured development of colonies of both fibroblast and epithelial type.Conclusions. Proliferative Amniocar medium is efficient for mass cultivation of various cells of mesenchymal origin and can be used for diagnostic purposes in medical genetics, oncology, etc.
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Human airway xenograft models of epithelial cell regeneration
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Puchelle Edith
2000-10-01
Full Text Available Abstract Regeneration and restoration of the airway epithelium after mechanical, viral or bacterial injury have a determinant role in the evolution of numerous respiratory diseases such as chronic bronchitis, asthma and cystic fibrosis. The study in vivo of epithelial regeneration in animal models has shown that airway epithelial cells are able to dedifferentiate, spread, migrate over the denuded basement membrane and progressively redifferentiate to restore a functional respiratory epithelium after several weeks. Recently, human tracheal xenografts have been developed in immunodeficient severe combined immunodeficiency (SCID and nude mice. In this review we recall that human airway cells implanted in such conditioned host grafts can regenerate a well-differentiated and functional human epithelium; we stress the interest in these humanized mice in assaying candidate progenitor and stem cells of the human airway mucosa.
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Sphingolipid trafficking and protein sorting in epithelial cells
Slimane, TA; Hoekstra, D
2002-01-01
Sphingolipids represent a minor, but highly dynamic subclass of lipids in all eukaryotic cells. They are involved in functions that range from structural protection to signal transduction and protein sorting, and participate in lipid raft assembly. In polarized epithelial cells, which display an
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γδ T cells in homeostasis and host defence of epithelial barrier tissues.
Nielsen, Morten M; Witherden, Deborah A; Havran, Wendy L
2017-12-01
Epithelial surfaces line the body and provide a crucial interface between the body and the external environment. Tissue-resident epithelial γδ T cells represent a major T cell population in the epithelial tissues and are ideally positioned to carry out barrier surveillance and aid in tissue homeostasis and repair. In this Review, we focus on the intraepithelial γδ T cell compartment of the two largest epithelial tissues in the body - namely, the epidermis and the intestine - and provide a comprehensive overview of the crucial contributions of intraepithelial γδ T cells to tissue integrity and repair, host homeostasis and protection in the context of the symbiotic relationship with the microbiome and during pathogen clearance. Finally, we describe epithelium-specific butyrophilin-like molecules and briefly review their emerging role in selectively shaping and regulating epidermal and intestinal γδ T cell repertoires.
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Massingham, Lauren J; Johnson, Kirby L; Scholl, Thomas M; Slonim, Donna K; Wick, Heather C; Bianchi, Diana W
2014-09-01
Turner syndrome is a sex chromosome aneuploidy with characteristic malformations. Amniotic fluid, a complex biological material, could contribute to the understanding of Turner syndrome pathogenesis. In this pilot study, global gene expression analysis of cell-free RNA in amniotic fluid supernatant was utilized to identify specific genes/organ systems that may play a role in Turner syndrome pathophysiology. Cell-free RNA from amniotic fluid of five mid-trimester Turner syndrome fetuses and five euploid female fetuses matched for gestational age was extracted, amplified, and hybridized onto Affymetrix(®) U133 Plus 2.0 arrays. Significantly differentially regulated genes were identified using paired t tests. Biological interpretation was performed using Ingenuity Pathway Analysis and BioGPS gene expression atlas. There were 470 statistically significantly differentially expressed genes identified. They were widely distributed across the genome. XIST was significantly down-regulated (p Turner syndrome transcriptome from other aneuploidies we previously studied. Manual curation of the differentially expressed gene list identified genes of possible pathologic significance, including NFATC3, IGFBP5, and LDLR. Transcriptomic differences in the amniotic fluid of Turner syndrome fetuses are due to genome-wide dysregulation. The hematologic/immune system differences may play a role in early-onset autoimmune dysfunction. Other genes identified with possible pathologic significance are associated with cardiac and skeletal systems, which are known to be affected in females with Turner syndrome. The discovery-driven approach described here may be useful in elucidating novel mechanisms of disease in Turner syndrome.
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Alveolar type II epithelial cell dysfunction in rat experimental hepatopulmonary syndrome (HPS.
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Wenli Yang
Full Text Available The hepatopulmonary syndrome (HPS develops when pulmonary vasodilatation leads to abnormal gas exchange. However, in human HPS, restrictive ventilatory defects are also observed supporting that the alveolar epithelial compartment may also be affected. Alveolar type II epithelial cells (AT2 play a critical role in maintaining the alveolar compartment by producing four surfactant proteins (SPs, SP-A, SP-B, SP-C and SP-D which also facilitate alveolar repair following injury. However, no studies have evaluated the alveolar epithelial compartment in experimental HPS. In this study, we evaluated the alveolar epithelial compartment and particularly AT2 cells in experimental HPS induced by common bile duct ligation (CBDL. We found a significant reduction in pulmonary SP production associated with increased apoptosis in AT2 cells after CBDL relative to controls. Lung morphology showed decreased mean alveolar chord length and lung volumes in CBDL animals that were not seen in control models supporting a selective reduction of alveolar airspace. Furthermore, we found that administration of TNF-α, the bile acid, chenodeoxycholic acid, and FXR nuclear receptor activation (GW4064 induced apoptosis and impaired SP-B and SP-C production in alveolar epithelial cells in vitro. These results imply that AT2 cell dysfunction occurs in experimental HPS and is associated with alterations in the alveolar epithelial compartment. Our findings support a novel contributing mechanism in experimental HPS that may be relevant to humans and a potential therapeutic target.
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Dietary antioxidants protect gut epithelial cells from oxidant-induced apoptosis
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Bobrowski Paul
2001-12-01
Full Text Available Abstract Background The potential of ascorbic acid and two botanical decoctions, green tea and cat's claw, to limit cell death in response to oxidants were evaluated in vitro. Methods Cultured human gastric epithelial cells (AGS or murine small intestinal epithelial cells (IEC-18 were exposed to oxidants – DPPH (3 μM, H2O2 (50 μM, peroxynitrite (300 μM – followed by incubation for 24 hours, with antioxidants (10 μg/ml administered as a 1 hour pretreatment. Cell number (MTT assay and death via apoptosis or necrosis (ELISA, LDH release was determined. The direct interactions between antioxidants and DPPH (100 μM or H2O2 (50 μM were evaluated by spectroscopy. Results The decoctions did not interact with H2O2, but quenched DPPH although less effectively than vitamin C. In contrast, vitamin C was significantly less effective in protecting human gastric epithelial cells (AGS from apoptosis induced by DPPH, peroxynitrite and H2O2 (P 2O2, but green tea was more effective than cat's claw in reducing DPPH-induced apoptosis (P 2O2, and was attenuated both by cat's claw and green tea (P Conclusions These results indicate that dietary antioxidants can limit epithelial cell death in response to oxidant stress. In the case of green tea and cat's claw, the cytoprotective response exceed their inherent ability to interact with the injurious oxidant, suggestive of actions on intracellular pathways regulating cell death.
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Directory of Open Access Journals (Sweden)
Maryam Parvini
2013-10-01
Full Text Available Pluripotent stem cells as the cells with a capacity for self-renewal and differentiation into various specificcell types have been highly regarded in regenerative medicine studies. To repair the eye disease damages, thedifferentiation into retinal pigment epithelial cells of pluripotent stem cells has gained great importance inrecent decades because the inappropriate function of these cells is the main cause of degenerative diseases suchas the age-related macular degeneration. Millions of people in the world suffer this disease.To restore the damaged cells and, finally, to improve the vision, numerous studies have been conducted on usingpluripotent stem cells, their differentiation into retinal pigment epithelial cells, and finally, their applicationin cell therapy. Based on this, many researchers have attempted to produce highly efficient retinal pigmentepithelial cells, such that they show a proper function after transplant, along with the host cells. In this reviewarticle, the importance and the role of pigment epithelial cells, as well as, the studies on the in vitro productionof these cells were examined
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Cigarette smoke alters the secretome of lung epithelial cells.
Mossina, Alessandra; Lukas, Christina; Merl-Pham, Juliane; Uhl, Franziska E; Mutze, Kathrin; Schamberger, Andrea; Staab-Weijnitz, Claudia; Jia, Jie; Yildirim, Ali Ö; Königshoff, Melanie; Hauck, Stefanie M; Eickelberg, Oliver; Meiners, Silke
2017-01-01
Cigarette smoke is the most relevant risk factor for the development of lung cancer and chronic obstructive pulmonary disease. Many of its more than 4500 chemicals are highly reactive, thereby altering protein structure and function. Here, we used subcellular fractionation coupled to label-free quantitative MS to globally assess alterations in the proteome of different compartments of lung epithelial cells upon exposure to cigarette smoke extract. Proteomic profiling of the human alveolar derived cell line A549 revealed the most pronounced changes within the cellular secretome with preferential downregulation of proteins involved in wound healing and extracellular matrix organization. In particular, secretion of secreted protein acidic and rich in cysteine, a matricellular protein that functions in tissue response to injury, was consistently diminished by cigarette smoke extract in various pulmonary epithelial cell lines and primary cells of human and mouse origin as well as in mouse ex vivo lung tissue cultures. Our study reveals a previously unrecognized acute response of lung epithelial cells to cigarette smoke that includes altered secretion of proteins involved in extracellular matrix organization and wound healing. This may contribute to sustained alterations in tissue remodeling as observed in lung cancer and chronic obstructive pulmonary disease. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.
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Comparative effects of metal oxide nanoparticles on human airway epithelial cells and macrophages
International Nuclear Information System (INIS)
Rotoli, Bianca Maria; Bussolati, Ovidio; Costa, Anna Luisa; Blosi, Magda; Di Cristo, Luisana; Zanello, Pier Paolo; Bianchi, Massimiliano G.; Visigalli, Rossana; Bergamaschi, Enrico
2012-01-01
Among nanomaterials of industrial relevance, metal-based nanoparticles (NPs) are widely used, but their effects on airway cells are relatively poorly characterized. To compare the effects of metal NPs on cells representative of the lung-blood barrier, Calu-3 epithelial cells and Raw264.7 macrophages were incubated with three industrially relevant preparations of TiO 2 NPs (size range 4–33 nm), two preparations of CeO 2 NPs (9–36 nm) and CuO NPs (25 nm). While Raw264.7 were grown on standard plasticware, Calu-3 cells were seeded on permeable filters, where they form a high-resistance monolayer, providing an in vitro model of the airway barrier. Metal NPs, obtained from industrial sources, were characterized under the conditions adopted for the biological tests. Cytotoxicity was assessed with resazurin method in both epithelial and macrophage cells, while epithelial barrier permeability was monitored measuring the trans-epithelial electrical resistance (TEER). In macrophages, titania and ceria had no significant effect on viability in the whole range of nominal doses tested (15–240 μg/cm 2 of monolayer), while CuO NPs produced a marked viability loss. Moreover, only CuO NPs, but not the other NPs, lowered TEER of Calu-3 monolayers, pointing to the impairment of the epithelial barrier. TEER decreased by 30 % at the dose of 10 μg/cm 2 of CuO NPs, compared to untreated control, and was abolished at doses ≥80 μg/cm 2 , in strict correlation with changes in cell viability. These results indicate that (1) CuO NPs increase airway epithelium permeability even at relatively low doses and are significantly toxic for macrophages and airway epithelial cells, likely through the release of Cu ions in the medium; (2) TiO 2 and CeO 2 NPs do not affect TEER and exhibit little acute toxicity for airway epithelial cells and macrophages; and (3) TEER measurement can provide a simple method to assess the impairment of in vitro airway epithelial barrier model by manufactured
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Comparative effects of metal oxide nanoparticles on human airway epithelial cells and macrophages
Rotoli, Bianca Maria; Bussolati, Ovidio; Costa, Anna Luisa; Blosi, Magda; Di Cristo, Luisana; Zanello, Pier Paolo; Bianchi, Massimiliano G.; Visigalli, Rossana; Bergamaschi, Enrico
2012-09-01
Among nanomaterials of industrial relevance, metal-based nanoparticles (NPs) are widely used, but their effects on airway cells are relatively poorly characterized. To compare the effects of metal NPs on cells representative of the lung-blood barrier, Calu-3 epithelial cells and Raw264.7 macrophages were incubated with three industrially relevant preparations of TiO2 NPs (size range 4-33 nm), two preparations of CeO2 NPs (9-36 nm) and CuO NPs (25 nm). While Raw264.7 were grown on standard plasticware, Calu-3 cells were seeded on permeable filters, where they form a high-resistance monolayer, providing an in vitro model of the airway barrier. Metal NPs, obtained from industrial sources, were characterized under the conditions adopted for the biological tests. Cytotoxicity was assessed with resazurin method in both epithelial and macrophage cells, while epithelial barrier permeability was monitored measuring the trans-epithelial electrical resistance (TEER). In macrophages, titania and ceria had no significant effect on viability in the whole range of nominal doses tested (15-240 μg/cm2 of monolayer), while CuO NPs produced a marked viability loss. Moreover, only CuO NPs, but not the other NPs, lowered TEER of Calu-3 monolayers, pointing to the impairment of the epithelial barrier. TEER decreased by 30 % at the dose of 10 μg/cm2 of CuO NPs, compared to untreated control, and was abolished at doses ≥80 μg/cm2, in strict correlation with changes in cell viability. These results indicate that (1) CuO NPs increase airway epithelium permeability even at relatively low doses and are significantly toxic for macrophages and airway epithelial cells, likely through the release of Cu ions in the medium; (2) TiO2 and CeO2 NPs do not affect TEER and exhibit little acute toxicity for airway epithelial cells and macrophages; and (3) TEER measurement can provide a simple method to assess the impairment of in vitro airway epithelial barrier model by manufactured nanomaterials.
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Human epithelial cells increase their rigidity with ageing in vitro: direct measurements
International Nuclear Information System (INIS)
Berdyyeva, Tamara K; Woodworth, Craig D; Sokolov, Igor
2005-01-01
The decrease in elasticity of epithelial tissues with ageing contributes to many human diseases. This change was previously attributed to increased crosslinking of extracellular matrix proteins. Here we show that individual human epithelial cells also become significantly more rigid during ageing in vitro. Using atomic force microscopy (AFM), we found that the Young's modulus of viable cells was consistently increased two- to four-fold in older versus younger cells. Direct visualization of the cytoskeleton using a novel method involving the AFM suggested that increased rigidity of ageing cells was due to a higher density of cytoskeletal fibres. Our results identify a unique mechanism that might contribute to the age-related loss of elasticity in epithelial tissues
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Role of Echogenic Amniotic Fluid Particles and Optical Density in ...
African Journals Online (AJOL)
This study was aimed to correlate echogenic amniotic fluid particle size (AFPS) in late third trimester to fetal lung maturity and amniotic fluid optical density (AFOD) at labor. AFPS were measured with specified criteria by real time transabdominal USG (3.5MHz) while Amniotic Fluid Index (AFI) was measured during routine ...
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Directory of Open Access Journals (Sweden)
Dhananjay M Nawandar
2015-10-01
Full Text Available Epstein-Barr virus (EBV is a human herpesvirus associated with B-cell and epithelial cell malignancies. EBV lytically infects normal differentiated oral epithelial cells, where it causes a tongue lesion known as oral hairy leukoplakia (OHL in immunosuppressed patients. However, the cellular mechanism(s that enable EBV to establish exclusively lytic infection in normal differentiated oral epithelial cells are not currently understood. Here we show that a cellular transcription factor known to promote epithelial cell differentiation, KLF4, induces differentiation-dependent lytic EBV infection by binding to and activating the two EBV immediate-early gene (BZLF1 and BRLF1 promoters. We demonstrate that latently EBV-infected, telomerase-immortalized normal oral keratinocyte (NOKs cells undergo lytic viral reactivation confined to the more differentiated cell layers in organotypic raft culture. Furthermore, we show that endogenous KLF4 expression is required for efficient lytic viral reactivation in response to phorbol ester and sodium butyrate treatment in several different EBV-infected epithelial cell lines, and that the combination of KLF4 and another differentiation-dependent cellular transcription factor, BLIMP1, is highly synergistic for inducing lytic EBV infection. We confirm that both KLF4 and BLIMP1 are expressed in differentiated, but not undifferentiated, epithelial cells in normal tongue tissue, and show that KLF4 and BLIMP1 are both expressed in a patient-derived OHL lesion. In contrast, KLF4 protein is not detectably expressed in B cells, where EBV normally enters latent infection, although KLF4 over-expression is sufficient to induce lytic EBV reactivation in Burkitt lymphoma cells. Thus, KLF4, together with BLIMP1, plays a critical role in mediating lytic EBV reactivation in epithelial cells.
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Células-tronco do líquido amniótico Amniotic fluid stem cells
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Sergio P. Bydlowski
2009-05-01
Full Text Available Desde o primeiro isolamento e cultivo de células-tronco embrionárias humanas, há mais de 10 anos, seu uso na pesquisa e terapia foi inibida por considerações éticas complexas e pelo risco de transformação maligna destas células indiferenciadas após transplante no paciente. As células-tronco adultas são eticamente aceitas e o risco de transformação maligna é muito baixo. Entretanto, seu potencial de diferenciação e sua capacidade proliferativa são limitados. Cerca de 6 anos atrás, a descoberta de célulastronco no líquido amniótico que expressavam Oct-4, um marcador específico de pluripotencialidade, com alta capacidade de proliferação e diferenciação, iniciou um novo campo promissor na área das células-tronco. Estas células têm potencial de se diferenciar em células dos três folhetos germinativos. Não formam tumores in vivo e não levantam os questionamentos éticos associados com as células-tronco embrionárias humanas. Futuras investigações revelarão se as células-tronco do líquido amniótico realmente irão representar um tipo intermediário com vantagens em relação tanto às células-tronco embrionárias quanto às adultas. Este artigo faz uma revisão acerca destes tópicos e das características biológicas das células-tronco do líquido amniótico.Since the first successful isolation and cultivation of human embryonic stem cells about 10 years ago, their use for research and therapy has been constrained by complex ethical considerations as well as by the risk of development of malignancies of undifferentiated embryonic stem cells after transplantation into the patient. Adult stem cells are ethically acceptable and the risk of tumor development is low. However, their differentiation potential and proliferative capacity are limited. About 6 years ago, the discovery of Oct-4 expressing amniotic fluid stem cells, a specific marker of pluripotency, with a high proliferative capacity, and multilineage
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International Nuclear Information System (INIS)
Matsuzaki-Horibuchi, Shiori; Yasuda, Takeshi; Sakaguchi, Nagako; Yamaguchi, Yoshihiro; Akashi, Makoto
2015-01-01
One of the important mechanisms for gastrointestinal (GI) injury following high-dose radiation exposure is apoptosis of epithelial cells. X-linked inhibitor of apoptosis (XIAP) and cellular IAP2 (cIAP2) are intrinsic cellular inhibitors of apoptosis. In order to study the effects of exogenously added IAPs on apoptosis in intestinal epithelial cells, we constructed bacterial expression plasmids containing genes of XIAP (full-length, BIR2 domain and BIR3-RING domain with and without mutations of auto-ubiquitylation sites) and cIAP2 proteins fused to a protein-transduction domain (PTD) derived from HIV-1 Tat protein (TAT) and purified these cell-permeable recombinant proteins. When the TAT-conjugated IAPs were added to rat intestinal epithelial cells IEC6, these proteins were effectively delivered into the cells and inhibited apoptosis, even when added after irradiation. Our results suggest that PTD-mediated delivery of IAPs may have clinical potential, not only for radioprotection but also for rescuing the GI system from radiation injuries. (author)
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Urokinase and the intestinal mucosa: evidence for a role in epithelial cell turnover
Gibson, P; Birchall, I; Rosella, O; Albert, V; Finch, C; Barkla, D; Young, G
1998-01-01
Background—The functions of urokinase in intestinal epithelia are unknown. Aims—To determine the relation of urokinase expressed by intestinal epithelial cells to their position in the crypt-villus/surface axis and of mucosal urokinase activity to epithelial proliferative kinetics in the distal colon. Methods—Urokinase expression was examined immunohistochemically in human intestinal mucosa. Urokinase activity was measured colorimetrically in epithelial cells isolated sequ...
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DEFF Research Database (Denmark)
Kiilgaard, Jens Folke; Prause, Jan U; Prause, Michala
2007-01-01
PURPOSE: To study topographical differences in porcine retinal pigment epithelial (RPE) cell proliferation (1) in vivo, after experimental central surgical subretinal injury, and (2) in vitro. METHODS: Domestic pigs underwent either experimental RPE debridement (n = 5), subretinal amniotic membrane...... cells. This observation was true of both types of experimental surgery performed. In vitro, RPE isolates from the pre-equatorial region consistently yielded higher cell densities than did RPE cell isolates from more central parts of the epithelium. This was apparent through the three first passages...
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Directory of Open Access Journals (Sweden)
Dagher Hayat
2010-02-01
Full Text Available Abstract Background Transforming growth factor β1 (TGF-β1-mediated epithelial mesenchymal transition (EMT of alveolar epithelial cells (AEC may contribute to lung fibrosis. Since PPARγ ligands have been shown to inhibit fibroblast activation by TGF-β1, we assessed the ability of the thiazolidinediones rosiglitazone (RGZ and ciglitazone (CGZ to regulate TGF-β1-mediated EMT of A549 cells, assessing changes in cell morphology, and expression of cell adhesion molecules E-cadherin (epithelial cell marker and N-cadherin (mesenchymal cell marker, and collagen 1α1 (COL1A1, CTGF and MMP-2 mRNA. Methods Serum-deprived A549 cells (human AEC cell line were pre-incubated with RGZ and CGZ (1 - 30 μM in the absence or presence of the PPARγ antagonist GW9662 (10 μM before TGFβ-1 (0.075-7.5 ng/ml treatment for up to 72 hrs. Changes in E-cadherin, N-cadherin and phosphorylated Smad2 and Smad3 levels were analysed by Western blot, and changes in mRNA levels including COL1A1 assessed by RT-PCR. Results TGFβ-1 (2.5 ng/ml-induced reductions in E-cadherin expression were associated with a loss of epithelial morphology and cell-cell contact. Concomitant increases in N-cadherin, MMP-2, CTGF and COL1A1 were evident in predominantly elongated fibroblast-like cells. Neither RGZ nor CGZ prevented TGFβ1-induced changes in cell morphology, and PPARγ-dependent inhibitory effects of both ligands on changes in E-cadherin were only evident at submaximal TGF-β1 (0.25 ng/ml. However, both RGZ and CGZ inhibited the marked elevation of N-cadherin and COL1A1 induced by TGF-β1 (2.5 ng/ml, with effects on COL1A1 prevented by GW9662. Phosphorylation of Smad2 and Smad3 by TGF-β1 was not inhibited by RGZ or CGZ. Conclusions RGZ and CGZ inhibited profibrotic changes in TGF-β1-stimulated A549 cells independently of inhibition of Smad phosphorylation. Their inhibitory effects on changes in collagen I and E-cadherin, but not N-cadherin or CTGF, appeared to be PPAR
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Directory of Open Access Journals (Sweden)
Jason Bennett
2016-04-01
Full Text Available Epithelial-mesenchymal transition (EMT, a process which describes the trans-differentiation of epithelial cells into motile mesenchymal cells, is pivotal in stem cell behavior, development and wound healing, as well as contributing to disease processes including fibrosis and cancer progression. Maintenance immunosuppression with calcineurin inhibitors (CNIs has become routine management for renal transplant patient, but unfortunately the nephrotoxicity of these drugs has been well documented. HK-2 cells were exposed to Tacrolimus (FK506 and EMT markers were assessed by RT PCR and western blot. FK506 effects on TGF-β mRNA were assessed by RT PCR and TGF-β secretion was measured by ELISA. The impact of increased TGF-β secretion on Smad signaling pathways was investigated. The impact of inhibition of TGF-β signaling on EMT processes was assessed by scratch-wound assay. The results presented in this study suggest that FK506 initiates EMT processes in the HK-2 cell line, with altered expression of epithelial and myofibroblast markers evident. Additionally, the study demonstrates that FK506 activation of the TGF-β/ SMAD pathways is an essential step in the EMT process. Overall the results demonstrate that EMT is heavily involved in renal fibrosis associated with CNI nephrotoxicity.
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Zhang, Wenyao; Li, Xuezhong; Xu, Tong; Ma, Mengru; Zhang, Yong; Gao, Ming-Qing
2016-11-15
Hypernomic secretion of epithelial cytokines has several effects on stromal cells. The contributions of inflammatory epithelial cells to stromal fibroblasts in bovine mammary glands with mastitis remain poorly understood. Here, we established an inflammatory epithelial cell model of bovine mastitis with gram-negative lipopolysaccharide (LPS) and gram-positive lipoteichoic acid (LTA) bacterial cell wall components. We characterized immune responses of mammary stromal fibroblasts induced by inflammatory epithelial cells. Our results showed that inflammatory epithelial cells affected stromal fibroblast characteristics by increasing inflammatory mediator expression, elevating extracellular matrix protein deposition, decreasing proliferation capacity, and enhancing migration ability. The changes in stromal fibroblast proliferation and migration abilities were mediated by signal molecules, such as WNT signal pathway components. LPS- and LTA-induced inflammatory epithelial cells triggered different immune responses in stromal fibroblasts. Thus, in mastitis, bovine mammary gland stromal fibroblasts were affected by inflammatory epithelial cells and displayed inflammation-specific changes, suggesting that fibroblasts play crucial roles in bovine mastitis. Copyright © 2016 Elsevier Inc. All rights reserved.
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Mechanism research of miR-181 regulating human lens epithelial cell apoptosis
Directory of Open Access Journals (Sweden)
Yu Qin
2015-05-01
Full Text Available AIM: To investigate the expression of miR-181 in the lens tissue of cataract and the regulating mechanism of miR-181 on apoptosis of human lens epithelial cell.METHODS:Real time q-PCR was used to measure the expression of miR-181 in the anterior lens capsules of age-related cataract and human lens epithelial cell apoptosis model. miR-181 mimic and inhibitor were transfected using Lipofectamine 2 000 to regulate the expression of miR-181, and then Real time q-PCR was used to verify transfection efficiency. Flow cytometry was used to detect the change of cell apoptosis rate. RESULTS: Compared with control group, the expression of miR-181 was significantly higher in both the anterior lens capsules of age-related cataract and human lens epithelial cell apoptosis model; the relative expression of miR-181 in lens epithelial cells transfected with miR-181 mimic was increased, whereas decreased in cells transfected with miR-181 inhibitor; the apoptosis rate of cells transfected with miR-181 mimic was increased, while reduced in miR-181 inhibitor group. Each result was statistically significant(PCONCLUSION: High expression of miR-181 is detected in anterior lens capsule of age-related cataract. miR-181 might play a certain role in the pathogenesis of cataract via promoting human lens epithelial cell apoptosis. miR-181 probably becomes a new approach for the nonoperative treatment of cataract, but the concrete mechanism still needs to be further studied.
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Psychobiology of the amniotic environment.
Benassi, Luigi; Accorsi, Francesca; Marconi, Lorenza; Benassi, Gianluca
2004-01-01
Water, basic element of amniotic fluid (A.F.), is closely related to Life, Fertility and Motherhood in several cultures and religions. Through material evidences of an essential growth medium and useful diagnostic source, a new concept grow up: the fluid as a first real environment in which fetus lives and acts. Many studies confirm that in A.F. fetus starts his character-building, his memory and his intelligence. The fluid seems to be the first means of learning and acknowledgement. Sounds, smells and tastes are perceived as well as emotions and fears. Urinoterapy and staminal cells sampling shows how A.F. can be considered as an additional terapeutic resource.
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Nesfatin-1 inhibits ovarian epithelial carcinoma cell proliferation in vitro
International Nuclear Information System (INIS)
Xu, Yang; Pang, Xiaoyan; Dong, Mei; Wen, Fang; Zhang, Yi
2013-01-01
Highlights: •Nesfatin-1 inhibits the proliferation and growth of HO-8910 cells by G1 phase arrest. •Nesfatin-1 enhances HO-8910 cell apoptosis. •Nesfatin-1 inhibits HO-8910 cell proliferation via mTOR and RhoA/ROCK signaling pathway. •The first report of nesfatin-1-mediated proliferation in ovarian epithelial carcinoma. -- Abstract: Nesfatin-1, an 82-amino-acid peptide derived from a 396-amino-acid precursor protein nucleobindin 2 (NUCB2), was originally identified in hypothalamic nuclei involved in the regulation of food intake. It was recently reported that nesfatin-1 is a novel depot specific adipokine preferentially produced by subcutaneous tissue, with obesity- and food deprivation-regulated expression. Although a relation between ovarian cancer mortality and obesity has been previously established, a role of nesfatin-1 in ovarian epithelial carcinoma remains unknown. The aim of the present study is to examine the effect of nesfatin-1 on ovary carcinoma cells proliferation. We found that nesfatin-1 inhibits the proliferation and growth of HO-8910 cells by G1 phase arrest, this inhibition could be abolished by nesfatin-1 neutralizing antibody. Nesfatin-1 enhances HO-8910 cell apoptosis, activation of mammalian target of rapamycin (mTOR) and RhoA/ROCK signaling pathway block the effects of nesfatin-1-induced apoptosis, therefore reverses the inhibition of HO-8910 cell proliferation by nesfatin-1. In conclusion, the present study demonstrated that nesfatin-1 can inhibit the proliferation in human ovarian epithelial carcinoma cell line HO-8910 cells through inducing apoptosis via mTOR and RhoA/ROCK signaling pathway. This study provides a novel regulatory signaling pathway of nesfatin-1-regulated ovarian epithelial carcinoma growth and may contribute to ovarian cancer prevention and therapy, especially in obese patients
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Nesfatin-1 inhibits ovarian epithelial carcinoma cell proliferation in vitro
Energy Technology Data Exchange (ETDEWEB)
Xu, Yang; Pang, Xiaoyan; Dong, Mei; Wen, Fang, E-mail: wenfang64@hotmail.com; Zhang, Yi, E-mail: syzi960@yahoo.com
2013-11-01
Highlights: •Nesfatin-1 inhibits the proliferation and growth of HO-8910 cells by G1 phase arrest. •Nesfatin-1 enhances HO-8910 cell apoptosis. •Nesfatin-1 inhibits HO-8910 cell proliferation via mTOR and RhoA/ROCK signaling pathway. •The first report of nesfatin-1-mediated proliferation in ovarian epithelial carcinoma. -- Abstract: Nesfatin-1, an 82-amino-acid peptide derived from a 396-amino-acid precursor protein nucleobindin 2 (NUCB2), was originally identified in hypothalamic nuclei involved in the regulation of food intake. It was recently reported that nesfatin-1 is a novel depot specific adipokine preferentially produced by subcutaneous tissue, with obesity- and food deprivation-regulated expression. Although a relation between ovarian cancer mortality and obesity has been previously established, a role of nesfatin-1 in ovarian epithelial carcinoma remains unknown. The aim of the present study is to examine the effect of nesfatin-1 on ovary carcinoma cells proliferation. We found that nesfatin-1 inhibits the proliferation and growth of HO-8910 cells by G1 phase arrest, this inhibition could be abolished by nesfatin-1 neutralizing antibody. Nesfatin-1 enhances HO-8910 cell apoptosis, activation of mammalian target of rapamycin (mTOR) and RhoA/ROCK signaling pathway block the effects of nesfatin-1-induced apoptosis, therefore reverses the inhibition of HO-8910 cell proliferation by nesfatin-1. In conclusion, the present study demonstrated that nesfatin-1 can inhibit the proliferation in human ovarian epithelial carcinoma cell line HO-8910 cells through inducing apoptosis via mTOR and RhoA/ROCK signaling pathway. This study provides a novel regulatory signaling pathway of nesfatin-1-regulated ovarian epithelial carcinoma growth and may contribute to ovarian cancer prevention and therapy, especially in obese patients.
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TLR-dependent human mucosal epithelial cell responses to microbial pathogens.
Directory of Open Access Journals (Sweden)
Paola eMassari
2014-08-01
Full Text Available AbstractToll-Like Receptor (TLR signaling represents one of the best studied pathways to implement defense mechanisms against invading microbes in humans as well as in animals. TLRs respond to specific microbial ligands and to danger signals produced by the host during infection, and initiate downstream cascades that activate both innate and adaptive immunity. TLRs are expressed by professional immune cells and by the large majority of non-hematopoietic cells, including epithelial cells. In epithelial tissues, TLR functions are particularly important because these sites are constantly exposed to microorganisms, due to their location at the host interface with the environment. While at these sites, specific defense mechanisms and inflammatory responses are initiated via TLR signaling against pathogens, suppression or lack of TLR activation is also observed in response to the commensal microbiota. The mechanisms by which TLR signaling is regulated in mucosal epithelial cells include differential expression and levels of TLRs (and their signaling partners, their cellular localization and positioning within the tissue in a fashion that favors responses to pathogens while dampening responses to commensals and maintaining tissue homeostasis in physiologic conditions. In this review, the expression and activation of TLRs in mucosal epithelial cells of several sites of the human body are examined. Specifically, the oral cavity, the ear canal and eye, the airways, the gut and the reproductive tract are discussed, along with how site-specific host defense mechanisms are implemented via TLR signaling.
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Feldkamp, Marcia L; Ward, Diane M; Pysher, Theodore J; Chambers, Christina T
2017-07-17
Vacuolated amniotic epithelium with lipid droplets in gastroschisis placentas is an unusual finding. Mass spectrometry of lipid droplets identified triglycerides, ester-linked to an unusual pattern of fatty acids. We hypothesize that these findings result from a Chlamydia trachomatis infection during the periconceptional period. The rising incidence of chlamydia infections has paralleled the increasing prevalence of gastroschisis among women less than 25 years of age. Histologically, young women are at greatest risk for a chlamydia infection due to their immature columnar epithelium, the preferential site for attachment of Chlamydia trachomatis infectious particle (elementary body). Chlamydia trachomatis survive in an inclusion, relying on its host to acquire essential nutrients, amino acids, and nucleotides for survival and replication. If essential nutrients are not available, the bacteria cannot replicate and may be trafficked to the lysosome for degradation or remain quiescent, within the inclusion, subverting innate immunologic clearance. Chlamydiae synthesize several lipids (phosphatidylethanolamine, phosphatidylserine, and phosphoatidylglycerol); however, their lipid content reveal eukaryotic lipids (sphingomyelin, cholesterol, phosphatidylcholine, and phosphatidylinositol), evidence that chlamydiae "hijack" host lipids for expansion and replication. The abnormal amniotic epithelial findings are supported by experimental evidence of the trafficking of host lipids into the chlamydiae inclusion. If not lethal, what harm will elementary bodies inflict to the developing embryo? Do these women have a greater pro-inflammatory response to an environmental exposure, whether cigarette smoking, change in partner, or a pathogen? Testing the hypothesis that Chlamydia trachomatis is responsible for amniotic epithelium vacuoles will be a critical first step. Birth Defects Research 109:1003-1010, 2017. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.
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2016-09-01
AWARD NUMBER: W811XWH-13-1-0310 TITLE: Acceleration of Regeneration of Large-Gap Peripheral Nerve Injuries Using Acellular Nerve Allografts...plus amniotic Fluid Derived Stem Cells (AFS). PRINCIPAL INVESTIGATOR: Zhongyu Li, MD, PhD RECIPIENT: Wake Forest University Health Sciences...REPORT DATE September 2016 2. REPORT TYPE Annual 3. DATES COVERED 1Sep2015 - 31Aug2016 4. TITLE AND SUBTITLE Acceleration of Regeneration of Large
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Vaginal epithelial cells regulate membrane adhesiveness to co-ordinate bacterial adhesion.
Younes, Jessica A; Klappe, Karin; Kok, Jan Willem; Busscher, Henk J; Reid, Gregor; van der Mei, Henny C
2016-04-01
Vaginal epithelium is colonized by different bacterial strains and species. The bacterial composition of vaginal biofilms controls the balance between health and disease. Little is known about the relative contribution of the epithelial and bacterial cell surfaces to bacterial adhesion and whether and how adhesion is regulated over cell membrane regions. Here, we show that bacterial adhesion forces with cell membrane regions not located above the nucleus are stronger than with regions above the nucleus both for vaginal pathogens and different commensal and probiotic lactobacillus strains involved in health. Importantly, adhesion force ratios over membrane regions away from and above the nucleus coincided with the ratios between numbers of adhering bacteria over both regions. Bacterial adhesion forces were dramatically decreased by depleting the epithelial cell membrane of cholesterol or sub-membrane cortical actin. Thus, epithelial cells can regulate membrane regions to which bacterial adhesion is discouraged, possibly to protect the nucleus. © 2015 John Wiley & Sons Ltd.
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Vitreous Humor Changes Expression of Iron-Handling Proteins in Lens Epithelial Cells
Goralska, Malgorzata; Fleisher, Lloyd N.; McGahan, M. Christine
2017-01-01
Purpose In humans, vitrectomy is associated with development of nuclear cataracts. Iron catalyzes free radical formation causing oxidative damage, which is implicated in cataract formation. This study was designed to determine if vitreous humor, which can initiate differentiation of lens epithelial cells, would have an effect on iron-handling proteins. Methods Cultured canine lens epithelial cells were treated with collected canine vitreous humor. Lysates of treated and control cells were separated by SDS-PAGE. Ferritin H- and L-chains, transferrin receptor 1, and aquaporin 0 were immunodetected and quantitated with specific antibodies. Morphologic changes in treated cells were assessed. Results Treatment of lens epithelial cells with a 33% (vol/vol) solution of vitreous humor changed the morphology of lens cells and induced expression of aquaporin 0, a marker of fiber cell differentiation that was undetectable in control cells. Treatment did not modify the size of iron-handling proteins but significantly increased content of ferritin from 2.9- to 8.8-fold over control and decreased levels of transferrin receptor by 37% to 59%. Conclusions Vitreous humor may significantly limit iron uptake by transferrin/transferrin receptor pathway, and by increasing ferritin levels could profoundly increase the iron-storage capacity of ferritin in lens cells. Vitreous humor may play a significant protective role against iron-catalyzed oxidative damage of lens epithelial cells and therefore in the formation of cataracts. PMID:28245299
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Ulm, Ashley; Mayhew, Christopher N; Debley, Jason; Khurana Hershey, Gurjit K; Ji, Hong
2016-03-10
Nasal epithelial cells (NECs) are the part of the airways that respond to air pollutants and are the first cells infected with respiratory viruses. They are also involved in many airway diseases through their innate immune response and interaction with immune and airway stromal cells. NECs are of particular interest for studies in children due to their accessibility during clinical visits. Human induced pluripotent stem cells (iPSCs) have been generated from multiple cell types and are a powerful tool for modeling human development and disease, as well as for their potential applications in regenerative medicine. This is the first protocol to lay out methods for successful generation of iPSCs from NECs derived from pediatric participants for research purposes. It describes how to obtain nasal epithelial cells from children, how to generate primary NEC cultures from these samples, and how to reprogram primary NECs into well-characterized iPSCs. Nasal mucosa samples are useful in epidemiological studies related to the effects of air pollution in children, and provide an important tool for studying airway disease. Primary nasal cells and iPSCs derived from them can be a tool for providing unlimited material for patient-specific research in diverse areas of airway epithelial biology, including asthma and COPD research.
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Smith, I M; Baker, A; Arneborg, N; Jespersen, L
2015-11-01
The human gastrointestinal epithelium makes up the largest barrier separating the body from the external environment. Whereas invasive pathogens cause epithelial barrier disruption, probiotic micro-organisms modulate tight junction regulation and improve epithelial barrier function. In addition, probiotic strains may be able to reduce epithelial barrier disruption caused by pathogenic species. The aim of this study was to explore non-Saccharomyces yeast modulation of epithelial cell barrier function in vitro. Benchmarking against established probiotic strains, we evaluated the ability of four nonpathogenic yeast species to modulate transepithelial electrical resistance (TER) across a monolayer of differentiated human colonocytes (Caco-2 cells). Further, we assessed yeast modulation of a Salmonella Typhimurium-induced epithelial cell barrier function insult. Our findings demonstrate distinct patterns of non-Saccharomyces yeast modulation of epithelial cell barrier function. While the established probiotic yeast Saccharomyces boulardii increased TER across a Caco-2 monolayer by 30%, Kluyveromyces marxianus exhibited significantly stronger properties of TER enhancement (50% TER increase). In addition, our data demonstrate significant yeast-mediated modulation of Salmonella-induced epithelial cell barrier disruption and identify K. marxianus and Metschnikowia gruessii as two non-Saccharomyces yeasts capable of protecting human epithelial cells from pathogen invasion. This study demonstrates distinct patterns of non-Saccharomyces yeast modulation of epithelial cell barrier function in vitro. Further, our data demonstrate significant yeast-mediated modulation of Salmonella Typhimurium-induced epithelial cell barrier disruption and identify Kluyveromyces marxianus and Metschnikowia gruessii as two non-Saccharomyces yeasts capable of protecting human epithelial cells from pathogen invasion. This study is the first to demonstrate significant non-Saccharomyces yeast
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Yamamoto, Mizuki; Sakane, Kota; Tominaga, Kana; Gotoh, Noriko; Niwa, Takayoshi; Kikuchi, Yasuko; Tada, Keiichiro; Goshima, Naoki; Semba, Kentaro; Inoue, Jun-Ichiro
2017-06-01
Epithelial-mesenchymal transition (EMT) and its reverse process, mesenchymal-epithelial transition MET, are crucial in several stages of cancer metastasis. Epithelial-mesenchymal transition allows cancer cells to move to proximal blood vessels for intravasation. However, because EMT and MET processes are dynamic, mesenchymal cancer cells are likely to undergo MET transiently and subsequently re-undergo EMT to restart the metastatic process. Therefore, spatiotemporally coordinated mutual regulation between EMT and MET could occur during metastasis. To elucidate such regulation, we chose HCC38, a human triple-negative breast cancer cell line, because HCC38 is composed of epithelial and mesenchymal populations at a fixed ratio even though mesenchymal cells proliferate significantly more slowly than epithelial cells. We purified epithelial and mesenchymal cells from Venus-labeled and unlabeled HCC38 cells and mixed them at various ratios to follow EMT and MET. Using this system, we found that the efficiency of EMT is approximately an order of magnitude higher than that of MET and that the two populations significantly enhance the transition of cells from the other population to their own. In addition, knockdown of Zinc finger E-box-binding homeobox 1 (ZEB1) or Zinc finger protein SNAI2 (SLUG) significantly suppressed EMT but promoted partial MET, indicating that ZEB1 and SLUG are crucial to EMT and MET. We also show that primary breast cancer cells underwent EMT that correlated with changes in expression profiles of genes determining EMT status and breast cancer subtype. These changes were very similar to those observed in EMT in HCC38 cells. Consequently, we propose HCC38 as a suitable model to analyze EMT-MET dynamics that could affect the development of triple-negative breast cancer. © 2017 The Authors. Cancer Science published by John Wiley & Sons Australia, Ltd on behalf of Japanese Cancer Association.
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International Nuclear Information System (INIS)
Jang, Soo Hwa; Choi, Changsun; Hong, Seong-Geun; Yarishkin, Oleg V.; Bae, Young Min; Kim, Jae Gon; O'Grady, Scott M.; Yoon, Kyong-Ah; Kang, Kyung-Sun; Ryu, Pan Dong; Lee, So Yeong
2009-01-01
Potassium channel activity has been shown to facilitate cell proliferation in cancer cells. In the present study, the role of Kv4.1 channels in immortal and tumorigenic human mammary epithelial cells was investigated. Kv4.1 protein expression was positively correlated with tumorigenicity. Moreover, transfection with siRNAs targeting Kv4.1 mRNA suppressed proliferation of tumorigenic mammary epithelial cells. Experiments using mRNA isolated from human breast cancer tissues revealed that the level of Kv4.1 mRNA expression varied depending on the stage of the tumor. Kv4.1 protein expression increased during stages T2 and T3 compared to normal tissue. These results demonstrated that Kv4.1 plays a role in proliferation of tumorigenic human mammary epithelial cells. In addition, elevated Kv4.1 expression may be useful as a diagnostic marker for staging mammary tumors and selective blockers of Kv4.1 may serve to suppress tumor cell proliferation.
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Wnt is necessary for mesenchymal to epithelial transition in colorectal cancer cells.
Schwab, Renate H M; Amin, Nancy; Flanagan, Dustin J; Johanson, Timothy M; Phesse, Toby J; Vincan, Elizabeth
2018-03-01
Metastasis underlies most colorectal cancer mortality. Cancer cells spread through the body as single cells or small clusters of cells that have an invasive, mesenchymal, nonproliferative phenotype. At the secondary site, they revert to a proliferative "tumor constructing" epithelial phenotype to rebuild a tumor. We previously developed a unique in vitro three-dimensional model, called LIM1863-Mph, which faithfully recapitulates these reversible transitions that underpin colorectal cancer metastasis. Wnt signaling plays a key role in these transitions and is initiated by the coupling of extracellular Wnt to Frizzled (FZD). Using the LIM1863-Mph model system we demonstrated that the Wnt receptor FZD7 is necessary for mesenchymal to epithelial transition (MET). Here we investigate the role of Wnt in MET. Wnt secretion is dependent on palmitoylation by Porcupine (PORC). A PORC inhibitor (IWP2) that prevents Wnt secretion, blocked the epithelial transition of mesenchymal LIM1863-Mph cells. Wnt gene array analysis identified several Wnts that are upregulated in epithelial compared with mesenchymal LIM1863-Mph cells, suggesting these ligands in MET. Wnt2B was the most abundant differentially expressed Wnt gene. Indeed, recombinant Wnt2B could overcome the IWP2-mediated block in epithelial transition of mesenchymal LIM1863-Mph cells. Wnt2B co-operates with Frizzled7 to mediate MET in colorectal cancer. Developmental Dynamics 247:521-530, 2018. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.
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Nuclear Nox4 Role in Stemness Power of Human Amniotic Fluid Stem Cells
Directory of Open Access Journals (Sweden)
Tullia Maraldi
2015-01-01
Full Text Available Human amniotic fluid stem cells (AFSC are an attractive source for cell therapy due to their multilineage differentiation potential and accessibility advantages. However the clinical application of human stem cells largely depends on their capacity to expand in vitro, since there is an extensive donor-to-donor heterogeneity. Reactive oxygen species (ROS and cellular oxidative stress are involved in many physiological and pathophysiological processes of stem cells, including pluripotency, proliferation, differentiation, and stress resistance. The mode of action of ROS is also dependent on the localization of their target molecules. Thus, the modifications induced by ROS can be separated depending on the cellular compartments they affect. NAD(PH oxidase family, particularly Nox4, has been known to produce ROS in the nucleus. In the present study we show that Nox4 nuclear expression (nNox4 depends on the donor and it correlates with the expression of transcription factors involved in stemness regulation, such as Oct4, SSEA-4, and Sox2. Moreover nNox4 is linked with the nuclear localization of redox sensitive transcription factors, as Nrf2 and NF-κB, and with the differentiation potential. Taken together, these results suggest that nNox4 regulation may have important effects in stem cell capability through modulation of transcription factors and DNA damage.
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Directory of Open Access Journals (Sweden)
Liara M Gonzalez
Full Text Available Significant advances in intestinal stem cell biology have been made in murine models; however, anatomical and physiological differences between mice and humans limit mice as a translational model for stem cell based research. The pig has been an effective translational model, and represents a candidate species to study intestinal epithelial stem cell (IESC driven regeneration. The lack of validated reagents and epithelial culture methods is an obstacle to investigating IESC driven regeneration in a pig model. In this study, antibodies against Epithelial Adhesion Molecule 1 (EpCAM and Villin marked cells of epithelial origin. Antibodies against Proliferative Cell Nuclear Antigen (PCNA, Minichromosome Maintenance Complex 2 (MCM2, Bromodeoxyuridine (BrdU and phosphorylated Histone H3 (pH3 distinguished proliferating cells at various stages of the cell cycle. SOX9, localized to the stem/progenitor cells zone, while HOPX was restricted to the +4/'reserve' stem cell zone. Immunostaining also identified major differentiated lineages. Goblet cells were identified by Mucin 2 (MUC2; enteroendocrine cells by Chromogranin A (CGA, Gastrin and Somatostatin; and absorptive enterocytes by carbonic anhydrase II (CAII and sucrase isomaltase (SIM. Transmission electron microscopy demonstrated morphologic and sub-cellular characteristics of stem cell and differentiated intestinal epithelial cell types. Quantitative PCR gene expression analysis enabled identification of stem/progenitor cells, post mitotic cell lineages, and important growth and differentiation pathways. Additionally, a method for long-term culture of porcine crypts was developed. Biomarker characterization and development of IESC culture in the porcine model represents a foundation for translational studies of IESC-driven regeneration of the intestinal epithelium in physiology and disease.
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Management of Amniotic Sheet with a Hammock-like Placenta
Directory of Open Access Journals (Sweden)
Liangcheng Wang
2016-09-01
Full Text Available An amniotic sheet is a septation in the amniotic cavity with a perforation that allows amniotic fluid to pass through. Although the incidence of abnormal placental implantation is higher in such cases, the management recommendations remain unclear. We report a case of an amniotic sheet with a hammock-like placenta located in the center of the uterine cavity. A 25-year-old woman with a history of two dilation and curettage procedures was found to have an amniotic cavity separated by a septum that contained part of the placenta. At gestational Week 32, magnetic resonance images revealed that the placenta was attached from the anterior to posterior uterine walls and resembled a hammock hanging in the center of the uterus. Subsequently, continuous intravenous administration of ritodrine hydrochloride and magnesium sulfate were given. The pregnancy was extended to Week 36. Elective cesarean section was performed, and a 3212-g female infant was delivered. Thus, owing to the risk of umbilical cord complications and placental injury secondary to premature rupture of membranes, aggressive and careful perinatal management is required in such cases.
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Adenovirus E1A/E1B Transformed Amniotic Fluid Cells Support Human Cytomegalovirus Replication
Directory of Open Access Journals (Sweden)
Natascha Krömmelbein
2016-02-01
Full Text Available The human cytomegalovirus (HCMV replicates to high titers in primary human fibroblast cell cultures. A variety of primary human cells and some tumor-derived cell lines do also support permissive HCMV replication, yet at low levels. Cell lines established by transfection of the transforming functions of adenoviruses have been notoriously resistant to HCMV replication and progeny production. Here, we provide first-time evidence that a permanent cell line immortalized by adenovirus type 5 E1A and E1B (CAP is supporting the full HCMV replication cycle and is releasing infectious progeny. The CAP cell line had previously been established from amniotic fluid cells which were likely derived from membranes of the developing fetus. These cells can be grown under serum-free conditions. HCMV efficiently penetrated CAP cells, expressed its immediate-early proteins and dispersed restrictive PML-bodies. Viral DNA replication was initiated and viral progeny became detectable by electron microscopy in CAP cells. Furthermore, infectious virus was released from CAP cells, yet to lower levels compared to fibroblasts. Subviral dense bodies were also secreted from CAP cells. The results show that E1A/E1B expression in transformed cells is not generally repressive to HCMV replication and that CAP cells may be a good substrate for dense body based vaccine production.
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File list: NoD.Utr.50.AllAg.Fallopian_tube_secretory_epithelial_cell [Chip-atlas[Archive
Lifescience Database Archive (English)
Full Text Available NoD.Utr.50.AllAg.Fallopian_tube_secretory_epithelial_cell hg19 No description Uterus Fallopian tube secret...ory epithelial cell http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/NoD.Utr.50.AllAg.Fallopian_tube_secretory_epithelial_cell.bed ...
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File list: NoD.Utr.05.AllAg.Fallopian_tube_secretory_epithelial_cell [Chip-atlas[Archive
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Full Text Available NoD.Utr.05.AllAg.Fallopian_tube_secretory_epithelial_cell hg19 No description Uterus Fallopian tube secret...ory epithelial cell http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/NoD.Utr.05.AllAg.Fallopian_tube_secretory_epithelial_cell.bed ...
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File list: NoD.Utr.20.AllAg.Fallopian_tube_secretory_epithelial_cell [Chip-atlas[Archive
Lifescience Database Archive (English)
Full Text Available NoD.Utr.20.AllAg.Fallopian_tube_secretory_epithelial_cell hg19 No description Uterus Fallopian tube secret...ory epithelial cell http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/NoD.Utr.20.AllAg.Fallopian_tube_secretory_epithelial_cell.bed ...
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File list: NoD.Utr.10.AllAg.Fallopian_tube_secretory_epithelial_cell [Chip-atlas[Archive
Lifescience Database Archive (English)
Full Text Available NoD.Utr.10.AllAg.Fallopian_tube_secretory_epithelial_cell hg19 No description Uterus Fallopian tube secret...ory epithelial cell http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/NoD.Utr.10.AllAg.Fallopian_tube_secretory_epithelial_cell.bed ...
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Pup retrieval and maternal attraction to canine amniotic fluids.
Dunbar, I; Ranson, E; Buehler, M
1981-10-01
Three purebred female beagles were observed with both their first and second litters. Dams were given three separate simultaneous-choice retrieval tests: In addition, the maternal response to amniotic fluids was observed when one pup in the litter was treated with amniotic fluids and all other pups were treated with water. There was no evidence to suggest that pups were retrieved preferentially on the basis of their sex. One female retrieved her own but not alien pups, whereas another female made no such discrimination and readily retrieved alien pups in addition to her own. Retrieval behavior developed at the time of each whelping and normally lasted until the pups were about 5 days old, although a bitch would retrieve younger (alien) pups up to 14 days post partum. Maternal bitches were strongly attracted towards amniotic fluids: they investigated pups daubed with amniotic fluids to a significantly greater extent than control pups treated with water. The attraction of maternal females towards amniotic fluids developed at the time of each whelping and persisted for up to 30 days, well beyond the time that a whelping bitch would normally be exposed to her own fetal fluids. A possible role for amniotic fluids in the development of maternal behaviour and the establishment of the maternal/puppy bond is discussed. Copyright © 1981. Published by Elsevier B.V.
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The human airway epithelial basal cell transcriptome.
Directory of Open Access Journals (Sweden)
Neil R Hackett
2011-05-01
Full Text Available The human airway epithelium consists of 4 major cell types: ciliated, secretory, columnar and basal cells. During natural turnover and in response to injury, the airway basal cells function as stem/progenitor cells for the other airway cell types. The objective of this study is to better understand human airway epithelial basal cell biology by defining the gene expression signature of this cell population.Bronchial brushing was used to obtain airway epithelium from healthy nonsmokers. Microarrays were used to assess the transcriptome of basal cells purified from the airway epithelium in comparison to the transcriptome of the differentiated airway epithelium. This analysis identified the "human airway basal cell signature" as 1,161 unique genes with >5-fold higher expression level in basal cells compared to differentiated epithelium. The basal cell signature was suppressed when the basal cells differentiated into a ciliated airway epithelium in vitro. The basal cell signature displayed overlap with genes expressed in basal-like cells from other human tissues and with that of murine airway basal cells. Consistent with self-modulation as well as signaling to other airway cell types, the human airway basal cell signature was characterized by genes encoding extracellular matrix components, growth factors and growth factor receptors, including genes related to the EGF and VEGF pathways. Interestingly, while the basal cell signature overlaps that of basal-like cells of other organs, the human airway basal cell signature has features not previously associated with this cell type, including a unique pattern of genes encoding extracellular matrix components, G protein-coupled receptors, neuroactive ligands and receptors, and ion channels.The human airway epithelial basal cell signature identified in the present study provides novel insights into the molecular phenotype and biology of the stem/progenitor cells of the human airway epithelium.
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Directory of Open Access Journals (Sweden)
Zuraw Bruce L
2009-10-01
Full Text Available Abstract Background Chronic persistent asthma is characterized by ongoing airway inflammation and airway remodeling. The processes leading to airway remodeling are poorly understood, and there is increasing evidence that even aggressive anti-inflammatory therapy does not completely prevent this process. We sought to investigate whether TGFβ1 stimulates bronchial epithelial cells to undergo transition to a mesenchymal phenotype, and whether this transition can be abrogated by corticosteroid treatment or enhanced by the pro-inflammatory cytokine IL-1β. Methods BEAS-2B and primary normal human bronchial epithelial cells were stimulated with TGFβ1 and expression of epithelial and mesenchymal markers assessed by quantitative real-time PCR, immunoblotting, immunofluorescence microscopy and zymography. In some cases the epithelial cells were also incubated with corticosteroids or IL-1β. Results were analyzed using non-parametric statistical tests. Results Treatment of BEAS-2B or primary human bronchial epithelial cells with TGFβ1 significantly reduced the expression level of the epithelial adherence junction protein E-cadherin. TGFβ1 then markedly induced mesenchymal marker proteins such as collagen I, tenascin C, fibronectin and α-smooth muscle actin mRNA in a dose dependant manner. The process of mesenchymal transition was accompanied by a morphological change towards a more spindle shaped fibroblast cell type with a more motile and invasive phenotype. Corticosteroid pre-treatment did not significantly alter the TGFβ1 induced transition but IL-1β enhanced the transition. Conclusion Our results indicate, that TGFβ1 can induce mesenchymal transition in the bronchial epithelial cell line and primary cells. Since asthma has been strongly associated with increased expression of TGFβ1 in the airway, epithelial to mesenchymal transition may contribute to the contractile and fibrotic remodeling process that accompanies chronic asthma.
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DEFF Research Database (Denmark)
Alviano, Francesco; Fossati, Valentina; Marchionni, Cosetta
2007-01-01
of CD34 and von Willebrand Factor positive cells. CONCLUSION: The current study suggests that AM-hMSCs may emerge as a remarkable tool for the cell therapy of multiple diseased tissues. AM-hMSCs may potentially assist both bone and cartilage repair, nevertheless, due to their angiogenic potential......BACKGROUND: Term Amniotic membrane (AM) is a very attractive source of Mesenchymal Stem Cells (MSCs) due to the fact that this fetal tissue is usually discarded without ethical conflicts, leading to high efficiency in MSC recovery with no intrusive procedures. Here we confirmed that term AM......, as previously reported in the literature, is an abundant source of hMSCs; in particular we further investigated the AM differentiation potential by assessing whether these cells may also be committed to the angiogenic fate. In agreement with the recommendation of the International Society for Cellular Therapy...
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Inhibition of PTP1B disrupts cell–cell adhesion and induces anoikis in breast epithelial cells
Hilmarsdottir, Bylgja; Briem, Eirikur; Halldorsson, Skarphedinn; Kricker, Jennifer; Ingthorsson, Sævar; Gustafsdottir, Sigrun; Mælandsmo, Gunhild M; Magnusson, Magnus K; Gudjonsson, Thorarinn
2017-01-01
Protein tyrosine phosphatase 1B (PTP1B) is a well-known inhibitor of insulin signaling pathways and inhibitors against PTP1B are being developed as promising drug candidates for treatment of obesity. PTP1B has also been linked to breast cancer both as a tumor suppressor and as an oncogene. Furthermore, PTP1B has been shown to be a regulator of cell adhesion and migration in normal and cancer cells. In this study, we analyzed the PTP1B expression in normal breast tissue, primary breast cells and the breast epithelial cell line D492. In normal breast tissue and primary breast cells, PTP1B is widely expressed in both epithelial and stromal cells, with highest expression in myoepithelial cells and fibroblasts. PTP1B is widely expressed in branching structures generated by D492 when cultured in 3D reconstituted basement membrane (3D rBM). Inhibition of PTP1B in D492 and another mammary epithelial cell line HMLE resulted in reduced cell proliferation and induction of anoikis. These changes were seen when cells were cultured both in monolayer and in 3D rBM. PTP1B inhibition affected cell attachment, expression of cell adhesion proteins and actin polymerization. Moreover, epithelial to mesenchymal transition (EMT) sensitized cells to PTP1B inhibition. A mesenchymal sublines of D492 and HMLE (D492M and HMLEmes) were more sensitive to PTP1B inhibition than D492 and HMLE. Reversion of D492M to an epithelial state using miR-200c-141 restored resistance to detachment induced by PTP1B inhibition. In conclusion, we have shown that PTP1B is widely expressed in the human breast gland with highest expression in myoepithelial cells and fibroblasts. Inhibition of PTP1B in D492 and HMLE affects cell–cell adhesion and induces anoikis-like effects. Finally, cells with an EMT phenotype are more sensitive to PTP1B inhibitors making PTP1B a potential candidate for further studies as a target for drug development in cancer involving the EMT phenotype. PMID:28492548
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DEFF Research Database (Denmark)
Wiencke, Anne Katrine
2001-01-01
ophthalmology, age-related macular degeneration, transplantation, retinal pigment epithelial cells, treatment......ophthalmology, age-related macular degeneration, transplantation, retinal pigment epithelial cells, treatment...
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DEFF Research Database (Denmark)
Wiencke, Anne Katrine
2001-01-01
ophthalmology, age-related macular degeneration, retinal pigment epithelial cells, transplantation, treatment......ophthalmology, age-related macular degeneration, retinal pigment epithelial cells, transplantation, treatment...
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Sakai, Norihiko; Chun, Jerold; Duffield, Jeremy S; Lagares, David; Wada, Takashi; Luster, Andrew D; Tager, Andrew M
2017-03-01
The expansion of the fibroblast pool is a critical step in organ fibrosis, but the mechanisms driving expansion remain to be fully clarified. We previously showed that lysophosphatidic acid (LPA) signaling through its receptor LPA 1 expressed on fibroblasts directly induces the recruitment of these cells. Here we tested whether LPA-LPA 1 signaling drives fibroblast proliferation and activation during the development of renal fibrosis. LPA 1 -deficient (LPA 1 -/- ) or -sufficient (LPA 1 +/+ ) mice were crossed to mice with green fluorescent protein expression (GFP) driven by the type I procollagen promoter (Col-GFP) to identify fibroblasts. Unilateral ureteral obstruction-induced increases in renal collagen were significantly, though not completely, attenuated in LPA 1 -/- Col-GFP mice, as were the accumulations of both fibroblasts and myofibroblasts. Connective tissue growth factor was detected mainly in tubular epithelial cells, and its levels were suppressed in LPA 1 -/- Col-GFP mice. LPA-LPA 1 signaling directly induced connective tissue growth factor expression in primary proximal tubular epithelial cells, through a myocardin-related transcription factor-serum response factor pathway. Proximal tubular epithelial cell-derived connective tissue growth factor mediated renal fibroblast proliferation and myofibroblast differentiation. Administration of an inhibitor of myocardin-related transcription factor/serum response factor suppressed obstruction-induced renal fibrosis. Thus, targeting LPA-LPA 1 signaling and/or myocardin-related transcription factor/serum response factor-induced transcription could be promising therapeutic strategies for renal fibrosis. Copyright © 2016 International Society of Nephrology. Published by Elsevier Inc. All rights reserved.
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Egea, Laia; McAllister, Christopher S.; Lakhdari, Omar; Minev, Ivelina; Shenouda, Steve; Kagnoff, Martin F.
2012-01-01
GM-CSF is a growth factor that promotes the survival and activation of macrophages and granulocytes, and dendritic cell (DC) differentiation and survival in vitro. The mechanism by which exogenous GM-CSF ameliorates the severity of Crohn’s disease in humans and colitis in murine models has been considered mainly to reflect its activity on myeloid cells. We used GM-CSF deficient (GM-CSF−/−) mice to probe the functional role of endogenous host-produced GM-CSF in a colitis model induced after injury to the colon epithelium. Dextran sodium sulfate (DSS) at doses that resulted in little epithelial damage and mucosal ulceration in wild type (WT) mice resulted in marked colon ulceration and delayed ulcer healing in GM-CSF−/− mice. Colon crypt epithelial cell proliferation in vivo was significantly decreased in GM-CSF−/− mice at early times after DSS injury. This was paralleled by decreased expression of crypt epithelial cell genes involved in cell cycle, proliferation, and wound healing. Decreased crypt cell proliferation and delayed ulcer healing in GM-CSF−/− mice were rescued by exogenous GM-CSF, indicating the lack of a developmental abnormality in the epithelial cell proliferative response in those mice. Non-hematopoietic cells and not myeloid cells produced the GM-CSF important for colon epithelial proliferation after DSS-induced injury as revealed by bone marrow chimera and DC depletion experiments, with colon epithelial cells being the cellular source of GM-CSF. Endogenous epithelial cell produced GM-CSF has a novel non-redundant role in facilitating epithelial cell proliferation and ulcer healing in response to injury of the colon crypt epithelium. PMID:23325885
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Noruddin, Nur Adelina Ahmad; Saim, Aminuddin B; Chua, Kien Hui; Idrus, Ruszymah
2007-12-01
To compare a co-culture system with a conventional dispase-dissociation method for obtaining functional human respiratory epithelial cells from the nasal turbinates for tissue engineering application. Human respiratory epithelial cells were serially passaged using a co-culture system and a conventional dispase-dissociation technique. The growth kinetics and gene expression levels of the cultured respiratory epithelial cells were compared. Four genes were investigated, namely cytokeratin-18, a marker for ciliated and secretory epithelial cells; cytokeratin-14, a marker for basal epithelial cells; MKI67, a proliferation marker; and MUC5B, a marker for mucin secretion. Immunocytochemical analysis was performed using monoclonal antibodies against the high molecular-weight cytokeratin 34 beta E12, cytokeratin 18, and MUC5A to investigate the protein expression from cultured respiratory epithelial cells. Respiratory epithelial cells cultured using both methods maintained polygonal morphology throughout the passages. At passage 1, co-cultured respiratory epithelial showed a 2.6-times higher growth rate compared to conventional dispase dissociation technique, and 7.8 times higher at passage 2. Better basal gene expression was observed by co-cultured respiratory epithelial cells compared to dispase dissociated cells. Immunocytochemical analyses were positive for the respiratory epithelial cells cultured using both techniques. Co-culture system produced superior quality of cultured human respiratory epithelial cells from the nasal turbinates as compared to dispase dissociation technique.
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Energy Technology Data Exchange (ETDEWEB)
Person, Rachel J.; Olive Ngalame, Ntube N.; Makia, Ngome L.; Bell, Matthew W.; Waalkes, Michael P.; Tokar, Erik J., E-mail: tokare@niehs.nih.gov
2015-07-01
Inorganic arsenic is a human lung carcinogen. We studied the ability of chronic inorganic arsenic (2 μM; as sodium arsenite) exposure to induce a cancer phenotype in the immortalized, non-tumorigenic human lung peripheral epithelial cell line, HPL-1D. After 38 weeks of continuous arsenic exposure, secreted matrix metalloproteinase-2 (MMP2) activity increased to over 200% of control, levels linked to arsenic-induced cancer phenotypes in other cell lines. The invasive capacity of these chronic arsenic-treated lung epithelial (CATLE) cells increased to 320% of control and colony formation increased to 280% of control. CATLE cells showed enhanced proliferation in serum-free media indicative of autonomous growth. Compared to control cells, CATLE cells showed reduced protein expression of the tumor suppressor gene PTEN (decreased to 26% of control) and the putative tumor suppressor gene SLC38A3 (14% of control). Morphological evidence of epithelial-to-mesenchymal transition (EMT) occurred in CATLE cells together with appropriate changes in expression of the EMT markers vimentin (VIM; increased to 300% of control) and e-cadherin (CDH1; decreased to 16% of control). EMT is common in carcinogenic transformation of epithelial cells. CATLE cells showed increased KRAS (291%), ERK1/2 (274%), phosphorylated ERK (p-ERK; 152%), and phosphorylated AKT1 (p-AKT1; 170%) protein expression. Increased transcript expression of metallothioneins, MT1A and MT2A and the stress response genes HMOX1 (690%) and HIF1A (247%) occurred in CATLE cells possibly in adaptation to chronic arsenic exposure. Thus, arsenic induced multiple cancer cell characteristics in human peripheral lung epithelial cells. This model may be useful to assess mechanisms of arsenic-induced lung cancer. - Highlights: • Chronic arsenic exposure transforms a human peripheral lung epithelia cell line. • Cells acquire characteristics in common with human lung adenocarcinoma cells. • These transformed cells provide a
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The plasticity of human breast carcinoma cells is more than epithelial to mesenchymal conversion
International Nuclear Information System (INIS)
William Petersen, Ole; Lind Nielsen, Helga; Gudjonsson, Thorarinn; Villadsen, René; Rønnov-Jessen, Lone; Bissell, Mina J
2001-01-01
The human breast comprises three lineages: the luminal epithelial lineage, the myoepithelial lineage, and the mesenchymal lineage. It has been widely accepted that human breast neoplasia pertains only to the luminal epithelial lineage. In recent years, however, evidence has accumulated that neoplastic breast epithelial cells may be substantially more plastic in their differentiation repertoire than previously anticipated. Thus, along with an increasing availability of markers for the myoepithelial lineage, at least a partial differentiation towards this lineage is being revealed frequently. It has also become clear that conversions towards the mesenchymal lineage actually occur, referred to as epithelial to mesenchymal transitions. Indeed, some of the so-called myofibroblasts surrounding the tumor may have an epithelial origin rather than a mesenchymal origin. Because myoepithelial cells, epithelial to mesenchymal transition-derived cells, genuine stromal cells and myofibroblasts share common markers, we now need to define a more ambitious set of markers to distinguish these cell types in the microenvironment of the tumors. This is necessary because the different microenvironments may confer different clinical outcomes. The aim of this commentary is to describe some of the inherent complexities in defining cellular phenotypes in the microenvironment of breast cancer and to expand wherever possible on the implications for tumor suppression and progression
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The plasticity of human breast carcinoma cells is more than epithelial to mesenchymal conversion
Energy Technology Data Exchange (ETDEWEB)
Petersen, Ole William; Nielsen, Helga Lind; Gudjonsson, Thorarinn; Villadsen, Ren& #233;
; Ronnov-Jessen, Lone; Bissell, Mina J. 2001-05-12
The human breast comprises three lineages: the luminal epithelial lineage, the myoepithelial lineage, and the mesenchymal lineage. It has been widely accepted that human breast neoplasia pertains only to the luminal epithelial lineage. In recent years, however, evidence has accumulated that neoplastic breast epithelial cells may be substantially more plastic in their differentiation repertoire than previously anticipated. Thus, along with an increasing availability of markers for the myoepithelial lineage, at least a partial differentiation towards this lineage is being revealed frequently. It has also become clear that conversions towards the mesenchymal lineage actually occur, referred to as epithelial to mesenchymal transitions. Indeed, some of the so-called myofibroblasts surrounding the tumor may indeed have an epithelial origin rather than a mesenchymal origin. Because myoepithelial cells, epithelial to mesenchymal transition-derived cells, genuine stromal cells and myofibroblasts share common markers, we now need to define a more ambitious set of markers to distinguish these cell types in the microenvironment of the tumors. This is necessary because the different microenvironments may confer different clinical outcomes. The aim of this commentary is to describe some of the inherent complexities in defining cellular phenotypes in the microenvironment of breast cancer and to expand wherever possible on the implications for tumor suppression and progression.
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Sumitomo, Tomoko; Nakata, Masanobu; Yamaguchi, Masaya; Terao, Yutaka; Kawabata, Shigetada
2012-01-01
Streptococcus pneumoniae is a major pathogen of respiratory infections that utilizes platelet-activating factor receptor (PAFR) for firm adherence to host cells. The mucolytic agent S-carboxymethylcysteine (S-CMC) has been shown to exert inhibitory effects against infection by several respiratory pathogens including S. pneumoniae in vitro and in vivo. Moreover, clinical studies have implicated the benefits of S-CMC in preventing exacerbation of chronic obstructive pulmonary disease, which is considered to be related to respiratory infections. In this study, to assess whether the potency of S-CMC is attributable to inhibition of pneumococcal adherence to host cells, an alveolar epithelial cell line stimulated with interleukin-1α was used as a model of inflamed epithelial cells. Despite upregulation of PAFR by inflammatory activation, treatment with S-CMC efficiently inhibited pneumococcal adherence to host epithelial cells. In order to gain insight into the inhibitory mechanism, the effects of S-CMC on PAFR expression were also investigated. Following treatment with S-CMC, PAFR expression was reduced at both mRNA and post-transcriptional levels. Interestingly, S-CMC was also effective in inhibiting pneumococcal adherence to cells transfected with PAFR small interfering RNAs. These results indicate S-CMC as a probable inhibitor targeting numerous epithelial receptors that interact with S. pneumoniae.
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Membrane associated ion transport enzymes in normal and transformed fibroblasts and epithelial cells
International Nuclear Information System (INIS)
Borek, C.
1982-01-01
In an effort to evaluate membrane changes associated with neoplastic transformation of fibroblasts and epithelial cells by radiation and chemicals, alterations in membrane-associated (Na + + K + )-ATPase and 5'-nucleotidase activities were investigated. Cell cultures consisted of normal and radiation transformed hamster embryo fibroblasts (HE) and mouse C3H 10T 1/2 fibroblasts, normal and chemically transformed adult rat liver epithelial cells (ARL), as well as hepatocarcinoma cells induced by the liver transformants. Transformed fibroblasts demonstrated a 1-2 fold increase in (Na + + K + )-ATPase activity over the normal, while the transformed liver epithelial cells and carcinoma cells showed a 60% and 40% decrease in activity compared to the normal values, respectively. The 5'-nucleotidase activity was 2 to 3 times higher in the transformed fibroblasts
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Wnt-10b promotes differentiation of skin epithelial cells in vitro
International Nuclear Information System (INIS)
Ouji, Yukiteru; Yoshikawa, Masahide; Shiroi, Akira; Ishizaka, Shigeaki
2006-01-01
To evaluate the role of Wnt-10b in epithelial differentiation, we investigated the effects of Wnt-10b on adult mouse-derived primary skin epithelial cells (MPSEC). Recombinant Wnt-10b protein (rWnt-10b) was prepared using a gene engineering technique and MPSEC were cultured in its presence, which resulted in morphological changes from cuboidal to spindle-shaped and inhibited their proliferation. Further, involvement of the canonical Wnt signal pathway was also observed. MPSEC treated with rWnt-10b showed characteristics of the hair shaft and inner root sheath of the hair follicle, in results of Ayoub Shklar staining and immunocytochemistry. Further, the cells expressed mRNA for differentiated epithelial cells, including keratin 1, keratin 2, loricrin, mHa5, and mHb5, in association with a decreased expression of the basal cell marker keratin 5. These results suggest that Wnt-10b promotes the differentiation of MPSEC
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Directory of Open Access Journals (Sweden)
Naoki Tajiri
Full Text Available We recently reported isolation of viable rat amniotic fluid-derived stem (AFS cells [1]. Here, we tested the therapeutic benefits of AFS cells in a rodent model of ischemic stroke. Adult male Sprague-Dawley rats received a 60-minute middle cerebral artery occlusion (MCAo. Thirty-five days later, animals exhibiting significant motor deficits received intravenous transplants of rat AFS cells or vehicle. At days 60-63 post-MCAo, significant recovery of motor and cognitive function was seen in stroke animals transplanted with AFS cells compared to vehicle-infused stroke animals. Infarct volume, as revealed by hematoxylin and eosin (H&E staining, was significantly reduced, coupled with significant increments in the cell proliferation marker, Ki67, and the neuronal marker, MAP2, in the dentate gyrus (DG [2] and the subventricular zone (SVZ of AFS cell-transplanted stroke animals compared to vehicle-infused stroke animals. A significantly higher number of double-labeled Ki67/MAP2-positive cells and a similar trend towards increased Ki67/MAP2 double-labeling were observed in the DG and SVZ of AFS cell-transplanted stroke animals, respectively, compared to vehicle-infused stroke animals. This study reports the therapeutic potential of AFS cell transplantation in stroke animals, possibly via enhancement of endogenous repair mechanisms.
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Son, Eun Suk; Kim, Se-Hee; Ryter, Stefan W; Yeo, Eui-Ju; Kyung, Sun Young; Kim, Yu Jin; Jeong, Sung Hwan; Lee, Chang Soo; Park, Jeong-Woong
2018-04-01
Recent studies demonstrate that the autophagy-dependent turnover of mitochondria (mitophagy) mediates pulmonary epithelial cell death in response to cigarette smoke extract (CSE) exposure, and contributes to emphysema development in vivo during chronic cigarette smoke (CS)-exposure, although the underlying mechanisms remain unclear. Here, we investigated the role of mitophagy in regulating apoptosis in CSE-exposed human lung bronchial epithelial cells. Furthermore, we investigated the potential of the polymethoxylated flavone antioxidant quercetogetin (QUE) to inhibit CSE-induced mitophagy-dependent apoptosis. Our results demonstrate that CSE induces mitophagy in epithelial cells via mitochondrial dysfunction, and causes increased expression levels of the mitophagy-regulator protein PTEN-induced putative kinase-1 (PINK1) and the mitochondrial fission protein dynamin-1-like protein (DRP-1). CSE induced epithelial cell death and increased the expression of the apoptosis-related proteins cleaved caspase-3, -8 and -9. Caspase-3 activity was significantly increased in Beas-2B cells exposed to CSE, and decreased by siRNA-dependent knockdown of DRP-1. Treatment of epithelial cells with QUE inhibited CSE-induced mitochondrial dysfunction and mitophagy by inhibiting phospho (p)-DRP-1 and PINK1 expression. QUE suppressed mitophagy-dependent apoptosis by inhibiting the expression of cleaved caspase-3, -8 and -9 and downregulating caspase activity in human bronchial epithelial cells. These findings suggest that QUE may serve as a potential therapeutic in CS-induced pulmonary diseases. Copyright © 2018 Elsevier Ltd. All rights reserved.
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Limbal Stem Cell Deficiency and Treatment with Stem Cell Transplantation.
Barut Selver, Özlem; Yağcı, Ayşe; Eğrilmez, Sait; Gürdal, Mehmet; Palamar, Melis; Çavuşoğlu, Türker; Ateş, Utku; Veral, Ali; Güven, Çağrı; Wolosin, Jose Mario
2017-10-01
The cornea is the outermost tissue of the eye and it must be transparent for the maintenance of good visual function. The superficial epithelium of the cornea, which is renewed continuously by corneal stem cells, plays a critical role in the permanence of this transparency. These stem cells are localized at the cornea-conjunctival transition zone, referred to as the limbus. When this zone is affected/destroyed, limbal stem cell deficiency ensues. Loss of limbal stem cell function allows colonization of the corneal surface by conjunctival epithelium. Over 6 million people worldwide are affected by corneal blindness, and limbal stem cell deficiency is one of the main causes. Fortunately, it is becoming possible to recover vision by autologous transplantation of limbal cells obtained from the contralateral eye in unilateral cases. Due to the potential risks to the donor eye, only a small amount of tissue can be obtained, in which only 1-2% of the limbal epithelial cells are actually limbal stem cells. Vigorous attempts are being made to expand limbal stem cells in culture to preserve or even enrich the stem cell population. Ex vivo expanded limbal stem cell treatment in limbal stem cell deficiency was first reported in 1997. In the 20 years since, various protocols have been developed for the cultivation of limbal epithelial cells. It is still not clear which method promotes effective stem cell viability and this remains a subject of ongoing research. The most preferred technique for limbal cell culture is the explant culture model. In this approach, a small donor eye limbal biopsy is placed as an explant onto a biocompatible substrate (preferably human amniotic membrane) for expansion. The outgrowth (cultivated limbal epithelial cells) is then surgically transferred to the recipient eye. Due to changing regulations concerning cell-based therapy, the implementation of cultivated limbal epithelial transplantation in accordance with Good Laboratory Practice using
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International Nuclear Information System (INIS)
Castleman, W.L.; Dungworth, D.L.; Schwartz, L.W.; Tyler, W.S.
1980-01-01
The pathogenesis of acute respiratory bronchiolitis was examined in Rhesus monkeys exposed to 0.8 ppM ozone for 4 to 50 hours. Epithelial injury and renewal were qualitatively and quantitatively characterized by correlated techniques of scanning and transmission electron microscopy as well as by light-microscopic autoradiography following labeling with tritiated thymidine. Extensive degeneration and necrosis of Type 1 epithelial cells occurred on the respiratory bronchiolar wall during the initial 4 to 12 hours of exposure. Increased numbers of labeled epithelial cells were present in this region after 18 hours of exposure, and the highest labeling index (18%) was measured after 50 hours of exposure. Most (67 to 80%) of the labeled cells and all the mitotic epithelial cells (22) observed ultrastructurally were cuboidal bronchiolar epithelial cells. Of the labeled epithelial cells, 20 to 33% were Type 2 epithelial cells. After 50 hours of exposure the respiratory bronchiolar epithelium was hyperplastic. The predominant inflammatory cell in respiratory bronchiolar exudate was the alveolar macrophage. Monkeys that were exposed for 50 hours and allowed to recover in unozonized air for 7 days had incomplete resolution of respiratory bronchiolar epithelial hyperplasia. The results indicate that Type 1 epithelial cells lining respiratory bronchioles are the cell types most sensitive to injury and that both cuboidal bronchiolar epithelial cells and Type 2 epithelial cells function as stem cells in epithelial renewal
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Mulder, Daniel J; Pooni, Aman; Mak, Nanette; Hurlbut, David J; Basta, Sameh; Justinich, Christopher J
2011-02-01
Professional antigen-presenting cells (APCs) play a crucial role in initiating immune responses. Under pathological conditions, epithelial cells at mucosal surfaces act as nonprofessional APCs, thereby regulating immune responses at the site of exposure. Epithelial cells in the esophagus may contribute to the pathogenesis of eosinophilic esophagitis (EoE) by presenting antigens on the major histocompatibility complex (MHC) class II. Our goal was to demonstrate the ability of esophageal epithelial cells to process and present antigens on the MHC class II system and to investigate the contribution of epithelial cell antigen presentation to EoE. Immunohistochemistry detected HLA-DR, CD80, and CD86 expression and enzyme-linked immunosorbent assay detected interferon-γ (IFNγ) in esophageal biopsies. Antigen presentation was studied using the human esophageal epithelial cell line HET-1A by reverse transcriptase-PCR, flow cytometry, and confocal microscopy. T helper cell lymphocyte proliferation was assessed by flow cytometry and IL-2 secretion. IFNγ and MHC class II were increased in mucosa of patients with EoE. IFNγ increased mRNA of HLA-DP, HLA-DQ, HLA-DR, and CIITA in HET-1A cells. HET-1A engulfed cell debris and processed ovalbumin. HET-1A cells expressed HLA-DR after IFNγ treatment. HET-1A stimulated T helper cell activation. In this study, we demonstrated the ability of esophageal epithelial cells to act as nonprofessional APCs in the presence of IFNγ. Esophageal epithelial cell antigen presentation may contribute to the pathophysiology of eosinophilic esophagitis. Copyright © 2011 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.
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Lesko, Alyssa C.; Goss, Kathleen H.; Yang, Frank F.; Schwertner, Adam; Hulur, Imge; Onel, Kenan; Prosperi, Jenifer R.
2015-01-01
The Adenomatous Polyposis Coli (APC) tumor suppressor has been previously implicated in the control of apical-basal polarity; yet, the consequence of APC loss-of-function in epithelial polarization and morphogenesis has not been characterized. To test the hypothesis that APC is required for the establishment of normal epithelial polarity and morphogenesis programs, we generated APC-knockdown epithelial cell lines. APC depletion resulted in loss of polarity and multi-layering on permeable supports, and enlarged, filled spheroids with disrupted polarity in 3D culture. Importantly, these effects of APC knockdown were independent of Wnt/β-catenin signaling, but were rescued with either full-length or a carboxy (c)-terminal segment of APC. Moreover, we identified a gene expression signature associated with APC knockdown that points to several candidates known to regulate cell-cell and cell-matrix communication. Analysis of epithelial tissues from mice and humans carrying heterozygous APC mutations further support the importance of APC as a regulator of epithelial behavior and tissue architecture. These data also suggest that the initiation of epithelial-derived tumors as a result of APC mutation or gene silencing may be driven by loss of polarity and dysmorphogenesis. PMID:25578398
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Nerve Invasion by Epithelial Cells in Benign Breast Diseases
Directory of Open Access Journals (Sweden)
Yu-Jan Chan
2009-03-01
Full Text Available Nerve invasion by glandular epithelial cells in a lesion is usually regarded as invasive carcinoma. However, some benign conditions in the pancreas, prostate, breast and other organs may show involvement of nerve bundles by benign epithelial cells. We report an 18-year-old female with nerve invasion in benign breast disease. The lesion in her right breast revealed fibrocystic changes with ductal hyperplasia and stromal sclerosis. Perineural and intraneural involvement by bland-looking small ducts lined by 2 layers of cells including an outer layer of myoepithelial cells were found, suggestive of benign nerve invasion. There was no evidence of malignant cells in any of the sections. The patient remains well after 31 months of follow-up. About 44 cases of nerve invasion in benign breast diseases have been reported in the literature. It is necessary to carefully evaluate nerve involvement in breast lesions to avoid over-diagnosis and inappropriate operation.
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Hypoxia is a key regulator of limbal epithelial stem cell growth and differentiation
DEFF Research Database (Denmark)
Bath, Chris; Yang, Sufang; Muttuvelu, Danson
2013-01-01
The aim of this study was to determine whether the growth and differentiation of limbal epithelial stem cell cultures could be controlled through manipulation of the oxygen tension. Limbal epithelial cells were isolated from corneoscleral disks, and cultured using either feeder cells in a growth......, progression through cell cycle, colony forming efficiency (CFE), and expression of stem cell (ABCG2 and p63α) and differentiation (CK3) markers was determined throughout the culture period of up to 18 days. Low oxygen levels favored a stem cell phenotype with a lower proliferative rate, high CFE......, and a relatively higher expression of ABCG2 and p63α, while higher levels of oxygen led not only to decreased CFE but also to increased proportion of differentiated cells positive for CK3. Hypoxic cultures may thus potentially improve stem cell grafts for cultured limbal epithelial transplantation (CLET)....
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Chromosomal changes in cultured human epithelial cells transformed by low- and high-LET radiation
Energy Technology Data Exchange (ETDEWEB)
Yang, Tracy Chui-hsu; Craise, L.M; Prioleau, J.C.; Stampfer, M.R.; Rhim, J.S.
1990-11-01
For a better assessment of radiation risk in space, an understanding of the responses of human cells, especially the epithelial cells, to low- and high-LET radiation is essential. In our laboratory, we have successfully developed techniques to study the neoplastic transformation of two human epithelial cell systems by ionizing radiation. These cell systems are human mammary epithelial cells (H184B5) and human epidermal keratinocytes (HEK). Both cell lines are immortal, anchorage dependent for growth, and nontumorigenic in athymic nude nice. Neoplastic transformation was achieved by irradiation cells successively. Our results showed that radiogenic cell transformation is a multistep process and that a single exposure of ionizing radiation can cause only one step of transformation. It requires, therefore, multihits to make human epithelial cells fully tumorigenic. Using a simple karyotyping method, we did chromosome analysis with cells cloned at various stages of transformation. We found no consistent large terminal deletion of chromosomes in radiation-induced transformants. Some changes of total number of chromosomes, however, were observed in the transformed cells. These transformants provide an unique opportunity for further genetic studies at a molecular level. 15 refs., 9 figs., 2 tabs.
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Chromosomal changes in cultured human epithelial cells transformed by low- and high-LET radiation
International Nuclear Information System (INIS)
Yang, Tracy Chui-hsu; Craise, L.M; Prioleau, J.C.; Stampfer, M.R.; Rhim, J.S.
1990-11-01
For a better assessment of radiation risk in space, an understanding of the responses of human cells, especially the epithelial cells, to low- and high-LET radiation is essential. In our laboratory, we have successfully developed techniques to study the neoplastic transformation of two human epithelial cell systems by ionizing radiation. These cell systems are human mammary epithelial cells (H184B5) and human epidermal keratinocytes (HEK). Both cell lines are immortal, anchorage dependent for growth, and nontumorigenic in athymic nude nice. Neoplastic transformation was achieved by irradiation cells successively. Our results showed that radiogenic cell transformation is a multistep process and that a single exposure of ionizing radiation can cause only one step of transformation. It requires, therefore, multihits to make human epithelial cells fully tumorigenic. Using a simple karyotyping method, we did chromosome analysis with cells cloned at various stages of transformation. We found no consistent large terminal deletion of chromosomes in radiation-induced transformants. Some changes of total number of chromosomes, however, were observed in the transformed cells. These transformants provide an unique opportunity for further genetic studies at a molecular level. 15 refs., 9 figs., 2 tabs
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Lee, C A; Silva, M; Siber, A M; Kelly, A J; Galyov, E; McCormick, B A
2000-10-24
In response to Salmonella typhimurium, the intestinal epithelium generates an intense inflammatory response consisting largely of polymorphonuclear leukocytes (neutrophils, PMN) migrating toward and ultimately across the epithelial monolayer into the intestinal lumen. It has been shown that bacterial-epithelial cell interactions elicit the production of inflammatory regulators that promote transepithelial PMN migration. Although S. typhimurium can enter intestinal epithelial cells, bacterial internalization is not required for the signaling mechanisms that induce PMN movement. Here, we sought to determine which S. typhimurium factors and intestinal epithelial signaling pathways elicit the production of PMN chemoattractants by enterocytes. Our results suggest that S. typhimurium activates a protein kinase C-dependent signal transduction pathway that orchestrates transepithelial PMN movement. We show that the type III effector protein, SipA, is not only necessary but is sufficient to induce this proinflammatory response in epithelial cells. Our results force us to reconsider the long-held view that Salmonella effector proteins must be directly delivered into host cells from bacterial cells.
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Barrera, C; Ye, G; Espejo, R; Gunasena, S; Almanza, R; Leary, J; Crowe, S; Ernst, P; Reyes, V E
2001-10-01
Helicobacter pylori infection is linked to chronic gastritis, peptic ulcer and gastric carcinoma. During H. pylori infection, class II MHC expression by the gastric epithelium increases, as does the number of local CD4(+) T cells, which appear to be important in the associated pathogenesis. These observations suggested that the epithelium might present antigens to T cells. Thus, we sought to determine whether gastric epithelial cells process antigens to establish their function as local antigen presenting cells (APC). We examined a panel of gastric epithelial cell lines for expression of the antigen processing cathepsins B (CB), L (CL), S (CS), and D (CD). The mRNA for these enzymes were detected by RT-PCR and the enzymes in the gastric epithelial cells were identified by various independent methods. We corroborated the expression of CB and CD on gastric epithelial cells from human biopsy samples. The functions of these proteases were confirmed by assessing their ability to digest ovalbumin, a conventional dietary antigen, and proteins from H. pylori. In summary, multiple lines of evidence suggest gastric epithelial cells process antigens for presentation to CD4(+) T cells. To our knowledge, these are the first studies to document the antigen processing capacity of human gastric epithelial cells.
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Jang, Jun-Ho; Chand, Hitendra S; Bruse, Shannon; Doyle-Eisele, Melanie; Royer, Christopher; McDonald, Jacob; Qualls, Clifford; Klingelhutz, Aloysius J; Lin, Yong; Mallampalli, Rama; Tesfaigzi, Yohannes; Nyunoya, Toru
2017-04-01
The purpose of this study was to determine whether expression of connective tissue growth factor (CTGF) protein in chronic obstructive pulmonary disease (COPD) is consistent in humans and animal models of COPD and to investigate the role of this protein in lung epithelial cells. CTGF in lung epithelial cells of ex-smokers with COPD was compared with ex-smokers without COPD by immunofluorescence. A total of twenty C57Bl/6 mice and sixteen non-human primates (NHPs) were exposed to cigarette smoke (CS) for 4 weeks. Ten mice of these CS-exposed mice and eight of the CS-exposed NHPs were infected with H3N2 influenza A virus (IAV), while the remaining ten mice and eight NHPs were mock-infected with vehicle as control. Both mRNA and protein expression of CTGF in lung epithelial cells of mice and NHPs were determined. The effects of CTGF overexpression on cell proliferation, p16 protein, and senescence-associated β-galactosidase (SA-β-gal) activity were examined in cultured human bronchial epithelial cells (HBECs). In humans, CTGF expression increased with increasing COPD severity. We found that protein expression of CTGF was upregulated in lung epithelial cells in both mice and NHPs exposed to CS and infected with IAV compared to those exposed to CS only. When overexpressed in HBECs, CTGF accelerated cellular senescence accompanied by p16 accumulation. Both CTGF and p16 protein expression in lung epithelia are positively associated with the severity of COPD in ex-smokers. These findings show that CTGF is consistently expressed in epithelial cells of COPD lungs. By accelerating lung epithelial senescence, CTGF may block regeneration relative to epithelial cell loss and lead to emphysema.
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Aquaporin 2 promotes cell migration and epithelial morphogenesis.
Chen, Ying; Rice, William; Gu, Zhizhan; Li, Jian; Huang, Jianmin; Brenner, Michael B; Van Hoek, Alfred; Xiong, Jianping; Gundersen, Gregg G; Norman, Jim C; Hsu, Victor W; Fenton, Robert A; Brown, Dennis; Lu, Hua A Jenny
2012-09-01
The aquaporin 2 (AQP2) water channel, expressed in kidney collecting ducts, contributes critically to water homeostasis in mammals. Animals lacking or having significantly reduced levels of AQP2, however, have not only urinary concentrating abnormalities but also renal tubular defects that lead to neonatal mortality from renal failure. Here, we show that AQP2 is not only a water channel but also an integrin-binding membrane protein that promotes cell migration and epithelial morphogenesis. AQP2 expression modulates the trafficking and internalization of integrin β1, facilitating its turnover at focal adhesions. In vitro, disturbing the interaction between AQP2 and integrin β1 by mutating the RGD motif led to reduced endocytosis, retention of integrin β1 at the cell surface, and defective cell migration and tubulogenesis. Similarly, in vivo, AQP2-null mice exhibited significant retention of integrin β1 at the basolateral membrane and had tubular abnormalities. In summary, these data suggest that the water channel AQP2 interacts with integrins to promote renal epithelial cell migration, contributing to the structural and functional integrity of the mammalian kidney.
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Directory of Open Access Journals (Sweden)
Amie L. Holmes
2013-11-01
Full Text Available Nickel is a well-known human lung carcinogen with the particulate form being the most potent; however, the carcinogenic mechanism remains largely unknown. Few studies have investigated the genotoxicity and carcinogenicity of nickel in its target cell, human bronchial epithelial cells. Thus, the goal of this study was to investigate the effects of particulate nickel in human lung epithelial cells. We found that nickel subsulfide induced concentration- and time-dependent increases in both cytotoxicity and genotoxicity in human lung epithelial cells (BEP2D. Chronic exposure to nickel subsulfide readily induced cellular transformation, inducing 2.55, 2.9 and 2.35 foci per dish after exposure to 1, 2.5 and 5 μg/cm2 nickel subsulfide, respectively. Sixty-one, 100 and 70 percent of the foci isolated from 1, 2.5, and 5 μg/cm2 nickel subsulfide treatments formed colonies in soft agar and the degree of soft agar colony growth increased in a concentration-dependent manner. Thus, chronic exposure to particulate nickel induces genotoxicity and cellular transformation in human lung epithelial cells.
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Lange, Alexander W.; Sridharan, Anusha; Xu, Yan; Stripp, Barry R.; Perl, Anne-Karina; Whitsett, Jeffrey A.
2015-01-01
The Hippo/Yap pathway is a well-conserved signaling cascade that regulates cell proliferation and differentiation to control organ size and stem/progenitor cell behavior. Following airway injury, Yap was dynamically regulated in regenerating airway epithelial cells. To determine the role of Hippo signaling in the lung, the mammalian Hippo kinases, Mst1 and Mst2, were deleted in epithelial cells of the embryonic and mature mouse lung. Mst1/2 deletion in the fetal lung enhanced proliferation and inhibited sacculation and epithelial cell differentiation. The transcriptional inhibition of cell proliferation and activation of differentiation during normal perinatal lung maturation were inversely regulated following embryonic Mst1/2 deletion. Ablation of Mst1/2 from bronchiolar epithelial cells in the adult lung caused airway hyperplasia and altered differentiation. Inhibitory Yap phosphorylation was decreased and Yap nuclear localization and transcriptional targets were increased after Mst1/2 deletion, consistent with canonical Hippo/Yap signaling. YAP potentiated cell proliferation and inhibited differentiation of human bronchial epithelial cells in vitro. Loss of Mst1/2 and expression of YAP regulated transcriptional targets controlling cell proliferation and differentiation, including Ajuba LIM protein. Ajuba was required for the effects of YAP on cell proliferation in vitro. Hippo/Yap signaling regulates Ajuba and controls proliferation and differentiation of lung epithelial progenitor cells. PMID:25480985
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Lin, Louis M; Huang, George T-J; Rosenberg, Paul A
2007-08-01
There is continuing controversy regarding the potential for inflammatory apical cysts to heal after nonsurgical endodontic therapy. Molecular cell biology may provide answers to a series of related questions. How are the epithelial cell rests of Malassez stimulated to proliferate? How are the apical cysts formed? How does the lining epithelium of apical cysts regress after endodontic therapy? Epithelial cell rests are induced to divide and proliferate by inflammatory mediators, proinflammatory cytokines, and growth factors released from host cells during periradicular inflammation. Quiescent epithelial cell rests can behave like restricted-potential stem cells if stimulated to proliferate. Formation of apical cysts is most likely caused by the merging of proliferating epithelial strands from all directions to form a three-dimensional ball mass. After endodontic therapy, epithelial cells in epithelial strands of periapical granulomas and the lining epithelium of apical cysts may stop proliferating because of a reduction in inflammatory mediators, proinflammatory cytokines, and growth factors. Epithelial cells will also regress because of activation of apoptosis or programmed cell death through deprivation of survival factors or by receiving death signals during periapical wound healing.
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ELF5 in epithelial ovarian carcinoma tissues and biological behavior in ovarian carcinoma cells.
Yan, Hongchao; Qiu, Linglin; Xie, Xiaolei; Yang, He; Liu, Yongli; Lin, Xiaoman; Huang, Hongxiang
2017-03-01
The expression of E74-like factor 5 (ELF5) in epithelial ovarian carcinoma tissues and its effects on biological behavior in ovarian carcinoma cells were assessed in search for a new approach for gene treatment of epithelial ovarian carcinoma. RT-PCR technology was applied to detect the expression of ELF5 mRNA in epithelial ovarian carcinoma (n=49), borderline ovarian epithelial tumor (n=19), benign ovarian epithelial tumor (n=31) and normal ovarian tissues (n=40). Then, we transfected recombinant plasmid pcDNA3.1‑ELF5+EGFP into human ovarian carcinoma SKOV3 cells (recombinant plasmid group) in vitro and screened out stably transfected cells to conduct multiplication culture. Western blot analysis was performed to detect the expression of ELF5 protein in the different groups. Flow cytometry was employed to detect cell apoptosis and cycles. ELF5 mRNA in epithelial ovarian carcinoma and borderline ovarian epithelial tumor tissues were significantly lower (Pepithelial tumor and normal ovarian tissues. ELF5 protein expression in the cells of recombinant plasmid group was significantly higher compared with empty plasmid and blank control groups. The capacity of cell reproductive recombinant plasmid group at each time point decreased (P<0.05). Flow cytometry detection showed that 67.03% of cells in recombinant plasmid group was blocked in G0/G1 phase (P<0.05), compared with empty plasmid group (37.17%) and blank control group (38.24%). Apoptotic rate of recombinant plasmid group was significantly lower (31.4±1.9%; P<0.05), compared with that of empty plasmid group (9.1±2.2%) and blank control group (8.7±1.5%), and the differences were statistically significant. In conclusion, ELF5 interfered with cell cycle of human ovarian carcinoma SKOV3 cells and promoted apoptosis of human ovarian carcinoma SKOV3 cells inhibiting their growth and invasive capacity; and thus providing a new approach to gene treatment of ovarian carcinoma.
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Liu, Daqing; Yovchev, Mladen I.; Zhang, Jinghang; Alfieri, Alan A.; Tchaikovskaya, Tatyana; Laconi, Ezio; Dabeva, Mariana D.
2016-01-01
In normal rat liver, thymocyte antigen 1 (Thy1) is expressed in fibroblasts/myofibroblasts and in some blood progenitor cells. Thy1-expressing cells also accumulate in the liver during impaired liver regeneration. The origin and nature of these cells are not well understood. By using RT-PCR analysis and immunofluorescence microscopy, we describe the presence of rare Thy1+ cells in the liver lobule of normal animals, occasionally forming small collections of up to 20 cells. These cells constitute a small portion (1.7% to 1.8%) of nonparenchymal cells and reveal a mixed mesenchymal-epithelial phenotype, expressing E-cadherin, cytokeratin 18, and desmin. The most potent mitogens for mesenchymal-epithelial Thy1+ cells in vitro are the inflammatory cytokines interferon γ, IL-1, and platelet-derived growth factor-BB, which are not produced by Thy1+ cells. Thy1+ cells express all typical mesenchymal stem cell and hepatic progenitor cell markers and produce growth factor and cytokine mRNA (Hgf, Il6, Tgfa, and Tweak) for proteins that maintain oval cell growth and differentiation. Under appropriate conditions, mesenchymal-epithelial cells differentiate in vitro into hepatocyte-like cells. In this study, we show that the adult rat liver harbors a small pool of endogenous mesenchymal-epithelial cells not recognized previously. In the quiescent state, these cells express both mesenchymal and epithelial cell markers. They behave like hepatic stem cells/progenitors with dual phenotype, exhibiting high plasticity and long-lasting proliferative activity. PMID:25447047
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Apoptosis induction of human endometriotic epithelial and stromal cells by noscapine
Directory of Open Access Journals (Sweden)
Mohammad Rasoul Khazaei
2016-09-01
Full Text Available Objective(s: Endometriosis is a complex gynecologic disease with unknown etiology. Noscapine has been introduced as a cancer cell suppressor. Endometriosis was considered as a cancer like disorder, The aim of present study was to investigate noscapine apoptotic effect on human endometriotic epithelial and stromal cells in vitro. Materials and Methods:In this in vitro study, endometrial biopsies from endometriosis patients (n=9 were prepared and digested by an enzymatic method (collagenase I, 2 mg/ml. Stromal and epithelial cells were separated by sequential filtration through a cell strainer and ficoll layering. The cells of each sample were divided into five groups: control (0, 10, 25, 50 and 100 micromole/liter (µM concentration of noscapine and were cultured for three different periods of times; 24, 48 and 72 hr. Cell viability was assessed by colorimetric assay. Nitric oxide (NO concentration was measured by Griess reagent. Cell death was analyzed by Acridine Orange (AO–Ethidium Bromide (EB double staining and Terminal deoxynucleotidyl transferase (TdT dUTP Nick-End Labeling (TUNEL assay. Data were analyzed by one-way ANOVA. Results: Viability of endometrial epithelial and stromal cells significantly decreased in 10, 25, 50 and 100 µM noscapine concentration in 24, 48, 72 hr (P
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α-Crystallin localizes to the leading edges of migrating lens epithelial cells
International Nuclear Information System (INIS)
Maddala, Rupalatha; Vasantha Rao, P.
2005-01-01
α-crystallin (αA and αB) is a major lens protein, which belongs to the small heat-shock family of proteins and binds to various cytoskeletal proteins including actin, vimentin and desmin. In this study, we investigated the cellular localization of αA and αB-crystallins in migrating epithelial cells isolated from porcine lens. Immunofluorescence localization and confocal imaging of αB-crystallin in confluent and in migrating subconfluent cell cultures revealed a distinct pattern of subcellular distribution. While αB-crystallin localization was predominantly cytoplasmic in confluent cultures, it was strongly localized to the leading edges of cell membrane or the lamellipodia in migrating cells. In accordance with this pattern, we found abundant levels of αB-crystallin in membrane fractions compared to cytosolic and nuclear fractions in migrating lens epithelial cells. αA-crystallin, which has 60% sequence identity to αB-crystallin, also exhibited a distribution profile localizing to the leading edge of the cell membrane in migrating lens epithelial cells. Localization of αB-crystallin to the lamellipodia appears to be dependent on phosphorylation of residue serine-59. An inhibitor of p38 MAP kinase (SB202190), but not the ERK kinase inhibitor PD98059, was found to diminish localization of αB-crystallin to the lamellipodia, and this effect was found to be associated with reduced levels of Serine-59 phosphorylated αB-crystallin in SB202190-treated migrating lens epithelial cells. αB-crystallin localization to the lamellipodia was also altered by the treatment with RGD (Arg-Ala-Asp) peptide, dominant negative N17 Rac1 GTPase, cytochalasin D and Src kinase inhibitor (PP2), but not by the Rho kinase inhibitor Y-27632 or the myosin II inhibitor, blebbistatin. Additionally, in migrating lens epithelial cells, αB-crystallin exhibited a clear co-localization with the actin meshwork, β-catenin, WAVE-1, a promoter of actin nucleation, Abi-2, a component of WAVE
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Jin, Ying; Li, Yuwan; Zhang, Chenghao; Wu, Shuhong; Cheng, Daixiong; Liu, Yi
2016-02-01
To discuss whether human amniotic mesenchymal stem cells (hAMSCs) possesses the characteristic of mesenchymal stem cells, and could differentiate into ligament cells in vitro after induction. The hAMSCs were separated through enzyme digestion, and the phenotypic characteristics of hAMSCs were tested through flow cytometry. The cells at passage 3 were cultured with L-DMEM/F12 medium containing transforming growth factor beta1 (TGF-beta1) + basic fibroblast growth factor (bFGF) (group A), containing hyaluronic acid (HA) (group B), containing TGF-beta1+bFGF+HA (group C), and simple L-DMEM/F12 medium (group D) as control group. The morphology changes of cells in each group were observed by inverted phase contrast microscope at 21 days after induction; the cellular activities and proliferation were examined by sulforhodamine (SRB) colorimetric method; and specific mRNA and protein expressions of ligament including collagen type I, collagen type III, and tenascin C (TNC) were measured by real-time fluorescence quantitative PCR and immunohistochemical staining. The flow cytometry result indicated that hAMSCs expressed mesenchymal stem cell phenotype. After 21 days of induction, the cells in groups A, B, and C grew like spindle-shaped fibroblasts under inverted phase contrast microscope, and cells showed single shape, obvious directivity, and compact arrangement in group C. The SRB result indicated that the cells in each group reached the peak of growth curve at 6 days; the cellular activities of groups A, B, and C were significantly higher than that of group D at 6 days after induction. Also, the immunohistochemical staining results showed that no expressions of TNC were detected in 4 groups at 7 days; expressions of collagen type I in groups A, B, and C were significantly higher than that in group D at 7, 14, and 21 days (Pligament specific genes can be up-regulated and the synthesis of ligament specific proteins can be also strengthened. As a result, it can be used as
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Protective effects of trehalose on the corneal epithelial cells.
Aragona, Pasquale; Colosi, Pietro; Rania, Laura; Colosi, Francesca; Pisani, Antonina; Puzzolo, Domenico; Micali, Antonio
2014-01-01
Aim of the present work was to evaluate the effects of the trehalose on the corneal epithelium undergoing alcohol delamination. Twelve patients undergoing laser subepithelial keratomileusis (LASEK) were consecutively included in the study. The right eyes were pretreated with 3% trehalose eye drops, whilst left eyes were used as control. Epithelial specimens were processed for cells vitality assessment, apoptosis, and light and transmission electron microscopy; a morphometric analysis was performed in both groups. In both trehalose-untreated eyes (TUE) and trehalose-treated eyes (TTE), the percentage of vital cells was similar and no apoptotic cells were observed. In TUE, the corneal epithelium showed superficial cells with reduced microfolds, wing cells with vesicles and dilated intercellular spaces, and dark basal cells with vesicles and wide clefts. In TTE, superficial and wing cells were better preserved, and basal cells were generally clear with intracytoplasmatic vesicles. The morphometric analysis showed statistically significant differences between the two groups: the TTE epithelial height was higher, the basal cells showed larger area and clearer cytoplasm. The distribution of desmosomes and hemidesmosomes was significantly different between the groups. Trehalose administration better preserved morphological and morphometric features of alcohol-treated corneal epithelium, when compared to controls.
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TCDD alters medial epithelial cell differentiation during palatogenesis
International Nuclear Information System (INIS)
Abbott, B.D.; Birnbaum, L.S.
1989-01-01
2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is a widely distributed, persistent environmental contaminant that is teratogenic in mice, where it induces hydronephrosis and cleft palate. The incidence of clefting has been shown to be dose dependent after exposure on either gestation Day (GD) 10 or 12, although the embryo is more susceptible on GD 12. TCDD-exposed palatal shelves meet but do not fuse, and programmed cell death of the medial epithelial cells is inhibited. The mechanism of action through which TCDD alters the program of medial cell development has not been examined in earlier studies, and it is not known whether the mechanism is the same regardless of the dose or developmental stage of exposure. In this study, C57BL/6N mice, a strain sensitive to TCDD, were dosed orally on GD 10 or 12 with 0, 6, 12, 24, or 30 micrograms/kg body wt, in 10 ml corn oil/kg. Embryonic palatal shelves were examined on GD 14, 15, or 16. The degree of palatal closure, epithelial surface morphology, and cellular ultrastructure, the incorporation of [3H]TdR, the expression of EGF receptors, and the binding of 125I-EGF were assessed. After exposure on GD 10 or 12, TCDD altered the differentiation pathway of the medial epithelial cells. The palatal shelves were of normal size and overall morphology, but fusion of the medial epithelia of the opposing shelves did not occur. TCDD prevented programmed cell death of the medial peridermal cells. The expression of EGF receptors by medial cells continued through Day 16 and the receptors were able to bind ligand. The medial cells differentiated into a stratified, squamous, keratinizing epithelium. The shift in phenotype to an oral-like epithelium occurred after exposure on either GD 10 or 12. At the lower dose (6 micrograms/kg), fewer cleft palates were produced, but those shelves which did respond had a fully expressed shift in differentiation
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Transfection of normal human bronchial epithelial cells with the bcl-2 oncogene
Energy Technology Data Exchange (ETDEWEB)
Kennedy, C.H.; Kenyon, K.D.; Tesfaigzi, J. [and others
1995-12-01
In vitro, studies examining the transformation of virus-immortalized human bronchial epithelial (HBE) cells after exposure to chemical and physical carcinogens have contributed to our understanding of the mechanisms that underlie the development of lung cancer. Virus-immortalized HBE cells have been used because of both the limited life span of normal human bronchial epithelial (NHBE) cells in culture (approximately 30-35 population doublins) and their resistance to in vitro malignant transformation. For example, human papillomavirus (HPV)-immortalized HBE cells have been used to study the genetic changes that occur after exposure to {alpha}-particles in vitro. Although this model may prove to be useful for studying the 18% or less of bronchogenic carcinomas found to contain HPV sequences, it is not an appropriate model for studying the majority of lung epithelial malignancies in which HPV DNA is not detected. This view is supported by the fact that HPV-immortalized cell lines commonly exhibit aneuploidy. This results of this study suggest that: (1) NHBE cells can be transiently transfected with the pCMV{Beta} vector; and (2) the antibiotic hygromycin-resistant transfected cells.
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Transfection of normal human bronchial epithelial cells with the bcl-2 oncogene
International Nuclear Information System (INIS)
Kennedy, C.H.; Kenyon, K.D.; Tesfaigzi, J.
1995-01-01
In vitro, studies examining the transformation of virus-immortalized human bronchial epithelial (HBE) cells after exposure to chemical and physical carcinogens have contributed to our understanding of the mechanisms that underlie the development of lung cancer. Virus-immortalized HBE cells have been used because of both the limited life span of normal human bronchial epithelial (NHBE) cells in culture (approximately 30-35 population doublins) and their resistance to in vitro malignant transformation. For example, human papillomavirus (HPV)-immortalized HBE cells have been used to study the genetic changes that occur after exposure to α-particles in vitro. Although this model may prove to be useful for studying the 18% or less of bronchogenic carcinomas found to contain HPV sequences, it is not an appropriate model for studying the majority of lung epithelial malignancies in which HPV DNA is not detected. This view is supported by the fact that HPV-immortalized cell lines commonly exhibit aneuploidy. This results of this study suggest that: (1) NHBE cells can be transiently transfected with the pCMVΒ vector; and (2) the antibiotic hygromycin-resistant transfected cells
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Directory of Open Access Journals (Sweden)
Ricardo Chaves Vilela
2013-02-01
Full Text Available Trichomonas vaginalis and Tritrichomonas foetus are parasitic protists of the human and bovine urogenital tracts, respectively. Several studies have described the cytotoxic effects of trichomonads on urogenital tract epithelial cells. However, little is known about the host cell response against trichomonads. The aim of this study was to determine whether T. foetus and T. vaginalis stimulated the release of the cytokine interleukin (IL-10 from cultured bovine epithelial cells. To characterise the inflammatory response induced by these parasites, primary cultures of bovine oviduct epithelial cells were exposed to either T. vaginalis or T. foetus. Within 12 h after parasite challenge, supernatants were collected and cytokine production was analysed. Large amounts of IL-10 were detected in the supernatants of cultures that had been stimulated with T. foetus. Interestingly, T. vaginalis induced only a small increase in the release of IL-10 upon exposure to the same bovine cells. Thus, the inflammatory response of the host cell is species-specific. Only T. foetus and not T. vaginalis induced the release of IL-10 by bovine oviduct epithelial cells.
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Directory of Open Access Journals (Sweden)
Fiona L Cousins
Full Text Available BACKGROUND: In women dynamic changes in uterine tissue architecture occur during each menstrual cycle. Menses, characterised by the shedding of the upper functional layer of the endometrium, is the culmination of a cascade of irreversible changes in tissue function including stromal decidualisation, inflammation and production of degradative enzymes. The molecular mechanisms that contribute to the rapid restoration of tissue homeostasis at time of menses are poorly understood. METHODOLOGY: A modified mouse model of menses was developed to focus on the events occurring within the uterine lining during endometrial shedding/repair. Decidualisation, vaginal bleeding, tissue architecture and cell proliferation were evaluated at 4, 8, 12, and 24 hours after progesterone (P4 withdrawal; mice received a single injection of bromodeoxyuridine (BrdU 90 mins before culling. Expression of genes implicated in the regulation of mesenchymal to epithelial transition (MET was determined using a RT2 PCR profiler array, qRTPCR and bioinformatic analysis. PRINCIPAL FINDINGS: Mice exhibited vaginal bleeding between 4 and 12 hours after P4 withdrawal, concomitant with detachment of the decidualised cell mass from the basal portion of the endometrial lining. Immunostaining for BrdU and pan cytokeratin revealed evidence of epithelial cell proliferation and migration. Cells that appeared to be in transition from a mesenchymal to an epithelial cell identity were identified within the stromal compartment. Analysis of mRNAs encoding genes expressed exclusively in the epithelial or stromal compartments, or implicated in MET, revealed dynamic changes in expression, consistent with a role for reprogramming of mesenchymal cells so that they could contribute to re-epithelialisation. CONCLUSIONS/SIGNIFICANCE: These studies have provided novel insights into the cellular processes that contribute to re-epithelialisation post-menses implicating both epithelial cell migration and
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Cousins, Fiona L; Murray, Alison; Esnal, Arantza; Gibson, Douglas A; Critchley, Hilary O D; Saunders, Philippa T K
2014-01-01
In women dynamic changes in uterine tissue architecture occur during each menstrual cycle. Menses, characterised by the shedding of the upper functional layer of the endometrium, is the culmination of a cascade of irreversible changes in tissue function including stromal decidualisation, inflammation and production of degradative enzymes. The molecular mechanisms that contribute to the rapid restoration of tissue homeostasis at time of menses are poorly understood. A modified mouse model of menses was developed to focus on the events occurring within the uterine lining during endometrial shedding/repair. Decidualisation, vaginal bleeding, tissue architecture and cell proliferation were evaluated at 4, 8, 12, and 24 hours after progesterone (P4) withdrawal; mice received a single injection of bromodeoxyuridine (BrdU) 90 mins before culling. Expression of genes implicated in the regulation of mesenchymal to epithelial transition (MET) was determined using a RT2 PCR profiler array, qRTPCR and bioinformatic analysis. Mice exhibited vaginal bleeding between 4 and 12 hours after P4 withdrawal, concomitant with detachment of the decidualised cell mass from the basal portion of the endometrial lining. Immunostaining for BrdU and pan cytokeratin revealed evidence of epithelial cell proliferation and migration. Cells that appeared to be in transition from a mesenchymal to an epithelial cell identity were identified within the stromal compartment. Analysis of mRNAs encoding genes expressed exclusively in the epithelial or stromal compartments, or implicated in MET, revealed dynamic changes in expression, consistent with a role for reprogramming of mesenchymal cells so that they could contribute to re-epithelialisation. These studies have provided novel insights into the cellular processes that contribute to re-epithelialisation post-menses implicating both epithelial cell migration and mesenchymal cell differentiation in restoration of an intact epithelial cell layer. These
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DEFF Research Database (Denmark)
Water, Jorrit Jeroen; Smart, Simon; Franzyk, Henrik
2015-01-01
intracellularly in Calu-3 epithelial cells and in THP-1 cells, whereas A549 cells did not show significant uptake of nanoparticles. Overall, encapsulation of plectasin into PLGA-based nanoparticles appears to be a viable strategy to improve the efficacy of plectasin against infections in epithelial tissues....... epithelial cells might thus be a promising approach to combat such infections. In this work, plectasin, which is a cationic AMP of the defensin class, was encapsulated into poly(lactic-co-glycolic acid) (PLGA) nanoparticles using the double emulsion solvent evaporation method. The nanoparticles displayed...... high plectasin encapsulation efficiency (71-90%) and mediated release of the peptide over 24h. The antimicrobial efficacy of the peptide-loaded nanoparticles was investigated using bronchiolar epithelial Calu-3 cell monolayers infected with S. aureus. The plectasin-loaded nanoparticles displayed...
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Regulation of defensive function on gingival epithelial cells can prevent periodontal disease.
Fujita, Tsuyoshi; Yoshimoto, Tetsuya; Kajiya, Mikihito; Ouhara, Kazuhisa; Matsuda, Shinji; Takemura, Tasuku; Akutagawa, Keiichi; Takeda, Katsuhiro; Mizuno, Noriyoshi; Kurihara, Hidemi
2018-05-01
Periodontal disease is a bacterial biofilm-associated inflammatory disease that has been implicated in many systemic diseases. A new preventive method for periodontal disease needs to be developed in order to promote the health of the elderly in a super-aged society. The gingival epithelium plays an important role as a mechanical barrier against bacterial invasion and a part of the innate immune response to infectious inflammation in periodontal tissue. The disorganization of cell-cell interactions and subsequent inflammation contribute to the initiation of periodontal disease. These make us consider that regulation of host defensive functions, epithelial barrier and neutrophil activity, may become novel preventive methods for periodontal inflammation. Based on this concept, we have found that several agents regulate the barrier function of gingival epithelial cells and suppress the accumulation of neutrophils in the gingival epithelium. We herein introduce the actions of irsogladine maleate, azithromycin, amphotericin B, and Houttuynia cordata (dokudami in Japanese), which is commonly used in traditional medicine, on the epithelial barrier and neutrophil migration in gingival epithelial cells in vivo and in vitro , in order to provide support for the clinical application of these agents to the prevention of periodontal inflammation.
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The regenerative potential of parietal epithelial cells in adult mice
Berger, K.; Schulte, K.; Boor, P.; Kuppe, C.; Kuppevelt, T.H. van; Floege, J.; Smeets, B.; Moeller, M.J.
2014-01-01
Previously, we showed that some podocytes in juvenile mice are recruited from cells lining Bowman's capsule, suggesting that parietal epithelial cells (PECs) are a progenitor cell population for podocytes. To investigate whether PECs also replenish podocytes in adult mice, PECs were genetically
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Lee, Yeo Song; Lee, Do Yeon; Yu, Da Yeon; Kim, Shin; Lee, Yong Chan
2014-12-01
Chronic infection with Helicobacter pylori (H. pylori) is causally linked with gastric carcinogenesis. Virulent H. pylori strains deliver bacterial CagA into gastric epithelial cells. Induction of high motility and an elongated phenotype is considered to be CagA-dependent process. Casein kinase 2 plays a critical role in carcinogenesis through signaling pathways related to the epithelial mesenchymal transition. This study was aimed to investigate the effect of H. pylori infection on the casein kinase 2-mediated migration and invasion in gastric epithelial cells. AGS or MKN28 cells as human gastric epithelial cells and H. pylori strains Hp60190 (ATCC 49503, CagA(+)) and Hp8822 (CagA(-)) were used. Cells were infected with H. pylori at multiplicity of infection of 100 : 1 for various times. We measured in vitro kinase assay to examine casein kinase 2 activity and performed immunofluorescent staining to observe E-cadherin complex. We also examined β-catenin transactivation through promoter assay and MMP7 expression by real-time PCR and ELISA. H. pylori upregulates casein kinase 2 activity and inhibition of casein kinase 2 in H. pylori-infected cells profoundly suppressed cell invasiveness and motility. We confirmed that casein kinase 2 mediates membranous α-catenin depletion through dissociation of the α-/β-catenin complex in H. pylori-infected cells. We also found that H. pylori induces β-catenin nuclear translocation and increases MMP7 expressions mediated through casein kinase 2. We show for the first time that CagA(+) H. pylori upregulates cellular invasiveness and motility through casein kinase 2. The demonstration of a mechanistic interplay between H. pylori and casein kinase 2 provides important insights into the role of CagA(+) H. pylori in the gastric cancer invasion and metastasis. © 2014 John Wiley & Sons Ltd.
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Thumann, G; Stöcker, M; Maltusch, C; Salz, A K; Barth, S; Walter, P; Johnen, S
2010-02-01
Transplantation of pigment epithelial cells in patients with age-related macular degeneration and Parkinson's disease has the potential to improve functional rehabilitation. Genetic modification of cells before transplantation may allow the delivery of neuroprotective factors to achieve functional improvement. As transplantation of cells modified using viral vectors is complicated by the possible dissemination of viral particles and severe immune reactions, we have explored non-viral methods to insert genetic material in pigment epithelial cells. Using lipofection or nucleofection ARPE-19 cells, freshly isolated and primary retinal and iris pigment epithelial (IPE) cells were transfected with plasmids encoding green fluorescent protein (GFP) and with three plasmids encoding recombinant pigment epithelium-derived factor (PEDF) and GFP. Transfection efficiency was evaluated by fluorescence microscopy and stability of protein expression by immunoblotting. Pigment epithelial cells were successfully transfected with plasmid encoding GFP. Expression of GFP in ARPE-19 was transient, but was observed for up to 1 year in IPE cells. Analysis of pigment epithelial cells transfected with PEDF plasmids revealed that PEDF fusion proteins were successfully expressed and functionally active. In conclusion, efficient transfer of genetic information in pigment epithelial cells can be achieved using non-viral transfection protocols.
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Cigarette Smoke Enhances the Expression of Profibrotic Molecules in Alveolar Epithelial Cells.
Directory of Open Access Journals (Sweden)
Marco Checa
Full Text Available Idiopathic pulmonary fibrosis (IPF is a progressive and lethal disease of unknown etiology. A growing body of evidence indicates that it may result from an aberrant activation of alveolar epithelium, which induces the expansion of the fibroblast population, their differentiation to myofibroblasts and the excessive accumulation of extracellular matrix. The mechanisms that activate the alveolar epithelium are unknown, but several studies indicate that smoking is the main environmental risk factor for the development of IPF. In this study we explored the effect of cigarette smoke on the gene expression profile and signaling pathways in alveolar epithelial cells. Lung epithelial cell line from human (A549, was exposed to cigarette smoke extract (CSE for 1, 3, and 5 weeks at 1, 5 and 10% and gene expression was evaluated by complete transcriptome microarrays. Signaling networks were analyzed with the Ingenuity Pathway Analysis software. At 5 weeks of exposure, alveolar epithelial cells acquired a fibroblast-like phenotype. At this time, gene expression profile revealed a significant increase of more than 1000 genes and deregulation of canonical signaling pathways such as TGF-β and Wnt. Several profibrotic genes involved in EMT were over-expressed, and incomplete EMT was observed in these cells, and corroborated in mouse (MLE-12 and rat (RLE-6TN epithelial cells. The secretion of activated TGF-β1 increased in cells exposed to cigarette smoke, which decreased when the integrin alpha v gene was silenced. These findings suggest that the exposure of alveolar epithelial cells to CSE induces the expression and release of a variety of profibrotic genes, and the activation of TGF-β1, which may explain at least partially, the increased risk of developing IPF in smokers.
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Energy Technology Data Exchange (ETDEWEB)
Zhang, Wenyao; Li, Xuezhong; Xu, Tong; Ma, Mengru [College of Veterinary Medicine, Northwest A& F University, Yangling 712100, Shaanxi (China); Zhang, Yong, E-mail: zhangyong1956@nwsuaf.edu.cn [College of Veterinary Medicine, Northwest A& F University, Yangling 712100, Shaanxi (China); Key Laboratory of Animal Biotechnology, Ministry of Agriculture, Northwest A& F University, Yangling 712100, Shaanxi (China); Gao, Ming-Qing, E-mail: gaomingqing@nwsuaf.edu.cn [College of Veterinary Medicine, Northwest A& F University, Yangling 712100, Shaanxi (China); Key Laboratory of Animal Biotechnology, Ministry of Agriculture, Northwest A& F University, Yangling 712100, Shaanxi (China)
2016-11-15
Hypernomic secretion of epithelial cytokines has several effects on stromal cells. The contributions of inflammatory epithelial cells to stromal fibroblasts in bovine mammary glands with mastitis remain poorly understood. Here, we established an inflammatory epithelial cell model of bovine mastitis with gram-negative lipopolysaccharide (LPS) and gram-positive lipoteichoic acid (LTA) bacterial cell wall components. We characterized immune responses of mammary stromal fibroblasts induced by inflammatory epithelial cells. Our results showed that inflammatory epithelial cells affected stromal fibroblast characteristics by increasing inflammatory mediator expression, elevating extracellular matrix protein deposition, decreasing proliferation capacity, and enhancing migration ability. The changes in stromal fibroblast proliferation and migration abilities were mediated by signal molecules, such as WNT signal pathway components. LPS- and LTA-induced inflammatory epithelial cells triggered different immune responses in stromal fibroblasts. Thus, in mastitis, bovine mammary gland stromal fibroblasts were affected by inflammatory epithelial cells and displayed inflammation-specific changes, suggesting that fibroblasts play crucial roles in bovine mastitis. - Highlights: • Inflammatory BMEs affect the properties of BMFs during mastitis. • BMEs inhibited the proliferation and promoted the migration of BMFs. • BMEs enhanced secretion of inflammatory mediators and deposition of ECM in BMFs. • Changes of the properties of BMFs were mediated by specific signal molecules.
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International Nuclear Information System (INIS)
Zhang, Wenyao; Li, Xuezhong; Xu, Tong; Ma, Mengru; Zhang, Yong; Gao, Ming-Qing
2016-01-01
Hypernomic secretion of epithelial cytokines has several effects on stromal cells. The contributions of inflammatory epithelial cells to stromal fibroblasts in bovine mammary glands with mastitis remain poorly understood. Here, we established an inflammatory epithelial cell model of bovine mastitis with gram-negative lipopolysaccharide (LPS) and gram-positive lipoteichoic acid (LTA) bacterial cell wall components. We characterized immune responses of mammary stromal fibroblasts induced by inflammatory epithelial cells. Our results showed that inflammatory epithelial cells affected stromal fibroblast characteristics by increasing inflammatory mediator expression, elevating extracellular matrix protein deposition, decreasing proliferation capacity, and enhancing migration ability. The changes in stromal fibroblast proliferation and migration abilities were mediated by signal molecules, such as WNT signal pathway components. LPS- and LTA-induced inflammatory epithelial cells triggered different immune responses in stromal fibroblasts. Thus, in mastitis, bovine mammary gland stromal fibroblasts were affected by inflammatory epithelial cells and displayed inflammation-specific changes, suggesting that fibroblasts play crucial roles in bovine mastitis. - Highlights: • Inflammatory BMEs affect the properties of BMFs during mastitis. • BMEs inhibited the proliferation and promoted the migration of BMFs. • BMEs enhanced secretion of inflammatory mediators and deposition of ECM in BMFs. • Changes of the properties of BMFs were mediated by specific signal molecules.
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Renal epithelial cell growth can occur in absence of Na+-H+ exchanger activity
International Nuclear Information System (INIS)
Mohrmann, M.; Cantiello, H.F.; Ausiello, D.A.
1987-01-01
An electroneutral Na+-H+ exchange system has been described in a variety of tissues and cell types, including those of renal origin, and has been proposed to play a role in the activation of growth. We have recently characterized the presence of this ubiquitous transporter in the apical domain of confluent epithelial LLC-PK1 cells. Because most apical membrane proteins appear late in cell growth, accompanying epithelial cell polarization, we determined whether the Na+-H+ exchanger is required for the growth of LLC-PK1 cells. The studies reported here show that there is no obligatory requirement for increased H+ efflux or Na+ entry via the Na+-H+ exchanger for the initiation of cell growth in this epithelial cell line. We used 22 Na+ influx, acid extrusion, and intracellular pH determinations to show that onset of cell growth, as measured by DNA content, precedes the activity of the Na+-H+ exchanger in exponentially growing cells, whereas confluent monolayers express Na+-H+ exchanger activity. When confluent cells are replated at low density, Na+-H+ exchanger activity disappears within 8 h in contrast to high-density replated cells. The fact that Na+-H+ exchanger activity is only present in confluent monolayers suggests that the development of tight junctions and polar differentiation play a role in the expression of the Na+-H+ exchanger and that this exchanger is more important to the polar epithelial cell for transepithelial transport than for the maintenance of intracellular pH
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Meconium concentration and amniotic fluid index influence the outcome of amnioinfusion.
Puertas, A; Carrillo, M P; çlvarez, M; Cañizares, J M; Miño, M; Malde, J
2001-10-01
To determine the usefulness of amnioinfusion as a function of meconium concentration and amniotic fluid index. This was a prospective study of 206 pregnant women in whom amniotic fluid was moderately or heavily stained with meconium, according to subjective evaluation. The women were assigned randomly to receive amnioinfusion (n=103) or no amnioinfusion (control group, n=103). The results were compared in women with =/15 % meconium in the amniotic fluid (measured by centrifugation), and in women in whom the amniotic fluid index calculated 60 min after insertion of the amnioinfusion catheter was 10. In women with >15% meconium, amnioinfusion decreased the rate of cesarian sections motivated by fetal distress (2.5% vs 22.2%), and in women with =/amnioinfusion decreased the presence of meconium below the vocal cords (6.4% vs 25.9%). Greater benefits after amnioinfusion were seen in women with an amniotic fluid index =/>10: the rate of cesarian sections was lower (1.3% vs 13.3%), as was the frequency of meconium below the vocal cords (10.1% vs 33.3%). Beneficial effects of amnioinfusion were seen in women with high and low concentrations of meconium, and with high and low amniotic fluid indexes. These criteria should therefore not be used to decide whether amnioinfusion is indicated when the amniotic fluid is moderately or heavily stained with meconium.
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The Long and Complicated Relationship between Epstein-Barr Virus and Epithelial Cells.
Hutt-Fletcher, Lindsey M
2017-01-01
The roles of epithelial cells in infection and persistence of the Epstein-Barr virus (EBV) have long been difficult to resolve. However, recent developments have reinforced the conclusion that these cells are a major site of virus replication and raised the possibility that, like papillomaviruses, EBV has evolved to take advantage of epithelial differentiation to ensure survival, persistence, and spread. Copyright © 2016 American Society for Microbiology.
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Energy Technology Data Exchange (ETDEWEB)
Zhou, Xiangjun [Department of Urology, Taihe Hospital, Hubei University of Medicine, Hubei (China); Yao, Qisheng, E-mail: yymcyqs@126.com [Department of Urology, Taihe Hospital, Hubei University of Medicine, Hubei (China); Sun, Xinbo; Gong, Xiaoxin; Yang, Yong; Chen, Congbo [Department of Urology, Taihe Hospital, Hubei University of Medicine, Hubei (China); Shan, Guang [Department of Urology, Renmin Hospital of Wuhan University, Hubei (China)
2017-03-01
Hypoxic acute kidney injury (AKI) is often incompletely repaired and leads to chronic kidney disease (CKD), which is characterized by tubulointerstitial inflammation and fibrosis. The Slit2 family of secreted glycoproteins is expressed in the kidney, it has been shown to exert an anti-inflammatory activity and prevent ischemic renal injury in vivo. However, whether Slit2 reduces renal fibrosis and inflammation after hypoxic and inflammatory epithelial cells injury in vitro remains unknown. In this study, we aimed to evaluate whether Slit2 ameliorated fibrosis and inflammation in two renal epithelial cells line challenged with hypoxia and lipopolysaccharide (LPS). Renal epithelial cells were treated with hypoxia and LPS to induce cell injury. Hoechst staining and Western blot analysis was conducted to examine epithelial cells injury. Immunofluorescence staining and Western blot analysis was performed to evaluate tubulointerstitial fibrosis. Real-time polymerase chain reaction (PCR) tested the inflammatory factor interleukin (IL)−1β and tumor necrosis factor (TNF)-α, and Western blot analysis determined the hypoxia-inducible factor (HIF)−1α, Toll-like receptor 4 (TLR4) and nuclear factor (NF)-κB. Results revealed that hypoxia induced epithelial cells apoptosis, inflammatory factor IL-1β and TNF-α release and tubulointerstitial fibrosis. LPS could exacerbate hypoxia -induced epithelial cells apoptosis, IL-1β and TNF-α release and fibrosis. Slit2 reduced the expression of fibronectin, the rate of epithelial cell apoptosis, and the expression of inflammatory factor. Slit2 could also inhibit the expression of TLR4 and NF-κB, but not the expression of HIF-1α. Therefore, Slit2 attenuated inflammation and fibrosis after LPS- and hypoxia-induced epithelial cells injury via the TLR4/NF-κB signaling pathway, but not depending on the HIF-1α signaling pathway. - Highlights: • Slit2 ameliorates inflammation after hypoxia-and LPS-induced epithelial cells injury
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Energy Technology Data Exchange (ETDEWEB)
Xin, Ying [Stem Cell Institute, James Brown Cancer Center, University of Louisville School of Medicine, 301 E. Muhammad Ali Blvd., Louisville, KY 40202 (United States); Department of Ophthalmology and Visual Sciences, University of Louisville School of Medicine, 301 E. Muhammad Ali Blvd., Louisville, KY 40202 (United States); Lu, Qingxian [Tumor Immunobiology Group, James Brown Cancer Center, University of Louisville School of Medicine, 301 E. Muhammad Ali Blvd., Louisville, KY 40202 (United States); Department of Ophthalmology and Visual Sciences, University of Louisville School of Medicine, 301 E. Muhammad Ali Blvd., Louisville, KY 40202 (United States); Department of Biochemistry and Molecular Biology, University of Louisville School of Medicine, 301 E. Muhammad Ali Blvd., Louisville, KY 40202 (United States); Li, Qiutang, E-mail: q.li@louisville.edu [Stem Cell Institute, James Brown Cancer Center, University of Louisville School of Medicine, 301 E. Muhammad Ali Blvd., Louisville, KY 40202 (United States); Department of Ophthalmology and Visual Sciences, University of Louisville School of Medicine, 301 E. Muhammad Ali Blvd., Louisville, KY 40202 (United States)
2010-02-19
14-3-3{sigma} (also called stratifin) is specifically expressed in the stratified squamous epithelium and its function was recently shown to be linked to epidermal stratification and differentiation in the skin. In this study, we investigated its role in corneal epithelium cell proliferation and differentiation. We showed that the 14-3-3{sigma} mutation in repeated epilation (Er) mutant mice results in a dominant negative truncated protein. Primary corneal epithelial cells expressing the dominant negative protein failed to undergo high calcium-induced cell cycle arrest and differentiation. We further demonstrated that blocking endogenous 14-3-3{sigma} activity in corneal epithelial cells by overexpressing dominative negative 14-3-3{sigma} led to reduced Notch activity and Notch1/2 transcription. Significantly, expression of the active Notch intracellular domain overcame the block in epithelial cell differentiation in 14-3-3{sigma} mutant-expressing corneal epithelial cells. We conclude that 14-3-3{sigma} is critical for regulating corneal epithelial proliferation and differentiation by regulating Notch signaling activity.
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International Nuclear Information System (INIS)
Xin, Ying; Lu, Qingxian; Li, Qiutang
2010-01-01
14-3-3σ (also called stratifin) is specifically expressed in the stratified squamous epithelium and its function was recently shown to be linked to epidermal stratification and differentiation in the skin. In this study, we investigated its role in corneal epithelium cell proliferation and differentiation. We showed that the 14-3-3σ mutation in repeated epilation (Er) mutant mice results in a dominant negative truncated protein. Primary corneal epithelial cells expressing the dominant negative protein failed to undergo high calcium-induced cell cycle arrest and differentiation. We further demonstrated that blocking endogenous 14-3-3σ activity in corneal epithelial cells by overexpressing dominative negative 14-3-3σ led to reduced Notch activity and Notch1/2 transcription. Significantly, expression of the active Notch intracellular domain overcame the block in epithelial cell differentiation in 14-3-3σ mutant-expressing corneal epithelial cells. We conclude that 14-3-3σ is critical for regulating corneal epithelial proliferation and differentiation by regulating Notch signaling activity.
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Regulation of epithelial and lymphocyte cell adhesion by adenosine deaminase-CD26 interaction.
Ginés, Silvia; Mariño, Marta; Mallol, Josefa; Canela, Enric I; Morimoto, Chikao; Callebaut, Christian; Hovanessian, Ara; Casadó, Vicent; Lluis, Carmen; Franco, Rafael
2002-01-01
The extra-enzymic function of cell-surface adenosine deaminase (ADA), an enzyme mainly localized in the cytosol but also found on the cell surface of monocytes, B cells and T cells, has lately been the subject of numerous studies. Cell-surface ADA is able to transduce co-stimulatory signals in T cells via its interaction with CD26, an integral membrane protein that acts as ADA-binding protein. The aim of the present study was to explore whether ADA-CD26 interaction plays a role in the adhesion of lymphocyte cells to human epithelial cells. To meet this aim, different lymphocyte cell lines (Jurkat and CEM T) expressing endogenous, or overexpressing human, CD26 protein were tested in adhesion assays to monolayers of colon adenocarcinoma human epithelial cells, Caco-2, which express high levels of cell-surface ADA. Interestingly, the adhesion of Jurkat and CEM T cells to a monolayer of Caco-2 cells was greatly dependent on CD26. An increase by 50% in the cell-to-cell adhesion was found in cells containing higher levels of CD26. Incubation with an anti-CD26 antibody raised against the ADA-binding site or with exogenous ADA resulted in a significant reduction (50-70%) of T-cell adhesion to monolayers of epithelial cells. The role of ADA-CD26 interaction in the lymphocyte-epithelial cell adhesion appears to be mediated by CD26 molecules that are not interacting with endogenous ADA (ADA-free CD26), since SKW6.4 (B cells) that express more cell-surface ADA showed lower adhesion than T cells. Adhesion stimulated by CD26 and ADA is mediated by T cell lymphocyte function-associated antigen. A role for ADA-CD26 interaction in cell-to-cell adhesion was confirmed further in integrin activation assays. FACS analysis revealed a higher expression of activated integrins on T cell lines in the presence of increasing amounts of exogenous ADA. Taken together, these results suggest that the ADA-CD26 interaction on the cell surface has a role in lymphocyte-epithelial cell adhesion. PMID
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Directory of Open Access Journals (Sweden)
Jeanette Wagener
Full Text Available C. albicans is one of the most common fungal pathogen of humans, causing local and superficial mucosal infections in immunocompromised individuals. Given that the key structure mediating host-C. albicans interactions is the fungal cell wall, we aimed to identify features of the cell wall inducing epithelial responses and be associated with fungal pathogenesis. We demonstrate here the importance of cell wall protein glycosylation in epithelial immune activation with a predominant role for the highly branched N-glycosylation residues. Moreover, these glycan moieties induce growth arrest and apoptosis of epithelial cells. Using an in vitro model of oral candidosis we demonstrate, that apoptosis induction by C. albicans wild-type occurs in early stage of infection and strongly depends on intact cell wall protein glycosylation. These novel findings demonstrate that glycosylation of the C. albicans cell wall proteins appears essential for modulation of epithelial immunity and apoptosis induction, both of which may promote fungal pathogenesis in vivo.
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Effect of amniotic fluid on the in vitro culture of human corneal endothelial cells.
Feizi, Sepehr; Soheili, Zahra-Soheila; Bagheri, Abouzar; Balagholi, Sahar; Mohammadian, Azam; Rezaei-Kanavi, Mozhgan; Ahmadieh, Hamid; Samiei, Shahram; Negahban, Kambiz
2014-05-01
The present study was designed to evaluate the effects of human amniotic fluid (HAF) on the growth of human corneal endothelial cells (HCECs) and to establish an in vitro method for expanding HCECs. HCECs were cultured in DMEM-F12 supplemented with 20% fetal bovine serum (FBS). Confluent monolayer cultures were trypsinized and passaged using either FBS- or HAF-containing media. Cell proliferation and cell death ELISA assays were performed to determine the effect of HAF on cell growth and viability. The identity of the cells cultured in 20% HAF was determined using immunocytochemistry (ICC) and real-time reverse transcription polymerase chain reaction (RT-PCR) techniques to evaluate the expression of factors that are characteristic of HCECs, including Ki-67, Vimentin, Na+/K+-ATPase and ZO-1. HCEC primary cultures were successfully established using 20% HAF-containing medium, and these cultures demonstrated rapid cell proliferation according to the cell proliferation and death ELISA assay results. The ICC and real time RT-PCR results indicated that there was a higher expression of Na+/K+-ATPase and ZO-1 in the 20% HAF cell cultures compared with the control (20% FBS) (P < 0.05). The 20% HAF-containing medium exhibited a greater stimulatory effect on HCEC growth and could represent a potential enriched supplement for HCEC regeneration studies. Copyright © 2014 Elsevier Ltd. All rights reserved.
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Directory of Open Access Journals (Sweden)
Bhavani S. Kowtharapu
2018-05-01
Full Text Available In the cornea, healing of the wounded avascular surface is an intricate process comprising the involvement of epithelial, stromal and neuronal cell interactions. These interactions result to the release of various growth factors that play prominent roles during corneal wound healing response. Bone morphogenetic proteins (BMPs are unique multi-functional potent growth factors of the transforming growth factor-beta (TGF-β superfamily. Treatment of corneal epithelial cells with substance P and nerve growth factor resulted to an increase in the expression of BMP7 mRNA. Since BMP7 is known to modulate the process of corneal wound healing, in this present study, we investigated the influence of exogenous rhBMP7 on human corneal epithelial cell and stromal cell (SFs function. To obtain a high-fidelity expression profiling of activated biomarkers and pathways, transcriptome-wide gene-level expression profiling of epithelial cells in the presence of BMP7 was performed. Gene ontology analysis shows BMP7 stimulation activated TGF-β signaling and cell cycle pathways, whereas biological processes related to cell cycle, microtubule and intermediate filament cytoskeleton organization were significantly impacted in corneal epithelial cells. Scratch wound healing assay showed increased motility and migration of BMP7 treated epithelial cells. BMP7 stimulation studies show activation of MAPK cascade proteins in epithelial cells and SFs. Similarly, a difference in the expression of claudin, Zink finger E-box-binding homeobox 1 was observed along with phosphorylation levels of cofilin in epithelial cells. Stimulation of SFs with BMP7 activated them with increased expression of α-smooth muscle actin. In addition, an elevated phosphorylation of epidermal growth factor receptor following BMP7 stimulation was also observed both in corneal epithelial cells and SFs. Based on our transcriptome analysis data on epithelial cells and the results obtained in SFs, we
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INFLUENCE OF AMNIOTIC FLUID INDEX ON FOETAL OUTCOME
Directory of Open Access Journals (Sweden)
Raja Lakshmi
2015-03-01
Full Text Available BACKGROUND AND OBJECTIVE: In these days of smaller families and the obstetrician having to share the onus of giving a healthy child capable of independent existence as well as to ensure that the population is limited for further progress of this developing country, the estimation o f foetal maturity assumes greatest practical importance. As means to achieving the end, estimates of foetal maturity have been done by various clinical and laboratory methods of which assessment of amniotic fluid index assumes importance. The objective is to study the correlation of amniotic fluid index on foetal outcome at term gestation . MATERIALS AND METHODS: The study was carried out on two hundred antenatal women who attended the institute of obstetrics and gynaecology at a Government Hospital for Wome n and Children in Visakhapatnam from Jan 2014 to Jan 2015. It is a comparative prospective study comparing 100 cases of Oligohydramnios (amniotic fluid index 5 cm as control group. RESU LTS : Perinatal outcome was inferred by noting the various parameters and Statistical Analysis was done by applying the chisquare (x2 test and the value of probability was taken . CONCLUSION: The goal of antepartum fetal surveillance is to identify the fetu s at increased risk. Amniotic fluid volume has been proved as an indirect measure of feto - placental function and hence the estimation of amniotic fluid volume assists the obstetrician in risk assessment
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Hydrogen sulfide lowers proliferation and induces protective autophagy in colon epithelial cells.
Directory of Open Access Journals (Sweden)
Ya C Wu
Full Text Available Hydrogen sulfide (H(2S is a gaseous bacterial metabolite that reaches high levels in the large intestine. In the present study, the effect of H(2S on the proliferation of normal and cancerous colon epithelial cells was investigated. An immortalized colon epithelial cell line (YAMC and a panel of colon cancer cell lines (HT-29, SW1116, HCT116 were exposed to H(2S at concentrations similar to those found in the human colon. H(2S inhibited normal and cancerous colon epithelial cell proliferation as measured by MTT assay. The anti-mitogenic effect of H(2S was accompanied by G(1-phase cell cycle arrest and the induction of the cyclin-dependent kinase inhibitor p21(Cip. Moreover, exposure to H(2S led to features characteristic of autophagy, including increased formation of LC3B(+ autophagic vacuoles and acidic vesicular organelles as determined by immunofluorescence and acridine orange staining, respectively. Abolition of autophagy by RNA interference targeting Vps34 or Atg7 enhanced the anti-proliferative effect of H(2S. Further mechanistic investigation revealed that H(2S stimulated the phosphorylation of AMP-activated protein kinase (AMPK and inhibited the phosphorylation of mammalian target of rapamycin (mTOR and S6 kinase. Inhibition of AMPK significantly reversed H(2S-induced autophagy and inhibition of cell proliferation. Collectively, we demonstrate that H(2S inhibits colon epithelial cell proliferation and induces protective autophagy via the AMPK pathway.
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Synergic production of neutrophil chemotactic activity by colonic epithelial cells and eosinophils.
Dent, Gordon; Loweth, Sam C; Hasan, Anwar Matar; Leslie, Fiona M
2014-10-01
The presence of eosinophils in the lumen and mucosa of the intestine is characteristic of both ulcerative colitis (UC) and Crohn's disease (CD). There is evidence of eosinophil activation in the intestine during acute inflammatory episodes of these diseases; these episodes are also characterized by an influx of neutrophils, which have the potential to cause extensive tissue damage. We undertook a study to determine whether eosinophils in contact with colonic epithelial cells produce factors that may attract neutrophils in response to immunological stimulation. Neutrophil chemotactic activity (NCA) and concentrations of three neutrophil-attracting CXC chemokines - CXCL1 (Groα), CXCL5 (Ena78) and CXCL8 (IL8) - were measured in supernatants of T84 colonic epithelial cells and blood eosinophils or eosinophil-like myeloid leukaemia cells (AML14.3D10), alone or in combination. Cells were stimulated with serum-opsonized zymosan (OZ) particles. NCA (Peosinophil co-cultures were significantly higher than in the supernatants of either cell type alone. Release of CXCL1 (Peosinophils but not higher than from OZ-stimulated epithelial cells. Eosinophils and colonic epithelial cells exhibit synergy in production of neutrophil chemoattractants in response to immunological stimulation. This may represent a mechanism for exaggerated recruitment of neutrophils to the intestine in response to acute infection in conditions that are characterized by the presence of eosinophils in the bowel. Copyright © 2014 Elsevier GmbH. All rights reserved.
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Jin, Liting; Qu, Ying; Gomez, Liliana J; Chung, Stacey; Han, Bingchen; Gao, Bowen; Yue, Yong; Gong, Yiping; Liu, Xuefeng; Amersi, Farin; Dang, Catherine; Giuliano, Armando E; Cui, Xiaojiang
2018-02-20
Conditional reprogramming methods allow for the inexhaustible in vitro proliferation of primary epithelial cells from human tissue specimens. This methodology has the potential to enhance the utility of primary cell culture as a model for mammary gland research. However, few studies have systematically characterized this method in generating in vitro normal human mammary epithelial cell models. We show that cells derived from fresh normal breast tissues can be propagated and exhibit heterogeneous morphologic features. The cultures are composed of CK18, desmoglein 3, and CK19-positive luminal cells and vimentin, p63, and CK14-positive myoepithelial cells, suggesting the maintenance of in vivo heterogeneity. In addition, the cultures contain subpopulations with different CD49f and EpCAM expression profiles. When grown in 3D conditions, cells self-organize into distinct structures that express either luminal or basal cell markers. Among these structures, CK8-positive cells enclosing a lumen are capable of differentiation into milk-producing cells in the presence of lactogenic stimulus. Furthermore, our short-term cultures retain the expression of ERα, as well as its ability to respond to estrogen stimulation. We have investigated conditionally reprogrammed normal epithelial cells in terms of cell type heterogeneity, cellular marker expression, and structural arrangement in two-dimensional (2D) and three-dimensional (3D) systems. The conditional reprogramming methodology allows generation of a heterogeneous culture from normal human mammary tissue in vitro . We believe that this cell culture model will provide a valuable tool to study mammary cell function and malignant transformation.
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Ruscetti, Marcus; Quach, Bill; Dadashian, Eman L.; Mulholland, David J.; Wu, Hong
2015-01-01
The epithelial-mesenchymal transition (EMT) has been postulated as a mechanism by which cancer cells acquire the invasive and stem-like traits necessary for distant metastasis. However, direct in vivo evidence for the role of EMT in the formation of cancer stem-like cells (CSC) and the metastatic cascade remains lacking. Here we report the first isolation and characterization of mesenchymal and EMT tumor cells, which harbor both epithelial and mesenchymal characteristics, in an autochthonous murine model of prostate cancer. By crossing the established Pb-Cre+/−;PtenL/L;KrasG12D/+ prostate cancer model with a vimentin-GFP reporter strain, generating CPKV mice, we were able to isolate epithelial, EMT and mesenchymal cancer cells based on expression of vimentin and EpCAM. CPKV mice (but not mice with Pten deletion alone) exhibited expansion of cells with EMT (EpCAM+/Vim-GFP+) and mesenchymal (EpCAM−/Vim-GFP+) characteristics at the primary tumor site and in circulation. These EMT and mesenchymal tumor cells displayed enhanced stemness and invasive character compared to epithelial tumor cells. Moreover, they displayed an enriched tumor-initiating capacity and could regenerate epithelial glandular structures in vivo, indicative of epithelia-mesenchyme plasticity. Interestingly, while mesenchymal tumor cells could persist in circulation and survive in the lung following intravenous injection, only epithelial and EMT tumor cells could form macrometastases. Our work extends the evidence that mesenchymal and epithelial states in cancer cells contribute differentially to their capacities for tumor initiation and metastatic seeding, respectively, and that EMT tumor cells exist with plasticity that can contribute to multiple stages of the metastatic cascade. PMID:25948589
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Oral Rehabilitation for Amniotic Band Syndrome: An Unusual Presentation.
Hotwani, Kavita; Sharma, Krishna
2015-01-01
Amniotic band syndrome (ABS) is a congenital disorder caused by entrapment of fetal parts in fibrous amniotic bands while in utero. The syndrome is underdiagnosed and its presentation is variable. The syndrome has been well described in the pediatric, orthopedic and obstetric literature; however, despite the discernable craniomaxillofacial involvement, ABS has not been reported in the dental literature very often. The present report describes a case of a patient with ABS and concomitant dental findings. How to cite this article: Hotwani K, Sharma K. Oral Rehabilitation for Amniotic Band Syndrome: An Unusual Presentation. Int J Clin Pediatr Dent 2015;8(1):55-57.
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Fetal- and uterine-specific antigens in human amniotic fluid.
Sutcliffe, R G; Brock, D J; Nicholson, L V; Dunn, E
1978-09-01
Removal of the major maternal serum proteins from second trimester amniotic fluid by antibody affinity chromatography revealed various soluble tissue antigens, of which two were fetal-specific skin proteins and another, of alpha2-mobility, was specific to the uterus, and was therefore designated alpha-uterine protein (AUP). These proteins could not be detected in maternal serum by antibody-antigen crossed electrophoresis. The concentration of AUP in amniotic fluid reached a maximum between 10 and 20 weeks of gestation, suggesting that there is an influx of uterine protein into the amniotic fluid at this stage of pregnancy.
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LENUS (Irish Health Repository)
Corcionivoschi, N
2012-02-01
The host cell environment can alter bacterial pathogenicity. We employed a combination of cellular and molecular techniques to study the expression of Campylobacter jejuni polysaccharides cocultured with HCT-8 epithelial cells. After two passages, the amount of membrane-bound high-molecular-weight polysaccharide was considerably reduced. Microarray profiling confirmed significant downregulation of capsular polysaccharide (CPS) locus genes. Experiments using conditioned media showed that sugar depletion occurred only when the bacterial and epithelial cells were cocultured. CPS depletion occurred when C. jejuni organisms were exposed to conditioned media from a different C. jejuni strain but not when exposed to conditioned media from other bacterial species. Proteinase K or heat treatment of conditioned media under coculture conditions abrogated the effect on the sugars, as did formaldehyde fixation and cycloheximide treatment of host cells or chloramphenicol treatment of the bacteria. However, sugar depletion was not affected in flagellar export (fliQ) and quorum-sensing (luxS) gene mutants. Passaged C. jejuni showed reduced invasiveness and increased serum sensitivity in vitro. C. jejuni alters its surface polysaccharides when cocultured with epithelial cells, suggesting the existence of a cross talk mechanism that modulates CPS expression during infection.
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Epigenetic influences of low-dose bisphenol A in primary human breast epithelial cells
International Nuclear Information System (INIS)
Weng, Yu-I; Hsu, Pei-Yin; Liyanarachchi, Sandya; Liu, Joseph; Deatherage, Daniel E.; Huang Yiwen; Zuo Tao; Rodriguez, Benjamin; Lin, Ching-Hung; Cheng, Ann-Lii; Huang, Tim H.-M.
2010-01-01
Substantial evidence indicates that exposure to bisphenol A (BPA) during early development may increase breast cancer risk later in life. The changes may persist into puberty and adulthood, suggesting an epigenetic process being imposed in differentiated breast epithelial cells. The molecular mechanisms by which early memory of BPA exposure is imprinted in breast progenitor cells and then passed onto their epithelial progeny are not well understood. The aim of this study was to examine epigenetic changes in breast epithelial cells treated with low-dose BPA. We also investigated the effect of BPA on the ERα signaling pathway and global gene expression profiles. Compared to control cells, nuclear internalization of ERα was observed in epithelial cells preexposed to BPA. We identified 170 genes with similar expression changes in response to BPA. Functional analysis confirms that gene suppression was mediated in part through an ERα-dependent pathway. As a result of exposure to BPA or other estrogen-like chemicals, the expression of lysosomal-associated membrane protein 3 (LAMP3) became epigenetically silenced in breast epithelial cells. Furthermore, increased DNA methylation in the LAMP3 CpG island was this repressive mark preferentially occurred in ERα-positive breast tumors. These results suggest that the in vitro system developed in our laboratory is a valuable tool for exposure studies of BPA and other xenoestrogens in human cells. Individual and geographical differences may contribute to altered patterns of gene expression and DNA methylation in susceptible loci. Combination of our exposure model with epigenetic analysis and other biochemical assays can give insight into the heritable effect of low-dose BPA in human cells.
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International Nuclear Information System (INIS)
Katano, Takahito; Ootani, Akifumi; Mizoshita, Tsutomu; Tanida, Satoshi; Tsukamoto, Hironobu; Ozeki, Keiji; Ebi, Masahide; Mori, Yoshinori; Kataoka, Hiromi; Kamiya, Takeshi; Toda, Shuji; Joh, Takashi
2013-01-01
Highlights: ► We established a 3D culture system to allow long-term culture of stomach cells. ► In this culture system, gastric epithelial cells grew for about 3 months. ► The cultured cells differentiated into multi-units of the stomach. ► This culture method should be useful for elucidating the cause of gastric diseases. -- Abstract: Compared to the small intestine and colon, little is known about stem cells in the stomach because of a lack of specific stem cell markers and an in vitro system that allows long-term culture. Here we describe a long-term three-dimensional (3D) primary gastric culture system within the stem cell niche. Glandular stomach cells from neonatal mice cultured in collagen gel yielded expanding sphere-like structures for 3 months. The wall of the gastrospheres consisted of a highly polarized epithelial monolayer with an outer lining of myofibroblasts. The epithelial cells showed a tall columnar cell shape, basal round nuclei, and mucus-filled cytoplasm as well as expression of MUC5AC, indicating differentiation into gastric surface mucous cells. These cells demonstrated the features of fully differentiated gastric surface mucous cells such as microvilli, junctional complexes, and glycogen and secretory granules. Fewer than 1% of cultured epithelial cells differentiated into enteroendocrine cells. Active proliferation of the epithelial cells and many apoptotic cells in the inner lumen revealed the rapid cell turnover in gastrospheres in vitro. This method enables us to investigate the role of signaling between cell–cell and epithelial–mesenchymal interactions in an environment that is extremely similar to the in vivo environment
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Energy Technology Data Exchange (ETDEWEB)
Katano, Takahito [Department of Gastroenterology and Metabolism, Nagoya City University Graduate School of Medical Sciences, 1 Kawasumi, Mizuho-cho, Mizuho-ku, Nagoya 467-8601 (Japan); Ootani, Akifumi [Department of Gastroenterology and GI Endoscopy Center, Shin-Kokura Hospital, Federation of National Public Service Personnel Mutual Aid Associations, 1-3-1 Kanada, Kokurakita-ku, Kitakyushu 803-0816 (Japan); Department of Internal Medicine, Faculty of Medicine, Saga University, 5-1-1 Nabeshima, Saga 849-8501 (Japan); Mizoshita, Tsutomu, E-mail: tmizoshi@med.nagoya-cu.ac.jp [Department of Gastroenterology and Metabolism, Nagoya City University Graduate School of Medical Sciences, 1 Kawasumi, Mizuho-cho, Mizuho-ku, Nagoya 467-8601 (Japan); Tanida, Satoshi; Tsukamoto, Hironobu; Ozeki, Keiji; Ebi, Masahide; Mori, Yoshinori; Kataoka, Hiromi; Kamiya, Takeshi [Department of Gastroenterology and Metabolism, Nagoya City University Graduate School of Medical Sciences, 1 Kawasumi, Mizuho-cho, Mizuho-ku, Nagoya 467-8601 (Japan); Toda, Shuji [Department of Pathology and Microbiology, Faculty of Medicine, Saga University, 5-1-1 Nabeshima, Saga 849-8501 (Japan); Joh, Takashi [Department of Gastroenterology and Metabolism, Nagoya City University Graduate School of Medical Sciences, 1 Kawasumi, Mizuho-cho, Mizuho-ku, Nagoya 467-8601 (Japan)
2013-03-22
Highlights: ► We established a 3D culture system to allow long-term culture of stomach cells. ► In this culture system, gastric epithelial cells grew for about 3 months. ► The cultured cells differentiated into multi-units of the stomach. ► This culture method should be useful for elucidating the cause of gastric diseases. -- Abstract: Compared to the small intestine and colon, little is known about stem cells in the stomach because of a lack of specific stem cell markers and an in vitro system that allows long-term culture. Here we describe a long-term three-dimensional (3D) primary gastric culture system within the stem cell niche. Glandular stomach cells from neonatal mice cultured in collagen gel yielded expanding sphere-like structures for 3 months. The wall of the gastrospheres consisted of a highly polarized epithelial monolayer with an outer lining of myofibroblasts. The epithelial cells showed a tall columnar cell shape, basal round nuclei, and mucus-filled cytoplasm as well as expression of MUC5AC, indicating differentiation into gastric surface mucous cells. These cells demonstrated the features of fully differentiated gastric surface mucous cells such as microvilli, junctional complexes, and glycogen and secretory granules. Fewer than 1% of cultured epithelial cells differentiated into enteroendocrine cells. Active proliferation of the epithelial cells and many apoptotic cells in the inner lumen revealed the rapid cell turnover in gastrospheres in vitro. This method enables us to investigate the role of signaling between cell–cell and epithelial–mesenchymal interactions in an environment that is extremely similar to the in vivo environment.
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Directory of Open Access Journals (Sweden)
Edgar Corneille Ontsouka
Full Text Available BACKGROUND: Mammary cell cultures are convenient tools for in vitro studies of mammary gland biology. However, the heterogeneity of mammary cell types, e.g., glandular milk secretory epithelial or myoepithelial cells, often complicates the interpretation of cell-based data. The present study was undertaken to determine the relevance of bovine primary mammary epithelial cells isolated from American Holstein (bMEC US or Swiss Holstein-Friesian (bMEC CH cows, and of primary bovine mammary alveolar epithelial cells stably transfected with simian virus-40 (SV-40 large T-antigen (MAC-T for in vitro analyses. This was evaluated by testing their expression pattern of cytokeratin (CK 7, 18, 19, vimentin, and α-smooth muscle actin (α-SMA. RESULTS: The expression of the listed markers was assessed using real-time quantitative PCR, flow cytometry and immunofluorescence microscopy. Characteristic markers of the mesenchymal (vimentin, myoepithelial (α-SMA and glandular secretory cells (CKs showed differential expression among the studied cell cultures, partly depending on the analytical method used. The relative mRNA expression of vimentin, CK7 and CK19, respectively, was lower (P < 0.05 in immortalized than in primary mammary cell cultures. The stain index (based on flow cytometry of CK7 and CK19 protein was lower (P < 0.05 in MAC-T than in bMECs, while the expression of α-SMA and CK18 showed an inverse pattern. Immunofluorescence microscopy analysis mostly confirmed the mRNA data, while partly disagreed with flow cytometry data (e.g., vimentin level in MAC-T. The differential expression of CK7 and CK19 allowed discriminating between immortal and primary mammary cultures. CONCLUSIONS: The expression of the selected widely used cell type markers in primary and immortalized MEC cells did not allow a clear preference between these two cell models for in vitro analyses studying aspects of milk composition. All tested cell models exhibited to a variable
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Mechanisms of Cell Polarity-Controlled Epithelial Homeostasis and Immunity in the Intestine
Klunder, Leon J.; Faber, Klaas Nico; Dijkstra, Gerard; van IJzendoorn, Sven C. D.
Intestinal epithelial cell polarity is instrumental to maintain epithelial homeostasis and balance communications between the gut lumen and bodily tissue, thereby controlling the defense against gastrointestinal pathogens and maintenance of immune tolerance to commensal bacteria. In this review, we
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Histamine Induces Bovine Rumen Epithelial Cell Inflammatory Response via NF-κB Pathway
Directory of Open Access Journals (Sweden)
Xudong Sun
2017-06-01
Full Text Available Background/Aims: Subacute ruminal acidosis (SARA is a common disease in high-producing lactating cows. Rumenitis is the initial insult of SARA and is associated with the high concentrations of histamine produced in the rumen of dairy cows during SARA. However, the exact mechanism remains unclear. The objective of the current study is to investigate whether histamine induces inflammation of rumen epithelial cells and the underlying mechanism of this process. Methods: Bovine rumen epithelial cells were cultured and treated with different concentrations of histamine and pyrrolidine dithiocarbamate (PDTC, an NF-κB inhibitor cultured in different pH medium (pH 7.2 or 5.5. qRT-PCR, Western-blotting, ELISA and immunocytofluorescence were used to evaluate whether histamine activated the NF-κB pathway and inflammatory cytokines. Results: The results showed that histamine significantly increased the activity of IKK β and the phosphorylation levels of IκB α, as well as upregulated the mRNA and protein expression levels of NF-κB p65 in the rumen epithelial cells cultured in neutral (pH=7.2 and acidic (pH=5.5 medium. Furthermore, histamine treatment also significantly increased the transcriptional activity of NF-κB p65. High expression and transcriptional activity of NF-κB p65 significantly increased the mRNA expressions and concentrations of inflammatory cytokines, tumor necrosis factor alpha (TNF-α, interleukin 6 (IL-6 and interleukin 1 beta (IL-1β, thereby inducing the inflammatory response in bovine rumen epithelial cells. However, inhibition of NF-κB p65 by PDTC significantly decreased the expressions and concentrations of the inflammatory cytokines induced by histamine in the rumen epithelial cells cultured in the neutral and acidic medium. Conclusion: The present data indicate that histamine induces the inflammatory response of bovine rumen epithelial cells through the NF-κB pathway.
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Hardy, Katharine M.; Garriock, Robert J.; Yatskievych, Tatiana A.; D'Agostino, Susan L.; Antin, Parker B.; Krieg, Paul A.
2008-01-01
Knowledge of the molecular mechanisms regulating cell ingression, epithelial-mesenchymal transition and migration movements during amniote gastrulation is steadily improving. In the frog and fish embryo, Wnt5 and Wnt11 ligands are expressed around the blastopore and play an important role in regulating cell movements associated with gastrulation. In the chicken embryo, although Wnt5a and Wnt5b are expressed in the primitive streak, the known Wnt11 gene is expressed in paraxial and intermediat...
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Directory of Open Access Journals (Sweden)
Yeo Jin Jeon
Full Text Available Fabry disease is a lysosomal storage disorder caused by deficiency of alpha-galactosidase A (α-gal A, which results in the deposition of globotriaosylceramide (Gb3 in the vascular endothelium. Globotriaosylsphingosine (lyso-Gb3, a deacylated Gb3, is also increased in the plasma of patients with Fabry disease. Renal fibrosis is a key feature of advanced Fabry disease patients. Therefore, we evaluated the association of Gb3 and lyso-Gb3 accumulation and the epithelial-mesenchymal transition (EMT on tubular epithelial cells of the kidney. In HK2 cells, exogenous treatments of Gb3 and lyso-Gb3 increased the expression of TGF-β, EMT markers (N-cadherin and α-SMA, and phosphorylation of PI3K/AKT, and decreased the expression of E-cadherin. Lyso-Gb3, rather than Gb3, strongly induced EMT in HK2 cells. In the mouse renal mesangial cell line, SV40 MES 13 cells, Gb3 strongly induced phenotype changes. The EMT induced by Gb3 was inhibited by enzyme α-gal A treatment, but EMT induced by lyso-Gb3 was not abrogated by enzyme treatment. However, TGF-β receptor inhibitor (TRI, SB525334 inhibited the activation of TGF-β and EMT markers in HK2 cells with Gb3 and lyso-Gb3 treatments. This study suggested that increased plasma lyso-Gb3 has a crucial role in the development of renal fibrosis through the cell-specific induction of the EMT in Fabry disease, and that TRI treatment, alongside enzyme replacement therapy, could be a potential therapeutic option for patients with Fabry disease.
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Wiggins, Rebecca; Horner, Patrick J; Whittington, Kate; Holmes, Christopher H
2009-01-01
Abstract Background Epithelial cells in first catch urine (FCU) specimens from 87 men with and without urethritis were quantified. Epithelial cells were broadly categorised into transitional and squamous populations using morphological characteristics and immunostaining with anti-pan leukocyte and anti-cytokeratin monoclonal antibodies. Findings The majority (77/87 = 89%) of samples contained both transitional (76/87 = 87%; range 1 × 104 – 6 × 105, median 6 × 104) and squamous (57/87 = 66%; r...
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Generation of folliculogenic human epithelial stem cells from induced pluripotent stem cells
Yang, Ruifeng; Zheng, Ying; Burrows, Michelle; Liu, Shujing; Wei, Zhi; Nace, Arben; Guo, Wei; Kumar, Suresh; Cotsarelis, George; Xu, Xiaowei
2014-01-01
Epithelial stem cells (EpSCs) in the hair follicle bulge are required for hair follicle growth and cycling. The isolation and propagation of human EpSCs for tissue engineering purposes remains a challenge. Here we develop a strategy to differentiate human iPSCs (hiPSCs) into CD200+/ITGA6+ EpSCs that can reconstitute the epithelial components of the hair follicle and interfollicular epidermis. The hiPSC-derived CD200+/ITGA6+ cells show a similar gene expression signature as EpSCs directly isolated from human hair follicles. Human iPSC-derived CD200+/ITGA6+ cells are capable of generating all hair follicle lineages including the hair shaft, and the inner and outer root sheaths in skin reconstitution assays. The regenerated hair follicles possess a KRT15+ stem cell population and produce hair shafts expressing hair-specific keratins. These results suggest an approach for generating large numbers of human EpSCs for tissue engineering and new treatments for hair loss, wound healing and other degenerative skin disorders.
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Directory of Open Access Journals (Sweden)
Elham Hashemi
2017-01-01
Full Text Available As the key producer of cerebrospinal fluid (CSF, the choroid plexus (CP provides a unique protective system in the central nervous system. CSF components are not invariable and they can change based on the pathological conditions of the central nervous system. The purpose of the present study was to assess the effects of non-traumatic and traumatic CSF on the differentiation of multipotent stem-like cells of CP into the neural and/or glial cells. CP epithelial cells were isolated from adult male rats and treated with human non-traumatic and traumatic CSF. Alterations in mRNA expression of Nestin and microtubule-associated protein (MAP2, as the specific markers of neurogenesis, and astrocyte marker glial fibrillary acidic protein (GFAP in cultured CP epithelial cells were evaluated using quantitative real-time PCR. The data revealed that treatment with CSF (non-traumatic and traumatic led to increase in mRNA expression levels of MAP2 and GFAP. Moreover, the expression of Nestin decreased in CP epithelial cells treated with non-traumatic CSF, while treatment with traumatic CSF significantly increased its mRNA level compared to the cells cultured only in DMEM/F12 as control. It seems that CP epithelial cells contain multipotent stem-like cells which are inducible under pathological conditions including exposure to traumatic CSF because of its compositions.
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Radiation studies on sensitivity and repair of human mammary epithelial cells
International Nuclear Information System (INIS)
Tracy Chuihsu Yang; Stampfer, M.R.; Tobias, C.A.
1989-01-01
The authors present results indicating that normal breast epithelial cells and fibroblasts respond to X-rays similarly, lacking significant repair of sublethal damage when 2 Gy was used as the conditioning dose. Epithelial cells from tumor and from parenchymal tissue peripheral to the tumor, however, did show an efficient repair of sublethal damage. The reasons for this difference is unknown. Heavy-ion studies suggest energetic carbon and neon particles can be more effective in killing normal and tumour cells. The RBE for normal cells, however, appeared to be slightly less than for tumor cells. The repair of sublethal damage in tumor cells was less for neon particles than for X-rays. These findings suggest that heavy ions might be more advantageous than X-rays in treating breast tumors. (author)
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Aziz, Faisal; Yang, Xuesong; Wen, Qingping; Yan, Qiu
2015-08-01
At present, biopsy specimens, cancer cell lines and tissues obtained by gastric surgery are used in the study and analysis of gastric cancer, including the molecular mechanisms and proteomics. However, fibroblasts and other tissue components may interfere with these techniques. Therefore, the present study aimed to develop a procedure for the isolation of viable human gastric epithelial cells from gastric surgical tissues. A method was developed to culture human gastric epithelial cells using fresh, surgically excised tissues and was evaluated using immunocytochemistry, periodic acid-Schiff (PAS) staining and cell viability assays. Low cell growth was observed surrounding the gastric tissue on the seventh day of tissue explant culture. Cell growth subsequently increased, and at 12 days post-explant a high number of pure epithelial cells were detected. The gastric cancer cells exhibited rapid growth with a doubling time of 13-52 h, as compared to normal cells, which had a doubling time of 20-53 h. Immunocytochemical analyses of primary gastric cells revealed positive staining for cytokeratin 18 and 19, which indicated that the culture was comprised of pure epithelial cells and contained no fibroblasts. Furthermore, PAS staining demonstrated that the cultured gastric cells produced neutral mucin. Granulin and carbohydrate antigen 724 staining confirmed the purity of gastric cancer and normal cells in culture. This method of cell culture indicated that the gastric cells in primary culture consisted of mucin-secreting gastric epithelial cells, which may be useful for the study of gastric infection with Helicobacter pylori and gastric cancer.
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Androgen-Sensitized Apoptosis of HPr-1AR Human Prostate Epithelial Cells.
Directory of Open Access Journals (Sweden)
Congcong Chen
Full Text Available Androgen receptor (AR signaling is crucial to the development and homeostasis of the prostate gland, and its dysregulation mediates common prostate pathologies. The mechanisms whereby AR regulates growth suppression and differentiation of luminal epithelial cells in the prostate gland and proliferation of malignant versions of these cells have been investigated in human and rodent adult prostate. However, the cellular stress response of human prostate epithelial cells is not well understood, though it is central to prostate health and pathology. Here, we report that androgen sensitizes HPr-1AR and RWPE-AR human prostate epithelial cells to cell stress agents and apoptotic cell death. Although 5α-dihydrotestosterone (DHT treatment alone did not induce cell death, co-treatment of HPr-1AR cells with DHT and an apoptosis inducer, such as staurosporine (STS, TNFt, or hydrogen peroxide, synergistically increased cell death in comparison to treatment with each apoptosis inducer by itself. We found that the synergy between DHT and apoptosis inducer led to activation of the intrinsic/mitochondrial apoptotic pathway, which is supported by robust cleavage activation of caspase-9 and caspase-3. Further, the dramatic depolarization of the mitochondrial membrane potential that we observed upon co-treatment with DHT and STS is consistent with increased mitochondrial outer membrane permeabilization (MOMP in the pro-apoptotic mechanism. Interestingly, the synergy between DHT and apoptosis inducer was abolished by AR antagonists and inhibitors of transcription and protein synthesis, suggesting that AR mediates pro-apoptotic synergy through transcriptional regulation of MOMP genes. Expression analysis revealed that pro-apoptotic genes (BCL2L11/BIM and AIFM2 were DHT-induced, whereas pro-survival genes (BCL2L1/BCL-XL and MCL1 were DHT-repressed. Hence, we propose that the net effect of these AR-mediated expression changes shifts the balance of BCL2-family proteins
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Yolk sac development in lizards (Lacertilia: Scincidae): New perspectives on the egg of amniotes.
Stewart, James R; Thompson, Michael B
2017-04-01
Embryos of oviparous reptiles develop on the surface of a large mass of yolk, which they metabolize to become relatively large hatchlings. Access to the yolk is provided by tissues growing outward from the embryo to cover the surface of the yolk. A key feature of yolk sac development is a dedicated blood vascular system to communicate with the embryo. The best known model for yolk sac development and function of oviparous amniotes is based on numerous studies of birds, primarily domestic chickens. In this model, the vascular yolk sac forms the perimeter of the large yolk mass and is lined by a specialized epithelium, which takes up, processes and transports yolk nutrients to the yolk sac blood vessels. Studies of lizard yolk sac development, dating to more than 100 years ago, report characteristics inconsistent with this model. We compared development of the yolk sac from oviposition to near hatching in embryonic series of three species of oviparous scincid lizards to consider congruence with the pattern described for birds. Our findings reinforce results of prior studies indicating that squamate reptiles mobilize and metabolize the large yolk reserves in their eggs through a process unknown in other amniotes. Development of the yolk sac of lizards differs from birds in four primary characteristics, migration of mesoderm, proliferation of endoderm, vascular development and cellular diversity within the yolk sac cavity. Notably, all of the yolk is incorporated into cells relatively early in development and endodermal cells within the yolk sac cavity align along blood vessels which course throughout the yolk sac cavity. The pattern of uptake of yolk by endodermal cells indicates that the mechanism of yolk metabolism differs between lizards and birds and that the evolution of a fundamental characteristic of embryonic nutrition diverged in these two lineages. Attributes of the yolk sac of squamates reveal the existence of phylogenetic diversity among amniote lineages
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DEFF Research Database (Denmark)
Nielsen, Helga Lind; Gudjonsson, Thorarinn; Villadsen, René
2004-01-01
Mesenchymal-like cells in the stroma of breast cancer may arise as a consequence of plasticity within the epithelial compartment, also referred to as epithelial-mesenchymal transition, or by recruitment of genuine mesenchymal cells from the peritumoral stroma. Cells of both the epithelial...... compartment and the stromal compartment express alpha smooth muscle actin (alpha-sm actin) as part of a myoepithelial or a myofibroblastic differentiation program, respectively. Moreover, because both epithelial- and mesenchymal-derived cells are nontumorigenic, other means of discrimination are warranted....... Here, we describe the contraction of hydrated collagen gels as a rapid functional assay for the distinction between epithelial- and mesenchymal-derived stromal-like cells irrespective of the status of alpha-sm actin expression. Three epithelial-derived cell lines and three genuine mesenchymal...
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DEFF Research Database (Denmark)
Dabelsteen, Erik; Grøn, B.; Mandel, U.
1998-01-01
Alterations in cell to cell adhesion are necessary to enable the type of cell movements that are associated with epithelial wound healing and malignant invasion. Several studies of transformed cells have related epithelial cell movement to changes in the cell surface expression of the carbohydrate......-T antigen. The changes induced by wounding in the expression of collagen IV, laminin gamma2-chain (laminin-5), and laminin alpha5-chain were similar to those found in skin wounds and served to define the region of epithelial movement. This region was found to show a marked increase in staining for both...... epithelium, a pattern of expression similar to K16, which was also strongly upregulated in both the outgrowth and the adjacent nonwounded epithelium. These findings provide further support for an influence of such carbohydrate structures on the migratory behavior of epithelial cells....
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Ethane dimethanesulfonate (EDS) perturbs epididymal epithelial cell function in vitro
International Nuclear Information System (INIS)
Klinefelter, G.
1990-01-01
The formation of sperm granulomas in the epididymis following exposure to EDS, a Leydig cell toxicant, was reported by Cooper and Jackson in 1970. Recent work suggests that EDS may effect the epididymis directly. An in vitro system was developed to determine the nature of any direct effect. The caput epididymis from adult rats was dissected free of connective tissue and small pieces of the tissue were enzymatically digested until plaques of epididymal epithelial cells were obtained. Plaques were cultured on an extracellular matrix gelled on top of a semipermeable filter creating dual-compartment environments. The epithelial cells maintained typical morphology and protein secretion in this culture system for several days. Beginning on day 3, EDS (1 mM) was added to the basal compartment, with or without 35 S-methionine. After 24 hours, 35 S-labelled culture medium was taken from the apical compartment and analyzed by SDS-PAGE and fluorography. EDS caused decreased secretion of several proteins, including a 39 Kd molecule. Interestingly, a 39 Kd protein was also shown to disappear from sperm taken from the caput epididymidis following in vivo exposure to EDS. Unlabelled cultures were fixed and processed for light microscopy. No alterations in morphological integrity were observed. Thus, epididymal epithelial cell function is directly altered by EDS exposure
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Ewald, Andrew J.; Huebner, Robert J.; Palsdottir, Hildur; Lee, Jessie K.; Perez, Melissa J.; Jorgens, Danielle M.; Tauscher, Andrew N.; Cheung, Kevin J.; Werb, Zena; Auer, Manfred
2012-01-01
Normal mammary morphogenesis involves transitions between simple and multilayered epithelial organizations. We used electron microscopy and molecular markers to determine whether intercellular junctions and apico-basal polarity were maintained in the multilayered epithelium. We found that multilayered elongating ducts had polarized apical and basal tissue surfaces both in three-dimensional culture and in vivo. However, individual cells were only polarized on surfaces in contact with the lumen or extracellular matrix. The basolateral marker scribble and the apical marker atypical protein kinase C zeta localized to all interior cell membranes, whereas PAR3 displayed a cytoplasmic localization, suggesting that the apico-basal polarity was incomplete. Despite membrane localization of E-cadherin and β-catenin, we did not observe a defined zonula adherens connecting interior cells. Instead, interior cells were connected through desmosomes and exhibited complex interdigitating membrane protrusions. Single-cell labeling revealed that individual cells were both protrusive and migratory within the epithelial multilayer. Inhibition of Rho kinase (ROCK) further reduced intercellular adhesion on apical and lateral surfaces but did not disrupt basal tissue organization. Following morphogenesis, segregated membrane domains were re-established and junctional complexes re-formed. We observed similar epithelial organization during mammary morphogenesis in organotypic culture and in vivo. We conclude that mammary epithelial morphogenesis involves a reversible, spatially limited, reduction in polarity and intercellular junctions and active individualistic cell migration. Our data suggest that reductions in polarity and adhesion during breast cancer progression might reflect partial recapitulation of a normal developmental program. PMID:22344263
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21 CFR 862.1455 - Lecithin/sphingomyelin ratio in amniotic fluid test system.
2010-04-01
... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Lecithin/sphingomyelin ratio in amniotic fluid... Clinical Chemistry Test Systems § 862.1455 Lecithin/sphingomyelin ratio in amniotic fluid test system. (a) Identification. A lecithin/sphingomyelin ratio in amniotic fluid test system is a device intended to measure the...
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Oxidative stress biomarkers in amniotic fluid of pregnant women with hypothyroidism.
Novakovic, Tanja R; Dolicanin, Zana C; Djordjevic, Natasa Z
2017-11-15
Hypothyroidism in pregnancy is the serious state that may lead to fetal morbidity and mortality. Oxidative stress biomarkers in the amniotic fluid can provide important information on the health, development and maturation of the fetus during pregnancy. In this study, we examined whether maternal hypothyroidism contributes to increased oxidative stress biomarkers in the amniotic fluid during the first trimester of pregnancy. The study was conducted on healthy pregnant women and pregnant women with hypothyroidism (gestational age: 16-18 weeks). Oxidative stress biomarkers, such as superoxide anion (O 2 •- ), hydrogen peroxide (H 2 O 2 ), nitric oxide (NO), peroxynitrite (ONOO - ), lipid peroxide (LPO), reduced glutathione (GSH) and oxidized glutathione (GSSG) were assayed in the amniotic fluid. The results of this study indicated that concentrations of O 2 •- and NO are significantly higher, while the concentration of H 2 O 2 is significantly lower in the amniotic fluid of pregnant women with hypothyroidism in comparison to healthy pregnant women. There were no differences in concentrations of LPO, GSH and GSSG among tested groups. Also, we found that amniotic fluid concentration of O 2 •- is negatively correlated with the body weight and Apgar score values of the newborns. These results suggest that pregnancy hypothyroidism is characterized by the amniotic fluid oxidative stress. Incorporation of the oxidative stress biomarkers measurement in the amniotic fluid may be of clinical importance in the management of pregnancy hypothyroidism.
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Cysteine cathepsins B and X promote epithelial-mesenchymal transition of tumor cells.
Mitrović, Ana; Pečar Fonović, Urša; Kos, Janko
2017-09-01
Cathepsins B and X are lysosomal cysteine carboxypeptidases suggested as having a redundant role in cancer. They are involved in a number of processes leading to tumor progression but their role in the epithelial-mesenchymal transition (EMT) remains unknown. We have investigated the contribution of both cathepsins B and X in EMT using tumor cell lines differing in their expression of epithelial and mesenchymal markers and cell morphology. Higher levels of both cathepsins are shown to promote EMT and are associated with the mesenchymal-like cell phenotype. Moreover, simultaneous knockdown of the two peptidases triggers a reverse, mesenchymal to epithelial transition. Of the two cathepsins, cathepsin B appears to be the stronger promotor of EMT. Furthermore, we evaluated the involvement of cathepsin B and X in the transforming growth factor-β1 (TGF-β1) signaling pathway, one of the key signaling mechanisms triggering EMT in cancer. In MCF-7 cells the expression of cathepsin B was shown to depend on their activation with TGF-β1 while, for cathepsin X, a TGF-β1 independent mechanism of induction during EMT is indicated. EMT is thus shown to be another mechanism linking cathepsins B and X with tumor progression. With silencing of their expression or inhibition of enzymatic activity, the tumor cells could be reverted to less aggressive epithelial-like phenotype. Copyright © 2017 Elsevier GmbH. All rights reserved.
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The world of epithelial sheets.
Honda, Hisao
2017-06-01
An epithelium is a layer of closely connected cells covering the body or lining a body cavity. In this review, several fundamental questions are addressed regarding the epithelium. (i) While an epithelium functions as barrier against the external environment, how is barrier function maintained during its construction? (ii) What determines the apical and basal sides of epithelial layer? (iii) Is there any relationship between the apical side of the epithelium and the apical membrane of an epithelial cell? (iv) Why are hepatocytes (liver cells) called epithelial, even though they differ completely from column-like shape of typical epithelial cells? Keeping these questions in mind, multiple shapes of epithelia were considered, extracting a few of their elemental processes, and constructing a virtual world of epithelia by combining them. Epithelial cells were also classified into several types based on the number of apical domains of each cell. In addition, an intracellular organelle was introduced within epithelial cells, the vacuolar apical compartment (VAC), which is produced within epithelial cells surrounded by external cell matrix (ECM). The VAC interacts with areas of cell-cell contact of the cell surface membrane and is converted to apical membrane. The properties of VACs enable us to answer the initial questions posed above. Finally, the genetic and molecular mechanisms of epithelial morphogenesis are discussed. © 2017 Japanese Society of Developmental Biologists.
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Trichostatin A inhibits beta-casein expression in mammary epithelial cells
International Nuclear Information System (INIS)
Pujuguet, Philippe; Radisky, Derek; Levy, Dinah; Lacza, Charlemagne; Bissell, Mina J.
2002-01-01
Many aspects of cellular behavior are affected by information derived from association of the extracellular matrix (ECM) and with cell membrane receptors. When cultured in the presence of laminin-containing ECM and prolactin (Prl), normal mammary epithelial cells express the milk protein beta-casein. Previously, we defined the minimal ECM- and Prl-responsive enhancer element BCE-1 from the upstream region of the beta-casein gene. We also found that BCE-1 was only active when stably integrated into chromatin, and that trichostatin A (TSA), a reagent that leads to alterations in chromatin structure, was able to activate the integrated enhancer element. We now show that endogenous b-casein gene, which is controlled by a genetic assembly that is highly similar to that of BCE-1 and which is also activated by incubation in ECM and Prl, is instead inhibited by TSA. We provide evidence that the differing response of b-casein and BCE-1 to TSA is neither due to an unusual effect of TSA on mammary epithelial cells, nor to secondary consequences from the expression of a separate gene, nor to a particular property of the BCE-1 construct. As a component of this investigation, we also showed that ECM could mediate rapid histone deacetylation in mammary epithelial cells. These results are discussed in combination with previous work showing that TSA mediates the differentiation of many types of cancer cells but inhibits differentiation of some nonmalignant cell types
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McMurtry, Vanity; Saavedra, Joseph E; Nieves-Alicea, René; Simeone, Ann-Marie; Keefer, Larry K; Tari, Ana M
2011-04-01
Targeted therapy with reduced side effects is a major goal in cancer research. We investigated the effects of JS-K, a nitric oxide (NO) prodrug designed to release high levels of NO when suitably activated, on human breast cancer cell lines, on non-transformed human MCF-10A mammary cells, and on normal human mammary epithelial cells (HMECs). Cell viability assay, flow cytometry, electron microscopy, and Western blot analysis were used to study the effects of JS-K on breast cancer and on mammary epithelial cells. After a 3-day incubation, the IC50s of JS-K against the breast cancer cells ranged from 0.8 to 3 µM. However, JS-K decreased the viability of the MCF-10A cells by only 20% at 10-µM concentration, and HMECs were unaffected by 10 µM JS-K. Flow cytometry indicated that JS-K increased the percentages of breast cancer cells under-going apoptosis. Interestingly, flow cytometry indicated that JS-K increased acidic vesicle organelle formation in breast cancer cells, suggesting that JS-K induced autophagy in breast cancer cells. Electron microscopy confirmed that JS-K-treated breast cancer cells underwent autophagic cell death. Western blot analysis showed that JS-K induced the expression of microtubule light chain 3-II, another autophagy marker, in breast cancer cells. However, JS-K did not induce apoptosis or autophagy in normal human mammary epithelial cells. These data indicate that JS-K selectively induces programmed cell death in breast cancer cells while sparing normal mammary epithelial cells under the same conditions. The selective anti-tumor activity of JS-K warrants its further investigation in breast tumors.
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Protective Effects of Trehalose on the Corneal Epithelial Cells
Directory of Open Access Journals (Sweden)
Pasquale Aragona
2014-01-01
Full Text Available Purpose. Aim of the present work was to evaluate the effects of the trehalose on the corneal epithelium undergoing alcohol delamination. Methods. Twelve patients undergoing laser subepithelial keratomileusis (LASEK were consecutively included in the study. The right eyes were pretreated with 3% trehalose eye drops, whilst left eyes were used as control. Epithelial specimens were processed for cells vitality assessment, apoptosis, and light and transmission electron microscopy; a morphometric analysis was performed in both groups. Results. In both trehalose-untreated eyes (TUE and trehalose-treated eyes (TTE, the percentage of vital cells was similar and no apoptotic cells were observed. In TUE, the corneal epithelium showed superficial cells with reduced microfolds, wing cells with vesicles and dilated intercellular spaces, and dark basal cells with vesicles and wide clefts. In TTE, superficial and wing cells were better preserved, and basal cells were generally clear with intracytoplasmatic vesicles. The morphometric analysis showed statistically significant differences between the two groups: the TTE epithelial height was higher, the basal cells showed larger area and clearer cytoplasm. The distribution of desmosomes and hemidesmosomes was significantly different between the groups. Conclusions. Trehalose administration better preserved morphological and morphometric features of alcohol-treated corneal epithelium, when compared to controls.
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Kv7.1 surface expression is regulated by epithelial cell polarization
DEFF Research Database (Denmark)
Andersen, Martin N; Olesen, Søren-Peter; Rasmussen, Hanne Borger
2011-01-01
The potassium channel K(V)7.1 is expressed in the heart where it contributes to the repolarization of the cardiac action potential. In addition, K(V)7.1 is expressed in epithelial tissues where it plays a role in salt and water transport. Mutations in the kcnq1 gene can lead to long QT syndrome...... and deafness, and several mutations have been described as trafficking mutations. To learn more about the basic mechanisms that regulate K(V)7.1 surface expression, we have investigated the trafficking of K(V)7.1 during the polarization process of the epithelial cell line Madin-Darby Canine Kidney (MDCK) using...... is regulated by signaling mechanisms involved in epithelial cell polarization in particular signaling cascades involving protein kinase C and PI3K....
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Massie, Isobel; Levis, Hannah J; Daniels, Julie T
2014-10-01
Limbal epithelial stem cell deficiency can cause blindness but may be treated by human limbal epithelial cell (hLE) transplantation, normally on human amniotic membrane. Clinical outcomes using amnion can be unreliable and so we have developed an alternative tissue equivalent (TE), RAFT (Real Architecture for 3D Tissue), which supports hLE expansion, and stratification when airlifted. Human limbal fibroblasts (hLF) may be incorporated into RAFT TEs, where they support overlying hLE and improve phenotype. However, the impact of neither airlifting nor hLF on hLE function has been investigated. hLE on RAFT TEs (±hLF and airlifting) were wounded using heptanol and re-epithelialisation (fluorescein diacetate staining), and percentage putative stem cell marker p63α and proliferative marker Ki67 expression (wholemount immunohistochemistry), measured. Airlifted, hLF- RAFT TEs were unable to close the wound and p63α expression was 7 ± 0.2% after wounding. Conversely, non-airlifted, hLF- RAFT TEs closed the wound within 9 days and p63α expression was higher at 22 ± 5% (p < 0.01). hLE on both hLF- and hLF+ RAFT TEs (non-airlifted) closed the wound and p63α expression was 26 ± 8% and 36 ± 3% respectively (ns). Ki67 expression by hLE increased from 1.3 ± 0.5% before wounding to 7.89 ± 2.53% post-wounding for hLF- RAFT TEs (p < 0.01), and 0.8 ± 0.08% to 17.68 ± 10.88% for hLF+ RAFT TEs (p < 0.05), suggesting that re-epithelialisation was a result of proliferation. These data suggest that neither airlifting nor hLF are necessarily required to maintain a functional epithelium on RAFT TEs, thus simplifying and shortening the production process. This is important when working towards clinical application of regenerative medicine products. Copyright © 2014 Elsevier Ltd. All rights reserved.
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Dong, Ji; Hu, Yuqiong; Fan, Xiaoying; Wu, Xinglong; Mao, Yunuo; Hu, Boqiang; Guo, Hongshan; Wen, Lu; Tang, Fuchou
2018-03-14
Organogenesis is crucial for proper organ formation during mammalian embryonic development. However, the similarities and shared features between different organs and the cellular heterogeneity during this process at single-cell resolution remain elusive. We perform single-cell RNA sequencing analysis of 1916 individual cells from eight organs and tissues of E9.5 to E11.5 mouse embryos, namely, the forebrain, hindbrain, skin, heart, somite, lung, liver, and intestine. Based on the regulatory activities rather than the expression patterns, all cells analyzed can be well classified into four major groups with epithelial, mesodermal, hematopoietic, and neuronal identities. For different organs within the same group, the similarities and differences of their features and developmental paths are revealed and reconstructed. We identify mutual interactions between epithelial and mesenchymal cells and detect epithelial cells with prevalent mesenchymal features during organogenesis, which are similar to the features of intermediate epithelial/mesenchymal cells during tumorigenesis. The comprehensive transcriptome at single-cell resolution profiled in our study paves the way for future mechanistic studies of the gene-regulatory networks governing mammalian organogenesis.
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File list: ALL.Utr.20.AllAg.Fallopian_tube_secretory_epithelial_cell [Chip-atlas[Archive
Lifescience Database Archive (English)
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File list: ALL.Utr.50.AllAg.Fallopian_tube_secretory_epithelial_cell [Chip-atlas[Archive
Lifescience Database Archive (English)
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File list: ALL.Utr.05.AllAg.Fallopian_tube_secretory_epithelial_cell [Chip-atlas[Archive
Lifescience Database Archive (English)
Full Text Available ALL.Utr.05.AllAg.Fallopian_tube_secretory_epithelial_cell hg19 All antigens Uterus Fallopian tube secret...hg19/assembled/ALL.Utr.05.AllAg.Fallopian_tube_secretory_epithelial_cell.bed ...
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Two-layer tissue engineered urethra using oral epithelial and muscle derived cells.
Mikami, Hiroshi; Kuwahara, Go; Nakamura, Nobuyuki; Yamato, Masayuki; Tanaka, Masatoshi; Kodama, Shohta
2012-05-01
We fabricated novel tissue engineered urethral grafts using autologously harvested oral cells. We report their viability in a canine model. Oral tissues were harvested by punch biopsy and divided into mucosal and muscle sections. Epithelial cells from mucosal sections were cultured as epithelial cell sheets. Simultaneously muscle derived cells were seeded on collagen mesh matrices to form muscle cell sheets. At 2 weeks the sheets were joined and tubularized to form 2-layer tissue engineered urethras, which were autologously grafted to surgically induced urethral defects in 10 dogs in the experimental group. Tissue engineered grafts were not applied to the induced urethral defect in control dogs. The dogs were followed 12 weeks postoperatively. Urethrogram and histological examination were done to evaluate the grafting outcome. We successfully fabricated 2-layer tissue engineered urethras in vitro and transplanted them in dogs in the experimental group. The 12-week complication-free rate was significantly higher in the experimental group than in controls. Urethrogram confirmed urethral patency without stricture in the complication-free group at 12 weeks. Histologically urethras in the transplant group showed a stratified epithelial layer overlying well differentiated submucosa. In contrast, urethras in controls showed severe fibrosis without epithelial layer formation. Two-layer tissue engineered urethras were engineered using cells harvested by minimally invasive oral punch biopsy. Results suggest that this technique can encourage regeneration of a functional urethra. Copyright © 2012 American Urological Association Education and Research, Inc. Published by Elsevier Inc. All rights reserved.
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Directory of Open Access Journals (Sweden)
Sashaina E Fanibunda
Full Text Available BACKGROUND: During sexual transmission of HIV in women, the virus breaches the multi-layered CD4 negative stratified squamous epithelial barrier of the vagina, to infect the sub-epithelial CD4 positive immune cells. However the mechanisms by which HIV gains entry into the sub-epithelial zone is hitherto unknown. We have previously reported human mannose receptor (hMR as a CD4 independent receptor playing a role in HIV transmission on human spermatozoa. The current study was undertaken to investigate the expression of hMR in vaginal epithelial cells, its HIV gp120 binding potential, affinity constants and the induction of matrix metalloproteinases (MMPs downstream of HIV gp120 binding to hMR. PRINCIPAL FINDINGS: Human vaginal epithelial cells and the immortalized vaginal epithelial cell line Vk2/E6E7 were used in this study. hMR mRNA and protein were expressed in vaginal epithelial cells and cell line, with a molecular weight of 155 kDa. HIV gp120 bound to vaginal proteins with high affinity, (Kd = 1.2±0.2 nM for vaginal cells, 1.4±0.2 nM for cell line and the hMR antagonist mannan dose dependently inhibited this binding. Both HIV gp120 binding and hMR exhibited identical patterns of localization in the epithelial cells by immunofluorescence. HIV gp120 bound to immunopurified hMR and affinity constants were 2.9±0.4 nM and 3.2±0.6 nM for vaginal cells and Vk2/E6E7 cell line respectively. HIV gp120 induced an increase in MMP-9 mRNA expression and activity by zymography, which could be inhibited by an anti-hMR antibody. CONCLUSION: hMR expressed by vaginal epithelial cells has high affinity for HIV gp120 and this binding induces production of MMPs. We propose that the induction of MMPs in response to HIV gp120 may lead to degradation of tight junction proteins and the extracellular matrix proteins in the vaginal epithelium and basement membrane, leading to weakening of the epithelial barrier; thereby facilitating transport of HIV across the
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Danelishvili, Lia; McGarvey, Jeffery; Li, Yong-Jun; Bermudez, Luiz E
2003-09-01
Mycobacterium tuberculosis interacts with macrophages and epithelial cells in the alveolar space of the lung, where it is able to invade and replicate in both cell types. M. tuberculosis-associated cytotoxicity to these cells has been well documented, but the mechanisms of host cell death are not well understood. We examined the induction of apoptosis and necrosis of human macrophages (U937) and type II alveolar epithelial cells (A549) by virulent (H37Rv) and attenuated (H37Ra) M. tuberculosis strains. Apoptosis was determined by both enzyme-linked immunosorbent assay (ELISA) and TdT-mediated dUTP nick end labelling (TUNEL) assay, whereas necrosis was evaluated by the release of lactate dehydrogenase (LDH). Both virulent and attenuated M. tuberculosis induced apoptosis in macrophages; however, the attenuated strain resulted in significantly more apoptosis than the virulent strain after 5 days of infection. In contrast, cytotoxicity of alveolar cells was the result of necrosis, but not apoptosis. Although infection with M. tuberculosis strains resulted in apoptosis of 14% of the cells on the monolayer, cell death associated with necrosis was observed in 59% of alveolar epithelial cells after 5 days of infection. Infection with M. tuberculosis suppressed apoptosis of alveolar epithelial cells induced by the kinase inhibitor, staurosporine. Because our findings suggest that M. tuberculosis can modulate the apoptotic response of macrophages and epithelial cells, we carried out an apoptosis pathway-specific cDNA microarray analysis of human macrophages and alveolar epithelial cells. Whereas the inhibitors of apoptosis, bcl-2 and Rb, were upregulated over 2.5-fold in infected (48 h) alveolar epithelial cells, the proapoptotic genes, bad and bax, were downregulated. The opposite was observed when U937 macrophages were infected with M. tuberculosis. Upon infection of alveolar epithelial cells with M. tuberculosis, the generation of apoptosis, as determined by the
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Xiong, Jimin; Gronthos, Stan; Bartold, P Mark
2013-10-01
Periodontitis is a highly prevalent inflammatory disease that results in damage to the tooth-supporting tissues, potentially leading to tooth loss. Periodontal tissue regeneration is a complex process that involves the collaboration of two hard tissues (cementum and alveolar bone) and two soft tissues (gingiva and periodontal ligament). To date, no periodontal-regenerative procedures provide predictable clinical outcomes. To understand the rational basis of regenerative procedures, a better understanding of the events associated with the formation of periodontal components will help to establish reliable strategies for clinical practice. An important aspect of this is the role of the Hertwig's epithelial root sheath in periodontal development and that of its descendants, the epithelial cell rests of Malassez, in the maintenance of the periodontium. An important structure during tooth root development, the Hertwig's epithelial root sheath is not only a barrier between the dental follicle and dental papilla cells but is also involved in determining the shape, size and number of roots and in the development of dentin and cementum, and may act as a source of mesenchymal progenitor cells for cementoblasts. In adulthood, the epithelial cell rests of Malassez are the only odontogenic epithelial population in the periodontal ligament. Although there is no general agreement on the functions of the epithelial cell rests of Malassez, accumulating evidence suggests that the putative roles of the epithelial cell rests of Malassez in adult periodontal ligament include maintaining periodontal ligament homeostasis to prevent ankylosis and maintain periodontal ligament space, to prevent root resorption, to serve as a target during periodontal ligament innervation and to contribute to cementum repair. Recently, ovine epithelial cell rests of Malassez cells have been shown to harbor clonogenic epithelial stem-cell populations that demonstrate similar properties to mesenchymal stromal
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Directory of Open Access Journals (Sweden)
Jeroen P Vanderhoeven
2014-03-01
Full Text Available Early events leading to intrauterine infection remain poorly defined, but may hold the key to preventing preterm delivery. To determine molecular pathways within fetal membranes (chorioamnion associated with early choriodecidual infection that may progress to preterm premature rupture of membranes (PPROM, we examined the effects of a Group B Streptococcus (GBS choriodecidual infection on chorioamnion in a nonhuman primate model. Ten chronically catheterized pregnant monkeys (Macaca nemestrina at 118-125 days gestation (term = 172 days received choriodecidual inoculation of either GBS (n = 5 or saline (n = 5. Cesarean section was performed in the first week after GBS or saline inoculation. RNA extracted from chorioamnion (inoculation site was profiled by microarray. Single gene, Gene Set, and Ingenuity Pathway Analysis results were validated using qRT-PCR (chorioamnion, Luminex (amniotic fluid, AF, immunohistochemistry, and transmission electron microscopy (TEM. Despite uterine quiescence in most cases, significant elevations of AF cytokines (TNF-α, IL-8, IL-1β, IL-6 were detected in GBS versus controls (p2-fold change, p<0.05. Remarkably, GBS exposure was associated with significantly downregulated expression of multiple cytokeratin (CK and other cytoskeletal genes critical for maintenance of tissue tensile strength. Immunofluorescence revealed highly significant changes in the CK network within amniocytes with dense CK aggregates and retraction from the cell periphery (all p = 0.006. In human pregnancies affected by PPROM, there was further evidence of CK network retraction with significantly shorter amniocyte foot processes (p = 0.002. These results suggest early choriodecidual infection results in decreased cellular membrane integrity and tensile strength via dysfunction of CK networks. Downregulation of CK expression and perturbations in the amniotic epithelial cell intermediate filament network occur after GBS
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Berrocal, Liliana; Fuentes, Juan A; Trombert, A Nicole; Jofré, Matías R; Villagra, Nicolás A; Valenzuela, Luis M; Mora, Guido C
2015-07-07
Salmonella enterica serovar Typhi (S. Typhi) stg operon, encoding a chaperone/usher fimbria (CU), contributes to an increased adherence to human epithelial cells. However, one report suggests that the presence of the Stg fimbria impairs the monocyte--bacteria association, as deduced by the lower level of invasion to macrophage-like cells observed when the stg fimbrial cluster was overexpressed. Nevertheless, since other CU fimbrial structures increase the entry of S. Typhi into macrophages, and considering that transcriptomic analyses revealed that stg operon is indeed expressed in macrophages, we reassessed the role of the stg operon in the interaction between S. Typhi strain STH2370 and human cells, including macrophage-like cells and mononuclear cells directly taken from human peripheral blood. We compared S. Typhi STH2370 WT, a Chilean clinical strain, and the S. Typhi STH2370 Δstg mutant with respect to association and invasion using epithelial and macrophage-like cells. We observed that deletion of stg operon reduced the association and invasion of S. Typhi, in both cellular types. The presence of the cloned stg operon restored the WT phenotype in all the cases. Moreover, we compared Salmonella enterica sv. Typhimurium 14028s (S. Typhimurium, a serovar lacking stg operon) and S. Typhimurium heterologously expressing S. Typhi stg. We found that the latter presents an increased cell disruption of polarized epithelial cells and an increased association in both epithelial and macrophage-like cells. S. Typhi stg operon encodes a functional adhesin that participates in the interaction bacteria-eukaryotic cells, including epithelial cells and macrophages-like cells. The phenotypes associated to stg operon include increased association and consequent invasion in bacteria-eukaryotic cells, and cell disruption.
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DEFF Research Database (Denmark)
Smith, Ida Mosbech; Baker, A; Arneborg, Nils
2015-01-01
distinct patterns of non-Saccharomyces yeast modulation of epithelial cell barrier function. While the established probiotic yeast Saccharomyces boulardii increased TER across a Caco-2 monolayer by 30%, Kluyveromyces marxianus exhibited significantly stronger properties of TER enhancement (50% TER increase....... In addition, probiotic strains may be able to reduce epithelial barrier disruption caused by pathogenic species. The aim of this study was to explore non-Saccharomyces yeast modulation of epithelial cell barrier function in vitro. Benchmarking against established probiotic strains, we evaluated the ability......). In addition, our data demonstrate significant yeast-mediated modulation of Salmonella-induced epithelial cell barrier disruption and identify K. marxianus and Metschnikowia gruessii as two non-Saccharomyces yeasts capable of protecting human epithelial cells from pathogen invasion. SIGNIFICANCE AND IMPACT...
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Lefevre, James G; Chiu, Han S; Combes, Alexander N; Vanslambrouck, Jessica M; Ju, Ali; Hamilton, Nicholas A; Little, Melissa H
2017-03-15
Human pluripotent stem cells, after directed differentiation in vitro , can spontaneously generate complex tissues via self-organisation of the component cells. Self-organisation can also reform embryonic organ structure after tissue disruption. It has previously been demonstrated that dissociated embryonic kidneys can recreate component epithelial and mesenchymal relationships sufficient to allow continued kidney morphogenesis. Here, we investigate the timing and underlying mechanisms driving self-organisation after dissociation of the embryonic kidney using time-lapse imaging, high-resolution confocal analyses and mathematical modelling. Organotypic self-organisation sufficient for nephron initiation was observed within a 24 h period. This involved cell movement, with structure emerging after the clustering of ureteric epithelial cells, a process consistent with models of random cell movement with preferential cell adhesion. Ureteric epithelialisation rapidly followed the formation of ureteric cell clusters with the reformation of nephron-forming niches representing a later event. Disruption of P-cadherin interactions was seen to impair this ureteric epithelial cell clustering without affecting epithelial maturation. This understanding could facilitate improved regulation of patterning within organoids and facilitate kidney engineering approaches guided by cell-cell self-organisation. © 2017. Published by The Company of Biologists Ltd.
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Directory of Open Access Journals (Sweden)
Kai-Hung Wang
2016-01-01
Full Text Available We have previously reported the isolation and culture of a human breast epithelial cell type with stem cell characteristics (Type I HBEC from reduction mammoplasty using the MSU-1 medium. Subsequently, we have developed several different normal human adult stem cell types from different tissues using the K-NAC medium. In this study, we determined whether this low calcium K-NAC medium with antioxidants (N-acetyl-L-cysteine and L-ascorbic acid-2-phosphate is a better medium to grow human breast epithelial cells. The results clearly show that the K-NAC medium is a superior medium for prolonged growth (cumulative population doubling levels ranged from 30 to 40 of normal breast epithelial cells that expressed stem cell phenotypes. The characteristics of these mammary stem cells include deficiency in gap junctional intercellular communication, expression of Oct-4, and the ability to differentiate into basal epithelial cells and to form organoid showing mammary ductal and terminal end bud-like structures. Thus, this new method of growing Type I HBECs will be very useful in future studies of mammary development, breast carcinogenesis, chemoprevention, and cancer therapy.
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Cultured alveolar epithelial cells from septic rats mimic in vivo septic lung.
Directory of Open Access Journals (Sweden)
Taylor S Cohen
2010-06-01
Full Text Available Sepsis results in the formation of pulmonary edema by increasing in epithelial permeability. Therefore we hypothesized that alveolar epithelial cells isolated from septic animals develop tight junctions with different protein composition and reduced barrier function relative to alveolar epithelial cells from healthy animals. Male rats (200-300 g were sacrificed 24 hours after cecal ligation and double puncture (2CLP or sham surgery. Alveolar epithelial cells were isolated and plated on fibronectin-coated flexible membranes or permeable, non-flexible transwell substrates. After a 5 day culture period, cells were either lysed for western analysis of tight junction protein expressin (claudin 3, 4, 5, 7, 8, and 18, occludin, ZO-1, and JAM-A and MAPk (JNK, ERK, an p38 signaling activation, or barrier function was examined by measuring transepithelial resistance (TER or the flux of two molecular tracers (5 and 20 A. Inhibitors of JNK (SP600125, 20 microM and ERK (U0126, 10 microM were used to determine the role of these pathways in sepsis induced epithelial barrier dysfunction. Expression of claudin 4, claudin 18, and occludin was significantly lower, and activation of JNK and ERK signaling pathways was significantly increased in 2CLP monolayers, relative to sham monolayers. Transepithelial resistance of the 2CLP monolayers was reduced significantly compared to sham (769 and 1234 ohm-cm(2, respectively, however no significant difference in the flux of either tracer was observed. Inhibition of ERK, not JNK, significantly increased TER and expression of claudin 4 in 2CLP monolayers, and prevented significant differences in claudin 18 expression between 2CLP and sham monolayers. We conclude that alveolar epithelial cells isolated from septic animals form confluent monolayers with impaired barrier function compared to healthy monolayers, and inhibition of ERK signaling partially reverses differences between these monolayers. This model provides a unique
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CD44 plays a functional role in Helicobacter pylori-induced epithelial cell proliferation.
Directory of Open Access Journals (Sweden)
Nina Bertaux-Skeirik
2015-02-01
Full Text Available The cytotoxin-associated gene (Cag pathogenicity island is a strain-specific constituent of Helicobacter pylori (H. pylori that augments cancer risk. CagA translocates into the cytoplasm where it stimulates cell signaling through the interaction with tyrosine kinase c-Met receptor, leading cellular proliferation. Identified as a potential gastric stem cell marker, cluster-of-differentiation (CD CD44 also acts as a co-receptor for c-Met, but whether it plays a functional role in H. pylori-induced epithelial proliferation is unknown. We tested the hypothesis that CD44 plays a functional role in H. pylori-induced epithelial cell proliferation. To assay changes in gastric epithelial cell proliferation in relation to the direct interaction with H. pylori, human- and mouse-derived gastric organoids were infected with the G27 H. pylori strain or a mutant G27 strain bearing cagA deletion (∆CagA::cat. Epithelial proliferation was quantified by EdU immunostaining. Phosphorylation of c-Met was analyzed by immunoprecipitation followed by Western blot analysis for expression of CD44 and CagA. H. pylori infection of both mouse- and human-derived gastric organoids induced epithelial proliferation that correlated with c-Met phosphorylation. CagA and CD44 co-immunoprecipitated with phosphorylated c-Met. The formation of this complex did not occur in organoids infected with ∆CagA::cat. Epithelial proliferation in response to H. pylori infection was lost in infected organoids derived from CD44-deficient mouse stomachs. Human-derived fundic gastric organoids exhibited an induction in proliferation when infected with H. pylori that was not seen in organoids pre-treated with a peptide inhibitor specific to CD44. In the well-established Mongolian gerbil model of gastric cancer, animals treated with CD44 peptide inhibitor Pep1, resulted in the inhibition of H. pylori-induced proliferation and associated atrophic gastritis. The current study reports a unique
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Anti-inflammatory effects of embelin in A549 cells and human asthmatic airway epithelial tissues.
Lee, In-Seung; Cho, Dong-Hyuk; Kim, Ki-Suk; Kim, Kang-Hoon; Park, Jiyoung; Kim, Yumi; Jung, Ji Hoon; Kim, Kwanil; Jung, Hee-Jae; Jang, Hyeung-Jin
2018-02-01
Allergic asthma is the most common type in asthma, which is defined as a chronic inflammatory disease of the lung. In this study, we investigated whether embelin (Emb), the major component of Ardisia japonica BL. (AJB), exhibits anti-inflammatory effects on allergic asthma via inhibition of NF-κB activity using A549 cells and asthmatic airway epithelial tissues. Inflammation was induced in A549 cells, a human airway epithelial cell line, by IL-1β (10 ng/ml) treatment for 4 h. The effects of Emb on NF-κB activity and COX-2 protein expression in inflamed airway epithelial cells and human asthmatic airway epithelial tissues were analyzed via western blot. The secretion levels of NF-κB-mediated cytokines/chemokines, including IL-4, 6, 9, 13, TNF-α and eotaxin, were measured by a multiplex assay. Emb significantly blocked NF-κB activity in IL-1β-treated A549 cells and human asthmatic airway epithelial tissues. COX-2 expression was also reduced in both IL-1β-treated A549 cells and asthmatic tissues Emb application. Emb significantly reduced the secretion of IL-4, IL-6 and eotaxin in human asthmatic airway epithelial tissues by inhibiting activity of NF-κB. The results of this study suggest that Emb may be used as an anti-inflammatory agent via inhibition of NF-κB and related cytokines.
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Catanese, Muriel; Popovici, Cornel; Proust, Hélène; Hoffart, Louis; Matonti, Frédéric; Cochereau, Isabelle; Conrath, John; Gabison, Eric E
2011-02-22
To assess corneal epithelial cell survival after keratoplasty. Corneal impression cytology (CIC) was performed on sex-mismatched corneal transplants. Fluorescent in situ hybridization (FISH) with sex chromosome-specific probes was performed to identify epithelial cell mosaicism and therefore allocate the donor or recipient origin of the cells. Twenty-four samples of corneal epithelial cells derived from 21 transplanted patients were analyzed. All patients received post-operative treatment using dexamethasone eye drops, with progressive tapering over 18 months, and nine patients also received 2% cyclosporine eye drops. Out of the 24 samples reaching quality criteria, sex mosaicism was found in 13, demonstrating the presence of donor-derived cells at the center of the graft for at least 211 days post keratoplasty. Kaplan-Meier analysis established a median survival of donor corneal epithelial cells of 385 days. Although not statistically significant, the disappearance of donor cells seemed to be delayed and the average number of persistent cells appeared to be greater when 2% cyclosporine was used topically as an additional immunosuppressive therapy. The combination of corneal impressions and FISH analysis is a valuable tool with negligible side effects to investigate the presence of epithelial cell mosaicism in sex-mismatched donor transplants. Epithelial cells survived at the center of the graft with a median survival of more than one year, suggesting slower epithelial turnover than previously described.
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Kahounová, Zuzana; Kurfürstová, Daniela; Bouchal, Jan; Kharaishvili, Gvantsa; Navrátil, Jiří; Remšík, Ján; Šimečková, Šárka; Študent, Vladimír; Kozubík, Alois; Souček, Karel
2017-04-06
The identification of fibroblasts and cancer-associated fibroblasts from human cancer tissue using surface markers is difficult, especially because the markers used currently are usually not expressed solely by fibroblasts, and the identification of fibroblast-specific surface molecules is still under investigation. It was aimed to compare three commercially available antibodies in the detection of different surface epitopes of fibroblasts (anti-fibroblast, fibroblast activation protein α, and fibroblast surface protein). The specificity of their expression, employing fibroblast cell lines and tumor-derived fibroblasts from breast and prostate tissues was investigated. Both the established fibroblast cell line HFF-1 and ex vivo primary fibroblasts isolated from breast and prostate cancer tissues expressed the tested surface markers to different degrees. Surprisingly, those markers were expressed also by permanent cell lines of epithelial origin, both benign and cancer-derived (breast-cell lines MCF 10A, HMLE and prostate-cell lines BPH-1, DU 145, and PC-3). The expression of fibroblast activation protein α increased on the surface of previously described models of epithelial cells undergoing epithelial-to-mesenchymal transition in response to treatment with TGF-β1. To prove the co-expression of the fibroblast markers on cells of epithelial origin, we used freshly dissociated human prostate and breast cancer tissues. The results confirmed the co-expression of anti-fibroblast and fibroblast surface protein on CD31/CD45-negative/EpCAM-positive epithelial cells. In summary, our data support the findings that the tested fibroblast markers are not fibroblast specific and may be expressed also by cells of epithelial origin (e.g., cells undergoing EMT). Therefore, the expression of these markers should be interpreted with caution, and the combination of several epitopes for both positive (anti-fibroblast or fibroblast activation protein α) and negative (Ep
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UV-induced changes in cell cycle and gene expression within rabbit lens epithelial cells
International Nuclear Information System (INIS)
Sidjanin, D.; Grdina, D.; Woloschak, G.E.
1996-01-01
Damage to lens epithelial cells is a probable initiation process in cataract formation mediated by UV radiation. In these experiments, we investigated the effects of exposure to 254 nm radiation on cell cycle progression in the rabbit lens epithelial cell line N/N1003A. The RNA was harvested at various times following exposure to UV (254 nm) radiation and analyzed by dot-blot and northern blot hybridizations. These results revealed that during the first 6 h following exposure of the cells to UV, there was, associated with decreasing dose, a decrease in accumulation of transcripts specific for histones H3 and H4 and an increase in the mRNA encoding protein kinase C and β- and γ-actin. Using flow cytometry, we detected an accumulation of cells in G1/S phase of the cell cycle 1 h following exposure to 254 nm radiation. The observed changes in gene expression, especially the decreased accumulation of histone transcripts reported here, may play a role in UV-induced inhibition of cell cycle progression. (Author)
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Directory of Open Access Journals (Sweden)
Pamela Di Tomo
Full Text Available Amniotic fluid-derived stem (AFS cells have been identified as a promising source for cell therapy applications in bone traumatic and degenerative damage. Calcium Sensing Receptor (CaSR, a G protein-coupled receptor able to bind calcium ions, plays a physiological role in regulating bone metabolism. It is expressed in different kinds of cells, as well as in some stem cells. The bone CaSR could potentially be targeted by allosteric modulators, in particular by agonists such as calcimimetic R-568, which may potentially be helpful for the treatment of bone disease. The aim of our study was first to investigate the presence of CaSR in ovine Amniotic Fluid Mesenchymal Stem Cells (oAFMSCs and then the potential role of calcimimetics in in vitro osteogenesis. oAFMSCs were isolated, characterized and analyzed to examine the possible presence of CaSR by western blotting and flow cytometry analysis. Once we had demonstrated CaSR expression, we worked out that 1 µM R-568 was the optimal and effective concentration by cell viability test (MTT, cell number, Alkaline Phosphatase (ALP and Alizarin Red S (ARS assays. Interestingly, we observed that basal diffuse CaSR expression in oAFMSCs increased at the membrane when cells were treated with R-568 (1 µM, potentially resulting in activation of the receptor. This was associated with significantly increased cell mineralization (ALP and ARS staining and augmented intracellular calcium and Inositol trisphosphate (IP3 levels, thus demonstrating a potential role for calcimimetics during osteogenic differentiation. Calhex-231, a CaSR allosteric inhibitor, totally reversed R-568 induced mineralization. Taken together, our results demonstrate for the first time that CaSR is expressed in oAFMSCs and that calcimimetic R-568, possibly through CaSR activation, can significantly improve the osteogenic process. Hence, our study may provide useful information on the mechanisms regulating osteogenesis in oAFMSCs, perhaps
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... Supplements Videos & Tools Español You Are Here: Home → Medical Encyclopedia → Amniotic band sequence URL of this page: //medlineplus.gov/ency/ ... birth. The baby should be delivered in a medical center that has specialists experienced in caring for babies ... or partial loss of function of a body part. Congenital bands affecting large parts of the body cause the ...
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Chehelcheraghi, Farzaneh; Abbaszadeh, Abolfazl; Tavafi, Magid
2018-03-06
Skin flap procedures are employed in plastic surgery, but failure can lead to necrosis of the flap. Studies have used bone marrow mesenchymal stem cells (BM-MSCs) to improve flap viability. BM-MSCs and acellular amniotic membrane (AAM) have been introduced as alternatives. The objective of this study was to evaluate the effect of BM-MSCs and AAM on mast cells of random skin flaps (RSF) in rats. RSFs (80 × 30 mm) were created on 40 rats that were randomly assigned to one of four groups, including (I) AAM, (II) BM-MSCs, (III) BM-MSCs/AAM, and (IV) saline (control). Transplantation was carried out during the procedure (zero day). Flap necrosis was observed on day 7, and skin samples were collected from the transition line of the flap to evaluate the total number and types of mast cells. The development and the total number of mast cells were related to the development of capillaries. The results of one-way ANOVA indicated that there was no statistically significant difference between the mean numbers of mast cell types for different study groups. However, the difference between the total number of mast cells in the study groups was statistically significant (p = 0.001). The present study suggests that the use of AAM/BM-MSCs can improve the total number of mast cells and accelerate the growth of capillaries at the transient site in RSFs in rats.
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Directory of Open Access Journals (Sweden)
Marta Barba
2012-01-01
Full Text Available Multipotent mesenchymal stem cells with extensive self-renewal properties can be easily isolated and rapidly expanded in culture from small volumes of amniotic fluid. These cells, namely, amniotic fluid-stromal cells (AFSCs, can be regarded as an attractive source for tissue engineering purposes, being phenotypically and genetically stable, plus overcoming all the safety and ethical issues related to the use of embryonic/fetal cells. LMP3 is a novel osteoinductive molecule acting upstream to the main osteogenic pathways. This study is aimed at delineating the basic molecular events underlying LMP3-induced osteogenesis, using AFSCs as a cellular model to focus on the molecular features underlying the multipotency/differentiation switch. For this purpose, AFSCs were isolated and characterized in vitro and transfected with a defective adenoviral vector expressing the human LMP3. LMP3 induced the successful osteogenic differentiation of AFSC by inducing the expression of osteogenic markers and osteospecific transcription factors. Moreover, LMP3 induced an early repression of the kruppel-like factor-4, implicated in MSC stemness maintenance. KLF4 repression was released upon LMP3 silencing, indicating that this event could be reasonably considered among the basic molecular events that govern the proliferation/differentiation switch during LMP3-induced osteogenic differentiation of AFSC.
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Gong, Baolan; Yue, Yan; Wang, Renxiao; Zhang, Yi; Jin, Quanfang; Zhou, Xi
2017-06-01
The epithelial-mesenchymal transition is the key process driving cancer metastasis. MicroRNA-194 inhibits epithelial-mesenchymal transition in several cancers and its downregulation indicates a poor prognosis in human endometrial carcinoma. Self-renewal factor Sox3 induces epithelial-mesenchymal transition at gastrulation and is also involved epithelial-mesenchymal transition in several cancers. We intended to determine the roles of Sox3 in inducing epithelial-mesenchymal transition in endometrial cancer stem cells and the possible role of microRNA-194 in controlling Sox3 expression. Firstly, we found that Sox3 and microRNA-194 expressions were associated with the status of endometrial cancer stem cells in a panel of endometrial carcinoma tissue, the CD133+ cell was higher in tumorsphere than in differentiated cells, and overexpression of microRNA-194 would decrease CD133+ cell expression. Silencing of Sox3 in endometrial cancer stem cell upregulated the epithelial marker E-cadherin, downregulated the mesenchymal marker vimentin, and significantly reduced cell invasion in vitro; overexpression of Sox3 reversed these phenotypes. Furthermore, we discovered that the expression of Sox3 was suppressed by microRNA-194 through direct binding to the Sox3 3'-untranslated region. Ectopic expression of microRNA-194 in endometrial cancer stem cells induced a mesenchymal-epithelial transition by restoring E-cadherin expression, decreasing vimentin expression, and inhibiting cell invasion in vitro. Moreover, overexpression of microRNA-194 inhibited endometrial cancer stem cell invasion or metastasis in vivo by injection of adenovirus microRNA-194. These findings demonstrate the novel mechanism by which Sox3 contributes to endometrial cancer stem cell invasion and suggest that repression of Sox3 by microRNA-194 may have therapeutic potential to suppress endometrial carcinoma metastasis. The cancer stem cell marker, CD133, might be the surface marker of endometrial cancer stem
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CDDO-Me protects normal lung and breast epithelial cells but not cancer cells from radiation.
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Mariam El-Ashmawy
Full Text Available Although radiation therapy is commonly used for treatment for many human diseases including cancer, ionizing radiation produces reactive oxygen species that can damage both cancer and healthy cells. Synthetic triterpenoids, including CDDO-Me, act as anti-inflammatory and antioxidant modulators primarily by inducing the transcription factor Nrf2 to activate downstream genes containing antioxidant response elements (AREs. In the present series of experiments, we determined if CDDO-Me can be used as a radioprotector in normal non-cancerous human lung and breast epithelial cells, in comparison to lung and breast cancer cell lines. A panel of normal non-cancerous, partially cancer progressed, and cancer cell lines from both lung and breast tissue was exposed to gamma radiation with and without pre-treatment with CDDO-Me. CDDO-Me was an effective radioprotector when given ∼18 hours before radiation in epithelial cells (average dose modifying factor (DMF = 1.3, and Nrf2 function was necessary for CDDO-Me to exert these radioprotective effects. CDDO-Me did not protect cancer lines tested from radiation-induced cytotoxicity, nor did it protect experimentally transformed human bronchial epithelial cells (HBECs with progressive oncogenic manipulations. CDDO-Me also protected human lymphocytes against radiation-induced DNA damage. A therapeutic window exists in which CDDO-Me protects normal cells from radiation by activating the Nrf2 pathway, but does not protect experimentally transformed or cancer cell lines. This suggests that use of this oral available, non-toxic class of drug can protect non-cancerous healthy cells during radiotherapy, resulting in better outcomes and less toxicity for patients.
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Normal morphogenesis of epithelial tissues and progression of epithelial tumors
Wang, Chun-Chao; Jamal, Leen; Janes, Kevin A.
2011-01-01
Epithelial cells organize into various tissue architectures that largely maintain their structure throughout the life of an organism. For decades, the morphogenesis of epithelial tissues has fascinated scientists at the interface of cell, developmental, and molecular biology. Systems biology offers ways to combine knowledge from these disciplines by building integrative models that are quantitative and predictive. Can such models be useful for gaining a deeper understanding of epithelial morphogenesis? Here, we take inventory of some recurring themes in epithelial morphogenesis that systems approaches could strive to capture. Predictive understanding of morphogenesis at the systems level would prove especially valuable for diseases such as cancer, where epithelial tissue architecture is profoundly disrupted. PMID:21898857
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Amniotic Tissues for the Treatment of Chronic Plantar Fasciosis and Achilles Tendinosis
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Bruce Werber
2015-01-01
Full Text Available Introduction. Allogeneic amniotic tissue and fluid may be used to treat chronic plantar fasciosis and Achilles tendinosis. This innovative approach involves delivering a unique allograft of live human cells in a nonimmunogenic structural tissue matrix to treat chronic tendon injury. These tissues convey very positive regenerative attributes; procurement is performed with maternal consent during elective caesarian birth. Materials and Methods. In the present investigation all patients were unresponsive to multiple standard therapies for a minimum of 6 months and were treated with one implantation of PalinGen SportFLOW around the plantar fascia and/or around the Achilles paratenon. The patients were given a standard protocol for postimplant active rehabilitation. Results. The analogue pretreatment pain score (VAS of 8. By the fourth week after treatment, all patients had significantly reduced self-reported pain. Twelve weeks following the procedure the average pain level had reduced to only 2. No adverse reactions were reported in any of the patients. Conclusion. All patients in this study experienced heel or Achilles pain, unresponsive to standard therapy protocols. After treatment all patients noted significant pain reduction, indicating that granulized amniotic membrane and amniotic fluid can be successfully used to treat both chronic plantar fasciosis and Achilles tendinosis.
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Establishment and evaluation of a stable cattle type II alveolar epithelial cell line.
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Feng Su
Full Text Available Macrophages and dendritic cells are recognized as key players in the defense against mycobacterial infection. Recent research has confirmed that alveolar epithelial cells (AECs also play important roles against mycobacterium infections. Thus, establishing a stable cattle AEC line for future endogenous immune research on bacterial invasion is necessary. In the present study, we first purified and immortalized type II AECs (AEC II cells by transfecting them with a plasmid containing the human telomerase reverse trancriptase gene. We then tested whether or not the immortalized cells retained the basic physiological properties of primary AECs by reverse-transcription polymerase chain reaction and Western blot. Finally, we tested the secretion capacity of immortalized AEC II cells upon stimulation by bacterial invasion. The cattle type II alveolar epithelial cell line (HTERT-AEC II that we established retained lung epithelial cell characteristics: the cells were positive for surfactants A and B, and they secreted tumor necrosis factor-α and interleukin-6 in response to bacterial invasion. Thus, the cell line we established is a potential tool for research on the relationship between AECs and Mycobacterium tuberculosis.
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High Glucose Increases Metallothionein Expression in Renal Proximal Tubular Epithelial Cells
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Daisuke Ogawa
2011-01-01
Full Text Available Metallothionein (MT is an intracellular metal-binding, cysteine-rich protein, and is a potent antioxidant that protects cells and tissues from oxidative stress. Although the major isoforms MT-1 and -2 (MT-1/-2 are highly inducible in many tissues, the distribution and role of MT-1/-2 in diabetic nephropathy are poorly understood. In this study, diabetes was induced in adult male rats by streptozotocin, and renal tissues were stained with antibodies for MT-1/-2. MT-1/-2 expression was also evaluated in mProx24 cells, a mouse renal proximal tubular epithelial cell line, stimulated with high glucose medium and pretreated with the antioxidant vitamin E. MT-1/-2 expression was gradually and dramatically increased, mainly in the proximal tubular epithelial cells and to a lesser extent in the podocytes in diabetic rats, but was hardly observed in control rats. MT-1/-2 expression was also increased by high glucose stimulation in mProx24 cells. Because the induction of MT was suppressed by pretreatment with vitamin E, the expression of MT-1/-2 is induced, at least in part, by high glucose-induced oxidative stress. These observations suggest that MT-1/-2 is induced in renal proximal tubular epithelial cells as an antioxidant to protect the kidney from oxidative stress, and may offer a novel therapeutic target against diabetic nephropathy.
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Cell Extrusion: A Stress-Responsive Force for Good or Evil in Epithelial Homeostasis.
Ohsawa, Shizue; Vaughen, John; Igaki, Tatsushi
2018-02-05
Epithelial tissues robustly respond to internal and external stressors via dynamic cellular rearrangements. Cell extrusion acts as a key regulator of epithelial homeostasis by removing apoptotic cells, orchestrating morphogenesis, and mediating competitive cellular battles during tumorigenesis. Here, we delineate the diverse functions of cell extrusion during development and disease. We emphasize the expanding role for apoptotic cell extrusion in exerting morphogenetic forces, as well as the strong intersection of cell extrusion with cell competition, a homeostatic mechanism that eliminates aberrant or unfit cells. While cell competition and extrusion can exert potent, tumor-suppressive effects, dysregulation of either critical homeostatic program can fuel cancer progression. Copyright © 2018 Elsevier Inc. All rights reserved.
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Cellular toxicity of calf blood extract on human corneal epithelial cells in vitro.
Park, Young Min; Kim, Su Jin; Han, Young Sang; Lee, Jong Soo
2015-01-01
To investigate the biologic effects of the calf blood extract on corneal epithelial cells in vitro. The effects on corneal epithelial cells were evaluated after 1, 4, 12, and 24 h of exposure to various concentrations of calf blood extract (3, 5, 8 and 16%). The MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide) assay was performed to measure levels of cellular metabolic activity. The lactate dehydrogenase (LDH) assay was performed to determine the extent of cellular damage. Cellular morphology was examined using phase-contrast microscopy. The scratch wound assay was performed to quantify the migration of corneal epithelial cells. At the 3 and 5% concentrations of calf blood extract, MTT values were similar to those observed in the control group. However, at a concentration of 8 and 16%, cellular metabolic activity was significantly decreased after 4 h of exposure to calf blood extract. After 12 h of exposure to 8 and 16% concentrations of calf blood extract, LDH activity and cellular morphological damage to the corneal epithelial cells were significantly increased. There was no evidence of cellular migration after 12 h exposure to 5% or higher concentration of calf blood extract because of cellular toxicity. Compared with normal corneal epithelial cells, the cellular activity was decreased, and toxicity was increased after over 12 h of exposure to more than 5% concentration of calf blood extract. Further clinical studies will be necessary to determine the optimal concentration and exposure time for the topical application of eye drops containing calf blood extract.
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Nomeda Girnius
2017-11-01
Full Text Available Summary: Developmental morphogenesis, tissue injury, and oncogenic transformation can cause the detachment of epithelial cells. These cells are eliminated by a specialized form of apoptosis (anoikis. While the processes that contribute to this form of cell death have been studied, the underlying mechanisms remain unclear. Here, we tested the role of the cJUN NH2-terminal kinase (JNK signaling pathway using murine models with compound JNK deficiency in mammary and kidney epithelial cells. These studies demonstrated that JNK is required for efficient anoikis in vitro and in vivo. Moreover, JNK-promoted anoikis required pro-apoptotic members of the BCL2 family of proteins. We show that JNK acts through a BAK/BAX-dependent apoptotic pathway by increasing BIM expression and phosphorylating BMF, leading to death of detached epithelial cells. : Developmental morphogenesis, tissue injury, and oncogenic transformation can cause epithelial cell detachment. These cells are eliminated by a specialized form of apoptosis termed anoikis. Girnius and Davis show that anoikis is mediated by the cJUN NH2-terminal kinase (JNK, which increases BIM expression and phosphorylates BMF to engage BAK/BAX-dependent apoptosis. Keywords: apoptosis, anoikis, epithelial cell, mammary gland, JNK, BAX, BAK, BH3-only protein, BIM, BMF
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Trichomonas vaginalis perturbs the junctional complex in epithelial cells
Institute of Scientific and Technical Information of China (English)
无
2005-01-01
Trichomonas vaginalis, a protist parasite of the urogenital tract in humans, is the causative agent of trichomonosis,which in recent years have been associated with the cervical cancer development. In the present study we analyzed the modifications at the junctional complex level of Caco-2 cells after interaction with two isolates of T. vaginalis and the influence of the iron concentration present in the parasite's culture medium on the interaction effects. Our results show that T. vaginalis adheres to the epithelial cell causing alterations in the junctional complex, such as: (a) a decrease in transepithelial electrical resistance; (b) alteration in the pattern of junctional complex proteins distribution as obseryed for E-cadherin, occludin and ZO-1; and (c) enlargement of the spaces between epithelial cells. These effects were dependent on (a) the degree of the parasite virulence isolate, (b) the iron concentration in the culture medium, and (c) the expression of adhesin proteins on the parasite surface.
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Histamine Induces Bovine Rumen Epithelial Cell Inflammatory Response via NF-κB Pathway.
Sun, Xudong; Yuan, Xue; Chen, Liang; Wang, Tingting; Wang, Zhe; Sun, Guoquan; Li, Xiaobing; Li, Xinwei; Liu, Guowen
2017-01-01
Subacute ruminal acidosis (SARA) is a common disease in high-producing lactating cows. Rumenitis is the initial insult of SARA and is associated with the high concentrations of histamine produced in the rumen of dairy cows during SARA. However, the exact mechanism remains unclear. The objective of the current study is to investigate whether histamine induces inflammation of rumen epithelial cells and the underlying mechanism of this process. Bovine rumen epithelial cells were cultured and treated with different concentrations of histamine and pyrrolidine dithiocarbamate (PDTC, an NF-κB inhibitor) cultured in different pH medium (pH 7.2 or 5.5). qRT-PCR, Western-blotting, ELISA and immunocytofluorescence were used to evaluate whether histamine activated the NF-κB pathway and inflammatory cytokines. The results showed that histamine significantly increased the activity of IKK β and the phosphorylation levels of IκB α, as well as upregulated the mRNA and protein expression levels of NF-κB p65 in the rumen epithelial cells cultured in neutral (pH=7.2) and acidic (pH=5.5) medium. Furthermore, histamine treatment also significantly increased the transcriptional activity of NF-κB p65. High expression and transcriptional activity of NF-κB p65 significantly increased the mRNA expressions and concentrations of inflammatory cytokines, tumor necrosis factor alpha (TNF-α), interleukin 6 (IL-6) and interleukin 1 beta (IL-1β), thereby inducing the inflammatory response in bovine rumen epithelial cells. However, inhibition of NF-κB p65 by PDTC significantly decreased the expressions and concentrations of the inflammatory cytokines induced by histamine in the rumen epithelial cells cultured in the neutral and acidic medium. The present data indicate that histamine induces the inflammatory response of bovine rumen epithelial cells through the NF-κB pathway. © 2017 The Author(s). Published by S. Karger AG, Basel.
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Selective interactions between epithelial tumour cells and bone marrow mesenchymal stem cells
Hombauer, H; Minguell, J J
2000-01-01
This work is a comparative study on the features displayed by an epithelial metastatic breast cancer cell line (MCF-7) when set in co-culture with human bone marrow mesenchymal stem cells (MSC) or a feeder layer of 3T3 fibroblasts. MSC, a subset of non-haematopoietic cells in the marrow stroma, display a potential for self-renewal, proliferation and differentiation into precursors for bone, cartilage, connective and muscular tissue. Adhesion of MCF-7 cells to monolayers of MSC or 3T3 was high...
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File list: His.Utr.05.AllAg.Fallopian_tube_secretory_epithelial_cell [Chip-atlas[Archive
Lifescience Database Archive (English)
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Kayastha, Forum; Madhu, Hardik; Vasavada, Abhay; Johar, Kaid
2014-11-01
Lens epithelial cell proliferation, migration, and transdifferentiation are involved in the development of subcapsular cataracts and postoperative capsular opacification (PCO). PI3K/Akt pathway is involved in the proliferation and migration of lens epithelial cells. Andrographolide is the main bioactive component of Andrographis paniculata and is known to possess anti-proliferative and anti-migratory activities. The purpose of this study is to evaluate the effect of andrographolide on proliferation and migration induced by growth factors (TGF-β and bFGF) in the lens epithelial cell line, FHL 124. We have also evaluated the role of the PI3K/Akt pathway and its alteration by andrographolide during proliferation and migration of lens epithelial cells. The results showed that andrographolide significantly inhibited proliferation in a dose and time dependent manner. The growth factors, TGF-β and bFGF, induced migration of lens epithelial cells, which was lowered by andrographolide. The growth factors also up regulated phosphorylated Akt (Ser473) and Akt (Thr308), which was abolished by simultaneous treatment of andrographolide. Similar changes were also observed with the PI3K inhibitor, LY290042. Our findings suggest that andrographolide reduces proliferation, migration, and phosphorylated Akt levels in lens epithelial cells. Hence andrographolide can be utilized for the prevention of PCO. Copyright © 2014 Elsevier Ltd. All rights reserved.
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Parietal epithelial cells and podocytes in glomerular diseases
Smeets, B.; Moeller, M.J.
2012-01-01
In recent years, it has become apparent that parietal epithelial cells (PECs) play an important role within the renal glomerulus, in particular in diseased conditions. In this review, we examine current knowledge about the role of PECs and their interactions with podocytes in development and under
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Suppression of ICE and Apoptosis in Mammary Epithelial Cells by Extracellular Matrix
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Boudreau, Nancy; Sympson, C. J.; Werb, Zena; Bissell, Mina J.
1994-12-01
Apoptosis (programmed cell death) plays a major role in development and tissue regeneration. Basement membrane extracellular matrix (ECM), but not fibronectin or collagen, was shown to suppress apoptosis of mammary epithelial cells in tissue culture and in vivo. Apoptosis was induced by antibodies to beta 1 integrins or by overexpression of stromelysin-1, which degrades ECM. Expression of interleukin-1 beta converting enzyme (ICE) correlated with the loss of ECM, and inhibitors of ICE activity prevented apoptosis. These results suggest that ECM regulates apoptosis in mammary epithelial cells through an integrin-dependent negative regulation of ICE expression.
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Gingerich, Aaron; Pang, Lan; Hanson, Jarod; Dlugolenski, Daniel; Streich, Rebecca; Lafontaine, Eric R; Nagy, Tamás; Tripp, Ralph A; Rada, Balázs
2016-01-01
Our aim was to study whether an extracellular, oxidative antimicrobial mechanism inherent to tracheal epithelial cells is capable of inactivating influenza H1N2 virus. Epithelial cells were isolated from tracheas of male Sprague-Dawley rats. Both primary human and rat tracheobronchial epithelial cells were differentiated in air-liquid interface cultures. A/swine/Illinois/02860/09 (swH1N2) influenza A virions were added to the apical side of airway cells for 1 h in the presence or absence of lactoperoxidase or thiocyanate. Characterization of rat epithelial cells (morphology, Duox expression) occurred via western blotting, PCR, hydrogen peroxide production measurement and histology. The number of viable virions was determined by plaque assays. Statistical difference of the results was analyzed by ANOVA and Tukey's test. Our data show that rat tracheobronchial epithelial cells develop a differentiated, polarized monolayer with high transepithelial electrical resistance, mucin production and expression of dual oxidases. Influenza A virions are inactivated by human and rat epithelial cells via a dual oxidase-, lactoperoxidase- and thiocyanate-dependent mechanism. Differentiated air-liquid interface cultures of rat tracheal epithelial cells provide a novel model to study airway epithelium-influenza interactions. The dual oxidase/lactoperoxidase/thiocyanate extracellular oxidative system producing hypothiocyanite is a fast and potent anti-influenza mechanism inactivating H1N2 viruses prior to infection of the epithelium.
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Deep RNA-Seq analysis reveals unexpected features of human prostate basal epithelial cells
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Dingxiao Zhang
2016-03-01
Full Text Available Prostate cancer is the second leading cause of cancer-related deaths among American men [1]. The prostate gland mainly contains basal and luminal cells, which are constructed as a pseudostratified epithelium. Annotation of prostate epithelial transcriptomes provides a foundation for discoveries that can impact disease understanding and treatment. Here, for the first time, we describe a whole-genome transcriptome analysis of human benign prostatic basal and luminal populations by using deep RNA sequencing (GSE67070 [2]. Combined with comprehensive molecular and biological characterizations, we show that the differential gene expression profiles account for their distinct functional phenotypes. Strikingly, in contrast to luminal cells, basal cells preferentially express gene categories associated with stem cells, neural and neuronal development, and RNA processing. Of clinical relevance, the treatment failed castration-resistant and anaplastic prostate cancers molecularly resemble a basal-like phenotype. We also identified genes associated with patient clinical outcome. Therefore, we provide a gene expression resource for understanding human prostate epithelial lineages, and link the cell-type specific gene signatures to subtypes of prostate cancer development. Keywords: Prostate epithelial cells, Basal cells, Luminal cells, RNA-seq
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Testosterone determination in amniotic fluid for sec diagnosis
International Nuclear Information System (INIS)
Quiroa C, M.M.
1985-11-01
This study was carried on 50 samples of amniotic fluid obtained from pregnant patients with gestation old of 38 to 40 weeks; diagnosis of the foetus sex was made by measuring the testosterone levels by radioimmunoassay technique. It was found that 94% of the cases were correctly diagnosed. The testosterone levels found in the amniotic fluid of male and female foetus were significantly different (L<0.01) these confirm the efficacy of the method. (author)
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He, Guiliang; Ma, Mengru; Yang, Wei; Wang, Hao; Zhang, Yong; Gao, Ming-Qing
2017-01-01
Fibroblasts constitute the majority of the stromal cells within bovine mammary gland, yet the functional contributions of these cells to mastitis and fibrosis and the mechanism are poorly understood. In this study, we demonstrate that inflammation-associated fibroblasts (INFs) extracted from bovine mammary glands with clinical mastitis had different expression pattern regarding to several extracellular matrix (ECM) proteins, chemokines and cytokines compared to normal fibroblasts (NFs) from dairy cows during lactation. The INFs induced epithelial-mesenchymal transition (EMT) and inflammatory responses of mammary epithelial cells in a vitro co-culture model. These functional contributions of INFs to normal epithelial cells were mediated through their ability to secrete stromal cell-derived factor 1 (SDF-1). SDF-1 was highly secreted/expressed by INFs, lipopolysaccharide (LPS) -treated NFs, lipoteichoic acid (LTA) -treated NFs, as well as mastitic tissue compared to their counterparts. Exogenous SDF-1 promoted EMT on epithelial cells through activating NF-κB pathway, induced inflammation response and inhibited proliferation of epithelial cells. In addition, SDF-1 was able to induce mastitis and slight fibrosis of mouse mammary gland, which was attenuated by a specific inhibitor of the receptor of SDF-1. Our findings indicate that stromal fibroblasts within mammary glands with mastitis contribute to EMT and inflammatory responses of epithelial cells through the secretion of SDF-1, which could result in the inflammation spread and fibrosis within mammary gland.
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Prohibitin 1 modulates mitochondrial stress-related autophagy in human colonic epithelial cells.
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Arwa S Kathiria
Full Text Available Autophagy is an adaptive response to extracellular and intracellular stress by which cytoplasmic components and organelles, including damaged mitochondria, are degraded to promote cell survival and restore cell homeostasis. Certain genes involved in autophagy confer susceptibility to Crohn's disease. Reactive oxygen species and pro-inflammatory cytokines such as tumor necrosis factor α (TNFα, both of which are increased during active inflammatory bowel disease, promote cellular injury and autophagy via mitochondrial damage. Prohibitin (PHB, which plays a role in maintaining normal mitochondrial respiratory function, is decreased during active inflammatory bowel disease. Restoration of colonic epithelial PHB expression protects mice from experimental colitis and combats oxidative stress. In this study, we investigated the potential role of PHB in modulating mitochondrial stress-related autophagy in intestinal epithelial cells.We measured autophagy activation in response to knockdown of PHB expression by RNA interference in Caco2-BBE and HCT116 WT and p53 null cells. The effect of exogenous PHB expression on TNFα- and IFNγ-induced autophagy was assessed. Autophagy was inhibited using Bafilomycin A(1 or siATG16L1 during PHB knockdown and the affect on intracellular oxidative stress, mitochondrial membrane potential, and cell viability were determined. The requirement of intracellular ROS in siPHB-induced autophagy was assessed using the ROS scavenger N-acetyl-L-cysteine.TNFα and IFNγ-induced autophagy inversely correlated with PHB protein expression. Exogenous PHB expression reduced basal autophagy and TNFα-induced autophagy. Gene silencing of PHB in epithelial cells induces mitochondrial autophagy via increased intracellular ROS. Inhibition of autophagy during PHB knockdown exacerbates mitochondrial depolarization and reduces cell viability.Decreased PHB levels coupled with dysfunctional autophagy renders intestinal epithelial cells
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Role of epithelial cells in idiopathic pulmonary fibrosis: from innocent targets to serial killers.
Selman, Moisés; Pardo, Annie
2006-06-01
Idiopathic pulmonary fibrosis (IPF), a progressive and relentless lung scarring of unknown etiology, has been recognized as the most lethal interstitial lung disease. Despite the growing interest in IPF, the precise molecular mechanisms underlying the development of fibrosis and leading to the irreversible destruction of the lung are still unknown. Recently, it has been proposed that IPF, instead of being a chronic inflammatory disorder, results from multiple cycles of epithelial cell injury and activation. In turn, active alveolar epithelial cells provoke the migration, proliferation, and activation of mesenchymal cells with the formation of fibroblastic/myofibroblastic foci and the exaggerated accumulation of extracellular matrix, mirroring abnormal wound repair. In this article, some characteristics of the alveolar epithelium are briefly outlined, and the fibrogenic mechanisms specifically operated by active abnormal epithelial cells are examined.
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Directory of Open Access Journals (Sweden)
Heni Susilowati
2017-01-01
Full Text Available Pseudomonas aeruginosa is a Gram-negative bacterium and causes respiratory infection especially in elderly patients. Royal jelly has been used worldwide as a traditional remedy and as a nutrient; however, the effect against P. aeruginosa is unclear. The aim of this study was to analyze antibacterial, antiadherent, and anti-inflammatory effects of royal jelly against P. aeruginosa. Wild-type strain PAO1 and clinical isolates of P. aeruginosa were used for antibacterial assay and antiadherent assay to abiotic surface and epithelial cells, which are pharynx (Detroit 562 and lung (NCI-H292 epithelial cells. In anti-inflammatory assay, epithelial cells were pretreated with royal jelly before bacterial exposure to investigate its inhibitory effect on interleukin (IL-8 and macrophage inflammatory protein-3α/CCL20 overproduction. Although royal jelly did not have antibacterial activity at concentration of 50% w/v, antiadherent activity was confirmed on the abiotic surface and epithelial cells under concentration of 25%. Pretreatment with royal jelly significantly inhibited overproduction of IL-8 and CCL20 from both cells. These results demonstrated that royal jelly inhibits P. aeruginosa adherence and protects epithelial cells from excessive inflammatory responses against P. aeruginosa infection. Our findings suggested that royal jelly may be a useful supplement as complementary and alternative medicine for preventing respiratory infection caused by P. aeruginosa.