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Sample records for progerin-positive fibroblasts localize

  1. Tumor-associated fibroblasts predominantly come from local and not circulating precursors.

    Science.gov (United States)

    Arina, Ainhoa; Idel, Christian; Hyjek, Elizabeth M; Alegre, Maria-Luisa; Wang, Ying; Bindokas, Vytautas P; Weichselbaum, Ralph R; Schreiber, Hans

    2016-07-05

    Fibroblasts are common cell types in cancer stroma and lay down collagen required for survival and growth of cancer cells. Although some cancer therapy strategies target tumor fibroblasts, their origin remains controversial. Multiple publications suggest circulating mesenchymal precursors as a source of tumor-associated fibroblasts. However, we show by three independent approaches that tumor fibroblasts derive primarily from local, sessile precursors. First, transplantable tumors developing in a mouse expressing green fluorescent reporter protein (EGFP) under control of the type I collagen (Col-I) promoter (COL-EGFP) had green stroma, whereas we could not find COL-EGFP(+) cells in tumors developing in the parabiotic partner lacking the fluorescent reporter. Lack of incorporation of COL-EGFP(+) cells from the circulation into tumors was confirmed in parabiotic pairs of COL-EGFP mice and transgenic mice developing autochthonous intestinal adenomas. Second, transplantable tumors developing in chimeric mice reconstituted with bone marrow cells from COL-EGFP mice very rarely showed stromal fibroblasts expressing EGFP. Finally, cancer cells injected under full-thickness COL-EGFP skin grafts transplanted in nonreporter mice developed into tumors containing green stromal cells. Using multicolor in vivo confocal microscopy, we found that Col-I-expressing fibroblasts constituted approximately one-third of the stromal mass and formed a continuous sheet wrapping the tumor vessels. In summary, tumors form their fibroblastic stroma predominantly from precursors present in the local tumor microenvironment, whereas the contribution of bone marrow-derived circulating precursors is rare.

  2. Local fibroblast proliferation but not influx is responsible for synovial hyperplasia in a murine model of rheumatoid arthritis

    Energy Technology Data Exchange (ETDEWEB)

    Matsuo, Yusuke; Mizoguchi, Fumitaka; Saito, Tetsuya [Department of Rheumatology, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University (TMDU), 1-5-45 Yushima, Bunkyo-ku, Tokyo, 113-8519 (Japan); Japan Science and Technology Agency (JST), Core Research for Evolutional Science and Technology (CREST) Program, Sanbancho, Chiyoda-ku, Tokyo, 102-0075 (Japan); Kawahata, Kimito [Department of Rheumatology, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University (TMDU), 1-5-45 Yushima, Bunkyo-ku, Tokyo, 113-8519 (Japan); Ueha, Satoshi; Matsushima, Kouji [Japan Science and Technology Agency (JST), Core Research for Evolutional Science and Technology (CREST) Program, Sanbancho, Chiyoda-ku, Tokyo, 102-0075 (Japan); Department of Molecular Preventive Medicine, Graduate School of Medicine, University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo, 113-0033 (Japan); Inagaki, Yutaka [Japan Science and Technology Agency (JST), Core Research for Evolutional Science and Technology (CREST) Program, Sanbancho, Chiyoda-ku, Tokyo, 102-0075 (Japan); Center for Matrix Biology and Medicine, Graduate School of Medicine and the Institute of Medical Sciences, Tokai University, 143 Shimo-kasuya, Isehara, Kanagawa, 259-1193 (Japan); Miyasaka, Nobuyuki [Department of Rheumatology, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University (TMDU), 1-5-45 Yushima, Bunkyo-ku, Tokyo, 113-8519 (Japan); Kohsaka, Hitoshi, E-mail: kohsaka.rheu@tmd.ac.jp [Department of Rheumatology, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University (TMDU), 1-5-45 Yushima, Bunkyo-ku, Tokyo, 113-8519 (Japan); Japan Science and Technology Agency (JST), Core Research for Evolutional Science and Technology (CREST) Program, Sanbancho, Chiyoda-ku, Tokyo, 102-0075 (Japan)

    2016-02-12

    Synovial fibroblasts play crucial roles in inflammation and joint destruction in rheumatoid arthritis (RA). How they accumulate in the RA joints remains unclear. This study was conducted to discern whether cellular influx from the outside of the joints and local proliferation are responsible for synovial fibroblast accumulation in an animal model of RA. We found that synovial fibroblasts were identified as GFP+ cells using collagen type I alpha 2 (Col1a2)-GFP transgenic reporter mice. Then, bone marrow transplantation and parabiosis techniques were utilized to study the cellular influx. Irradiated wild-type mice were transplanted with bone marrow from Col1a2-GFP mice. Col1a2-GFP and wild-type mice were conjoined for parabiosis. The transplanted mice and the parabionts were subjected to collagen antibody-induced arthritis (CAIA). We found no GFP+ cells in the hyperplastic synovial tissues from the transplanted mice with CAIA and from the wild-type parabionts with CAIA. Furthermore, normal and CAIA synovial tissues from Col1a2-GFP mice and from fluorescent ubiquitination-based cell cycle indicator (Fucci) transgenic mice, in which cells in S/G{sub 2}/M phases of the cell cycle express Azami-Green, were studied for Ki67, a cellular proliferation marker, and vimentin, a fibroblast marker, expression. The percentages of Ki67+/GFP+ and Azami-Green+/vimentin+ cells in the CAIA synovial tissues were higher than those in the untreated synovial tissues (34% vs. 0.40% and 19% vs. 0.26%, respectively). These findings indicate that local fibroblast proliferation but not cellular influx is responsible for the synovial hyperplasia in CAIA. Suppression of proliferation of the local synovial fibroblasts should be a promising treatment for RA. - Highlights: • We studied how synovial fibroblasts accumulate in joints in a murine model of RA. • Bone marrow-derived cells did not accumulate in arthritic joints. • Synovial fibroblasts did not accumulate in arthritic joints via

  3. Immunohistochemical localization of basic fibroblast growth factor within the mouse uterus.

    Science.gov (United States)

    Wordinger, R J; Moss, A E; Lockard, T; Gray, D; Chang, I F; Jackson, T L

    1992-09-01

    Uterine samples were either rapidly frozen in liquid nitrogen or placed in Bouin's fixative. A commercial primary polyclonal antibody made in rabbits against human recombinant basic fibroblast growth factor (bFGF) was used. Western blot analysis indicated that the antibody was specific for bFGF and did not react with acidic FGF. The primary antibody was followed by either goat anti-rabbit immunoglobulin G (IgG) conjugated to the fluorescent phycobiliprotein tracer phycoerythrin or biotinylated goat anti-rabbit IgG and a biotin-avidin-peroxidase complex. Specificity controls using adjacent sections were carried out by (i) substituting normal rabbit sera for the primary antisera, (ii) omitting the primary antisera or (iii) extracting sections with NaCl (2 mol l-1) prior to the immunochemical procedures. No binding of the antibody was observed with any of the specificity control sections. The connective tissue stroma and the basal lamina associated with uterine glandular and surface epithelial layers were positive for bFGF. Localization was not observed within surface or glandular epithelial cells. The basal lamina and endothelial cells associated with blood vessels within the uterus and the smooth muscle cells of the myometrium were positive for bFGF. There were no differences in uterine localization patterns or intensity during the oestrous cycle or after ovariectomy and steroid hormone supplementation. These studies demonstrate the specific localization of bFGF within the mouse uterus.

  4. Distribution and localization of fibroblast growth factor-8 in rat brain and nerve cells during neural stem/progenitor cell differentiation.

    Science.gov (United States)

    Lu, Jiang; Li, Dongsheng; Lu, Kehuan

    2012-07-05

    The present study explored the distribution and localization of fibroblast growth factor-8 and its potential receptor, fibroblast growth factor receptor-3, in adult rat brain in vivo and in nerve cells during differentiation of neural stem/progenitor cells in vitro. Immunohistochemistry was used to examine the distribution of fibroblast growth factor-8 in adult rat brain in vivo. Localization of fibroblast growth factor-8 and fibroblast growth factor receptor-3 in cells during neural stem/progenitor cell differentiation in vitro was detected by immunofluorescence. Flow cytometry and immunofluorescence were used to evaluate the effect of an anti-fibroblast growth factor-8 antibody on neural stem/progenitor cell differentiation and expansion in vitro. Results from this study confirmed that fibroblast growth factor-8 was mainly distributed in adult midbrain, namely the substantia nigra, compact part, dorsal tier, substantia nigra and reticular part, but was not detected in the forebrain comprising the caudate putamen and striatum. Unusual results were obtained in retrosplenial locations of adult rat brain. We found that fibroblast growth factor-8 and fibroblast growth factor receptor-3 were distributed on the cell membrane and in the cytoplasm of nerve cells using immunohistochemistry and immunofluorescence analyses. We considered that the distribution of fibroblast growth factor-8 and fibroblast growth factor receptor-3 in neural cells corresponded to the characteristics of fibroblast growth factor-8, a secretory factor. Addition of an anti-fibroblast growth factor-8 antibody to cultures significantly affected the rate of expansion and differentiation of neural stem/progenitor cells. In contrast, addition of recombinant fibroblast growth factor-8 to differentiation medium promoted neural stem/progenitor cell differentiation and increased the final yields of dopaminergic neurons and total neurons. Our study may help delineate the important roles of fibroblast growth

  5. Local induction of pacemaking activity in a monolayer of electrically coupled quiescent NRK fibroblasts

    NARCIS (Netherlands)

    Dernison, M.M.; Kusters, J.M.A.M.; Peters, P.H.J.; Meerwijk, W.P. van; Ypey, D.L.; Gielen, C.C.A.M.; Zoelen, E.J.J. van; Theuvenet, A.P.R.

    2008-01-01

    Cultures of normal rat kidney (NRK) fibroblasts may display spontaneous calcium action potentials which propagate throughout the cellular monolayer. Pacemaking activity of NRK cells was studied by patch clamp electrophysiology and vital calcium imaging, using a new experimental approach in which a

  6. Corneal Fibroblasts as Sentinel Cells and Local Immune Modulators in Infectious Keratitis

    Directory of Open Access Journals (Sweden)

    Ken Fukuda

    2017-08-01

    Full Text Available The cornea serves as a barrier to protect the eye against external insults including microbial pathogens and antigens. Bacterial infection of the cornea often results in corneal melting and scarring that can lead to severe visual impairment. Not only live bacteria but also their components such as lipopolysaccharide (LPS of Gram-negative bacteria contribute to the development of inflammation and subsequent corneal damage in infectious keratitis. We describe the important role played by corneal stromal fibroblasts (activated keratocytes as sentinel cells, immune modulators, and effector cells in infectious keratitis. Corneal fibroblasts sense bacterial infection through Toll-like receptor (TLR–mediated detection of a complex of LPS with soluble cluster of differentiation 14 (CD14 and LPS binding protein present in tear fluid. The cells then initiate innate immune responses including the expression of chemokines and adhesion molecules that promote the recruitment of inflammatory cells necessary for elimination of the infecting bacteria. Infiltrated neutrophils are activated by corneal stromal collagen and release mediators that stimulate the production of pro–matrix metalloproteinases by corneal fibroblasts. Elastase produced by Pseudomonas aeruginosa (P. aeruginosa activates these released metalloproteinases, resulting in the degradation of stromal collagen. The modulation of corneal fibroblast activation and of the interaction of these cells with inflammatory cells and bacteria is thus important to minimize corneal scarring during treatment of infectious keratitis. Pharmacological agents that are able to restrain such activities of corneal fibroblasts without allowing bacterial growth represent a potential novel treatment option for prevention of excessive scarring and tissue destruction in the cornea.

  7. Synthesis and intracellular localization of chick acid alpha-glucosidase in chick erythrocyte-human fibroblast heterokaryons. A model system for the study of lysosomal enzyme synthesis.

    Science.gov (United States)

    Sips, H J; Reuser, A J; van der Veer, E

    1986-02-01

    The synthesis and localization of chick acid alpha-glucosidase has been studied in chick erythrocyte-human fibroblast heterokaryons. Monospecific antibodies raised against purified chick liver acid alpha-glucosidase were used. It was found that the acid alpha-glucosidase in the heterokaryons is of chick origin, and is localized in the same lysosomes as the human lysosomal enzymes. It is concluded that chick erythrocyte-human fibroblast heterokaryons provide a useful model system for the study of lysosomal enzyme synthesis and routing.

  8. Fibroblastic rheumatism

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    Jyoti Ranjan Parida

    2017-01-01

    Full Text Available Fibroblastic rheumatism (FR is a rare dermoarthopathy reported from different parts of the world since 1980. Although the exact cause is unknown, few reports implicate infection may be a triggering event. Patients usually present with multiple skin nodules and polyarthropathy with progressive skin contractures. Laboratory parameters including acute phase reactants are usually normal. The confirmatory diagnosis is based on histopathologic study of skin nodules, which demonstrate fibroblastic proliferation, thickened collagen fibers, dermal fibrosis, and decreased number of elastic fibers. Immunoreactivity for b-catenin, smooth muscle actin, and the monoclonal antibody HHF35 show myofibroblastic differentiation. Treatments with oral prednisolone and other disease-modifying drugs such as methotrexate, infliximab, and interferon have been tried with variable success. In general, skin lesions respond more aptly than joint symptoms indicating that skin fibroblast is more amenable to treatment than synovial fibroblasts. Awareness regarding this orphan disease among clinicians and pathologists will help in more reporting of such cases and finding out optimal treatment regimen.

  9. Co-localization and regulation of basic fibroblast growth factor and arginine vasopressin in neuroendocrine cells of the rat and human brain

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    Gonzalez Ana M

    2010-08-01

    Full Text Available Abstract Background Adult rat hypothalamo-pituitary axis and choroid plexus are rich in basic fibroblast growth factor (FGF2 which likely has a role in fluid homeostasis. Towards this end, we characterized the distribution and modulation of FGF2 in the human and rat central nervous system. To ascertain a functional link between arginine vasopressin (AVP and FGF2, a rat model of chronic dehydration was used to test the hypothesis that FGF2 expression, like that of AVP, is altered by perturbed fluid balance. Methods Immunohistochemistry and confocal microscopy were used to examine the distribution of FGF2 and AVP neuropeptides in the normal human brain. In order to assess effects of chronic dehydration, Sprague-Dawley rats were water deprived for 3 days. AVP neuropeptide expression and changes in FGF2 distribution in the brain, neural lobe of the pituitary and kidney were assessed by immunohistochemistry, and western blotting (FGF2 isoforms. Results In human hypothalamus, FGF2 and AVP were co-localized in the cytoplasm of supraoptic and paraventricular magnocellular neurons and axonal processes. Immunoreactive FGF2 was associated with small granular structures distributed throughout neuronal cytoplasm. Neurohypophysial FGF2 immunostaining was found in axonal processes, pituicytes and Herring bodies. Following chronic dehydration in rats, there was substantially-enhanced FGF2 staining in basement membranes underlying blood vessels, pituicytes and other glia. This accompanied remodeling of extracellular matrix. Western blot data revealed that dehydration increased expression of the hypothalamic FGF2 isoforms of ca. 18, 23 and 24 kDa. In lateral ventricle choroid plexus of dehydrated rats, FGF2 expression was augmented in the epithelium (Ab773 as immunomarker but reduced interstitially (Ab106 immunostaining. Conclusions Dehydration altered FGF2 expression patterns in AVP-containing magnocellular neurons and neurohypophysis, as well as in choroid

  10. Fibroblast growth factor 2 is of prognostic value for patients with locally advanced squamous cell carcinoma of the head and neck

    Energy Technology Data Exchange (ETDEWEB)

    Rades, D.; Seibold, N.D. [University of Luebeck, Department of Radiation Oncology, Luebeck (Germany); Gebhard, M.P.; Noack, F. [University of Luebeck, Institute of Pathology, Luebeck (Germany); Schild, S.E. [Mayo Clinic Scottsdale, Department of Radiation Oncology, Scottsdale (United States)

    2014-01-15

    Patients with locally advanced SCCHN have a poor prognosis. This study investigated the prognostic value of the tumor cell expression of the fibroblast growth factor 2 (FGF-2) in patients treated with surgery followed by radiotherapy. The impact of FGF-2-expression and 11 additional potential prognostic factors on loco-regional control (LRC), metastases-free survival (MFS), and overall survival (OS) was retrospectively evaluated in 146 patients. Additional factors included age, gender, performance status, pre-radiotherapy hemoglobin levels, tumor site, histologic grade, T-category, N-category, human papilloma virus (HPV) status, extent of resection, and chemotherapy. Univariate analyses were performed with the Kaplan-Meier method and the log-rank test, multivariate analyses with the Cox proportional hazard model. On multivariate analysis, improved LRC was significantly associated with FGF-2-negativity [risk ratio (RR): 7.33; 95 %-confidence interval (CI): 2.88-19.05; p < 0.001], lower T-category (RR: 2.42; 95 %-CI: 1.47-4.33; p < 0.001), lower N-category (RR: 12.36; 95 %-CI: 3.48-78.91; p < 0.001), and pre-radiotherapy hemoglobin levels ≥12 g/dl (RR: 4.18; 95 %-CI: 1.73-10.53; p = 0.002). No factor was significantly associated with improved MFS. Lower T-category showed a trend (RR: 1.59; 95 %-CI: 0.97-2.82; p = 0.069). Better OS was significantly associated with FGF-2-negativity (RR: 5.10; 2.22-11.80; p < 0.001), lower T-category (RR: 2.17; 95 %-CI: 1.38-3.68; p < 0.001), lower N-category (RR: 3.86; 95 %-CI: 1.60-10.85; p = 0.002), and pre-radiotherapy hemoglobin levels ≥12 g/dl (RR: 3.20; 95 %-CI: 1.46-7.30; p = 0.004). HPV-positivity showed a trend (RR: 2.36; 95 %-CI: n.a.; p = 0.054). Tumor cell expression of FGF-2 proved to be an independent prognostic factor for LRC and OS. This factor can help personalize treatment and stratify patients in future trials. (orig.)

  11. Localization of DNA probes with tight linkage to the cystic fibrosis locus by in situ hybridization using fibroblasts with a 7q22 deletion

    NARCIS (Netherlands)

    van der Hout, A. H.; van der Veen, A. Y.; Aten, J. A.; Buys, C. H.

    1988-01-01

    Five DNA probes known to originate from the region 7q22-q31 were sublocalized by in situ hybridization to metaphase preparations of fibroblasts having besides a normal chromosome 7, a homologue 7 with an apparent interstitial deletion of a large part of band q22. A flow cytometric chromosome

  12. Endothelial cells, fibroblasts and vasculitis.

    Science.gov (United States)

    Buckley, Christopher D; Rainger, G Ed; Nash, Gerard B; Raza, Karim

    2005-07-01

    One of the most important questions in vasculitis research is not why inflammation of blood vessels occurs but why it persists, often in a site-specific manner. In this review we illustrate how stromal cells, such as fibroblasts and pericytes, might play an important role in regulating the site at which vasculitis occurs. Smooth muscle cells and fibroblasts directly influence the behaviour of overlying vascular cells, amplifying the response of the endothelium to proinflammatory agents such as TNF-alpha and allowing enhanced and inappropriate leucocyte recruitment. An abnormal local vascular stromal environment can therefore influence local endothelial function and drive the persistence of local vascular inflammation. However, such local vascular inflammation can have distant effects on the systemic vascular system, leading to widespread endothelial cell dysfunction. Vascular endothelial dysfunction is common in a range of immune-mediated inflammatory diseases, is seen in multiple vascular beds, and is reversible following the induction of disease remission. The mechanisms that drive such systemic vascular endothelial dysfunction are unclear but factors such as TNF-alpha and CRP may play a role. Persistence of such widespread endothelial dysfunction in systemic vasculitis appears to have long-term consequences, leading to the acceleration of atherosclerosis and premature ischaemic heart disease. It may also underlie the accelerated atherosclerosis seen in other immune-mediated rheumatic diseases, such as rheumatoid arthritis.

  13. Tumor-secreted LOXL2 activates fibroblasts through FAK signaling

    DEFF Research Database (Denmark)

    Barker, Holly E; Bird, Demelza; Lang, Georgina

    2013-01-01

    models. Here, we discovered that tumor-derived LOXL2 directly activated stromal fibroblasts in the tumor microenvironment. Genetic manipulation or antibody inhibition of LOXL2 in orthotopically grown mammary tumors reduced the expression of α-smooth muscle actin (α-SMA). Using a marker for reticular...... fibroblasts, it was determined that expression of α-SMA was localized to fibroblasts recruited from the host tissue. This marker also revealed that the matrix present in tumors with reduced levels of LOXL2 was more scattered compared with control tumors which exhibited matrices with dense, parallel alignments....... Importantly, in vitro assays revealed that tumor-derived LOXL2 and a recombinant LOXL2 protein induced fibroblast branching on collagen matrices, as well as increased fibroblast-mediated collagen contraction and invasion of fibroblasts through extracellular matrix. Moreover, LOXL2 induced the expression of α-SMA...

  14. Cell-cycle-dependent localization of human cytomegalovirus UL83 phosphoprotein in the nucleolus and modulation of viral gene expression in human embryo fibroblasts in vitro.

    Science.gov (United States)

    Arcangeletti, Maria-Cristina; Rodighiero, Isabella; Mirandola, Prisco; De Conto, Flora; Covan, Silvia; Germini, Diego; Razin, Sergey; Dettori, Giuseppe; Chezzi, Carlo

    2011-01-01

    The nucleolus is a multifunctional nuclear compartment widely known to be involved in several cellular processes, including mRNA maturation and shuttling to cytoplasmic sites, control of the cell cycle, cell proliferation, and apoptosis; thus, it is logical that many viruses, including herpesvirus, target the nucleolus in order to exploit at least one of the above-mentioned functions. Recent studies from our group demonstrated the early accumulation of the incoming ppUL83 (pp65), the major tegument protein of human cytomegalovirus (HCMV), in the nucleolus. The obtained results also suggested that a functional relationship might exist between the nucleolar localization of pp65, rRNA synthesis, and the development of the lytic program of viral gene expression. Here we present new data which support the hypothesis of a potentially relevant role of HCMV pp65 and its nucleolar localization for the control of the cell cycle by HCMV (arrest of cell proliferation in G1-G1/S), and for the promotion of viral infection. We demonstrated that, although the incoming pp65 amount in the infected cells appears to be constant irrespective of the cell-cycle phase, its nucleolar accumulation is prominent in G1 and G1/S, but very poor in S or G2/M. This correlates with the observation that only cells in G1 and G1/S support an efficient development of the HCMV lytic cycle. We propose that HCMV pp65 might be involved in regulatory/signaling pathways related to nucleolar functions, such as the cell-cycle control. Co-immunoprecipitation experiments have permitted to identify nucleolin as one of the nucleolar partners of pp65.

  15. Fibroblasts in myocardial infarction: a role in inflammation and repair

    Science.gov (United States)

    Shinde, Arti V.; Frangogiannis, Nikolaos G.

    2014-01-01

    Fibroblasts do not only serve as matrix-producing reparative cells, but exhibit a wide range of functions in inflammatory and immune responses, angiogenesis and neoplasia. The adult mammalian myocardium contains abundant fibroblasts enmeshed within the interstitial and perivascular extracellular matrix. The current review manuscript discusses the dynamic phenotypic and functional alterations of cardiac fibroblasts following myocardial infarction. Extensive necrosis of cardiomyocytes in the infarcted heart triggers an intense inflammatory reaction. In the early stages of infarct healing, fibroblasts become pro-inflammatory cells, activating the inflammasome and producing cytokines, chemokines and proteases. Pro-inflammatory cytokines (such as Interleukin-1) delay myofibroblast transformation, until the wound is cleared from dead cells and matrix debris. Resolution of the inflammatory infiltrate is associated with fibroblast migration, proliferation, matrix protein synthesis and myofibroblast conversion. Growth factors and matricellular proteins play an important role in myofibroblast activation during the proliferative phase of healing. Formation of a mature cross-linked scar is associated with clearance of fibroblasts, as poorly-understood inhibitory signals restrain the fibrotic response. However, in the non-infarcted remodeling myocardium, local fibroblasts may remain activated in response to volume and pressure overload and may promote interstitial fibrosis. Considering their abundance, their crucial role in cardiac inflammation and repair, and their involvement in myocardial dysfunction and arrhythmogenesis, cardiac fibroblasts may be key therapeutic targets in cardiac remodeling. PMID:24321195

  16. Differential expression of wound fibrotic factors between facial and trunk dermal fibroblasts.

    Science.gov (United States)

    Kurita, Masakazu; Okazaki, Mutsumi; Kaminishi-Tanikawa, Akiko; Niikura, Mamoru; Takushima, Akihiko; Harii, Kiyonori

    2012-01-01

    Clinically, wounds on the face tend to heal with less scarring than those on the trunk, but the causes of this difference have not been clarified. Fibroblasts obtained from different parts of the body are known to show different properties. To investigate whether the characteristic properties of facial and trunk wound healing are caused by differences in local fibroblasts, we comparatively analyzed the functional properties of superficial and deep dermal fibroblasts obtained from the facial and trunk skin of seven individuals, with an emphasis on tendency for fibrosis. Proliferation kinetics and mRNA and protein expression of 11 fibrosis-associated factors were investigated. The proliferation kinetics of facial and trunk fibroblasts were identical, but the expression and production levels of profibrotic factors, such as extracellular matrix, transforming growth factor-β1, and connective tissue growth factor mRNA, were lower in facial fibroblasts when compared with trunk fibroblasts, while the expression of antifibrotic factors, such as collagenase, basic fibroblast growth factor, and hepatocyte growth factor, showed no clear trends. The differences in functional properties of facial and trunk dermal fibroblasts were consistent with the clinical tendencies of healing of facial and trunk wounds. Thus, the differences between facial and trunk scarring are at least partly related to the intrinsic nature of the local dermal fibroblasts.

  17. [Fibroblast growth factor-2].

    Science.gov (United States)

    Faitová, J

    2004-01-01

    Fibroblast growth factor-2 is a member of a large family of proteins that bind heparin and heparan sulfate and modulate the function of a wide range of cell types. FGF-2 occurs in several isoforms resulting from alternative initiations of traslation: an 18 kDa cytoplasmic isoform and four larger molecular weight nuclear isoforms (22, 22.5, 24 and 34 kDa). It acts mainly through a paracrine/autocrine mechanism involving high affinity transmembrane receptors and heparan sulfate proteoglycan low affinity receptors. It is expressed mostly in tissues of mesoderm and neuroectoderm origin, and plays an important role in mesoderm induction, stimulates the growth and development of the new blood vessels (angiogenesis), normal wound healing and tissue development. FGF-2 positively regulates hematopoiesis by acting on various cellular targets: stromal cells, early and committed hematopoietic progenitors and possibly some mature blood cells. FGF-2 is a potent hematopoietic growth factor that is likely to play an important role in physiological and pathological hematopoiesis.

  18. Endothelial cells, fibroblasts and vasculitis

    OpenAIRE

    Buckley, Christopher D.; Rainger, G.Ed; Nash, Gerard B; Raza, Karim

    2005-01-01

    One of the most important questions in vasculitis research is not why inflammation of blood vessels occurs but why it persists, often in a site-specific manner. In this review we illustrate how stromal cells, such as fibroblasts and pericytes, might play an important role in regulating the site at which vasculitis occurs. Smooth muscle cells and fibroblasts directly influence the behaviour of overlying vascular cells, amplifying the response of the endothelium to proinflammatory agents such a...

  19. Cancer associated fibroblasts (CAFs) are activated in cutaneous basal cell carcinoma and in the peritumoural skin

    DEFF Research Database (Denmark)

    Omland, Silje Haukali; Wettergren, Erika Elgstrand; Mourier, Tobias

    2017-01-01

    Background: Cutaneous basal cell carcinoma (BCC) is the commonest cancer worldwide. BCC is locally invasive and the surrounding stromal microenvironment is pivotal for tumourigenesis. Cancer associated fibroblasts (CAFs) in the microenvironment are essential for tumour growth in a variety...

  20. Cancer associated fibroblasts (CAFs) are activated in cutaneous basal cell carcinoma and in the peritumoural skin

    DEFF Research Database (Denmark)

    Omland, Silje Haukali; Wettergren, Erika Elgstrand; Mollerup, Sarah

    2017-01-01

    BACKGROUND: Cutaneous basal cell carcinoma (BCC) is the commonest cancer worldwide. BCC is locally invasive and the surrounding stromal microenvironment is pivotal for tumourigenesis. Cancer associated fibroblasts (CAFs) in the microenvironment are essential for tumour growth in a variety...

  1. Preparation of extracellular matrices produced by cultured and primary fibroblasts

    Science.gov (United States)

    Franco-Barraza, Janusz; Beacham, Dorothy A.; Amatangelo, Michael D.; Cukierman, Edna

    2016-01-01

    matrix (Support Protocol 3), the acquisition of an in vivo-like spindle-shaped morphology can also be measured (Support Protocol 4). To ascertain whether fibroblasts respond to the 3-D microenvironment when plated within specific (NIH-3T3) matrices, the phosphorylation level of non-receptor focal adhesion kinase (FAK) pY397 can be quantified by immunoblotting (see Support Protocol 6). In addition, the ability of tumor- or cancer-associated fibroblast-derived matrices to induce and maintain an active (i.e., desmoplastic) phenotype in quiescent naïve fibroblasts can be queried by calculating levels (via western blot and/or semi-quantitative immunofluorescence) and localization (using immunofluorescence) of myofibroblastic markers such as alpha-smooth muscle actin (see Support Protocols 5 and 6) This unit will also describe how to mechanically compress the fibroblast-derived 3-D matrices to obtain 2-D substrate controls (see Support Protocol 7). Moreover, a support protocol will illustrate how to solubilize the fibroblast-derived 3-D matrices to produce a matrix-derived protein mixture for additional 2-D coating controls and for subsequent biochemical analysis of the matrices (see Support Protocol 8 and Commentary). Lastly, there are two support protocols designed for the isolation of primary fibroblasts from fresh tissue samples to produce additional types of fibroblast-derived 3-D matrices (see Support Protocols 9 and 10). PMID:27245425

  2. Case report 511: Fibroblastic rheumatism

    Energy Technology Data Exchange (ETDEWEB)

    Hernandez, R.J.; Martel, W.; Headington, J.T.; Kaufman, R.A.

    1989-03-01

    We report a ten-year-old child with the newly described entity of fibroblastic rheumatism. This child developed rapid, progressive, symmetrical polyarthritis, similar to the radiographic appearance of juvenile rheumatoid arthritis, except for the rapidity of progression. The polyarthritis was preceded by the development of skin nodules with characteristic histological changes. (orig./GDG).

  3. Increased susceptibility of spinal muscular atrophy fibroblasts to camptothecin is p53-independent

    Directory of Open Access Journals (Sweden)

    Funanage Vicky L

    2009-05-01

    Full Text Available Abstract Background Deletion or mutation(s of the survival motor neuron 1 (SMN1 gene causes spinal muscular atrophy (SMA. The SMN protein is known to play a role in RNA metabolism, neurite outgrowth, and cell survival. Yet, it remains unclear how SMN deficiency causes selective motor neuron death and muscle atrophy seen in SMA. Previously, we have shown that skin fibroblasts from SMA patients are more sensitive to the DNA topoisomerase I inhibitor camptothecin, supporting a role for SMN in cell survival. Here, we examine the potential mechanism of camptothecin sensitivity in SMA fibroblasts. Results Camptothecin treatment reduced the DNA relaxation activity of DNA topoisomerase I in human fibroblasts. In contrast, kinase activity of DNA topoisomerase I was not affected by camptothecin, because levels of phosphorylated SR proteins were not decreased. Upon camptothecin treatment, levels of p53 were markedly increased. To determine if p53 plays a role in the increased sensitivity of SMA fibroblasts to camptothecin, we analyzed the sensitivity of SMA fibroblasts to another DNA topoisomerase I inhibitor, β-lapachone. This compound is known to induce death via a p53-independent pathway in several cancer cell lines. We found that β-lapachone did not induce p53 activation in human fibroblasts. In addition, SMA and control fibroblasts showed essentially identical sensitivity to this compound. By immunofluorescence staining, SMN and p53 co-localized in gems within the nucleus, and this co-localization was overall reduced in SMA fibroblasts. However, depletion of p53 by siRNA did not lessen the camptothecin sensitivity in SMA fibroblasts. Conclusion Even though p53 and SMN are associated, the increased sensitivity of SMA fibroblasts to camptothecin does not occur through a p53-dependent mechanism.

  4. Endoglin negatively regulates transforming growth factor beta1-induced profibrotic responses in intestinal fibroblasts.

    LENUS (Irish Health Repository)

    Burke, J P

    2012-02-01

    BACKGROUND: Fibroblasts isolated from strictures in Crohn\\'s disease (CD) exhibit reduced responsiveness to stimulation with transforming growth factor (TGF) beta1. TGF-beta1, acting through the smad pathway, is critical to fibroblast-mediated intestinal fibrosis. The membrane glycoprotein, endoglin, is a negative regulator of TGF-beta1. METHODS: Intestinal fibroblasts were cultured from seromuscular biopsies of patients undergoing intestinal resection for CD strictures or from control patients. Endoglin expression was assessed using confocal microscopy, flow cytometry and western blot. The effect of small interfering (si) RNA-mediated knockdown and plasmid-mediated overexpression of endoglin on fibroblast responsiveness to TGF-beta1 was assessed by examining smad phosphorylation, smad binding element (SBE) promoter activity, connective tissue growth factor (CTGF) expression and ability to contract collagen. RESULTS: Crohn\\'s stricture fibroblasts expressed increased constitutive cell-surface and whole-cell endoglin relative to control cells. Endoglin co-localized with filamentous actin. Fibroblasts treated with siRNA directed against endoglin exhibited enhanced TGF-beta1-mediated smad-3 phosphorylation, and collagen contraction. Cells transfected with an endoglin plasmid did not respond to TGF-beta1 by exhibiting SBE promoter activity or producing CTGF. CONCLUSION: Fibroblasts from strictures in CD express increased constitutive endoglin. Endoglin is a negative regulator of TGF-beta1 signalling in the intestinal fibroblast, modulating smad-3 phosphorylation, SBE promoter activity, CTGF production and collagen contraction.

  5. Podoplanin increases the migration of human fibroblasts and affects the endothelial cell network formation: A possible role for cancer-associated fibroblasts in breast cancer progression.

    Science.gov (United States)

    Suchanski, Jaroslaw; Tejchman, Anna; Zacharski, Maciej; Piotrowska, Aleksandra; Grzegrzolka, Jedrzej; Chodaczek, Grzegorz; Nowinska, Katarzyna; Rys, Janusz; Dziegiel, Piotr; Kieda, Claudine; Ugorski, Maciej

    2017-01-01

    podoplanin-mediated effects can affect tube formation by endothelial cells and participate in their pathological properties in the tumor context. Our experimental data were supported by clinical studies. First, when IDC and DCIS were analyzed by immunohistochemistry according to the presence of podoplanin-expressing cells, the numbers of cancer-associated fibroblasts with high expression of this glycoprotein were significantly higher in IDC than in DCIS cases. Second, using immunofluorescence, the co-localization of PDPN-positive CAFs with blood vessels stained with antibody directed against CD34 was observed in tumor stroma of IDC samples.

  6. Podoplanin increases the migration of human fibroblasts and affects the endothelial cell network formation: A possible role for cancer-associated fibroblasts in breast cancer progression.

    Directory of Open Access Journals (Sweden)

    Jaroslaw Suchanski

    time, that such podoplanin-mediated effects can affect tube formation by endothelial cells and participate in their pathological properties in the tumor context. Our experimental data were supported by clinical studies. First, when IDC and DCIS were analyzed by immunohistochemistry according to the presence of podoplanin-expressing cells, the numbers of cancer-associated fibroblasts with high expression of this glycoprotein were significantly higher in IDC than in DCIS cases. Second, using immunofluorescence, the co-localization of PDPN-positive CAFs with blood vessels stained with antibody directed against CD34 was observed in tumor stroma of IDC samples.

  7. AGE AND MULTIPLICATION OF FIBROBLASTS.

    Science.gov (United States)

    Carrel, A; Ebeling, A H

    1921-11-30

    Pure cultures of fibroblasts displayed marked differences in their activity in the plasma of young, middle aged, and old chickens. The rate of cell multiplication varied in inverse ratio to the age of the animal from which the plasma was taken. There was a definite relation between the age of the animal and the amount of new tissue produced in its plasma in a given time (Text-figs. 1 to 10). The chart obtained by plotting the rate of cell proliferation in ordinates, and the age of the animal in abscissae, showed that the rate of growth decreased more quickly than the age increased (Text-fig. 12). The decrease in the rate of growth was 50 per cent during the first 3 years of life, while in the following 6 years it was only 30 per cent. When the duration of the life of the cultures in the four plasmas was compared, a curve was obtained which showed about the same characteristics (Text-fig. 11). The duration of life of the fibroblasts in vitro varied in inverse ratio to the age of the animal, and decreased more quickly than the age increased. As the differences in the amount of new tissue produced in the plasma of young, middle aged, and old chickens were large, the growth of a pure culture of fibroblasts could be employed as a reagent for detecting certain changes occurring in the plasma under the influence of age. But the method possesses the necessary accuracy only when it is used as has already been described, and by technicians thoroughly trained in the details of its application. A comparative study of the growth of fibroblasts in media containing no serum, and serum under low and high concentrations, was made in order to ascertain whether the decreasing rate of cell multiplication was due to the loss of an accelerating factor, or to the increase See PDF for Structure of an inhibiting one. In high and low concentrations of the serum of young animals, no difference in the rate of multiplication of fibroblasts was observed. This showed that the serum of an

  8. mRNA encoding WAVE-Arp2/3-associated proteins is co-localized with foci of active protein synthesis at the leading edge of MRC5 fibroblasts during cell migration.

    Science.gov (United States)

    Willett, Mark; Brocard, Michele; Pollard, Hilary J; Morley, Simon J

    2013-05-15

    During cell spreading, mammalian cells migrate using lamellipodia formed from a large dense branched actin network which produces the protrusive force required for leading edge advancement. The formation of lamellipodia is a dynamic process and is dependent on a variety of protein cofactors that mediate their local regulation, structural characteristics and dynamics. In the present study, we show that mRNAs encoding some structural and regulatory components of the WAVE [WASP (Wiskott-Aldrich syndrome protein) verprolin homologous] complex are localized to the leading edge of the cell and associated with sites of active translation. Furthermore, we demonstrate that steady-state levels of ArpC2 and Rac1 proteins increase at the leading edge during cell spreading, suggesting that localized protein synthesis has a pivotal role in controlling cell spreading and migration.

  9. Caspase 3 activity in isolated fetal rat lung fibroblasts and rat periodontal ligament fibroblasts: cigarette smoke-induced alterations

    Directory of Open Access Journals (Sweden)

    James Elliot Scott

    2016-03-01

    Full Text Available Background Cigarette smoking is the leading cause of preventable death in the world. It has been implicated in the pathogenesis of pulmonary, oral and systemic diseases. Smoking during pregnancy is clearly a risk factor for the developing fetus and may be a major cause of infant mortality. Moreover, the oral cavity is the first site of exposure to cigarette smoke and may be a possible source for the spread of toxins to other organs of the body. Fibroblasts in general are morphologically heterogeneous connective tissue cells with diverse functions. Apoptosis or programmed cell death is a crucial process during embryogenesis and for the maintenance of homeostasis throughout life. Deregulation of apoptosis has been implicated in abnormal lung development in the fetus and disease progression in adults. Caspases, are proteases which belong to the family of cysteine aspartic acid proteases and are the key components for the downstream amplification of intra-cellular apoptotic signals. Of the 14 caspases known, caspase-3 is the key executioner of apoptosis. Fetal rat lung fibroblasts but not PDL viability is reduced by exposure to CSE. In addition Caspase 3 activity is elevated after CSE exposure in fetal lung fibroblasts but not in PDLs. Expression of caspase 3 is induced in CSE exposed lung fibroblasts but not in PDLs. Caspase 3 was localized to the cytoplasm in both cell types.

  10. Recombinant interferon regulation of lung fibroblast growth

    Energy Technology Data Exchange (ETDEWEB)

    Elias, J.A.; Freundlich, B.; Rossman, M.D.; Gustilo, K.; Jimenez, S.A.

    1986-03-01

    The authors characterized the effect on human lung fibroblast proliferation of recombinant interferons (gamma (..gamma..), alpha/sub A/ (..cap alpha../sub A/), alpha/sub D/ (..cap alpha../sub D/) and beta (..beta..)). Quiescent and log phase fibroblasts were used. Proliferation was assessed using /sup 3/HTdr uptake and cell counts. ..gamma.. stimulated the /sup 3/HTdr uptake of quiescent cells. Maximal stimulation was seen between 100 and 500 IU/ml. Fibroblasts incubated in 100 IU/ml incorporated 160 +/- 9% as much /sup 3/HTdr as quiescent controls (p < .04). In contrast, ..gamma.. caused a dose dependent inhibition of the /sup 3/HTdr uptake of log phase cells. Fibroblasts incubated in 1000 IU/ml incorporated 41 +/- 9% as much /sup 3/HTdr as log phase controls (p < .02). Unlike ..gamma.., ..cap alpha../sub A/, ..cap alpha../sub D/ and ..beta.. did not stimulate the proliferation of quiescent cells. However, they did inhibit the proliferation of log phase cells. Fibroblasts incubated in 1000 IU/ml of ..cap alpha../sub A/, ..cap alpha../sub D/ and ..beta.. incorporated 64 +/- 10, 81 +/- 4 and 62 +/- 7% as much /sup 3/HTdr as log phase controls (p < .03 for all). ..gamma.., ..cap alpha../sub A/, ..cap alpha../sub D/ and ..beta.. did not stimulate fibroblast prostaglandin E (PGE) production and incubating fibroblasts with indomethacin did not reverse the inhibition of fibroblast proliferation caused by the interferons. Thus, (1) ..gamma.. can stimulate or inhibit fibroblast growth, (2) ..cap alpha../sub A/, ..cap alpha../sub D/ and ..beta.. only inhibit fibroblast growth, (3) the inhibition of fibroblast growth caused by ..gamma.., ..cap alpha../sub A/, ..cap alpha../sub D/ and ..beta.. is largely independent of fibroblast PGE production.

  11. Apoptosis-Like Cell Death Induction and Aberrant Fibroblast Properties in Human Incisional Hernia Fascia

    Science.gov (United States)

    Diaz, Ramon; Quiles, Maria T.; Guillem-Marti, Jordi; Lopez-Cano, Manuel; Huguet, Pere; Ramon-y-Cajal, Santiago; Reventos, Jaume; Armengol, Manel; Arbos, Maria A.

    2011-01-01

    Incisional hernia often occurs following laparotomy and can be a source of serious problems. Although there is evidence that a biological cause may underlie its development, the mechanistic link between the local tissue microenvironment and tissue rupture is lacking. In this study, we used matched tissue-based and in vitro primary cell culture systems to examine the possible involvement of fascia fibroblasts in incisional hernia pathogenesis. Fascia biopsies were collected at surgery from incisional hernia patients and non-incisional hernia controls. Tissue samples were analyzed by histology and immunoblotting methods. Fascia primary fibroblast cultures were assessed at morphological, ultrastructural, and functional levels. We document tissue and fibroblast loss coupled to caspase-3 activation and induction of apoptosis-like cell-death mechanisms in incisional hernia fascia. Alterations in cytoskeleton organization and solubility were also observed. Incisional hernia fibroblasts showed a consistent phenotype throughout early passages in vitro, which was characterized by significantly enhanced cell proliferation and migration, reduced adhesion, and altered cytoskeleton properties, as compared to non-incisional hernia fibroblasts. Moreover, incisional hernia fibroblasts displayed morphological and ultrastructural alterations compatible with autophagic processes or lysosomal dysfunction, together with enhanced sensitivity to proapoptotic challenges. Overall, these data suggest an ongoing complex interplay of cell death induction, aberrant fibroblast function, and tissue loss in incisional hernia fascia, which may significantly contribute to altered matrix maintenance and tissue rupture in vivo. PMID:21641387

  12. Fibroblast spheroids as a model to study sustained fibroblast quiescence and their crosstalk with tumor cells

    Energy Technology Data Exchange (ETDEWEB)

    Salmenperä, Pertteli, E-mail: pertteli.salmenpera@helsinki.fi [Department of Virology, Medicum, Faculty of Medicine, University of Helsinki, P.O. Box 21, FIN-00014 (Finland); Karhemo, Piia-Riitta [Research Programs Unit, Translational Cancer Biology, and Institute of Biomedicine, University of Helsinki, P.O. Box 63, FIN-00014 (Finland); Räsänen, Kati [Department of Virology, Medicum, Faculty of Medicine, University of Helsinki, P.O. Box 21, FIN-00014 (Finland); Laakkonen, Pirjo [Research Programs Unit, Translational Cancer Biology, and Institute of Biomedicine, University of Helsinki, P.O. Box 63, FIN-00014 (Finland); Vaheri, Antti [Department of Virology, Medicum, Faculty of Medicine, University of Helsinki, P.O. Box 21, FIN-00014 (Finland)

    2016-07-01

    Stromal fibroblasts have an important role in regulating tumor progression. Normal and quiescent fibroblasts have been shown to restrict and control cancer cell growth, while cancer-associated, i. e. activated fibroblasts have been shown to enhance proliferation and metastasis of cancer cells. In this study we describe generation of quiescent fibroblasts in multicellular spheroids and their effects on squamous cell carcinoma (SCC) growth in soft-agarose and xenograft models. Quiescent phenotype of fibroblasts was determined by global down-regulation of expression of genes related to cell cycle and increased expression of p27. Interestingly, microarray analysis showed that fibroblast quiescence was associated with similar secretory phenotype as seen in senescence and they expressed senescence-associated-β-galactosidase. Quiescent fibroblasts spheroids also restricted the growth of RT3 SCC cells both in soft-agarose and xenograft models unlike proliferating fibroblasts. Restricted tumor growth was associated with marginally increased tumor cell senescence and cellular differentiation, showed with senescence-associated-β-galactosidase and cytokeratin 7 staining. Our results show that the fibroblasts spheroids can be used as a model to study cellular quiescence and their effects on cancer cell progression. - Highlights: • Fibroblasts acquire a sustained quiescence when grown as multicellular spheroids. • This quiescence is associated with drastic change in gene expression. • Fibroblasts spheroids secrete various inflammation-linked cytokines and chemokines. • Fibroblasts spheroids reduced growth of RT3 SCC cells in xenograft model.

  13. Expression and localization of fibroblast growth factor (FGF) family in buffalo ovarian follicle during different stages of development and modulatory role of FGF2 on steroidogenesis and survival of cultured buffalo granulosa cells.

    Science.gov (United States)

    Mishra, S R; Thakur, N; Somal, A; Parmar, M S; Reshma, R; Rajesh, G; Yadav, V P; Bharti, M K; Bharati, Jaya; Paul, A; Chouhan, V S; Sharma, G T; Singh, G; Sarkar, M

    2016-10-01

    The present study investigated the expression and localization of FGF and its functional receptors in the follicle of buffalo and the treatment of FGF2 on mRNA expression of CYP19A1 (aromatase), PCNA, and BAX (BCL-2 associated X protein) in cultured buffalo granulosa cells (GCs). Follicles were classified into four groups based on size and E2 level in follicular fluid (FF): F1, 4-6mm diameter, E214mm, E2>180ng/ml. The qPCR studies revealed that the mRNA expression of FGF1, FGF2 and FGF7 were maximum (P<0.05) in theca interna (TI) whereas the transcripts of FGFR1, FGFR2, FGFR2IIIB and FGFR2IIIC were up-regulated (P<0.05) in GCs of F4 follicles. Protein expression of most members were maximum (P<0.05) in F4 follicles except FGFR3 and FGFR4. All members were localized in GC and TI with a stage specific immunoreactivity. Primary culture of GCs with treatment of FGF2 at different dose-time combinations revealed that the mRNA expression and immunoreactivity of CYP19A1 and PCNA were maximum (P<0.05) whereas BAX was minimum (P<0.05) with 200ng/ml at 72h of incubation. The findings indicate that FGF family members are expressed in a regulated manner in buffalo ovarian follicles during different stages of development where FGF2 may promote steroidogenesis and GC survival through autocrine and paracrine manner. Copyright © 2016 Elsevier Ltd. All rights reserved.

  14. Altered fibroblast proteoglycan production in COPD.

    Science.gov (United States)

    Hallgren, Oskar; Nihlberg, Kristian; Dahlbäck, Magnus; Bjermer, Leif; Eriksson, Leif T; Erjefält, Jonas S; Löfdahl, Claes-Göran; Westergren-Thorsson, Gunilla

    2010-05-11

    Airway remodeling in COPD includes reorganization of the extracellular matrix. Proteoglycans play a crucial role in this process as regulators of the integrity of the extracellular matrix. Altered proteoglycan immunostaining has been demonstrated in COPD lungs and this has been suggested to contribute to the pathogenesis. The major cell type responsible for production and maintenance of ECM constituents, such as proteoglycans, are fibroblasts. Interestingly, it has been proposed that central airways and alveolar lung parenchyma contain distinct fibroblast populations. This study explores the hypothesis that altered depositions of proteoglycans in COPD lungs, and in particular versican and perlecan, is a result of dysregulated fibroblast proteoglycan production. Proliferation, proteoglycan production and the response to TGF-beta1 were examined in vitro in centrally and distally derived fibroblasts isolated from COPD patients (GOLD stage IV) and from control subjects. Phenotypically different fibroblast populations were identified in central airways and in the lung parenchyma. Versican production was higher in distal fibroblasts from COPD patients than from control subjects (p proteoglycan production in distally derived fibroblasts from COPD patients and control subjects. In contrast, centrally derived fibroblasts from COPD patients were less responsive to TGF-beta1 than those from control subjects. The results show that fibroblasts from COPD patients have alterations in proteoglycan production that may contribute to disease development. Distally derived fibroblasts from COPD patients have enhanced production of versican that may have a negative influence on the elastic recoil. In addition, a lower perlecan production in centrally derived fibroblasts from COPD patients may indicate alterations in bronchial basement membrane integrity in severe COPD.

  15. Altered fibroblast proteoglycan production in COPD

    Directory of Open Access Journals (Sweden)

    Erjefält Jonas S

    2010-05-01

    Full Text Available Abstract Background Airway remodeling in COPD includes reorganization of the extracellular matrix. Proteoglycans play a crucial role in this process as regulators of the integrity of the extracellular matrix. Altered proteoglycan immunostaining has been demonstrated in COPD lungs and this has been suggested to contribute to the pathogenesis. The major cell type responsible for production and maintenance of ECM constituents, such as proteoglycans, are fibroblasts. Interestingly, it has been proposed that central airways and alveolar lung parenchyma contain distinct fibroblast populations. This study explores the hypothesis that altered depositions of proteoglycans in COPD lungs, and in particular versican and perlecan, is a result of dysregulated fibroblast proteoglycan production. Methods Proliferation, proteoglycan production and the response to TGF-β1 were examined in vitro in centrally and distally derived fibroblasts isolated from COPD patients (GOLD stage IV and from control subjects. Results Phenotypically different fibroblast populations were identified in central airways and in the lung parenchyma. Versican production was higher in distal fibroblasts from COPD patients than from control subjects (p 1 triggered similar increases in proteoglycan production in distally derived fibroblasts from COPD patients and control subjects. In contrast, centrally derived fibroblasts from COPD patients were less responsive to TGF-β1 than those from control subjects. Conclusions The results show that fibroblasts from COPD patients have alterations in proteoglycan production that may contribute to disease development. Distally derived fibroblasts from COPD patients have enhanced production of versican that may have a negative influence on the elastic recoil. In addition, a lower perlecan production in centrally derived fibroblasts from COPD patients may indicate alterations in bronchial basement membrane integrity in severe COPD.

  16. Proteomic alterations of fibroblasts induced by ovarian cancer cells reveal potential cancer targets.

    Science.gov (United States)

    Zhang, X Y; Hong, S S; Zhang, M; Cai, Q Q; Zhang, M X; Xu, C J

    2017-08-31

    The common spread pattern of ovarian cancer is peritoneal implantation. The growth of the shed ovarian cancer cells in the peritoneal cavity is closely related to the tumor microenvironment. Cancer-associated fibroblasts are vital in the tumor microenvironment. It is not clearly defined that the protein expression alters during the activating process of fibroblasts. This study detected the protein alterations in fibroblasts induced by ovarian cancer cells and explored the potential biological relevance through two-dimensional gel electrophoresis and mass spectrometry. Our data showed that the level of CENPE, BAG2, SOD2, GDI2, CORO1C, CFL1, DSTN, CALD1, PHGDH, PDHA1, AKR1B1, TST and TBCA proteins were significantly up-regulated in the fibroblasts co-cultured with ovarian cancer cells, whereas HSPB1, P4HB and VIM were significantly down-regulated. However, only BAG2, SOD2 and CORO1C proteins were confirmed to be significantly increased by western blot analysis. The differentially expressed proteins were mainly involved in metabolic processes, cellular component organization, responses to stimulus, multicellular organismal processes, localization, protein depolymerization, cellular senescence and the mitotic pathway. These data demonstrated that fibroblasts had an altered protein expression pattern after being induced by ovarian cancer cells, and participated in multiple cell processes resulting in tumor progression. The differentially expressed proteins should be considered as targets for cancer treatment Keywords: ovarian neoplasms, fibroblast, stroma, biomarker, proteomics.

  17. Response of dupuytren fibroblasts to different oxygen environments.

    Science.gov (United States)

    Türker, Tolga; Murphy, Erin; Kaufman, Christina L; Kutz, Joseph E; Meister, Edward A; Hoying, James B

    2013-12-01

    It is thought that local ischemia and oxygen radicals are responsible for fibroblast-to-myofibroblast cell transformation and proliferation. We hypothesized that hypoxia could differentially activate the contractility of fibroblasts from normal human palmar fascia and from fibroblasts-myofibroblasts of Dupuytren cords. Normal palmar fascia from 5 patients with carpal tunnel syndrome and Dupuytren cords from 5 patients were harvested. Cells were cultured from all tissue samples, and collagen lattices were prepared containing these cells. Oxygen treatment subgroups were created and incubated under hypoxic (1% O(2), 5% CO(2), and 94% N(2)), normoxic (21% O(2), 5% CO(2), and 74% N(2)), and hyperoxic (100% oxygen using 2.4 atm pressure twice a day for 7 d) conditions. After 7 days, each subgroup was photographed, and lattices were released from dishes. Postrelease photographs were taken immediately, 5 minutes after release, and after 1 hour. Areas of the lattices at each time point were calculated using MetaMorph software. Actin staining and live/dead cell analysis was performed. Linear repeated measures analysis of variance was used for data analysis given that contraction levels were measured over 3 distinct time points. We found a statistically significant difference between normal samples and Dupuytren samples in mean contraction levels over time. There was no statistically significant difference between tissue groups over the 3 time periods based on the oxygen treatment received. Our results showed a greater degree of contractility in Dupuytren disease cells than normal fibroblasts. However, the contraction in either group was not affected by oxygen level. Future in vivo research is needed to better understand the nature of pathophysiology of Dupuytren disease. Copyright © 2013 American Society for Surgery of the Hand. Published by Elsevier Inc. All rights reserved.

  18. Spiral-Wave Dynamics in a Mathematical Model of Human Ventricular Tissue with Myocytes and Fibroblasts

    Science.gov (United States)

    Nayak, Alok Ranjan; Shajahan, T. K.; Panfilov, A. V.; Pandit, Rahul

    2013-01-01

    Cardiac fibroblasts, when coupled functionally with myocytes, can modulate the electrophysiological properties of cardiac tissue. We present systematic numerical studies of such modulation of electrophysiological properties in mathematical models for (a) single myocyte-fibroblast (MF) units and (b) two-dimensional (2D) arrays of such units; our models build on earlier ones and allow for zero-, one-, and two-sided MF couplings. Our studies of MF units elucidate the dependence of the action-potential (AP) morphology on parameters such as , the fibroblast resting-membrane potential, the fibroblast conductance , and the MF gap-junctional coupling . Furthermore, we find that our MF composite can show autorhythmic and oscillatory behaviors in addition to an excitable response. Our 2D studies use (a) both homogeneous and inhomogeneous distributions of fibroblasts, (b) various ranges for parameters such as , and , and (c) intercellular couplings that can be zero-sided, one-sided, and two-sided connections of fibroblasts with myocytes. We show, in particular, that the plane-wave conduction velocity decreases as a function of , for zero-sided and one-sided couplings; however, for two-sided coupling, decreases initially and then increases as a function of , and, eventually, we observe that conduction failure occurs for low values of . In our homogeneous studies, we find that the rotation speed and stability of a spiral wave can be controlled either by controlling or . Our studies with fibroblast inhomogeneities show that a spiral wave can get anchored to a local fibroblast inhomogeneity. We also study the efficacy of a low-amplitude control scheme, which has been suggested for the control of spiral-wave turbulence in mathematical models for cardiac tissue, in our MF model both with and without heterogeneities. PMID:24023798

  19. Effect of mechanical strain on the collagen VI pericellular matrix in anterior cruciate ligament fibroblasts.

    Science.gov (United States)

    Sardone, Francesca; Traina, Francesco; Tagliavini, Francesca; Pellegrini, Camilla; Merlini, Luciano; Squarzoni, Stefano; Santi, Spartaco; Neri, Simona; Faldini, Cesare; Maraldi, Nadir; Sabatelli, Patrizia

    2014-07-01

    Cell-extracellular matrix interaction plays a major role in maintaining the structural integrity of connective tissues and sensing changes in the biomechanical environment of cells. Collagen VI is a widely expressed non-fibrillar collagen, which regulates tissues homeostasis. The objective of the present investigation was to extend our understanding of the role of collagen VI in human ACL. This study shows that collagen VI is associated both in vivo and in vitro to the cell membrane of knee ACL fibroblasts, contributing to the constitution of a microfibrillar pericellular matrix. In cultured cells the localization of collagen VI at the cell surface correlated with the expression of NG2 proteoglycan, a major collagen VI receptor. The treatment of ACL fibroblasts with anti-NG2 antibody abolished the localization of collagen VI indicating that collagen VI pericellular matrix organization in ACL fibroblasts is mainly mediated by NG2 proteoglycan. In vitro mechanical strain injury dramatically reduced the NG2 proteoglycan protein level, impaired the association of collagen VI to the cell surface, and promoted cell cycle withdrawal. Our data suggest that the injury-induced alteration of specific cell-ECM interactions may lead to a defective fibroblast self-renewal and contribute to the poor regenerative ability of ACL fibroblasts. © 2013 Wiley Periodicals, Inc.

  20. Regulation of IL-6 and IL-8 production by reciprocal cell-to-cell interactions between tumor cells and stromal fibroblasts through IL-1α in ameloblastoma

    Energy Technology Data Exchange (ETDEWEB)

    Fuchigami, Takao [Department of Biochemistry and Genetics, Kagoshima University Graduate School of Medical and Dental Sciences, 8-35-1 Sakuragaoka, Kagoshima 890-8544 (Japan); Department of Oral and Maxillofacial Surgery, Kagoshima University Graduate School of Medical and Dental Sciences, 8-35-1 Sakuragaoka, Kagoshima 890-8544 (Japan); Kibe, Toshiro [Department of Oral and Maxillofacial Surgery, Kagoshima University Graduate School of Medical and Dental Sciences, 8-35-1 Sakuragaoka, Kagoshima 890-8544 (Japan); Koyama, Hirofumi; Kishida, Shosei; Iijima, Mikio [Department of Biochemistry and Genetics, Kagoshima University Graduate School of Medical and Dental Sciences, 8-35-1 Sakuragaoka, Kagoshima 890-8544 (Japan); Nishizawa, Yoshiaki [Kagoshima University Faculty of Medicine, 8-35-1 Sakuragaoka, Kagoshima 890-8544 (Japan); Hijioka, Hiroshi; Fujii, Tomomi [Department of Oral and Maxillofacial Surgery, Kagoshima University Graduate School of Medical and Dental Sciences, 8-35-1 Sakuragaoka, Kagoshima 890-8544 (Japan); Ueda, Masahiro [Natural Science Centre for Research and Education, Kagoshima University, 1-21-24 Koorimoto, Kagoshima 890-8580 (Japan); Nakamura, Norifumi [Department of Oral and Maxillofacial Surgery, Kagoshima University Graduate School of Medical and Dental Sciences, 8-35-1 Sakuragaoka, Kagoshima 890-8544 (Japan); Kiyono, Tohru [Department of Virology, National Cancer Center Research Institute, 5-1-1 Tsukiji, Chuouku, Tokyo 104-0045 (Japan); Kishida, Michiko, E-mail: kmichiko@m2.kufm.kagoshima-u.ac.jp [Department of Biochemistry and Genetics, Kagoshima University Graduate School of Medical and Dental Sciences, 8-35-1 Sakuragaoka, Kagoshima 890-8544 (Japan)

    2014-09-05

    Highlights: • We studied the interaction between tumor cells and fibroblasts in ameloblastoma. • AM-3 ameloblastoma cells secreted significantly high IL-1α levels. • IL-1α derived from AM-3 cells promoted IL-6 and IL-8 secretion of fibroblasts. • IL-6 and IL-8 activated the cellular motility and proliferation of AM-3 cells. - Abstract: Ameloblastoma is an odontogenic benign tumor that occurs in the jawbone, which invades bone and reoccurs locally. This tumor is treated by wide surgical excision and causes various problems, including changes in facial countenance and mastication disorders. Ameloblastomas have abundant tumor stroma, including fibroblasts and immune cells. Although cell-to-cell interactions are considered to be involved in the pathogenesis of many diseases, intercellular communications in ameloblastoma have not been fully investigated. In this study, we examined interactions between tumor cells and stromal fibroblasts via soluble factors in ameloblastoma. We used a human ameloblastoma cell line (AM-3 ameloblastoma cells), human fibroblasts (HFF-2 fibroblasts), and primary-cultured fibroblasts from human ameloblastoma tissues, and analyzed the effect of ameloblastoma-associated cell-to-cell communications on gene expression, cytokine secretion, cellular motility and proliferation. AM-3 ameloblastoma cells secreted higher levels of interleukin (IL)-1α than HFF-2 fibroblasts. Treatment with conditioned medium from AM-3 ameloblastoma cells upregulated gene expression and secretion of IL-6 and IL-8 of HFF-2 fibroblasts and primary-cultured fibroblast cells from ameloblastoma tissues. The AM3-stimulated production of IL-6 and IL-8 in fibroblasts was neutralized by pretreatment of AM-3 cells with anti-IL-1α antibody and IL-1 receptor antagonist. Reciprocally, cellular motility of AM-3 ameloblastoma cells was stimulated by HFF-2 fibroblasts in IL-6 and IL-8 dependent manner. In conclusion, ameloblastoma cells and stromal fibroblasts behave

  1. Fibroblasts in fibrosis: novel roles and mediators

    Directory of Open Access Journals (Sweden)

    Ryan Thomas Kendall

    2014-05-01

    Full Text Available Fibroblasts are the most common cell type of the connective tissues found throughout the body and the principal source of the extensive extracellular matrix (ECM characteristic of these tissues. They are also the central mediators of the pathological fibrotic accumulation of ECM and the cellular proliferation and differentiation that occurs in response to prolonged tissue injury and chronic inflammation. The transformation of the fibroblast cell lineage involves classical developmental signaling programs and includes a surprisingly diverse range of precursor cell types—most notably, myofibroblasts that are the apex of the fibrotic phenotype. Myofibroblasts display exaggerated ECM production; constitutively secrete and are hypersensitive to chemical signals such as cytokines, chemokines, and growth factors; and are endowed with a contractile apparatus allowing them to manipulate the ECM fibers physically to close open wounds. In addition to ECM production, fibroblasts have multiple concomitant biological roles, such as in wound healing, inflammation, and angiogenesis, which are each interwoven with the process of fibrosis. We now recognize many common fibroblast-related features across various physiological and pathological protracted processes. Indeed, a new appreciation has emerged for the role of noncancerous fibroblast interactions with tumors in cancer progression. Although the predominant current clinical treatments of fibrosis involve nonspecific immunosuppressive and anti-proliferative drugs, a variety of potential therapies under investigation specifically target fibroblast biology.

  2. Fibroblasts from different sites may promote or inhibit recruitment of flowing lymphocytes by endothelial cells.

    Science.gov (United States)

    McGettrick, Helen M; Smith, Emily; Filer, Andrew; Kissane, Stephen; Salmon, Michael; Buckley, Christopher D; Rainger, G Ed; Nash, Gerard B

    2009-01-01

    We examined the hypothesis that stromal fibroblasts modulate the ability of endothelial cells (EC) to recruit lymphocytes in a site-specific manner. PBL were perfused over HUVEC that had been cultured with fibroblasts isolated from the inflamed synovium or the skin of patients with rheumatoid arthritis or osteoarthritis, or from normal synovium, with or without exposure to the inflammatory cytokines TNF-alpha+IFN-gamma. Fibroblasts from inflamed synovium, but no others, caused unstimulated HUVEC to bind flowing lymphocytes. This adhesion was supported by alpha(4)beta(1)-VCAM-1 interaction and stabilised by activation of PBL through CXCR4-CXCL12. Antibody neutralisation of IL-6 during co-culture effectively abolished the ability of EC to bind lymphocytes. Cytokine-stimulated EC supported high levels of lymphocyte adhesion, through the presentation of VCAM-1, E-selectin and chemokine(s) acting through CXCR3. Interestingly, co-culture with dermal fibroblasts caused a marked reduction in cytokine-induced adhesion, while synovial fibroblasts had variable effects depending on their source. In the dermal co-cultures, neutralisation of IL-6 or TGF-beta caused partial recovery of cytokine-induced lymphocyte adhesion; this was complete when both were neutralised. Exogenous IL-6 was also found to inhibit response to TNF-alpha+IFN-gamma. Normal stromal fibroblasts appear to regulate the cytokine-sensitivity of vascular endothelium, while fibroblasts associated with chronic inflammation bypass this and develop a directly inflammatory phenotype. Actions of IL-6 might be pro-inflammatory or anti-inflammatory, depending on the local milieu.

  3. Fibroblasts from different sites may promote or inhibit recruitment of flowing lymphocytes by endothelial cells

    Science.gov (United States)

    McGettrick, Helen M; Smith, Emily; Filer, Andrew; Kissane, Stephen; Salmon, Michael; Buckley, Christopher D; Ed Rainger, G; Nash, Gerard B

    2009-01-01

    We examined the hypothesis that stromal fibroblasts modulate the ability of endothelial cells (EC) to recruit lymphocytes in a site-specific manner. PBL were perfused over HUVEC that had been cultured with fibroblasts isolated from the inflamed synovium or the skin of patients with rheumatoid arthritis or osteoarthritis, or from normal synovium, with or without exposure to the inflammatory cytokines TNF-α+IFN-γ. Fibroblasts from inflamed synovium, but no others, caused unstimulated HUVEC to bind flowing lymphocytes. This adhesion was supported by α4β1-VCAM-1 interaction and stabilised by activation of PBL through CXCR4–CXCL12. Antibody neutralisation of IL-6 during co-culture effectively abolished the ability of EC to bind lymphocytes. Cytokine-stimulated EC supported high levels of lymphocyte adhesion, through the presentation of VCAM-1, E-selectin and chemokine(s) acting through CXCR3. Interestingly, co-culture with dermal fibroblasts caused a marked reduction in cytokine-induced adhesion, while synovial fibroblasts had variable effects depending on their source. In the dermal co-cultures, neutralisation of IL-6 or TGF-β caused partial recovery of cytokine-induced lymphocyte adhesion; this was complete when both were neutralised. Exogenous IL-6 was also found to inhibit response to TNF-α+IFN-γ. Normal stromal fibroblasts appear to regulate the cytokine-sensitivity of vascular endothelium, while fibroblasts associated with chronic inflammation bypass this and develop a directly inflammatory phenotype. Actions of IL-6 might be pro-inflammatory or anti-inflammatory, depending on the local milieu. PMID:19130557

  4. N-cadherin is overexpressed in Crohn's stricture fibroblasts and promotes intestinal fibroblast migration.

    LENUS (Irish Health Repository)

    Burke, John P

    2012-02-01

    BACKGROUND: Intestinal fibroblasts mediate stricture formation in Crohn\\'s disease (CD). Transforming growth factor-beta (TGF-beta) is important in fibroblast activation, while cell attachment and migration is regulated by the adhesion molecule N-cadherin. The aim of this study was to investigate the expression and function of N-cadherin in intestinal fibroblasts in patients with fibrostenosing CD. METHODS: Intestinal fibroblasts were cultured from seromuscular biopsies from patients undergoing resection for terminal ileal fibrostenosing CD (n = 14) or controls patients (n = 8). N-cadherin expression was assessed using Western blot and quantitative reverse-transcription polymerase chain reaction (qRT-PCR). Fibroblasts were stimulated with TGF-beta and selective pathway inhibitors Y27632, PD98050, and LY294002 were used to examine the Rho\\/ROCK, ERK-1\\/2, and Akt signaling pathways, respectively. Cell migration was assessed using a scratch wound assay. N-cadherin was selectively overexpressed using a plasmid. RESULTS: Fibroblasts from fibrostenosing CD express increased constitutive N-cadherin mRNA and protein and exhibit enhanced basal cell migration relative to those from directly adjacent normal bowel. Control fibroblasts treated with TGF-beta induced N-cadherin in a dose-dependent manner which was inhibited by Rho\\/ROCK and Akt pathway modulation. Control fibroblasts exhibited enhanced cell migration in response to treatment with TGF-beta or transfection with an N-cadherin plasmid. CONCLUSIONS: Fibroblasts from strictures in CD express increased constitutive N-cadherin and exhibit enhanced basal cell migration. TGF-beta is a potent inducer of N-cadherin in intestinal fibroblasts resulting in enhanced cell migration. The TGF-beta-mediated induction of N-cadherin may potentiate Crohn\\'s stricture formation.

  5. Fibroblast reticular cells engineer a blastema extracellular network during digit tip regeneration in mice.

    Science.gov (United States)

    Marrero, Luis; Simkin, Jennifer; Sammarco, Mimi; Muneoka, Ken

    2017-04-01

    The regeneration blastema which forms following amputation of the mouse digit tip is composed of undifferentiated cells bound together by an organized network of fibers. A monoclonal antibody (ER-TR7) that identifies extracellular matrix (ECM) fibers produced by fibroblast reticular cells during lymphoid organogenesis was used to characterize the ECM of the digit, the blastema, and the regenerate. Digit fibroblast reticular cells produce an ER-TR7+ ECM network associated with different tissues and represent a subset of loose connective tissue fibroblasts. During blastema formation there is an upregulation of matrix production that returns to its pre-existing level and anatomical pattern in the endpoint regenerate. Co-localization studies demonstrate a strong spatial correlation between the ER-TR7 antigen and collagen type III (COL3) in histological sections. ER-TR7 and COL3 are co-induced in cultured digit fibroblasts following treatment with tumor necrosis factor alpha and a lymphotoxin beta receptor agonist. These results provide an initial characterization of the ECM during digit regeneration and identify a subpopulation of fibroblasts involved in producing the blastema provisional matrix that is remodeled during the regeneration response.

  6. Tumor fibroblast-derived epiregulin promotes growth of colitis-associated neoplasms through ERK.

    Science.gov (United States)

    Neufert, Clemens; Becker, Christoph; Türeci, Özlem; Waldner, Maximilian J; Backert, Ingo; Floh, Katharina; Atreya, Imke; Leppkes, Moritz; Jefremow, Andre; Vieth, Michael; Schneider-Stock, Regine; Klinger, Patricia; Greten, Florian R; Threadgill, David W; Sahin, Ugur; Neurath, Markus F

    2013-04-01

    Molecular mechanisms specific to colitis-associated cancers have been poorly characterized. Using comparative whole-genome expression profiling, we observed differential expression of epiregulin (EREG) in mouse models of colitis-associated, but not sporadic, colorectal cancer. Similarly, EREG expression was significantly upregulated in cohorts of patients with colitis-associated cancer. Furthermore, tumor-associated fibroblasts were identified as a major source of EREG in colitis-associated neoplasms. Functional studies showed that Ereg-deficient mice, although more prone to colitis, were strongly protected from colitis-associated tumors. Serial endoscopic studies revealed that EREG promoted tumor growth rather than initiation. Additionally, we demonstrated that fibroblast-derived EREG requires ERK activation to induce proliferation of intestinal epithelial cells (IEC) and tumor development in vivo. To demonstrate the functional relevance of EREG-producing tumor-associated fibroblasts, we developed a novel system for adoptive transfer of these cells via mini-endoscopic local injection. It was found that transfer of EREG-producing, but not Ereg-deficient, fibroblasts from tumors significantly augmented growth of colitis-associated neoplasms in vivo. In conclusion, our data indicate that EREG and tumor-associated fibroblasts play a crucial role in controlling tumor growth in colitis-associated neoplasms.

  7. Biochemical mechanisms of skin radiation burns inhibition and healing by the volumetric autotransplantation of fibroblasts and of keratinocytes with fibroblasts composition

    Directory of Open Access Journals (Sweden)

    L. V. Altukhova

    2015-09-01

    Full Text Available Mechanisms of influence of volumetric autotransplantation of fibroblasts and of the mixture of fibroblasts and keratinocytes on the development of the local 3rd degree X-ray burn and the radiation skin ulcer in guinea pigs were investigated. We used deepadministration into the irradiation zone on its perimeter of 6 doses, which contained (150–160×103 fibroblasts and (130–140×103 keratinocytes in 100 µl. It is shown that this autotransplantation carried out 1 hour after the irradiation, and then every 24 hours, reduces the area of burn on the 35th day, compared to the control by 63%. Radiation ulcer appears on the 10th day after irradiation and is completely healed on the 25th day. With the same regimen of administration of only fibroblasts containing (200–210×103 cells in 100 µl, these parameters of treatment were equal to 31% on 4th and 35th day, respectively. It is shown that as a result of radiation in the area of burn the level of gene expression of collagen types I and III, elastin, fibronectin, vinculin, decorin, hyaluronansynthases 1, 2, 3, matrix metalloproteinases 1, 2, 3, 7, 9 and hyaluronidase is reduced. Besides, in the burn area the level of gene expression of transforming growth factor α, fibroblast growth factors 1, 2, 8 and anti-inflammatory cytokines – interleukin 10 and transforming growth factor-β1 – is reduced, while the level of gene expression of proinflammatory cytokine (interleykin1β increases. Both types of autotransplantation cause the growth of the expression level of all the structural genes and regulatory proteins of biopolymers and decrease in the expression level of interleukin 1β, which leads to activation of tissue regeneration and healing of the burn wound. Reasonsfor the higher efficiency of autotransplantation using the mixture of fibroblasts and keratinocytes compared to autotransplantation by fibroblasts only are both the larger total number of live cells regularly replacing dead cells in

  8. Metallic nanoparticles reduce the migration of human fibroblasts in vitro

    Science.gov (United States)

    Vieira, Larissa Fernanda de Araújo; Lins, Marvin Paulo; Viana, Iana Mayane Mendes Nicácio; dos Santos, Jeniffer Estevão; Smaniotto, Salete; Reis, Maria Danielma dos Santos

    2017-03-01

    Nanoparticles have extremely wide applications in the medical and biological fields. They are being used in biosensors, local drug delivery, diagnostics, and medical therapy. However, the potential effects of nanoparticles on target cell and tissue function, apart from cytotoxicity, are not completely understood. Thus, the aim of this study was to investigate the in vitro effects of silver nanoparticles (AgNPs) and gold nanoparticles (AuNPs) on human fibroblasts with respect to their interaction with the extracellular matrix and in cell migration. Immunofluorescence analysis revealed that treatment with AgNPs or AuNPs decreased collagen and laminin production at all the concentrations tested (0.1, 1, and 10 μg/mL). Furthermore, cytofluorometric analysis showed that treatment with AgNPs reduced the percentage of cells expressing the collagen receptor very late antigen 2, α2β1 integrin (VLA-2) and the laminin receptor very late antigen 6, α6β1 integrin (VLA-6). In contrast, AuNP treatment increased and decreased the percentages of VLA-2-positive and VLA-6-positive cells, respectively, as compared to the findings for the controls. Analysis of cytoskeletal reorganization showed that treatment with both types of nanoparticles increased the formation of stress fibres and number of cell protrusions and impaired cell polarity. Fibroblasts exposed to different concentrations of AuNPs and AgNPs showed reduced migration through transwell chambers in the functional chemotaxis assay. These results demonstrated that metal nanoparticles may influence fibroblast function by negatively modulating the deposition of extracellular matrix molecules (ECM) and altering the expression of ECM receptors, cytoskeletal reorganization, and cell migration.

  9. Hedgehog signaling contributes to basic fibroblast growth factor-regulated fibroblast migration

    Energy Technology Data Exchange (ETDEWEB)

    Zhu, Zhong Xin [School of Pharmaceutical Sciences, Wenzhou Medical University, Wenzhou, Zhejiang (China); Sun, Cong Cong [School of Pharmaceutical Sciences, Wenzhou Medical University, Wenzhou, Zhejiang (China); Wenzhou People' s Hospital, Wenzhou, Zhejiang (China); Ting Zhu, Yu; Wang, Ying; Wang, Tao; Chi, Li Sha; Cai, Wan Hui [School of Pharmaceutical Sciences, Wenzhou Medical University, Wenzhou, Zhejiang (China); Zheng, Jia Yong [Wenzhou People' s Hospital, Wenzhou, Zhejiang (China); Zhou, Xuan [Ningbo First Hospital, Ningbo, Zhejiang (China); Cong, Wei Tao [School of Pharmaceutical Sciences, Wenzhou Medical University, Wenzhou, Zhejiang (China); Li, Xiao Kun, E-mail: proflxk@163.com [School of Pharmaceutical Sciences, Wenzhou Medical University, Wenzhou, Zhejiang (China); Jin, Li Tai, E-mail: jin_litai@126.com [School of Pharmaceutical Sciences, Wenzhou Medical University, Wenzhou, Zhejiang (China)

    2017-06-15

    Fibroblast migration is a central process in skin wound healing, which requires the coordination of several types of growth factors. bFGF, a well-known fibroblast growth factor (FGF), is able to accelerate fibroblast migration; however, the underlying mechanism of bFGF regulation fibroblast migration remains unclear. Through the RNA-seq analysis, we had identified that the hedgehog (Hh) canonical pathway genes including Smoothened (Smo) and Gli1, were regulated by bFGF. Further analysis revealed that activation of the Hh pathway via up-regulation of Smo promoted fibroblast migration, invasion, and skin wound healing, but which significantly reduced by GANT61, a selective antagonist of Gli1/Gli2. Western blot analyses and siRNA transfection assays demonstrated that Smo acted upstream of phosphoinositide 3-kinase (PI3K)-c-Jun N-terminal kinase (JNK)-β-catenin to promote cell migration. Moreover, RNA-seq and qRT-PCR analyses revealed that Hh pathway genes including Smo and Gli1 were under control of β-catenin, suggesting that β-catenin turn feedback activates Hh signaling. Taken together, our analyses identified a new bFGF-regulating mechanism by which Hh signaling regulates human fibroblast migration, and the data presented here opens a new avenue for the wound healing therapy. - Highlights: • bFGF regulates Hedgehog (Hh) signaling in fibroblasts. • The Smo and Gli two master regulators of Hh signaling positively regulate fibroblast migration. • Smo facilitates β-catenin nuclear translocation via activation PI3K/JNK/GSK3β. • β-catenin positively regulates fibroblast cell migration and the expression of Hh signaling genes including Smo and Gli.

  10. Effects of dynamic matrix remodelling on en masse migration of fibroblasts on collagen matrices.

    Science.gov (United States)

    Ozcelikkale, Altug; Dutton, J Craig; Grinnell, Frederick; Han, Bumsoo

    2017-10-01

    Fibroblast migration plays a key role during various physiological and pathological processes. Although migration of individual fibroblasts has been well studied, migration in vivo often involves simultaneous locomotion of fibroblasts sited in close proximity, so-called 'en masse migration', during which intensive cell-cell interactions occur. This study aims to understand the effects of matrix mechanical environments on the cell-matrix and cell-cell interactions during en masse migration of fibroblasts on collagen matrices. Specifically, we hypothesized that a group of migrating cells can significantly deform the matrix, whose mechanical microenvironment dramatically changes compared with the undeformed state, and the alteration of the matrix microenvironment reciprocally affects cell migration. This hypothesis was tested by time-resolved measurements of cell and extracellular matrix movement during en masse migration on collagen hydrogels with varying concentrations. The results illustrated that a group of cells generates significant spatio-temporal deformation of the matrix before and during the migration. Cells on soft collagen hydrogels migrate along tortuous paths, but, as the matrix stiffness increases, cell migration patterns become aligned with each other and show coordinated migration paths. As cells migrate, the matrix is locally compressed, resulting in a locally stiffened and dense matrix across the collagen concentration range studied. © 2017 The Author(s).

  11. Cryopreservation of canine ovarian and testicular fibroblasts.

    Science.gov (United States)

    Yu, Il-Jeoung; Leibo, S P; Songsasen, Nucharin; Dresser, Betsy L; Kim, In-Shik

    2009-01-01

    To derive a practical procedure to store canine somatic cells, fibroblasts isolated from testicular or ovarian tissues were cryopreserved in 1.2 M ethylene glycol or in 1.2 M dimethylsulfoxide prepared in Dulbecco's Modified Eagle Medium as cryoprotectants, and were frozen either in plastic straws or vials. Thawed cells were cultured for 24 hr at 38.5 degree C in a humidified atmosphere of 5 percent CO2 95 percent air, and then their membrane integrity was assayed with a double fluorescent stain, Fertilight. In addition, frozen-thawed fibroblasts were cultured for 4 days, and then their functional survival was measured after staining small colonies with trypan blue. After freezing and thawing, membrane integrity of testicular fibroblasts was 55-70 percent and functional survival ranged from 20-40 percent. With frozen-thawed ovarian cells, the average membrane integrity was 55-75 percent and the average functional survival was 35-40 percent. When frozen in ethylene glycol, functional survival of ovarian fibroblasts was significantly higher than that of testicular cells (P less than 0.05). These methods should prove useful to preserve cells collected from canids in the wild.

  12. Fibroblast growth factor 23 - et fosfatregulerende hormon

    DEFF Research Database (Denmark)

    Beck-Nielsen, Signe; Pedersen, Susanne Møller; Kassem, Moustapha

    2010-01-01

    Fibroblast growth factor 23 (FGF23) er et nyligt identificeret fosfatonin. FGF23's fysiologiske hovedfunktion er at opretholde normalt serumfosfat og at virke som et D-vitaminmodregulatorisk hormon. Sygdomme, der er koblet til forhøjet serum FGF23, er hypofosfatæmisk rakitis, fibrøs dysplasi og...

  13. Allogeneic fibroblasts in dermal substitutes induce inflammation and scar formation

    NARCIS (Netherlands)

    Lamme, Evert N.; van Leeuwen, Rene T. J.; Mekkes, Jan R.; Middelkoop, Esther

    2002-01-01

    In the present study, we compared the use of autologous versus allogeneic fibroblasts in dermal skin substitutes in a porcine wound model. The allogeneic fibroblast populations were isolated from female and a male pig (allo-1, - 2 and - 3) and the controls, autologous fibroblasts, from female

  14. HETEROGENIC SERUM, AGE, AND MULTIPLICATION OF FIBROBLASTS.

    Science.gov (United States)

    Carrel, A; Ebeling, A H

    1922-01-01

    The presence in a culture medium of heterogenic serum of various concentrations exerts a definite influence on the rate of multiplication of fibroblasts. Dog serum does not inhibit the growth of See PDF for Structure chicken fibroblasts markedly until its concentration reaches 15 per cent. Beyond this figure, each increase of the concentration brings about a rapid decrease in the rate of cell multiplication. When the concentration reaches from 30 to 45 per cent, no growth takes place. The inhibiting action of cat serum begins to manifest itself at a concentration of 25 per cent and prevents cell proliferation completely at a concentration of 55 and 60 per cent. The ratio, See PDF for Equation can be taken as expressing the action of the serum on fibroblast multiplication; that is, as the growth index of the serum. See PDF for Structure The inhibiting influence of heterogenic serum was found to vary in direct ratio to the age of the animal from which it was obtained. The rate of proliferation of chicken fibroblasts was studied comparatively in media containing varied concentrations of serum from young and old animals. For each concentration of serum, the rate of growth in the serum of the old animal was expressed in relation to the rate of growth in the serum of the young animal. When cat serum was used, the curve obtained in plotting this ratio in ordinates and the serum concentration in abscissae showed a rapid increase in the inhibiting action of the old serum as soon as the concentration reached 30 per cent. The same tests were repeated with the serum from young and old dogs. The general results were identical, although See PDF for Structure the quantitative inhibiting action of both sera was greater than that of cat serum. It may be concluded that under the conditions of the experiments: 1. Heterogenicsera inhibit and prevent the growth of chicken fibroblasts when their concentration is made to vary within certain limits. 2. A relation exists between the rate

  15. Fibroblast α11β1 integrin regulates tensional homeostasis in fibroblast/A549 carcinoma heterospheroids.

    Directory of Open Access Journals (Sweden)

    Ning Lu

    Full Text Available We have previously shown that fibroblast expression of α11β1 integrin stimulates A549 carcinoma cell growth in a xenograft tumor model. To understand the molecular mechanisms whereby a collagen receptor on fibroblast can regulate tumor growth we have used a 3D heterospheroid system composed of A549 tumor cells and fibroblasts without (α11+/+ or with a deletion (α11-/- in integrin α11 gene. Our data show that α11-/-/A549 spheroids are larger than α11+/+/A549 spheroids, and that A549 cell number, cell migration and cell invasion in a collagen I gel are decreased in α11-/-/A549 spheroids. Gene expression profiling of differentially expressed genes in fibroblast/A549 spheroids identified CXCL5 as one molecule down-regulated in A549 cells in the absence of α11 on the fibroblasts. Blocking CXCL5 function with the CXCR2 inhibitor SB225002 reduced cell proliferation and cell migration of A549 cells within spheroids, demonstrating that the fibroblast integrin α11β1 in a 3D heterospheroid context affects carcinoma cell growth and invasion by stimulating autocrine secretion of CXCL5. We furthermore suggest that fibroblast α11β1 in fibroblast/A549 spheroids regulates interstitial fluid pressure by compacting the collagen matrix, in turn implying a role for stromal collagen receptors in regulating tensional hemostasis in tumors. In summary, blocking stromal α11β1 integrin function might thus be a stroma-targeted therapeutic strategy to increase the efficacy of chemotherapy.

  16. Fibroblast-fibronectin patterning and network formation in 3D fibrin matrices.

    Science.gov (United States)

    Miron-Mendoza, Miguel; Graham, Eric; Manohar, Sujal; Petroll, W Matthew

    2017-06-07

    We previously reported that fibroblasts migrating within 3-D collagen matrices move independently, whereas fibroblasts within 3-D fibrin matrices form an interconnected network. Similar networks have been identified previously during in vivo corneal wound healing. In this study, we investigate the role of fibronectin in mediating this mechanism of collective cell spreading, migration and patterning. To assess cell spreading, corneal fibroblasts were plated within fibrillar collagen or fibrin matrices. To assess migration, compacted cell-populated collagen matrices were nested inside cell-free fibrin matrices. Constructs were cultured in serum-free media containing PDGF, with or without RGD peptide, anti-α5 or anti-fibronectin blocking antibodies. In some experiments, LifeAct and fluorescent fibronectin were used to allow dynamic assessment of cell-induced fibronectin reorganization. 3-D and 4-D imaging were used to assess cell mechanical behavior, connectivity, F-actin, α5 integrin and fibronectin organization. Corneal fibroblasts within 3-D fibrin matrices formed an interconnected network that was lined with cell-secreted fibronectin. Live cell imaging demonstrated that fibronectin tracks were formed at the leading edge of spreading and migrating cells. Furthermore, fibroblasts preferentially migrated through fibronectin tracks laid down by other cells. Interfering with cell-fibronectin binding with RGD, anti α5 integrin or anti fibronectin antibodies inhibited cell spreading and migration through fibrin, but did not affect cell behavior in collagen. In this study, a novel mode of cell patterning was identified in which corneal fibroblasts secrete and attach to fibronectin via α5β1 integrin to facilitate spreading and migration within 3-D fibrin matrices, resulting in the formation of localized fibronectin tracks. Other cells use these fibronectin tracks as conduits, resulting in an interconnected cell-fibronectin network. Copyright © 2017 International

  17. Quantitative analysis of chromatin accessibility in mouse embryonic fibroblasts.

    Science.gov (United States)

    Zhuo, Baowen; Yu, Juan; Chang, Luyuan; Lei, Jiafan; Wen, Zengqi; Liu, Cuifang; Mao, Guankun; Wang, Kehui; Shen, Jie; Xu, Xueqing

    2017-11-04

    Genomic DNA of eukaryotic cells is hierarchically packaged into chromatin by histones. The dynamic organization of chromatin fibers plays a critical role in the regulation of gene transcription and other DNA-associated biological processes. Recently, numerous approaches have been developed to map the chromatin organization by characterizing chromatin accessibilities in genome-wide. However, reliable methods to quantitatively map chromatin accessibility are not well-established, especially not on a genome-wide scale. Here, we developed a modified MNase-seq for mouse embryonic fibroblasts, wherein chromatin was partially digested at multiple digestion times using micrococcal nuclease (MNase), allowing quantitative analysis of local yet genome-wide chromatin compaction. Our results provide strong evidence that the chromatin accessibility at promoter regions are positively correlated with gene activity. In conclusion, our assay is an ideal tool for the quantitative study of gene regulation in the perspective of chromatin accessibility. Copyright © 2017 Elsevier Inc. All rights reserved.

  18. Iron status and fibroblast growth factor-23 in Gambian children

    Science.gov (United States)

    Braithwaite, Vickie; Jarjou, Landing M.A.; Goldberg, Gail R.; Prentice, Ann

    2012-01-01

    A relationship between iron and fibroblast growth factor-23 (FGF23) metabolic pathways has been proposed. Iron deficiency anaemia is prevalent in The Gambia and concentrations of fibroblast growth factor-23 FGF23 are elevated in a large percentage of Gambian children with rickets-like bone deformity. We speculate that low iron status may be involved in the aetiology of Gambian rickets. The aim of this study was to determine if there was a relationship between haemoglobin, as a marker of iron status, and FGF23 in samples from children with and without a history of rickets-like bone deformities in The Gambia. We conducted a retrospective analysis of studies carried out from 2006 to 2008 in children from a rural community in The Gambia where iron deficiency anaemia is endemic and where elevated circulating concentrations of FGF23 have been found. To investigate the relationship between circulating FGF23 and haemoglobin concentrations we used an age-adjusted linear regression model on data from children rickets-like bone deformity (BD) (n = 108) and from the local community (LC) (n = 382). We found that circulating concentration of FGF23 was inversely correlated with haemoglobin concentration. This effect was more pronounced in BD children compared with LC children (interaction: P ≤ 0.0001). Anaemia and elevated FGF23 were more prevalent in BD children compared to LC children (P = 0.0003 and P = 0.0001 respectively). In conclusion, there is a stronger relationship between FGF23 and haemoglobin in Gambian children with a history of rickets compared to local community children. This study provides support for the contention that iron may be involved in FGF23 metabolic pathways. PMID:22465847

  19. The effect of tranilast on fibroblast activation protein α (FAP-α expression in normal and keloid fibroblasts in vitro

    Directory of Open Access Journals (Sweden)

    Paweł P. Antończak

    2017-07-01

    Full Text Available Introduction . Tranilast (N-(3’,4’-demethoxycinnamoyl-anthranilic acid is an anti-allergic drug. Its mechanism of action is based on the inhibition of antigen-induced release of chemical mediators from mast cells and basophils. It also reveals antifibroproliferative activities. These properties of tranilast are used in the treatment of hypertrophic scars and keloids. Keloids are characterized by incorrect extracellular matrix components turnover. Fibroblasts derived from keloids reveal overproduction of collagen type I and decreased degradation of extracellular matrix in comparison with normal fibroblasts. Fibroblast activation protein α (FAP-α may play an important role in remodeling of extracellular matrix and the invasive properties of keloids. Objective . In the present study, the effect of tranilast on expression of FAP-α gene and its protein was evaluated in normal human dermal fibroblasts and fibroblasts derived from keloids cultured in vitro . Materials and methods. In the first stage of the study, the influence of tranilast on cell viability was estimated. The second stage of the study included the quantitative evaluation of FAP-α mRNA expression in normal and keloid fibroblasts treated with tranilast. The third stage of the study comprised fibroblast activation protein α expression analysis in the examined cells treated with tranilast. Results and conclusions . The expression of FAP-α gene and fibroblast activation protein α is higher in keloid fibroblasts. Tranilast at concentrations of 3 μM and 30 μM up-regulated mRNA FAP-α expression in normal fibroblasts but did not influence keloid fibroblasts. The drug, at concentrations of 30 μM and 300 μM up-regulated fibroblast activation protein α expression in normal fibroblasts and did not influence keloid fibroblasts. Tranilast antiproliferative effect is not associated with FAP-α expression in keloid fibroblasts.

  20. Acidic Fibroblast Growth Factor Promotes Vascular Repair

    Science.gov (United States)

    Bjornsson, Thorir D.; Dryjski, Maciej; Tluczek, John; Mennie, Robert; Ronan, John; Mellin, Theodore N.; Thomas, Kenneth A.

    1991-10-01

    Intravascular injury to arteries can result in thickening of the intimal smooth muscle layer adjacent to the lumen by migration and proliferation of cells from the underlying medial smooth muscle layer accompanied by deposition of extracellular matrix. This pathological response, which decreases lumen diameter, might, in part, be the result of the access of smooth muscle cells to plasma and platelet-derived growth factors as a consequence of denudation of the overlying confluent monolayer of vascular endothelial cells. Injured rat carotid arteries were treated by i.v. administration of acidic fibroblast growth factor, a heparin-binding protein that is chemotactic and mitogenic for vascular endothelial cells. The growth factor treatment resulted in dose-dependent inhibition of intimal thickening with parallel promotion of endothelial regeneration over the injured area. Therefore, acidic fibroblast growth factor might be efficacious in the prevention of restenosis caused by intimal thickening following angioplasty in humans.

  1. Posttranslational modification of β-catenin is associated with pathogenic fibroblastic changes in bronchopulmonary dysplasia.

    Science.gov (United States)

    Sucre, Jennifer M S; Vijayaraj, Preethi; Aros, Cody J; Wilkinson, Dan; Paul, Manash; Dunn, Bruce; Guttentag, Susan H; Gomperts, Brigitte N

    2017-02-01

    Bronchopulmonary dysplasia (BPD) is a common complication of premature birth. The histopathology of BPD is characterized by an arrest of alveolarization with fibroblast activation. The Wnt/β-catenin signaling pathway is important in early lung development. When Wnt signaling is active, phosphorylation of β-catenin by tyrosine kinases at activating sites, specifically at tyrosine 489 (Y489), correlates with nuclear localization of β-catenin. We examined fetal lung tissue, lung tissue from term newborns, and lung tissue from infants who died with BPD; we found nuclear β-catenin phosphorylation at Y489 in epithelial and mesenchymal cells in fetal tissue and BPD tissue, but not in the lungs of term infants. Using a 3D human organoid model, we found increased nuclear localization of β-catenin phosphorylated at Y489 (p-β-catenin(Y489)) after exposure to alternating hypoxia and hyperoxia compared with organoids cultured in normoxia. Exogenous stimulation of the canonical Wnt pathway in organoids was sufficient to cause nuclear localization of p-β-catenin(Y489) in normoxia and mimicked the pattern of α-smooth muscle actin (α-SMA) expression seen with fibroblastic activation from oxidative stress. Treatment of organoids with a tyrosine kinase inhibitor prior to cyclic hypoxia-hyperoxia inhibited nuclear localization of p-β-catenin(Y489) and prevented α-SMA expression by fibroblasts. Posttranslational phosphorylation of β-catenin is a transient feature of normal lung development. Moreover, the persistence of p-β-catenin(Y489) is a durable marker of fibroblast activation in BPD and may play an important role in BPD disease pathobiology. Copyright © 2017 the American Physiological Society.

  2. Fibroblasts as architects of cancer pathogenesis.

    Science.gov (United States)

    Marsh, Timothy; Pietras, Kristian; McAllister, Sandra S

    2013-07-01

    Studies of epithelial cancers (i.e., carcinomas) traditionally focused on transformation of the epithelium (i.e., the cancer cells) and how aberrant signaling within the cancer cells modulates the surrounding tissue of origin. In more recent decades, the normal cells, blood vessels, molecules, and extracellular components that surround the tumor cells, collectively known as the "tumor microenvironment" or "stroma", have received increasing attention and are now thought to be key regulators of tumor initiation and progression. Of particular relevance to the work reviewed herein are the fibroblasts, which make up the major cell type within the microenvironment of most carcinomas. Due to their inherent heterogeneity, plasticity, and function, it is perhaps not surprising that fibroblasts are ideal modulators of normal and cancerous epithelium; however, these aspects also present challenges if we are to interrupt their tumor-supportive functions. Here, we review the current body of knowledge and the many questions that still remain about the special entity known as the cancer-associated fibroblast. This article is part of a Special Issue entitled: Fibrosis: Translation of basic research to human disease. Copyright © 2012. Published by Elsevier B.V.

  3. Fibroblasts as maestros orchestrating tissue regeneration.

    Science.gov (United States)

    Costa-Almeida, Raquel; Soares, Raquel; Granja, Pedro L

    2017-01-20

    Fibroblasts constitute a dynamic and versatile population of cells of mesenchymal origin, implicated in both regenerative strategies and pathological conditions. Despite being frequently associated to disease development, particularly through the establishment of fibrotic tissue, fibroblasts hold great potential for tissue engineering and regenerative medicine applications. They are responsible for synthesizing and depositing extracellular matrix components, allowing other cells to settle and migrate along a three-dimensional support and thereby generating an organ-specific architecture. Additionally, they produce bioactive molecules that are involved in several physiological processes, including angiogenesis and tissue repair. Although there seems to be much still to unveil about these fascinating cells they have been attracting increasing interest and are now being intensively explored as a cell source to develop bioengineered tissue constructs or to improve stem cell-based technologies. This review intends to highlight the potential of fibroblasts in orchestrating tissue regeneration, as well as to contribute to uncover uncharted prospective applications of these cells. Copyright © 2017 John Wiley & Sons, Ltd. Copyright © 2017 John Wiley & Sons, Ltd.

  4. Rho Kinase Regulation of Fibroblast Migratory Mechanics in Fibrillar Collagen Matrices.

    Science.gov (United States)

    Zhou, Chengxin; Petroll, W Matthew

    2010-03-01

    Migration of activated corneal fibroblasts plays an important role in matrix patterning during embryonic development and wound repopulation following injury or refractive surgery. In this study, we investigate the role of Rho kinase in regulating fibroblast migration mechanics, by modifying a previously described nested collagen matrix model to facilitate dynamic imaging of cell-matrix interactions.Human corneal fibroblasts were cultured in nested matrices with media containing either 1% fetal bovine serum (FBS), or 1% FBS plus the Rho kinase inhibitor Y-27632. Time-lapse DIC imaging of cell and extracellular matrix (ECM) movements was performed for up to 72 hours. In addition, static confocal imaging was used to assess 3-D cell morphology and local matrix reorganization.In 1% FBS, significant tractional forces were generated during migration, as indicated by inward displacement and reorganization of collagen in front of cells. When Rho kinase was inhibited, cells became more elongated, and extended dendritic processes into the outer matrix. Interestingly, these dendritic cells were still able to generate tractional forces at their leading edge, whereas cell translocation was substantially reduced. Overall, the data suggests that Rho kinase impacts 3-D fibroblast migration by affecting morphology, polarization, and mechanical coordination between the leading and trailing edges of cells.

  5. Fibroblast and keratinocyte gene expression following exposure to extracts of neem plant (Azadirachta indica

    Directory of Open Access Journals (Sweden)

    Takao Someya

    2018-02-01

    Full Text Available This data article provides gene expression profiles, determined by using real-time PCR, of fibroblasts and keratinocytes treated with 0.01% and 0.001% extracts of neem plant (Azadirachta indica, local name “Kohomba” in Sri Lanka, harvested in Sri Lanka. For fibroblasts, the dataset includes expression profiles for genes encoding hyaluronan synthase 1 (HAS1, hyaluronan synthase 2 (HAS2, hyaluronidase-1 (HYAL1, hyaluronidase-2 (HYAL2, versican, aggrecan, CD44, collagen, type I, alpha 1 (COL1A1, collagen, type III, alpha 1 (COL3A1, collagen, type VII, alpha 1 (COL7A1, matrix metalloproteinase 1 (MMP1, acid ceramidase, basic fibroblast growth factor (bFGF, fibroblast growth factor-7 (FGF7, vascular endothelial growth factor (VEGF, interleukin-1 alpha (IL-1α, cyclooxygenase-2 (cox2, transforming growth factor beta (TGF-β, and aquaporin 3 (AQP3. For keratinocytes, the expression profiles are for genes encoding HAS1, HAS2, HYAL1, HYAL2, versican, CD44, IL-1α, cox2, TGF-β, AQP3, Laminin5, collagen, type XVII, alpha 1 (COL17A1, integrin alpha-6 (ITGA6, ceramide synthase 3 (CERS3, elongation of very long chain fatty acids protein 1 (ELOVL1, elongation of very long chain fatty acids protein 4 (ELOVL4, filaggrin (FLG, transglutaminase 1 (TGM1, and keratin 1 (KRT1. The expression profiles are provided as bar graphs.

  6. Alteration of Skin Properties with Autologous Dermal Fibroblasts

    Directory of Open Access Journals (Sweden)

    Rajesh L. Thangapazham

    2014-05-01

    Full Text Available Dermal fibroblasts are mesenchymal cells found between the skin epidermis and subcutaneous tissue. They are primarily responsible for synthesizing collagen and glycosaminoglycans; components of extracellular matrix supporting the structural integrity of the skin. Dermal fibroblasts play a pivotal role in cutaneous wound healing and skin repair. Preclinical studies suggest wider applications of dermal fibroblasts ranging from skin based indications to non-skin tissue regeneration in tendon repair. One clinical application for autologous dermal fibroblasts has been approved by the Food and Drug Administration (FDA while others are in preclinical development or various stages of regulatory approval. In this context, we outline the role of fibroblasts in wound healing and discuss recent advances and the current development pipeline for cellular therapies using autologous dermal fibroblasts. The microanatomic and phenotypic differences of fibroblasts occupying particular locations within the skin are reviewed, emphasizing the therapeutic relevance of attributes exhibited by subpopulations of fibroblasts. Special focus is provided to fibroblast characteristics that define regional differences in skin, including the thick and hairless skin of the palms and soles as compared to hair-bearing skin. This regional specificity and functional identity of fibroblasts provides another platform for developing regional skin applications such as the induction of hair follicles in bald scalp or alteration of the phenotype of stump skin in amputees to better support their prosthetic devices.

  7. Pericellular Versican Regulates the Fibroblast-Myofibroblast Transition

    Science.gov (United States)

    Hattori, Noriko; Carrino, David A.; Lauer, Mark E.; Vasanji, Amit; Wylie, James D.; Nelson, Courtney M.; Apte, Suneel S.

    2011-01-01

    The cell and its glycosaminoglycan-rich pericellular matrix (PCM) comprise a functional unit. Because modification of PCM influences cell behavior, we investigated molecular mechanisms that regulate PCM volume and composition. In fibroblasts and other cells, aggregates of hyaluronan and versican are found in the PCM. Dermal fibroblasts from Adamts5−/− mice, which lack a versican-degrading protease, ADAMTS5, had reduced versican proteolysis, increased PCM, altered cell shape, enhanced α-smooth muscle actin (SMA) expression and increased contractility within three-dimensional collagen gels. The myofibroblast-like phenotype was associated with activation of TGFβ signaling. We tested the hypothesis that fibroblast-myofibroblast transition in Adamts5−/− cells resulted from versican accumulation in PCM. First, we noted that versican overexpression in human dermal fibroblasts led to increased SMA expression, enhanced contractility, and increased Smad2 phosphorylation. In contrast, dermal fibroblasts from Vcan haploinsufficient (Vcanhdf/+) mice had reduced contractility relative to wild type fibroblasts. Using a genetic approach to directly test if myofibroblast transition in Adamts5−/− cells resulted from increased PCM versican content, we generated Adamts5−/−;Vcanhdf/+ mice and isolated their dermal fibroblasts for comparison with dermal fibroblasts from Adamts5−/− mice. In Adamts5−/− fibroblasts, Vcan haploinsufficiency or exogenous ADAMTS5 restored normal fibroblast contractility. These findings demonstrate that altering PCM versican content through proteolytic activity of ADAMTS5 profoundly influenced the dermal fibroblast phenotype and may regulate a phenotypic continuum between the fibroblast and its alter ego, the myofibroblast. We propose that a physiological function of ADAMTS5 in dermal fibroblasts is to maintain optimal versican content and PCM volume by continually trimming versican in hyaluronan-versican aggregates. PMID:21828051

  8. Nucleoplasmic LAP2alpha-lamin A complexes are required to maintain a proliferative state in human fibroblasts.

    NARCIS (Netherlands)

    Pekovic, V.; Harborth, J.; Broers, J.L.; Ramaekers, F.C.S.; Engelen, B.G.M. van; Lammens, M.M.Y.; Zglinicki, T. von; Foisner, R.; Hutchison, C.; Markiewicz, E.

    2007-01-01

    In human diploid fibroblasts (HDFs), expression of lamina-associated polypeptide 2 alpha (LAP2alpha) upon entry and exit from G(0) is tightly correlated with phosphorylation and subnuclear localization of retinoblastoma protein (Rb). Phosphoisoforms of Rb and LAP2alpha are down-regulated in G(0).

  9. Primary cell culture from human oral tissue: gingival keratinocytes,gingival fibroblasts and periodontal ligament fibroblasts

    Directory of Open Access Journals (Sweden)

    Supreya Wanichpakorn

    2010-08-01

    Full Text Available Primary cell culture of human oral tissue has many applications for oral biology research. There are two techniques in primary culture, which includes the enzymatic and direct explant technique. The objectives of this study were (1 to isolate and investigate the difference in percentage the success in culturing three cell types from human oral tissue: gingival keratinocytes, gingival fibroblasts and periodontal ligament fibroblasts by using the direct explant technique; (2 to compare the effect of sex and age on the success of tissue culturing. Twenty seven tissue samples were obtained from healthy human gingival tissue, 19 female and 8 male patients aged 14-67 years (37.7±17.5. The tissue was cut into 1x1 mm pieces and placed on plastic culture plates containing Dulbecco’s Modified Eagle’s Medium supplemented with 10% fetal calf serum, 100 U/ml penicillin, 100 µg/ml streptomycin and 1% amphotericin B. For the keratinocytes culture, after the epithelial cells started to multiply around the gingival origin and the diameter was 2-5 mm., the fibroblasts were liminated by mechanical removal under inverted microscope to prevent fibroblast overgrowth and the medium was changed to keratinocyte-SFM (Gibco, BRL supplemented with 5 µg/ml gentamycin. The results revealed that gingival fibroblast gave the highest success rate in culture (96.3%, followed by gingival keratinocytes (88.9% and periodontal ligament fibroblasts (81.5%. There was no significant difference in the success rate of cultivation between younger and older individuals, as between sex of the subjects (p>0.05. The risk of failure in culture techniques is mainly caused by microbiological contamination from the tissue samples.

  10. Mutant Parkin impairs mitochondrial function and morphology in human fibroblasts.

    Directory of Open Access Journals (Sweden)

    Anne Grünewald

    Full Text Available BACKGROUND: Mutations in Parkin are the most common cause of autosomal recessive Parkinson disease (PD. The mitochondrially localized E3 ubiquitin-protein ligase Parkin has been reported to be involved in respiratory chain function and mitochondrial dynamics. More recent publications also described a link between Parkin and mitophagy. METHODOLOGY/PRINCIPAL FINDINGS: In this study, we investigated the impact of Parkin mutations on mitochondrial function and morphology in a human cellular model. Fibroblasts were obtained from three members of an Italian PD family with two mutations in Parkin (homozygous c.1072delT, homozygous delEx7, compound-heterozygous c.1072delT/delEx7, as well as from two relatives without mutations. Furthermore, three unrelated compound-heterozygous patients (delEx3-4/duplEx7-12, delEx4/c.924C>T and delEx1/c.924C>T and three unrelated age-matched controls were included. Fibroblasts were cultured under basal or paraquat-induced oxidative stress conditions. ATP synthesis rates and cellular levels were detected luminometrically. Activities of complexes I-IV and citrate synthase were measured spectrophotometrically in mitochondrial preparations or cell lysates. The mitochondrial membrane potential was measured with 5,5',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazolylcarbocyanine iodide. Oxidative stress levels were investigated with the OxyBlot technique. The mitochondrial network was investigated immunocytochemically and the degree of branching was determined with image processing methods. We observed a decrease in the production and overall concentration of ATP coinciding with increased mitochondrial mass in Parkin-mutant fibroblasts. After an oxidative insult, the membrane potential decreased in patient cells but not in controls. We further determined higher levels of oxidized proteins in the mutants both under basal and stress conditions. The degree of mitochondrial network branching was comparable in mutants and

  11. Local Foods, Local Places

    Science.gov (United States)

    The Local Foods, Local Places technical assistance program protects human health and the environment, spurs revitalization, increases access to healthy foods, and creates economic opportunities by promoting local foods.

  12. The mutant form of lamin A that causes Hutchinson-Gilford progeria is a biomarker of cellular aging in human skin.

    Directory of Open Access Journals (Sweden)

    Dayle McClintock

    2007-12-01

    Full Text Available Hutchinson-Gilford progeria syndrome (HGPS, OMIM 176670 is a rare disorder characterized by accelerated aging and early death, frequently from stroke or coronary artery disease. 90% of HGPS cases carry the LMNA G608G (GGC>GGT mutation within exon 11 of LMNA, activating a splice donor site that results in production of a dominant negative form of lamin A protein, denoted progerin. Screening 150 skin biopsies from unaffected individuals (newborn to 97 years showed that a similar splicing event occurs in vivo at a low level in the skin at all ages. While progerin mRNA remains low, the protein accumulates in the skin with age in a subset of dermal fibroblasts and in a few terminally differentiated keratinocytes. Progerin-positive fibroblasts localize near the basement membrane and in the papillary dermis of young adult skin; however, their numbers increase and their distribution reaches the deep reticular dermis in elderly skin. Our findings demonstrate that progerin expression is a biomarker of normal cellular aging and may potentially be linked to terminal differentiation and senescence in elderly individuals.

  13. [Analysis of chromosome karyotype of oral carcinoma-associated fibroblasts].

    Science.gov (United States)

    Zheng, Xiao-Hui; Liu, Ying; Zhou, Hong-Mei; Chen, Qian-Ming; Li, Bing-Qi

    2005-04-01

    The purpose of this study was to investigate whether the fundamental genetic character of oral carcinoma-associated fibroblasts changes through contrasting and analyzing the oral carcinoma-associated fibroblasts and the normal fibroblasts of oral mucosa. The two kinds of cells were treated with colchicine and microsometic fluid, and then were expanded with cold acetic acid and formalized with methyl alcohol. The cells were observed under the oil microscope after Giemsa staining. The chromosome karyotype of the two kinds of cells was analyzed by Visus 2. 1. There were not obvious differences in the way of chromosome karyotype between the oral carcinoma-associated fibroblasts and the normal fibroblasts of oral mucosa. The basic genetic characteristics of the normal cells are conserved in the oral carcinoma-associated fibroblasts, which means the cells have no malignant changes.

  14. Low Dose Theophylline Showed an Inhibitory Effect on the Production of IL-6 and IL-8 in Primary Lung Fibroblast from Patients with COPD

    Science.gov (United States)

    Zhang, Jing; Feng, Ming-xiang; Qu, Jie-ming

    2012-01-01

    Chronic obstructive pulmonary disease (COPD) is characterized by the abnormal and chronic lung inflammation. We hypothesized that lung fibroblasts could contribute to the local inflammation and investigated whether low dose theophylline had a beneficial effect on fibroblasts inflammation. Subjects undergoing lobectomy for bronchial carcinoma were enrolled and divided into COPD and control groups according to spirometry. Primary human lung fibroblasts were cultured from peripheral lung tissue distant to tumor tissue. There was a significant increase in both the mRNA expressions and protein levels for IL-6 and IL-8 in fibroblasts in COPD group, and the values were negatively correlated with lung function (P theophylline treatment. In addition, theophylline at the dose of 5 μg/mL reduced the increased production of IL-6 and IL-8 induced by 1 μg/mL LPS in primary human lung fibroblasts. Our data suggest that lung fibroblasts participate in the chronic inflammation in COPD by releasing IL-6 and IL-8, and low dose theophylline can alleviate the proinflammatory mediators' production by fibroblasts. PMID:22363103

  15. HGF is released from buccal fibroblasts after smokeless tobacco stimulation

    DEFF Research Database (Denmark)

    Dabelsteen, S; Christensen, S; Gron, B

    2005-01-01

    To investigate the effect of smokeless tobacco (ST) on (1) HGF, KGF and GM-CSF expression by buccal fibroblasts and (2) on keratinocyte and fibroblast proliferation. Buccal fibroblasts were stimulated with different concentrations of ST extracts in a double dilution from 0.50% w/v to 0.03% w/v. S....... Keratinocytes and fibroblasts showed no increase in proliferation after stimulation with increased concentrations of ST. The results suggest that HGF and KGF may play an important role as a paracrine growth factor in epithelial hyperplasia in ST lesions....

  16. Hsp90 regulation of fibroblast activation in pulmonary fibrosis

    Science.gov (United States)

    Sontake, Vishwaraj; Wang, Yunguan; Kasam, Rajesh K.; Sinner, Debora; Reddy, Geereddy B.; Naren, Anjaparavanda P.; McCormack, Francis X.; Jegga, Anil G.; Madala, Satish K.

    2017-01-01

    Idiopathic pulmonary fibrosis (IPF) is a severe fibrotic lung disease associated with fibroblast activation that includes excessive proliferation, tissue invasiveness, myofibroblast transformation, and extracellular matrix (ECM) production. To identify inhibitors that can attenuate fibroblast activation, we queried IPF gene signatures against a library of small-molecule-induced gene-expression profiles and identified Hsp90 inhibitors as potential therapeutic agents that can suppress fibroblast activation in IPF. Although Hsp90 is a molecular chaperone that regulates multiple processes involved in fibroblast activation, it has not been previously proposed as a molecular target in IPF. Here, we found elevated Hsp90 staining in lung biopsies of patients with IPF. Notably, fibroblasts isolated from fibrotic lesions showed heightened Hsp90 ATPase activity compared with normal fibroblasts. 17-N-allylamino-17-demethoxygeldanamycin (17-AAG), a small-molecule inhibitor of Hsp90 ATPase activity, attenuated fibroblast activation and also TGF-β–driven effects on fibroblast to myofibroblast transformation. The loss of the Hsp90AB, but not the Hsp90AA isoform, resulted in reduced fibroblast proliferation, myofibroblast transformation, and ECM production. Finally, in vivo therapy with 17-AAG attenuated progression of established and ongoing fibrosis in a mouse model of pulmonary fibrosis, suggesting that targeting Hsp90 represents an effective strategy for the treatment of fibrotic lung disease. PMID:28239659

  17. Oral fibroblasts produce more HGF and KGF than skin fibroblasts in response to co-culture with keratinocytes

    DEFF Research Database (Denmark)

    Grøn, Birgitte; Stoltze, Kaj; Andersson, Anders

    2002-01-01

    The production of hepatocyte growth factor (HGF) and keratinocyte growth factor (KGF) in subepithelial fibroblasts from buccal mucosa, periodontal ligament, and skin was determined after co-culture with keratinocytes. The purpose was to detect differences between the fibroblast subpopulations...... that could explain regional variation in epithelial growth and wound healing. Normal human fibroblasts were cultured on polystyrene or maintained in collagen matrix and stimulated with keratinocytes cultured on membranes. The amount of HGF and KGF protein in the culture medium was determined every 24 h for 5...... days by ELISA. When cultured on polystyrene, the constitutive level of KGF and HGF in periodontal fibroblasts was higher than the level in buccal and skin fibroblasts. In the presence of keratinocytes, all three types of fibroblasts in general increased their HGF and KGF production 2-3 times. When...

  18. Immortalization of Werner syndrome and progeria fibroblasts

    Energy Technology Data Exchange (ETDEWEB)

    Saito, H.; Moses, R.E. (Baylor College of Medicine, Houston, TX (USA))

    1991-02-01

    Human fibroblast cells from two different progeroid syndromes, Werner syndrome (WS) and progeria, were established as immortalized cell lines by transfection with plasmid DNA containing the SV40 early region. The lineage of each immortalized cell line was confirmed by VNTR analysis. Each of the immortalized cell lines maintained its original phenotype of slow growth. DNA repair ability of these cells was also studied by measuring sensitivity to killing by uv or the DNA-damaging drugs methyl methansulfonate, bleomycin, and cis-dichlorodiamine platinum. The results showed that both WS and progeria cells have normal sensitivity to these agents.

  19. Gene Signature of Human Oral Mucosa Fibroblasts: Comparison with Dermal Fibroblasts and Induced Pluripotent Stem Cells.

    Science.gov (United States)

    Miyoshi, Keiko; Horiguchi, Taigo; Tanimura, Ayako; Hagita, Hiroko; Noma, Takafumi

    2015-01-01

    Oral mucosa is a useful material for regeneration therapy with the advantages of its accessibility and versatility regardless of age and gender. However, little is known about the molecular characteristics of oral mucosa. Here we report the first comparative profiles of the gene signatures of human oral mucosa fibroblasts (hOFs), human dermal fibroblasts (hDFs), and hOF-derived induced pluripotent stem cells (hOF-iPSCs), linking these with biological roles by functional annotation and pathway analyses. As a common feature of fibroblasts, both hOFs and hDFs expressed glycolipid metabolism-related genes at higher levels compared with hOF-iPSCs. Distinct characteristics of hOFs compared with hDFs included a high expression of glycoprotein genes, involved in signaling, extracellular matrix, membrane, and receptor proteins, besides a low expression of HOX genes, the hDFs-markers. The results of the pathway analyses indicated that tissue-reconstructive, proliferative, and signaling pathways are active, whereas senescence-related genes in p53 pathway are inactive in hOFs. Furthermore, more than half of hOF-specific genes were similarly expressed to those of hOF-iPSC genes and might be controlled by WNT signaling. Our findings demonstrated that hOFs have unique cellular characteristics in specificity and plasticity. These data may provide useful insight into application of oral fibroblasts for direct reprograming.

  20. Differences in motility pattern between human buccal fibroblasts and periodontal and skin fibroblasts

    DEFF Research Database (Denmark)

    Lepekhin, Eugene; Grøn, Birgitte; Berezin, Vladimir

    2002-01-01

    at these sites can be explained by differences in the motile behavior of their respective fibroblast populations. The migratory characteristics were studied in a two-dimensional culture system. The migration of single cells was time-lapse video recorded at intervals of 15 min for a period of 6 h using a computer...

  1. S100A12 Induced in the Epidermis by Reduced Hydration Activates Dermal Fibroblasts and Causes Dermal Fibrosis.

    Science.gov (United States)

    Zhao, Jingling; Zhong, Aimei; Friedrich, Emily E; Jia, Shengxian; Xie, Ping; Galiano, Robert D; Mustoe, Thomas A; Hong, Seok Jong

    2017-03-01

    Disruption of the barrier function of skin increases transepidermal water loss and up-regulates inflammatory pathways in the epidermis. Consequently, sustained expression of proinflammatory cytokines from the epidermis is associated with dermal scarring. We found increased expression of S100A12 in the epidermis of human hypertrophic and keloid scar. Exposing a stratified keratinocyte culture to a reduced-hydration environment increased the expression and secretion of S100A12 by nearly 70%, which in turn activated dermal fibroblasts in vitro. Direct treatment of fibroblasts with conditioned medium collected from stratified keratinocyte culture under reduced-hydration conditions activated fibroblasts, shown by up-regulation of α-smooth muscle actin, pro-collagen 1, and F-actin expression. However, this fibroblast activation was not found when S100A12 was knocked down by RNA interference in keratinocytes. Pharmacological blockade of S100A12 receptors, RAGE, or TLR4 inhibited S100A12-induced fibroblast activation. Local delivery of S100A12 resulted in a marked hypertrophic scar formation in a validated rabbit hypertrophic scar model compared with saline control. Our findings indicate that S100A12 functions as a proinflammatory cytokine and suggest that S100A12 is a potential therapeutic target for dermal scarring. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

  2. Replacement of murine fibroblasts by human fibroblasts irradiated in obtaining feeder layer for the culture of human keratinocytes

    Energy Technology Data Exchange (ETDEWEB)

    Yoshito, Daniele; Sufi, Bianca S.; Santin, Stefany P.; Mathor, Monica B. [Instituto de Pesquisas Energeticas e Nucleares (IPEN/CNEN-SP), Sao Paulo, SP (Brazil); Altran, Silvana C.; Isaac, Cesar [Universidade Sao Paulo (USP), Sao Paulo, SP (Brazil). Fac. de Medicina. Lab. de Microcirurgia Plastica; Esteves-Pedro, Natalia M. [Universidade Sao Paulo (USP), Sao Paulo, SP (Brazil). Fac. de Ciencias Farmaceuticas. Lab. de Controle Biologico; Herson, Marisa R. [DonorTissue Bank of Victoria (Australia)

    2011-07-01

    Human autologous epithelia cultivated in vitro, have been used successfully in treating damage to skin integrity. The methodology allowed the cultivation of these epithelia was described by Rheinwald and Green in 1975, this methodology consisted in seeding keratinocytes onto a feeder layer composed of lineage 3T3 murine fibroblasts, the proliferation rate is controlled through the action of ionizing radiation. However, currently there is a growing concern about the possibility of transmitting prions and murine viruses to transplanted patients. Taking into account this concern, in this present work, we replaced the feeder layer originally composed of murine fibroblasts by human fibroblasts. To obtain this new feeder layer was necessary to standardize the enough irradiation dose to inhibit the replication of human fibroblasts and the verification of effectiveness of the development of keratinocytes culture on a feeder layer thus obtained. According to the obtained results we can verify that the human fibroblasts irradiated at various tested doses (60, 70, 100, 200, 250 and 300 Gy) had their mitotic activity inactivated by irradiation, allowing the use of any of these doses to confection of the feeder layer, since these fibroblasts irradiated still showed viable until fourteen days of cultivation. In the test of colony formation efficiency was observed that keratinocytes seeded on irradiated human fibroblasts were able to develop satisfactorily, preserving their clonogenic potential. Therefore it was possible the replacement of murine fibroblasts by human fibroblasts in confection of the feeder layer, in order to eliminate this xenobiotic component of the keratinocytes culture. (author)

  3. Osteoblast, fibroblast and in vivo biological response to poly(vinylidene fluoride) based composite materials.

    Science.gov (United States)

    Costa, R; Ribeiro, C; Lopes, A C; Martins, P; Sencadas, V; Soares, R; Lanceros-Mendez, S

    2013-02-01

    Electroactive materials can be taken to advantage for the development of sensors and actuators as well as for novel tissue engineering strategies. Composites based on poly(vinylidene fluoride), PVDF, have been evaluated with respect to their biological response. Cell viability and proliferation were performed in vitro both with Mesenchymal Stem Cells differentiated to osteoblasts and Human Fibroblast Foreskin 1. In vivo tests were also performed using 6-week-old C57Bl/6 mice. It was concluded that zeolite and clay composites are biocompatible materials promoting cell response and not showing in vivo pro-inflammatory effects which renders both of them attractive for biological applications and tissue engineering, opening interesting perspectives to development of scaffolds from these composites. Ferrite and silver nanoparticle composites decrease osteoblast cell viability and carbon nanotubes decrease fibroblast viability. Further, carbon nanotube composites result in a significant increase in local vascularization accompanied an increase of inflammatory markers after implantation.

  4. Toll-like receptor 9 mediated responses in cardiac fibroblasts.

    Directory of Open Access Journals (Sweden)

    Ingrid Kristine Ohm

    Full Text Available Altered cardiac Toll-like receptor 9 (TLR9 signaling is important in several experimental cardiovascular disorders. These studies have predominantly focused on cardiac myocytes or the heart as a whole. Cardiac fibroblasts have recently been attributed increasing significance in mediating inflammatory signaling. However, putative TLR9-signaling through cardiac fibroblasts remains non-investigated. Thus, our aim was to explore TLR9-signaling in cardiac fibroblasts and investigate the consequence of such receptor activity on classical cardiac fibroblast cellular functions. Cultivated murine cardiac fibroblasts were stimulated with different TLR9 agonists (CpG A, B and C and assayed for the secretion of inflammatory cytokines (tumor necrosis factor α [TNFα], CXCL2 and interferon α/β. Expression of functional cardiac fibroblast TLR9 was proven as stimulation with CpG B and -C caused significant CXCL2 and TNFα-release. These responses were TLR9-specific as complete inhibition of receptor-stimulated responses was achieved by co-treatment with a TLR9-antagonist (ODN 2088 or chloroquine diphosphate. TLR9-stimulated responses were also found more potent in cardiac fibroblasts when compared with classical innate immune cells. Stimulation of cardiac fibroblasts TLR9 was also found to attenuate migration and proliferation, but did not influence myofibroblast differentiation in vitro. Finally, results from in vivo TLR9-stimulation with subsequent fractionation of specific cardiac cell-types (cardiac myocytes, CD45+ cells, CD31+ cells and cardiac fibroblast-enriched cell-fractions corroborated our in vitro data and provided evidence of differentiated cell-specific cardiac responses. Thus, we conclude that cardiac fibroblast may constitute a significant TLR9 responder cell within the myocardium and, further, that such receptor activity may impact important cardiac fibroblast cellular functions.

  5. Direct Conversion of Fibroblasts to Megakaryocyte Progenitors

    Directory of Open Access Journals (Sweden)

    Julian Pulecio

    2016-10-01

    Full Text Available Current sources of platelets for transfusion are insufficient and associated with risk of alloimmunization and blood-borne infection. These limitations could be addressed by the generation of autologous megakaryocytes (MKs derived in vitro from somatic cells with the ability to engraft and differentiate in vivo. Here, we show that overexpression of a defined set of six transcription factors efficiently converts mouse and human fibroblasts into MK-like progenitors. The transdifferentiated cells are CD41+, display polylobulated nuclei, have ploidies higher than 4N, form MK colonies, and give rise to platelets in vitro. Moreover, transplantation of MK-like murine progenitor cells into NSG mice results in successful engraftment and further maturation in vivo. Similar results are obtained using disease-corrected fibroblasts from Fanconi anemia patients. Our results combined demonstrate that functional MK progenitors with clinical potential can be obtained in vitro, circumventing the use of hematopoietic progenitors or pluripotent stem cells.

  6. Rac inhibition reverses the phenotype of fibrotic fibroblasts.

    Directory of Open Access Journals (Sweden)

    Shi-wen Xu

    Full Text Available BACKGROUND: Fibrosis, the excessive deposition of scar tissue by fibroblasts, is one of the largest groups of diseases for which there is no therapy. Fibroblasts from lesional areas of scleroderma patients possess elevated abilities to contract matrix and produce alpha-smooth muscle actin (alpha-SMA, type I collagen and CCN2 (connective tissue growth factor, CTGF. The basis for this phenomenon is poorly understood, and is a necessary prerequisite for developing novel, rational anti-fibrotic strategies. METHODS AND FINDINGS: Compared to healthy skin fibroblasts, dermal fibroblasts cultured from lesional areas of scleroderma (SSc patients possess elevated Rac activity. NSC23766, a Rac inhibitor, suppressed the persistent fibrotic phenotype of lesional SSc fibroblasts. NSC23766 caused a decrease in migration on and contraction of matrix, and alpha-SMA, type I collagen and CCN2 mRNA and protein expression. SSc fibroblasts possessed elevated Akt phosphorylation, which was also blocked by NSC23766. Overexpression of rac1 in normal fibroblasts induced matrix contraction and alpha-SMA, type I collagen and CCN2 mRNA and protein expression. Rac1 activity was blocked by PI3kinase/Akt inhibition. Basal fibroblast activity was not affected by NSC23766. CONCLUSION: Rac inhibition may be considered as a novel treatment for the fibrosis observed in SSc.

  7. Analysis of primary cilia in directional cell migration in fibroblasts

    DEFF Research Database (Denmark)

    Christensen, Søren Tvorup; Veland, Iben; Schwab, Albrecht

    2013-01-01

    Early studies of migrating fibroblasts showed that primary cilia orient in front of the nucleus and point toward the leading edge. Recent work has shown that primary cilia coordinate a series of signaling pathways critical to fibroblast cell migration during development and in wound healing. In p...

  8. Fibroblastic rheumatism: Scientific Letter | Kawtar | African Journal of ...

    African Journals Online (AJOL)

    Fibroblastic Rheumatism (FR) is a rare rheumatologic entity of unknown etiology. The pathophysiological mechanism involving fibroblast proliferation is characterized by symmetrical polyarthritis associated with sudden onset of cutaneous nodules, flexion contractures. Bone erosion can occur as the disease progresses and ...

  9. Growth modulation of fibroblasts by chitosan-polyvinyl pyrrolidone ...

    Indian Academy of Sciences (India)

    Wounds in adults and fetuses differ in their healing ability with respect to scar formation. In adults, wounds lacking the epidermis exhibit excess collagen production and scar formation. Fibroblasts synthesize and deposit a collagen rich extracellular matrix. The early migration and proliferation of fibroblasts in the wound area ...

  10. Myogenic conversion of bladder fibroblasts by construction and ...

    African Journals Online (AJOL)

    Gene therapy of detrusor underactivity, by autologous cells transplantation, is limited by the number of primary myogenic. The purpose of this study was to investigate whether Myod1 could induce primary bladder fibroblasts to undergo myogenic conversion. Primary bladder fibroblasts from Sprague-Daley rats were cultured ...

  11. Rho A and the Rho kinase pathway regulate fibroblast contraction: Enhanced contraction in constitutively active Rho A fibroblast cells

    Energy Technology Data Exchange (ETDEWEB)

    Nobe, Koji, E-mail: kojinobe@pharm.showa-u.ac.jp [Department of Pharmacology, School of Pharmaceutical Sciences, Showa University, Tokyo (Japan); Nobe, Hiromi [Department of Pharmacology, School of Pharmaceutical Sciences, Showa University, Tokyo (Japan); Department of Physical Therapy, Bunkyo-Gakuin University (Japan); Yoshida, Hiroko [Department of Pharmacology, School of Pharmaceutical Sciences, Showa University, Tokyo (Japan); Kolodney, Michael S. [Dermatology Division, Department of Medicine, UCLA, Los Angeles, CA (United States); Paul, Richard J. [Department of Molecular and Cellular Physiology, University of Cincinnati College of Medicine, Cincinnati, OH (United States); Honda, Kazuo [Department of Pharmacology, School of Pharmaceutical Sciences, Showa University, Tokyo (Japan)

    2010-08-20

    Research highlights: {yields} Mechanisms of fibroblast cell contraction in collagen matrix. {yields} Assessed an isometric force development using 3D-reconstituted-fibroblast fiber. {yields} Constitutively active Rho A induced the over-contraction of fibroblast cells. {yields} Rho A and Rho kinase pathway has a central role in fibroblast cell contraction. -- Abstract: Fibroblast cells play a central role in the proliferation phase of wound healing processes, contributing to force development. The intracellular signaling pathways regulating this non-muscle contraction are only partially understood. To study the relations between Rho A and contractile responses, constitutively active Rho A (CA-Rho A) fibroblast cells were reconstituted into fibers and the effects of calf serum (CS) on isometric force were studied. CS-induced force in CA-Rho A fibroblast fibers was twice as large as that in wild type (NIH 3T3) fibroblast fibers. During this response, the translocation of Rho A from the cytosol to the membrane was detected by Rho A activity assays and Western blot analysis. Pre-treatment with a Rho specific inhibitor (C3-exoenzyme) suppressed translocation as well as contraction. These results indicate that Rho A activation is essential for fibroblast contraction. The Rho kinase inhibitor ( (Y27632)) inhibited both NIH 3T3 and CA-Rho A fibroblast fiber contractions. Activation of Rho A is thus directly coupled with Rho kinase activity. We conclude that the translocation of Rho A from the cytosol to the membrane and the Rho kinase pathway can regulate wound healing processes mediated by fibroblast contraction.

  12. Influence of three laser wavelengths on human fibroblasts cell culture.

    Science.gov (United States)

    Crisan, Bogdan; Soritau, Olga; Baciut, Mihaela; Campian, Radu; Crisan, Liana; Baciut, Grigore

    2013-02-01

    Although experimental studies in vitro and vivo have been numerous, the effect of laser wavelength irradiation on human fibroblast cell culture is poorly understood. This emphasizes the need of additional cellular and molecular research into laser influence with low energy and power. The aim of this study was to assess the influence of three different laser wavelengths on the human skin fibroblasts cell culture. We wanted to evaluate if near infrared lasers had any influence in healing of wounds by stimulating mitochondrial activity of fibroblasts. The cells were irradiated using 830-, 980- and 2,940-nm laser wavelengths. The irradiated cells were incubated and their mitochondrial activity was assessed by the MTT assay at 24, 48 and 72 h. Simultaneously, an apoptosis assay was assessed on the irradiated fibroblasts. It can be concluded that laser light of the near-infrared region (830 and 980 nm) influences fibroblasts mitochondrial activity compared to the 2,940-nm wavelength which produces apoptosis.

  13. Hyaluronic acid production by irradiated human synovial fibroblasts

    Energy Technology Data Exchange (ETDEWEB)

    Yaron, M.; Yaron, I.; Levita, M.; Herzberg, M.

    1977-03-01

    Radioactive particles as well as x irradiation from an external source has been used in the treatment of rheumatoid arthritis, osteoarthritis, and ankylosing spondylitis. In order to clarify effects of ionizing irradiation on synovial cells, radioactive gold (/sup 198/Au) and yttrium (/sup 90/Y) were added to fibroblast cultures derived from human synovial membranes. Other cultures were irradiated by a Picker x-ray machine. Fibroblast growth and hyaluronic acid production were measured. Radioactive gold and yttrium particles induced a significant increase of hyaluronic acid synthesis rate (pg/cell/day) and inhibited fibroblast growth. Fibroblasts continued to overproduce hyaluronic acid and to show growth inhibition 3 weeks after irradiation with radioactive gold. Hydrocortisone inhibited hyaluronic acid overproduction induced by radioactive gold. Overproduction of hyaluronic acid induced by the x-ray machine was inhibited by hydrocortisone, actinomycin-D, and cycloheximide. Fibroblasts derived from normal and rheumatoid patients responded similarly to ionizing irradiation.

  14. Novel therapeutic strategies targeting fibroblasts and fibrosis in heart disease

    Science.gov (United States)

    Gourdie, Robert G.; Dimmeler, Stefanie; Kohl, Peter

    2016-01-01

    Our understanding of cardiac fibroblast functions has moved beyond their roles in heart structure and extracellular matrix generation, and now includes contributions to paracrine, mechanical and electrical signalling during ontogenesis and normal cardiac activity. Fibroblasts have central roles in pathogenic remodelling during myocardial ischaemia, hypertension and heart failure. As key contributors to scar formation, they are crucial for tissue repair after interventions including surgery and ablation. Novel experimental approaches targeting cardiac fibroblasts are promising potential therapies for heart disease. Indeed, several existing drugs act, at least partially, through effects on cardiac connective tissue. This Review outlines the origins and roles of fibroblasts in cardiac development, homeostasis and disease; illustrates the involvement of fibroblasts in current and emerging clinical interventions; and identifies future targets for research and development. PMID:27339799

  15. Bradykinin-induced asthmatic fibroblast/myofibroblast activities via bradykinin B2 receptor and different MAPK pathways

    NARCIS (Netherlands)

    Sabatini, Federica; Luppi, Fabrizio; Petecchia, Loredana; Stefano, Antonino Di; Longo, Anna M.; Eva, Alessandra; Vanni, Cristina; Hiemstra, Pieter S.; Sterk, Peter J.; Sorbello, Valentina; Fabbri, Leonardo M.; Rossi, Giovanni A.; Ricciardolo, Fabio L. M.

    2013-01-01

    Bradykinin drives normal lung fibroblasts into myofibroblasts, induces fibroblast proliferation and activates mitogen activated protein kinase pathways (MAPK) but its effects on bronchial fibroblasts from asthmatics (HBAFb) have not been yet studied. We studied bradykinin-induced fibroblast

  16. The impact of vascular endothelial growth factor and basic fibroblast growth factor on cardiac fibroblasts grown under altered gravity conditions

    DEFF Research Database (Denmark)

    Ulbrich, Claudia; Leder, Annekatrin; Pietsch, Jessica

    2010-01-01

    Myocardium is very sensitive to gravitational changes. During a spaceflight cardiovascular atrophy paired with rhythm problems and orthostatic intolerance can occur. The aim of this study was to investigate the impact of basic fibroblast growth factor (bFGF) and vascular endothelial growth factor...... (VEGF) on cardiac fibroblasts (CF) grown under altered gravity conditions....

  17. Nuclear targeting of IGF-1 receptor in orbital fibroblasts from Graves' disease: apparent role of ADAM17.

    Directory of Open Access Journals (Sweden)

    Neil Hoa

    Full Text Available Insulin-like growth factor-1 receptor (IGF-1R comprises two subunits, including a ligand binding domain on extra- cellular IGF-1Rα and a tyrosine phosphorylation site located on IGF-1Rβ. IGF-1R is over-expressed by orbital fibroblasts in the autoimmune syndrome, Graves' disease (GD. When activated by IGF-1 or GD-derived IgG (GD-IgG, these fibroblasts produce RANTES and IL-16, while those from healthy donors do not. We now report that IGF-1 and GD-IgG provoke IGF-1R accumulation in the cell nucleus of GD fibroblasts where it co-localizes with chromatin. Nuclear IGF-1R is detected with anti-IGF-1Rα-specific mAb and migrates to approximately 110 kDa, consistent with its identity as an IGF-1R fragment. Nuclear IGF-1R migrating as a 200 kDa protein and consistent with an intact receptor was undetectable when probed with either anti-IGF-1Rα or anti-IGF-1Rβ mAbs. Nuclear redistribution of IGF-1R is absent in control orbital fibroblasts. In GD fibroblasts, it can be abolished by an IGF-1R-blocking mAb, 1H7 and by physiological concentrations of glucocorticoids. When cell-surface IGF-1R is cross-linked with (125I IGF-1, (125I-IGF-1/IGF-1R complexes accumulate in the nuclei of GD fibroblasts. This requires active ADAM17, a membrane associated metalloproteinase, and the phosphorylation of IGF-1R. In contrast, virally encoded IGF-1Rα/GFP fusion protein localizes equivalently in nuclei in both control and GD fibroblasts. This result suggests that generation of IGF-1R fragments may limit the accumulation of nuclear IGF-1R. We thus identify a heretofore-unrecognized behavior of IGF-1R that appears limited to GD-derived fibroblasts. Nuclear IGF-1R may play a role in disease pathogenesis.

  18. Fibroblastic osteosarcoma in a lion (Panthera leo

    Directory of Open Access Journals (Sweden)

    L. Leonardi

    2014-01-01

    Full Text Available This report describes a case of spontaneous fibroblastic osteosarcoma in the humerus of a lion from a private park in Perugia, Italy. The tumor had an irregular, smooth, brown surface and a generally firm, rubbery consistence with gritty to hard areas interspersed. The mass was poorly vascularized with areas of necrosis at the periphery. The cut surface showed a multilobulated mass that had breached the humeral cortex, with periosteal production of reactive bone. The mass invaded the epiphysis, the synovial membrane, the joint capsule and ligaments. A mild hemorrhagic effusion appeared in the joint space. Clinical signs, gross and histopathologic findings are described in this rare case of a malignant bone tumor.

  19. A stromal address code defined by fibroblasts.

    Science.gov (United States)

    Parsonage, Greg; Filer, Andrew D; Haworth, Oliver; Nash, Gerard B; Rainger, G Ed; Salmon, Michael; Buckley, Christopher D

    2005-03-01

    To navigate into and within tissues, leukocytes require guidance cues that enable them to recognize which tissues to enter and which to avoid. Such cues are partly provided at the time of extravasation from blood by an endothelial address code on the luminal surface of the vascular endothelium. Here, we review the evidence that fibroblasts help define an additional stromal address code that directs leukocyte behaviour within tissues. We examine how this stromal code regulates site-specific leukocyte accumulation, differentiation and survival in a variety of physiological stromal niches, and how the aberrant expression of components of this code in the wrong tissue at the wrong time contributes to the persistence of chronic inflammatory diseases.

  20. Beryllium induces premature senescence in human fibroblasts.

    Science.gov (United States)

    Coates, Shannon S A; Lehnert, Bruce E; Sharma, Sunil; Kindell, Susan M; Gary, Ronald K

    2007-07-01

    After cells have completed a sufficient number of cell divisions, they exit the cell cycle and enter replicative senescence. Here, we report that beryllium causes proliferation arrest with premature expression of the principal markers of senescence. After young presenescent human fibroblasts were treated with 3 microM BeSO(4) for 24 h, p21 cyclin-dependent kinase inhibitor mRNA increased by >200%. Longer periods of exposure caused mRNA and protein levels to increase for both p21 and p16(Ink4a), a senescence regulator that prevents pRb-mediated cell cycle progression. BeSO(4) also caused dose-dependent induction of senescence-associated beta-galactosidase activity (SA-beta-gal). Untreated cells had 48 relative fluorescence units (RFU)/microg/h of SA-beta-gal, whereas 3 microM BeSO(4) caused activity to increase to 84 RFU/microg/h. In chromatin immunoprecipitation experiments, BeSO(4) caused p53 protein to associate with its DNA binding site in the promoter region of the p21 gene, indicating that p53 transcriptional activity is responsible for the large increase in p21 mRNA elicited by beryllium. Forced expression of human telomerase reverse transcriptase (hTERT) rendered HFL-1 cells incapable of normal replicative senescence. However, there was no difference in the responsiveness of normal HFL-1 fibroblasts (IC(50) = 1.9 microM) and hTERT-immortalized cells (IC(50) = 1.7 microM) to BeSO(4) in a 9-day proliferation assay. The effects of beryllium resemble those of histone deacetylase-inhibiting drugs, which also cause large increases in p21. However, beryllium produced no changes in histone acetylation, suggesting that Be(2+) acts as a novel and potent pharmacological inducer of premature senescence.

  1. Gene-specific mitochondria dysfunctions in human TARDBP and C9ORF72 fibroblasts.

    Science.gov (United States)

    Onesto, Elisa; Colombrita, Claudia; Gumina, Valentina; Borghi, Maria Orietta; Dusi, Sabrina; Doretti, Alberto; Fagiolari, Gigliola; Invernizzi, Federica; Moggio, Maurizio; Tiranti, Valeria; Silani, Vincenzo; Ratti, Antonia

    2016-05-05

    Dysregulation of RNA metabolism represents an important pathogenetic mechanism in both amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) due to the involvement of the DNA/RNA-binding proteins TDP-43 and FUS and, more recently, of C9ORF72. A potential link between dysregulation of RNA metabolism and mitochondrial dysfunction is recently emerged in TDP-43 disease models. To further investigate the possible relationship between these two pathogenetic mechanisms in ALS/FTD, we studied mitochondria functionality in human mutant TARDBP(p.A382T) and C9ORF72 fibroblasts grown in galactose medium to induce a switch from a glycolytic to an oxidative metabolism. In this condition we observed significant changes in mitochondria morphology and ultrastructure in both mutant cells with a fragmented mitochondria network particularly evident in TARDBP(p.A382T) fibroblasts. From analysis of the mitochondrial functionality, a decrease of mitochondria membrane potential with no alterations in oxygen consumption rate emerged in TARDBP fibroblasts. Conversely, an increased oxygen consumption and mitochondria hyperpolarization were observed in C9ORF72 fibroblasts in association to increased ROS and ATP content. We found evidence of autophagy/mitophagy in dynamic equilibrium with the biogenesis of novel mitochondria, particularly in mutant C9ORF72 fibroblasts where an increase of mitochondrial DNA content and mass, and of PGC1-α protein was observed. Our imaging and biochemical data show that wild-type and mutant TDP-43 proteins do not localize at mitochondria so that the molecular mechanisms responsible for such mitochondria impairment remain to be further elucidated. For the first time our findings assess a link between C9ORF72 and mitochondria dysfunction and indicate that mitochondria functionality is affected in TARDBP and C9ORF72 fibroblasts with gene-specific features in oxidative conditions. As in neuronal metabolism mitochondria are actively used for ATP

  2. Fibroblast growth factors as tissue repair and regeneration therapeutics

    Directory of Open Access Journals (Sweden)

    Quentin M. Nunes

    2016-01-01

    Full Text Available Cell communication is central to the integration of cell function required for the development and homeostasis of multicellular animals. Proteins are an important currency of cell communication, acting locally (auto-, juxta-, or paracrine or systemically (endocrine. The fibroblast growth factor (FGF family contributes to the regulation of virtually all aspects of development and organogenesis, and after birth to tissue maintenance, as well as particular aspects of organism physiology. In the West, oncology has been the focus of translation of FGF research, whereas in China and to an extent Japan a major focus has been to use FGFs in repair and regeneration settings. These differences have their roots in research history and aims. The Chinese drive into biotechnology and the delivery of engineered clinical grade FGFs by a major Chinese research group were important enablers in this respect. The Chinese language clinical literature is not widely accessible. To put this into context, we provide the essential molecular and functional background to the FGF communication system covering FGF ligands, the heparan sulfate and Klotho co-receptors and FGF receptor (FGFR tyrosine kinases. We then summarise a selection of clinical reports that demonstrate the efficacy of engineered recombinant FGF ligands in treating a wide range of conditions that require tissue repair/regeneration. Alongside, the functional reasons why application of exogenous FGF ligands does not lead to cancers are described. Together, this highlights that the FGF ligands represent a major opportunity for clinical translation that has been largely overlooked in the West.

  3. Regulation of corneal fibroblast morphology and collagen reorganization by extracellular matrix mechanical properties.

    Science.gov (United States)

    Karamichos, Dimitris; Lakshman, Neema; Petroll, W Matthew

    2007-11-01

    To investigate how extracellular matrix mechanical properties influence cell and matrix patterning in three-dimensional culture. Human corneal fibroblasts were seeded within 30 x 10 mm collagen matrices that were unconstrained (UN), fully constrained (CO) along the long axis by attaching the construct to two immobilized plastic bars, or partially constrained (PC) by allowing linear elastic displacement of one bar. After 24 hours, constructs were labeled with phalloidin and were imaged using fluorescent and reflected light (for collagen) confocal microscopy. Cell morphology and local collagen fibril density and alignment were measured using digital image processing. Corneal fibroblasts in UN matrices were less elongated (UN collagen density and the degree of cell/collagen coalignment were higher in constrained matrices (UN collagen were often observed between individual cells. These data suggest that cell spreading, alignment, and contractile force generation are directly influenced by the mechanical properties of the surrounding extracellular matrix (ECM). Corneal fibroblasts generally align and compact collagen parallel to the axis of greatest ECM stiffness. Mechanical cross-talk between adjacent cells leads to enhancement of matrix reorganization, and results in additional, more complex matrix patterning.

  4. Fatty acid transport protein 4 is the principal very long chain fatty acyl-CoA synthetase in skin fibroblasts.

    Science.gov (United States)

    Jia, Zhenzhen; Moulson, Casey L; Pei, Zhengtong; Miner, Jeffrey H; Watkins, Paul A

    2007-07-13

    Fatty acid transport protein 4 (FATP4) is a fatty acyl-CoA synthetase that preferentially activates very long chain fatty acid substrates, such as C24:0, to their CoA derivatives. To gain better insight into the physiological functions of FATP4, we established dermal fibroblast cell lines from FATP4-deficient wrinkle-free mice and wild type (w.t.) mice. FATP4 -/- fibroblasts had no detectable FATP4 protein by Western blot. Compared with w.t. fibroblasts, cells lacking FATP4 had an 83% decrease in C24:0 activation. Peroxisomal degradation of C24:0 was reduced by 58%, and rates of C24:0 incorporation into major phospholipid species (54-64% decrease), triacylglycerol (64% decrease), and cholesterol esters (58% decrease) were significantly diminished. Because these lipid metabolic processes take place in different subcellular organelles, we used immunofluorescence and Western blotting of subcellular fractions to investigate the distribution of FATP4 protein and measured enzyme activity in fractions from w.t. and FATP4 -/- fibroblasts. FATP4 protein and acyl-CoA synthetase activity localized to multiple organelles, including mitochondria, peroxisomes, endoplasmic reticulum, and the mitochondria-associated membrane fraction. We conclude that in murine skin fibroblasts, FATP4 is the major enzyme producing very long chain fatty acid-CoA for lipid metabolic pathways. Although FATP4 deficiency primarily affected very long chain fatty acid metabolism, mutant fibroblasts also showed reduced uptake of a fluorescent long chain fatty acid and reduced levels of long chain polyunsaturated fatty acids. FATP4-deficient cells also contained abnormal neutral lipid droplets. These additional defects indicate that metabolic abnormalities in these cells are not limited to very long chain fatty acids.

  5. (+)-Catechin protects dermal fibroblasts against oxidative stress-induced apoptosis

    Science.gov (United States)

    2014-01-01

    Background Oxidative stress has been suggested as a mechanism underlying skin aging, as it triggers apoptosis in various cell types, including fibroblasts, which play important roles in the preservation of healthy, youthful skin. Catechins, which are antioxidants contained in green tea, exert various actions such as anti-inflammatory, anti-bacterial, and anti-cancer actions. In this study, we investigated the effect of (+)-catechin on apoptosis induced by oxidative stress in fibroblasts. Methods Fibroblasts (NIH3T3) under oxidative stress induced by hydrogen peroxide (0.1 mM) were treated with either vehicle or (+)-catechin (0–100 μM). The effect of (+)-catechin on cell viability, apoptosis, phosphorylation of c-Jun terminal kinases (JNK) and p38, and activation of caspase-3 in fibroblasts under oxidative stress were evaluated. Results Hydrogen peroxide induced apoptotic cell death in fibroblasts, accompanied by induction of phosphorylation of JNK and p38 and activation of caspase-3. Pretreatment of the fibroblasts with (+)-catechin inhibited hydrogen peroxide-induced apoptosis and reduced phosphorylation of JNK and p38 and activation of caspase-3. Conclusion (+)-Catechin protects against oxidative stress-induced cell death in fibroblasts, possibly by inhibiting phosphorylation of p38 and JNK. These results suggest that (+)-catechin has potential as a therapeutic agent for the prevention of skin aging. PMID:24712558

  6. Early activation of fibroblasts during PDT treatment in leg ulcers.

    Science.gov (United States)

    Corsi, Alessandro; Lecci, Pier P; Bacci, Stefano; Cappugi, Pietro; Pimpinelli, Nicola

    2016-06-01

    This pilot study was aimed to assess the variations of some microscopical parameters in skin ulcers, caused by chronic venous insufficiency of the lower extremities (chronic leg ulcers), in 15 patients refractory to previous conventional treatments during photodynamic therapy (PDT). Samples of control, wounded and PDT treated skin were taken and analyzed by immunohistochemistry. The cellular infiltrate, as well as the thickness of epidermis, vascularization, mast cell and fibroblast numbers, were increased in chronic wounds as compared to healthy skin. After completion of PDT, fibroblasts appeared further increased in number. Mast cells, closely clustered with fibroblasts, also showed an increase in their numbers, degranulation index and expression of basic fibroblast growth factor. The present findings support a primary role of fibroblasts in the wound healing process upon PDT treatment, given their early and intense reaction to injury. Mast cells seem to play an accessory yet important role, on the basis of their number and degranulation index variations and expression of basic FGF. In addition, the clustering of mast cells with fibroblasts around blood vessels suggest that these cells may stimulate angiogenesis and, in parallel, fibroblasts to secrete extracellular matrix during PDT therapy.

  7. LIF Mediates Proinvasive Activation of Stromal Fibroblasts in Cancer

    Directory of Open Access Journals (Sweden)

    Jean Albrengues

    2014-06-01

    Full Text Available Signaling crosstalk between tumor cells and fibroblasts confers proinvasive properties to the tumor microenvironment. Here, we identify leukemia inhibitory factor (LIF as a tumor promoter that mediates proinvasive activation of stromal fibroblasts independent of alpha-smooth muscle actin (α-SMA expression. We demonstrate that a pulse of transforming growth factor β (TGF-β establishes stable proinvasive fibroblast activation by inducing LIF production in both fibroblasts and tumor cells. In fibroblasts, LIF mediates TGF-β-dependent actomyosin contractility and extracellular matrix remodeling, which results in collective carcinoma cell invasion in vitro and in vivo. Accordingly, carcinomas from multiple origins and melanomas display strong LIF upregulation, which correlates with dense collagen fiber organization, cancer cell collective invasion, and poor clinical outcome. Blockade of JAK activity by Ruxolitinib (JAK inhibitor counteracts fibroblast-dependent carcinoma cell invasion in vitro and in vivo. These findings establish LIF as a proinvasive fibroblast producer independent of α-SMA and may open novel therapeutic perspectives for patients with aggressive primary tumors.

  8. Ets2 in tumor fibroblasts promotes angiogenesis in breast cancer.

    Directory of Open Access Journals (Sweden)

    Julie A Wallace

    Full Text Available Tumor fibroblasts are active partners in tumor progression, but the genes and pathways that mediate this collaboration are ill-defined. Previous work demonstrates that Ets2 function in stromal cells significantly contributes to breast tumor progression. Conditional mouse models were used to study the function of Ets2 in both mammary stromal fibroblasts and epithelial cells. Conditional inactivation of Ets2 in stromal fibroblasts in PyMT and ErbB2 driven tumors significantly reduced tumor growth, however deletion of Ets2 in epithelial cells in the PyMT model had no significant effect. Analysis of gene expression in fibroblasts revealed a tumor- and Ets2-dependent gene signature that was enriched in genes important for ECM remodeling, cell migration, and angiogenesis in both PyMT and ErbB2 driven-tumors. Consistent with these results, PyMT and ErbB2 tumors lacking Ets2 in fibroblasts had fewer functional blood vessels, and Ets2 in fibroblasts elicited changes in gene expression in tumor endothelial cells consistent with this phenotype. An in vivo angiogenesis assay revealed the ability of Ets2 in fibroblasts to promote blood vessel formation in the absence of tumor cells. Importantly, the Ets2-dependent gene expression signatures from both mouse models were able to distinguish human breast tumor stroma from normal stroma, and correlated with patient outcomes in two whole tumor breast cancer data sets. The data reveals a key function for Ets2 in tumor fibroblasts in signaling to endothelial cells to promote tumor angiogenesis. The results highlight the collaborative networks that orchestrate communication between stromal cells and tumor cells, and suggest that targeting tumor fibroblasts may be an effective strategy for developing novel anti-angiogenic therapies.

  9. Bacterial lipopolysaccharide promotes profibrotic activation of intestinal fibroblasts.

    LENUS (Irish Health Repository)

    Burke, J P

    2012-02-01

    BACKGROUND: Fibroblasts play a critical role in intestinal wound healing. Lipopolysaccharide (LPS) is a cell wall component of commensal gut bacteria. The effects of LPS on intestinal fibroblast activation were characterized. METHODS: Expression of the LPS receptor, toll-like receptor (TLR) 4, was assessed in cultured primary human intestinal fibroblasts using flow cytometry and confocal microscopy. Fibroblasts were treated with LPS and\\/or transforming growth factor (TGF) beta1. Nuclear factor kappaB (NFkappaB) pathway activation was assessed by inhibitory kappaBalpha (IkappaBalpha) degradation and NFkappaB promoter activity. Fibroblast contractility was measured using a fibroblast-populated collagen lattice. Smad-7, a negative regulator of TGF-beta1 signalling, and connective tissue growth factor (CTGF) expression were assessed using reverse transcriptase-polymerase chain reaction and western blot. The NFkappaB pathway was inhibited by IkappaBalpha transfection. RESULTS: TLR-4 was present on the surface of intestinal fibroblasts. LPS treatment of fibroblasts induced IkappaBalpha degradation, enhanced NFkappaB promoter activity and increased collagen contraction. Pretreatment with LPS (before TGF-beta1) significantly increased CTGF production relative to treatment with TGF-beta1 alone. LPS reduced whereas TGF-beta1 increased smad-7 expression. Transfection with an IkappaBalpha plasmid enhanced basal smad-7 expression. CONCLUSION: Intestinal fibroblasts express TLR-4 and respond to LPS by activating NFkappaB and inducing collagen contraction. LPS acts in concert with TGF-beta1 to induce CTGF. LPS reduces the expression of the TGF-beta1 inhibitor, smad-7.

  10. Expression of factors involved in dental pulp physiopathological processes by nemotic human pulpal fibroblasts.

    Science.gov (United States)

    Le Clerc, J; Tricot-Doleux, S; Pellen-Mussi, P; Pérard, M; Jeanne, S; Pérez, F

    2017-03-10

    To investigate in human dental pulp fibroblasts (HDPF) the expression of factors involved in dental pulp physiopathological processes and in an experimental model of cell activation called nemosis, and to compare the behaviour of pulp cell activation with sound lung fibroblast MRC5, employed as a reference model for nemosis. Nemotic response was induced in three-dimensional cultures of HDPF and lung fibroblasts. The expressions of molecules involved in physiological (alkaline phosphatase, type I collagen) and in inflammatory processes (IL-6, CXCL8, CCL20, COX-2) were studied using real-time PCR. Concentrations of IL-6 and CXCL8 were analysed during 4 days with ELISA. Nonparametric tests were used to determine statistical differences between groups. A significant decrease (P MRC5 and HDPF nemotic responses. Although the amounts of mRNA differed between these cell types, there was an increase in CCL20, CXCL8 and COX-2 expression (P MRC5 spheroids displayed significant amounts of IL-6 concentrations and mRNA expression. Notably, increased concentrations of CXCL8 were recorded in all three-dimensional cultures compared with monolayers as a function of time (P < 0.05). Although the nemotic responses observed were not identical in the pulpal and lung fibroblasts, similarities occurred in the expression of chemokines and cyclooxygenase-2. Nemotic reactions and inflammatory processes in pulp diseases share similarities in terms of the expression of factors. Thus, this in vitro model could constitute a powerful tool to study intercellular relations within the dental pulp and to develop new local treatments to counteract the inflammatory reaction that occurs during pulpitis. © 2017 International Endodontic Journal. Published by John Wiley & Sons Ltd.

  11. Alzheimer skin fibroblasts show increased susceptibility to free radicals.

    Science.gov (United States)

    Tesco, G; Latorraca, S; Piersanti, P; Piacentini, S; Amaducci, L; Sorbi, S

    1992-11-01

    We have studied the response to toxic oxygen metabolites of fibroblasts derived from skin biopsies of 5 patients with familial (FAD) and 4 with sporadic (AD) Alzheimer's disease compared with those derived from 4 normal controls. Fibroblasts were damaged by the generation of oxygen metabolites during the enzymatic oxidation of acetaldehyde by 50 munits of xanthine-oxidase (Xo). To quantify cell damage we measured lactate dehydrogenase (LDH) activity in the culture medium and cell viability in fibroblast cultures. We found a significant increase in LDH activity in the FAD vs. controls and also in the AD vs. controls.

  12. Cancer-associated fibroblasts promote proliferation of endometrial cancer cells.

    Directory of Open Access Journals (Sweden)

    Kavita S Subramaniam

    Full Text Available Endometrial cancer is the most commonly diagnosed gynecologic malignancy worldwide; yet the tumor microenvironment, especially the fibroblast cells surrounding the cancer cells, is poorly understood. We established four primary cultures of fibroblasts from human endometrial cancer tissues (cancer-associated fibroblasts, CAFs using antibody-conjugated magnetic bead isolation. These relatively homogenous fibroblast cultures expressed fibroblast markers (CD90, vimentin and alpha-smooth muscle actin and hormonal (estrogen and progesterone receptors. Conditioned media collected from CAFs induced a dose-dependent proliferation of both primary cultures and cell lines of endometrial cancer in vitro (175% when compared to non-treated cells, in contrast to those from normal endometrial fibroblast cell line (51% (P<0.0001. These effects were not observed in fibroblast culture derived from benign endometrial hyperplasia tissues, indicating the specificity of CAFs in affecting endometrial cancer cell proliferation. To determine the mechanism underlying the differential fibroblast effects, we compared the activation of PI3K/Akt and MAPK/Erk pathways in endometrial cancer cells following treatment with normal fibroblasts- and CAFs-conditioned media. Western blot analysis showed that the expression of both phosphorylated forms of Akt and Erk were significantly down-regulated in normal fibroblasts-treated cells, but were up-regulated/maintained in CAFs-treated cells. Treatment with specific inhibitors LY294002 and U0126 reversed the CAFs-mediated cell proliferation (P<0.0001, suggesting for a role of these pathways in modulating endometrial cancer cell proliferation. Rapamycin, which targets a downstream molecule in PI3K pathway (mTOR, also suppressed CAFs-induced cell proliferation by inducing apoptosis. Cytokine profiling analysis revealed that CAFs secrete higher levels of macrophage chemoattractant protein (MCP-1, interleukin (IL-6, IL-8, RANTES and vascular

  13. Low Dose Theophylline Showed an Inhibitory Effect on the Production of IL-6 and IL-8 in Primary Lung Fibroblast from Patients with COPD

    Directory of Open Access Journals (Sweden)

    Jing Zhang

    2012-01-01

    Full Text Available Chronic obstructive pulmonary disease (COPD is characterized by the abnormal and chronic lung inflammation. We hypothesized that lung fibroblasts could contribute to the local inflammation and investigated whether low dose theophylline had a beneficial effect on fibroblasts inflammation. Subjects undergoing lobectomy for bronchial carcinoma were enrolled and divided into COPD and control groups according to spirometry. Primary human lung fibroblasts were cultured from peripheral lung tissue distant to tumor tissue. There was a significant increase in both the mRNA expressions and protein levels for IL-6 and IL-8 in fibroblasts in COPD group, and the values were negatively correlated with lung function (P<0.05. For COPD fibroblasts, the protein levels of IL-6 and IL-8 decreased from 993.0 ± 738.9 pg/mL to 650.1 ± 421.9 pg/mL (P=0.014 and from 703.1 ± 278.0 pg/mL to 492.0 ± 214.9 pg/mL (P=0.001, respectively, with 5 μg/mL theophylline treatment. In addition, theophylline at the dose of 5 μg/mL reduced the increased production of IL-6 and IL-8 induced by 1 μg/mL LPS in primary human lung fibroblasts. Our data suggest that lung fibroblasts participate in the chronic inflammation in COPD by releasing IL-6 and IL-8, and low dose theophylline can alleviate the proinflammatory mediators’ production by fibroblasts.

  14. PTCH1 +/- dermal fibroblasts isolated from healthy skin of Gorlin syndrome patients exhibit features of carcinoma associated fibroblasts.

    Directory of Open Access Journals (Sweden)

    Alexandre Valin

    Full Text Available Gorlin's or nevoid basal cell carcinoma syndrome (NBCCS causes predisposition to basal cell carcinoma (BCC, the commonest cancer in adult human. Mutations in the tumor suppressor gene PTCH1 are responsible for this autosomal dominant syndrome. In NBCCS patients, as in the general population, ultraviolet exposure is a major risk factor for BCC development. However these patients also develop BCCs in sun-protected areas of the skin, suggesting the existence of other mechanisms for BCC predisposition in NBCCS patients. As increasing evidence supports the idea that the stroma influences carcinoma development, we hypothesized that NBCCS fibroblasts could facilitate BCC occurence of the patients. WT (n = 3 and NBCCS fibroblasts bearing either nonsense (n = 3 or missense (n = 3 PTCH1 mutations were cultured in dermal equivalents made of a collagen matrix and their transcriptomes were compared by whole genome microarray analyses. Strikingly, NBCCS fibroblasts over-expressed mRNAs encoding pro-tumoral factors such as Matrix Metalloproteinases 1 and 3 and tenascin C. They also over-expressed mRNA of pro-proliferative diffusible factors such as fibroblast growth factor 7 and the stromal cell-derived factor 1 alpha, known for its expression in carcinoma associated fibroblasts. These data indicate that the PTCH1(+/- genotype of healthy NBCCS fibroblasts results in phenotypic traits highly reminiscent of those of BCC associated fibroblasts, a clue to the yet mysterious proneness to non photo-exposed BCCs in NBCCS patients.

  15. Fibroblasts express immune relevant genes and are important sentinel cells during tissue damage in rainbow trout (Oncorhynchus mykiss.

    Directory of Open Access Journals (Sweden)

    Hans-Christian Ingerslev

    Full Text Available Fibroblasts have shown to be an immune competent cell type in mammals. However, little is known about the immunological functions of this cell-type in lower vertebrates. A rainbow trout hypodermal fibroblast cell-line (RTHDF was shown to be responsive to PAMPs and DAMPs after stimulation with LPS from E. coli, supernatant and debris from sonicated RTHDF cells. LPS was overall the strongest inducer of IL-1beta, IL-8, IL-10, TLR-3 and TLR-9. IL-1beta and IL-8 were already highly up regulated after 1 hour of LPS stimulation. Supernatant stimuli significantly increased the expression of IL-1beta, TLR-3 and TLR-9, whereas the debris stimuli only increased expression of IL-1beta. Consequently, an in vivo experiment was further set up. By mechanically damaging the muscle tissue of rainbow trout, it was shown that fibroblasts in the muscle tissue of rainbow trout contribute to electing a highly local inflammatory response following tissue injury. The damaged muscle tissue showed a strong increase in the expression of the immune genes IL-1beta, IL-8 and TGF-beta already 4 hours post injury at the site of injury while the expression in non-damaged muscle tissue was not influenced. A weaker, but significant response was also seen for TLR-9 and TLR-22. Rainbow trout fibroblasts were found to be highly immune competent with a significant ability to express cytokines and immune receptors. Thus fish fibroblasts are believed to contribute significantly to local inflammatory reactions in concert with the traditional immune cells.

  16. The radiation response of human dermal fibroblasts

    Science.gov (United States)

    Mitchell, Stephen Andrew

    A clinically reliable predictive assay based on normal-tissue radiosensitivity may lead to improved tumour control through individualised dose prescriptions. In-vitro fibroblast radiosensitivity has been shown, in several studies, to correlate with late radiation morbidity. The aim of this study was to investigate some of the cellular mechanisms underlying the normal-tissue response. In this study, seventeen primary fibroblast strains were established by enzymatic disaggregation of skin biopsies obtained from patients. These comprised seven who experienced acute tissue reactions to radiotherapy, four patients with a normal response and six non-cancer volunteers. An AT cell line was included as a positive control for radiosensitivity. In-vitro radiosensitivity was measured using a clonogenic assay at both high (HDR: 1.6 Gymin-1) and low dose rate (LDR: 0.01 Gymin-1). The radiation parameter HDR SF2 was the most sensitive in discriminating the seven sensitive patients from the remaining ten normal patients (range 0.11-0.19 sensitive patients compared with 0.17-0.34 control patients: pDNA damage. However, a strong correlation was found between clonogenic survival and both residual DNA damage (measured over 10-70 Gy, allowing 4 h repair, correlation coefficient: 0.90, DNA damage, with the sensitive cell lines generally showing a higher level of residual DNA damage. Cell-cycle delays were found in all 18 cell strains in response to 2 Gy irradiation, but were not found to discriminate between sensitive and normal patients. Associated studies found no mutations of the ATM gene in the five radiosensitive patients studied. However, a coding sequence alteration was found in the XRCC1 gene in one of the radiosensitive patients. These findings indicate that a DNA repair defect may be partly responsible for the extreme reactions to radiotherapy observed in a small percentage of patients and that with further modifications, an assay based on measurement of residual DNA damage

  17. Relationship among expression of basic-fibroblast growth factor ...

    African Journals Online (AJOL)

    Relationship among expression of basic-fibroblast growth factor, MTDH/Astrocyte elevated gene-1, adenomatous polyposis coli, matrix metalloproteinase 9,and COX-2 markers with prognostic factors in prostate carcinomas.

  18. Eugenol Toxicity in Human Dental Pulp Fibroblasts of Primary Teeth.

    Science.gov (United States)

    Escobar-García, Maria; Rodríguez-Contreras, Karen; Ruiz-Rodríguez, Socorro; Pierdant-Pérez, Mauricio; Cerda-Cristerna, Bernardino; Pozos-Guillén, Amaury

    2016-01-01

    The aim of the study was to determine the eugenol concentrations at which toxicity occurs in human dental pulp fibroblasts of primary teeth. Samples of primary dental pulp tissue were taken. Tissue samples were seeded by means of explant technique and used in the 4(th)-5th pass. Single Cell Gel Electrophoresis (Comet), phenazine MeThoSulfate (MTS), LIVE/DEAD Cell Viability/Toxicity and trypan blue assays for evaluation of the cytotoxicity of increasing concentrations of eugenol (0.06 to 810 μM) were performed. The results of toxicity tests showed toxic effects on dental pulp fibroblasts, even at very low concentrations of eugenol (0.06 μM). Very low concentrations of eugenol produce high toxicity in human dental pulp fibroblasts. All of the concentrations of eugenol that we evaluated produced high toxicity in human dental pulp fibroblasts of primary teeth.

  19. Improved Fibroblast Functionalities by Microporous Pattern Fabricated by Microelectromechanical Systems

    Science.gov (United States)

    Wei, Hongbo; Zhao, Lingzhou; Chen, Bangdao; Bai, Shizhu; Zhao, Yimin

    2014-01-01

    Fibroblasts, which play an important role in biological seal formation and maintenance, determine the long-term success of percutaneous implants. In this study, well-defined microporous structures with micropore diameters of 10–60 µm were fabricated by microelectromechanical systems and their influence on the fibroblast functionalities was observed. The results show that the microporous structures with micropore diameters of 10–60 µm did not influence the initial adherent fibroblast number; however, those with diameters of 40 and 50 µm improved the spread, actin stress fiber organization, proliferation and fibronectin secretion of the fibroblasts. The microporous structures with micropore diameters of 40–50 µm may be promising for application in the percutaneous part of an implant. PMID:25054322

  20. Fibroblast behavior after titanium surfaces exposure.

    Science.gov (United States)

    Danza, Matteo; Zollino, Ilaria; Candotto, Valentina; Cura, Francesca; Carinci, Francesco

    2012-12-01

    The main requirements for a good material are its ability to promote attraction and adhesion of bone precursor cells and their proliferation and differentiation. Different biocompatible materials are currently employed as scaffold. Among these, titanium is considered a gold standard because of its biocompatibility and good corrosion resistance. The aim of this work was to compare two different AoN titanium layers (GR4 and GR5) to investigate which one had a greater osteoconductive power using human fibroblasts (HFb) culture at two different time-points. The expression levels of some adhesion and traction-resistance related genes (COL11A1, COL2A1, COL9A1, DSP, ELN, HAS1, and TFRC) were analyzed using real time reverse transcription-polymerase chain reaction. After 7 days of treatment with TiA 4GR, the only two up-regulated genes were COL2A1 and DSP. After 15 days of treatment, none of genes over expressed. Our preliminary results suggest that neither AoN 4GR nor AoN 5GR are able to promote the production of protein involved in cell-cell and cell-matrix adhesion and in stress-resistance, required for a good outcome in dental implantology.

  1. The Fibroblast Growth Factor signaling pathway

    Science.gov (United States)

    Ornitz, David M; Itoh, Nobuyuki

    2015-01-01

    The signaling component of the mammalian Fibroblast Growth Factor (FGF) family is comprised of eighteen secreted proteins that interact with four signaling tyrosine kinase FGF receptors (FGFRs). Interaction of FGF ligands with their signaling receptors is regulated by protein or proteoglycan cofactors and by extracellular binding proteins. Activated FGFRs phosphorylate specific tyrosine residues that mediate interaction with cytosolic adaptor proteins and the RAS-MAPK, PI3K-AKT, PLCγ, and STAT intracellular signaling pathways. Four structurally related intracellular non-signaling FGFs interact with and regulate the family of voltage gated sodium channels. Members of the FGF family function in the earliest stages of embryonic development and during organogenesis to maintain progenitor cells and mediate their growth, differentiation, survival, and patterning. FGFs also have roles in adult tissues where they mediate metabolic functions, tissue repair, and regeneration, often by reactivating developmental signaling pathways. Consistent with the presence of FGFs in almost all tissues and organs, aberrant activity of the pathway is associated with developmental defects that disrupt organogenesis, impair the response to injury, and result in metabolic disorders, and cancer. © 2015 Wiley Periodicals, Inc. PMID:25772309

  2. Rosmarinic acid potentiates carnosic acid induced apoptosis in lung fibroblasts

    OpenAIRE

    Bahri, Sana; Mies, Fr?d?rique; Ben Ali, Ridha; Mlika, Mona; Jameleddine, Saloua; Mc Entee, Kathleen; Shlyonsky, Vadim

    2017-01-01

    Pulmonary fibrosis is characterized by over-population and excessive activation of fibroblasts and myofibroblasts disrupting normal lung structure and functioning. Rosemary extract rich in carnosic acid (CA) and rosmarinic acid (RA) was reported to cure bleomycin-(BLM)-induced pulmonary fibrosis. We demonstrate that CA decreased human lung fibroblast (HLF) viability with IC50 value of 17.13?1.06 ?M, while RA had no cytotoxic effect. In the presence of 50 ?M of RA, dose-response for CA shifted...

  3. Abscisic acid ameliorates the systemic sclerosis fibroblast phenotype in vitro

    Energy Technology Data Exchange (ETDEWEB)

    Bruzzone, Santina, E-mail: santina.bruzzone@unige.it [Department of Experimental Medicine, Section of Biochemistry, University of Genova, Viale Benedetto XV 1, 16132 Genova (Italy); Centre of Excellence for Biomedical Research, University of Genova, Viale Benedetto XV 9, 16132 Genova (Italy); Advanced Biotechnology Center, Largo Rosanna Benzi 10, 16132 Genova (Italy); Battaglia, Florinda [Centre of Excellence for Biomedical Research, University of Genova, Viale Benedetto XV 9, 16132 Genova (Italy); Mannino, Elena [Department of Experimental Medicine, Section of Biochemistry, University of Genova, Viale Benedetto XV 1, 16132 Genova (Italy); Parodi, Alessia [Centre of Excellence for Biomedical Research, University of Genova, Viale Benedetto XV 9, 16132 Genova (Italy); Fruscione, Floriana [Department of Experimental Medicine, Section of Biochemistry, University of Genova, Viale Benedetto XV 1, 16132 Genova (Italy); Advanced Biotechnology Center, Largo Rosanna Benzi 10, 16132 Genova (Italy); Basile, Giovanna [Department of Experimental Medicine, Section of Biochemistry, University of Genova, Viale Benedetto XV 1, 16132 Genova (Italy); Salis, Annalisa; Sturla, Laura [Department of Experimental Medicine, Section of Biochemistry, University of Genova, Viale Benedetto XV 1, 16132 Genova (Italy); Centre of Excellence for Biomedical Research, University of Genova, Viale Benedetto XV 9, 16132 Genova (Italy); Negrini, Simone; Kalli, Francesca; Stringara, Silvia [Centre of Excellence for Biomedical Research, University of Genova, Viale Benedetto XV 9, 16132 Genova (Italy); Filaci, Gilberto [Centre of Excellence for Biomedical Research, University of Genova, Viale Benedetto XV 9, 16132 Genova (Italy); Department of Internal Medicine, Viale Benedetto XV 6, 16132 Genova (Italy); and others

    2012-05-25

    Highlights: Black-Right-Pointing-Pointer ABA is an endogenous hormone in humans, regulating different cell responses. Black-Right-Pointing-Pointer ABA reverts some of the functions altered in SSc fibroblasts to a normal phenotype. Black-Right-Pointing-Pointer UV-B irradiation increases ABA content in SSc cultures. Black-Right-Pointing-Pointer SSc fibroblasts could benefit from exposure to ABA and/or to UV-B. -- Abstract: The phytohormone abscisic acid (ABA) has been recently identified as an endogenous hormone in humans, regulating different cell functions, including inflammatory processes, insulin release and glucose uptake. Systemic sclerosis (SSc) is a chronic inflammatory disease resulting in fibrosis of skin and internal organs. In this study, we investigated the effect of exogenous ABA on fibroblasts obtained from healthy subjects and from SSc patients. Migration of control fibroblasts induced by ABA was comparable to that induced by transforming growth factor-{beta} (TGF-{beta}). Conversely, migration toward ABA, but not toward TGF-{beta}, was impaired in SSc fibroblasts. In addition, ABA increased cell proliferation in fibroblasts from SSc patients, but not from healthy subjects. Most importantly, presence of ABA significantly decreased collagen deposition by SSc fibroblasts, at the same time increasing matrix metalloproteinase-1 activity and decreasing the expression level of tissue inhibitor of metalloproteinase (TIMP-1). Thus, exogenously added ABA appeared to revert some of the functions altered in SSc fibroblasts to a normal phenotype. Interestingly, ABA levels in plasma from SSc patients were found to be significantly lower than in healthy subjects. UV-B irradiation induced an almost 3-fold increase in ABA content in SSc cultures. Altogether, these results suggest that the fibrotic skin lesions in SSc patients could benefit from exposure to high(er) ABA levels.

  4. Regulating Cancer-Associated Fibroblast Biology in Prostate Cancer

    Science.gov (United States)

    2017-10-01

    AWARD NUMBER: W81XWH-15-1-0512 TITLE: Regulating Cancer-Associated Fibroblast Biology in Prostate Cancer PRINCIPAL INVESTIGATOR: Andrew...SUBTITLE 5a. CONTRACT NUMBER Regulating Cancer-Associated Fibroblast Biology in Prostate Cancer 5b. GRANT NUMBER W81XWH-15-1-0512 5c. PROGRAM...blocked by the addition of Pim inhibitors. These results suggest that the Pim protein kinase can regulate stromal cell biology to modulate epithelial

  5. Regulating Cancer Associated Fibroblast Biology in Prostate Cancer

    Science.gov (United States)

    2016-10-01

    SUPPLEMENTARY NOTES 14. ABSTRACT There is an urgent need to develop both new approaches to the treatment of prostate cancer. Analysis of human prostate...AWARD NUMBER: W81XWH-15-1-0512 TITLE: Regulating Cancer-Associated Fibroblast Biology in Prostate Cancer PRINCIPAL INVESTIGATOR: Andrew...CONTRACT NUMBER Regulating Cancer-Associated Fibroblast Biology in Prostate Cancer 5b. GRANT NUMBER W81XWH-15-1-0512 5c. PROGRAM ELEMENT NUMBER 6

  6. Small molecules enable highly efficient neuronal conversion of human fibroblasts.

    Science.gov (United States)

    Ladewig, Julia; Mertens, Jerome; Kesavan, Jaideep; Doerr, Jonas; Poppe, Daniel; Glaue, Finnja; Herms, Stefan; Wernet, Peter; Kögler, Gesine; Müller, Franz-Josef; Koch, Philipp; Brüstle, Oliver

    2012-06-01

    Forced expression of proneural transcription factors has been shown to direct neuronal conversion of fibroblasts. Because neurons are postmitotic, conversion efficiencies are an important parameter for this process. We present a minimalist approach combining two-factor neuronal programming with small molecule-based inhibition of glycogen synthase kinase-3β and SMAD signaling, which converts postnatal human fibroblasts into functional neuron-like cells with yields up to >200% and neuronal purities up to >80%.

  7. SPATIAL ORGANIZATION OF FIBROBLAST NUCLEAR CHROMOCENTERS: COMPONENT TREE ANALYSIS

    OpenAIRE

    Snapp, Robert R.; Goveia, Elyse; Peet, Lindsay; Bouffard, Nicole A; Badger, Gary J.; Langevin, Helene M

    2013-01-01

    The nuclei of mouse connective tissue fibroblasts contain chromocenters which are well-defined zones of heterochromatin that can be used as positional landmarks to examine nuclear remodeling in response to a mechanical perturbation. This study used component tree analysis, an image segmentation algorithm that detects high intensity voxels that are topologically connected, to quantify the spatial organization of chromocenters in fibroblasts within whole mouse connective tissue fixed and staine...

  8. FIBROBLAST CYTOSKELETAL REMODELING INDUCED BY TISSUE STRETCH INVOLVES ATP SIGNALING

    OpenAIRE

    Langevin, HM; Fujita, T.; Bouffard, NA; Takano, T; Koptiuch, C; Badger, GJ; Nedergaard, M

    2013-01-01

    Fibroblasts in whole areolar connective tissue respond to static stretching of the tissue by expanding and remodeling their cytoskeleton within minutes both ex vivo and in vivo. This study tested the hypothesis that the mechanism of fibroblast expansion in response to tissue stretch involves extracellular ATP signaling. In response to tissue stretch ex vivo, ATP levels in the bath solution increased significantly, and this increase was sustained for 20 minutes, returning to baseline at 60 min...

  9. Autophagy is required for IL-2-mediated fibroblast growth

    Energy Technology Data Exchange (ETDEWEB)

    Kang, Rui [Department of Surgery, University of Pittsburgh, Pittsburgh, Pennsylvania 15219 (United States); Tang, Daolin, E-mail: tangd2@upmc.edu [Department of Surgery, University of Pittsburgh, Pittsburgh, Pennsylvania 15219 (United States); Lotze, Michael T., E-mail: lotzemt@upcm.edu [Department of Surgery, University of Pittsburgh, Pittsburgh, Pennsylvania 15219 (United States); Zeh III, Herbert J., E-mail: zehh@upmc.edu [Department of Surgery, University of Pittsburgh, Pittsburgh, Pennsylvania 15219 (United States)

    2013-02-15

    Autophagy is an evolutionarily conserved pathway responsible for delivery of cytoplasmic material into the lysosomal degradation pathway to enable vesicular exocytosis. Interleukin (IL)-2 is produced by T-cells and its activity is important for immunoregulation. Fibroblasts are an immune competent cell type, playing a critical role in wound healing, chronic inflammation, and tumor development. Although autophagy plays an important role in each of these processes, whether it regulates IL-2 activity in fibroblasts is unknown. Here, we show that autophagy is required for IL-2-induced cell growth in fibroblasts. IL-2 significantly induced autophagy in mouse embryonic fibroblasts (MEFs) and primary lung fibroblasts. Autophagy inhibitors (e.g., 3-methylamphetamine and bafilomycin A1) or knockdown of ATG5 and beclin 1 blocked clinical grade IL-2-induced autophagy. Moreover, IL-2 induced HMGB1 cytoplasmic translocation in MEFs and promoted interaction between HMGB1 and beclin1, which is required for autophagy induction. Pharmacological and genetic inhibition of autophagy inhibited IL-2-induced cell proliferation and enhanced IL-2-induced apoptosis. These findings suggest that autophagy is an important pro-survival regulator for IL-2-induced cell growth in fibroblasts.

  10. Analysis of carnitine esters by radio-high performance liquid chromatography in cultured skin fibroblasts from patients with mitochondrial fatty acid oxidation disorders

    NARCIS (Netherlands)

    Schmidt-Sommerfeld, E.; Bobrowski, P. J.; Penn, D.; Rhead, W. J.; Wanders, R. J.; Bennett, M. J.

    1998-01-01

    Acylcarnitines are important diagnostic markers for inborn errors of fatty acid oxidation, but their analysis in body fluids may not always be reliable. Recently, disease-specific acylcarnitine profiles generated by cultured skin fibroblasts were reported to facilitate the diagnosis by localizing a

  11. Oncogenes induce the cancer-associated fibroblast phenotype: metabolic symbiosis and "fibroblast addiction" are new therapeutic targets for drug discovery.

    Science.gov (United States)

    Lisanti, Michael P; Martinez-Outschoorn, Ubaldo E; Sotgia, Federica

    2013-09-01

    Metabolic coupling, between mitochondria in cancer cells and catabolism in stromal fibroblasts, promotes tumor growth, recurrence, metastasis, and predicts anticancer drug resistance. Catabolic fibroblasts donate the necessary fuels (such as L-lactate, ketones, glutamine, other amino acids, and fatty acids) to anabolic cancer cells, to metabolize via their TCA cycle and oxidative phosphorylation (OXPHOS). This provides a simple mechanism by which metabolic energy and biomass are transferred from the host microenvironment to cancer cells. Recently, we showed that catabolic metabolism and "glycolytic reprogramming" in the tumor microenvironment are orchestrated by oncogene activation and inflammation, which originates in epithelial cancer cells. Oncogenes drive the onset of the cancer-associated fibroblast phenotype in adjacent normal fibroblasts via paracrine oxidative stress. This oncogene-induced transition to malignancy is "mirrored" by a loss of caveolin-1 (Cav-1) and an increase in MCT4 in adjacent stromal fibroblasts, functionally reflecting catabolic metabolism in the tumor microenvironment. Virtually identical findings were obtained using BRCA1-deficient breast and ovarian cancer cells. Thus, oncogene activation (RAS, NFkB, TGF-β) and/or tumor suppressor loss (BRCA1) have similar functional effects on adjacent stromal fibroblasts, initiating "metabolic symbiosis" and the cancer-associated fibroblast phenotype. New therapeutic strategies that metabolically uncouple oxidative cancer cells from their glycolytic stroma or modulate oxidative stress could be used to target this lethal subtype of cancers. Targeting "fibroblast addiction" in primary and metastatic tumor cells may expose a critical Achilles' heel, leading to disease regression in both sporadic and familial cancers.

  12. The influence of genistein on free radicals in normal dermal fibroblasts and keloid fibroblasts examined by EPR spectroscopy.

    Science.gov (United States)

    Jurzak, Magdalena; Ramos, Paweł; Pilawa, Barbara

    2017-01-01

    Normal and keloid fibroblasts were examined using X-band (9.3 GHz) electron paramagnetic resonance spectroscopy. The effect of genistein on the concentration of free radicals in both normal dermal and keloid fibroblasts after ultraviolet irradiation was investigated. The highest concentration of free radicals was seen in keloid fibroblasts, with normal fibroblasts containing a lower concentration. The concentration of free radicals in both normal and keloid fibroblasts was altered in a concentration-dependent manner by the presence of genistein. The change in intra-cellular free radical concentration after the ultraviolet irradiation of both normal and keloid fibroblasts is also discussed. The antioxidant properties of genistein, using its 1,1-Diphenyl-2-picrylhydrazyl (DPPH) free radical-scavenging activity as a model, were tested, and the effect of ultraviolet irradiation on its interaction with free radicals was examined. The electron paramagnetic resonance spectra of DPPH showed quenching by genistein. The interaction of genistein with DPPH free radicals in the absence of ultraviolet irradiation was shown to be slow, but this interaction was much faster under ultraviolet irradiation. Ultraviolet irradiation enhanced the free radical-scavenging activity of genistein.

  13. Fibroblast growth factor signaling in metabolic regulation

    Directory of Open Access Journals (Sweden)

    Vera eNies

    2016-01-01

    Full Text Available The prevalence of obesity is a growing health problem. Obesity is strongly associated with several comorbidities, such as non-alcoholic fatty liver disease, certain cancers, insulin resistance and type 2 diabetes, which all reduce life expectancy and life quality. Several drugs have been put forward in order to treat these diseases, but many of them have detrimental side effects. The unexpected role of the family of fibroblast growth factors in the regulation of energy metabolism provides new approaches to the treatment of metabolic diseases, and offers a valuable tool to gain more insight into metabolic regulation. The known beneficial effects of FGF19 and FGF21 on metabolism, together with recently discovered similar effects of FGF1 suggest that FGFs and their derivatives carry great potential as novel therapeutics to treat metabolic conditions. To facilitate the development of new therapies with improved targeting and minimal side effects, a better understanding of the molecular mechanism of action of FGFs is needed.In this review we will discuss what is currently known about the physiological roles of FGF signaling in tissues important for metabolic homeostasis. In addition, we will discuss current concepts regarding their pharmacological properties and effector tissues in the context of metabolic disease. Also the recent progress in the development of FGF variants will be reviewed. Our goal is to provide a comprehensive overview of the current concepts and consensuses regarding FGF signaling in metabolic health and disease, and to provide starting points for the development of FGF-based therapies against metabolic conditions.

  14. Binding, uptake, and release of nicotine by human gingival fibroblasts

    Energy Technology Data Exchange (ETDEWEB)

    Hanes, P.J.; Schuster, G.S.; Lubas, S. (Medical College of Georgia, Augusta (USA))

    1991-02-01

    Previous studies of the effects of nicotine on fibroblasts have reported an altered morphology and attachment of fibroblasts to substrates and disturbances in protein synthesis and secretion. This altered functional and attachment response may be associated with changes in the cell membrane resulting from binding of the nicotine, or to disturbances in cell metabolism as a result of high intracellular levels of nicotine. The purpose of the present study, therefore, was to (1) determine whether gingival fibroblasts bound nicotine and if any binding observed was specific or non-specific in nature; (2) determine whether gingival fibroblasts internalized nicotine, and if so, at what rate; (3) determine whether gingival fibroblasts also released nicotine back into the extracellular environment; and (4) if gingival fibroblasts release nicotine intact or as a metabolite. Cultures of gingival fibroblasts were prepared from gingival connective tissue biopsies. Binding was evaluated at 4{degree}C using a mixture of {sup 3}H-nicotine and unlabeled nicotine. Specific binding was calculated as the difference between {sup 3}H-nicotine bound in the presence and absence of unlabeled nicotine. The cells bound 1.44 (+/- 0.42) pmols/10(6) cells in the presence of unlabeled nicotine and 1.66 (+/- 0.55) pmols/10(6) cells in the absence of unlabeled nicotine. The difference was not significant. Uptake of nicotine was measured at 37{degree}C after treating cells with {sup 3}H-nicotine for time periods up to 4 hours. Uptake in pmols/10(6) cells was 4.90 (+/- 0.34) at 15 minutes, 8.30 (+/- 0.75) at 30 minutes, 12.28 (+/- 2.62) at 1 hour and 26.31 (+/- 1.15) at 4 hours.

  15. Isolation of fibroblasts and epithelial cells in bronchoalveolar lavage (BAL).

    Science.gov (United States)

    Pollock, Kathryn; Albares, Luke; Wendt, Chris; Hubel, Allison

    2013-04-01

    The long-term outcome of lung transplants is poor with 60%-70% of patients developing chronic rejection. Chronic rejection is manifested histologically by obliterative bronchiolitis with bronchiolitis obliterans syndrome (BOS), the clinical surrogate. Recent studies suggest that fibroblasts and epithelial cells present in bronchoalveolar lavage (BAL) may be a clinically relevant biomarker for BOS. The goal of this investigation was to develop a fast, repeatable method to individually isolate these low-frequency cell types. Fibroblasts and epithelial cells were isolated from BAL using attachment methods and the phenotype of the cells confirmed using immunostaining for vimentin (fibroblasts) and epithelial cell adhesion molecule (EpCAM, epithelial cells). Both fibroblasts and epithelial cells were isolated in every sample of BAL processed with the frequency of fibroblasts ranging from 0.03% to 0.48% and epithelial cells ranging from 0.05% to 1.5% of the total sample. Additional studies were performed using cytospins of cells after macrophages were depleted; cells exhibiting characteristics of both fibroblasts and epithelial cells were observed. The frequency of the cells of interest suggests that conventional methods of immunomagnetic isolation will not be effective in isolating these subpopulations. Finally, some of the low-frequency cells isolated via cytospin exhibit characteristics of epithelial to mesenchymal transition (which was not observed in plating incubations), indicating that the epithelial to mesenchymal cell transition fibroblasts may be nonadherent. In future studies, this technique and dataset may be of use to statistically correlate low-frequency cell type abundance to the onset and development of BOS.

  16. Functional screen of paracrine signals in breast carcinoma fibroblasts.

    Directory of Open Access Journals (Sweden)

    Gui Su

    Full Text Available Stromal fibroblasts actively participate in normal mammary gland homeostasis and in breast carcinoma growth and progression by secreting paracrine factors; however, little is known about the identity of paracrine mediators in individual patients. The purpose of this study was to characterize paracrine signaling pathways between breast carcinoma cells and breast carcinoma-associated fibroblasts (CAF or normal mammary fibroblasts (NF, respectively. CAF and NF were isolated from breast carcinoma tissue samples and adjacent normal mammary gland tissue of 28 patients. The fibroblasts were grown in 3D collagen gel co-culture with T47D human breast carcinoma cells and T47D cell growth was measured. CAF stimulated T47D cell growth to a significantly greater degree than NF. We detected a considerable inter-individual heterogeneity of paracrine interactions but identified FGF2, HB-EGF, heparanase-1 and SDF1 as factors that were consistently responsible for the activity of carcinoma-associated fibroblasts. CAF from low-grade but not high-grade carcinomas required insulin-like growth factor 1 and transforming growth factor beta 1 to stimulate carcinoma growth. Paradoxically, blocking of membrane-type 1 matrix metalloprotease stimulated T47D cell growth in co-culture with NF. The results were largely mirrored by treating the fibroblasts with siRNA oligonucleotides prior to co-culture, implicating the fibroblasts as principal production site for the secreted mediators. In summary, we identify a paracrine signaling network with inter-individual commonalities and differences. These findings have significant implications for the design of stroma-targeted therapies.

  17. Macrophages and fibroblasts during inflammation and tissue repair in models of organ regeneration

    Science.gov (United States)

    2017-01-01

    Abstract This review provides a concise summary of the changing phenotypes of macrophages and fibroblastic cells during the local inflammatory response, the onset of tissue repair, and the resolution of inflammation which follow injury to an organ. Both cell populations respond directly to damage and present coordinated sequences of activation states which determine the reparative outcome, ranging from true regeneration of the organ to fibrosis and variable functional deficits. Recent work with mammalian models of organ regeneration, including regeneration of full‐thickness skin, hair follicles, ear punch tissues, and digit tips, is summarized and the roles of local immune cells in these systems are discussed. New investigations of the early phase of amphibian limb and tail regeneration, including the effects of pro‐inflammatory and anti‐inflammatory agents, are then briefly discussed, focusing on the transition from the normally covert inflammatory response to the initiation of the regeneration blastema by migrating fibroblasts and the expression of genes for limb patterning. PMID:28616244

  18. Investigating NF-κB signaling in lung fibroblasts in 2D and 3D culture systems.

    Science.gov (United States)

    Htwe, Su Su; Harrington, Helen; Knox, Alan; Rose, Felicity; Aylott, Jonathan; Haycock, John W; Ghaemmaghami, Amir M

    2015-12-01

    Inflammatory respiratory diseases are amongst major global health challenges. Lung fibroblasts have been shown to play a key role in lung inflammatory responses. However, their exact role in initiation and maintenance of lung diseases has remained elusive partly due to the limited availability of physiologically relevant in vitro models. Therefore, developing new tools that enable investigating the molecular pathways (e.g. nuclear factor-kappa B (NF-κB) activation) that underpin inflammatory responses in fibroblasts could be a valuable resource for scientists working in this area of research. In order to investigate NF-κB activation in response to pro-inflammatory stimuli in real-time, we first developed two detection systems based on nuclear localization of NF-κB by immunostaining and luciferase reporter assay system. Furthermore using electrospun porous scaffolds, with similar geometry to human lung extracellular matrix, we developed 3D cultures of lung fibroblasts allowing comparing NF-κB activation in response to pro-inflammatory stimuli (i.e. TNF-α) in 2D and 3D. Our data clearly show that the magnitude of NF-κB activation in 2D cultures is substantially higher than 3D cultures. However, unlike 2D cultures, cells in the 3D model remained responsive to TNF-α at higher concentrations. The more subdued and wider dynamic range of NF-κB responses in 3D culture system was associated with a different expression pattern for TNF receptor I in 3D versus 2D cultures collectively reflecting a more in vivo like TNF receptor I expression and NF-κB activation pattern in the 3D system. Our data suggest that lung fibroblasts are actively involved in the pathogenesis of lung inflammation by activation of NF-κB signaling pathway. The 3D culture detection system provides a sensitive and biologically relevant tool for investigating different pro-inflammatory events involving lung fibroblasts.

  19. Degranulating mast cells in fibrotic regions of human tumors and evidence that mast cell heparin interferes with the growth of tumor cells through a mechanism involving fibroblasts

    Directory of Open Access Journals (Sweden)

    Kanakubo Emi

    2005-09-01

    Full Text Available Abstract Background The purpose of this study was to test the hypothesis that mast cells that are present in fibrotic regions of cancer can suppress the growth of tumor cells through an indirect mechanism involving peri-tumoral fibroblasts. Methods We first immunostained a wide variety of human cancers for the presence of degranulated mast cells. In a subsequent series of controlled in vitro experiments, we then co-cultured UACC-812 human breast cancer cells with normal fibroblasts in the presence or absence of different combinations and doses of mast cell tryptase, mast cell heparin, a lysate of the human mast cell line HMC-1, and fibroblast growth factor-7 (FGF-7, a powerful, heparin-binding growth factor for breast epithelial cells. Results Degranulating mast cells were localized predominantly in the fibrous tissue of every case of breast cancer, head and neck cancer, lung cancer, ovarian cancer, non-Hodgkin's lymphoma, and Hodgkin's disease that we examined. Mast cell tryptase and HMC-1 lysate had no significant effect on the clonogenic growth of cancer cells co-cultured with fibroblasts. By contrast, mast cell heparin at multiple doses significantly reduced the size and number of colonies of tumor cells co-cultured with fibroblasts, especially in the presence of FGF-7. Neither heparin nor FGF-7, individually or in combination, produced any significant effect on the clonogenic growth of breast cancer cells cultured without fibroblasts. Conclusion Degranulating mast cells are restricted to peri-tumoral fibrous tissue, and mast cell heparin is a powerful inhibitor of clonogenic growth of tumor cells co-cultured with fibroblasts. These results may help to explain the well-known ability of heparin to inhibit the growth of primary and metastatic tumors.

  20. Site-Specific Keloid Fibroblasts Alter the Behaviour of Normal Skin and Normal Scar Fibroblasts through Paracrine Signalling

    Science.gov (United States)

    Bayat, Ardeshir

    2013-01-01

    Keloid disease (KD) is an abnormal cutaneous fibroproliferative disorder of unknown aetiopathogenesis. Keloid fibroblasts (KF) are implicated as mediators of elevated extracellular matrix deposition. Aberrant secretory behaviour by KF relative to normal skin fibroblasts (NF) may influence the disease state. To date, no previous reports exist on the ability of site-specific KF to induce fibrotic-like phenotypic changes in NF or normal scar fibroblasts (NS) by paracrine mechanisms. Therefore, the aim of this study was to investigate the influence of conditioned media from site-specific KF on the cellular and molecular behaviour of both NF and NS enabled by paracrine mechanisms. Conditioned media was collected from cultured primary fibroblasts during a proliferative log phase of growth including: NF, NS, peri-lesional keloid fibroblasts (PKF) and intra-lesional keloid fibroblasts (IKF). Conditioned media was used to grow NF, NS, PKF and IKF cells over 240 hrs. Cellular behavior was monitored through real time cell analysis (RTCA), proliferation rates and migration in a scratch wound assay. Fibrosis-associated marker expression was determined at both protein and gene level. PKF conditioned media treatment of both NF and NS elicited enhanced cell proliferation, spreading and viability as measured in real time over 240 hrs versus control conditioned media. Following PKF and IKF media treatments up to 240 hrs, both NF and NS showed significantly elevated proliferation rates (pmedia from growing marginal PKF elicited the strongest effects. In conclusion, primary NF and NS cells treated with PKF or IKF conditioned media exhibit enhanced expression of fibrosis-associated molecular markers and increased cellular activity as a result of keloid fibroblast-derived paracrine factors. PMID:24348987

  1. Orthodontic adhesives induce human gingival fibroblast toxicity and inflammation.

    Science.gov (United States)

    Huang, Tsui-Hsien; Liao, Pao-Hsin; Li, Han Yu; Ding, Shinn Jyh; Yen, Min; Kao, Chia-Tze

    2008-05-01

    To test the null hypothesis that the resin base and the resin hybrid glass ionomer base adhesives do not cause inflammation after contacting primary human gingival fibroblasts in vitro. The resin base and resin hybrid glass ionomer base adhesives were used to treat human gingival fibroblasts to evaluate the survival rate using MTT colorimetric assay to detect the level of cyclooxygenase-2 (COX-2) mRNA by reverse transcription polymerase chain reaction (RT-PCR) technique and COX-2 protein expression using Western blot analysis. The results were analyzed using one-way analysis of variance (ANOVA). Tests of differences of the treatments were analyzed using the Tukey test and a value of P adhesive and the liquid of glass ionomer adhesive showed decreasing survival rates after 24 hours of treatment (P adhesives induced COX-2 protein expression in human gingival fibroblasts. The exposure of quiescent human gingival fibroblasts to adhesives resulted in the induction of COX-2 mRNA expression. The investigations of the time-dependent COX-2 mRNA expression in adhesive-treated human gingival fibroblasts revealed different patterns. The hypothesis is rejected. For orthodontic patients with gingival inflammation, except for those with oral hygiene problems, the activation of COX-2 expression by orthodontic adhesive may be one of the potential mechanisms.

  2. Cultured Human Fibroblast Biostimulation Using a 940 nm Diode Laser.

    Science.gov (United States)

    Illescas-Montes, Rebeca; Melguizo-Rodríguez, Lucía; Manzano-Moreno, Francisco Javier; García-Martínez, Olga; Ruiz, Concepción; Ramos-Torrecillas, Javier

    2017-07-13

    Fibroblasts are the main cells involved in regeneration during wound healing. The objective was to determine the effect of 940 nm diode laser on cultured human fibroblasts using different irradiation regimens. The CCD-1064Sk human epithelial fibroblast cell line was treated with a 940 nm diode laser at different energy doses (power: 0.2-1 W and energy density: 1-7 J/cm²) using different transmission modes (continuous or pulsed). The effect on cell growth at 24 and 72 h post-treatment was examined by measuring the proliferative capacity, the impact on the cell cycle, and the effect on cell differentiation. fibroblast proliferative capacity was increased at 24 and 72 h post-treatment as a function of the energy dose. The greatest increase was observed with a power of 0.2 or 0.5 W and energy density between 1 and 4 J/cm²; no difference was observed between continuous and pulsed modes. There were no significant differences in cell cycle between treated groups and controls. α-actin expression was increased by treatment, indicating enhanced cell differentiation. The 940 nm diode laser has biostimulating effects on fibroblasts, stimulating proliferative capacity and cell differentiation without altering the cell cycle. Further researches are necessary to explore its potential clinical usefulness in wound healing.

  3. Androgen receptor characteristics in skin fibroblasts from hirsute women.

    Science.gov (United States)

    Eil, C; Cutler, G B; Loriaux, D L

    1985-01-01

    Hormonal measurements in some women with hirsutism often reveal little or no elevation in androgen levels to explain the disorder. Thus, it has been postulated that increased sensitivity of the hair follicle to androgen may contribute to the development of hirsutism in such patients. We, therefore, sought androgen receptor abnormalities in skin fibroblasts cultured from 10 hirsute women (ages 17-43) and normal or mildly elevated plasma testosterone levels (28-82 ng/dl). Androgen receptor content (Ro) and binding affinity (Kd) in cultured pubic skin fibroblasts were measured using a dispersed, whole cell assay. Ten such cell lines from these women were compared with 19 pubic skin cell lines from 9 normal volunteers (6 males and 3 females) and from 10 other subjects (males with gynecomastia or hypospadias). There was no statistically significant difference in the mean androgen receptor content (11,600 +/- 2700 (SE) sites/cell fibroblasts vs 7900 +/- 700 sites/cell or binding affinity (2.0 +/- 0.3 (SE) X 10(-9) M vs 1.5 +/- 0.2 X 10(-9) M, respectively) between the patients' fibroblasts and those of the controls. We conclude that hirsutism cannot be explained by abnormalities in fibroblast androgen receptor number or affinity. These observations do not exclude the possibility that other mechanisms might lead to increased peripheral androgen sensitivity in such patients.

  4. Dysregulated proinflammatory and fibrogenic phenotype of fibroblasts in cystic fibrosis.

    Directory of Open Access Journals (Sweden)

    François Huaux

    Full Text Available Morbi-mortality in cystic fibrosis (CF is mainly related to chronic lung infection and inflammation, uncontrolled tissue rearrangements and fibrosis, and yet the underlying mechanisms remain largely unknown. We evaluated inflammatory and fibrosis responses to bleomycin in F508del homozygous and wild-type mice, and phenotype of fibroblasts explanted from mouse lungs and skin. The effect of vardenafil, a cGMP-specific phosphodiesterase type 5 inhibitor, was tested in vivo and in culture. Responses of proinflammatory and fibrotic markers to bleomycin were enhanced in lungs and skin of CF mice and were prevented by treatment with vardenafil. Purified lung and skin fibroblasts from CF mice proliferated and differentiated into myofibroblasts more prominently and displayed higher sensitivity to growth factors than those recovered from wild-type littermates. Under inflammatory stimulation, mRNA and protein expression of proinflammatory mediators were higher in CF than in wild-type fibroblasts, in which CFTR expression reached similar levels to those observed in other non-epithelial cells, such as macrophages. Increased proinflammatory responses in CF fibroblasts were reduced by half with submicromolar concentrations of vardenafil. Proinflammatory and fibrogenic functions of fibroblasts are upregulated in CF and are reduced by vardenafil. This study provides compelling new support for targeting cGMP signaling pathway in CF pharmacotherapy.

  5. Cultured Human Fibroblast Biostimulation Using a 940 nm Diode Laser

    Directory of Open Access Journals (Sweden)

    Rebeca Illescas-Montes

    2017-07-01

    Full Text Available Background: Fibroblasts are the main cells involved in regeneration during wound healing. The objective was to determine the effect of 940 nm diode laser on cultured human fibroblasts using different irradiation regimens. Methods: The CCD-1064Sk human epithelial fibroblast cell line was treated with a 940 nm diode laser at different energy doses (power: 0.2–1 W and energy density: 1–7 J/cm2 using different transmission modes (continuous or pulsed. The effect on cell growth at 24 and 72 h post-treatment was examined by measuring the proliferative capacity, the impact on the cell cycle, and the effect on cell differentiation. Results: fibroblast proliferative capacity was increased at 24 and 72 h post-treatment as a function of the energy dose. The greatest increase was observed with a power of 0.2 or 0.5 W and energy density between 1 and 4 J/cm2; no difference was observed between continuous and pulsed modes. There were no significant differences in cell cycle between treated groups and controls. α-actin expression was increased by treatment, indicating enhanced cell differentiation. Conclusion: The 940 nm diode laser has biostimulating effects on fibroblasts, stimulating proliferative capacity and cell differentiation without altering the cell cycle. Further researches are necessary to explore its potential clinical usefulness in wound healing.

  6. Anatomic demarcation by positional variation in fibroblast gene expression programs.

    Directory of Open Access Journals (Sweden)

    John L Rinn

    2006-07-01

    Full Text Available Fibroblasts are ubiquitous mesenchymal cells with many vital functions during development, tissue repair, and disease. Fibroblasts from different anatomic sites have distinct and characteristic gene expression patterns, but the principles that govern their molecular specialization are poorly understood. Spatial organization of cellular differentiation may be achieved by unique specification of each cell type; alternatively, organization may arise by cells interpreting their position along a coordinate system. Here we test these models by analyzing the genome-wide gene expression profiles of primary fibroblast populations from 43 unique anatomical sites spanning the human body. Large-scale differences in the gene expression programs were related to three anatomic divisions: anterior-posterior (rostral-caudal, proximal-distal, and dermal versus nondermal. A set of 337 genes that varied according to these positional divisions was able to group all 47 samples by their anatomic sites of origin. Genes involved in pattern formation, cell-cell signaling, and matrix remodeling were enriched among this minimal set of positional identifier genes. Many important features of the embryonic pattern of HOX gene expression were retained in fibroblasts and were confirmed both in vitro and in vivo. Together, these findings suggest that site-specific variations in fibroblast gene expression programs are not idiosyncratic but rather are systematically related to their positional identities relative to major anatomic axes.

  7. Sporadic Alzheimer disease fibroblasts display an oxidative stress phenotype.

    Science.gov (United States)

    Ramamoorthy, Mahesh; Sykora, Peter; Scheibye-Knudsen, Morten; Dunn, Christopher; Kasmer, Cindy; Zhang, Yongqing; Becker, Kevin G; Croteau, Deborah L; Bohr, Vilhelm A

    2012-09-15

    Alzheimer disease (AD) is a major health problem in the United States, affecting one in eight Americans over the age of 65. The number of elderly suffering from AD is expected to continue to increase over the next decade, as the average age of the U.S. population increases. The risk factors for and etiology of AD are not well understood; however, recent studies suggest that exposure to oxidative stress may be a contributing factor. Here, microarray gene expression signatures were compared in AD-patient-derived fibroblasts and normal fibroblasts exposed to hydrogen peroxide or menadione (to simulate conditions of oxidative stress). Using the 23K Illumina cDNA microarray to screen expression of >14,000 human genes, we identified a total of 1017 genes that are chronically up- or downregulated in AD fibroblasts, 215 of which were also differentially expressed in normal human fibroblasts within 12h after exposure to hydrogen peroxide or menadione. Pathway analysis of these 215 genes and their associated pathways revealed cellular functions that may be critically dysregulated by oxidative stress and play a critical role in the etiology and/or pathology of AD. We then examined the AD fibroblasts for the presence of oxidative DNA damage and found increased accumulation of 8-oxo-guanine. These results indicate the possible role of oxidative stress in the gene expression profile seen in AD. Published by Elsevier Inc.

  8. Modeled Microgravity Affects Fibroblast Functions Related to Wound Healing

    Science.gov (United States)

    Cialdai, Francesca; Vignali, Leonardo; Morbidelli, Lucia; Colciago, Alessandra; Celotti, Fabio; Santi, Alice; Caselli, Anna; Cirri, Paolo; Monici, Monica

    2017-02-01

    Wound healing is crucial for the survival of an organism. Therefore, in the perspective of space exploration missions, it is important to understand if and how microgravity conditions affect the behavior of the cell populations involved in wound healing and the evolution of the process. Since fibroblasts are the major players in tissue repair, this study was focused on the behavior of fibroblasts in microgravity conditions, modeled by a RCCS. Cell cytoskeleton was studied by immunofluorescence microscopy, the ability to migrate was assessed by microchemotaxis and scratch assay, and the expression of markers of fibroblast activation, angiogenesis, and inflammation was assessed by western blot. Results revealed that after cell exposure to modeled microgravity conditions, a thorough rearrangement of microtubules occurred and α-SMA bundles were replaced by a tight network of faulty and disorganized filaments. Exposure to modeled microgravity induced a decrease in α-SMA and E-CAD expressions. Also, the expression of the pro-angiogenic protein VEGF decreased, while that of the inflammatory signal COX-2 increased. Fibroblast ability to adhere, migrate, and respond to chemoattractants (PRP), closely related to cytoskeleton integrity and membrane junctions, was significantly impaired. Nevertheless, PRP was able to partially restore fibroblast migration.

  9. Fibroblast cytoskeletal remodeling induced by tissue stretch involves ATP signaling.

    Science.gov (United States)

    Langevin, Helene M; Fujita, Takumi; Bouffard, Nicole A; Takano, Takahiro; Koptiuch, Cathryn; Badger, Gary J; Nedergaard, Maiken

    2013-09-01

    Fibroblasts in whole areolar connective tissue respond to static stretching of the tissue by expanding and remodeling their cytoskeleton within minutes both ex vivo and in vivo. This study tested the hypothesis that the mechanism of fibroblast expansion in response to tissue stretch involves extracellular ATP signaling. In response to tissue stretch ex vivo, ATP levels in the bath solution increased significantly, and this increase was sustained for 20 min, returning to baseline at 60 min. No increase in ATP was observed in tissue incubated without stretch or tissue stretched in the presence of the Rho kinase inhibitor Y27632. The increase in fibroblast cross sectional area in response to tissue stretch was blocked by both suramin (a purinergic receptor blocker) and apyrase (an enzyme that selectively degrades extracellular ATP). Furthermore, connexin channel blockers (octanol and carbenoxolone), but not VRAC (fluoxetine) or pannexin (probenecid) channel blockers, inhibited fibroblast expansion. Together, these results support a mechanism in which extracellular ATP signaling via connexin hemichannels mediate the active change in fibroblast shape that occurs in response to a static increase in tissue length. Copyright © 2013 Wiley Periodicals, Inc.

  10. Interaction of human gingival fibroblasts with PVA/gelatine sponges.

    Science.gov (United States)

    Moscato, Stefania; Mattii, Letizia; D'Alessandro, Delfo; Cascone, Maria Grazia; Lazzeri, Luigi; Serino, Lorenzo Pio; Dolfi, Amelio; Bernardini, Nunzia

    2008-07-01

    Tissue engineering scaffolds should be able to reproduce optimal microenvironments in order to support cell attachment, three-dimensional growth, migration and, regarding fibroblasts, must also promote extracellular matrix production. Various bioactive molecules are employed in the preparation of spongy scaffolds to obtain biomimetic matrices by either surface-coating or introducing them into the bulk composition of the biomaterial. The biomimetic properties of a spongy matrix composed of PVA combined with the natural component gelatine were evaluated by culturing human gingival fibroblasts on the scaffold. Cell adhesion, morphology and distribution within the scaffold were assessed by histology and electron microscopy; viability and metabolic activity as well as extracellular matrix production were analyzed by MTT assay, cytochemistry and immunocytochemistry. Fibroblasts interacted positively with PVA/gelatine. They adhered to the PVA/gelatine matrix in which they had good spreading activity and active metabolism; fibroblasts were also able to produce extracellular matrix molecules (type I collagen, fibronectin and laminin) compared to bi-dimensionally grown cells. The in situ creation of a biological matrix by human fibroblasts together with the ability to produce growth factor TGF-beta1 and the intracellular signal transduction molecule RhoA, suggests that this kind of PVA/gelatine sponge may represent a suitable support for in vitro extracellular matrix production and connective tissue regeneration.

  11. Keratinocyte growth factor mRNA expression in periodontal ligament fibroblasts

    DEFF Research Database (Denmark)

    Dabelsteen, S; Wandall, H H; Grøn, B

    1997-01-01

    Keratinocyte growth factor (KGF) is a fibroblast growth factor which mediates epithelial growth and differentiation. KGF is expressed in subepithelial fibroblasts, but generally not in fibroblasts of deep connective tissue, such as fascia and ligaments. Here we demonstrate that KGF mRNA is expres......Keratinocyte growth factor (KGF) is a fibroblast growth factor which mediates epithelial growth and differentiation. KGF is expressed in subepithelial fibroblasts, but generally not in fibroblasts of deep connective tissue, such as fascia and ligaments. Here we demonstrate that KGF m...

  12. Calcium oscillations in wounded fibroblast monolayers are spatially regulated through substrate mechanics

    Science.gov (United States)

    Lembong, Josephine; Sabass, Benedikt; Stone, Howard A.

    2017-08-01

    The maintenance of tissue integrity is essential for the life of multicellular organisms. Healing of a skin wound is a paradigm for how various cell types localize and repair tissue perturbations in an orchestrated fashion. To investigate biophysical mechanisms associated with wound localization, we focus on a model system consisting of a fibroblast monolayer on an elastic substrate. We find that the creation of an edge in the monolayer causes cytosolic calcium oscillations throughout the monolayer. The oscillation frequency increases with cell density, which shows that wound-induced calcium oscillations occur collectively. Inhibition of myosin II reduces the number of oscillating cells, demonstrating a coupling between actomyosin activity and calcium response. The spatial distribution of oscillating cells depends on the stiffness of the substrate. For soft substrates with a Young’s modulus E ~ 360 Pa, oscillations occur on average within 0.2 mm distance from the wound edge. Increasing substrate stiffness leads to an average localization of oscillations away from the edge (up to ~0.6 mm). In addition, we use traction force microscopy to determine stresses between cells and substrate. We find that an increase of substrate rigidity leads to a higher traction magnitude. For E    ~8 kPa, traction magnitude is on average almost uniform beneath the monolayer. Thus, the spatial occurrence of calcium oscillations correlates with the cell-substrate traction. Overall, the experiments with fibroblasts demonstrate a collective, chemomechanical localization mechanism at the edge of a wound with a potential physiological role.

  13. Fibroblasts from phenotypically normal palmar fascia exhibit molecular profiles highly similar to fibroblasts from active disease in Dupuytren's Contracture

    Directory of Open Access Journals (Sweden)

    Satish Latha

    2012-05-01

    Full Text Available Abstract Background Dupuytren's contracture (DC is a fibroproliferative disorder characterized by the progressive development of a scar-like collagen-rich cord that affects the palmar fascia of the hand and leads to digital flexion contractures. DC is most commonly treated by surgical resection of the diseased tissue, but has a high reported recurrence rate ranging from 27% to 80%. We sought to determine if the transcriptomic profiles of fibroblasts derived from DC-affected palmar fascia, adjacent phenotypically normal palmar fascia, and non-DC palmar fascial tissues might provide mechanistic clues to understanding the puzzle of disease predisposition and recurrence in DC. Methods To achieve this, total RNA was obtained from fibroblasts derived from primary DC-affected palmar fascia, patient-matched unaffected palmar fascia, and palmar fascia from non-DC patients undergoing carpal tunnel release (6 patients in each group. These cells were grown on a type-1 collagen substrate (to better mimic their in vivo environments. Microarray analyses were subsequently performed using Illumina BeadChip arrays to compare the transcriptomic profiles of these three cell populations. Data were analyzed using Significance Analysis of Microarrays (SAM v3.02, hierarchical clustering, concordance mapping and Venn diagram. Results We found that the transcriptomic profiles of DC-disease fibroblasts and fibroblasts from unaffected fascia of DC patients exhibited a much greater overlap than fibroblasts derived from the palmar fascia of patients undergoing carpal tunnel release. Quantitative real time RT-PCR confirmed the differential expression of select genes validating the microarray data analyses. These data are consistent with the hypothesis that predisposition and recurrence in DC may stem, at least in part, from intrinsic similarities in the basal gene expression of diseased and phenotypically unaffected palmar fascia fibroblasts. These data also demonstrate that

  14. Fibroblasts from phenotypically normal palmar fascia exhibit molecular profiles highly similar to fibroblasts from active disease in Dupuytren's Contracture

    Science.gov (United States)

    2012-01-01

    Background Dupuytren's contracture (DC) is a fibroproliferative disorder characterized by the progressive development of a scar-like collagen-rich cord that affects the palmar fascia of the hand and leads to digital flexion contractures. DC is most commonly treated by surgical resection of the diseased tissue, but has a high reported recurrence rate ranging from 27% to 80%. We sought to determine if the transcriptomic profiles of fibroblasts derived from DC-affected palmar fascia, adjacent phenotypically normal palmar fascia, and non-DC palmar fascial tissues might provide mechanistic clues to understanding the puzzle of disease predisposition and recurrence in DC. Methods To achieve this, total RNA was obtained from fibroblasts derived from primary DC-affected palmar fascia, patient-matched unaffected palmar fascia, and palmar fascia from non-DC patients undergoing carpal tunnel release (6 patients in each group). These cells were grown on a type-1 collagen substrate (to better mimic their in vivo environments). Microarray analyses were subsequently performed using Illumina BeadChip arrays to compare the transcriptomic profiles of these three cell populations. Data were analyzed using Significance Analysis of Microarrays (SAM v3.02), hierarchical clustering, concordance mapping and Venn diagram. Results We found that the transcriptomic profiles of DC-disease fibroblasts and fibroblasts from unaffected fascia of DC patients exhibited a much greater overlap than fibroblasts derived from the palmar fascia of patients undergoing carpal tunnel release. Quantitative real time RT-PCR confirmed the differential expression of select genes validating the microarray data analyses. These data are consistent with the hypothesis that predisposition and recurrence in DC may stem, at least in part, from intrinsic similarities in the basal gene expression of diseased and phenotypically unaffected palmar fascia fibroblasts. These data also demonstrate that a collagen

  15. Secretome Analysis of Human Primary Fibroblasts Undergoing Senescence

    DEFF Research Database (Denmark)

    Rogowska-Wrzesinska, Adelina; Micutkova, Lucia; Diener, Thomas

    Introduction Cultures of diploid human fibroblasts can replicate only a finite number of times; rapid proliferation is followed by decline in replicative frequency and finally cells become senescent and are incapable of further proliferation. Senescencent cells display altered growth, morphology......-degrading. In this study we use proteomic tools to characterise the secretome of young and senescent fibroblasts.   Methods Three independent preparations of primary human foreskin fibroblasts were grown to senescence. Young, rapidly proliferating cells at passage 11 and cells from passage 28 displaying senescent...... spectrometry; peptide count was used to estimate protein abundance.   Results 2DGE based analysis of secretion profiles of young and senescent cells derived from the same cell lineage revealed a number of protein spots differentially expressed. We have observed an increased secretion of matrix...

  16. Chemical Conversion of Human Fibroblasts into Functional Schwann Cells

    Directory of Open Access Journals (Sweden)

    Eva C. Thoma

    2014-10-01

    Full Text Available Direct transdifferentiation of somatic cells is a promising approach to obtain patient-specific cells for numerous applications. However, conversion across germ-layer borders often requires ectopic gene expression with unpredictable side effects. Here, we present a gene-free approach that allows efficient conversion of human fibroblasts via a transient progenitor stage into Schwann cells, the major glial cell type of peripheral nerves. Using a multikinase inhibitor, we transdifferentiated fibroblasts into transient neural precursors that were subsequently further differentiated into Schwann cells. The resulting induced Schwann cells (iSCs expressed numerous Schwann cell-specific proteins and displayed neurosupportive and myelination capacity in vitro. Thus, we established a strategy to obtain mature Schwann cells from human postnatal fibroblasts under chemically defined conditions without the introduction of ectopic genes.

  17. Systems-level modeling of cancer-fibroblast interaction.

    Directory of Open Access Journals (Sweden)

    Raymond C Wadlow

    2009-09-01

    Full Text Available Cancer cells interact with surrounding stromal fibroblasts during tumorigenesis, but the complex molecular rules that govern these interactions remain poorly understood thus hindering the development of therapeutic strategies to target cancer stroma. We have taken a mathematical approach to begin defining these rules by performing the first large-scale quantitative analysis of fibroblast effects on cancer cell proliferation across more than four hundred heterotypic cell line pairings. Systems-level modeling of this complex dataset using singular value decomposition revealed that normal tissue fibroblasts variably express at least two functionally distinct activities, one which reflects transcriptional programs associated with activated mesenchymal cells, that act either coordinately or at cross-purposes to modulate cancer cell proliferation. These findings suggest that quantitative approaches may prove useful for identifying organizational principles that govern complex heterotypic cell-cell interactions in cancer and other contexts.

  18. Effects of HEMA on type I collagen protein in human gingival fibroblasts.

    Science.gov (United States)

    Falconi, M; Teti, G; Zago, M; Pelotti, S; Breschi, L; Mazzotti, G

    2007-09-01

    The cytotoxicity of dental composites has been attributed to the release of residual monomers from polymerized adhesive systems due to degradation processes or the incomplete polymerization of materials. 2-Hydroxyethyl methacrylate (HEMA) is one of the major components released from dental adhesives. Cytotoxic effects due to high concentrations of HEMA have already been investigated, but the influence of minor toxic concentrations on specific proteins such as type I collagen has not been studied in depth. The objective of this project was to study the effect of minor toxic concentrations of HEMA on human gingival fibroblasts (HGFs), investigating modification in cell morphology, cell viability, and the influence on type I collagen protein. Primary lines of human gingival fibroblasts were exposed to 3 mmol/L HEMA for different periods of time (24 h, 72 h, 96 h). The cell vitality was determined by MTT assay, and high-resolution scanning electron microscopy analysis was performed to evaluate differences in cell morphology before and after treatment. The presence and localization of type I collagen was determined by immunofluorescence in HGFs treated with HEMA for the same period of time. The vitality of the cells decreased after 72 h of exposure. The HGFs grown in monolayer and observed by field emission in-lens scanning electron microscopy demonstrated a preserved surface morphology after 24 h of treatment, while they showed an altered morphology after 96 h of treatment. Immunofluorescence demonstrated a reduction of type I collagen due to HEMA exposure after 96 h. From these results, we conclude that low concentrations of HEMA can significantly alter the morphology of human gingival fibroblasts and interfere with the presence of type I collagen protein.

  19. Inhibition of human scleral fibroblast cell attachment to collagen type I by TGFBIp.

    Science.gov (United States)

    Shelton, Lilian; Rada, Jody A Summers

    2009-08-01

    Transforming growth factor beta-induced protein (TGFBIp; 68 kDa) is a secreted extracellular matrix (ECM) protein that has been demonstrated to regulate cell attachment in a variety of cell types. The sclera synthesizes and secretes TGFBIp, which may function to facilitate scleral ECM remodeling events associated with myopia development. Here the authors report that human scleral fibroblasts (HSFs) express TGFBI and that its protein product, TGFBIp, mediates an effect on cell attachment. TGFBI/TGFBIp expression was evaluated by RT-PCR and immunoblot of HSF lysates and culture supernatants. The effect of rTGFBIp (50 microg/mL) on cell attachment to collagen type I was determined with the use of fluid-phase cell attachment assays in HSFs, human foreskin fibroblasts (HFFs), and human corneal stroma fibroblasts (HCFs). Binding assays using biotinylated rTGFBIp were used to assess TGFBIp binding to the HSF surface. Flow cytometry and immunocytochemistry were used to determine both alphavbeta3 and alphavbeta5 expression and localization to the HSF cell surface. HSFs expressed TGFBI and secreted TGFBIp (approximately 833 ng/h). rTGFBIp significantly decreased (25 microg/mL; P collagen type I, whereas rTGFBIp did not significantly affect cell attachment of HFFs (P = 0.50) or HCFs (P = 0.24) to collagen compared with BSA. Integrins alphavbeta3 and alphavbeta5 were detected on the cell surface, and both anti-alphavbeta3 and anti-alphavbeta5 functionally blocked rTGFBIp binding to HSFs. TGFBIp plays an inhibitory role in HSF attachment to collagen type I in vitro through interactions with alphavbeta3 and alphavbeta5 integrin receptors. These results suggest that TGFBIp may modulate scleral cell-matrix interactions in vivo, thereby affecting scleral viscoelasticity.

  20. Regional Heterogeneity in Murine Lung Fibroblasts from Normal Mice or Mice Exposed Once to Cigarette Smoke

    Science.gov (United States)

    Preobrazhenska, Olena; Wright, Joanne L.; Churg, Andrew

    2012-01-01

    Chronic obstructive lung disease (COPD) is characterized by matrix deposition in the small airways but matrix loss from the parenchyma, phenomena which must depend on the ability of local fibroblasts to produce matrix after smoke exposure. To investigate this idea, we exposed C57Bl/6 mice once to cigarette smoke or to air (control) and prepared primary cultures of lung fibroblasts by microdissecting large airways (trachea, LAF), medium size airways (major bronchi, MAF) and parenchyma (PF). Control PF showed the lowest rate of wound closure and wound closure was depressed in all lines by a single in vivo smoke exposure. Gene expression of matrix proteins differed considerably among the sites; decorin, which may sequester TGFβ, was markedly higher in PF. PF showed higher intrinsic ratios of pSmad2/Smad2. Smoke caused much greater increases in secreted and matrix deposited collagens 1 and 3 in PF than in LAF or MAF. Expression of Thy-1, a gene that suppresses myofibroblast differentiation, was increased by smoke in PF. We conclude that there is considerable regional heterogeneity in murine lung fibroblasts in terms of matrix production, either basally or after in vivo smoke exposure; that PF have lower ability to repair wounds and higher intrinsic TGFβ signaling; and that a single exposure to smoke produces lasting changes in the pattern of matrix production and wound repair, changes that may be mediated in part by smoke-induced release of TGFβ. However, PF still retain the ability to repair by producing new matrix after a single in vivo smoke exposure. PMID:22761892

  1. Expression of the endocannabinoid system in fibroblasts and myofascial tissues.

    Science.gov (United States)

    McPartland, John M

    2008-04-01

    The endocannabinoid (eCB) system, like the better-known endorphin system, consists of cell membrane receptors, endogenous ligands and ligand-metabolizing enzymes. Two cannabinoid receptors are known: CB(1) is principally located in the nervous system, whereas CB(2) is primarily associated with the immune system. Two eCB ligands, anandamide (AEA) and 2-arachidonoylglycerol (2-AG), are mimicked by cannabis plant compounds. The first purpose of this paper was to review the eCB system in detail, highlighting aspects of interest to bodyworkers, especially eCB modulation of pain and inflammation. Evidence suggests the eCB system may help resolve myofascial trigger points and relieve symptoms of fibromyalgia. However, expression of the eCB system in myofascial tissues has not been established. The second purpose of this paper was to investigate the eCB system in fibroblasts and other fascia-related cells. The investigation used a bioinformatics approach, obtaining microarray data via the GEO database (www.ncbi.nlm.nih.gov/geo/). GEO data mining revealed that fibroblasts, myofibroblasts, chondrocytes and synoviocytes expressed CB(1), CB(2) and eCB ligand-metabolizing enzymes. Fibroblast CB(1) levels nearly equalled levels expressed by adipocytes. CB(1) levels upregulated after exposure to inflammatory cytokines and equiaxial stretching of fibroblasts. The eCB system affects fibroblast remodeling through lipid rafts associated with focal adhesions and dampens cartilage destruction by decreasing fibroblast-secreted metalloproteinase enzymes. In conclusion, the eCB system helps shape biodynamic embryological development, diminishes nociception and pain, reduces inflammation in myofascial tissues and plays a role in fascial reorganization. Practitioners wield several tools that upregulate eCB activity, including myofascial manipulation, diet and lifestyle modifications, and pharmaceutical approaches.

  2. Chloride transport in human fibroblasts is activated by hypotonic shock

    Energy Technology Data Exchange (ETDEWEB)

    Rugolo, M.; Mastocola, T.; Flamigni, A.; Lenaz, G. (Universita' di Bologna (Italy))

    1989-05-15

    Incubation of human skin fibroblasts in hypotonic media induced the activation of {sup 36}Cl- efflux which was roughly proportional to the decrease in the osmolality of the media. The efflux of {sup 36}Cl- was insensitive to DIDS plus furosemide and inhibited by addition of a Cl- channel blocker such as 5-nitro-2-(3-phenyl propylamino) benzoic acid (NPPB). We propose that a conductive pathway for Cl- transport, almost silent in isotonic conditions, is activated by exposing human fibroblasts to hypotonic shock, this conclusion being supported by evidence that also {sup 36}Cl- influx was enhanced by hypotonic medium.

  3. Application of Allogeneic Fibroblast Cells in Cellular Therapy of Recessive Dystrophic Epidermolysis Bullosa

    Directory of Open Access Journals (Sweden)

    Zare

    2015-09-01

    Full Text Available Context Connective tissue cells include fibroblasts, chondrocytes, adipocyte, and osteocytes. These cells are specialized for the secretion of collagenous extracellular matrix and are responsible for the architectural framework of the human body. Evidence Acquisition Connective tissue cells play a central role in supporting as well as repairing tissues and organs. Fibroblast cell therapy could be used for the treatment of burn wounds, scars, diabetic foot ulcers, acne scars and skin aging. This review focused on biology of fibroblasts and their role in cell therapy of recessive dystrophic epidermolysis bullosa (RDEB. Results Fibroblasts are known to play a pivotal role in skin structure and integrity, and dermal fibroblasts are believed to promote skin regeneration and rejuvenation via collagen production. Conclusions Fibroblasts can be used in transplantations to ameliorate an immune system response, in order to reduce antigen production. Human fibroblasts suppress ongoing mixed lymphocyte reactions (MLRs between lymphocyte cells from two individuals, and supernatant materials from fibroblast cultures suppress MLRs.

  4. Gingival fibroblast responsiveness is differentially affected by Porphyromonas gingivalis: implications for the pathogenesis of periodontitis

    NARCIS (Netherlands)

    Scheres, N.; Crielaard, W.

    2013-01-01

    In periodontitis, tissue damage results mainly from aberrant host responses to oral microorganisms. Fibroblasts can play an important role in this. Gingival fibroblasts do not develop tolerance against the lipopolysaccharide of Porphyromonas gingivalis, a keystone pathogen in periodontitis, which

  5. Cardiac fibroblast-derived microRNA passenger strand-enriched exosomes mediate cardiomyocyte hypertrophy

    National Research Council Canada - National Science Library

    Bang, Claudia; Batkai, Sandor; Dangwal, Seema; Gupta, Shashi Kumar; Foinquinos, Ariana; Holzmann, Angelika; Just, Annette; Remke, Janet; Zimmer, Karina; Zeug, Andre; Ponimaskin, Evgeni; Schmiedl, Andreas; Yin, Xiaoke; Mayr, Manuel; Halder, Rashi; Fischer, Andre; Engelhardt, Stefan; Wei, Yuanyuan; Schober, Andreas; Fiedler, Jan; Thum, Thomas

    2014-01-01

    ...; however, miRNAs have emerged recently as paracrine signaling mediators. Thus, we investigated a potential paracrine miRNA crosstalk between cardiac fibroblasts and cardiomyocytes and found that cardiac fibroblasts secrete miRNA-enriched exosomes...

  6. Phytanic acid alpha-oxidation in peroxisomal disorders: studies in cultured human fibroblasts

    NARCIS (Netherlands)

    Verhoeven, N. M.; Schor, D. S.; Roe, C. R.; Wanders, R. J.; Jakobs, C.

    1997-01-01

    We studied the alpha-oxidation of phytanic acid in human fibroblasts of controls and patients affected with classical Refsum disease, rhizomelic chondrodysplasia punctata, generalized peroxisomal disorders and peroxisomal bifunctional protein deficiency. Cultured fibroblasts were incubated with

  7. Water-filtered near-infrared influences collagen synthesis of keloid-fibroblasts in contrast to normal foreskin fibroblasts.

    Science.gov (United States)

    Zöller, Nadja; König, Anke; Butting, Manuel; Kaufmann, Roland; Bernd, August; Valesky, Eva; Kippenberger, Stefan

    2016-10-01

    Hypertrophic scar development is associated to impaired wound healing, imbalanced fibroblast proliferation and extracellular matrix synthesis. Stigmatization, physical restrictions and high recurrence rates are only some aspects that illustrate the severe influence impaired wound healing can have on patients' life. The treatment of hypertrophic scars especially keloids is still a challenge. In recent years water-filtered near-infrared irradiation (wIRA) composed of near-infrared (NIR) and a thermal component is applied for an increasing penal of clinical purposes. It is described to beneficially influence e.g. wound healing. But discrimination between the thermal and the NIR dependent components of these effects has not been conclusively elucidated. Aim of our study was therefore to investigate the influence of the light fraction on the thermal impact of wIRA irradiation in dermal cells. We concentrated our analysis on morphological properties and collagen synthesis. Foreskin fibroblasts and the keloid fibroblast cell line KF111 were exposed to temperatures between 37°C and 46°C with or without additional irradiation with 360J/cm(2) NIR. Our results show that viability was not influenced by irradiation. Independent of the analysed fibroblast species temperature dependent occurrence of spheric cells could be observed. These morphological changes were clearly counteracted by additional light exposure. Convective heat reduced collagen type I synthesis in both cell species depending on the applied temperature. Co-treatment with NIR significantly reversed this effect in keloid fibroblast cultures treated at 46°C whereas no difference could be observed in the foreskin fibroblasts. The observed influence on collagen type I synthesis was associated to a temperature dependent TGF-β1 secretion reduction. Co-stimulation of keloid cultures with NIR at 46°C completely abolished the temperature dependent TGF-β1 secretion reduction. In foreskin fibroblast cultures co

  8. The effect of local hyperglycemia on skin cells in vitro and on wound healing in euglycemic rats

    DEFF Research Database (Denmark)

    Kruse, Carla R; Singh, Mansher; Sørensen, Jens A

    2016-01-01

    BACKGROUND: Multiple previous studies have established that high systemic blood glucose concentration impairs skin wound healing. However, the effects of local hyperglycemia on wound healing are not well defined. Comprehensive animal studies and in vitro studies using both fibroblasts and keratin......BACKGROUND: Multiple previous studies have established that high systemic blood glucose concentration impairs skin wound healing. However, the effects of local hyperglycemia on wound healing are not well defined. Comprehensive animal studies and in vitro studies using both fibroblasts...

  9. On the mechanisms of cortical actin flow and its role in cytoskeletal organisation of fibroblasts.

    Science.gov (United States)

    Heath, J P; Holifield, B F

    1993-01-01

    In this review we discuss the organization of F-actin in motile fibroblasts in relation to the phenomenon of cortical actin flow; we review some of the mechanisms proposed to drive this process and then relate some of our new findings on the interactions of cortical flow and substratum adhesions in fibroblasts. It is our thesis that the centripetal flow of F-actin through the lamellipodium and leading lamella is the major determining factor organising the polarized stress fiber system and associated cell-substratum adhesions in crawling fibroblastic cells. The broad flattened region of cytoplasm anterior to the nucleus, called the leading lamella, is fringed with much thinner structures, the lamellipodia, which are the primary protrusive organelles of motile cells. Lamellipodia are filled with a criss-crossed network of actin filaments interspersed with small bundles or ribs. In the leading lamella, the actin cytoskeleton is largely confined to two cortical layers beneath the dorsal and ventral cell surfaces. The ventral cortex is engaged in cell adhesion; the dorsal cortex is made up of a circumferentially orientated sheet of actin filaments and bundles which we team the dorsal cortical microfilament sheath (DCMS). Stress fibers insert into the ventral adhesions and pass back through the lamella rising up to meet the DCMS. Cortical flow appears to be a constitutive process in most types of cells when they become motile. In fibroblasts a continuous centripetal flux of structure is seen flowing through the lamellipodium and into the more central regions of the lamella at approximately 0.1 micron per second. Some of the structures engaged in the flux pass back and merge into the DCMS which is also moving rearward at a slower rate of 1 to 5 microns per minute. We find that the formation of a stress fibre is a direct consequence of cortical flow. Initially, stress fibre formation involves the establishment of a focal adhesion between an F-actin bundle in the

  10. Increased autophagy in fibroblast-like synoviocytes leads to immune enhancement potential in rheumatoid arthritis.

    Science.gov (United States)

    Yang, Ru; Zhang, Yingzi; Wang, Lin; Hu, Ji; Wen, Jian; Xue, Leixi; Tang, Mei; Liu, Zhichun; Fu, Jinxiang

    2017-02-28

    The incidence of rheumatoid arthritis (RA) has been reported to be correlated with a disorder of immunregulation. Rheumatoid arthritis fibroblast-like synoviocytes (RA-FLSs) play an important role in regulating the local immune microenvironment. However, the potential mechanism of RA-FLS in regulating the immnue response is not clearly understood. In this study, we demonstrated that the expression of HIF-1α was significantly up-regulated in rheumatoid arthritis tissue which indicated that the hypoxia condition in the microenvironment. We also observed that RA-FLSs demonstrated the potential to up-regulate immune activation. Meanwhile, the level of autophagy increased in RA-FLSs compared with control group. Besides that, the expression of IL-6 was up-regulated not only in RA-FLSs but also in the fibroblasts that treated with hypoxia condition. Accordingly, we found that autophagy inhibitiors could effectively inhibit the immune activation function of RA-FLSs medicated by IL-6. Taken together, the results we demonstrated above indicated that the hypoxia microenvironment could effectively induce the incidence of autophagy and then lead to the immune activation function of RA-FLSs medicated by IL-6.

  11. Protective Effect of Modified Human Acidic Fibroblast Growth Factor ...

    African Journals Online (AJOL)

    Purpose: To investigate whether modified acidic fibroblast growth factor (MaFGF) can protect NRK52E cell against apoptotic death induced by actinomycin D (Act D) and the effect of MaFGF on PI3K/Akt signaling pathway. Methods: NRK52E cell apoptotic death was measured by several methods including cell morphologic ...

  12. Transient expression of acidic fibroblast growth factor in pea ( Pisum ...

    African Journals Online (AJOL)

    Nowadays, there are many therapeutic proteins produced in different host plants in transient easy to perform, short production cycle, efficient and inexpensive. In this study, the modified pea early browning virus (PEBV) vector containing GFP and acidic fibroblast growth factor (aFGF) was introduced into pea plants by leave ...

  13. Fibroblast growth factor 23 and dietary factors in renal disease

    NARCIS (Netherlands)

    da Cunha Baia, Leandro

    2015-01-01

    Omega-3 poly-unsaturated fatty acids and mineral metabolism: novel therapy for cardiovascular disease in renal patients? Deregulations in mineral metabolism, particularly related to phosphate and its regulating hormone fibroblast growth factor 23 (FGF23), are common in patients with chronic kidney

  14. Airway smooth muscle and fibroblasts in the pathogenesis of asthma

    NARCIS (Netherlands)

    Johnson, Peter R A; Burgess, Janette K

    2004-01-01

    Asthma is a disease characterized by marked structural changes within the airway wall. These changes include deposition of extracellular matrix proteins and an increase in the numbers of airway smooth muscle cells and subepithelial fibroblasts. Both these cell types possess properties that would

  15. Cancer-Associated Fibroblasts: Perspectives in Cancer Therapy

    NARCIS (Netherlands)

    Prakash, Jai

    2016-01-01

    The interplay between cancer cells and stromal cells is increasingly recognized as a main driver of tumor progression and metastasis. This Forum article highlights the role of cancer-associated stromal fibroblasts (CAFs) in tumorigenesis and discusses the potential for developing specific stromal

  16. Optimum microcurrent stimulation intensity for galvanotaxis in human fibroblasts.

    Science.gov (United States)

    Sugimoto, M; Maeshige, N; Honda, H; Yoshikawa, Y; Uemura, M; Yamamoto, M; Terashi, H

    2012-01-01

    In this study, we develop methods to measure galvanotaxis of fibroblasts and determined the optimum conditions of electrical stimulation. An inverted 35mm dish containing cell suspensions (3×105 primary human skin fibroblasts, DMEM, and 10% FBS) was placed on the centre of a 100mm dish. The 35mm dish was removed 24 hours later, and culture medium was added to the 100mm dish. Fibroblasts were randomised (double-blind) into three groups, where electrical stimulation was given at varying intensities: 0UA (control), 50UA, and 100UA. Electrical stimulation (frequency=0.3Hz) was conducted, for a duration of 4 hours, with platinum electrodes in a CO2 incubator. We took pictures immediately before and 20 hours after stimulation. We calculated the migration ratio to the negative pole by dividing the area of attached fibroblasts after stimulation with that before stimulation. The migration ratio to the negative pole was significantly higher in the 100UA group than in the control group (pmicrocurrent efficacy for pressure ulcer healing. Electrical stimulation based on our in vitro experiment might be important for the development of physical therapy for pressure ulcers.

  17. Fibroblast growth factor 23: a potential cause of cardiovascular ...

    African Journals Online (AJOL)

    Fibroblast growth factor 23 (FGF-23) has been identified as one of the risk factors for the development of cardiovascular diseases (CVDs) in chronic kidney disease (CKD) patients. Although FGF-23 is necessary for the maintenance of phosphate balance, it has been implicated in the pathogenesis of left ventricular ...

  18. Pulsed short-wave diathermy effects on human fibroblast proliferation.

    Science.gov (United States)

    Hill, Jonathan; Lewis, Martyn; Mills, Pauline; Kielty, Cay

    2002-06-01

    To investigate the influence of pulsed short-wave diathermy (PSWD) on fibroblast and chondrocyte cell proliferation rates and to establish the influences of different dosages applied. Four single-blind trials. Laboratory, in vitro study. Human adult dermal fibroblast and chondrocyte cells were plated at known concentrations and incubated for 5 days. Exposure to PSWD, twice daily, on days 2, 3, and 4. After crystal violet staining (day 5), optical density (cell number) was determined spectrophotometrically. PSWD, given at mean power of 48W for 10 minutes, increased fibroblast proliferation compared with control groups (P<.001). There was a relationship between cell proliferation and the amount of energy given (P<0.001). The optimal mean power for proliferation was estimated to be 13.8W. While keeping mean power constant at 6W, altering pulse duration and pulse repetition rate dosage parameters did not have a significant effect on proliferation (P=.519). Chondrocyte proliferation also increased with PSWD exposure of 6W at 10 minutes duration (P=.015). In addition, treatment time was significantly associated with chondrocyte proliferation (P<.001). PSWD is associated with increased rates of fibroblast and chondrocyte proliferation in vitro, which is dose dependent. These results contribute to an understanding of the physiologic mechanisms underlying the therapeutic effects of PSWD. Copyright 2002 by the American Congress of Rehabilitation Medicine and the American Academy of Physical Medicine and Rehabilitation

  19. The Living Scar – Cardiac Fibroblasts and the Injured Heart

    Science.gov (United States)

    Rog-Zielinska, Eva A; Norris, Russell A; Kohl, Peter; Markwald, Roger

    2015-01-01

    Cardiac scars, often perceived as “dead” tissue, are very much alive, with heterocellular activity ensuring the maintenance of structural and mechanical integrity following heart injury. To form a scar, non-myocytes such as fibroblasts, proliferate and are recruited from intra- and extra-cardiac sources. Fibroblasts perform important autocrine and paracrine signalling functions. They also establish mechanical and, as is increasingly evident, electrical junctions with other cells. While fibroblasts were previously thought to act simply as electrical insulators, they may be electrically connected among themselves and, under certain circumstances, to other cells, including cardiomyocytes. A better understanding of these interactions will help target scar structure and function and facilitate the development of novel therapies aimed at modifying scar properties for patient benefit. This review explores available insight and recent concepts on fibroblast integration in the heart, and highlights potential avenues for harnessing their roles to optimise scar function following heart injury such as infarction, and therapeutic interventions such as ablation. PMID:26776094

  20. Fibroblast Growth Factor-2 Alters the Nature of Extinction

    Science.gov (United States)

    Graham, Bronwyn M.; Richardson, Rick

    2011-01-01

    These experiments examined the effects of the NMDA-receptor (NMDAr) antagonist MK801 on reacquisition and re-extinction of a conditioned fear that had been previously extinguished before injection of fibroblast growth factor-2 (FGF2) or vehicle. Recent findings have shown that relearning and re-extinction, unlike initial learning and extinction,…

  1. Photodynamic inactivation of fibroblasts by a cationic porphyrin

    NARCIS (Netherlands)

    Lambrechts, Saskia A. G.; Schwartz, Kevin R.; Aalders, Maurice C. G.; Dankert, Jacob B.

    2005-01-01

    An important determinant of the clinical applicability and value of antimicrobial photodynamic inactivation (PDI) is the cytotoxicity of the treatment to human cells. We evaluated the in vitro cytotoxicity of PDI to human dermal fibroblasts using 5-phenyl-10,15,20-tris(N-methyl-4-pyridyl)porphyrin

  2. Constitutive Reprogramming of Fibroblast Mitochondrial Metabolism in Pulmonary Hypertension

    Czech Academy of Sciences Publication Activity Database

    Plecitá-Hlavatá, Lydie; Tauber, Jan; Li, M.; Zhang, H.; Flockton, A. R.; Pullamsetti, S. S.; Chelladurai, P.; D'Alessandro, A.; El Kasmi, K. C.; Ježek, Petr; Stenmark, K. R.

    2016-01-01

    Roč. 55, č. 1 (2016), s. 47-57 ISSN 1044-1549 R&D Projects: GA MŠk(CZ) LH11055; GA MŠk(CZ) LH15071 Institutional support: RVO:67985823 Keywords : mitochondria * complex I * oxidative metabolism * pulmonary hypertension * adventitial fibroblasts Subject RIV: ED - Physiology Impact factor: 4.100, year: 2016

  3. Spatial organization of fibroblast nuclear chromocenters: component tree analysis.

    Science.gov (United States)

    Snapp, Robert R; Goveia, Elyse; Peet, Lindsay; Bouffard, Nicole A; Badger, Gary J; Langevin, Helene M

    2013-09-01

    The nuclei of mouse connective tissue fibroblasts contain chromocenters which are well-defined zones of heterochromatin that can be used as positional landmarks to examine nuclear remodeling in response to a mechanical perturbation. This study used component tree analysis, an image segmentation algorithm that detects high intensity voxels that are topologically connected, to quantify the spatial organization of chromocenters in fibroblasts within whole mouse connective tissue fixed and stained with 4',6-diamidino-2-phenylindole (DAPI). The component tree analysis method was applied to confocal microscopy images of whole mouse areolar connective tissue incubated for 30 min ex vivo with or without static stretch. In stretched tissue, the mean distance between chromocenters within fibroblast nuclei was significantly greater (vs. non-stretched, P stretch and no stretch. Average chromocenter distance was positively correlated with nuclear cross-sectional area (r = 0.78, P Static stretching of mouse areolar connective tissue for 30 min resulted in substantially increased separation of nuclear chromocenters in connective tissue fibroblasts. This interior remodeling of the nucleus induced by tissue stretch may impact transcriptionally active euchromatin within the inter-chromocenter space. © 2013 Anatomical Society.

  4. fibroblast growth factor, MTDH/Astrocyte elevated gene-1

    African Journals Online (AJOL)

    2012-12-05

    Dec 5, 2012 ... Background: The etiopathogenesis of prostate cancer (PC) is still not clear, but hormonal, genetic, and environmental factors are thought to play a role in the tumor pathogenesis. ... many genetic and epigenetic alterations have been detected in human PC.[1]. Basic fibroblast growth factor (bFGF), also ...

  5. Involvement of the mitochondrial compartment in human NCL fibroblasts

    Energy Technology Data Exchange (ETDEWEB)

    Pezzini, Francesco; Gismondi, Floriana [Department of Neurological, Psychological, Morphological and Motor Sciences, Divisions of Neurology (Child Neurology) and Neuropathology, University of Verona Medical School, Verona (Italy); Tessa, Alessandra [IRCCS Fondazione Stella Maris-Molecular Medicine Unit, Pisa (Italy); Tonin, Paola [Department of Neurological, Psychological, Morphological and Motor Sciences, Divisions of Neurology (Child Neurology) and Neuropathology, University of Verona Medical School, Verona (Italy); Carrozzo, Rosalba [IRCCS Bambino Gesu Hospital-Molecular Medicine Unit, Roma (Italy); Mole, Sara E. [MRC Laboratory for Molecular Cell Biology, Molecular Medicines Unit, UCL Institute of Child Health and Department of Genetics, Evolution and Environment, University College London (United Kingdom); Santorelli, Filippo M. [IRCCS Fondazione Stella Maris-Molecular Medicine Unit, Pisa (Italy); Simonati, Alessandro, E-mail: alessandro.simonati@univr.it [Department of Neurological, Psychological, Morphological and Motor Sciences, Divisions of Neurology (Child Neurology) and Neuropathology, University of Verona Medical School, Verona (Italy)

    2011-12-09

    Highlights: Black-Right-Pointing-Pointer Mitochondrial reticulum fragmentation occurs in human CLN1 and CLN6 fibroblasts. Black-Right-Pointing-Pointer Likewise mitochondrial shift-to periphery and decreased mitochondrial density are seen. Black-Right-Pointing-Pointer Enhanced caspase-mediated apoptosis occurs following STS treatment in CLN1 fibroblasts. -- Abstract: Neuronal ceroid lipofuscinosis (NCL) are a group of progressive neurodegenerative disorders of childhood, characterized by the endo-lysosomal storage of autofluorescent material. Impaired mitochondrial function is often associated with neurodegeneration, possibly related to the apoptotic cascade. In this study we investigated the possible effects of lysosomal accumulation on the mitochondrial compartment in the fibroblasts of two NCL forms, CLN1 and CLN6. Fragmented mitochondrial reticulum was observed in all cells by using the intravital fluorescent marker Mitotracker, mainly in the perinuclear region. This was also associated with intense signal from the lysosomal markers Lysotracker and LAMP2. Likewise, mitochondria appeared to be reduced in number and shifted to the cell periphery by electron microscopy; moreover the mitochondrial markers VDCA and COX IV were reduced following quantitative Western blot analysis. Whilst there was no evidence of increased cell death under basal condition, we observed a significant increase in apoptotic nuclei following Staurosporine treatment in CLN1 cells only. In conclusion, the mitochondrial compartment is affected in NCL fibroblasts invitro, and CLN1 cells seem to be more vulnerable to the negative effects of stressed mitochondrial membrane than CLN6 cells.

  6. Fibroblastic osteosarcoma in a lion ( Panthera leo ) | Leonardi ...

    African Journals Online (AJOL)

    This report describes a case of spontaneous fibroblastic osteosarcoma in the humerus of a lion from a private park in Perugia, Italy. The tumor had an irregular, smooth, brown surface and a generally firm, rubbery consistence with gritty to hard areas interspersed. The mass was poorly vascularized with areas of necrosis at ...

  7. Adhesion, growth, and matrix production by fibroblasts on laminin substrates

    DEFF Research Database (Denmark)

    Couchman, J R; Höök, M; Rees, D A

    1983-01-01

    Human embryonic skin fibroblasts have been shown to attach and spread on laminin substrates in the absence of protein synthesis and presence of fibronectin-depleted serum and anti-fibronectin antibodies. Rates of attachment and the type of spreading are virtually identical on fibronectin and lami...

  8. Cellular electrophysiological principles that modulate secretion from synovial fibroblasts.

    Science.gov (United States)

    Clark, R B; Schmidt, T A; Sachse, F B; Boyle, D; Firestein, G S; Giles, W R

    2017-02-01

    Rheumatoid arthritis (RA) is a progressive disease that affects both pediatric and adult populations. The cellular basis for RA has been investigated extensively using animal models, human tissues and isolated cells in culture. However, many aspects of its aetiology and molecular mechanisms remain unknown. Some of the electrophysiological principles that regulate secretion of essential lubricants (hyaluronan and lubricin) and cytokines from synovial fibroblasts have been identified. Data sets describing the main types of ion channels that are expressed in human synovial fibroblast preparations have begun to provide important new insights into the interplay among: (i) ion fluxes, (ii) Ca2+ release from the endoplasmic reticulum, (iii) intercellular coupling, and (iv) both transient and longer duration changes in synovial fibroblast membrane potential. A combination of this information, knowledge of similar patterns of responses in cells that regulate the immune system, and the availability of adult human synovial fibroblasts are likely to provide new pathophysiological insights. © 2016 University of Calgary. The Journal of Physiology © 2016 The Physiological Society.

  9. High level expression of human basic fibroblast growth factor in ...

    African Journals Online (AJOL)

    USER

    2010-04-19

    Apr 19, 2010 ... High-level expression of recombinant human basic fibroblast growth factor in Escherichia coli presents research opportunities such as analysis ... The general agreement from the published data on heterologous gene ..... for protein expression (Casimiro et al., 1997; Gold et al.,. 1981; Hamdan et al., 2002; ...

  10. Basic FGF and PDGF-BB synergistically stimulate hyaluronan and IL-6 production by orbital fibroblasts

    NARCIS (Netherlands)

    S. Virakul (Sita); J.W. Heutz (Judith W.); V.A.S.H. Dalm (Virgil); R.P. Peeters (Robin); A.D.A. Paridaens (Dion); W.A. van den Bosch (Willem); N. Hirankarn (Nattiya); P.M. van Hagen (Martin); W.A. Dik (Willem)

    2016-01-01

    textabstractOrbital fibroblast activation is a central pathologic feature of Graves' Ophthalmopathy (GO). Basic fibroblast growth factor (bFGF) and vascular endothelial growth factor (VEGF) have been proposed to contribute to GO, but their effects on orbital fibroblasts are largely unknown.We found

  11. File list: DNS.Oth.05.AllAg.Fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available DNS.Oth.05.AllAg.Fibroblasts hg19 DNase-seq Others Fibroblasts SRX100966,SRX480646,...SRX480648,SRX135564,SRX089279,SRX480647,SRX089280,SRX480645,SRX100965 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/DNS.Oth.05.AllAg.Fibroblasts.bed ...

  12. File list: ALL.Oth.05.AllAg.Fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available ALL.Oth.05.AllAg.Fibroblasts hg19 All antigens Others Fibroblasts SRX396266,SRX4814...X651506,SRX480793,SRX480792,SRX396262 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/ALL.Oth.05.AllAg.Fibroblasts.bed ...

  13. File list: Unc.Oth.50.AllAg.Tail_fibroblast [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Unc.Oth.50.AllAg.Tail_fibroblast mm9 Unclassified Others Tail fibroblast SRX306689,...SRX306688,SRX306687,SRX306686,SRX306691,SRX306690 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/Unc.Oth.50.AllAg.Tail_fibroblast.bed ...

  14. File list: His.Oth.20.AllAg.Fibroblast [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available His.Oth.20.AllAg.Fibroblast mm9 Histone Others Fibroblast SRX024353,SRX024354,SRX02...4352 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/His.Oth.20.AllAg.Fibroblast.bed ...

  15. File list: ALL.Oth.20.AllAg.Fibroblast [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available ALL.Oth.20.AllAg.Fibroblast mm9 All antigens Others Fibroblast SRX191057,SRX024353,...SRX024354,SRX024355,SRX024352 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/ALL.Oth.20.AllAg.Fibroblast.bed ...

  16. File list: ALL.Oth.50.AllAg.Fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available ALL.Oth.50.AllAg.Fibroblasts hg19 All antigens Others Fibroblasts SRX338979,SRX4814...X396257,SRX396265,SRX396258,SRX396256 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/ALL.Oth.50.AllAg.Fibroblasts.bed ...

  17. File list: Unc.Oth.10.AllAg.Tail_fibroblast [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Unc.Oth.10.AllAg.Tail_fibroblast mm9 Unclassified Others Tail fibroblast SRX306689,...SRX306688,SRX306691,SRX306690,SRX306687,SRX306686 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/Unc.Oth.10.AllAg.Tail_fibroblast.bed ...

  18. Effect of β3-adrenoceptor on cardiac fibrosis in rat cardiac fibroblast ...

    African Journals Online (AJOL)

    Purpose: To investigate the effect of β3-adrenoceptors (β3-AR) up-regulation on fibrosis in cardiac fibroblast cells in rats and its potential mechanism. Methods: Cardiac fibroblast cells (CFB) were isolated and identified from rats' hearts. The β3-ARupregulated cardiac fibroblast cells were constructed by lentiviral transfection ...

  19. Enhanced ROCK1 dependent contractility in fibroblast from chronic obstructive pulmonary disease patients.

    Science.gov (United States)

    Hallgren, Oskar; Rolandsson, Sara; Andersson-Sjöland, Annika; Nihlberg, Kristian; Wieslander, Elisabet; Kvist-Reimer, Martina; Dahlbäck, Magnus; Eriksson, Leif; Bjermer, Leif; Erjefält, Jonas S; Löfdahl, Claes-Göran; Westergren-Thorsson, Gunilla

    2012-08-22

    During wound healing processes fibroblasts account for wound closure by adopting a contractile phenotype. One disease manifestation of COPD is emphysema which is characterized by destruction of alveolar walls and our hypothesis is that fibroblasts in the COPD lungs differentiate into a more contractile phenotype as a response to the deteriorating environment. Bronchial (central) and parenchymal (distal) fibroblasts were isolated from lung explants from COPD patients (n = 9) (GOLD stage IV) and from biopsies from control subjects and from donor lungs (n = 12). Tissue-derived fibroblasts were assessed for expression of proteins involved in fibroblast contraction by western blotting whereas contraction capacity was measured in three-dimensional collagen gels. The basal expression of rho-associated coiled-coil protein kinase 1 (ROCK1) was increased in both centrally and distally derived fibroblasts from COPD patients compared to fibroblasts from control subjects (p < 0.001) and (p < 0.01), respectively. Distally derived fibroblasts from COPD patients had increased contractile capacity compared to control fibroblasts (p < 0.01). The contraction was dependent on ROCK1 activity as the ROCK inhibitor Y27632 dose-dependently blocked contraction in fibroblasts from COPD patients. ROCK1-positive fibroblasts were also identified by immunohistochemistry in the alveolar parenchyma in lung tissue sections from COPD patients. Distally derived fibroblasts from COPD patients have an enhanced contractile phenotype that is dependent on ROCK1 activity. This feature may be of importance for the elastic dynamics of small airways and the parenchyma in late stages of COPD.

  20. File list: His.Oth.05.AllAg.Tail_fibroblast [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available His.Oth.05.AllAg.Tail_fibroblast mm9 Histone Others Tail fibroblast SRX335673,SRX33...5668,SRX335667,SRX335674,SRX335665 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/His.Oth.05.AllAg.Tail_fibroblast.bed ...

  1. File list: Pol.Oth.20.AllAg.Tail_fibroblast [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Pol.Oth.20.AllAg.Tail_fibroblast mm9 RNA polymerase Others Tail fibroblast SRX33566...6 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/Pol.Oth.20.AllAg.Tail_fibroblast.bed ...

  2. File list: ALL.Oth.05.AllAg.Tail_fibroblast [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available ALL.Oth.05.AllAg.Tail_fibroblast mm9 All antigens Others Tail fibroblast SRX306690,...5 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/ALL.Oth.05.AllAg.Tail_fibroblast.bed ...

  3. File list: His.Oth.05.AllAg.Fibroblast [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available His.Oth.05.AllAg.Fibroblast mm9 Histone Others Fibroblast SRX024353,SRX024354,SRX02...4352 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/His.Oth.05.AllAg.Fibroblast.bed ...

  4. File list: Pol.Oth.50.AllAg.Tail_fibroblast [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Pol.Oth.50.AllAg.Tail_fibroblast mm9 RNA polymerase Others Tail fibroblast SRX33566...6 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/Pol.Oth.50.AllAg.Tail_fibroblast.bed ...

  5. File list: Oth.Oth.10.AllAg.Fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Oth.Oth.10.AllAg.Fibroblasts hg19 TFs and others Others Fibroblasts SRX396266,SRX39...396255,SRX396268,SRX396252,SRX396257,SRX396260,SRX396256,SRX396253,SRX396265 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/Oth.Oth.10.AllAg.Fibroblasts.bed ...

  6. File list: DNS.Oth.20.AllAg.Fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available DNS.Oth.20.AllAg.Fibroblasts hg19 DNase-seq Others Fibroblasts SRX135564,SRX089279,...SRX480648,SRX100966,SRX480645,SRX480646,SRX089280,SRX100965,SRX480647 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/DNS.Oth.20.AllAg.Fibroblasts.bed ...

  7. File list: ALL.Oth.10.AllAg.Fibroblast [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available ALL.Oth.10.AllAg.Fibroblast mm9 All antigens Others Fibroblast SRX024353,SRX191057,...SRX024355,SRX024354,SRX024352 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/ALL.Oth.10.AllAg.Fibroblast.bed ...

  8. File list: His.Oth.05.AllAg.Fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available His.Oth.05.AllAg.Fibroblasts hg19 Histone Others Fibroblasts SRX651503,SRX651504,SR...X651505,SRX651506,SRX480793,SRX480792 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/His.Oth.05.AllAg.Fibroblasts.bed ...

  9. File list: Oth.Oth.50.AllAg.Fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Oth.Oth.50.AllAg.Fibroblasts hg19 TFs and others Others Fibroblasts SRX338979,SRX48...396255,SRX481483,SRX396260,SRX396266,SRX396257,SRX396265,SRX396258,SRX396256 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/Oth.Oth.50.AllAg.Fibroblasts.bed ...

  10. File list: His.Oth.10.AllAg.Fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available His.Oth.10.AllAg.Fibroblasts hg19 Histone Others Fibroblasts SRX651503,SRX651504,SR...X651505,SRX651506,SRX480793,SRX480792 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/His.Oth.10.AllAg.Fibroblasts.bed ...

  11. File list: Unc.Oth.05.AllAg.Tail_fibroblast [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Unc.Oth.05.AllAg.Tail_fibroblast mm9 Unclassified Others Tail fibroblast SRX306690,...SRX306689,SRX306688,SRX306691,SRX306687,SRX306686 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/Unc.Oth.05.AllAg.Tail_fibroblast.bed ...

  12. File list: DNS.Oth.10.AllAg.Fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available DNS.Oth.10.AllAg.Fibroblasts hg19 DNase-seq Others Fibroblasts SRX100966,SRX135564,...SRX480646,SRX480648,SRX089279,SRX480647,SRX480645,SRX089280,SRX100965 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/DNS.Oth.10.AllAg.Fibroblasts.bed ...

  13. File list: His.Oth.10.AllAg.Fibroblast [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available His.Oth.10.AllAg.Fibroblast mm9 Histone Others Fibroblast SRX024353,SRX024354,SRX02...4352 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/His.Oth.10.AllAg.Fibroblast.bed ...

  14. File list: Oth.Oth.05.AllAg.Fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Oth.Oth.05.AllAg.Fibroblasts hg19 TFs and others Others Fibroblasts SRX396266,SRX48...396251,SRX396255,SRX396257,SRX396268,SRX396260,SRX396253,SRX396256,SRX396265 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/Oth.Oth.05.AllAg.Fibroblasts.bed ...

  15. File list: ALL.Oth.10.AllAg.Tail_fibroblast [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available ALL.Oth.10.AllAg.Tail_fibroblast mm9 All antigens Others Tail fibroblast SRX335673,...5 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/ALL.Oth.10.AllAg.Tail_fibroblast.bed ...

  16. File list: His.Oth.10.AllAg.Tail_fibroblast [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available His.Oth.10.AllAg.Tail_fibroblast mm9 Histone Others Tail fibroblast SRX335673,SRX33...5668,SRX335667,SRX335674,SRX335665 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/His.Oth.10.AllAg.Tail_fibroblast.bed ...

  17. Local food:

    DEFF Research Database (Denmark)

    Sundbo, Donna Isabella Caroline

    2013-01-01

    Recently there has been more focus on food in general and local food in particular. But what is local food? And what are the perceptions of this concept according to theory and to providers and consumers of local food? This article first summarises and compares three different theoretical perspec...... the providers and consumers studied on what local food is. However, this may change in time.......Recently there has been more focus on food in general and local food in particular. But what is local food? And what are the perceptions of this concept according to theory and to providers and consumers of local food? This article first summarises and compares three different theoretical...... perspectives on local food, namely experience economy, local food systems and what is termed pro-industrialism. These have differing and sometimes opposite conceptualisations and aims for the concept of local food. Using the perspective of experience economy as theoretical background, the concept of local food...

  18. Effects of microelectrical current on migration of nasal fibroblasts.

    Science.gov (United States)

    Choi, Hyuk; Cho, Jung-Sun; Park, Il Ho; Yoon, Hu Geun; Lee, Heung-Man

    2011-01-01

    Migration of fibroblasts is critical in wound healing. The question of how wounded electric fields guide migration of nasal fibroblasts remains to be elucidated. This study was designed to determine morphology, directedness, and migration rate of nasal fibroblasts during microcurrent application, which is simulated by an endogenous electric field at the vicinity of the wound. Nasal fibroblasts were exposed to a microelectric field at 50, 100, and 250 mV/mm for 3 hours at 37°C. In this experiment, the field polarity was reversed for an additional 3 hours. During in vitro testing, the cells were incubated in a newly developed miniature, microcurrent generating chamber system, with 5% CO(2), at 37°C; the media was circulated by a pump system. A wound was created by scratching a cell-free area (∼150 μm wide) into a confluent monolayer. The average migration speed was calculated as the distance traveled by the cell divided by time. A microelectric field of 100 mV/mm or more induced significant cell migration in the direction of the cathode. Trajectory speeds at 50, 100, and 250 mV/mm were 9.8 ± 0.3, 11.8 ± 0.3, and 13.5 ± 0.9 μm/mm, respectively. A significant difference was observed between migratory rate of controls and that of 50 mV/mm (p < 0.05). Microelectric fields appear to have a crucial role in control of nasal fibroblast activity in the process of wound healing.

  19. Reprogramming of fibroblast nuclei after transfer into bovine oocytes.

    Science.gov (United States)

    De Sousa, P A; Winger, Q; Hill, J R; Jones, K; Watson, A J; Westhusin, M E

    1999-01-01

    Recent landmark achievements in animal cloning have demonstrated that the events of cell differentiation can, in principle, be reversed. This reversal necessarily requires large-scale genetic reprogramming, of which little is known. In the present study we characterized the extent to which blastocyst stage-specific mRNA expression would be conserved in bovine embryos produced by nuclear transfer (NT) using fetal fibroblasts as nuclei donors (FF NT). The mRNA pool of FF NT embryos was compared with that of NT embryos reconstructed from embryonic blastomeres (Emb NT), with embryos produced under in vivo or in vitro conditions, and finally with fibroblast cells. Embryo/cell-specific mRNA pools were contrasted using differential display methodology. Random oligonucleotide primer pair combinations were used to subfractionate mRNA populations and represent individual mRNAs as copy DNA (cDNA) bands ranging in size from 100 to 800 base pairs. Regardless of whether bovine blastocysts developed in vivo or in vitro, or were derived after nuclear transplantation with embryonic blastomeres or fetal fibroblasts, their mRNA profile was highly conserved and distinct from that of fetal fibroblast cells. There was approximately 95% conservation in cDNA banding patterns between FF NT, Emb NT, and in vivo derived blastocysts, when compared with in vitro derived blastocysts. In contrast, the cDNA banding in fibroblasts was only 67% conserved with in vitro derived blastocysts (p types to serve as nuclear donors for embryo reconstruction and provide information that can be used to improve the efficiency of cloning animals by nuclear transplantation.

  20. A novel role of EMMPRIN/CD147 in transformation of quiescent fibroblasts to cancer-associated fibroblasts by breast cancer cells

    Science.gov (United States)

    Xu, Jing; Lu, Yang; Qiu, Songbo; Chen, Zhi-Nan; Fan, Zhen

    2013-01-01

    We tested the novel hypothesis that EMMPRIN/CD147, a transmembrane glycoprotein overexpressed in breast cancer cells, has a previously unknown role in transforming fibroblasts to cancer-associated fibroblasts, and that cancer-associated fibroblasts in turn induce epithelial-to-mesenchymal transition of breast cancer cells. Co-culture of fibroblasts with breast cancer cells or treatment of fibroblasts with breast cancer cell conditioned culture medium or recombinant EMMPRIN/CD147 induced expression of α-SMA in the fibroblasts in an EMMPRIN/CD147-dependent manner and promoted epithelial-to-mesenchymal transition of breast cancer cells and enhanced cell migration potential. These findings support a novel role of EMMPRIN/CD147 in regulating the interaction between cancer and stroma. PMID:23474495

  1. Site-specific keloid fibroblasts alter the behaviour of normal skin and normal scar fibroblasts through paracrine signalling.

    Directory of Open Access Journals (Sweden)

    Kevin J Ashcroft

    Full Text Available Keloid disease (KD is an abnormal cutaneous fibroproliferative disorder of unknown aetiopathogenesis. Keloid fibroblasts (KF are implicated as mediators of elevated extracellular matrix deposition. Aberrant secretory behaviour by KF relative to normal skin fibroblasts (NF may influence the disease state. To date, no previous reports exist on the ability of site-specific KF to induce fibrotic-like phenotypic changes in NF or normal scar fibroblasts (NS by paracrine mechanisms. Therefore, the aim of this study was to investigate the influence of conditioned media from site-specific KF on the cellular and molecular behaviour of both NF and NS enabled by paracrine mechanisms. Conditioned media was collected from cultured primary fibroblasts during a proliferative log phase of growth including: NF, NS, peri-lesional keloid fibroblasts (PKF and intra-lesional keloid fibroblasts (IKF. Conditioned media was used to grow NF, NS, PKF and IKF cells over 240 hrs. Cellular behavior was monitored through real time cell analysis (RTCA, proliferation rates and migration in a scratch wound assay. Fibrosis-associated marker expression was determined at both protein and gene level. PKF conditioned media treatment of both NF and NS elicited enhanced cell proliferation, spreading and viability as measured in real time over 240 hrs versus control conditioned media. Following PKF and IKF media treatments up to 240 hrs, both NF and NS showed significantly elevated proliferation rates (p<0.03 and migration in a scratch wound assay (p<0.04. Concomitant up-regulation of collagen I, fibronectin, α-SMA, PAI-1, TGF-β and CTGF (p<0.03 protein expression were also observed. Corresponding qRT-PCR analysis supported these findings (P<0.03. In all cases, conditioned media from growing marginal PKF elicited the strongest effects. In conclusion, primary NF and NS cells treated with PKF or IKF conditioned media exhibit enhanced expression of fibrosis-associated molecular markers

  2. Caveolae/lipid rafts in fibroblast-like synoviocytes: ectopeptidase-rich membrane microdomains

    DEFF Research Database (Denmark)

    Riemann, D; Hansen, Gert Helge; Niels-Christiansen, L

    2001-01-01

    Membrane peptidases play important roles in cell activation, proliferation and communication. Human fibroblast-like synoviocytes express considerable amounts of aminopeptidase N/CD13, dipeptidyl peptidase IV/CD26, and neprilysin/CD10, transmembrane proteins previously proposed to be involved...... level, aminopeptidase N/CD13 and dipeptidyl peptidase IV/CD26 were found in caveolae, in particular in intracellular yet surface-connected vesicle-like structures and 'rosettes' made up of several caveolae. In addition, clusters of peptidases were seen at the cell surface in flat patches ranging in size...... from about 60 to 160 nm. Cholesterol depletion of synoviocytes by methyl-beta-cyclodextrin disrupted >90% of the caveolae and reduced the raft localization of aminopeptidase N/CD13 without affecting Ala-p-nitroanilide-cleaving activity of confluent cell cultures. In co-culture experiments with T...

  3. Expression of RANKL in osteolytic membranes: association with fibroblastic cell markers.

    Science.gov (United States)

    Ramage, Samuel C; Urban, Nicole H; Jiranek, William A; Maiti, Aparna; Beckman, Matthew J

    2007-04-01

    Aseptic loosening is often mentioned as the primary reason for costly revision of total joint arthroplasties. Receptor activator of nuclear factor-kappaB ligand (RANKL) appears to be a major factor in the bone resorption observed in periprosthetic osteolysis. RANKL plays an essential role in the recruitment, differentiation, and survival of the osteoclasts implicated in periprosthetic osteolysis. This study was performed in an effort to identify the cell type in the periprosthetic membrane responsible for expression of RANKL. Tissues harvested from osteolytic lesions in nine patients undergoing total joint revision were serially sectioned for immunohistochemical analysis. Intercellular adhesion molecule-1 (ICAM-1) and prolyl 4-hydroxylase (5B5) antibodies were used to detect fibroblasts, and anti-CD-163 (Ber-MAC3) was used to detect macrophages. In addition, antibodies to osteoprotegerin (OPG), RANKL, and receptor activator of nuclear factor-kappaB (RANK) were utilized. The binding pattern of these antibodies was then viewed with confocal microscopy with the use of only secondary antibodies as method controls. Histological analysis was confined to areas of the membrane where cells were detected with use of Hoechst 34580 nuclear stain. In the membrane specimens from all nine patients, diffuse RANKL staining was localized to areas lacking cells and more intense staining was seen in areas containing nucleated cells. There was strong colocalization between RANKL and OPG, and there was weak but specific colocalization between RANKL and both 5B5 and ICAM-1. In contrast, there was complete separation of antibody staining of Ber-MAC3 and RANKL, indicating only generalized overlap of the myeloid markers with the RANKL. RANKL expression was localized to cells that stained positively for fibroblast markers. The data also indicated that there is an intact RANKL/RANK/OPG system in the periprosthetic membrane that could regulate focalized bone resorption in osteolysis

  4. Differential Expression of Matrix Metalloproteases in Human Fibroblasts with Different Origins

    Directory of Open Access Journals (Sweden)

    Diana Lindner

    2012-01-01

    Full Text Available Fibroblasts are widely distributed cells and are responsible for the deposition of extracellular matrix (ECM components but also secrete ECM-degrading matrix metalloproteases. A finely balanced equilibrium between deposition and degradation of ECM is essential for structural integrity of tissues. In the past, fibroblasts have typically been understood as a uniform cell population with comparable functions regardless of their origin. Here, we determined growth curves of fibroblasts derived from heart, skin, and lung and clearly show the lowest proliferation rate for cardiac fibroblasts. Furthermore, we examined basal expression levels of collagen and different MMPs in these three types of fibroblasts and compared these concerning their site of origin. Interestingly, we found major differences in basal mRNA expression especially for MMP1 and MMP3. Moreover, we treated fibroblasts with TNF-α and observed different alterations under these proinflammatory conditions. In conclusion, fibroblasts show different properties in proliferation and MMP expression regarding their originated tissue.

  5. Multiple Directional Differentiation Difference of Neonatal Rat Fibroblasts from Six Organs

    Directory of Open Access Journals (Sweden)

    Yuqiao Chang

    2016-06-01

    Full Text Available Background/Aims: Fibroblasts are abundantly distributed throughout connective tissues in the body and are very important in maintaining the structural and functional integrity. Recent reports have proved that fibroblasts and mesenchymal stem cells share much more in common than previously recognized. The aim of this study was to investigate comparative studies in fibroblasts on the differences in the expression of molecular markers and differentiation capacity from different organs. Methods: Combined trypsin/collagenase enzymes digestion method was used to isolate and culture the fibroblasts derived from heart, liver, spleen, lung, kidney and skin. Cell activity was determined by methyl thiazolyl tetrazolium (MTT assay. Common molecular markers for fibroblasts such as vimentin, DDR2 and FSP1, stem cell markers nanog, c-kit and sca-1 were detected by RT-PCR, immunofluorescence and western blotting. The osteogenic, adipogenic and cardiogenic differentiations of fibroblasts were performed by inductive culture in special mediums, and analyzed by Alizarin red, Oil red O and immunofluorescence staining of cTnT respectively. Results: The proliferation rate of fibroblasts in lung was faster than in other five organs. Common molecular markers for fibroblasts were expressed differently in different organs. DDR2 was strongly expressed in fibroblasts in the heart, partly expressed in the heart, skin, liver and spleen. Interestingly, no expression of DDR2 was detected in liver and kidney. However, vimentin and FSP1 were consistently expressed in fibroblasts from skin, liver, kidney, spleen and lung. nanog expression in fibroblasts from lung was less than that from heart, skin, liver and spleen (P . c-kit expression in fibroblasts from heart, skin and kidney was higher than that from spleen (P , while the c-kit positive fibroblasts from liver was obviously higher than that from spleen (P . But sca-1 expression in fibroblasts from lung was the lowest among six

  6. Local Content

    CSIR Research Space (South Africa)

    Gibberd, Jeremy

    2016-10-01

    Full Text Available is also delineated in order to demonstrate the implications of local content on building design, construction and operation. The advantages and disadvantages of local content approaches are discussed and illustrated through examples. Finally, broad...

  7. RNA-Guided Activation of Pluripotency Genes in Human Fibroblasts

    DEFF Research Database (Denmark)

    Xiong, Kai; Zhou, Yan; Blichfeld, Kristian Aabo

    2017-01-01

    -associated protein 9 (dCas9)-VP64 (CRISPRa) alone, or a combination of dCas9-VP64 and MS2-P65-HSF1 [synergistic activation mediator (SAM) system] mediated activation of five pluripotency genes: KLF4 (K), LIN28 (L), MYC (M), OCT4 (O), and SOX2 (S) in human cells (HEK293T, HeLa, HepG2, and primary fibroblasts...... could be obtained from these SAM fibroblasts. In conclusion, our study showed that CRISPR/Cas9-based ATFs are potent to activate and maintain transcription of endogenous human pluripotent genes. However, future improvements of the system are still required to improve activation efficiency and cellular...

  8. Bio-artificial pleura using an autologous dermal fibroblast sheet

    Science.gov (United States)

    Kanzaki, Masato; Takagi, Ryo; Washio, Kaoru; Kokubo, Mami; Yamato, Masayuki

    2017-10-01

    Air leaks (ALs) are observed after pulmonary resections, and without proper treatment, can produce severe complications. AL prevention is a critical objective for managing patients after pulmonary resection. This study applied autologous dermal fibroblast sheets (DFS) to close ALs. For sealing ALs in a 44-year-old male human patient with multiple bullae, a 5 × 15-mm section of skin was surgically excised. From this skin specimen, primary dermal fibroblasts were isolated and cultured for 4 weeks to produce DFSs that were harvested after a 10-day culture. ALs were completely sealed using surgical placement of these autologous DFSs. DFS were found to be a durable long-term AL sealant, exhibiting requisite flexibility, elasticity, durability, biocompatibility, and usability, resulting reliable AL closure. DFS should prove to be an extremely useful tissue-engineered pleura substitute.

  9. Extracellular depolymerization of hyaluronic acid in cultured human skin fibroblasts

    Energy Technology Data Exchange (ETDEWEB)

    Nakamura, T.; Takagaki, K.; Kubo, K.; Morikawa, A.; Tamura, S.; Endo, M. (Hirosaki Univ. School of Medicine (Japan))

    1990-10-15

    The chain length of ({sup 3}H)hyaluronic acid synthesized by cultivating human skin fibroblasts in the presence of ({sup 3}H)glucosamine was investigated. ({sup 3}H)Hyaluronic acid obtained from the matrix fraction was excluded from a Sepharose CL-2B column irrespective of the incubation period, whereas that from the medium was depolymerized into a constant chain length (Mr = 40,000). The reducing and non-reducing terminals of the depolymerized hyaluronic acid were N-acetylglucosamine and glucuronic acid, respectively. Prolonged incubation produced no oligosaccharides as shown by examination of hyaluronidase digests, suggesting the presence of a novel endo-beta-N-acetylglucosaminidase in cultured human skin fibroblasts.

  10. Local complications

    NARCIS (Netherlands)

    van den Akker, H.P.; Baart, J.A.; Baart, J.A.; Brand, H.S.

    2017-01-01

    Local anaesthesia is frequently used in dentistry and seldom leads to serious local complications. Nevertheless, it is of great importance to be aware of the causes of each local complication and – if necessary – implement correct treatment. The patient must be informed extensively and, if

  11. Fibroblasts and the extracellular matrix in right ventricular disease.

    Science.gov (United States)

    Frangogiannis, Nikolaos G

    2017-10-01

    Right ventricular failure predicts adverse outcome in patients with pulmonary hypertension (PH), and in subjects with left ventricular heart failure and is associated with interstitial fibrosis. This review manuscript discusses the cellular effectors and molecular mechanisms implicated in right ventricular fibrosis. The right ventricular interstitium contains vascular cells, fibroblasts, and immune cells, enmeshed in a collagen-based matrix. Right ventricular pressure overload in PH is associated with the expansion of the fibroblast population, myofibroblast activation, and secretion of extracellular matrix proteins. Mechanosensitive transduction of adrenergic signalling and stimulation of the renin-angiotensin-aldosterone cascade trigger the activation of right ventricular fibroblasts. Inflammatory cytokines and chemokines may contribute to expansion and activation of macrophages that may serve as a source of fibrogenic growth factors, such as transforming growth factor (TGF)-β. Endothelin-1, TGF-βs, and matricellular proteins co-operate to activate cardiac myofibroblasts, and promote synthesis of matrix proteins. In comparison with the left ventricle, the RV tolerates well volume overload and ischemia; whether the right ventricular interstitial cells and matrix are implicated in these favourable responses remains unknown. Expansion of fibroblasts and extracellular matrix protein deposition are prominent features of arrhythmogenic right ventricular cardiomyopathies and may be implicated in the pathogenesis of arrhythmic events. Prevailing conceptual paradigms on right ventricular remodelling are based on extrapolation of findings in models of left ventricular injury. Considering the unique embryologic, morphological, and physiologic properties of the RV and the clinical significance of right ventricular failure, there is a need further to dissect RV-specific mechanisms of fibrosis and interstitial remodelling. Published on behalf of the European Society of

  12. Rosmarinic acid potentiates carnosic acid induced apoptosis in lung fibroblasts.

    Science.gov (United States)

    Bahri, Sana; Mies, Frédérique; Ben Ali, Ridha; Mlika, Mona; Jameleddine, Saloua; Mc Entee, Kathleen; Shlyonsky, Vadim

    2017-01-01

    Pulmonary fibrosis is characterized by over-population and excessive activation of fibroblasts and myofibroblasts disrupting normal lung structure and functioning. Rosemary extract rich in carnosic acid (CA) and rosmarinic acid (RA) was reported to cure bleomycin-(BLM)-induced pulmonary fibrosis. We demonstrate that CA decreased human lung fibroblast (HLF) viability with IC50 value of 17.13±1.06 μM, while RA had no cytotoxic effect. In the presence of 50 μM of RA, dose-response for CA shifted to IC50 value of 11.70±1.46 μM, indicating synergic action. TGFβ-transformed HLF, rat lung fibroblasts and L929 cells presented similar sensitivity to CA and CA+RA (20μM+100μM, respectively) treatment. Rat alveolar epithelial cells died only under CA+RA treatment, while A549 cells were not affected. Annexin V staining and DNA quantification suggested that HLF are arrested in G0/G1 cell cycle phase and undergo apoptosis. CA caused sustained activation of phospho-Akt and phospho-p38 expression and inhibition of p21 protein.Addition of RA potentiated these effects, while RA added alone had no action.Only triple combination of inhibitors (MAPK-p38, pan-caspase, PI3K/Akt/autophagy) partially attenuated apoptosis; this suggests that cytotoxicity of CA+RA treatment has a complex mechanism involving several parallel signaling pathways. The in vivo antifibrotic effect of CA and RA was compared with that of Vitamine-E in BLM-induced fibrosis model in rats. We found comparable reduction in fibrosis score by CA, RA and CA+RA, attenuation of collagen deposition and normalization of oxidative stress markers. In conclusion, antifibrotic effect of CA+RA is due to synergistic pro-apoptotic action on lung fibroblasts and myofibroblasts.

  13. Inhibition of normal human lung fibroblast growth by beryllium.

    Science.gov (United States)

    Lehnert, N M; Gary, R K; Marrone, B L; Lehnert, B E

    2001-03-07

    Inhalation of particulate beryllium (Be) and its compounds causes chronic Be disease (CBD) in a relatively small subset ( approximately 1-6%) of exposed individuals. Hallmarks of this pulmonary disease include increases in several cell types, including lung fibroblasts, that contribute to the fibrotic component of the disorder. In this regard, enhancements in cell proliferation appear to play a fundamental role in CBD development and progression. Paradoxically, however, some existing evidence suggests that Be actually has antiproliferative effects. In order to gain further information about the effects of Be on cell growth, we: (1) assessed cell proliferation and cell cycle effects of low concentrations of Be in normal human diploid fibroblasts, and (2) investigated the molecular pathway(s) by which the cell cycle disturbing effects of Be may be mediated. Treatment of human lung and skin fibroblasts with Be added in the soluble form of BeSO(4) (0.1-100 microM) caused inhibitions of their growth in culture in a concentration-dependent manner. Such growth inhibition was found to persist, even after cells were further cultured in Be(2+)-free medium. Flow cytometric analyses of cellular DNA labeled with the DNA-binding fluorochrome DAPI revealed that Be causes a G(0)-G(1)/pre-S phase arrest. Western blot analyses indicated that the Be-induced G(0)-G(1)/pre-S phase arrest involves elevations in TP53 (p53) and the cyclin-dependent kinase inhibitor CDKN1A (p21(Waf-1,Cip1)). That Be at low concentrations inhibits the growth of normal human fibroblasts suggests the possibility of the existence of abnormal cell cycle inhibitory responses to Be in individuals who are sensitive to the metal and ultimately develop CBD.

  14. Rosmarinic acid potentiates carnosic acid induced apoptosis in lung fibroblasts.

    Directory of Open Access Journals (Sweden)

    Sana Bahri

    Full Text Available Pulmonary fibrosis is characterized by over-population and excessive activation of fibroblasts and myofibroblasts disrupting normal lung structure and functioning. Rosemary extract rich in carnosic acid (CA and rosmarinic acid (RA was reported to cure bleomycin-(BLM-induced pulmonary fibrosis. We demonstrate that CA decreased human lung fibroblast (HLF viability with IC50 value of 17.13±1.06 μM, while RA had no cytotoxic effect. In the presence of 50 μM of RA, dose-response for CA shifted to IC50 value of 11.70±1.46 μM, indicating synergic action. TGFβ-transformed HLF, rat lung fibroblasts and L929 cells presented similar sensitivity to CA and CA+RA (20μM+100μM, respectively treatment. Rat alveolar epithelial cells died only under CA+RA treatment, while A549 cells were not affected. Annexin V staining and DNA quantification suggested that HLF are arrested in G0/G1 cell cycle phase and undergo apoptosis. CA caused sustained activation of phospho-Akt and phospho-p38 expression and inhibition of p21 protein.Addition of RA potentiated these effects, while RA added alone had no action.Only triple combination of inhibitors (MAPK-p38, pan-caspase, PI3K/Akt/autophagy partially attenuated apoptosis; this suggests that cytotoxicity of CA+RA treatment has a complex mechanism involving several parallel signaling pathways. The in vivo antifibrotic effect of CA and RA was compared with that of Vitamine-E in BLM-induced fibrosis model in rats. We found comparable reduction in fibrosis score by CA, RA and CA+RA, attenuation of collagen deposition and normalization of oxidative stress markers. In conclusion, antifibrotic effect of CA+RA is due to synergistic pro-apoptotic action on lung fibroblasts and myofibroblasts.

  15. Antioxidants successfully reduce ROS production in propionic acidemia fibroblasts.

    Science.gov (United States)

    Gallego-Villar, Lorena; Pérez, Belén; Ugarte, Magdalena; Desviat, Lourdes R; Richard, Eva

    2014-09-26

    Propionic acidemia (PA), caused by a deficiency of the mitochondrial biotin dependent enzyme propionyl-CoA carboxylase (PCC) is one of the most frequent organic acidurias in humans. Most PA patients present in the neonatal period with metabolic acidosis and hyperammonemia, developing different neurological symptoms, movement disorders and cardiac complications. There is strong evidence indicating that oxidative damage could be a pathogenic factor in neurodegenerative, mitochondrial and metabolic diseases. Recently, we identified an increase in ROS levels in PA patients-derived fibroblasts. Here, we analyze the capability of seven antioxidants to scavenge ROS production in PA patients' cells. Tiron, trolox, resveratrol and MitoQ significantly reduced ROS content in patients and controls' fibroblasts. In addition, changes in the expression of two antioxidant enzymes, superoxide dismutase and glutathione peroxidase, were observed in PA patients-derived fibroblasts after tiron and resveratrol treatment. Our results in PA cellular models establish the proof of concept of the potential of antioxidants as an adjuvant therapy for PA and pave the way for future assessment of antioxidant strategies in the murine model of PA. Copyright © 2014 Elsevier Inc. All rights reserved.

  16. Effect of periodontal dressings on human gingiva fibroblasts in vitro

    Energy Technology Data Exchange (ETDEWEB)

    Eber, R.M.; Shuler, C.F.; Buchanan, W.; Beck, F.M.; Horton, J.E. (Ohio State Univ. College of Dentistry, Columbus (USA))

    1989-08-01

    In vitro cytotoxicity studies of periodontal dressings have not generally produced a result consistent with in vivo observations. These prior in vitro studies have not used human intraoral cell lines. We tested the effects of two eugenol containing and two non-eugenol periodontal dressings on cultured human gingival fibroblasts (HGF) (ATCC No. 1292). Replicate HGF cultures grown in microtiter plates were exposed to stock, 1:4 and 1:16 dilutions of extracts made from each of the four periodontal dressings. The HGF cultures were pulse labelled with tritiated thymidine (3HTdR) after 24, 48, and 72 hours. Incorporations of the labelled thymidine were measured using liquid scintillation counting and expressed as counts per minute. The results showed that undiluted extracts from all four periodontal dressings totally inhibited 3HTdR uptake (P less than 0.05). The 1:4 dilution of eugenol dressings inhibited 3HTdR uptake significantly more than non-eugenol dressings (P less than 0.05). Interestingly, at 72 hours the 1:16 dilution of the non-eugenol dressings caused significantly increased 3HTdR uptake which was not observed with the eugenol dressings. The present results suggest that the use of a human fibroblastic cell line for testing the effects of periodontal dressings may provide information about the relative biological effects of these dressings. Using this cell line, we have found that eugenol dressings inhibit fibroblast proliferation to a greater extent than non-eugenol dressings.

  17. Effect of phenytoin and age on gingival fibroblast enzymes.

    Directory of Open Access Journals (Sweden)

    Surena Vahabi

    2014-06-01

    Full Text Available The alteration of cytokine balance is stated to exert greater influence on gingival overgrowth compared to the direct effect of the drug on the regulation of extracellular matrix metabolism. The current study evaluated the effect of phenytoin on the regulation of collagen, lysyl oxidase and elastin in gingival fibroblasts.Normal human gingival fibroblasts (HGFs were obtained from 4 healthy children and 4 adults. Samples were cultured with phenytoin. MTT test was used to evaluate the proliferation and ELISA was performed to determine the level of IL1β and PGE2 production by HGFs. Total RNA of gingival fibroblasts was extracted and RT-PCR was performed on samples. Mann-Whitney U test was used to analyze the data with an alpha error level less than 0.05.There was a significant difference in the expression of elastin between the controls and treated samples in both adult and pediatric groups and also in the lysyl oxidase expression of adult controls and treated adults. No significant difference was found between collagen expression in adults.The significant difference in elastin and lysyl oxidase expression between adult and pediatric samples indicates the significant effect of age on their production.

  18. Structural alterations in fibroblast monolayers caused by mast cell degranulation.

    Science.gov (United States)

    Ginsburg, H; Amira, M; Padawer, J; Davidson, S

    1989-06-01

    Populations of mature, long-lived, nondividing mast cells develop on embryonic fibroblast monolayers after 1 mo growth of lymph node cells taken from mice immunized with horse serum. Total mast cell degranulation with 80-90% histamine release has been obtained by monoclonal anti-2,4-dinitrophenol (anti-DNP) IgE and the antigen. This degranulation process was studied by time-lapse cinematography and scanning electron microscopy. Excitation of the mast cells began as early as 10 s after addition of the antigen and lasted for about 15 s. Consequently, the fibroblast cytoplasm was displaced by short 5-10 s movements. Before degranulation, due to an extracellular film that coated the cells and the extracellular fibers, the monolayer appeared as a continuous, uninterrupted layer. After degranulation and fibroblast cytoplasm displacement, the fibrous network was exposed. Several inhibitors and antagonists of mast cell mediators were introduced to the cultures prior to addition of the antigen. So far, only with soybean trypsin inhibitor was the cytoplasm dislocation inhibited. Histamine H1 and H2 and serotonin receptor antagonists, as well as indomethacin, cortisol, aprotinin, and phenylmethylsulfonyl fluoride, did not inhibit. These results suggest that chymase, which constitutes the greater part of the mast cell granule protein, is the causative agent.

  19. Potensi Terapeutik Fibroblast Growth Factor 21 terhadap Resistensi Insulin

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    Kurniasari Kurniasari

    2015-12-01

    Full Text Available Fibroblast growth factor 21 (FGF21 merupakan salah satu dari anggota FGF yang berperansebagai faktor endokrin. Hepar dan jaringan adiposa merupakan tempat kerja utama FGF21.Ekspresi FGF21 di hepar diatur oleh peroxisome proliferator activated receptor alpha (PPARαsedangkan di jaringan adiposa diatur oleh peroxisome proliferator activated receptor gamma(PPARγ. Kedua faktor transkripsi tersebut terlibat dalam metabolisme karbohidrat dan lipid. Padaresistensi insulin terdapat hiperglikemia, hiperinsulinemia, dan dislipidemia. Pemberian FGF21pada berbagai studi in vivo dan in vitro telah menunjukan potensi FGF21 dalam mengatasi kelainanakibat resistensi insulin sekaligus meningkatkan sensitivitas jaringan terhadap insulin. Kata kunci: FGF21, PPARγ, PPARα, resistensi insulin Fibroblast Growth Factor 21 (FGF21 Potension in InsulinResistance Treatment Abstract Fibroblast growth factor 21 (FGF21 is a member of FGF family that plays a role as endocrinefactor. Liver and adipose tissue are major target of FGF21. The expression of FGF21 in liveris regulated by peroxisome proliferator activated receptor alpha (PPARα, while peroxisomeproliferator activated receptor gamma (PPARγ regulate FGF21 expression in adipose tissue.Both transcription factors are involved in carbohydrate and lipid metabolism. Hyperglycemia,hyperinsulinemia, and dyslipidemia are observed in insulin resistance. Treatment with FGF21 inin vitro and in vivo study showed that FGF21 have the potential to overcome insulin resistance aswell as increasing tissue’s sensitivity towards insulin. Keywords: FGF21, PPARγ, PPARα, insulin resistance Normal 0 false false false IN X-NONE X-NONE

  20. Comparative proteomic analysis of fibrosarcoma and skin fibroblast cell lines.

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    Meral, Ogunc; Uysal, Hamdi

    2015-02-01

    Comparative proteomic analysis of normal and cancer cell lines provides for a better understanding of the molecular mechanism of cancer development and is essential for developing more effective strategies for new biomarker or drug target discovery. The purpose of this study is to compare protein expression levels between fibrosarcoma and fibroblast cell lines. In our study, two-dimensional polyacrylamide gel electrophoresis (2-D PAGE) and liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) techniques were carried out to compare the protein profile between fibrosarcoma and fibroblast cell lines. We prepared cell lysate samples to analyze intracellular proteins and secretome samples to analyze extracellular proteins in both cell lines. Our results revealed 13 upregulated proteins and 1 downregulated protein of which all of them identified in fibrosarcoma cell line after the comparison with fibroblast cell line cell lysates. When comparing secretome profiles of both cell lines, we found and identified 13 proteins only expressed in fibrosarcoma cell line. These identified proteins have common functions such as cell proliferation, cell differentiation, invasion, metastasis, and apoptosis in cancer. The data obtained from this study indicates that these proteins have importance on understanding the molecular mechanism of fibrosarcoma. These proteins may serve as candidate biomarkers and drug targets for future clinical studies.

  1. Hierarchical mechanisms for direct reprogramming of fibroblasts to neurons.

    Science.gov (United States)

    Wapinski, Orly L; Vierbuchen, Thomas; Qu, Kun; Lee, Qian Yi; Chanda, Soham; Fuentes, Daniel R; Giresi, Paul G; Ng, Yi Han; Marro, Samuele; Neff, Norma F; Drechsel, Daniela; Martynoga, Ben; Castro, Diogo S; Webb, Ashley E; Südhof, Thomas C; Brunet, Anne; Guillemot, Francois; Chang, Howard Y; Wernig, Marius

    2013-10-24

    Direct lineage reprogramming is a promising approach for human disease modeling and regenerative medicine, with poorly understood mechanisms. Here, we reveal a hierarchical mechanism in the direct conversion of fibroblasts into induced neuronal (iN) cells mediated by the transcription factors Ascl1, Brn2, and Myt1l. Ascl1 acts as an "on-target" pioneer factor by immediately occupying most cognate genomic sites in fibroblasts. In contrast, Brn2 and Myt1l do not access fibroblast chromatin productively on their own; instead, Ascl1 recruits Brn2 to Ascl1 sites genome wide. A unique trivalent chromatin signature in the host cells predicts the permissiveness for Ascl1 pioneering activity among different cell types. Finally, we identified Zfp238 as a key Ascl1 target gene that can partially substitute for Ascl1 during iN cell reprogramming. Thus, a precise match between pioneer factors and the chromatin context at key target genes is determinative for transdifferentiation to neurons and likely other cell types. Copyright © 2013 Elsevier Inc. All rights reserved.

  2. HCMV Induces Macropinocytosis for Host Cell Entry in Fibroblasts.

    Science.gov (United States)

    Hetzenecker, Stefanie; Helenius, Ari; Krzyzaniak, Magdalena Anna

    2016-04-01

    Human cytomegalovirus (HCMV) is an important and widespread pathogen in the human population. While infection by this β-herpesvirus in endothelial, epithelial and dendritic cells depends on endocytosis, its entry into fibroblasts is thought to occur by direct fusion of the viral envelope with the plasma membrane. To characterize individual steps during entry in primary human fibroblasts, we employed quantitative assays as well as electron, fluorescence and live cell microscopy in combination with a variety of inhibitory compounds. Our results showed that while infectious entry was pH- and clathrin-independent, it required multiple, endocytosis-related factors and processes. The virions were found to undergo rapid internalization into large vacuoles containing internalized fluid and endosome markers. The characteristics of the internalization process fulfilled major criteria for macropinocytosis. Moreover, we found that soon after addition to fibroblasts the virus rapidly triggered the formation of circular dorsal ruffles in the host cell followed by the generation of large macropinocytic vacuoles. This distinctive form of macropinocytosis has been observed especially in primary cells but has not previously been reported in response to virus stimulation. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  3. Effects of blue light irradiation on human dermal fibroblasts.

    Science.gov (United States)

    Opländer, Christian; Hidding, Sarah; Werners, Frauke B; Born, Matthias; Pallua, Norbert; Suschek, Christoph V

    2011-05-03

    Previous studies have reported that separately from UV-radiation also blue light influences cellular physiology in different cell types. However, little is known about the blue light action spectrum. The purpose of this study was to investigate effects of blue light at distinct wavelengths (410, 420, 453, 480 nm) emitted by well defined light-emitting-diodes on viability, proliferation and antioxidative capacity of human dermal fibroblasts. We found that irradiation with blue light (410, 420 nm) led to intracellular oxidative stress and toxic effects in a dose and wavelength dependent manner. No toxicity was observed using light at 453 nm and 480 nm. Furthermore, blue light (410, 420, 453 nm) at low doses reduced the antioxidative capacity of fibroblasts. At non-toxic doses, irradiations at 410, 420 and 453 nm reduced proliferation indicating a higher susceptibility of proliferating fibroblasts to blue light. Our results show that blue light at different wavelengths may induce varying degrees of intracellular oxidative stress with different physiological outcome, which could contribute to premature skin photoaging. On the other hand, the use of blue light due to its antiproliferative and toxic properties may represent a new approach in treatment and prevention of keloids, hypertrophic scars and fibrotic skin diseases. Copyright © 2011 Elsevier B.V. All rights reserved.

  4. Activity-dependent neuronal control of gap-junctional communication in fibroblasts.

    Science.gov (United States)

    Zeng, Yan; Lv, Xiaohua; Zeng, Shaoqun; Shi, Jing

    2009-07-14

    Intercellular communication through gap junctions plays an important role in fibroblast physiology. Here we report an activity-dependent neuronal control of gap-junctional communication (GJC) in rat dermal fibroblasts, which is associated with N-methyl-D-aspartate (NMDA) glutamate receptor-mediated elevations in intracellular Ca(2+). Both spontaneous and induced activation of dorsal root ganglion (DRG) neurons, manifested by action potentials and TTX-dependent inward currents, significantly reduced fibroblast GJC in Neuron/Fibroblast (N/F) cocultures. Reduced fibroblast GJC by DRG neurons was prevented by blockade of NMDA receptors and decrease of intra- and intercellular Ca(2+). Immunocytochemistry showed that the NR1 subunit of the NMDA receptor coexisted with CX43 at the plasma membrane of fibroblasts. Moreover, glutamate applied to fibroblast cultures triggered NMDA receptor-dependent intracellular Ca(2+) elevations and decrease of GJC. These data demonstrate that NMDA receptor activation contributes to downregulation of GJC in fibroblasts. Since fibroblasts have been shown to facilitate DRG neurite growth and survival, our findings provide a glimpse at the impact that neuronal activation has on fibroblast networks.

  5. A FTIR imaging characterization of fibroblasts stimulated by various breast cancer cell lines.

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    Saroj Kumar

    Full Text Available It is well known that the microenvironment plays a major role in breast cancer progression. Yet, the mechanism explaining the transition from normal fibroblasts to cancer-stimulated fibroblasts remains to be elucidated. Here we report a FTIR imaging study of the effects of three different breast cancer cell lines on normal fibroblasts in culture. Fibroblast activation process was monitored by FTIR imaging and spectra compared by multivariate statistical analyses. Principal component analysis evidenced that the fibroblasts stimulated by these cancer cell lines grouped together and remained distinctly separated from normal fibroblasts indicating a modified different chemical composition in the cancer-stimulated fibroblasts. Similar changes in fibroblasts were induced by the various breast cancer cell lines belonging to different sub-types. Most significant changes were observed in the region of 2950 and 1230 cm(-1, possibly related to changes in lipids and in the 1230 cm(-1 area assigned to phosphate vibrations (nucleotides. Interestingly, the cancer-cell induced changes in the fibroblasts also occurred when there was no possible direct contact between the two cell lines in the co-culture. When contact was possible, the spectral changes were similar, suggesting that soluble factors but not direct cell-cell interactions were responsible for fibroblast activation. Overall, the results indicate that IR imaging could be used in the future for analyzing the microenvironment of breast tumors.

  6. Fibroblast adhesion and activation onto micro-machined titanium surfaces.

    Science.gov (United States)

    Guillem-Marti, J; Delgado, L; Godoy-Gallardo, M; Pegueroles, M; Herrero, M; Gil, F J

    2013-07-01

    Surface modifications performed at the neck of dental implants, in the manner of micro-grooved surfaces, can reduce fibrous tissue encapsulation and prevent bacterial colonization, thereby improving fibrointegration and the formation of a biological seal. However, the applied procedures are technically complex and/or time consuming methods. The aim of this study was to analyse the fibroblast behaviour on modified titanium surfaces obtained, applying a simple and low-cost method. An array of titanium surfaces was obtained using a commercial computerized numerical control lathe, modifying the feed rate and the cutting depth. To elucidate the potential ability of the generated surfaces to activate connective tissue cells, a thorough gene (by real time - qPCR) and protein (by western blot or zymography) expression and cellular response characterization (cell morphology, cell adhesion and cell activation by secreting extracellular matrix (ECM) components and their enzyme regulators) was performed. Micro-grooved surfaces have statistically significant differences in the groove's width (approximately 10, 50 and 100 μm) depending on the applied advancing fixed speed. Field emission scanning electron microscopy images showed that fibroblasts oriented along the generated grooves, but they were only entirely accommodated on the wider grooves (≥50 μm). Micro-grooved surfaces exhibited an earlier cell attachment and activation, as seen by collagen Iα1 and fibronectin deposition and activation of ECM remodelling enzymes, compared with the other surfaces. However, fibroblasts could remain in an activated state on narrower surfaces (micro-grooved surfaces could improve implant integration at the gingival site with respect to polished surfaces. Micro-grooved surfaces enhance early fibroblast adhesion and activation, which could be critical for the formation of a biological seal and finally promote tissue integration. Surfaces with wider grooves (≥50 μm) seem to be more

  7. Human gingival fibroblasts display a non-fibrotic phenotype distinct from skin fibroblasts in three-dimensional cultures.

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    Wesley Mah

    Full Text Available Scar formation following skin injury can be a major psychosocial and physiological problem. However, the mechanisms of scar formation are still not completely understood. Previous studies have shown that wound healing in oral mucosa is faster, associates with a reduced inflammatory response and results to significantly reduced scar formation compared with skin wounds. In the present study, we hypothesized that oral mucosal fibroblasts from human gingiva are inherently distinct from fibroblasts from breast and abdominal skin, two areas prone to excessive scar formation, which may contribute to the preferential wound healing outcome in gingiva. To this end, we compared the phenotype of human gingival and skin fibroblasts cultured in in vivo-like three-dimensional (3D cultures that mimic the cells' natural extracellular matrix (ECM niche. To establish 3D cultures, five parallel fibroblast lines from human gingiva (GFBLs and breast skin (SFBLs were seeded in high density, and cultured for up to 21 days in serum and ascorbic acid containing medium to induce expression of wound-healing transcriptome and ECM deposition. Cell proliferation, morphology, phenotype and expression of wound healing and scar related genes were analyzed by real-time RT-PCR, Western blotting and immunocytochemical methods. The expression of a set of genes was also studied in three parallel lines of human abdominal SFBLs. Findings showed that GFBLs displayed morphologically distinct organization of the 3D cultures and proliferated faster than SFBLs. GFBLs expressed elevated levels of molecules involved in regulation of inflammation and ECM remodeling (MMPs while SFBLs showed significantly higher expression of TGF-β signaling, ECM and myofibroblast and cell contractility-related genes. Thus, GFBLs display an inherent phenotype conducive for fast resolution of inflammation and ECM remodeling, characteristic for scar-free wound healing, while SFBLs have a profibrotic, scar

  8. Lithium Attenuates TGF-β 1-Induced Fibroblasts to Myofibroblasts Transition in Bronchial Fibroblasts Derived from Asthmatic Patients

    Science.gov (United States)

    Michalik, Marta; Wójcik, Katarzyna Anna; Jakieła, Bogdan; Szpak, Katarzyna; Pierzchalska, Małgorzata; Sanak, Marek; Madeja, Zbigniew; Czyż, Jarosław

    2012-01-01

    Bronchial asthma is a chronic disorder accompanied by phenotypic transitions of bronchial epithelial cells, smooth muscle cells, and fibroblasts. Human bronchial fibroblasts (HBFs) derived from patients with diagnosed asthma display predestination towards TGF-β-induced phenotypic switches. Since the interference between TGF-β and GSK-3β signaling contributes to pathophysiology of chronic lung diseases, we investigated the effect of lithium, a nonspecific GSK-3β inhibitor, on TGF-β 1-induced fibroblast to myofibroblast transition (FMT) in HBF and found that the inhibition of GSK-3β attenuates TGF-β 1-induced FMT in HBF populations derived from asthmatic but not healthy donors. Cytoplasmically sequestrated β-catenin, abundant in TGF-β 1/LiCl-stimulated asthmatic HBFs, most likely interacts with and inhibits the nuclear accumulation and signal transduction of Smad proteins. These data indicate that the specific cellular context determines FMT-related responses of HBFs to factors interfering with the TGF-β signaling pathway. They may also provide a mechanistic explanation for epidemiological data revealing coincidental remission of asthmatic syndromes and their recurrence upon the discontinuation of lithium therapy in certain psychiatric diseases. PMID:22988467

  9. Lithium Attenuates TGF-β1-Induced Fibroblasts to Myofibroblasts Transition in Bronchial Fibroblasts Derived from Asthmatic Patients

    Directory of Open Access Journals (Sweden)

    Marta Michalik

    2012-01-01

    Full Text Available Bronchial asthma is a chronic disorder accompanied by phenotypic transitions of bronchial epithelial cells, smooth muscle cells, and fibroblasts. Human bronchial fibroblasts (HBFs derived from patients with diagnosed asthma display predestination towards TGF-β-induced phenotypic switches. Since the interference between TGF-β and GSK-3β signaling contributes to pathophysiology of chronic lung diseases, we investigated the effect of lithium, a nonspecific GSK-3β inhibitor, on TGF-β1-induced fibroblast to myofibroblast transition (FMT in HBF and found that the inhibition of GSK-3β attenuates TGF-β1-induced FMT in HBF populations derived from asthmatic but not healthy donors. Cytoplasmically sequestrated β-catenin, abundant in TGF-β1/LiCl-stimulated asthmatic HBFs, most likely interacts with and inhibits the nuclear accumulation and signal transduction of Smad proteins. These data indicate that the specific cellular context determines FMT-related responses of HBFs to factors interfering with the TGF-β signaling pathway. They may also provide a mechanistic explanation for epidemiological data revealing coincidental remission of asthmatic syndromes and their recurrence upon the discontinuation of lithium therapy in certain psychiatric diseases.

  10. Lithium Attenuates TGF-β(1)-Induced Fibroblasts to Myofibroblasts Transition in Bronchial Fibroblasts Derived from Asthmatic Patients.

    Science.gov (United States)

    Michalik, Marta; Wójcik, Katarzyna Anna; Jakieła, Bogdan; Szpak, Katarzyna; Pierzchalska, Małgorzata; Sanak, Marek; Madeja, Zbigniew; Czyż, Jarosław

    2012-01-01

    Bronchial asthma is a chronic disorder accompanied by phenotypic transitions of bronchial epithelial cells, smooth muscle cells, and fibroblasts. Human bronchial fibroblasts (HBFs) derived from patients with diagnosed asthma display predestination towards TGF-β-induced phenotypic switches. Since the interference between TGF-β and GSK-3β signaling contributes to pathophysiology of chronic lung diseases, we investigated the effect of lithium, a nonspecific GSK-3β inhibitor, on TGF-β(1)-induced fibroblast to myofibroblast transition (FMT) in HBF and found that the inhibition of GSK-3β attenuates TGF-β(1)-induced FMT in HBF populations derived from asthmatic but not healthy donors. Cytoplasmically sequestrated β-catenin, abundant in TGF-β(1)/LiCl-stimulated asthmatic HBFs, most likely interacts with and inhibits the nuclear accumulation and signal transduction of Smad proteins. These data indicate that the specific cellular context determines FMT-related responses of HBFs to factors interfering with the TGF-β signaling pathway. They may also provide a mechanistic explanation for epidemiological data revealing coincidental remission of asthmatic syndromes and their recurrence upon the discontinuation of lithium therapy in certain psychiatric diseases.

  11. The impact of vascular endothelial growth factor and basic fibroblast growth factor on cardiac fibroblasts grown under altered gravity conditions.

    Science.gov (United States)

    Ulbrich, Claudia; Leder, Annekatrin; Pietsch, Jessica; Flick, Burkhard; Wehland, Markus; Grimm, Daniela

    2010-01-01

    Myocardium is very sensitive to gravitational changes. During a spaceflight cardiovascular atrophy paired with rhythm problems and orthostatic intolerance can occur. The aim of this study was to investigate the impact of basic fibroblast growth factor (bFGF) and vascular endothelial growth factor (VEGF) on cardiac fibroblasts (CF) grown under altered gravity conditions. We examined the influence of exposure to a Random Positioning Machine (RPM) on CF, derived from porcine hearts. We focused on growth, extracellular matrix protein (ECMP) synthesis and apoptosis. When cultured on a RPM, CF began to form 3D spheroids within 24h, irrespective of growth factor treatment. Exposure to RPM induced an increased synthesis of ECMP and also resulted in elevated apoptosis in adherent CF as measured by terminal deoxynucleotidyl transferase-mediated dUTP digoxigenin nick end labeling (TUNEL) analysis, 4',6-diamidino-2-phenylindole (DAPI) staining, and caspase-3 detection. bFGF and VEGF significantly decreased the amount of ECMP (collagen type I, III, chondroitin sulfate) in 1g and RPM cultures, and also significantly reduced the amount of apoptotic CF as well as caspase-3. Altered gravity conditions on a RPM induced 3D growth, elevated ECMP synthesis and apoptosis in cardiac fibroblasts. Growth factor treatment attenuated programmed cell death and ECMP secretion. Copyright © 2010 S. Karger AG, Basel.

  12. Hypoxia-Induced Collagen Synthesis of Human Lung Fibroblasts by Activating the Angiotensin System

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    Shan-Shan Liu

    2013-12-01

    Full Text Available The exact molecular mechanism that mediates hypoxia-induced pulmonary fibrosis needs to be further clarified. The aim of this study was to explore the effect and underlying mechanism of angiotensin II (Ang II on collagen synthesis in hypoxic human lung fibroblast (HLF cells. The HLF-1 cell line was used for in vitro studies. Angiotensinogen (AGT, angiotensin converting enzyme (ACE, angiotensin II type 1 receptor (AT1R and angiotensin II type 2 receptor (AT2R expression levels in human lung fibroblasts were analysed using real-time polymerase chain reaction (RT-PCR after hypoxic treatment. Additionally, the collagen type I (Col-I, AT1R and nuclear factor κappaB (NF-κB protein expression levels were detected using Western blot analysis, and NF-κB nuclear translocation was measured using immunofluorescence localization analysis. Ang II levels in HLF-1 cells were measured with an enzyme-linked immunosorbent assay (ELISA. We found that hypoxia increased Col-I mRNA and protein expression in HLF-1 cells, and this effect could be inhibited by an AT1R or AT2R inhibitor. The levels of NF-κB, RAS components and Ang II production in HLF-1 cells were significantly increased after the hypoxia exposure. Hypoxia or Ang II increased NF-κB-p50 protein expression in HLF-1 cells, and the special effect could be inhibited by telmisartan (TST, an AT1R inhibitor, and partially inhibited by PD123319, an AT2R inhibitor. Importantly, hypoxia-induced NF-κB nuclear translocation could be nearly completely inhibited by an AT1R or AT2R inhibitor. Furthermore pyrrolidine dithiocarbamate (PDTC, a NF-κB blocker, abolished the expression of hypoxia-induced AT1R and Col-I in HLF-1 cells. Our results indicate that Ang II-mediated NF-κB signalling via ATR is involved in hypoxia-induced collagen synthesis in human lung fibroblasts.

  13. Local Helioseismology

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    Gizon Laurent

    2005-11-01

    Full Text Available We review the current status of local helioseismology, covering both theoretical and observational results. After a brief introduction to solar oscillations and wave propagation through inhomogeneous media, we describe the main techniques of local helioseismology: Fourier-Hankel decomposition, ring-diagram analysis, time-distance helioseismology, helioseismic holography, and direct modeling. We discuss local helioseismology of large-scale flows, the solar-cycle dependence of these flows, perturbations associated with regions of magnetic activity, and solar supergranulation.

  14. Pluripotent State Induction in Mouse Embryonic Fibroblast Using mRNAs of Reprogramming Factors

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    Ahmed Kamel El-Sayed

    2014-11-01

    Full Text Available Reprogramming of somatic cells has great potential to provide therapeutic treatments for a number of diseases as well as provide insight into mechanisms underlying early embryonic development. Improvement of induced Pluripotent Stem Cells (iPSCs generation through mRNA-based methods is currently an area of intense research. This approach provides a number of advantages over previously used methods such as DNA integration and insertional mutagenesis. Using transfection of specifically synthesized mRNAs of various pluripotency factors, we generated iPSCs from mouse embryonic fibroblast (MEF cells. The genetic, epigenetic and functional properties of the iPSCs were evaluated at different times during the reprogramming process. We successfully introduced synthesized mRNAs, which localized correctly inside the cells and exhibited efficient and stable translation into proteins. Our work demonstrated a robust up-regulation and a gradual promoter de-methylation of the pluripotency markers, including non-transfected factors such as Nanog, SSEA-1 (stage-specific embryonic antigen 1 and Rex-1 (ZFP-42, zinc finger protein 42. Using embryonic stem cells (ESCs conditions to culture the iPS cells resulted in formation of ES-like colonies after approximately 12 days with only five daily repeated transfections. The colonies were positive for alkaline phosphatase and pluripotency-specific markers associated with ESCs. This study revealed the ability of pluripotency induction and generation of mouse mRNA induced pluripotent stem cells (mRNA iPSCs using transfection of specifically synthesized mRNAs of various pluripotency factors into mouse embryonic fibroblast (MEF cells. These generated iPSCs exhibited molecular and functional properties similar to ESCs, which indicate that this method is an efficient and viable alternative to ESCs and can be used for further biological, developmental and therapeutic investigations.

  15. Growth properties and growth factor responsiveness in skin fibroblasts from centenarians.

    Science.gov (United States)

    Tesco, G; Vergelli, M; Grassilli, E; Salomoni, P; Bellesia, E; Sikora, E; Radziszewska, E; Barbieri, D; Latorraca, S; Fagiolo, U; Santacaterina, S; Amaducci, L; Tiozzo, R; Franceschi, C; Sorbi, S

    1998-03-27

    Human fibroblast cultures, which have a finite replicative lifespan in vitro, are the most widely used model for the study of senescence at the cellular level. An inverse relationship between replicative capability and donor age has been reported in human fibroblast strains. We studied the growth capacity of fibroblast primary cultures derived from people whose lifespan was as closer as possible to the expected maximum human lifespan, i.e. people over one hundred. Our data suggest that outgrowth of fibroblasts from biopsies, growth kinetics at different population doubling levels, capability to respond to a classical mitogenic stimulus (such as 20% serum) and a variety of growth factors, were remarkably similar in fibroblasts from centenarians and young controls. On the whole, our data challenge the tenet of a simple and strict relationship between in vivo aging and in vitro proliferative capability of human fibroblasts, at least at the individual level.

  16. Presence of arylsulfatase A (ARS A) in multiple sulfatase deficiency disorder fibroblasts.

    Science.gov (United States)

    Fluharty, A L; Stevens, R L; Davis, L L; Shapiro, L J; Kihara, H

    1978-05-01

    Multiple deficiency disorder fibroblasts cultured in MEM-CO2 showed deficiencies of arylsulfatase A(ARS A) comparable to the deficiency in metachromatic leukodystrophy fibroblasts. However, the MSDD fibroblasts cultured in MEM-HEPES contained near normal levels of ARS A. Moreover, the enzyme from the latter fibroblasts was indistinguishable from ARS A of control fibroblasts on DEAE-cellulose chromatography, ratio of activity with several substrates, thermal inactivation, sensitivity to inhibitors, and precipitation by antiserum to human ARS A. These data support the conclusion that the ARS A genome is intact in MSDD fibroblasts and, by extension, in MSDD patients. Other sulfatases were present at levels ranging from mildly deficient to near normal but never as low as seen in the corresponding specific sulfatase deficient disorders.

  17. Chromatin-modifying agents convert fibroblasts to OCT4+ and VEGFR-2+ capillary tube-forming cells.

    Science.gov (United States)

    Wary, Anita; Wary, Neil; Baruah, Jugajyoti; Mastej, Victoria; Wary, Kishore K

    2017-01-01

    The human epigenome is plastic. The goal of this study was to address if fibroblast cells can be epigenetically modified to promote neovessel formation. Here, we used highly abundant human adult dermal fibroblast cells (hADFCs) that were treated with the chromatin-modifying agents 5-aza-2'-deoxycytidine and trichostatin A, and subsequently subjected to differentiation by activating Wnt signaling. Our results show that these epigenetically modified hADFCs increasingly expressed β-catenin, pluripotency factor octamer-binding transcription factor-4 (OCT4, also known as POU5F1), and endothelial cell (EC) marker called vascular endothelial growth factor receptor-2 (VEGFR-2, also known as Fetal Liver Kinase-1). In microscopic analysis, β-catenin localized to cell-cell contact points, while OCT4 was found to be localized primarily to the nucleus of these cells. Furthermore, in a chromatin immunoprecipitation experiment, OCT4 bound to the VEGFR-2/FLK1 promoter. Finally, these modified hADFCs also transduced Wnt signaling. Importantly, on a two-dimensional (2D) gel substrate, a subset of the converted cells formed vascular network-like structures in the presence of VEGF. Chromatin-modifying agents converted hADFCs to OCT4+ and VEGFR-2+ capillary tube-forming cells in a 2D matrix in VEGF-dependent manner.

  18. Transforming growth factor type-β inhibits Mas receptor expression in fibroblasts but not in myoblasts or differentiated myotubes; Relevance to fibrosis associated to muscular dystrophies.

    Science.gov (United States)

    Cofre, Catalina; Acuña, María José; Contreras, Osvaldo; Morales, María Gabriela; Riquelme, Cecilia; Cabello-Verrugio, Claudio; Brandan, Enrique

    2015-01-01

    Duchenne muscular dystrophy is a genetic disorder characterized by myofiber degeneration, muscle weakness, and increased fibrosis. Transforming growth factor type-β (TGF-β), a central mediator of fibrosis, is upregulated in fibrotic diseases. Angiotensin-(1-7) [Ang-(1-7)] is a peptide with actions that oppose those of angiotensin-II (Ang II). Ang-(1-7) effects are mediated by the Mas receptor. Treatment with Ang-(1-7) produce positive effects in the mdx mouse, normalizing skeletal muscle architecture, decreasing local fibrosis, and fibroblasts, and improving muscle function. Mdx mice deficient for the Mas receptor showed the opposite effects. To identify the cell type(s) responsible for Mas receptor expression, and to characterize whether profibrotic effectors had any effect on its expression, we determined the effect of profibrotic agents on Mas expression. TGF-β, but not connective tissue growth factor or Ang-II, reduced the expression of Mas receptor in fibroblasts isolated from skeletal muscle cells and fibroblasts from two established cell lines. In contrast, no effects were observed in myoblasts and differentiated myotubes. This inhibition was mediated by the Smad-dependent (canonical) and the PI3K and MEK1/2 (noncanonical) TGF-β signaling pathways. When both canonical and noncanonical inhibitors of the TGF-β-dependent pathways were added together, the inhibitory effect of TGF-β on Mas expression was lost. The decrease in Mas receptor induced by TGF-β in fibroblasts reduced the Ang-(1-7) mediated stimulation of phosphorylation of AKT pathway proteins. These results suggest that reduction of Mas receptor in fibroblasts, by TGF-β, could increase the fibrotic phenotype observed in dystrophic skeletal muscle decreasing the beneficial effect of Ang-(1-7). © 2015 International Union of Biochemistry and Molecular Biology.

  19. Cathepsin B Regulates Collagen Expression by Fibroblasts via Prolonging TLR2/NF-?B Activation

    OpenAIRE

    Xue Li; Zhou Wu; Junjun Ni; Yicong Liu; Jie Meng; Weixian Yu; Hiroshi Nakanishi; Yanmin Zhou

    2016-01-01

    Fibroblasts are essential for tissue repair due to producing collagens, and lysosomal proteinase cathepsin B (CatB) is involved in promoting chronic inflammation. We herein report that CatB regulates the expression of collagens III and IV by fibroblasts in response to a TLR2 agonist, lipopolysaccharide from Porphyromonas gingivalis (P.g. LPS). In cultured human BJ fibroblasts, mRNA expression of CatB was significantly increased, while that of collagens III and IV was significantly decreased a...

  20. Allogeneic human dermal fibroblasts are viable in peripheral blood mononuclear co-culture

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    Restu Syamsul Hadi

    2014-08-01

    Full Text Available Background Transplanted allogeneic dermal fibroblasts retain stem cell subpopulations, and are easily isolated, expanded and stored using standard techniques. Their potential for regenerative therapy of chronic wounds should be evaluated. The aim of this study was to determine allogeneic fibroblast viability in the presence of peripheral blood mononuclear cells (PBMC. Methods In this experimental study, fibroblasts were isolated from foreskin explants, expanded in the presence of serum, and stored using slow-freezing. We used one intervention group of allogeneic fibroblasts co-cultured with PBMC and 2 control groups of separate fibroblast and PBMC cultures.Fibroblasts were characterized by their collagen secretion and octamer-binding transcription factor 4 (OCT4 expression. Viability was evaluated using water soluble tetrazolium-1 (WST-1 proliferation assay. Absorbances were measured at 450 nm. Data analysis was performed by student’s paired t-test. Results Dermal fibroblasts were shown to secrete collagen, express OCT4, be recoverable after cryopreservation, and become attached to the culture dish in a co-culture with PBMC. Co-cultured and control fibroblasts had no significantly different cell viabilities (p>0.05. Calculated viable cell numbers increased 1.8 and 5.1-fold, respectively, at days 2 and 4 in vitro. Both groups showed comparable doubling times at days 2 and 4 in vitro. PBMC did not interfere with allogeneic fibroblast viability and proliferative capacity Conclusions Allogeneic fibroblasts remain viable and proliferate in the presence of host PBMC. Future research should evaluate allogeneic human dermal fibroblast competency in clinical settings. Dermal fibroblasts are a potential source for cell therapy in chronic wound management.

  1. Allogeneic human dermal fibroblasts are viable in peripheral blood mononuclear co-culture

    Directory of Open Access Journals (Sweden)

    Restu Syamsul Hadi

    2015-12-01

    Full Text Available BACKGROUND Transplanted allogeneic dermal fibroblasts retain stem cell subpopulations, and are easily isolated, expanded and stored using standard techniques. Their potential for regenerative therapy of chronic wounds should be evaluated. The aim of this study was to determine allogeneic fibroblast viability in the presence of peripheral blood mononuclear cells (PBMC. METHODS In this experimental study, fibroblasts were isolated from foreskin explants, expanded in the presence of serum, and stored using slow-freezing. We used one intervention group of allogeneic fibroblasts co-cultured with PBMC and 2 control groups of separate fibroblast and PBMC cultures.Fibroblasts were characterized by their collagen secretion and octamer-binding transcription factor 4 (OCT4 expression. Viability was evaluated using water soluble tetrazolium-1 (WST-1 proliferation assay. Absorbances were measured at 450 nm. Data analysis was performed by student’s paired t-test. RESULTS Dermal fibroblasts were shown to secrete collagen, express OCT4, be recoverable after cryopreservation, and become attached to the culture dish in a co-culture with PBMC. Co-cultured and control fibroblasts had no significantly different cell viabilities (p>0.05. Calculated viable cell numbers increased 1.8 and 5.1- fold, respectively, at days 2 and 4 in vitro. Both groups showed comparable doubling times at days 2 and 4 in vitro. PBMC did not interfere with allogeneic fibroblast viability and proliferative capacity CONCLUSIONS Allogeneic fibroblasts remain viable and proliferate in the presence of host PBMC. Future research should evaluate allogeneic human dermal fibroblast competency in clinical settings. Dermal fibroblasts are a potential source for cell therapy in chronic wound management.

  2. Lidocaine Impairs Proliferative and Biosynthetic Functions of Aged Human Dermal Fibroblasts.

    Science.gov (United States)

    Bentov, Itay; Damodarasamy, Mamatha; Spiekerman, Charles; Reed, May J

    2016-09-01

    The aged are at increased risk of postoperative wound healing complications. Because local anesthetics are infiltrated commonly into the dermis of surgical wounds, we sought to determine whether local anesthetics adversely affect proliferative and biosynthetic functions of dermal fibroblasts. We also evaluated the effect of local anesthetics on insulin-like growth factor-1 (IGF-1) and transforming growth factor-β1 (TGF-β1), growth factors that are important regulators of wound healing. Human dermal fibroblasts (HFB) from aged and young donors were exposed to local anesthetic agents at clinically relevant concentrations. We screened the effects of lidocaine, bupivacaine, mepivacaine, and ropivacaine on proliferation of HFB. Lidocaine was most detrimental to proliferation in HFB. We then evaluated the effect of lidocaine on expression and function of the growth factors, IGF-1 and TGF-β1. Lastly, concurrent exposure to lidocaine and IGF-1 or TGF-β1 was evaluated for their effects on proliferation and expression of dermal collagens, respectively. Lidocaine and mepivacaine inhibited proliferation in aged HFB (for lidocaine 88% of control, 95% confidence interval [CI], 80%-98%, P = .009 and for mepivacaine 90% of control, 95% CI, 81%-99%, P = .032) but not in young HFB. Ropivacaine and bupivacaine did not inhibit proliferation. Because of the clinical utility of lidocaine relative to mepivacaine, we focused on lidocaine. Lidocaine decreased proliferation in aged HFB, which was abrogated by IGF-1. Lidocaine inhibited transcripts for IGF-1 and insulin-like growth factor-1 receptor (IGF1R) in fibroblasts from aged donors (IGF-1, log2 fold-change -1.25 [42% of control, 95% CI, 19%-92%, P = .035] and IGF1R, log2 fold-change -1.00 [50% of control, 95% CI, 31%-81%, P = .014]). In contrast, lidocaine did not affect the expression of IGF-1 or IGF1R transcripts in the young HFB. Transcripts for collagen III were decreased after lidocaine exposure in aged and young HFB (log2

  3. Localized Scleroderma

    Science.gov (United States)

    ... scleroderma, or who are experts at examining the skin, can arrive at the diagnosis without much difficulty after a careful examination. In ... especially in patients who continue to have active skin problems or develop new ... Diagnosis of Localized Scleroderma? The diagnosis of localized scleroderma ...

  4. Net Locality

    DEFF Research Database (Denmark)

    de Souza e Silva, Adriana Araujo; Gordon, Eric

    Provides an introduction to the new theory of Net Locality and the profound effect on individuals and societies when everything is located or locatable. Describes net locality as an emerging form of location awareness central to all aspects of digital media, from mobile phones, to Google Maps...... of emerging technologies, from GeoCities to GPS, Wi-Fi, Wiki Me, and Google Android....

  5. Platelets stimulate fibroblast-mediated contraction of collagen gels

    Directory of Open Access Journals (Sweden)

    Lundahl Joachim

    2003-10-01

    Full Text Available Abstract Background Platelets are thought to play a role in a variety of inflammatory conditions in the lung, some of which may lead to fibrosis. In the current study we tested the hypothesis that whole platelets and platelet lysate can mediate remodelling of extracellular matrix in vitro by affecting fibroblast-mediated contraction of a collagen gel. We also sought to determine to what extent platelet-derived growth factor (PDGF and transforming growth factor-β (TGF-β contribute to this effect. Methods Washed platelets, isolated from healthy blood donors, and platelet lysate (freezing and thawing, were cast together with human lung fibroblasts in three-dimensional collagen gels. The gels were then released and cultured for four days. PDGF and TGF-β1 concentrations were measured in culture supernatants by ELISA. Results Both platelets and platelet lysate augmented fibroblast-mediated gel contraction in a time and concentration dependent manner (19.9% ± 0.1 (mean ± SEM of initial area vs. 48.0% ± 0.4 at 48 hours; P 1 and PDGF-AA/AB were released in co-culture. PDGF-AA/AB had a maximum release at 24 hours whereas TGF-β1 release increased with longer culture periods. Neutralising antibodies to these mediators partially inhibited platelet-induced gel contraction. Conclusion We conclude that platelets may promote remodelling of extracellular matrix in vitro and that PDGF and TGF-β partially mediate this effect, also indicating a role for other mediators. The findings may be an important mechanism in regulating repair processes after injury.

  6. Reprogramming mouse embryo fibroblasts to functional islets without genetic manipulation.

    Science.gov (United States)

    Chandravanshi, Bhawna; Bhonde, Ramesh

    2018-02-01

    The constant quest for generation of large number of islets aimed us to explore the differentiation potential of mouse embryo fibroblast cells. Mouse embryo fibroblast cells isolated from 12- to 14-day-old pregnant mice were characterized for their surface markers and tri-lineage differentiation potential. They were subjected to serum-free media containing a cocktail of islet differentiating reagents and analyzed for the expression of pancreatic lineage transcripts. The islet-like cell aggregates (ICAs) was confirmed for their pancreatic properties via immunofluorecence for C-peptide, glucagon, and somatostain. They were positive for CD markers-Sca1, CD44, CD73, and CD90 and negative for hematopoietic markers-CD34 and CD45 at both transcription and translational levels. The transcriptional analysis of the ICAs at different day points exhibited up-regulation of islet markers (Insulin, PDX1, HNF3, Glucagon, and Somatostatin) and down-regulation of MSC-markers (Vimentin and Nestin). They positively stained for dithizone, C-peptide, insulin, glucagon, and somatostatin indicating intact insulin producing machinery. In vitro glucose stimulation assay revealed three-fold increase in insulin secretion as compared to basal glucose with insulin content being the same in both the conditions. The preliminary in vivo data on ICA transplantation showed reversal of diabetes in streptozotocin induced diabetic mice. Our results demonstrate for the first time that mouse embryo fibroblast cells contain a population of MSC-like cells which could differentiate into insulin producing cell aggregates. Hence, our study could be extrapolated for isolation of MSC-like cells from human, medically terminated pregnancies to generate ICAs for treating type 1 diabetic patients. © 2017 Wiley Periodicals, Inc.

  7. The synthetic pathway for glucosylsphingosine in cultured fibroblasts.

    Science.gov (United States)

    Yamaguchi, Y; Sasagasako, N; Goto, I; Kobayashi, T

    1994-09-01

    The synthesis of glucosylsphingosine (GlcSph), a glucosylceramide (GlcCer) analogue devoid of fatty acids, in cultured fibroblasts was studied by using conduritol beta epoxide (CBE), an inhibitor of beta-glucosidase, and 1-phenyl-2-decanoylamino-3-morpholino-1-propanol (PDMP), an inhibitor of glucosylceramide (GlcCer) synthase (glucosyltransferase). When CBE was added to the culture medium, the intracellular beta-glucosidase activity decreased, and both GlcCer and GlcSph accumulated in the cells. After the addition of PDMP, the concentration of GlcCer decreased, while the content of GlcSph increased. When CBE and PDMP were added together, the intracellular accumulation of GlcSph to decreased to less than when CBE alone was added. Based on these results, the synthetic pathway for GlcSph was thus considered to not only be through the glucosylation of sphingosine, but also through the deacylation of GlcCer. When GlcCer (d18:1, C12:0) was added to the culture medium, the intracellular accumulation of GlcSph (d18:1) was evident, and it was also more pronounced in the presence of CBE. In addition, when GlcCer (d18:0, C12:0) was used, apparent accumulation of GlcSph (d18:0) was also observed. In order to determine whether or not the deacylase of GlcCer is identical to acid ceramidase, a deacylase of ceramide, the same experiments were carried out using fibroblasts from two patients with Farber disease, in which acid ceramidase is genetically deficient. The accumulation of GlcSph in the Farber disease fibroblasts after the loading of GlcCer for 7 days was found to be one-fifth of the control level.(ABSTRACT TRUNCATED AT 250 WORDS)

  8. EPAC expression and function in cardiac fibroblasts and myofibroblasts

    Energy Technology Data Exchange (ETDEWEB)

    Olmedo, Ivonne; Muñoz, Claudia; Guzmán, Nancy; Catalán, Mabel; Vivar, Raúl; Ayala, Pedro; Humeres, Claudio; Aránguiz, Pablo [Departamento de Química Farmacológica y Toxicológica, Facultad de Ciencias Químicas y Farmacéuticas, Universidad de Chile (Chile); García, Lorena [Departamento de Bioquímica y Biología Molecular, Facultad de Ciencias Químicas y Farmacéuticas, Universidad de Chile (Chile); Velarde, Victoria [Departamento de Ciencias Fisiológicas, Facultad de Ciencias Biológicas, Pontificia Universidad Católica de Chile (Chile); Díaz-Araya, Guillermo, E-mail: gadiaz@ciq.uchile.cl [Departamento de Química Farmacológica y Toxicológica, Facultad de Ciencias Químicas y Farmacéuticas, Universidad de Chile (Chile)

    2013-10-15

    In the heart, cardiac fibroblasts (CF) and cardiac myofibroblasts (CMF) are the main cells responsible for wound healing after cardiac insult. Exchange protein activated by cAMP (EPAC) is a downstream effector of cAMP, and it has been not completely studied on CF. Moreover, in CMF, which are the main cells responsible for cardiac healing, EPAC expression and function are unknown. We evaluated in both CF and CMF the effect of transforming growth factor β1 (TGF-β1) on EPAC-1 expression. We also studied the EPAC involvement on collagen synthesis, adhesion, migration and collagen gel contraction. Method: Rat neonatal CF and CMF were treated with TGF-β1 at different times and concentrations. EPAC-1 protein levels and Rap1 activation were measured by western blot and pull down assay respectively. EPAC cellular functions were determined by adhesion, migration and collagen gel contraction assay; and collagen expression was determined by western blot. Results: TGF-β1 through Smad and JNK significantly reduced EPAC-1 expression in CF, while in CMF this cytokine increased EPAC-1 expression through ERK1/2, JNK, p38, AKT and Smad3. EPAC activation was able to induce higher Rap1-GTP levels in CMF than in CF. EPAC and PKA, both cAMP effectors, promoted CF and CMF adhesion on fibronectin, as well as CF migration; however, this effect was not observed in CMF. EPAC but not PKA activation mediated collagen gel contraction in CF, while in CMF both PKA and EPAC mediated collagen gel contraction. Finally, the EPAC and PKA activation reduced collagen synthesis in CF and CMF. Conclusion: TGF-β1 differentially regulates the expression of EPAC in CF and CMF; and EPAC regulates differentially CF and CMF functions associated with cardiac remodeling. - Highlights: • TGF-β1 regulates EPAC-1 expression in cardiac fibroblast and myofibroblast. • Rap-1GTP levels are higher in cardiac myofibroblast than fibroblast. • EPAC-1 controls adhesion, migration and collagen synthesis in cardiac

  9. Quercetin inhibits hexose transport in a human diploid fibroblast

    Energy Technology Data Exchange (ETDEWEB)

    Salter, D.W.; Custead-Jones, S.; Cook, J.S.

    1978-01-01

    The flavonol quercetin, a phloretin analog, inhibits transport of 2-deoxyglucose and 3-O-methylglucose in a cultured human diploid fibroblast. This inhibition is related to transport itself and not to the reported effects of flavonoids on membrane-bound ATPases. From concentration-inhibition curves at several pH's we conclude that uncharged (acid) quercetin (pH = 7.65) is the inhibitory form of the molecule (K/sub I/ = 10 ..mu..m). Quercetin, unlike phloretin, is rapidly degraded in 0.1 N NaOH; the degradation products are weakly inhibitory to hexose transport.

  10. Cancer Associated Fibroblasts and Tumor Growth: Focus on Multiple Myeloma

    Energy Technology Data Exchange (ETDEWEB)

    De Veirman, Kim, E-mail: kdeveirm@vub.ac.be [Department of Hematology and Immunology, Myeloma Center Brussels, Vrije Universiteit Brussel (VUB), Brussels 1090 (Belgium); Rao, Luigia [Department of Hematology and Immunology, Myeloma Center Brussels, Vrije Universiteit Brussel (VUB), Brussels 1090 (Belgium); Department of Biomedical Sciences and Human Oncology, Section of Internal Medicine, University of Bari Medical School, Bari I-70124 (Italy); De Bruyne, Elke; Menu, Eline; Van Valckenborgh, Els [Department of Hematology and Immunology, Myeloma Center Brussels, Vrije Universiteit Brussel (VUB), Brussels 1090 (Belgium); Van Riet, Ivan [Department of Hematology and Immunology, Myeloma Center Brussels, Vrije Universiteit Brussel (VUB), Brussels 1090 (Belgium); Stem Cell Laboratory, Division of Clinical Hematology, Universitair Ziekenhuis Brussel (UZ Brussel), Brussels 1090 (Belgium); Frassanito, Maria Antonia [Department of Biomedical Sciences and Human Oncology, Section of General Pathology, University of Bari Medical School, Bari I-70124 (Italy); Di Marzo, Lucia; Vacca, Angelo [Department of Biomedical Sciences and Human Oncology, Section of Internal Medicine, University of Bari Medical School, Bari I-70124 (Italy); Vanderkerken, Karin, E-mail: kdeveirm@vub.ac.be [Department of Hematology and Immunology, Myeloma Center Brussels, Vrije Universiteit Brussel (VUB), Brussels 1090 (Belgium)

    2014-06-27

    Cancer associated fibroblasts (CAFs) comprise a heterogeneous population that resides within the tumor microenvironment. They actively participate in tumor growth and metastasis by production of cytokines and chemokines, and the release of pro-inflammatory and pro-angiogenic factors, creating a more supportive microenvironment. The aim of the current review is to summarize the origin and characteristics of CAFs, and to describe the role of CAFs in tumor progression and metastasis. Furthermore, we focus on the presence of CAFs in hypoxic conditions in relation to multiple myeloma disease.

  11. Synthesis of multilayered alginate microcapsules for the sustained release of fibroblast growth factor-1

    Science.gov (United States)

    Khanna, Omaditya; Moya, Monica L; Opara, Emmanuel C; Brey, Eric M

    2010-01-01

    Alginate microcapsules coated with a permselective poly-L-ornithine (PLO) membrane have been investigated for the encapsulation and transplantation of islets as a treatment for type 1 diabetes. The therapeutic potential of this approach could be improved through local stimulation of microvascular networks in order to meet mass transport demands of the encapsulated cells. Fibroblast growth factor-1 (FGF-1) is a potent angiogenic factor with optimal effect occurring when it is delivered in a sustained manner. In this paper, a technique is described for the generation of multilayered alginate microcapsules with an outer alginate layer that can be used for the delivery of FGF-1. The influence of alginate concentration and composition (high mannuronic acid (M) or guluronic acid (G) content) on outer layer size and stability, protein encapsulation efficiency, and release kinetics was investigated. The technique results in a stable outer layer of alginate with a mean thickness between 113–164 µm, increasing with alginate concentration and G-content. The outer layer was able to encapsulate and release FGF-1 for up to thirty days, with 1.25% of high G alginate displaying the most sustained release. The released FGF-1 retained its biologic activity in the presence of heparin, and the addition of the outer layer did not alter the permselectivity of the PLO coat. This technique could be used to generate encapsulation systems that deliver proteins to stimulate local neovascularization around encapsulated islets. PMID:20725969

  12. Cell Toxicity in Fibroblasts, Tenocytes, and Human Mesenchymal Stem Cells-A Comparison of Necrosis and Apoptosis-Inducing Ability in Ropivacaine, Bupivacaine, and Triamcinolone.

    Science.gov (United States)

    Zhang, Anja Z; Ficklscherer, Andreas; Gülecyüz, Mehmet F; Paulus, Alexander C; Niethammer, Thomas R; Jansson, Volkmar; Müller, Peter E

    2017-04-01

    To analyze the ability of ropivacaine, bupivacaine, and triamcinolone to induce apoptosis and necrosis in fibroblasts, tenocytes, and human mesenchymal stem cells. Human dermal fibroblasts, adipose-derived human mesenchymal stem cells (hMSCs), and tenocytes gained from the rotator cuff tendon were seeded with a cell density of 0.5 × 10(4)/cm(2). One specimen of ropivacaine, bupivacaine, and triamcinolone was tested separately on the cells with separate concentrations of 0.5%, 0.25%, and 0.125% for each specimen. The negative control received no agent, only a change of medium. The incubation period for each agent was 30 minutes. After a change of medium and 1 hour, 24 hours, and 7 days of incubation, 10(4) cells were harvested and analyzed via fluorescence-activated cell sorting with double-staining with annexin V and propidium iodide. Statistical analysis to determine significant difference (P necrosis-inducing effects on fibroblasts and tenocytes, with the necrotic effect peaking at 0.5% and 0.25%. Ropivacaine and triamcinolone caused no significant necrosis. Compared with fibroblasts and tenocytes, hMSCs did not show significant necrotic or apoptotic effects after exposure to bupivacaine. Overall, no significant differences in apoptosis were detected between different cell lines, varying concentrations, or time measurements. Bupivacaine 0.5% and 0.25% have the most necrosis-inducing effects on fibroblasts and tenocytes. Ropivacaine caused less necrosis than bupivaine. Compared with fibroblasts and tenocytes, hMSCs were not affected by necrosis using any of the tested agents. A significant apoptosis-inducing effect could not be detected for the different cell lines. Possible cell toxicity raises questions of concern for intra-articular injections using local anesthetics and corticosteroids. The present study demonstrates the necrotic and apoptotic effects of ropivacaine, bupivacaine, and triamcinolone and may give recommendations for intra-articular use of

  13. Ganglioside GM3 sialidase activity in fibroblasts of normal individuals and of patients with sialidosis and mucolipidosis IV. Subcellular distribution and and some properties.

    Science.gov (United States)

    Lieser, M; Harms, E; Kern, H; Bach, G; Cantz, M

    1989-01-01

    Sensitive assays for the determination of the ganglioside sialidase activity of fibroblast homogenates were established using ganglioside GM3, 3H-labelled in the sphingosine moiety, as a substrate. Ganglioside GM3 sialidase activity was greatly stimulated by the presence of the non-ionic detergent Triton X-100 and was further enhanced by salts such as NaCl; the optimal pH was 4.5. The subcellular localization of this activity was determined by fractionation using free-flow electrophoresis and found to be exclusively associated with the marker for the plasma membrane, but not with that for lysosomes. This Triton-stimulated ganglioside sialidase activity was selectively inhibited by preincubating intact cells in the presence of millimolar concentrations of Cu2+, suggesting that the activity resides on the external surface of the plasma membrane. In normal fibroblasts homogenates, ganglioside GM3 sialidase was also greatly stimulated by sodium cholate. In contrast to the Triton X-100-activated reaction, however, it was not diminished by prior incubation of intact cells in the presence of Cu2+. Only after cell lysis was Cu2+ inhibitory. the cholate-stimulated ganglioside sialidase activity thus paralleled the behaviour of the lysosomal 4-methylumbelliferyl-alpha-D-N-acetylneuraminic acid (4-MU-NeuAc) sialidase. In fibroblasts from sialidosis patients, the cholate-stimulated ganglioside GM3 sialidase activity, but not that of the Triton-activated enzyme, was profoundly diminished. In fibroblasts from patients with mucolipidosis IV (ML IV), both the Triton X-100- and the cholate-stimulated ganglioside GM3 sialidase activities were in the range of normal controls. The Triton-activated enzyme was associated with the plasma membrane in the same manner as in normal cells. Our findings suggest that, in human fibroblasts, there exist two sialidases that degrade ganglioside GM3: one on the external surface of the plasma membrane, and another that is localized in lysosomes and

  14. Fibroblast growth factor 2 (FGF2) is present in human spermatozoa and is related with sperm motility. The use of recombinant FGF2 to improve motile sperm recovery.

    Science.gov (United States)

    Garbarino Azúa, D J; Saucedo, L; Giordana, S; Magri, M L; Buffone, M G; Neuspiller, F; Vazquez-Levin, M H; Marín-Briggiler, C I

    2017-09-01

    Fibroblast growth factors (FGFs) and their receptors (FGFRs) regulate several functions of somatic cells. In a previous work, we reported FGFR expression in human spermatozoa and their involvement in motility. This study aimed to evaluate the presence and localization of fibroblast growth factor 2 (FGF2) in human spermatozoa, to determine the relationship of FGF2 levels with conventional semen parameters and to assess the effect of recombinant FGF2 (rFGF2) on sperm recovery in a selection procedure. Western immunoblotting analysis using an antibody against FGF2 revealed an 18-kDa band in sperm protein extracts. The protein was immunolocalized in the sperm flagellum and acrosomal region, as well as in all germ cells. Sperm FGF2 levels, assessed by flow cytometry, showed a positive (p recoveries, and increased (p recovery in selection techniques. © 2017 American Society of Andrology and European Academy of Andrology.

  15. Fibroblast growth factor receptor 4 regulates tumor invasion by coupling fibroblast growth factor signaling to extracellular matrix degradation

    DEFF Research Database (Denmark)

    Sugiyama, Nami; Varjosalo, Markku; Meller, Pipsa

    2010-01-01

    Aberrant expression and polymorphism of fibroblast growth factor receptor 4 (FGFR4) has been linked to tumor progression and anticancer drug resistance. We describe here a novel mechanism of tumor progression by matrix degradation involving epithelial-to-mesenchymal transition in response...... substantiated by reduced MT1-MMP content and in vivo growth of prostate carcinoma cells after the FGFR4-R388 gene silencing. In contrast, knockdown of the alternative FGFR4-G388 allele enhanced MT1-MMP and invasive tumor cell growth in vivo and within three-dimensional collagen. These results will help...

  16. Encapsulation of fibroblasts causes accelerated alginate hydrogel degradation.

    Science.gov (United States)

    Hunt, N C; Smith, A M; Gbureck, U; Shelton, R M; Grover, L M

    2010-09-01

    Calcium-alginate hydrogel has been widely studied as a material for cell encapsulation for tissue engineering. At present, the effect that cells have on the degradation of alginate hydrogel is largely unknown. We have shown that fibroblasts encapsulated at a density of 7.5 x 10(5) cells ml(-1) in both 2% and 5% w/v alginate remain viable for at least 60 days. Rheological analysis was used to study how the mechanical properties exhibited by alginate hydrogel changed during 28 days in vitro culture. Alginate degradation was shown to occur throughout the study but was greatest within the first 7 days of culture for all samples, which correlated with a sharp release of calcium ions from the construct. Fibroblasts were shown to increase the rate of degradation during the first 7 days when compared with acellular samples in both 2% and 5% w/v gels, but after 28 days both acellular and cell-encapsulating samples retained disc-shaped morphologies and gel-like spectra. The results demonstrate that although at an early stage cells influence the mechanical properties of encapsulating alginate, over a longer period of culture, the hydrogels retain sufficient mechanical integrity to exhibit gel-like properties. This allows sustained immobilization of the cells at the desired location in vivo where they can produce extracellular matrix and growth factors to expedite the healing process. 2010 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

  17. An Expandable, Inducible Hemangioblast State Regulated by Fibroblast Growth Factor

    Directory of Open Access Journals (Sweden)

    David T. Vereide

    2014-12-01

    Full Text Available During development, the hematopoietic and vascular lineages are thought to descend from common mesodermal progenitors called hemangioblasts. Here we identify six transcription factors, Gata2, Lmo2, Mycn, Pitx2, Sox17, and Tal1, that “trap” murine cells in a proliferative state and endow them with a hemangioblast potential. These “expandable” hemangioblasts (eHBs are capable, once released from the control of the ectopic factors, to give rise to functional endothelial cells, multilineage hematopoietic cells, and smooth muscle cells. The eHBs can be derived from embryonic stem cells, from fetal liver cells, or poorly from fibroblasts. The eHBs reveal a central role for fibroblast growth factor, which not only promotes their expansion, but also facilitates their ability to give rise to endothelial cells and leukocytes, but not erythrocytes. This study serves as a demonstration that ephemeral progenitor states can be harnessed in vitro, enabling the creation of tractable progenitor cell lines.

  18. CEMP1 Induces Transformation in Human Gingival Fibroblasts.

    Directory of Open Access Journals (Sweden)

    Mercedes Bermúdez

    Full Text Available Cementum Protein 1 (CEMP1 is a key regulator of cementogenesis. CEMP1 promotes cell attachment, differentiation, deposition rate, composition, and morphology of hydroxyapatite crystals formed by human cementoblastic cells. Its expression is restricted to cementoblasts and progenitor cell subpopulations present in the periodontal ligament. CEMP1 transfection into non-osteogenic cells such as adult human gingival fibroblasts results in differentiation of these cells into a "mineralizing" cell phenotype. Other studies have shown evidence that CEMP1 could have a therapeutic potential for the treatment of bone defects and regeneration of other mineralized tissues. To better understand CEMP1's biological effects in vitro we investigated the consequences of its expression in human gingival fibroblasts (HGF growing in non-mineralizing media by comparing gene expression profiles. We identified several mRNAs whose expression is modified by CEMP1 induction in HGF cells. Enrichment analysis showed that several of these newly expressed genes are involved in oncogenesis. Our results suggest that CEMP1 causes the transformation of HGF and NIH3T3 cells. CEMP1 is overexpressed in cancer cell lines. We also determined that the region spanning the CEMP1 locus is commonly amplified in a variety of cancers, and finally we found significant overexpression of CEMP1 in leukemia, cervix, breast, prostate and lung cancer. Our findings suggest that CEMP1 exerts modulation of a number of cellular genes, cellular development, cellular growth, cell death, and cell cycle, and molecules associated with cancer.

  19. Capturing the Regenerative Potential of Periodontal Ligament Fibroblasts

    Directory of Open Access Journals (Sweden)

    Christina Springstead Scanlon

    2011-01-01

    Full Text Available The cell population within the periodontal ligament (PDL tissue is remarkably heterogeneous1. Fibroblasts, a mixed population of cells, are the main cellular component of the PDL and the cell type most often studied for periodontal regeneration. Osteoblasts and osteoclasts are found on the bone side, while fibroblasts, macrophages, undifferentiated adult/mesenchymal stem cells, neural elements, and endothelial cells are found throughout the PDL. Epithelial rests of Malassez cells and cementoblasts are focused near the root surface. PDL tissue also includes loose connective tissue between dense fiber bundles that contain branches of the periodontal blood vessels and nerves2. The complexity of the PDL tissue, with its various cell types and cell progenitor components, explains the challenges involved in therapies to restore tissue following periodontal disease. Cementoblasts, osteoblasts, and endothelial cells must migrate, differentiate, and coordinately interact with a variety of soluble mediators to regenerate the periodontium3. Stem cells located in the PDL tissue are key contributors to this process4. Stem cells in the PDL are important not only for formation and maintenance of the tissue but also for repair, remodeling, and regeneration of adjacent alveolar bone and cementum5. Our laboratory has shown that progenitor cells isolated from PDL tissue by selection with cell surface markers STRO-1+ and CD146+ are capable of differentiating into chondrogenic, osteogenic, and adipogenic phenotypes under appropriate culture conditions6.

  20. Cytotoxic effects of nickel nanowires in human fibroblasts

    KAUST Repository

    Felix Servin, Laura P.

    2016-03-09

    The increasing interest in the use of magnetic nanostructures for biomedical applications necessitates rigorous studies to be carried out in order to determine their potential toxicity. This work attempts to elucidate the cytotoxic effects of nickel nanowires (NWs) in human fibroblasts WI-38 by a colorimetric assay (MTT) under two different parameters: NW concentration and exposure time. This was complemented with TEM and confocal images to assess the NWs internalization and to identify any changes in the cell morphology. Ni NWs were fabricated by electrodeposition using porous alumina templates. Energy dispersive X-Ray analysis, scanning electron microscopy and transmission electron microscopy imaging were used for NW characterization. The results showed decreased cell metabolic activity for incubation times longer than 24 hours and no negative effects for exposure times shorter than that. The cytotoxicity effects for human fibroblasts were then compared with those reported for HCT 116 cells, and the findings point out that it is relevant to consider the cellular size. In addition, the present study compares the toxic effects of equivalent amounts of nickel in the form of its salt to those of NWs and shows that the NWs are more toxic than the salts. Internalized NWs were found in vesicles inside of the cells where their presence induced inflammation of the endoplasmic reticulum.

  1. Aromatase activity in human skin fibroblasts grown in cell culture.

    Science.gov (United States)

    Berkovitz, G D; Brown, T R; Fujimoto, M

    1987-01-01

    Recent studies in this laboratory have described an unusual kindred in which gynecomastia resulted from abnormally elevated levels of extraglandular aromatase activity. In order to better understand the molecular mechanisms responsible for the abnormal aromatase activity in these and other patients, we explored the aromatase activity of genital skin fibroblasts. Our studies demonstrate that the kinetic parameters for aromatase in skin are similar to those of other cultured cells and suggest that skin is an important site of extraglandular aromatase activity. These cells also contain 5 alpha-reductase activity and androgen receptors and are, therefore, a model for androgen action and metabolism. For example, they provided a system for the study of the potency and specificity of the aromatase inhibitors 4-OHA and MDL 18,962. Finally, the influence of DEX on aromatase in genital skin fibroblasts differs in some important respects from the pattern of control observed in adipose tissue stromal-vascular cells. These findings suggest that investigating the molecular mechanisms for the regulation of aromatase in skin may provide unique information about the control of the enzyme.

  2. Differential Gene Expression of Fibroblasts: Keloid versus Normal

    Directory of Open Access Journals (Sweden)

    Michael F. Angel

    2002-11-01

    Full Text Available Abstract: This study investigated gene regulation and unique gene products in both keloid (KDF and normal (NDF dermal fibroblasts in established cell lines. For gene regulation, NDF versus KDF were compared using Clontech's Atlas™ Human cDNA Expression Array while unique gene products were studied using RNA Fingerprinting Kit. RNA from each sample was converted to cDNA using oligo-dT primers. Down-regulated genes using Atlas Array in KDF were 1 60 S ribosomal protein, 2 Thioredoxin dependent peroxidase, 3 Nuclease sensitive element DNA binding protein, 4 c-myc purine-binding transcription factor, 5 c-AMP dependent protein kinase, and, 6 Heat Shock Protein 90 kDa. Genes that are up regulated in KDF were 1 Tubulin and 2 Heat Shock Protein 27 kDa. With the differential display, we found 17 bands unique to both KDF and NDF. The specific gene and the manner in which they were differentially regulated have direct implications to understanding keloid fibroblast proliferation.

  3. Proteases induce secretion of collagenase and plasminogen activator by fibroblasts

    Energy Technology Data Exchange (ETDEWEB)

    Werb, Z.; Aggeler, J.

    1978-04-01

    We have observed that treatment of rabbit synovial fibroblasts with proteolytic enzymes can induce secretion of collagenase (EC 3.4.24.7) and plasminogen activator (EC 3.4.21.-). Cells treated for 2 to 24 hr with plasmin, trypsin, chymotrypsin, pancreatic elastase, papain, bromelain, thermolysin, or ..cap alpha..-protease but not with thrombin or neuraminidase secreted detectable amounts of collagenase within 16 to 48 hr. Treatment of fibroblasts with trypsin also induced secretion of plasminogen activator. Proteases initiated secretion of collagenase (up to 20 units per 10/sup 6/ cells per 24 hr) only when treatment produced decreased cell adhesion. Collagenase production did not depend on continued presence of proteolytic activity or on subsequent cell adhesion, spreading, or proliferation. Routine subculturing with crude trypsin also induced collagenase secretion by cells. Secretion of collagenase was prevented and normal spreading was obtained if the trypsinized cells were placed into medium containing fetal calf serum. Soybean trypsin inhibitor, ..cap alpha../sub 1/-antitrypsin, bovine serum albumin, collagen, and fibronectin did not inhibit collagenase production. Although proteases that induced collagenase secretion also removed surface glycoprotein, the kinetics of induction of cell protease secretion were different from those for removal of fibronectin. Physiological inducers of secretion of collagenase and plasminogen activator by cells have not been identified. These results suggest that extracellular proteases in conjunction with plasma proteins may govern protease secretion by cells.

  4. Effect of Arctium lappa (burdock) extract on canine dermal fibroblasts.

    Science.gov (United States)

    Pomari, Elena; Stefanon, Bruno; Colitti, Monica

    2013-12-15

    Although the biological activities of Arctium lappa (burdock) have been already investigated in human and other species, data evaluating the molecular mechanisms have not been reported in the dog. In this study we analyzed for the first time the effect of a root extract of burdock on molecular responses in canine dermal fibroblasts with H2O2 stimulation (H group), with burdock treatment (B group) and with H2O2 stimulation and burdock treatment (BH group), using RNAseq technology. Differentially expressed genes (P<0.05) of H, B and BH groups in comparison to the untreated sample (negative control, C group) were identified with MeV software and were functional annotated and monitored for signaling pathways and candidate biomarkers using the Ingenuity Pathways Analysis (IPA). The expression profile of canine dermal fibroblasts treated with burdock extract with or without H2O2 stimulation, showed an up-regulation of mitochondrial superoxide dismutase (SOD2), disheveled 3 (DVL3) and chondroitin sulfate N-acetylgalactosaminyltransferase 2 (CSGALNACT2). The data suggested that burdock has implications in cell adhesion and gene expression with the modulation of Wnt/β catenin signaling and Chondroitin Sulphate Biosynthesis that are particularly important for the wound healing process. © 2013 Elsevier B.V. All rights reserved.

  5. Transdifferentiation of mouse fibroblasts and hepatocytes to functional neurons.

    Science.gov (United States)

    Marro, Samuele; Yang, Nan

    2014-01-01

    Nuclear reprogramming by defined transcription factors became of broad interest in 2006 with the work of Takahashi and Yamanaka (Cell 126:663-676, 2006), but the first example of cell fate reshaping via ectopic expression of transcription factor was provided back in 1987 when Davis and colleagues induced features of a muscle cell in fibroblast using the muscle transcription factor MyoD (Davis et al., Cell 51:987-1000, 1987). In 2010 our laboratory described how forced expression of the three neuronal transcription factors Ascl1, Brn2, and Myt1l rapidly converts mouse fibroblasts into neuronal cells that exhibit biochemical and electrophysiological properties of neurons. We named these cells induced neuronal cells (iN cells) (Vierbuchen et al., Nature 463:1035-1041, 2010; Vierbuchen and Wernig, Nat Biotechnol 29:892-907, 2011). Interestingly, iN cells can also be derived from defined endodermal cells such as primary hepatocytes, suggesting the existence of a more general reprogramming paradigm (Marro et al., Cell Stem Cell 9:374-382, 2011). In this chapter we describe the detailed methods used to attain the direct conversion.

  6. From mechanotransduction to extracellular matrix gene expression in fibroblasts.

    Science.gov (United States)

    Chiquet, Matthias; Gelman, Laurent; Lutz, Roman; Maier, Silke

    2009-05-01

    Tissue mechanics provide an important context for tissue growth, maintenance and function. On the level of organs, external mechanical forces largely influence the control of tissue homeostasis by endo- and paracrine factors. On the cellular level, it is well known that most normal cell types depend on physical interactions with their extracellular matrix in order to respond efficiently to growth factors. Fibroblasts and other adherent cells sense changes in physical parameters in their extracellular matrix environment, transduce mechanical into chemical information, and integrate these signals with growth factor derived stimuli to achieve specific changes in gene expression. For connective tissue cells, production of the extracellular matrix is a prominent response to changes in mechanical load. We will review the evidence that integrin-containing cell-matrix adhesion contacts are essential for force transmission from the extracellular matrix to the cytoskeleton, and describe novel experiments indicating that mechanotransduction in fibroblasts depends on focal adhesion adaptor proteins that might function as molecular springs. We will stress the importance of the contractile actin cytoskeleton in balancing external with internal forces, and describe new results linking force-controlled actin dynamics directly to the expression of specific genes, among them the extracellular matrix protein tenascin-C. As assembly lines for diverse signaling pathways, matrix adhesion contacts are now recognized as the major sites of crosstalk between mechanical and chemical stimuli, with important consequences for cell growth and differentiation.

  7. A Marfan syndrome gene expression phenotype in cultured skin fibroblasts

    Directory of Open Access Journals (Sweden)

    Emond Mary

    2007-09-01

    Full Text Available Abstract Background Marfan syndrome (MFS is a heritable connective tissue disorder caused by mutations in the fibrillin-1 gene. This syndrome constitutes a significant identifiable subtype of aortic aneurysmal disease, accounting for over 5% of ascending and thoracic aortic aneurysms. Results We used spotted membrane DNA macroarrays to identify genes whose altered expression levels may contribute to the phenotype of the disease. Our analysis of 4132 genes identified a subset with significant expression differences between skin fibroblast cultures from unaffected controls versus cultures from affected individuals with known fibrillin-1 mutations. Subsequently, 10 genes were chosen for validation by quantitative RT-PCR. Conclusion Differential expression of many of the validated genes was associated with MFS samples when an additional group of unaffected and MFS affected subjects were analyzed (p-value -6 under the null hypothesis that expression levels in cultured fibroblasts are unaffected by MFS status. An unexpected observation was the range of individual gene expression. In unaffected control subjects, expression ranges exceeding 10 fold were seen in many of the genes selected for qRT-PCR validation. The variation in expression in the MFS affected subjects was even greater.

  8. Establishment, Culture, and Characterization of Guinea Pig Fetal Fibroblast Cell

    Directory of Open Access Journals (Sweden)

    Davood Mehrabani

    2014-01-01

    Full Text Available Establishment of Guinea pig fetal fibroblast cells and their biological evaluation before and after cryopreservation were the main purposes of this study. After determination of the proper age of pregnancy by ultrasonography, 30 days old fetuses of Guinea pigs were recovered. Their skins were cut into small pieces (1 mm2 and were cultured. When reaching 80–90% confluence, the cells were passaged. Cells of the second and eighth passages were cultured in 24-well plates (4×104 cells/well for 6 days and three wells per day were counted. The average cell counts at each time point were then plotted against time and the population doubling time (PDT was determined. Then, vials of cells (2×106 cells/mL were cryopreserved for 1 month and after thawing, the cell viability was evaluated. The PDT of the second passage was about 23 h and for the eighth passage was about 30 h. The viability of the cultures was 95% in the second passage and 74.5% in the eighth passage. It was shown that the Guinea pig fetal fibroblast cell culture can be established using the adherent culture method while, after freezing, the viability indices of these cells were favorable.

  9. Biological activities of frankincense essential oil in human dermal fibroblasts

    Directory of Open Access Journals (Sweden)

    Xuesheng Han

    2017-06-01

    Full Text Available Although frankincense essential oil (FREO has become increasingly popular in skin care, research on its biological activities in human skin cells is scarce, if not completely absent. In the current study, we explored the biological activities of FREO in pre-inflamed human dermal fibroblasts by analyzing the levels of 17 important protein biomarkers pertinent to inflammation and tissue remodeling. FREO exhibited robust anti-proliferative activity in these skin cells. It also significantly inhibited collagen III, interferon gamma-induced protein 10, and intracellular cell adhesion molecule 1. We also studied its effect in regulating genome-wide gene expression. FREO robustly modulated global gene expression. Furthermore, Ingenuity® Pathway Analysis showed that FREO affected many important signaling pathways that are closely related to inflammation, immune response, and tissue remodeling. This study provides the first evidence of the biological activities of FREO in human dermal fibroblasts. Consistent with existing studies in other models, the current study suggests that FREO possesses promising potential to modulate the biological processes of inflammation and tissue remodeling in human skin. Further research into the biological mechanisms of action of FREO and its major active components is recommended.

  10. Adenoviral-mediated correction of methylmalonyl-CoA mutase deficiency in murine fibroblasts and human hepatocytes

    Directory of Open Access Journals (Sweden)

    Korson Mark

    2007-04-01

    Full Text Available Abstract Background Methylmalonic acidemia (MMA, a common organic aciduria, is caused by deficiency of the mitochondrial localized, 5'deoxyadenosylcobalamin dependent enzyme, methylmalonyl-CoA mutase (MUT. Liver transplantation in the absence of gross hepatic dysfunction provides supportive therapy and metabolic stability in severely affected patients, which invites the concept of using cell and gene delivery as future treatments for this condition. Methods To assess the effectiveness of gene delivery to restore the defective metabolism in this disorder, adenoviral correction experiments were performed using murine Mut embryonic fibroblasts and primary human methylmalonyl-CoA mutase deficient hepatocytes derived from a patient who harbored two early truncating mutations, E224X and R228X, in the MUT gene. Enzymatic and expression studies were used to assess the extent of functional correction. Results Primary hepatocytes, isolated from the native liver after removal subsequent to a combined liver-kidney transplantation procedure, or Mut murine fibroblasts were infected with a second generation recombinant adenoviral vector that expressed the murine methylmalonyl-CoA mutase as well as eGFP from distinct promoters. After transduction, [1-14C] propionate macromolecular incorporation studies and Western analysis demonstrated complete correction of the enzymatic defect in both cell types. Viral reconstitution of enzymatic expression in the human methylmalonyl-CoA mutase deficient hepatocytes exceeded that seen in fibroblasts or control hepatocytes. Conclusion These experiments provide proof of principle for viral correction in methylmalonic acidemia and suggest that hepatocyte-directed gene delivery will be an effective therapeutic treatment strategy in both murine models and in human patients. Primary hepatocytes from a liver that was unsuitable for transplantation provided an important resource for these studies.

  11. Oral pirfenidone protects against fibrosis by inhibiting fibroblast proliferation and TGF-β signaling in a murine colitis model.

    Science.gov (United States)

    Li, Guanwei; Ren, Jianan; Hu, Qiongyuan; Deng, Youming; Chen, Guopu; Guo, Kun; Li, Ranran; Li, Yuan; Wu, Lei; Wang, Gefei; Gu, Guosheng; Li, Jieshou

    2016-10-01

    Inflammatory bowel disease (IBD), particularly Crohn's disease, frequently causes intestinal fibrosis that ultimately leads to formation of strictures requiring bowel resection. Currently there is no effective antifibrotic therapy available for this disease. Pirfenidone is a small compound that has a broad spectrum of antifibrogenic effect and has been used for the treatment of fibrotic diseases in various organs. The present study aimed to investigate the antifibrogenic effect of pirfenidone in a dextran sulfate sodium (DSS)-induced murine colitis model. C57BL/6 mice were used and animals were randomly divided into groups receiving pirfenidone or vehicle by oral or transanal routes. Inflammation- and fibrosis-related indexes including body weight, colon length, disease activity, histological change, mRNA expression of pro-inflammatory and pro-fibrogenic cytokines were assessed. Furthermore, we performed in vitro analysis using CCD18-Co fibroblasts to evaluate cell proliferation, transdifferentiation, and viability after the cells were cultured with pirfenidone. It was found that oral administration of pirfenidone reduced deposition of collagen in colitis-associated fibrosis, and significantly suppressed the mRNA expression of col1a2, col3a1, and TGF-β. Moreover, pirfenidone inhibited the activation of TGF-β-related smad and MAPK pathways both in vitro and in vivo. Clinical and histological evaluation demonstrated that pirfenidone had no anti-inflammatory effect. The antifibrogenic effect was reduced when pirfenidone was administered in a delayed manner and was unobserved if given locally. Pirfenidone suppressed fibroblast proliferation and transdifferentiation without observed toxicity. Altogether, our results suggested that oral pirfenidone protects against fibrosis of DSS-induced colitis through inhibiting the proliferation of colonic fibroblasts and TGF-β signaling pathways. Copyright © 2016 Elsevier Inc. All rights reserved.

  12. TAT-Mediated Acidic Fibroblast Growth Factor Delivery to the Dermis Improves Wound Healing of Deep Skin Tissue in Rat.

    Directory of Open Access Journals (Sweden)

    Long Zheng

    Full Text Available The definition of deep tissue injury was derived from multiple clinical cases as "A purple or maroon localized area of discolored intact skin or blood-filled blister due to damage of underlying soft tissue from pressure and/or shear". Acidic fibroblast growth factor (aFGF significantly improves wound healing under diabetic conditions. However, to date, the therapeutic application of aFGF has been limited, due to its low delivery efficiency and short half-life.Using an animal model of magnet-induced pressure ulcers, transactivator of transcription protein (TAT-aFGF was evaluated for transdermal delivery and wound healing. Immunohistochemistry and Western blotting were also performed to determine the expression of transforming growth factor (TGF-β1, α-smooth muscle actin (α-SMA, CD68, proliferating cell nuclear antigen (PCNA and TGF-β-receptor II (TGF- βRII in cultured human dermal fibroblasts. We found that that mice treated with TAT-aFGF had higher accumulation of aFGF in both dermis and subcutaneous tissues compared with mice treated with aFGF alone. In the remodeling phase, TAT-aFGF treatment decreased the expression of α-SMA to normal levels, thereby facilitating normal wound healing processes and abrogating hypertrophic scarring. In human dermal fibroblasts, TAT-aFGF reversed the suppressive effect of TNF-α on α-SMA expression and restored TGF-βRII and TGF-β1 expression.Our results demonstrate that TAT-aFGF has a favorable therapeutic effect on the healing of subcutaneous deep tissue injury.

  13. ALS-Related Mutant FUS Protein Is Mislocalized to Cytoplasm and Is Recruited into Stress Granules of Fibroblasts from Asymptomatic FUS P525L Mutation Carriers.

    Science.gov (United States)

    Lo Bello, Margherita; Di Fini, Francesca; Notaro, Antonietta; Spataro, Rossella; Conforti, Francesca L; La Bella, Vincenzo

    2017-10-17

    Amyotrophic lateral sclerosis (ALS) shows a strong genetic basis, with SOD1, FUS, TARDBP, and C9ORF72 being the genes most frequently involved. This has allowed identification of asymptomatic mutation carriers, which may be of help in understanding the molecular changes preceding disease onset. We studied the cellular expression of FUS protein and the effect of heat-shock- and dithiothreitol-induced stress in fibroblasts from FUS P525L mutation carriers, healthy controls, and patients with sporadic ALS. Western blots and immunocytochemistry were performed to study the subcellular localization of FUS protein. Control and stressed cells were double stained with FUS and the stress marker TIA-R. Fibroblasts from healthy controls and sporadic ALS cases showed a prominent nuclear FUS expression. In the 2 FUS P525L mutation carriers, instead, most cells showed a protein localization in both nucleus and cytoplasm, or exclusively in the cytoplasm. Stress prompted the formation of cytoplasmic granules in all subjects and in sporadic ALS FUS mislocalization to the cytoplasm. Cytoplasmic FUS was recruited into stress granules, which showed a time-dependent decrease in all subjects. However, in the FUS P525L fibroblasts, the granules persisted longer, and they were more numerous than those detected in the cells from controls and sporadic ALS patients. We show that in fibroblasts of FUS P525L mutation carriers, FUS mislocalized to the cytoplasm where it redistributed into stress granules with likely a dose effect, i.e. a higher number of cells with granules, which persist longer, than in controls and ALS cases. These data represent an early molecular change occurring before ALS onset, suggesting a transient preaggregative state. © 2017 S. Karger AG, Basel.

  14. Anti-fibroblast antibodies detected by cell-based ELISA in systemic sclerosis enhance the collagenolytic activity and matrix metalloproteinase-1 production in dermal fibroblasts

    National Research Council Canada - National Science Library

    Fineschi, S; Cozzi, F; Burger, D; Dayer, J.-M; Meroni, P. L; Chizzolini, C

    2007-01-01

    ... to induce collagen deposition or degradation. Results. Fibroblasts stimulated with AFA-positive but not with AFA-negative and control IgG showed an increased capacity to digest collagen matrix and produce metalloproteinase-1 (MMP-1...

  15. Adipose tissue-derived stromal cells inhibit TGF-β1-induced differentiation of human dermal fibroblasts and keloid scar-derived fibroblasts in a paracrine fashion.

    Science.gov (United States)

    Spiekman, Maroesjka; Przybyt, Ewa; Plantinga, Josée A; Gibbs, Susan; van der Lei, Berend; Harmsen, Martin C

    2014-10-01

    Adipose tissue-derived stromal cells augment wound healing and skin regeneration. It is unknown whether and how they can also influence dermal scarring. The authors hypothesized that adipose tissue-derived stromal cells inhibit adverse differentiation of dermal fibroblasts induced by the pivotal factor in scarring, namely, transforming growth factor (TGF)-β. TGF-β1-treated adult human dermal fibroblasts and keloid scar-derived fibroblasts were incubated with adipose tissue-derived stromal cell-conditioned medium and assessed for proliferation and differentiation, particularly the production of collagen, expression of SM22α, and development of hypertrophy and contractility. TGF-β1-induced proliferation of adult human dermal fibroblasts was abolished by adipose tissue-derived stromal cell-conditioned medium. Simultaneously, the medium reduced SM22α gene and protein expression of TGF-β1-treated adult human dermal fibroblasts, and their contractility was reduced also. Furthermore, the medium strongly reduced transcription of collagen I and III genes and their corresponding proteins. In contrast, it tipped the balance of matrix turnover to degradation through stimulating gene expression of matrix metalloproteinase (MMP)-1, MMP-2, and MMP-14, whereas MMP-2 activity was up-regulated also. Even in end-stage myofibroblasts (i.e., keloid scar-derived fibroblasts), adipose tissue-derived stromal cell-conditioned medium suppressed TGF-β1-induced myofibroblast contraction and collagen III gene expression. The authors show that adipose tissue-derived stromal cells inhibit TGF-β1-induced adverse differentiation and function of adult human dermal fibroblasts and TGF-β1-induced contraction in keloid scar-derived fibroblasts, in a paracrine fashion.

  16. Fibroblast Growth Factor 23 in Postrenal Transplant: An Often Forgotten Hormone.

    Science.gov (United States)

    Amiri, Fateme Shamekhi; Khatami, Mohammad Reza

    2016-12-01

    Fibroblast growth factor 23 is likely to be the most important regulator of phosphate homeostasis, which mediates its functions through fibroblast growth factor receptors and the coreceptor Klotho. In addition to reducing expression of the sodium-phosphate cotransporters NPT2a and NPT2c in the proximal tubules, fibroblast growth factor 23 inhibits renal 1α-hydroxylase and stimulates 24-hydroxylase and appears to reduce parathyroid hormone secretion in short-term studies. Fibroblast growth factor 23 synthesis and secretion by osteocytes and osteoblasts are upregulated through 1,25-dihydroxyvitamin D3 and through an increased dietary phosphate intake. Recent studies have indicated that a low-protein diet and calcium deficiency reduce circulating fibroblast growth factor 23 levels, but magnesium deficiency increases fibroblast growth factor levels. Drugs such as phosphate binders, bisphosphonate, and estrogens have various effects on circulating fibroblast growth factor 23 levels. The high cardiovascular disease event rates and mortality associated with elevated levels of this hormone may be due to various effects on the cardiovascular system, including left ventricular hypertrophy, arterial stiffness, vascular calcifications, endothelial dysfunction, and increased levels of inflammatory markers. In addition, elevated levels of this hormone may contribute to mineral bone metabolism disorders and to patient and allograft survival after renal transplant. Here, we discuss the effects of fibroblast growth factor 23 on adverse renal, bone, and cardiovascular outcomes after kidney transplant.

  17. Effect of interleukin-17 on gene expression profile of fibroblasts from Crohn's disease patients

    NARCIS (Netherlands)

    Kerami, Zohra; Duijvis, Nicolette W.; Vogels, Esther W.; van Dooren, Faas H.; Moerland, Perry D.; te Velde, Anje A.

    2014-01-01

    The expression of interleukin (IL)-17 is upregulated in inflammatory bowel disease (IBD). Since fibroblasts are known to be responsive to IL-17, they may play a role in the modulation of inflammatory responses in IBD. Here, the effects of IL-17 on ileum and colon fibroblasts from Crohn's disease

  18. Fibroblast growth factor receptor 3 protein is overexpressed in oral and oropharyngeal squamous cell carcinoma

    NARCIS (Netherlands)

    Koole, Koos; van Kempen, Pauline M W; Swartz, Justin E|info:eu-repo/dai/nl/413983390; Peeters, Ton; van Diest, Paul J|info:eu-repo/dai/nl/075281775; Koole, Ron|info:eu-repo/dai/nl/123508126; van Es, Robert J. J.|info:eu-repo/dai/nl/216460646; Willems, Stefan M|info:eu-repo/dai/nl/33189582X

    Fibroblast growth factor receptor 3 (FGFR3) is a member of the fibroblast growth factor receptor tyrosine kinase family. It has been identified as a promising therapeutic target in multiple types of cancer. We have investigated FGFR3 protein expression and FGFR3 gene copy-numbers in a single

  19. High inorganic phosphate causes DNMT1 phosphorylation and subsequent fibrotic fibroblast activation

    Energy Technology Data Exchange (ETDEWEB)

    Tan, Xiaoying [Department of Nephrology and Rheumatology, Göttingen University Medical Center, Georg August University, Göttingen (Germany); Department of Cardiology and Pneumology, Göttingen University Medical Center, Georg August University, Göttingen (Germany); Xu, Xingbo [Department of Cardiology and Pneumology, Göttingen University Medical Center, Georg August University, Göttingen (Germany); Zeisberg, Elisabeth M. [Department of Cardiology and Pneumology, Göttingen University Medical Center, Georg August University, Göttingen (Germany); German Center for Cardiovascular Research (DZHK), Göttingen (Germany); Zeisberg, Michael, E-mail: mzeisberg@med.uni-goettingen.de [Department of Nephrology and Rheumatology, Göttingen University Medical Center, Georg August University, Göttingen (Germany); German Center for Cardiovascular Research (DZHK), Göttingen (Germany)

    2016-04-08

    Phosphate is an essential constituent of critical cellular functions including energy metabolism, nucleic acid synthesis and phosphorylation-dependent cell signaling. Increased plasma phosphate levels are an independent risk factor for lowered life-expectancy as well as for heart and kidney failure. Nevertheless, direct cellular effects of elevated phosphate concentrations within the microenvironment are poorly understood and have been largely neglected in favor of phosphor-regulatory hormones. Because interstitial fibrosis is the common determinant of chronic progressive kidney disease, and because fibroblasts are major mediators of fibrogenesis, we here explored the effect of high extracellular phosphate levels on renal fibroblasts. We demonstrate that high inorganic phosphate directly induces fibrotic fibroblast activation associated with increased proliferative activity, increased expression of α-smooth muscle actin and increased synthesis of type I collagen. We further demonstrate that such fibroblast activation is dependent on phosphate influx, aberrant phosphorylation of DNA methyltransferase DNMT1 and aberrant CpG island promoter methylation. In summary, our studies demonstrate that elevated phosphate concentrations induce pro-fibrotic fibroblast activation independent of phospho-regulatory hormones. - Highlights: • We exposed human kidney fibroblasts to media containing 1 mM or 3 mM phosphate. • Increased phosphate influx causes phosphorylation of DNA methyltransferase Dnmt1. • Phosphorylated Dnmt1 causes promoter methylation and transcriptional silencing of RASAL1. • Depletion of RASAL1 causes increased intrinsic Ras-GTP activity and fibroblast activation. • Inorganic phosphate causes fibroblast activation independent of phospho-regulatory hormones.

  20. Mitochondrial Bioenergetics Is Altered in Fibroblasts from Patients with Sporadic Alzheimer's Disease

    Directory of Open Access Journals (Sweden)

    María J. Pérez

    2017-10-01

    Full Text Available The identification of an early biomarker to diagnose Alzheimer's disease (AD remains a challenge. Neuropathological studies in animal and AD patients have shown that mitochondrial dysfunction is a hallmark of the development of the disease. Current studies suggest the use of peripheral tissues, like skin fibroblasts as a possibility to detect the early pathological alterations present in the AD brain. In this context, we studied mitochondrial function properties (bioenergetics and morphology in cultured fibroblasts obtained from AD, aged-match and young healthy patients. We observed that AD fibroblasts presented a significant reduction in mitochondrial length with important changes in the expression of proteins that control mitochondrial fusion. Moreover, AD fibroblasts showed a distinct alteration in proteolytic processing of OPA1, a master regulator of mitochondrial fusion, compared to control fibroblasts. Complementary to these changes AD fibroblasts showed a dysfunctional mitochondrial bioenergetics profile that differentiates these cells from aged-matched and young patient fibroblasts. Our findings suggest that the human skin fibroblasts obtained from AD patients could replicate mitochondrial impairment observed in the AD brain. These promising observations suggest that the analysis of mitochondrial bioenergetics could represent a promising strategy to develop new diagnostic methods in peripheral tissues of AD patients.

  1. Breast Cancer-Associated Fibroblasts: Where We Are and Where We Need to Go

    Directory of Open Access Journals (Sweden)

    Rachel J. Buchsbaum

    2016-01-01

    Full Text Available Cancers are heterogeneous tissues comprised of multiple components, including tumor cells and microenvironment cells. The tumor microenvironment has a critical role in tumor progression. The tumor microenvironment is comprised of various cell types, including fibroblasts, macrophages and immune cells, as well as extracellular matrix and various cytokines and growth factors. Fibroblasts are the predominant cell type in the tumor microenvironment. However, neither the derivation of tissue-specific cancer-associated fibroblasts nor markers of tissue-specific cancer-associated fibroblasts are well defined. Despite these uncertainties it is increasingly apparent that cancer-associated fibroblasts have a crucial role in tumor progression. In breast cancer, there is evolving evidence showing that breast cancer-associated fibroblasts are actively involved in breast cancer initiation, proliferation, invasion and metastasis. Breast cancer-associated fibroblasts also play a critical role in metabolic reprogramming of the tumor microenvironment and therapy resistance. This review summarizes the current understanding of breast cancer-associated fibroblasts.

  2. In vitro effects of pirfenidone on cardiac fibroblasts: proliferation, myofibroblast differentiation, migration and cytokine secretion.

    Directory of Open Access Journals (Sweden)

    Qiang Shi

    Full Text Available Cardiac fibroblasts (CFs are the primary cell type responsible for cardiac fibrosis during pathological myocardial remodeling. Several studies have illustrated that pirfenidone (5-methyl-1-phenyl-2-[1H]-pyridone attenuates cardiac fibrosis in different animal models. However, the effects of pirfenidone on cardiac fibroblast behavior have not been examined. In this study, we investigated whether pirfenidone directly modulates cardiac fibroblast behavior that is important in myocardial remodeling such as proliferation, myofibroblast differentiation, migration and cytokine secretion. Fibroblasts were isolated from neonatal rat hearts and bioassays were performed to determine the effects of pirfenidone on fibroblast function. We demonstrated that treatment of CFs with pirfenidone resulted in decreased proliferation, and attenuated fibroblast α-smooth muscle actin expression and collagen contractility. Boyden chamber assay illustrated that pirfenidone inhibited fibroblast migration ability, probably by decreasing the ratio of matrix metalloproteinase-9 to tissue inhibitor of metalloproteinase-1. Furthermore, pirfenidone attenuated the synthesis and secretion of transforming growth factor-β1 but elevated that of interleukin-10. These direct and pleiotropic effects of pirfenidone on cardiac fibroblasts point to its potential use in the treatment of adverse myocardial remodeling.

  3. Rho Kinase Regulation of Fibroblast Migratory Mechanics in Fibrillar Collagen Matrices

    OpenAIRE

    Zhou, Chengxin; Petroll, W. Matthew

    2010-01-01

    Migration of activated corneal fibroblasts plays an important role in matrix patterning during embryonic development and wound repopulation following injury or refractive surgery. In this study, we investigate the role of Rho kinase in regulating fibroblast migration mechanics, by modifying a previously described nested collagen matrix model to facilitate dynamic imaging of cell-matrix interactions.

  4. Printing-induced cell injury evaluation during laser printing of 3T3 mouse fibroblasts.

    Science.gov (United States)

    Zhang, Zhengyi; Chai, Wenxuan; Xiong, Ruitong; Zhou, Lei; Huang, Yong

    2017-06-20

    Three-dimensional bioprinting has emerged as a promising solution for the freeform fabrication of living cellular constructs, which can be used for tissue/organ transplantation and tissue models. During bioprinting, some living cells are unavoidably injured and may become necrotic or apoptotic cells. This study aims to investigate the printing-induced cell injury and evaluates injury types of post-printing cells using the annexin V/7-aminoactinomycin D and FAM-DEVD-FMK/propidium iodide assays during laser printing of NIH 3T3 mouse fibroblasts. As observed, the percentage of post-printing early apoptotic mouse fibroblasts increases with the incubation time, indicating that post-printing apoptotic mouse fibroblasts have different initiation lag times of apoptosis due to different levels of mechanical stress exerted during laser printing. Post-printing necrotic mouse fibroblasts can be detected immediately after printing, while post-printing early apoptotic mouse fibroblasts need time to develop into a late apoptotic stage. The minimum time needed for post-printing early apoptotic mouse fibroblasts to complete their apoptosis pathway and transition into late apoptotic mouse fibroblasts is from 4 h to 5 h post-printing. The resulting knowledge of the evolution of different apoptotic post-printing mouse fibroblasts will help better design future experiments to quantitatively determine, model, and mitigate the post-printing cell injury based on molecular signal pathway modeling.

  5. Enhanced ROCK1 dependent contractility in fibroblast from chronic obstructive pulmonary disease patients

    Directory of Open Access Journals (Sweden)

    Hallgren Oskar

    2012-08-01

    Full Text Available Abstract Background During wound healing processes fibroblasts account for wound closure by adopting a contractile phenotype. One disease manifestation of COPD is emphysema which is characterized by destruction of alveolar walls and our hypothesis is that fibroblasts in the COPD lungs differentiate into a more contractile phenotype as a response to the deteriorating environment. Methods Bronchial (central and parenchymal (distal fibroblasts were isolated from lung explants from COPD patients (n = 9 (GOLD stage IV and from biopsies from control subjects and from donor lungs (n = 12. Tissue-derived fibroblasts were assessed for expression of proteins involved in fibroblast contraction by western blotting whereas contraction capacity was measured in three-dimensional collagen gels. Results The basal expression of rho-associated coiled-coil protein kinase 1 (ROCK1 was increased in both centrally and distally derived fibroblasts from COPD patients compared to fibroblasts from control subjects (p  Conclusions Distally derived fibroblasts from COPD patients have an enhanced contractile phenotype that is dependent on ROCK1 activity. This feature may be of importance for the elastic dynamics of small airways and the parenchyma in late stages of COPD.

  6. Living skin substitutes: survival and function of fibroblasts seeded in a dermal substitute in experimental wounds

    NARCIS (Netherlands)

    Lamme, E. N.; van Leeuwen, R. T.; Jonker, A.; van Marle, J.; Middelkoop, E.

    1998-01-01

    The healing of full-thickness skin defects requires extensive synthesis and remodeling of dermal and epidermal components. Fibroblasts play an important role in this process and are being incorporated in the latest generation of artificial dermal substitutes. We studied the fate of fibroblasts

  7. Persistence of high-replicative capacity in cultured fibroblasts from nonagenarians

    NARCIS (Netherlands)

    Maier, Andrea B.; Le Cessie, Saskia; De Koning-treurniet, Corine; Blom, Johanna; Westendorp, Rudi G.J.; Van Heemst, Diana

    Earlier studies on human fibroblast cultures have demonstrated an inverse relationship between the total number of population doublings (PDs) and donor age. As more recent studies were unable to replicate these findings, we set out to analyze growth characteristics of fibroblast cultures from

  8. Relation between replicative senescence of human fibroblasts and life history characteristics

    NARCIS (Netherlands)

    Maier, Andrea B.; Westendorp, Rudi G J

    Replicative ageing of fibroblasts in vitro has often been used as a model for organismal ageing. The general assumption that the ageing process is mirrored by cellular senescence in vitro is based on lower replicative capacity of human fibroblasts from patients with accelerated ageing syndromes,

  9. Impaired response of fibroblasts from patients with hyperapobetalipoproteinemia to acylation-stimulating protein.

    OpenAIRE

    Cianflone, K M; Maslowska, M H; Sniderman, A D

    1990-01-01

    Acylation-stimulating protein (ASP) is a small, basic, human plasma protein that markedly stimulates triglyceride synthesis in human adipocytes and cultured human skin fibroblasts. The present studies examine the response to ASP of cultured skin fibroblasts from normal subjects patients with hyperapobetalipoproteinemia, patients with familial hypercholesterolemia, and patients with hypertriglyceridemia without hyperapobetalipoproteinemia. Triglyceride synthesis induced by ASP did not differ s...

  10. Versican V1 Overexpression Induces a Myofibroblast-Like Phenotype in Cultured Fibroblasts.

    Directory of Open Access Journals (Sweden)

    Jon M Carthy

    Full Text Available Versican, a chondroitin sulphate proteoglycan, is one of the key components of the provisional extracellular matrix expressed after injury. The current study evaluated the hypothesis that a versican-rich matrix alters the phenotype of cultured fibroblasts.The full-length cDNA for the V1 isoform of human versican was cloned and the recombinant proteoglycan was expressed in murine fibroblasts. Versican expression induced a marked change in fibroblast phenotype. Functionally, the versican-expressing fibroblasts proliferated faster and displayed enhanced cell adhesion, but migrated slower than control cells. These changes in cell function were associated with greater N-cadherin and integrin β1 expression, along with increased FAK phosphorylation. The versican-expressing fibroblasts also displayed expression of smooth muscle α-actin, a marker of myofibroblast differentiation. Consistent with this observation, the versican fibroblasts displayed increased synthetic activity, as measured by collagen III mRNA expression, as well as a greater capacity to contract a collagen lattice. These changes appear to be mediated, at least in part, by an increase in active TGF-β signaling in the versican expressing fibroblasts, and this was measured by phosphorylation and nuclear accumulation of SMAD2.Collectively, these data indicate versican expression induces a myofibroblast-like phenotype in cultured fibroblasts.

  11. Mitochondrial Bioenergetics Is Altered in Fibroblasts from Patients with Sporadic Alzheimer's Disease

    Science.gov (United States)

    Pérez, María J.; Ponce, Daniela P.; Osorio-Fuentealba, Cesar; Behrens, Maria I.; Quintanilla, Rodrigo A.

    2017-01-01

    The identification of an early biomarker to diagnose Alzheimer's disease (AD) remains a challenge. Neuropathological studies in animal and AD patients have shown that mitochondrial dysfunction is a hallmark of the development of the disease. Current studies suggest the use of peripheral tissues, like skin fibroblasts as a possibility to detect the early pathological alterations present in the AD brain. In this context, we studied mitochondrial function properties (bioenergetics and morphology) in cultured fibroblasts obtained from AD, aged-match and young healthy patients. We observed that AD fibroblasts presented a significant reduction in mitochondrial length with important changes in the expression of proteins that control mitochondrial fusion. Moreover, AD fibroblasts showed a distinct alteration in proteolytic processing of OPA1, a master regulator of mitochondrial fusion, compared to control fibroblasts. Complementary to these changes AD fibroblasts showed a dysfunctional mitochondrial bioenergetics profile that differentiates these cells from aged-matched and young patient fibroblasts. Our findings suggest that the human skin fibroblasts obtained from AD patients could replicate mitochondrial impairment observed in the AD brain. These promising observations suggest that the analysis of mitochondrial bioenergetics could represent a promising strategy to develop new diagnostic methods in peripheral tissues of AD patients. PMID:29056898

  12. TRIF promotes angiotensin II-induced cross-talk between fibroblasts and macrophages in atrial fibrosis

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Xiao-Qing; Zhang, Dao-Liang [Department of Cardiology, Shanghai Chest Hospital, Shanghai Jiaotong University, Shanghai (China); Zhang, Ming-Jian; Guo, Meng; Zhan, Yang-Yang; Liu, Fang [National Key Laboratory of Medical Immunology, Second Military Medical University, Shanghai (China); Jiang, Wei-Feng; Zhou, Li [Department of Cardiology, Shanghai Chest Hospital, Shanghai Jiaotong University, Shanghai (China); Zhao, Liang, E-mail: zhaol_zg@163.com [Department of Cardiology, Shanghai Chest Hospital, Shanghai Jiaotong University, Shanghai (China); Wang, Quan-Xing, E-mail: wqxejd@126.com [National Key Laboratory of Medical Immunology, Second Military Medical University, Shanghai (China); Liu, Xu, E-mail: liuxu_xk@163.com [Department of Cardiology, Shanghai Chest Hospital, Shanghai Jiaotong University, Shanghai (China)

    2015-08-14

    Aims: Atrial fibroblasts and macrophages have long been thought to participate in atrial fibrillation (AF). However, which specific mediator may regulate the interaction between them remains unclear. Methods and results: We provided the evidence for the involvement of Toll/IL-1 receptor domain-containing adaptor inducing IFN-β (TRIF), an important inflammation-related molecule, in the pathophysiology of AF. Patients with AF showed higher levels of angiotensin II (AngII) and TRIF expression and larger number of macrophages infiltration in left atria appendage than individuals with sinus rhythm (SR). In the cell study, AngII induced chemokines expressions in mouse atrial fibroblasts and AngII-stimulated atrial fibroblasts induced the chemotaxis of macrophages, which were reduced by losartan and TRIF siRNA. Meanwhile, AngII-stimulated atrial fibroblasts proliferation was enhanced by macrophages. Conclusions: Our data demonstrated that TRIF may be a crucial factor promoting the interaction between atrial fibroblasts and macrophages, leading to atrial fibrosis. - Highlights: • Compared with SR, AF showed higher TRIF expression in left atrial appendage. • TRIF siRNA reversed macrophage chemotaxis induced by AngII-treated fibroblast. • TRIF siRNA reversed chemokines expressions induced by AngII in fibroblast. • AngII-stimulated atrial fibroblast proliferation was enhanced by macrophage.

  13. Local Foods, Local Places Toolkit

    Science.gov (United States)

    Toolkit to help communities that want to use local foods to spur revitalization. The toolkit gives step-by-step instructions to help communities plan and host a workshop and create an action plan to implement.

  14. Instability of spiral and scroll waves in the presence of a gradient in the fibroblast density: the effects of fibroblast-myocyte coupling

    Science.gov (United States)

    Zimik, Soling; Pandit, Rahul

    2016-12-01

    Fibroblast-myocyte coupling can modulate electrical-wave dynamics in cardiac tissue. In diseased hearts, the distribution of fibroblasts is heterogeneous, so there can be gradients in the fibroblast density (henceforth we call this GFD) especially from highly injured regions, like infarcted or ischemic zones, to less-wounded regions of the tissue. Fibrotic hearts are known to be prone to arrhythmias, so it is important to understand the effects of GFD in the formation and sustenance of arrhythmic re-entrant waves, like spiral or scroll waves. Therefore, we investigate the effects of GFD on the stability of spiral and scroll waves of electrical activation in a state-of-the-art mathematical model for cardiac tissue in which we also include fibroblasts. By introducing GFD in controlled ways, we show that spiral and scroll waves can be unstable in the presence of GFDs because of regions with varying spiral- or scroll-wave frequency ω, induced by the GFD. We examine the effects of the resting membrane potential of the fibroblast and the number of fibroblasts attached to the myocytes on the stability of these waves. Finally, we show that the presence of GFDs can lead to the formation of spiral waves at high-frequency pacing.

  15. Ethanol exposure induces the cancer-associated fibroblast phenotype and lethal tumor metabolism: Implications for breast cancer prevention

    National Research Council Canada - National Science Library

    Sanchez-Alvarez, Rosa; Martinez-Outschoorn, Ubaldo E; Lin, Zhao; Lamb, Rebecca; Hulit, James; Howell, Anthony; Sotgia, Federica; Rubin, Emanuel; Lisanti, Michael P

    2013-01-01

    ...) and hTERT-immortalized fibroblasts. Here, we show that ethanol treatment (100 mM) promotes ROS production and oxidative stress in cancer-associated fibroblasts, which is sufficient to induce myofibroblastic differentiation...

  16. [Effect of Capparis spinosa on fibroblast proliferation and type I collagen production in progressive systemic sclerosis].

    Science.gov (United States)

    Cao, Yue-Lan; Li, Xin; Zheng, Min

    2008-03-01

    To investigate the effects of ethanolic extract from Capparis spinosa (ECS) on the fibroblast proliferation and type I collagen production in normal and progressive systemic sclerosis (PSS). Cellular activity was determined by the MTT method. Apoptosis was detected by flow cytometry analysis of Annexin V-stained cells. The expression levels of type I collagen messenger RNA and protein were analyzed by RT-PCR and western blot analysis. ECS could significantly inhibit the proliferation of fibroblast and reduced the expression of alpha2 (I) collagen mRNA and type I collagen protein in PSS in a dose-and time-dependent manner. ECS did not affect the proliferation of fibroblast and expression of type I collagen mRNA and protein in normal human. ECS could counteract the harmful effects on fibroblast by H2O2. ECS can effectively inhibit the fibroblast proliferation and type I collagen production in PSS.

  17. Stromal-epithelial interactions in aging and cancer: Senescent fibroblasts alter epithelial cell differentiation

    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.

  18. [Effect of PRX-2 gene transferred by lipofectamine on the proliferation of human skin fibroblasts].

    Science.gov (United States)

    Song, Hui-feng; Chai, Jia-ke; Lin, Zi-hao

    2011-10-11

    To explore the effects of PRX-2 gene transferred by lipofectamine on the human skin fibroblasts. Normal human skin fibroblasts were cultured and PRX-2 gene was transferred by lipofectamine. The proliferation of fibroblasts was detected by MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide) and flow cytometry. The proliferation of PRX-2-transfected fibroblasts was stronger than that of normal counterparts. There were fewer cells during G0-G1 period and more cells during S and G2-M periods. The proliferative index increased. The proliferation of fibroblasts may be modified by transfected PRX-2. Thus PRX-2 plays an important role during the healing of human skin wound.

  19. Hypoxia Enhances Direct Reprogramming of Mouse Fibroblasts to Cardiomyocyte-Like Cells.

    Science.gov (United States)

    Wang, Yanyan; Shi, Shujun; Liu, Huiwen; Meng, Li

    2016-02-01

    Recent work has shown that mouse and human fibroblasts can be reprogrammed to cardiomyocyte-like cells with a combination of transcription factors. Current research has focused on improving the efficiency and mechanisms for fibroblast reprogramming. Previously, it has been reported that hypoxia enhances fibroblast cell reprogramming to pluripotent stem cells. In this study, we observed that 6 h of hypoxic conditions (2% oxygen) on newborn mouse dermal fibroblasts can improve the efficiency of reprogramming to cardiomyocyte-like cells. Expression of cardiac-related genes and proteins increased at 4 weeks after transfer of three transcription factors (Gata4/Mef2c/Tbx5 [GMT]). However, beating cardiomyocyte cells were not detected. The epigenetic mechanism of hypoxia-induced fibroblast reprogramming to cardiomyocyte cells requires further study.

  20. Inefficient reprogramming of fibroblasts into cardiomyocytes using Gata4, Mef2c, Tbx5

    Science.gov (United States)

    Chen, J.X.; Krane, M.; Deutsch, M. A.; Wang, L.; Rav-Acha, M.; Gregoire, S.; Engels, M. C.; Rajarajan, K.; Karra, R.; Abel, E. D.; Wu, J. C.; Milan, D.; Wu, S. M.

    2012-01-01

    Rationale Direct reprogramming of fibroblasts into cardiomyocytes is a novel strategy for cardiac regeneration. However, the key determinants involved in this process are unknown. Objective To assess the efficiency of direct fibroblast reprogramming via viral overexpression of GATA4, Mef2c, and Tbx5 (GMT). Methods and Results We induced GMT overexpression in murine tail tip fibroblasts (TTFs) and cardiac fibroblasts (CFs) from multiple lines of transgenic mice carrying different cardiomyocyte lineage reporters. We found that the induction of GMT overexpression in TTFs and CFs is inefficient at inducing molecular and electrophysiological phenotypes of mature cardiomyocytes. In addition, transplantation of GMT infected CFs into injured mouse hearts resulted in decreased cell survival with minimal induction of cardiomyocyte genes. Conclusions Significant challenges remain in our ability to convert fibroblasts into cardiomyocyte-like cells and a greater understanding of cardiovascular epigenetics is needed to increase the translational potential of this strategy. PMID:22581928

  1. Locals Collection

    Directory of Open Access Journals (Sweden)

    Stephen Hastings-King

    2010-03-01

    Full Text Available A locals collection is a set of parameters that are used to delimit data-mining operations. This piece uses a collection of locals from around Essex Massachusetts to shape and delimit an interrogation of post-reality in contemporary America. It explores the notion of crisis, the possibility of a crisis of empire that may or may not emerge in a media-space that does not allow crisis of empire to be mentioned and relations this maybe-crisis to the various levels of economic dysfunction that have become evident since late 2008. But mostly this piece explores ways in which particular stories about particular people do and do not link/link to these larger-scale narratives. This is the first of a potential series of locals collections that will mine the American post-real.

  2. Desarrollo local

    Directory of Open Access Journals (Sweden)

    Luciano Palmitesta

    2001-12-01

    Full Text Available Tras explicar la génesis del concepto de desarrollo y a quien se le aplica la necesidad de desarrollarse, el autor de este ensayo hace un recorrido por las escuelas más representativas de pensamiento sobre el tema. A continuación, entra a reflexionar el concepto de lo local y de sociedad local y sus elementos. En la segunda parte, se analiza la Economía Local y se hace un intento de aplicar esta teoría a la realidad nicaragüense. En esta parte se examinan las diferentes formas empresariales más comunes en Nicaragua. Igualmente, se estudian las dificultades y ventajas de este tipo de empresas en un mundo globalizado y se plantean algunos posibles mecanismos para afrontar los problemas. El trabajo cierra con algunas breves conclusiones.

  3. The Expressions of TGF-β1 and IL-10 in Cultured Fibroblasts after ALA-IPL Photodynamic Treatment

    Science.gov (United States)

    Byun, Ji Yeon; Lee, Ga Youn; Choi, Hae Young; Myung, Ki Bum

    2011-01-01

    Background Topical photodynamic therapy (PDT) with 5-aminolevulinic acid (ALA) was originally used for treating superficial skin tumors. The application of PDT to other inflammatory dermatoses like acne vulgaris, psoriasis, granuloma annulare, localized scleroderma and lichen sclerosus has recently been introduced. However, the underlying mechanisms are not well understood. We've previously reported the induction of tumor growth factor (TGF)-β1 and interleukin (IL)-10 after PDT with ALA and intense pulsed light (IPL) in cultured HaCaT cells. Objective The purpose of this study was to investigate the expressions of TGF-β1 and IL-10 in cultured fibroblasts after PDT with using ALA and IPL. Methods Cultured fibroblasts were treated with ALA-IPL PDT (1µmol/L of ALA; 0, 4, 8 and 12 J/cm2 of IPL). The expressions of TGF-β1 and IL-10 were investigated by reverse transcription-polymerase chain reaction and enzyme linked immunosorbent assay. Results TGF-β1 mRNA and protein were reduced down to 0.52- and 0.63-fold, respectively, after PDT and the IL-10 protein was increased up to 2.74-fold after PDT. Conclusion The reduction of TGF-β1 was prominent after PDT and so an antisclerotic effect can be expected after PDT. The induction of IL-10 may contribute to the anti-inflammatory effect, which explains the therapeutic benefit of PDT for inflammatory dermatoses. PMID:21738358

  4. The Expressions of TGF-β(1) and IL-10 in Cultured Fibroblasts after ALA-IPL Photodynamic Treatment.

    Science.gov (United States)

    Byun, Ji Yeon; Lee, Ga Youn; Choi, Hae Young; Myung, Ki Bum; Choi, You Won

    2011-02-01

    Topical photodynamic therapy (PDT) with 5-aminolevulinic acid (ALA) was originally used for treating superficial skin tumors. The application of PDT to other inflammatory dermatoses like acne vulgaris, psoriasis, granuloma annulare, localized scleroderma and lichen sclerosus has recently been introduced. However, the underlying mechanisms are not well understood. We've previously reported the induction of tumor growth factor (TGF)-β(1) and interleukin (IL)-10 after PDT with ALA and intense pulsed light (IPL) in cultured HaCaT cells. The purpose of this study was to investigate the expressions of TGF-β(1) and IL-10 in cultured fibroblasts after PDT with using ALA and IPL. Cultured fibroblasts were treated with ALA-IPL PDT (1µmol/L of ALA; 0, 4, 8 and 12 J/cm(2) of IPL). The expressions of TGF-β(1) and IL-10 were investigated by reverse transcription-polymerase chain reaction and enzyme linked immunosorbent assay. TGF-β(1) mRNA and protein were reduced down to 0.52- and 0.63-fold, respectively, after PDT and the IL-10 protein was increased up to 2.74-fold after PDT. The reduction of TGF-β(1) was prominent after PDT and so an antisclerotic effect can be expected after PDT. The induction of IL-10 may contribute to the anti-inflammatory effect, which explains the therapeutic benefit of PDT for inflammatory dermatoses.

  5. Epigenetic and phenotypic profile of fibroblasts derived from induced pluripotent stem cells.

    Directory of Open Access Journals (Sweden)

    Kyle J Hewitt

    2011-02-01

    Full Text Available Human induced pluripotent stem (hiPS cells offer a novel source of patient-specific cells for regenerative medicine. However, the biological potential of iPS-derived cells and their similarities to cells differentiated from human embryonic stem (hES cells remain unclear. We derived fibroblast-like cells from two hiPS cell lines and show that their phenotypic properties and patterns of DNA methylation were similar to that of mature fibroblasts and to fibroblasts derived from hES cells. iPS-derived fibroblasts (iPDK and their hES-derived counterparts (EDK showed similar cell morphology throughout differentiation, and patterns of gene expression and cell surface markers were characteristic of mature fibroblasts. Array-based methylation analysis was performed for EDK, iPDK and their parental hES and iPS cell lines, and hierarchical clustering revealed that EDK and iPDK had closely-related methylation profiles. DNA methylation analysis of promoter regions associated with extracellular matrix (ECM-production (COL1A1 by iPS- and hESC-derived fibroblasts and fibroblast lineage commitment (PDGFRβ, revealed promoter demethylation linked to their expression, and patterns of transcription and methylation of genes related to the functional properties of mature stromal cells were seen in both hiPS- and hES-derived fibroblasts. iPDK cells also showed functional properties analogous to those of hES-derived and mature fibroblasts, as seen by their capacity to direct the morphogenesis of engineered human skin equivalents. Characterization of the functional behavior of ES- and iPS-derived fibroblasts in engineered 3D tissues demonstrates the utility of this tissue platform to predict the capacity of iPS-derived cells before their therapeutic application.

  6. Epigenetic and phenotypic profile of fibroblasts derived from induced pluripotent stem cells.

    Science.gov (United States)

    Hewitt, Kyle J; Shamis, Yulia; Hayman, Ryan B; Margvelashvili, Mariam; Dong, Shumin; Carlson, Mark W; Garlick, Jonathan A

    2011-02-28

    Human induced pluripotent stem (hiPS) cells offer a novel source of patient-specific cells for regenerative medicine. However, the biological potential of iPS-derived cells and their similarities to cells differentiated from human embryonic stem (hES) cells remain unclear. We derived fibroblast-like cells from two hiPS cell lines and show that their phenotypic properties and patterns of DNA methylation were similar to that of mature fibroblasts and to fibroblasts derived from hES cells. iPS-derived fibroblasts (iPDK) and their hES-derived counterparts (EDK) showed similar cell morphology throughout differentiation, and patterns of gene expression and cell surface markers were characteristic of mature fibroblasts. Array-based methylation analysis was performed for EDK, iPDK and their parental hES and iPS cell lines, and hierarchical clustering revealed that EDK and iPDK had closely-related methylation profiles. DNA methylation analysis of promoter regions associated with extracellular matrix (ECM)-production (COL1A1) by iPS- and hESC-derived fibroblasts and fibroblast lineage commitment (PDGFRβ), revealed promoter demethylation linked to their expression, and patterns of transcription and methylation of genes related to the functional properties of mature stromal cells were seen in both hiPS- and hES-derived fibroblasts. iPDK cells also showed functional properties analogous to those of hES-derived and mature fibroblasts, as seen by their capacity to direct the morphogenesis of engineered human skin equivalents. Characterization of the functional behavior of ES- and iPS-derived fibroblasts in engineered 3D tissues demonstrates the utility of this tissue platform to predict the capacity of iPS-derived cells before their therapeutic application.

  7. Modulation of the effects of alveolar macrophages on lung fibroblast collagen production rate

    Energy Technology Data Exchange (ETDEWEB)

    Clark, J.G.; Greenberg, J.

    1987-01-01

    Alveolar macrophages (AM) may function as effector cells that can either stimulate or inhibit lung fibroblast collagen production. However, conditions that determine the predominant effect of AM on fibroblasts are not well understood. To delineate factors that modulate the effects of AM on lung fibroblasts, we studied the interaction of AM products and fibroblasts in vitro. The AM were obtained by bronchoalveolar lavage of hamsters with bleomycin-induced pulmonary fibrosis. Conditioned medium (CM) from the AM cultures was incubated in varying amounts with lung fibroblast (IMR-90) cultures. After metabolic labeling with (/sup 3/H)proline, fibroblast collagen production based on procollagen-specific radioactivity was determined. Macrophage CM in concentrations greater than 5% suppressed collagen production, an event attributed to the macrophage-derived suppressive factor that we have previously characterized. Macrophages were also determined to produce PGE2 in culture. Authentic PGE2 at concentrations found in CM was found to suppress fibroblast collagen production, indicating that AM-derived PGE2 contributes to the suppressive activity in CM. To examine possible stimulatory factors in CM, the fibroblasts were preincubated with indomethacin. This approach was based on our previous observation that AM-derived suppressive factor increases endogenous fibroblast PGE2 and that its activity can be blocked by indomethacin. Macrophage CM in a concentration of 20% did not suppress the collagen production of indomethacin-treated fibroblasts. However, CM concentrations of 5 and 10% increased collagen production (173 and 143% of control values, respectively), indicating the presence of stimulatory factor(s) in macrophage-conditioned medium.

  8. Connexin43 Controls the Myofibroblastic Differentiation of Bronchial Fibroblasts from Patients with Asthma.

    Science.gov (United States)

    Paw, Milena; Borek, Izabela; Wnuk, Dawid; Ryszawy, Damian; Piwowarczyk, Katarzyna; Kmiotek, Katarzyna; Wójcik-Pszczoła, Katarzyna A; Pierzchalska, Małgorzata; Madeja, Zbigniew; Sanak, Marek; Błyszczuk, Przemysław; Michalik, Marta; Czyż, Jarosław

    2017-07-01

    Pathologic accumulation of myofibroblasts in asthmatic bronchi is regulated by extrinsic stimuli and by the intrinsic susceptibility of bronchial fibroblasts to transforming growth factor-β (TGF-β). The specific function of gap junctions and connexins in this process has remained unknown. Here, we investigated the role of connexin43 (Cx43) in TGF-β-induced myofibroblastic differentiation of fibroblasts derived from bronchoscopic biopsy specimens of patients with asthma and donors without asthma. Asthmatic fibroblasts expressed considerably higher levels of Cx43 and were more susceptible to TGF-β1-induced myofibroblastic differentiation than were their nonasthmatic counterparts. TGF-β1 efficiently up-regulated Cx43 levels and activated the canonical Smad pathway in asthmatic cells. Ectopic Cx43 expression in nonasthmatic (Cx43low) fibroblasts increased their predilection to TGF-β1-induced Smad2 activation and fibroblast-myofibroblast transition. Transient Cx43 silencing in asthmatic (Cx43high) fibroblasts by Cx43 small interfering RNA attenuated the TGF-β1-triggered Smad2 activation and myofibroblast formation. Direct interactions of Smad2 and Cx43 with β-tubulin were demonstrated by co-immunoprecipitation assay, whereas the sensitivity of these interactions to TGF-β1 signaling was confirmed by Förster Resonance Energy Transfer analyses. Furthermore, inhibition of the TGF-β1/Smad pathway attenuated TGF-β1-triggered Cx43 up-regulation and myofibroblast differentiation of asthmatic fibroblasts. Chemical inhibition of gap junctional intercellular communication with 18 α-glycyrrhetinic acid did not affect the initiation of fibroblast-myofibroblast transition in asthmatic fibroblasts but interfered with the maintenance of their myofibroblastic phenotype. Collectively, our data identified Cx43 as a new player in the feedback mechanism regulating TGF-β1/Smad-dependent differentiation of bronchial fibroblasts. Thus, our observations point to Cx43 as a novel

  9. CTRP6 inhibits fibrogenesis in TGF-β1-stimulated human dermal fibroblasts

    Energy Technology Data Exchange (ETDEWEB)

    Fan, Rong-hui, E-mail: fan_ronghuixa@163.com [Department of Burn and Plastic Surgery, Shaanxi Provincial People’s Hospital, Xi’an 710068 (China); Zhu, Xiu-mei; Sun, Yao-wen [Department of Burn and Plastic Surgery, Shaanxi Provincial People’s Hospital, Xi’an 710068 (China); Peng, Hui-zi [Department of Cosmetology Plastic Surgery, The First Affiliated Hospital of Xi’an Jiaotong University, Xi’an, Shaanxi 710061 (China); Wu, Hang-li; Gao, Wen-jie [Department of Burn and Plastic Surgery, Shaanxi Provincial People’s Hospital, Xi’an 710068 (China)

    2016-07-08

    Skin fibrosis is characterized by excessive proliferation of fibroblasts and overproduction of extracellular matrix (ECM). C1q/tumor necrosis factor-related protein 6 (CTRP6), a member of CTRPs, has been involved in the development of cardiac fibrosis. However, the function and detailed regulatory mechanism of CTRP6 in skin fibrosis remain unclear. The aim of this study was to investigate the effect of CTRP6 on the activation of human dermal fibroblasts. Our results showed that CTRP6 was lowly expressed in scar tissues and transforming growth factor-β1 (TGF-β1)-treated dermal fibroblasts. CTRP6 overexpression significantly inhibited the proliferation of dermal fibroblasts, as well as suppressed the expression of ECM in TGF-β1-treated dermal fibroblasts. Furthermore, CTRP6 overexpression markedly inhibited TGF-β1-induced phosphorylation of Smad3 in dermal fibroblasts. In conclusion, the data reported here demonstrate that CTRP6 is able to inhibit the proliferation and ECM expression in human dermal fibroblasts through suppressing the TGF-β1/Smad3 signaling pathway. These findings suggest that CTRP6 may be a potential therapeutic target for the prevention of skin fibrosis. -- Highlights: •CTRP6 expression was decreased in scar tissues and TGF-β1-treated dermal fibroblasts. •CTRP6 inhibits TGF-β1-induced the proliferation of dermal fibroblasts. •CTRP6 inhibits expression of collagen type I and α-SMA. •CTRP6 inhibits the activation of TGF-β1/Smad3 signaling pathway in dermal fibroblasts.

  10. Identification of a transitional fibroblast function in very early rheumatoid arthritis

    Science.gov (United States)

    Filer, Andrew; Ward, Lewis S C; Kemble, Samuel; Davies, Christopher S; Munir, Hafsa; Rogers, Rebekah; Raza, Karim; Buckley, Christopher Dominic; Nash, Gerard B; McGettrick, Helen M

    2017-01-01

    Objectives Synovial fibroblasts actively regulate the inflammatory infiltrate by communicating with neighbouring endothelial cells (EC). Surprisingly, little is known about how the development of rheumatoid arthritis (RA) alters these immunomodulatory properties. We examined the effects of phase of RA and disease outcome (resolving vs persistence) on fibroblast crosstalk with EC and regulation of lymphocyte recruitment. Methods Fibroblasts were isolated from patients without synovitis, with resolving arthritis, very early RA (VeRA; symptom ≤12 weeks) and established RA undergoing joint replacement (JRep) surgery. Endothelial-fibroblast cocultures were formed on opposite sides of porous filters. Lymphocyte adhesion from flow, secretion of soluble mediators and interleukin 6 (IL-6) signalling were assessed. Results Fibroblasts from non-inflamed and resolving arthritis were immunosuppressive, inhibiting lymphocyte recruitment to cytokine-treated endothelium. This effect was lost very early in the development of RA, such that fibroblasts no longer suppressed recruitment. Changes in IL-6 and transforming growth factor beta 1 (TGF-β1) signalling appeared critical for the loss of the immunosuppressive phenotype. In the absence of exogenous cytokines, JRep, but not VeRA, fibroblasts activated endothelium to support lymphocyte. Conclusions In RA, fibroblasts undergo two distinct changes in function: first a loss of immunosuppressive responses early in disease development, followed by the later acquisition of a stimulatory phenotype. Fibroblasts exhibit a transitional functional phenotype during the first 3 months of symptoms that contributes to the accumulation of persistent infiltrates. Finally, the role of IL-6 and TGF-β1 changes from immunosuppressive in resolving arthritis to stimulatory very early in the development of RA. Early interventions targeting ‘pathogenic’ fibroblasts may be required in order to restore protective regulatory processes. PMID:28847766

  11. Increased fibroblast telomerase expression precedes myofibroblast α-smooth muscle actin expression in idiopathic pulmonary fibrosis

    Directory of Open Access Journals (Sweden)

    Daniel Reis Waisberg

    2012-09-01

    Full Text Available OBJECTIVE: This study sought to identify the relationship between fibroblast telomerase expression, myofibroblasts, and telomerase-mediated regulatory signals in idiopathic pulmonary fibrosis. METHODS: Thirty-four surgical lung biopsies, which had been obtained from patients with idiopathic pulmonary fibrosis and histologically classified as usual interstitial pneumonia, were examined. Immunohistochemistry was used to evaluate fibroblast telomerase expression, myofibroblast α-smooth muscle actin expression and the tissue expression of inter leu kin-4, transforming growth factor-β, and basic fibroblast growth factor. The point-counting technique was used to quantify the expression of these markers in unaffected, collapsed, mural fibrosis, and honeycombing areas. The results were correlated to patient survival. RESULTS: Fibroblast telomerase expression and basic fibroblast growth factor tissue expression were higher in collapsed areas, whereas myofibroblast expression and interleukine-4 tissue expression were higher in areas of mural fibrosis. Transforming growth factor-β expression was higher in collapsed, mural fibrosis and honeycombing areas in comparison to unaffected areas. Positive correlations were found between basic fibroblast growth factor tissue expression and fibroblast telomerase expression and between interleukin-4 tissue expression and myofibroblast α-smooth muscle actin expression. Negative correlations were observed between interleukin-4 expression and basic fibroblast growth factor tissue expression in areas of mural fibrosis. Myofibroblast α-smooth muscle actin expression and interleukin-4 tissue expression in areas of mural fibrosis were negatively associated with patient survival. CONCLUSION: Fibroblast telomerase expression is higher in areas of early remodeling in lung tissues demonstrating typical interstitial pneumonia, whereas myofibroblast α-smooth muscle actin expression predominates in areas of late remodeling

  12. Stretch-Induced Mitogen-Activated Protein Kinase Activation in Lung Fibroblasts Is Independent of Receptor Tyrosine Kinases

    OpenAIRE

    Boudreault, Francis; Tschumperlin, Daniel J.

    2009-01-01

    Lung growth and remodeling are modulated by mechanical stress, with fibroblasts thought to play a leading role. Little mechanistic information is available about how lung fibroblasts respond to mechanical stress. We exposed cultured lung fibroblasts to tonic stretch and measured changes in phosphorylation status of mitogen-activated protein kinases (MAPKs), selected receptor tyrosine kinases (RTKs), and phospholipase Cγ1 (PLCγ1) and activation of the small G-protein Ras. Human lung fibroblast...

  13. Tattoo ink nanoparticles in skin tissue and fibroblasts.

    Science.gov (United States)

    Grant, Colin A; Twigg, Peter C; Baker, Richard; Tobin, Desmond J

    2015-01-01

    Tattooing has long been practised in various societies all around the world and is becoming increasingly common and widespread in the West. Tattoo ink suspensions unquestionably contain pigments composed of nanoparticles, i.e., particles of sub-100 nm dimensions. It is widely acknowledged that nanoparticles have higher levels of chemical activity than their larger particle equivalents. However, assessment of the toxicity of tattoo inks has been the subject of little research and ink manufacturers are not obliged to disclose the exact composition of their products. This study examines tattoo ink particles in two fundamental skin components at the nanometre level. We use atomic force microscopy and light microscopy to examine cryosections of tattooed skin, exploring the collagen fibril networks in the dermis that contain ink nanoparticles. Further, we culture fibroblasts in diluted tattoo ink to explore both the immediate impact of ink pigment on cell viability and also to observe the interaction between particles and the cells.

  14. Tattoo ink nanoparticles in skin tissue and fibroblasts

    Directory of Open Access Journals (Sweden)

    Colin A. Grant

    2015-05-01

    Full Text Available Tattooing has long been practised in various societies all around the world and is becoming increasingly common and widespread in the West. Tattoo ink suspensions unquestionably contain pigments composed of nanoparticles, i.e., particles of sub-100 nm dimensions. It is widely acknowledged that nanoparticles have higher levels of chemical activity than their larger particle equivalents. However, assessment of the toxicity of tattoo inks has been the subject of little research and ink manufacturers are not obliged to disclose the exact composition of their products. This study examines tattoo ink particles in two fundamental skin components at the nanometre level. We use atomic force microscopy and light microscopy to examine cryosections of tattooed skin, exploring the collagen fibril networks in the dermis that contain ink nanoparticles. Further, we culture fibroblasts in diluted tattoo ink to explore both the immediate impact of ink pigment on cell viability and also to observe the interaction between particles and the cells.

  15. PDGFRaa Signaling Is Regulated through the Primary Cilium in Fibroblasts

    DEFF Research Database (Denmark)

    Schneider, Linda; Clement, Christian Alexandro; Teilmann, S.C.

    2005-01-01

    Recent findings show that cilia are sensory organelles that display specific receptors and ion channels, which transmit signals from the extracellular environment via the cilium to the cell to control tissue homeostasis and function [ [1] , [2] , [3] , [4] , [5] and [6] ]. Agenesis of primary cilia...... or mislocation of ciliary signal components affects human pathologies, such as polycystic kidney disease [ 7 ] and disorders associated with Bardet-Biedl syndrome [ 8 ]. Primary cilia are essential for hedgehog ligand-induced signaling cascade regulating growth and patterning [ [9] and [10] ]. Here, we show...... is followed by activation of Akt and the Mek1/2-Erk1/2 pathways, with Mek1/2 being phosphorylated within the cilium and at the basal body. Fibroblasts derived from Tg737orpk mutants fail to form normal cilia and to upregulate the level of PDGFRa; PDGF-AA fails to activate PDGFRaa and the Mek1/2-Erk1/2 pathway...

  16. Fibroblast Growth Factor 23 in Long-Duration Spaceflight

    Science.gov (United States)

    Bokhari, R.; Zwart, S. R.; Fields, E.; Heer, M.; Sibonga, J.; Smith, S. M.

    2015-01-01

    Many nutritional factors influence bone, from the basics of calcium and vitamin D, to factors which influence bone through acid/base balance, including protein, sodium, and more. Fibroblast growth factor 23 (FGF23) is a recently identified factor, secreted from osteocytes, which is involved in classic (albeit complex) feedback loops controlling phosphorus homeostasis through both vitamin D and parathyroid hormone (PTH) (1, 2). As osteocytes are gravity sensing cells, it is important to determine if there are changes in FGF23 during spaceflight. In extreme cases, such as chronic kidney disease, FGF23 levels are highly elevated. FGF23 imbalances, secondary to dietary influences, may contribute to skeletal demineralization and kidney stone risk during spaceflight.

  17. Collagen matrix as a tool in studying fibroblastic cell behavior

    Science.gov (United States)

    Kanta, Jiří

    2015-01-01

    Type I collagen is a fibrillar protein, a member of a large family of collagen proteins. It is present in most body tissues, usually in combination with other collagens and other components of extracellular matrix. Its synthesis is increased in various pathological situations, in healing wounds, in fibrotic tissues and in many tumors. After extraction from collagen-rich tissues it is widely used in studies of cell behavior, especially those of fibroblasts and myofibroblasts. Cells cultured in a classical way, on planar plastic dishes, lack the third dimension that is characteristic of body tissues. Collagen I forms gel at neutral pH and may become a basis of a 3D matrix that better mimics conditions in tissue than plastic dishes. PMID:25734486

  18. Fibroblast activation protein (FAP) as a novel metabolic target

    DEFF Research Database (Denmark)

    Sánchez-Garrido, Miguel Angel; Habegger, Kirk M; Clemmensen, Christoffer

    2016-01-01

    to block FAP enzymatic activity. RESULTS: TB administration to diet-induced obese (DIO) animals led to profound decreases in body weight, reduced food consumption and adiposity, increased energy expenditure, improved glucose tolerance and insulin sensitivity, and lowered cholesterol levels. Total...... on body weight or any other measures of metabolism. In support of these results we observed no enzymatic degradation of human FGF21 at either end of the protein when FAP was inhibited in vitro by TB. CONCLUSIONS: We conclude that pharmacological inhibition of FAP enhances levels of FGF21 in obese mice......OBJECTIVE: Fibroblast activation protein (FAP) is a serine protease belonging to a S9B prolyl oligopeptidase subfamily. This enzyme has been implicated in cancer development and recently reported to regulate degradation of FGF21, a potent metabolic hormone. Using a known FAP inhibitor, talabostat...

  19. Fibroblast Growth Factors: Biology, Function, and Application for Tissue Regeneration

    Directory of Open Access Journals (Sweden)

    Ye-Rang Yun

    2010-01-01

    Full Text Available Fibroblast growth factors (FGFs that signal through FGF receptors (FGFRs regulate a broad spectrum of biological functions, including cellular proliferation, survival, migration, and differentiation. The FGF signal pathways are the RAS/MAP kinase pathway, PI3 kinase/AKT pathway, and PLCγ pathway, among which the RAS/MAP kinase pathway is known to be predominant. Several studies have recently implicated the in vitro biological functions of FGFs for tissue regeneration. However, to obtain optimal outcomes in vivo, it is important to enhance the half-life of FGFs and their biological stability. Future applications of FGFs are expected when the biological functions of FGFs are potentiated through the appropriate use of delivery systems and scaffolds. This review will introduce the biology and cellular functions of FGFs and deal with the biomaterials based delivery systems and their current applications for the regeneration of tissues, including skin, blood vessel, muscle, adipose, tendon/ligament, cartilage, bone, tooth, and nerve tissues.

  20. [Fibroblast growth factors and their effects in pancreas organogenesis].

    Science.gov (United States)

    Gnatenko, D A; Kopantzev, E P; Sverdlov, E D

    2017-05-01

    Fibroblast growth factors (FGF) - growth factors that regulate many important biological processes, including proliferation and differentiation of embryonic cells during organogenesis. In this review, we will summarize current information about the involvement of FGFs in the pancreas organogenesis. Pancreas organogenesis is a complex process, which involves constant signaling from mesenchymal tissue. This orchestrates the activation of various regulator genes at specific stages, determining the specification of progenitor cells. Alterations in FGF/FGFR signaling pathway during this process lead to incorrect activation of the master genes, which leads to different pathologies during pancreas development. Understanding the full picture about role of FGF factors in pancreas development will make it possible to more accurately understand their role in other pathologies of this organ, including carcinogenesis.

  1. File list: InP.PSC.05.AllAg.hESC_derived_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available InP.PSC.05.AllAg.hESC_derived_fibroblasts hg19 Input control Pluripotent stem cell hESC derived... fibroblasts http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/InP.PSC.05.AllAg.hESC_derived_fibroblasts.bed ...

  2. File list: DNS.PSC.10.AllAg.iPS_derived_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  3. File list: NoD.PSC.05.AllAg.iPS_derived_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  4. File list: Pol.PSC.20.AllAg.hESC_derived_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Pol.PSC.20.AllAg.hESC_derived_fibroblasts hg19 RNA polymerase Pluripotent stem cell hESC derived... fibroblasts http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/Pol.PSC.20.AllAg.hESC_derived_fibroblasts.bed ...

  5. File list: Unc.PSC.05.AllAg.hESC_derived_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Unc.PSC.05.AllAg.hESC_derived_fibroblasts hg19 Unclassified Pluripotent stem cell hESC derived... fibroblasts http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/Unc.PSC.05.AllAg.hESC_derived_fibroblasts.bed ...

  6. File list: DNS.PSC.05.AllAg.hESC_derived_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available DNS.PSC.05.AllAg.hESC_derived_fibroblasts hg19 DNase-seq Pluripotent stem cell hESC derived... fibroblasts http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/DNS.PSC.05.AllAg.hESC_derived_fibroblasts.bed ...

  7. File list: Pol.PSC.20.AllAg.iPS_derived_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Pol.PSC.20.AllAg.iPS_derived_fibroblasts hg19 RNA polymerase Pluripotent stem cell iPS derived... fibroblasts http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/Pol.PSC.20.AllAg.iPS_derived_fibroblasts.bed ...

  8. File list: DNS.PSC.05.AllAg.iPS_derived_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available DNS.PSC.05.AllAg.iPS_derived_fibroblasts hg19 DNase-seq Pluripotent stem cell iPS derived... fibroblasts http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/DNS.PSC.05.AllAg.iPS_derived_fibroblasts.bed ...

  9. File list: Pol.PSC.10.AllAg.hESC_derived_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Pol.PSC.10.AllAg.hESC_derived_fibroblasts hg19 RNA polymerase Pluripotent stem cell hESC derived... fibroblasts http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/Pol.PSC.10.AllAg.hESC_derived_fibroblasts.bed ...

  10. File list: Oth.PSC.20.AllAg.hESC_derived_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Oth.PSC.20.AllAg.hESC_derived_fibroblasts hg19 TFs and others Pluripotent stem cell hESC derived... fibroblasts http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/Oth.PSC.20.AllAg.hESC_derived_fibroblasts.bed ...

  11. File list: NoD.PSC.50.AllAg.hESC_derived_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available NoD.PSC.50.AllAg.hESC_derived_fibroblasts hg19 No description Pluripotent stem cell hESC derived... fibroblasts http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/NoD.PSC.50.AllAg.hESC_derived_fibroblasts.bed ...

  12. File list: Pol.PSC.50.AllAg.iPS_derived_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Pol.PSC.50.AllAg.iPS_derived_fibroblasts hg19 RNA polymerase Pluripotent stem cell iPS derived... fibroblasts http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/Pol.PSC.50.AllAg.iPS_derived_fibroblasts.bed ...

  13. File list: InP.Oth.20.AllAg.3T3_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available InP.Oth.20.AllAg.3T3_fibroblasts mm9 Input control Others 3T3 fibroblasts SRX099261...,SRX099262 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/InP.Oth.20.AllAg.3T3_fibroblasts.bed ...

  14. File list: His.PSC.10.AllAg.iPS_derived_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available His.PSC.10.AllAg.iPS_derived_fibroblasts hg19 Histone Pluripotent stem cell iPS der...ived fibroblasts http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/His.PSC.10.AllAg.iPS_derived_fibroblasts.bed ...

  15. File list: InP.PSC.05.AllAg.iPS_derived_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available InP.PSC.05.AllAg.iPS_derived_fibroblasts hg19 Input control Pluripotent stem cell iPS... derived fibroblasts http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/InP.PSC.05.AllAg.iPS_derived_fibroblasts.bed ...

  16. File list: Pol.PSC.10.AllAg.iPS_derived_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Pol.PSC.10.AllAg.iPS_derived_fibroblasts hg19 RNA polymerase Pluripotent stem cell iPS... derived fibroblasts http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/Pol.PSC.10.AllAg.iPS_derived_fibroblasts.bed ...

  17. File list: Oth.PSC.20.AllAg.iPS_derived_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Oth.PSC.20.AllAg.iPS_derived_fibroblasts hg19 TFs and others Pluripotent stem cell iPS... derived fibroblasts http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/Oth.PSC.20.AllAg.iPS_derived_fibroblasts.bed ...

  18. File list: Oth.PSC.10.AllAg.iPS_derived_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Oth.PSC.10.AllAg.iPS_derived_fibroblasts hg19 TFs and others Pluripotent stem cell iPS... derived fibroblasts http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/Oth.PSC.10.AllAg.iPS_derived_fibroblasts.bed ...

  19. File list: InP.PSC.10.AllAg.iPS_derived_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available InP.PSC.10.AllAg.iPS_derived_fibroblasts hg19 Input control Pluripotent stem cell iPS... derived fibroblasts http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/InP.PSC.10.AllAg.iPS_derived_fibroblasts.bed ...

  20. File list: His.PSC.05.AllAg.iPS_derived_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available His.PSC.05.AllAg.iPS_derived_fibroblasts hg19 Histone Pluripotent stem cell iPS der...ived fibroblasts http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/His.PSC.05.AllAg.iPS_derived_fibroblasts.bed ...

  1. File list: His.PSC.20.AllAg.iPS_derived_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available His.PSC.20.AllAg.iPS_derived_fibroblasts hg19 Histone Pluripotent stem cell iPS der...ived fibroblasts http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/His.PSC.20.AllAg.iPS_derived_fibroblasts.bed ...

  2. The initiation of embryonic-like collagen fibrillogenesis by adult human tendon fibroblasts when cultured under tension

    DEFF Research Database (Denmark)

    Bayer, Monika L; Yeung, Chin-Yan C; Kadler, Karl E

    2010-01-01

    Tendon fibroblasts synthesize collagen and form fibrils during embryonic development, but to what extent mature fibroblasts are able to recapitulate embryonic development and develop normal tendon structure is unknown. The present study examined the capability of mature human tendon fibroblasts t...

  3. File list: His.Oth.50.AllAg.3T3_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available His.Oth.50.AllAg.3T3_fibroblasts mm9 Histone Others 3T3 fibroblasts SRX099255,SRX10...5582,SRX099259,SRX099260 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/His.Oth.50.AllAg.3T3_fibroblasts.bed ...

  4. File list: ALL.Oth.20.AllAg.3T3_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available ALL.Oth.20.AllAg.3T3_fibroblasts mm9 All antigens Others 3T3 fibroblasts SRX099255,...SRX105582,SRX099259,SRX099260,SRX099261,SRX099262 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/ALL.Oth.20.AllAg.3T3_fibroblasts.bed ...

  5. File list: ALL.Oth.10.AllAg.3T3_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available ALL.Oth.10.AllAg.3T3_fibroblasts mm9 All antigens Others 3T3 fibroblasts SRX105582,...SRX099255,SRX099259,SRX099260,SRX099261,SRX099262 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/ALL.Oth.10.AllAg.3T3_fibroblasts.bed ...

  6. File list: His.Oth.20.AllAg.3T3_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available His.Oth.20.AllAg.3T3_fibroblasts mm9 Histone Others 3T3 fibroblasts SRX099255,SRX10...5582,SRX099259,SRX099260 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/His.Oth.20.AllAg.3T3_fibroblasts.bed ...

  7. File list: ALL.Oth.50.AllAg.3T3_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available ALL.Oth.50.AllAg.3T3_fibroblasts mm9 All antigens Others 3T3 fibroblasts SRX099255,...SRX105582,SRX099259,SRX099260,SRX099261,SRX099262 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/ALL.Oth.50.AllAg.3T3_fibroblasts.bed ...

  8. SWAP-70 contributes to spontaneous transformation of mouse embryo fibroblasts

    Energy Technology Data Exchange (ETDEWEB)

    Chang, Yu-Tzu; Shu, Chung-Li; Lai, Jing-Yang; Lin, Ching-Yu; Chuu, Chih-Pin [Institute of Cellular and System Medicine National Health Research Institute, Zhunan Town 35053, Miaoli County, Taiwan, ROC (China); Morishita, Kazuhiro; Ichikawa, Tomonaga [Division of Tumor and Cellular Biochemistry Department of Medical Sciences Faculty of Medicine University of Miyazaki, 5200 Kihara, Kiyotake, Miyazaki-shi, Miyazaki 889-1692 Japan (Japan); Jessberger, Rolf [Faculty of Medicine Carl Gustav Carus, Institute of Physiological Chemistry, Dresden University of Technology, Dresden (Germany); Fukui, Yasuhisa, E-mail: 990412@nhri.org.tw [Institute of Cellular and System Medicine National Health Research Institute, Zhunan Town 35053, Miaoli County, Taiwan, ROC (China)

    2016-07-15

    Mouse embryo fibroblasts (MEFs) grow slowly after cultivation from animals, however, after an extended period of cultivation, their growth accelerates. We found that SWAP-70 deficient MEFs failed to increase growth rates. They maintain normal growth rates and proliferation cycles for at least 5 years. Complementing SWAP-70 deficiency in one of these MEF clones, MEF1F2, by expressing human SWAP-70 resulted in fast growth of the cells after further cultivation for a long period. The resulting cells show a transformation phenotype, since they grow on top of each other and do not show contact inhibition. This phenotype was reverted when sanguinarine, a putative SWAP-70 inhibitor, was added. Two SWAP-70 expressing clones were examined in detail. Even after cell density became very high their cdc2 and NFκB were still activated suggesting that they do not stop growing. One of the clones formed colonies in soft agar and formed tumors in nude mice. Lately, one more clone became transformed being able to make colonies in soft agar. We maintain 4 human SWAP-70 expressing MEF1F2 cell lines. Three out of 4 clones exhibited transforming phenotypes. The mouse SWAP-70 gene also promoted transformation of MEFs. Taken together our data suggest that SWAP-70 is not a typical oncogene, but is required for spontaneous transformation of MEFs. - Highlights: • Mouse embryo fibroblasts (MEFs) lacking SWAP-70 do not cause spontaneous transform. • Adding back of SWAP-70 to SWAP-70-deficient MEFs induces spontaneous transformation. • SWAP-70 is required for spontaneous transformation of MEFs.

  9. Tumor-produced, active Interleukin-1 {beta} regulates gene expression in carcinoma-associated fibroblasts

    Energy Technology Data Exchange (ETDEWEB)

    Dudas, Jozsef, E-mail: Jozsef.Dudas@i-med.ac.at [Department of Otorhinolaryngology, Medical University Innsbruck, Anichstrasse 35, A-6020 Innsbruck (Austria); Fullar, Alexandra, E-mail: fullarsz@gmail.com [Department of Otorhinolaryngology, Medical University Innsbruck, Anichstrasse 35, A-6020 Innsbruck (Austria); 1st Institute of Pathology and Experimental Cancer Research, Semmelweis University, Ulloei ut 26, H-1085 Budapest (Hungary); Bitsche, Mario, E-mail: Mario.Bitsche@i-med.ac.at [Department of Otorhinolaryngology, Medical University Innsbruck, Anichstrasse 35, A-6020 Innsbruck (Austria); Schartinger, Volker, E-mail: Volker.Schartinger@i-med.ac.at [Department of Otorhinolaryngology, Medical University Innsbruck, Anichstrasse 35, A-6020 Innsbruck (Austria); Kovalszky, Ilona, E-mail: koval@korb1.sote.hu [1st Institute of Pathology and Experimental Cancer Research, Semmelweis University, Ulloei ut 26, H-1085 Budapest (Hungary); Sprinzl, Georg Mathias, E-mail: Georg.Sprinzl@i-med.ac.at [Department of Otorhinolaryngology, Medical University Innsbruck, Anichstrasse 35, A-6020 Innsbruck (Austria); Riechelmann, Herbert, E-mail: Herbert.Riechelmann@i-med.ac.at [Department of Otorhinolaryngology, Medical University Innsbruck, Anichstrasse 35, A-6020 Innsbruck (Austria)

    2011-09-10

    Recently we described a co-culture model of periodontal ligament (PDL) fibroblasts and SCC-25 lingual squamous carcinoma cells, which resulted in conversion of normal fibroblasts into carcinoma-associated fibroblasts (CAFs), and in epithelial-mesenchymal transition (EMT) of SCC-25 cells. We have found a constitutive high interleukin-1{beta} (IL1-{beta}) expression in SCC-25 cells in normal and in co-cultured conditions. In our hypothesis a constitutive IL1-{beta} expression in SCC-25 regulates gene expression in fibroblasts during co-culture. Co-cultures were performed between PDL fibroblasts and SCC-25 cells with and without dexamethasone (DEX) treatment; IL1-{beta} processing was investigated in SCC-25 cells, tumor cells and PDL fibroblasts were treated with IL1-{beta}. IL1-{beta} signaling was investigated by western blot and immunocytochemistry. IL1-{beta}-regulated genes were analyzed by real-time qPCR. SCC-25 cells produced 16 kD active IL1-{beta}, its receptor was upregulated in PDL fibroblasts during co-culture, which induced phosphorylation of interleukin-1 receptor-associated kinase-1 (IRAK-1), and nuclear translocalization of NF{kappa}B{alpha}. Several genes, including interferon regulatory factor 1 (IRF1) interleukin-6 (IL-6) and prostaglandin-endoperoxide synthase 2 (COX-2) were induced in CAFs during co-culture. The most enhanced induction was found for IL-6 and COX-2. Treatment of PDL fibroblasts with IL1-{beta} reproduced a time- and dose-dependent upregulation of IL1-receptor, IL-6 and COX-2. A further proof was achieved by DEX inhibition for IL1-{beta}-stimulated IL-6 and COX-2 gene expression. Constitutive expression of IL1-{beta} in the tumor cells leads to IL1-{beta}-stimulated gene expression changes in tumor-associated fibroblasts, which are involved in tumor progression. -- Graphical abstract: SCC-25 cells produce active, processed IL1-{beta}. PDL fibroblasts possess receptor for IL1-{beta}, and its expression is increased 4.56-times in the

  10. LXA{sub 4} actions direct fibroblast function and wound closure

    Energy Technology Data Exchange (ETDEWEB)

    Herrera, Bruno S. [Department of Applied Oral Sciences, Center for Periodontology, The Forsyth Institute, Cambridge, MA (United States); Microbiology Branch, US Army Dental and Trauma Research Detachment, Institute of Surgical Research, JBSA Fort Sam Houston, TX (United States); Kantarci, Alpdogan; Zarrough, Ahmed; Hasturk, Hatice [Department of Applied Oral Sciences, Center for Periodontology, The Forsyth Institute, Cambridge, MA (United States); Leung, Kai P., E-mail: kai.p.leung.civ@mail.mil [Microbiology Branch, US Army Dental and Trauma Research Detachment, Institute of Surgical Research, JBSA Fort Sam Houston, TX (United States); Van Dyke, Thomas E., E-mail: tvandyke@forsyth.org [Department of Applied Oral Sciences, Center for Periodontology, The Forsyth Institute, Cambridge, MA (United States)

    2015-09-04

    Timely resolution of inflammation is crucial for normal wound healing. Resolution of inflammation is an active biological process regulated by specialized lipid mediators including the lipoxins and resolvins. Failure of resolution activity has a major negative impact on wound healing in chronic inflammatory diseases that is manifest as excess fibrosis and scarring. Lipoxins, including Lipoxin A{sub 4} (LXA{sub 4}), have known anti-fibrotic and anti-scarring properties. The goal of this study was to elucidate the impact of LXA{sub 4} on fibroblast function. Mouse fibroblasts (3T3 Mus musculus Swiss) were cultured for 72 h in the presence of TGF-β1, to induce fibroblast activation. The impact of exogenous TGF-β1 (1 ng/mL) on LXA{sub 4} receptor expression (ALX/FPR2) was determined by flow cytometry. Fibroblast proliferation was measured by bromodeoxyuridine (BrdU) labeling and migration in a “scratch” assay wound model. Expression of α-smooth muscle actin (α-SMA), and collagen types I and III were measured by Western blot. We observed that TGF-β1 up-regulates LXA{sub 4} receptor expression, enhances fibroblast proliferation, migration and scratch wound closure. α-SMA levels and Collagen type I and III deposition were also enhanced. LXA{sub 4} slowed fibroblast migration and scratch wound closure at early time points (24 h), but wound closure was equal to TGF-β1 alone at 48 and 72 h. LXA{sub 4} tended to slow fibroblast proliferation at both concentrations, but had no impact on α-SMA or collagen production by TGF-β1 stimulated fibroblasts. The generalizability of the actions of resolution molecules was examined in experiments repeated with resolvin D2 (RvD2) as the agonist. The activity of RvD2 mimicked the actions of LXA{sub 4} in all assays, through an as yet unidentified receptor. The results suggest that mediators of resolution of inflammation enhance wound healing and limit fibrosis in part by modulating fibroblast function. - Highlights: • TGF

  11. C-type natriuretic peptide ameliorates pulmonary fibrosis by acting on lung fibroblasts in mice.

    Science.gov (United States)

    Kimura, Toru; Nojiri, Takashi; Hino, Jun; Hosoda, Hiroshi; Miura, Koichi; Shintani, Yasushi; Inoue, Masayoshi; Zenitani, Masahiro; Takabatake, Hiroyuki; Miyazato, Mikiya; Okumura, Meinoshin; Kangawa, Kenji

    2016-02-19

    Pulmonary fibrosis has high rates of mortality and morbidity; however, no effective pharmacological therapy has been established. C-type natriuretic peptide (CNP), a member of the natriuretic peptide family, selectively binds to the transmembrane guanylyl cyclase (GC)-B receptor and exerts anti-inflammatory and anti-fibrotic effects in various organs through vascular endothelial cells and fibroblasts that have a cell-surface GC-B receptor. Given the pathophysiological importance of fibroblast activation in pulmonary fibrosis, we hypothesized that the anti-fibrotic and anti-inflammatory effects of exogenous CNP against bleomycin (BLM)-induced pulmonary fibrosis were exerted in part by the effect of CNP on pulmonary fibroblasts. C57BL/6 mice were divided into two groups, CNP-treated (2.5 μg/kg/min) and vehicle, to evaluate BLM-induced (1 mg/kg) pulmonary fibrosis and inflammation. A periostin-CNP transgenic mouse model exhibiting CNP overexpression in fibroblasts was generated and examined for the anti-inflammatory and anti-fibrotic effects of CNP via fibroblasts in vivo. Additionally, we assessed CNP attenuation of TGF-β-induced differentiation into myofibroblasts by using immortalized human lung fibroblasts stably expressing GC-B receptors. Furthermore, to investigate whether CNP acts on human lung fibroblasts in a clinical setting, we obtained primary-cultured fibroblasts from surgically resected lungs of patients with lung cancer and analyzed levels of GC-B mRNA transcription. CNP reduced mRNA levels of the profibrotic cytokines interleukin (IL)-1β and IL-6, as well as collagen deposition and the fibrotic area in lungs of mice with bleomycin-induced pulmonary fibrosis. Furthermore, similar CNP effects were observed in transgenic mice exhibiting fibroblast-specific CNP overexpression. In cultured-lung fibroblasts, CNP treatment attenuated TGF-β-induced phosphorylation of Smad2 and increased mRNA and protein expression of α-smooth muscle actin and SM22

  12. Skin-derived fibroblasts for the treatment of refractory Achilles tendinosis: preliminary short-term results.

    Science.gov (United States)

    Obaid, Haron; Clarke, Andrew; Rosenfeld, Peter; Leach, Christopher; Connell, David

    2012-02-01

    Chronic Achilles tendinosis is a common musculoskeletal disorder often refractory to conservative management. Our study aimed to assess the safety and efficacy of the use of autologous skin-derived collagen-producing cells in the treatment of refractory Achilles tendinosis. We conducted a randomized, double-blind study on forty Achilles tendons in thirty-two patients (eight with bilateral involvement) who had a clinical and radiographic diagnosis of Achilles tendinosis. The patients ranged from twenty-two to sixty-seven years old and had a mean age of 45.2 years. The patients with unilateral involvement were randomized into the treatment group (twelve patients) and control group (twelve patients). The eight patients with bilateral involvement were individually randomized into treatment and control groups, with eight Achilles tendons in each group. Achilles tendons in the treatment group were injected under ultrasound guidance with laboratory-expanded, skin-derived fibroblasts suspended in autologous plasma. The control group received ultrasound-guided injection of a local anesthetic and physiotherapy. The Victorian Institute of Sport Assessment (VISA) questionnaire and visual analog scale (VAS) scores were used as the main outcome measures for both groups. Significant differences in the mean VISA and VAS scores were detected between the treatment and the control groups for the patients with unilateral involvement at six months (p tendinosis is safe. However, larger studies with a longer duration of follow-up are required to determine the long-term effectiveness before wider clinical application is considered.

  13. Modulation of the Senescence-Associated Inflammatory Phenotype in Human Fibroblasts by Olive Phenols

    Science.gov (United States)

    Menicacci, Beatrice; Cipriani, Caterina; Margheri, Francesca

    2017-01-01

    Senescent cells display an increase in the secretion of growth factors, inflammatory cytokines and proteolytic enzymes, termed the “senescence-associated-secretory-phenotype” (SASP), playing a major role in many age-related diseases. The phenolic compounds present in extra-virgin olive oil are inhibitors of oxidative damage and have been reported to play a protective role in inflammation-related diseases. Particularly, hydroxytyrosol and oleuropein are the most abundant and more extensively studied. Pre-senescent human lung (MRC5) and neonatal human dermal (NHDF) fibroblasts were used as cellular model to evaluate the effect of chronic (4–6 weeks) treatment with 1 μM hydroxytyrosol (HT) or 10 μM oleuropein aglycone (OLE) on senescence/inflammation markers. Both phenols were effective in reducing β-galactosidase-positive cell number and p16 protein expression. In addition, senescence/inflammation markers such as IL-6 and metalloprotease secretion, and Ciclooxigenase type 2 (COX-2) and α-smooth-actin levels were reduced by phenol treatments. In NHDF, COX-2 expression, Nuclear Factor κ-light-chain-enhancer of activated B cells (NFκB) protein level and nuclear localization were augmented with culture senescence and decreased by OLE and HT treatment. Furthermore, the inflammatory effect of Tumor Necrosis Factor α (TNFα) exposure was almost completely abolished in OLE- and HT-pre-treated NHDF. Thus, the modulation of the senescence-associated inflammatory phenotype might be an important mechanism underlying the beneficial effects of olive oil phenols. PMID:29084133

  14. Heparan sulfate dependent signaling of fibroblast growth factor (FGF) 18 by chondrocyte-derived perlecan

    Science.gov (United States)

    Chuang, Christine Y.; Lord, Megan S.; Melrose, James; Rees, Martin D.; Knox, Sarah M.; Freeman, Craig; Iozzo, Renato V.; Whitelock, John M.

    2010-01-01

    Perlecan is a large multi-domain proteoglycan which is essential for normal cartilage development. In this study perlecan was localized in the pericellular matrix of hypertrophic chondrocytes in developing human cartilage rudiments. Perlecan immunopurified from medium conditioned by cultured human fetal chondrocytes was found to be substituted with heparan sulfate (HS), chondroitin sulfate (CS) and keratan sulfate (KS). Ligand and carbohydrate engagement (LACE) assays demonstrated that immunopurified chondrocyte-derived perlecan formed HS dependent ternary complexes with fibroblast growth factors (FGF) 2 and either FGFR receptors (FGFRs) 1 or 3, however these complexes were not biologically active in the BaF32 cell system. Chondrocyte-derived perlecan also formed HS dependent ternary complexes with FGF18 and FGFR3. The proliferation of BaF32 cells expressing FGFR3 was promoted by chondrocyte-derived perlecan in the presence of FGF18 and this activity was reduced by digesting the HS with either heparinase III or mammalian heparanase. These data suggest that FGF2 and 18 bind to discrete structures on the HS chains attached to chondrocyte-derived perlecan which modulate the growth factor activities. The presence and activity of mammalian heparanase may be important in the turnover of HS and subsequent signaling required for the establishment and maintenance of functional osteo-chondral junctions in long bone growth. PMID:20507176

  15. Reprogramming of human fibroblasts to pluripotent stem cells using mRNA of four transcription factors

    Energy Technology Data Exchange (ETDEWEB)

    Yakubov, Eduard [Department of Molecular Cell Biology, Weizmann Institute of Science, 76100 Rehovot (Israel); Rechavi, Gidi [Cancer Research Center, Chaim Sheba Medical Center, Tel-Hashomer and Sackler School of Medicine, Tel-Aviv University, Tel-Aviv (Israel); Rozenblatt, Shmuel [Department of Molecular Microbiology and Biotechnology, Tel-Aviv University, Tel-Aviv (Israel); Givol, David, E-mail: david.givol@weizmann.ac.il [Department of Molecular Cell Biology, Weizmann Institute of Science, 76100 Rehovot (Israel)

    2010-03-26

    Reprogramming of differentiated cells into induced pluripotent cells (iPS) was accomplished in 2006 by expressing four, or less, embryonic stem cell (ESC)-specific transcription factors. Due to the possible danger of DNA damage and the potential tumorigenicity associated with such DNA damage, attempts were made to minimize DNA integration by the vectors involved in this process without complete success. Here we present a method of using RNA transfection as a tool for reprogramming human fibroblasts to iPS. We used RNA synthesized in vitro from cDNA of the same reprogramming four transcription factors. After transfection of the RNA, we show intracellular expression and nuclear localization of the respective proteins in at least 70% of the cells. We used five consecutive transfections to support continuous protein expression resulting in the formation of iPS colonies that express alkaline phosphatase and several ESC markers and that can be expanded. This method completely avoids DNA integration and may be developed to replace the use of DNA vectors in the formation of iPS.

  16. Upregulation of ABC transporters contributes to chemoresistance of sphingosine 1-phosphate lyase-deficient fibroblasts.

    Science.gov (United States)

    Ihlefeld, Katja; Vienken, Hans; Claas, Ralf Frederik; Blankenbach, Kira; Rudowski, Agnes; ter Braak, Michael; Koch, Alexander; Van Veldhoven, Paul P; Pfeilschifter, Josef; Meyer zu Heringdorf, Dagmar

    2015-01-01

    Sphingosine 1-phosphate (S1P) is an extra- and intracellular mediator that regulates cell growth, survival, migration, and adhesion in many cell types. S1P lyase is the enzyme that irreversibly cleaves S1P and thereby constitutes the ultimate step in sphingolipid catabolism. It has been reported previously that embryonic fibroblasts from S1P lyase-deficient mice (Sgpl1(-/-)-MEFs) are resistant to chemotherapy-induced apoptosis through upregulation of B cell lymphoma 2 (Bcl-2) and Bcl-2-like 1 (Bcl-xL). Here, we demonstrate that the transporter proteins Abcc1/MRP1, Abcb1/MDR1, Abca1, and spinster-2 are upregulated in Sgpl1(-/-)-MEFs. Furthermore, the cells efficiently sequestered the substrates of Abcc1 and Abcb1, fluo-4 and doxorubicin, in subcellular compartments. In line with this, Abcb1 was localized mainly at intracellular vesicular structures. After 16 h of incubation, wild-type MEFs had small apoptotic nuclei containing doxorubicin, whereas the nuclei of Sgpl1(-/-)-MEFs appeared unchanged and free of doxorubicin. A combined treatment with the inhibitors of Abcb1 and Abcc1, zosuquidar and MK571, respectively, reversed the compartmentalization of doxorubicin and rendered the cells sensitive to doxorubicin-induced apoptosis. It is concluded that upregulation of multidrug resistance transporters contributes to the chemoresistance of S1P lyase-deficient MEFs. Copyright © 2015 by the American Society for Biochemistry and Molecular Biology, Inc.

  17. Streptococcus mutans and Streptococcus sobrinus are able to adhere and invade human gingival fibroblast cell line.

    Science.gov (United States)

    Berlutti, F; Catizone, A; Ricci, G; Frioni, A; Natalizi, T; Valenti, P; Polimeni, A

    2010-01-01

    Streptococcus mutans and Streptococcus sobrinus, the principal etiologic agents of caries decay of teeth, are generally acquired in oral cavity at the moment of tooth eruption. However, as S. mutans has been detected in oral cavity of predentate children, the eruption of teeth seems not to be a necessary prerequisite, suggesting that this species may be not confined to dental plaque. Here, we evaluate the ability of S. mutans and S. sobrinus in planktonic and biofilm lifestyle to adhere, invade and survive within human gingival fibroblast (HGF-1) cells. Planktonic and biofilm streptococci adhered and invaded host cells to different extents, showing higher efficiencies of biofilm than planktonic counterparts. Moreover, planktonic and biofilm streptococci showed the same percentage of survival within host cells. Transmission electron and confocal microscopy observations confirmed intracellular localization of planktonic and biofilm bacteria. The adhesion, invasion and survival abilities within human oral cells may be considered S. mutans and S. sobrinus virulence mechanisms to colonize and persist in the oral cavity in the absence of tooth surface.

  18. Murine fibroblasts transfection by laser optoporation: preliminary notes on the use of a visible wavelength

    Science.gov (United States)

    Palumbo, Giuseppe; Tecce, Mario F.; Roberti, Giuseppe; Colasanti, Alberto

    1994-12-01

    A novel approach to gene trasfection, i.e. the introduction of an extraneous gene into a host cell, by the 'non-contact forces' of a laser microbeam, is presented here. By means of a large magnification (100x) objective, the blue microbeam of an Argon laser (488 nm) has been focused on the cell membrane in culture in the presence of a pH indicator (namely the phenol-red), which is an usual component of culture media. Due to the local high light absorption of phenol red, which shows an absorption peak at 475 nm, at the site of the beam impact the cell membrane melts forming small circular holes. Throughout the holes, DNA purposely added to culture medium, may penetrate the cytoplasm. The wall damage, whose extension may be regulated by controlling the irradiation time, disappear spontaneously (membrane repair) within 1-2 minutes. By this technique, thereafter indicated as 'optoporation', we have successfully transfected into murine NIH3T3 fibroblasts (beta) -galactosidase and chloramphenicol-acetyltransferase bacterial genes. These conditions of transfection by means of 'non contact' forces, are very mild and do not require any addition of extraneous, potentially toxic chemicals. In addition, since the radiation used is in the visible region, where nucleic acids and most proteins do not absorb, no further deleterious effects for the cell are expected.

  19. Partial correction of sensitivity to oxidant stress in Friedreich ataxia patient fibroblasts by frataxin-encoding adeno-associated virus and lentivirus vectors.

    Science.gov (United States)

    Fleming, Jane; Spinoulas, Afroditi; Zheng, Maolin; Cunningham, Sharon C; Ginn, Samantha L; McQuilty, Robert C; Rowe, Peter B; Alexander, Ian E

    2005-08-01

    Peripheral nervous system (PNS) sensory neurons are directly involved in the pathophysiology of a number of debilitating inherited and acquired neurological conditions. The lack of effective treatments for many such conditions provides a strong rationale for exploring novel therapeutic approaches, including gene therapy. Friedreich ataxia (FRDA), a sensory neuropathy, is a progressive neurodegenerative disease associated with a loss of large sensory neurons from the dorsal root ganglia. Because a mouse model for this well-characterized disease has been generated, we elected to use FRDA as a model disease. In previous studies we achieved efficient and sustained delivery of a reporter gene to PNS sensory neurons, using recombinant adeno-associated viral (AAV) and lentiviral (LV) vectors. In the current study, AAV and LV vectors encoding the human frataxin cDNA were constructed and assessed for frataxin expression and function in primary FRDA patient fibroblast cell lines. FRDA fibroblasts have been shown to exhibit subtle biochemical changes, including increased mitochondrial iron and sensitivity to oxidant stress. Despite the inherent difficulty in working with primary cells, transduction of patient fibroblasts with either vector resulted in the expression of appropriately localized frataxin and partial reversal of phenotype.

  20. Localized shocks

    Energy Technology Data Exchange (ETDEWEB)

    Roberts, Daniel A. [Center for Theoretical Physics and Department of Physics,Massachusetts Institute of Technology, Cambridge, MA (United States); Stanford, Douglas [Stanford Institute for Theoretical Physics and Department of Physics,Stanford University, Stanford, CA (United States); School of Natural Sciences, Institute for Advanced Study,Princeton, NJ (United States); Susskind, Leonard [Stanford Institute for Theoretical Physics and Department of Physics,Stanford University, Stanford, CA (United States)

    2015-03-10

    We study products of precursors of spatially local operators, W{sub x{sub n}}(t{sub n})…W{sub x{sub 1}}(t{sub 1}), where W{sub x}(t)=e{sup −iHt}W{sub x}e{sup iHt}. Using chaotic spin-chain numerics and gauge/gravity duality, we show that a single precursor fills a spatial region that grows linearly in t. In a lattice system, products of such operators can be represented using tensor networks. In gauge/gravity duality, they are related to Einstein-Rosen bridges supported by localized shock waves. We find a geometrical correspondence between these two descriptions, generalizing earlier work in the spatially homogeneous case.

  1. Toothbrushing promotes gingival fibroblast proliferation more effectively than removal of dental plaque.

    Science.gov (United States)

    Horiuchi, Masazumi; Yamamoto, Tatsuo; Tomofuji, Takaaki; Ishikawa, Akira; Morita, Manabu; Watanabe, Tatsuo

    2002-09-01

    Removal of dental plaque is an essential element of periodontal treatment. However, there have also been studies of the effects of the mechanical stimulation provided by toothbrushing on gingival host-defense mechanisms. The aim of the study was to evaluate the effects of toothbrushing on gingival fibroblast proliferation in dogs over time, compared to effects of plaque removal without brushing. The mouths of six mongrel dogs were divided into four quadrants: two for daily toothbrushing, and two for daily plaque removal with a curette. After 1, 3 and 5 weeks of treatment, histometrical analyses were performed to assess inflammatory cell infiltration, proliferating cell nuclear antigen (PCNA)-positive fibroblasts, procollagen type I-positive fibroblasts in the subepithelial connective tissue of junctional epithelium. Toothbrushing increased the number of PCNA-positive fibroblasts in the first week, increased the number of type I procollagen-positive fibroblasts at the fifth week, and reduced inflammatory cell infiltration at the third week. These findings suggest that mechanically stimulated fibroblasts begin proliferating within a week, and this cell division results in an increased number of fibroblasts at the third week. It takes 5 weeks before differences in collagen synthesis between brushing and plaque removal areas are detectable.

  2. Ear fibroblasts derived from Taiwan yellow cattle are more heat resistant than those from Holstein cattle.

    Science.gov (United States)

    Wu, Hung-Yi; Peng, Shao-Yu; Li, Hung; Lee, Jai-Wei; Kesorn, Piyawit; Wu, Hsi-Hsun; Ju, Jyh-Cherng; Shen, Perng-Chih

    2017-05-01

    The objective of this study was to compare the thermotolerances of ear fibroblasts derived from Holstein (H) and Taiwan yellow cattle (Y) and their apoptosis-related protein expressions with (1, 3, 6, 12, and 24h) or without heat shock treatment. The results showed that the vaginal temperatures of Y (38.4-38.5°C) were (Pderived from Y (6h: 1.1%; 12h: 1.6%; 24h: 2.6%) were lower (Pderived from H (6h: 1.8%; 12h: 4.0%; 24h: 6.9%), respectively, after heat shock (42°C). The expression level of apoptosis inducing factor (AIF) in ear fibroblasts derived from H was higher (Pderived from Y after the heat shock treatment for 6h and 12h, respectively. The level of cytochrome c of ear fibroblasts derived from H was higher (Pderived from Y after the heat shock treatment for 1-12h, respectively. The abundances of Caspase-3, Caspase-8 and Caspase-9 of ear fibroblasts derived from H were higher (Pderived from Y after 12h and 24h of heat shock, respectively; the Bcl-2/Bax ratios of ear fibroblasts derived from H were lower (Pderived fibroblasts after heated for 1-24h. The expression level of HSP-70 of Y-derived ear fibroblasts was also higher (Pderived from Taiwan yellow cattle was better than that of cells derived from Holstein cattle. Copyright © 2017 Elsevier Ltd. All rights reserved.

  3. Differential effect of extracellular matrix derived from papillary and reticular fibroblasts on epidermal development in vitro.

    Science.gov (United States)

    Janson, David; Rietveld, Marion; Mahé, Christian; Saintigny, Gaëlle; El Ghalbzouri, Abdoelwaheb

    2017-06-01

    Papillary and reticular fibroblasts have different effects on keratinocyte proliferation and differentiation. The aim of this study was to investigate whether these effects are caused by differential secretion of soluble factors or by differential generation of extracellular matrix from papillary and reticular fibroblasts. To study the effect of soluble factors, keratinocyte monolayer cultures were grown in papillary or reticular fibroblast-conditioned medium. To study the effect of extracellular matrix, keratinocytes were grown on papillary or reticular-derived matrix. Conditioned medium from papillary or reticular fibroblasts did not differentially affect keratinocyte viability or epidermal development. However, keratinocyte viability was increased when grown on matrix derived from papillary, compared with reticular, fibroblasts. In addition, the longevity of the epidermis was increased when cultured on papillary fibroblast-derived matrix skin equivalents compared with reticular-derived matrix skin equivalents. The findings indicate that the matrix secreted by papillary and reticular fibroblasts is the main causal factor to account for the differences in keratinocyte growth and viability observed in our study. Differences in response to soluble factors between both populations were less significant. Matrix components specific to the papillary dermis may account for the preferential growth of keratinocytes on papillary dermis.

  4. Effect of eosinophils activated with Alternaria on the production of extracellular matrix from nasal fibroblasts.

    Science.gov (United States)

    Shin, Seung-Heon; Ye, Mi-Kyung; Choi, Sung-Yong; Kim, Yee-Hyuk

    2016-06-01

    Eosinophils and fibroblasts are known to play major roles in the pathogenesis of nasal polyps. Fungi are commonly found in nasal secretion and are associated with airway inflammation. To investigate whether activated eosinophils by airborne fungi can influence the production of extracellular matrix (ECM) from nasal fibroblasts. Inferior turbinate and nasal polyp fibroblasts were stimulated with Alternaria or Aspergillus, respectively, for 24 hours and ECM messenger RNA (mRNA) and protein expressions were measured. Eosinophils isolated from healthy volunteers were stimulated with Alternaria or Aspergillus for 4 hours then superoxide, eosinophil peroxidase, and transforming growth factor β1 were measured. Then activated eosinophils were cocultured with nasal fibroblasts for 24 hours, and ECM mRNA expressions were measured. Alternaria strongly enhanced ECM mRNA expression and protein production from nasal fibroblasts. Alternaria also induced the production of superoxide, eosinophil peroxidase, and transforming growth factor β1 from eosinophils, and activated eosinophils enhanced ECM mRNA expression when they were cocultured without the Transwell insert system. Eosinophils activated with Alternaria enhanced ECM mRNA expression from nasal polyp fibroblasts. Alternaria plays an important role in tissue fibrosis in the pathogenesis of nasal polyps by directly or indirectly influencing the production of ECM from nasal fibroblasts. Copyright © 2016 American College of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.

  5. Differential effect of hypoxia and acidity on lung cancer cell and fibroblast metabolism.

    Science.gov (United States)

    Giatromanolaki, Alexandra; Liousia, Maria; Arelaki, Stella; Kalamida, Dimitra; Pouliliou, Stamatia; Mitrakas, Achilleas; Tsolou, Avgi; Sivridis, Efthimios; Koukourakis, Michael

    2017-06-01

    This study examined the metabolic response of lung cancer cells and normal lung fibroblasts to hypoxia and acidity. GLUT1 and HXKII mRNA/protein expression was up-regulated under hypoxia in the MRC5 fibroblasts and in the A549 and H1299 lung cancer cell lines, indicating intensified glucose absorption and glycolysis. Under hypoxia, the LDHA mRNA and LDH5 protein levels increased in the cancer cells but not in the fibroblasts. Acidity suppressed the above-mentioned hypoxia effect. PDH-kinase-1 (PDK1 mRNA and protein) and inactive phosphorylated-PDH protein levels were induced under hypoxia in the cancer cells, whereas these were reduced in the MRC5 lung fibroblasts. In human tissue sections, the prevalent expression patterns supported the contrasting metabolic behavior of cancer cells vs. tumor fibroblasts. The monocarboxylate/lactate transporter 1 (MCT1) was up-regulated in all the cell lines under hypoxic conditions, but it was suppressed under acidic conditions. The mitochondrial DNA (mtDNA) content per cell decreased significantly in the A549 cancer cell line under hypoxia, but it increased in the MRC5 fibroblasts. Taking into account these findings, we suggest that, under hypoxia, cancer cells intensify the anaerobic direction in glycolysis, while normal fibroblasts prefer to seek energy by intensifying the aerobic use of the available oxygen.

  6. Curcumin inhibits transforming growth factor β induced differentiation of mouselung fibroblasts to myofibroblasts

    Directory of Open Access Journals (Sweden)

    Daishun Liu

    2016-11-01

    Full Text Available Transforming growth factor β (TGFβ induced differentiation of lung fibroblasts to myofibroblasts is a key event in the pathogenesis of pulmonary fibrosis. This study aimed to evaluate the effect of curcumin on TGFβ induced differentiation of lung fibroblasts to myofibroblasts and explore the underlying mechanism. Mouse lung fibroblasts were cultured and treated with TGFβ2 and curcumin or rosiglitazone. Cell vitality was examined by MTT assay. The secretion of collagen-1 was assessed by ELISA. α smooth muscle actin (αSMA was visualized by immunofluorescence technique. The expression of peroxisome proliferator activated receptor γ (PPARγ and platelet derived growth factor R β (PDGFRβ was detected by PCR and Western blot analysis. We found that curcumin and rosiglitazone inhibited the proliferation and TGFβ induced differentiation of mouse lung fibroblasts. In addition, curcumin and rosiglitazone inhibited collagen-1 secretion and αSMA expression in mouse lung fibroblasts. Furthermore, curcumin and rosiglitazone upregulated PPARγ and downregulated PDGFRβ expression in mouse lung fibroblasts. In conclusion, our study reveals novel mechanism by which curcumin inhibits TGFβ2 driven differentiation of lung fibroblasts to myofibroblasts. Curcumin could potentially be used for effective treatment of pulmonary fibrosis.

  7. Defective alterations in the collagen network to prostacyclin in COPD lung fibroblasts

    Directory of Open Access Journals (Sweden)

    Larsson-Callerfelt Anna-Karin

    2013-02-01

    Full Text Available Abstract Background Prostacyclin analogs are potent vasodilators and possess anti-inflammatory properties. However, the effect of prostacyclin on extracellular matrix (ECM in COPD is not well known. Collagen fibrils and proteoglycans are essential ECM components in the lung and fibroblasts are key players in regulating the homeostasis of ECM proteins. The aim was to study the synthesis of prostacyclin and its effect on fibroblast activity and ECM production, and in particular collagen I and the collagen-associated proteoglycans biglycan and decorin. Methods Parenchymal lung fibroblasts were isolated from lungs from COPD patients (GOLD stage IV and from lungs and transbronchial biopsies from control subjects. The prostacyclin analog iloprost was used to study the effect of prostacyclin on ECM protein synthesis, migration, proliferation and contractile capacity of fibroblasts. Results TGF-β1 stimulation significantly increased prostacyclin synthesis in fibroblasts from COPD patients (p  Conclusions Iloprost reduced collagen I synthesis and fibroblast contractility but did not affect the collagen-associated proteoglycans or proliferation rate in fibroblasts from COPD patients. Enhanced prostacyclin production could lead to improper collagen network fibrillogenesis and a more emphysematous lung structure in severe COPD patients.

  8. Fibroblast Growth Factor 10-Fibroblast Growth Factor Receptor 2b Mediated Signaling Is Not Required for Adult Glandular Stomach Homeostasis

    Science.gov (United States)

    Sala, Frederic G.; Ford, Henri R.; Bellusci, Saverio; Grikscheit, Tracy C.

    2012-01-01

    The signaling pathways that are essential for gastric organogenesis have been studied in some detail; however, those that regulate the maintenance of the gastric epithelium during adult homeostasis remain unclear. In this study, we investigated the role of Fibroblast growth factor 10 (FGF10) and its main receptor, Fibroblast growth factor receptor 2b (FGFR2b), in adult glandular stomach homeostasis. We first showed that mouse adult glandular stomach expressed Fgf10, its receptors, Fgfr1b and Fgfr2b, and most of the other FGFR2b ligands (Fgf1, Fgf7, Fgf22) except for Fgf3 and Fgf20. Fgf10 expression was mesenchymal whereas FGFR1 and FGFR2 expression were mostly epithelial. Studying double transgenic mice that allow inducible overexpression of Fgf10 in adult mice, we showed that Fgf10 overexpression in normal adult glandular stomach increased epithelial proliferation, drove mucous neck cell differentiation, and reduced parietal and chief cell differentiation. Although a similar phenotype can be associated with the development of metaplasia, we found that Fgf10 overexpression for a short duration does not cause metaplasia. Finally, investigating double transgenic mice that allow the expression of a soluble form of Fgfr2b, FGF10's main receptor, which acts as a dominant negative, we found no significant changes in gastric epithelial proliferation or differentiation in the mutants. Our work provides evidence, for the first time, that the FGF10-FGFR2b signaling pathway is not required for epithelial proliferation and differentiation during adult glandular stomach homeostasis. PMID:23133671

  9. Fibroblast growth factor 10-fibroblast growth factor receptor 2b mediated signaling is not required for adult glandular stomach homeostasis.

    Directory of Open Access Journals (Sweden)

    Allison L Speer

    Full Text Available The signaling pathways that are essential for gastric organogenesis have been studied in some detail; however, those that regulate the maintenance of the gastric epithelium during adult homeostasis remain unclear. In this study, we investigated the role of Fibroblast growth factor 10 (FGF10 and its main receptor, Fibroblast growth factor receptor 2b (FGFR2b, in adult glandular stomach homeostasis. We first showed that mouse adult glandular stomach expressed Fgf10, its receptors, Fgfr1b and Fgfr2b, and most of the other FGFR2b ligands (Fgf1, Fgf7, Fgf22 except for Fgf3 and Fgf20. Fgf10 expression was mesenchymal whereas FGFR1 and FGFR2 expression were mostly epithelial. Studying double transgenic mice that allow inducible overexpression of Fgf10 in adult mice, we showed that Fgf10 overexpression in normal adult glandular stomach increased epithelial proliferation, drove mucous neck cell differentiation, and reduced parietal and chief cell differentiation. Although a similar phenotype can be associated with the development of metaplasia, we found that Fgf10 overexpression for a short duration does not cause metaplasia. Finally, investigating double transgenic mice that allow the expression of a soluble form of Fgfr2b, FGF10's main receptor, which acts as a dominant negative, we found no significant changes in gastric epithelial proliferation or differentiation in the mutants. Our work provides evidence, for the first time, that the FGF10-FGFR2b signaling pathway is not required for epithelial proliferation and differentiation during adult glandular stomach homeostasis.

  10. CULTURES OF FIBROBLAST-LIKE SYNOVIAL CELLS FROM PATIENTS WITH RHEUMATOID ARTHRITIS: PROPERTIES AND OPPORTUNITIES

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    M. A. Schneider

    2016-01-01

    Full Text Available The present review contains data from literature concerning the in vivo structure of synovial membranes in healthy people and patients with rheumatoid arthritis (RA. The properties of in vitro cultured fibroblast-like synovial cells (FLS from RA patients are considered, including FLS morphology, phenotype and function. A standard protocol of in vitro FLS culturing is described. Notably, the FLS are characterized by autonomic functioning, ability for invasive growth/migration, e.g., into non-affected joints. These FLS properties may a reason of multiple joint involvement typical to RA. Special attention is drawn to characterization of stable phenotypic profile of FLS which results from certain epigenetic disturbances, i.e., changes of the DNA methylation, histone acetylation, and micro-RNA effects.The FLS from RA patients are characterized with stable and extensive hypomethylation of genes which occurs in vivo and persists after repeated culture passages. Some promoters of genes involved into RA pathogenesis (for example, CXCL12, IL-6 are hypomethylated. By contrary, some other gene promoters (e.g., the death receptor 3 gene are shown to be hypermethylated. An increased histone acetylation of genes encoding proinflammatory mediators (such as MMP1 may be an important mechanism of persistent inflammation in RA. Changes in histone acetylation in FLS are related to high levels of ubiquitin-like SUMO-1 protein and concurrent decrease in specific protease SENP1activity. A role of histone acetylation in RA pathogenesis is supported by efficacy of a histone deacetylase inhibitor (Trichostatin A in collagen-induced murine arthritis. Local concentrations of micro RNA-155, micro-RNA-146а, and micro-RNA-203 are permanently increased in FLS cultures, synovial tissues, and PBMC of the RA patients. Expression of micro RNA-124а is decreased in FLS from RA, as compared with OA FLS.One may conclude that the fibroblast-like synovial cells are key cellular

  11. SIRT-1 regulates TGF-β-induced dermal fibroblast migration via modulation of Cyr61 expression.

    Science.gov (United States)

    Kwon, Eun-Jeong; Park, Eun-Jung; Yu, Hyeran; Huh, Jung-Sik; Kim, Jinseok; Cho, Moonjae

    2017-07-27

    SIRT1 is a NAD-dependent protein deacetylase that participates in cellular regulation. The increased migration of fibroblasts is an important phenotype in fibroblast activation. The role of SIRT1 in cell migration remains controversial as to whether SIRT1 acts as an activator or suppressor of cell migration. Therefore, we have established the role of SIRT1 in the migration of human dermal fibroblasts and explored targets of SIRT1 during dermal fibroblast migration. SIRT1 and Cyr61 were expressed in human dermal fibroblasts and the stimulation with TGF-β further induced their expression. Treatment with resveratrol (RSV), a SIRT1 agonist, or overexpression of SIRT1 also promoted the expression Cyr61 in human dermal fibroblasts, whereas the inhibition of SIRT1 activity by nicotinamide or knockdown of SIRT1 decreased the level of Cyr61, as well as TGF-β or RSV-induced Cyr61 expression. Blocking of ERK signaling by PD98509 reduced the expression of Cyr61 induced by TGF-β or RSV. TGF-β, RSV, or SIRT1 overexpression enhanced β-catenin as well as Cyr61 expression. This stimulation was reduced by the Wnt inhibitor XAV939. RSV increased migration and nicotinamide attenuated RSV-induced migration of human dermal fibroblasts. Furthermore, SIRT1 overexpression promoted cell migration, whereas blocking Cyr61 attenuated SIRT1-stimulated migration of human dermal fibroblasts. SIRT1 increased cell migration by stimulating Cyr61 expression and the ERK and Wnt/β-catenin signaling. SIRT1-induced Cyr61 activity is very important for human dermal fibroblasts migration.

  12. Enhanced keratinocyte proliferation and migration in co-culture with fibroblasts.

    Directory of Open Access Journals (Sweden)

    Zhenxiang Wang

    Full Text Available Wound healing is primarily controlled by the proliferation and migration of keratinocytes and fibroblasts as well as the complex interactions between these two cell types. To investigate the interactions between keratinocytes and fibroblasts and the effects of direct cell-to-cell contact on the proliferation and migration of keratinocytes, keratinocytes and fibroblasts were stained with different fluorescence dyes and co-cultured with or without transwells. During the early stage (first 5 days of the culture, the keratinocytes in contact with fibroblasts proliferated significantly faster than those not in contact with fibroblasts, but in the late stage (11(th to 15(th day, keratinocyte growth slowed down in all cultures unless EGF was added. In addition, keratinocyte migration was enhanced in co-cultures with fibroblasts in direct contact, but not in the transwells. Furthermore, the effects of the fibroblasts on keratinocyte migration and growth at early culture stage correlated with heparin-binding EGF-like growth factor (HB-EGF, IL-1α and TGF-β1 levels in the cultures where the cells were grown in direct contact. These effects were inhibited by anti-HB-EGF, anti-IL-1α and anti-TGF-β1 antibodies and anti-HB-EGF showed the greatest inhibition. Co-culture of keratinocytes and IL-1α and TGF-β1 siRNA-transfected fibroblasts exhibited a significant reduction in HB-EGF production and keratinocyte proliferation. These results suggest that contact with fibroblasts stimulates the migration and proliferation of keratinocytes during wound healing, and that HB-EGF plays a central role in this process and can be up-regulated by IL-1α and TGF-β1, which also regulate keratinocyte proliferation differently during the early and late stage.

  13. Metabolic cooperation between co-cultured lung cancer cells and lung fibroblasts.

    Science.gov (United States)

    Koukourakis, Michael I; Kalamida, Dimitra; Mitrakas, Achilleas G; Liousia, Maria; Pouliliou, Stamatia; Sivridis, Efthimios; Giatromanolaki, Alexandra

    2017-11-01

    Cooperation of cancer cells with stromal cells, such as cancer-associated fibroblasts (CAFs), has been revealed as a mechanism sustaining cancer cell survival and growth. In the current study, we focus on the metabolic interactions of MRC5 lung fibroblasts with lung cancer cells (A549 and H1299) using co-culture experiments and studying changes of the metabolic protein expression profile and of their growth and migration abilities. Using western blotting, confocal microscopy and RT-PCR, we observed that in co-cultures MRC5 respond by upregulating pyruvate dehydrogenase (PDH) and the monocarboxylate transporter MCT1. In contrast, cancer cells increase the expression of glucose transporters (GLUT1), LDH5, PDH kinase and the levels of phosphorylated/inactivated pPDH. H1299 cells growing in the same culture medium with fibroblasts exhibit a 'metastasis-like' phenomenon by forming nests within the fibroblast area. LDH5 and pPDH were drastically upregulated in these nests. The growth rate of both MRC5 and cancer cells increased in co-cultures. Suppression of LDHA or PDK1 in cancer cells abrogates the stimulatory signal from cancer cells to fibroblasts. Incubation of MRC5 fibroblasts with lactate resulted in an increase of LDHB and of PDH expression. Silencing of PDH gene in fibroblasts, or silencing of PDK1 or LDHA gene in tumor cells, impedes cancer cell's migration ability. Overall, a metabolic cooperation between lung cancer cells and fibroblasts has been confirmed in the context of direct Warburg effect, thus the fibroblasts reinforce aerobic metabolism to support the intensified anaerobic glycolytic pathways exploited by cancer cells.

  14. Chemokine release from human rhinovirus-infected airway epithelial cells promotes fibroblast migration.

    Science.gov (United States)

    Shelfoon, Christopher; Shariff, Sami; Traves, Suzanne L; Kooi, Cora; Leigh, Richard; Proud, David

    2016-07-01

    Thickening of the lamina reticularis, a feature of remodeling in the asthmatic airways, is now known to be present in young children who wheeze. Human rhinovirus (HRV) infection is a common trigger for childhood wheezing, which is a risk factor for subsequent asthma development. We hypothesized that HRV-infected epithelial cells release chemoattractants to recruit fibroblasts that could potentially contribute to thickening of the lamina reticularis. We sought to investigate whether conditioned medium from HRV-infected epithelial cells can trigger directed migration of fibroblasts. Human bronchial epithelial cells were exposed to medium alone or infected with HRV-16. Conditioned medium from both conditions were tested as chemoattractants for human bronchial fibroblasts in the xCELLigence cell migration apparatus. HRV-conditioned medium was chemotactic for fibroblasts. Treatment of fibroblasts with pertussis toxin, an inhibitor of Gαi-coupled receptors, prevented their migration. Production of epithelial chemoattractants required HRV replication. Multiplex analysis of epithelial supernatants identified CXCL10, CXCL8, and CCL5 as Gαi-coupled receptor agonists of potential interest. Subsequent analysis confirmed that fibroblasts express CXCR3 and CXCR1 receptors and that CXCL10 and, to a lesser extent, CXCL8, but not CCL5, are major contributors to fibroblast migration caused by HRV-conditioned medium. CXCL10 and CXCL8 produced from HRV-infected epithelial cells are chemotactic for fibroblasts. This raises the possibility that repeated HRV infections in childhood could contribute to the initiation and progression of airway remodeling in asthmatic patients by recruiting fibroblasts that produce matrix proteins and thicken the lamina reticularis. Copyright © 2016 American Academy of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.

  15. Fibroblasts Express Immune Relevant Genes and Are Important Sentinel Cells during Tissue Damage in Rainbow Trout (Oncorhynchus mykiss)

    DEFF Research Database (Denmark)

    Ingerslev, Hans-Christian; Ossum, Carlo Gunnar; Lindenstrom, Thomas

    2010-01-01

    from E. coli, supernatant and debris from sonicated RTHDF cells. LPS was overall the strongest inducer of IL-1 beta, IL-8, IL-10, TLR-3 and TLR-9. IL-1 beta and IL-8 were already highly up regulated after 1 hour of LPS stimulation. Supernatant stimuli significantly increased the expression of IL-1 beta......, TLR-3 and TLR-9, whereas the debris stimuli only increased expression of IL-1 beta. Consequently, an in vivo experiment was further set up. By mechanically damaging the muscle tissue of rainbow trout, it was shown that fibroblasts in the muscle tissue of rainbow trout contribute to electing a highly...... local inflammatory response following tissue injury. The damaged muscle tissue showed a strong increase in the expression of the immune genes IL-1 beta, IL-8 and TGF-beta already 4 hours post injury at the site of injury while the expression in non-damaged muscle tissue was not influenced. A weaker...

  16. Distinct Fibroblasts in the Papillary and Reticular Dermis: Implications for Wound Healing.

    Science.gov (United States)

    Woodley, David T

    2017-01-01

    Human skin wounds heal largely by reparative wound healing rather than regenerative wound healing. Human skin wounds heal with scarring and without pilosebaceous units or other appendages. Dermal fibroblasts come from 2 distinct lineages of cells that have distinct cell markers and, more importantly, distinct functional abilities. Human skin wound healing largely involves the dermal fibroblast lineage from the reticular dermis and not the papillary dermis. If scientists could find a way to stimulate the dermal fibroblast lineages from the papillary dermis in early wound healing, perhaps human skin wounds could heal without scarring and with skin appendages. Copyright © 2016 Elsevier Inc. All rights reserved.

  17. Emdogain-regulated gene expression in palatal fibroblasts requires TGF-βRI kinase signaling.

    OpenAIRE

    Stähli, Alexandra Beatrice; Bosshardt, Dieter; Sculean, Anton; Gruber, Reinhard

    2014-01-01

    Genome-wide microarrays have suggested that Emdogain regulates TGF-β target genes in gingival and palatal fibroblasts. However, definitive support for this contention and the extent to which TGF-β signaling contributes to the effects of Emdogain has remained elusive. We therefore studied the role of the TGF-β receptor I (TGF-βRI) kinase to mediate the effect of Emdogain on palatal fibroblasts. Palatal fibroblasts were exposed to Emdogain with and without the inhibitor for TGF-βRI kinase, SB43...

  18. Extracellular Matrix Metalloproteinase Inducer (EMMPRIN) promotes lung fibroblast proliferation, survival and differentiation to myofibroblasts.

    Science.gov (United States)

    Hasaneen, Nadia A; Cao, Jian; Pulkoski-Gross, Ashleigh; Zucker, Stanley; Foda, Hussein D

    2016-02-17

    Idiopathic pulmonary fibrosis (IPF) is a chronic progressively fatal disease. Extracellular Matrix Metalloproteinase Inducer (EMMPRIN) is a glycosylated transmembrane protein that induces the expression of some matrix metalloproteinase (MMP) in neighboring stromal cells through direct epithelial-stromal interactions. EMMPRIN is highly expressed in type II alveolar epithelial cells at the edges of the fibrotic areas in IPF lung sections. However, the exact role of EMMPRIN in IPF is unknown. To determine if EMMPRIN contributes to lung fibroblast proliferation, resistance to apoptosis, and differentiation to myofibroblasts, normal Human lung fibroblasts (NHLF) transiently transfected with either EMMPRIN/GFP or GFP were treated with TGF- β1 from 0 to 10 ng/ml for 48 h and examined for cell proliferation (thymidine incorporation), apoptosis (FACS analysis and Cell Death Detection ELISA assay), cell migration (Modified Boyden chamber) and differentiation to myofibroblasts using Western blot for α-smooth actin of cell lysates. The effect of EMMPRIN inhibition on NHLF proliferation, apoptosis, migration and differentiation to myofibroblasts after TGF- β1 treatment was examined using EMMPRIN blocking antibody. We examined the mechanism by which EMMPRIN induces its effects on fibroblasts by studying the β-catenin/canonical Wnt signaling pathway using Wnt luciferase reporter assays and Western blot for total and phosphorylated β-catenin. Human lung fibroblasts overexpressing EMMPRIN had a significant increase in cell proliferation and migration compared to control fibroblasts. Furthermore, EMMPRIN promoted lung fibroblasts resistance to apoptosis. Lung fibroblasts overexpressing EMMPRIN showed a significantly increased expression of α- smooth muscle actin, a marker of differentiation to myofibroblasts compared to control cells. TGF-β1 increased the expression of EMMPRIN in lung fibroblasts in a dose-dependent manner. Attenuation of EMMPRIN expression with the use of an

  19. Cytotoxic Effects of Nickel Nanowires in Human Fibroblasts

    KAUST Repository

    Felix Servin, Laura P.

    2014-04-01

    There is an increasing interest for the use of nanostructures as potential tools in areas that include biology and medicine, for applications spanning from cell separation to treatments of diseases. Magnetic nanoparticles (MNPs) have been the most widely studied and utilized nanostructures in biomedical applications. Despite their popularity, the regular shape of MNPs limits their potential for certain applications. Studies have shown that magnetic nanowires (MNWs), due to their high-­‐aspect ratio and specific magnetic properties, might provide improved performance for some biomedical applications. As a consequence, MNWs have received increasing attention from researchers in the last years. However, as with any other nanostructure intended for biomedical applications, rigorous studies must be carried out to determine their potential toxicity and adverse effects before they can be successfully incorporated in clinical applications. This work attempts to elucidate the cytotoxic effects of nickel NWs (Ni NWs) in human fibroblasts by measuring cell viability under different parameters. Ni NWs of three different lengths (0.86 ± 0.02 μm, 1.1 ± 0.1 μm and 6.1 ± 0.6 μm) were fabricated by electrodeposition using porous aluminum oxide (PAO) membranes as templates. Energy dispersive X-­‐Ray analysis (EDAX) and X-­‐Ray diffraction (XRD) were used for the chemical characterization of the Ni NWs. Their physical characterization was done using scanning electron microscopy (SEM) and transmission electron microscopy (TEM) imaging. MTT assays were performed to assess cell viability of human fibroblasts in the presence of Ni NWs. NW length, NW/cell ratio and exposure time were changed throughout the experiments to elucidate their effects on cell viability. The results showed that NWs length has a strong effect on internalization and cytotoxicity. Smaller NWs showed higher toxicity levels at earlier times while longer NWs had stronger effects on cell viability at

  20. Localized shocks

    OpenAIRE

    Stanford, Douglas; Susskind, Leonard; Roberts, Daniel Adam

    2014-01-01

    We study products of precursors of spatially local operators, W[subscript xn](tn)⋅⋅⋅W[subscript x1](t[subscript 1]), where W [subscript x] (t) = e [superscript − iHt] W [subscript x] e [superscript iHt]. Using chaotic spin-chain numerics and gauge/gravity duality, we show that a single precursor fills a spatial region that grows linearly in t. In a lattice system, products of such operators can be represented using tensor networks. In gauge/gravity duality, they are related to Einstein-Rosen ...

  1. ERK5 regulates basic fibroblast growth factor-induced type 1 plasminogen activator inhibitor expression and cell proliferation in lung fibroblasts.

    Science.gov (United States)

    Han, Jung-Hwa; Hwang, Ae-Rang; Nam, Dae-Hwan; Kim, Suji; Choi, Hyoung Chul; Lim, Jae Hyang; Woo, Chang-Hoon

    2015-08-15

    bFGF is a potent mitogen of cells associated with fibrosis. Although ERK5 has been reported to play roles in the development of fibrosis, its roles in regulating bFGF-induced fibrotic responses are not understood, especially in lung fibroblasts. The authors investigated the role of ERK5 in bFGF induction of cell proliferation and in induction of PAI-1, a critical regulator of the pathological features of fibrosis, in lung fibroblasts. The role played by ERK5 in bFGF-induced PAI-1 expression was elucidated by perturbing the ERK5 signaling pathway using a specific chemical inhibitor and siRNA of ERK5. The effects of ERK5 signal perturbation on PAI-1 expression were measured at multiple levels by Q-PCR, immunoblotting, ELISA, and reporter gene analysis. The role of MEF2 in bFGF-induced activation of PAI-1 promoter activity via ERK5 was measured using a biotin-labeled DNA pull-down assay, and the effects of ERK5 on the mitogenic effects of bFGF were assessed using a MTT assay. In both primary human lung fibroblast and lung fibroblast cell lines, inhibition of ERK5 blocked bFGF-induced PAI-1 expression at both mRNA and protein levels and inhibited bFGF-induced PAI-1 promoter activity induction by bFGF. Upon stimulation with bFGF, MEF2 directly bound to the consensus sequence of the MEF2 binding site in the PAI-1 promoter. In addition, bFGF-induced PAI-1 up-regulation was inhibited by MEF2 siRNA, and bFGF-induced fibroblast proliferation was blocked by inhibiting ERK5. This study reveals a novel role for the ERK5-MEF2 cascade, linking bFGF-induced PAI-1 expression and subsequent mitogenic processes in lung fibroblasts. Copyright © 2015 Elsevier Inc. All rights reserved.

  2. [A method for the primary culture of fibroblasts isolated from human airway granulation tissues].

    Science.gov (United States)

    Chen, Nan; Zhang, Jie; Xu, Min; Wang, Yu-ling; Pei, Ying-hua

    2013-04-01

    To establish a feasible method to culture primary fibroblasts isolated from human airway granulation tissues, and therefore to provide experimental data for the investigation of the pathogenesis of benign airway stenosis. The granulation tissues were collected from 6 patients during routine bronchoscopy at our department of Beijing Tiantan Hospital from April to June 2011. Primary fibroblasts were obtained by culturing the explanted tissues. Cell growth was observed under inverted microscope. All of these 6 primary cultures were successful. Fibroblast-like cells were observed to migrate from the tissue pieces 3 d after inoculation. After 9-11 d of culture, cells reached to 90% confluence and could be sub-cultured. After passage, the cells were still in a typical elongated spindle-shape and grew well. The cells could be sub-cultured further when they formed a monolayer. Explant culture is a reliable method for culturing primary fibroblasts from human airway granulation tissues.

  3. Senescence-associated β-galactosidase activity in the in vitro ovarian stromal fibroblasts

    National Research Council Canada - National Science Library

    Lilian Chuaire-Noack; Cristian García-Morcote; Sandra Rocío Ramírez-Clavijo

    2011-01-01

    ... the in vitro growth of stromal fibroblasts from the ovarian cortex and their β-galactosidase activity at pH 6,enzyme whose expression is considered as a marker of replicativesenescence. Methods...

  4. Degradable PLGA Scaffolds with Basic Fibroblast Growth Factor: Experimental Studies in Myocardial Revascularization

    OpenAIRE

    Wang, Ying; Liu, Xiao-Cheng; Zhao, Jian; Kong, Xiang-Rong; Shi, Rong-Fang; Zhao, Xiao-Bin; Song, Cun-Xian; Liu, Tian-Jun; Lu, Feng

    2009-01-01

    Our goal was to investigate the efficacy of degradable poly(D,L-lactic-coglycolic acid) (PLGA) scaffolds loaded with basic fibroblast growth factor (bFGF) in inducing cardiac neovascularization, increasing perfusion, and improving cardiac function.

  5. Functional characteristics of vaginal fibroblastic cells from premenopausal women with pelvic organ prolapse

    NARCIS (Netherlands)

    Ruiz-Zapata, A.M.; Kerkhof, M.H.; Zandieh-Doulabi, B.; Brölmann, H.A.M.; Smit, T.H.; Helder, M.N.

    2014-01-01

    Pelvic organ prolapse (POP) remains a great therapeutic challenge with no optimal treatment available. Tissue maintenance and remodelling are performed by fibroblasts, therefore altered cellular functionality may influence tissue quality. In this study, we evaluated functional characteristics of

  6. Inhibition of proliferation and migration of stricture fibroblasts by epithelial cell-conditioned media

    Directory of Open Access Journals (Sweden)

    Nilima Nath

    2015-01-01

    Conclusion: These results demonstrate the ability of ECCM to inhibit the proliferation and migration of stricture fibroblasts and present it as an effective adjunct in urethroplasty, which may influence stricture wound healing and inhibit the recurrence of stricture.

  7. Estrogen modulates the influence of cardiac inflammatory cells on function of cardiac fibroblasts

    Directory of Open Access Journals (Sweden)

    McLarty JL

    2013-08-01

    Full Text Available Jennifer L McLarty,1 Jianping Li,2 Scott P Levick,3 Joseph S Janicki2 1Department of Pathology, University of Alabama at Birmingham, Birmingham, AL, USA; 2Department of Cell Biology and Anatomy, School of Medicine, University of South Carolina, Columbia, SC, USA; 3Department of Pharmacology and Toxicology, Cardiovascular Center, Medical College of Wisconsin, Milwaukee, WI, USA Background: Inflammatory cells play a major role in the pathology of heart failure by stimulating cardiac fibroblasts to regulate the extracellular matrix in an adverse way. In view of the fact that inflammatory cells have estrogen receptors, we hypothesized that estrogen provides cardioprotection by decreasing the ability of cardiac inflammatory cells to influence fibroblast function. Methods: Male rats were assigned to either an untreated or estrogen-treated group. In the treated group, estrogen was delivered for 2 weeks via a subcutaneous implanted pellet containing 17β-estradiol. A mixed population of cardiac inflammatory cells, including T-lymphocytes (about 70%, macrophages (about 12%, and mast cells (about 12%, was isolated from each rat and cultured in a Boyden chamber with cardiac fibroblasts from untreated adult male rats for 24 hours. To examine if tumor necrosis factor-alpha (TNF-α produced by inflammatory cells represents a mechanism contributing to the stimulatory effects of inflammatory cells on cardiac fibroblasts, inflammatory cells from the untreated group were incubated with cardiac fibroblasts in a Boyden chamber system for 24 hours in the presence of a TNF-α -neutralizing antibody. Cardiac fibroblasts were also incubated with 5 ng/mL of TNF-α for 24 hours. Fibroblast proliferation, collagen synthesis, matrix metalloproteinase activity, β1 integrin protein levels, and the ability of fibroblasts to contract collagen gels were determined in all groups and statistically compared via one-way analysis of variance. Results: Inflammatory cells from the

  8. Glycogen synthase kinase-3 (GSK-3) regulates TGF-1-induced differentiation of pulmonary fibroblasts

    NARCIS (Netherlands)

    Baarsma, Hoeke A.; Engelbertink, Lilian H. J. M.; van Hees, Lonneke J.; Menzen, Mark H.; Meurs, Herman; Timens, Wim; Postma, Dirkje S.; Kerstjens, Huib A. M.; Gosens, Reinoud

    Background Chronic lung diseases such as asthma, COPD and pulmonary fibrosis are characterized by abnormal extracellular matrix (ECM) turnover. TGF- is a key mediator stimulating ECM production by recruiting and activating lung fibroblasts and initiating their differentiation process into more

  9. Cell-surface glycosyltransferases in cultured fibroblasts: increased activity and release during serum stimulation of growth.

    Science.gov (United States)

    LaMont, J T; Gammon, M T; Isselbacher, K J

    1977-01-01

    Cell-surface galactosyltransferase was studied in suspensions of intact baby hamster kidney fibroblasts with both endogenous and exogenous glycoprotein acceptors. The cell-surface location of galactosyltransferase was demonstrated in experiments with the enzyme modifier alpha-lactalbumin, which does not enter the cell. The addition of alpha-lactalbumin to the assay medium for galactosyltransferase resulted in accumulation of lactose in the medium but not in the cells. There was no detectable hydrolysis of UDP-galactose to free galactose by these cells, nor did a 100-fold molar excess of free galactose inhibit cell-surface galactosyltransferase. There was a marked increase in specific activity of cell-surface exogenous galactosyltransferase in serum-stimulated as compared to resting fibroblasts. Dividing but not resting fibroblasts released galactosyltransferase, but not sialyl- or fucosyltransferase, in soluble form into the tissue culture medium. The release of galactosyltransferase was greater from virally transformed than from nontransformed fibroblasts. PMID:191828

  10. Effect of ALDH2 on High Glucose-Induced Cardiac Fibroblast Oxidative Stress, Apoptosis, and Fibrosis

    National Research Council Canada - National Science Library

    Xiaoyu Gu; Tingting Fang; Pinfang Kang; Junfeng Hu; Ying Yu; Zhenghong Li; Xiangyang Cheng; Qin Gao

    2017-01-01

    Our study aimed firstly to observe whether ALDH2 was expressed in neonate rat cardiac fibroblasts, then to investigate the effect of activation of ALDH2 on oxidative stress, apoptosis, and fibrosis...

  11. Properties of sulfatases in cultured skin fibroblasts of multiple sulfatase deficient patients.

    Science.gov (United States)

    Yutaka, T; Okada, S; Kato, T; Inui, K; Yabuuchi, H

    1981-10-01

    Various sulfatase activities were assayed in cultured skin fibroblasts from patients with multiple sulfatase deficiency (MSD). MSD cell lines displayed deficiencies of arylsulfatase A and iduronate sulfatase, but activities of arylsulfatase B, N-acetylgalactosamine 6-sulfate sulfatase and N-acetylglucosamine 6-sulfate sulfatase were within normal ranges, but not consistently. Arylsulfatase A, minor anionic arylsulfatase and N-acetylgalactosamine 6-sulfate sulfatase in MSD cell lines had similar Km, pH optima, inhibitory or activator sensitivity to that of normal skin fibroblasts. Arylsulfatase B in MSD cell lines also had properties similar to that of normal skin fibroblasts, except an abnormal heat stability. From our results, we conclude that properties of arylsulfatase A, minor anionic arylsulfatase and N-acetylgalactosamine 6-sulfate sulfatase in MSD fibroblasts were intact. On the other hand, arylsulfatase B in MSD might be a functionally abnormal enzyme.

  12. Agonists of fibroblast growth factor receptor induce neurite outgrowth and survival of cerebellar granule neurons

    DEFF Research Database (Denmark)

    Li, Shizhong; Christensen, Claus; Køhler, Lene B

    2009-01-01

    Fibroblast growth factor receptor (FGFR) signaling is pivotal in the regulation of neurogenesis, neuronal differentiation and survival, and synaptic plasticity both during development and in adulthood. In order to develop low molecular weight agonists of FGFR, seven peptides, termed hexafins...

  13. Impact of Antibodies and Strain Polymorphisms on Cytomegalovirus Entry and Spread in Fibroblasts and Epithelial Cells.

    Science.gov (United States)

    Cui, Xiaohong; Freed, Daniel C; Wang, Dai; Qiu, Ping; Li, Fengsheng; Fu, Tong-Ming; Kauvar, Lawrence M; McVoy, Michael A

    2017-07-01

    Cytomegalovirus (CMV) entry into fibroblasts differs from entry into epithelial cells. CMV also spreads cell to cell and can induce syncytia. To gain insights into these processes, 27 antibodies targeting epitopes in CMV virion glycoprotein complexes, including glycoprotein B (gB), gH/gL, and the pentamer, were evaluated for their effects on viral entry and spread. No antibodies inhibited CMV spread in fibroblasts, including those with potent neutralizing activity against fibroblast entry, while all antibodies that neutralized epithelial cell entry also inhibited spread in epithelial cells and a correlation existed between the potencies of these two activities. This suggests that exposure of virions to the cell culture medium is obligatory during spread in epithelial cells but not in fibroblasts. In fibroblasts, the formation of syncytiumlike structures was impaired not only by antibodies to gB or gH/gL but also by antibodies to the pentamer, suggesting a potential role for the pentamer in promoting fibroblast fusion. Four antibodies reacted with linear epitopes near the N terminus of gH, exhibited strain specificity, and neutralized both epithelial cell and fibroblast entry. Five other antibodies recognized conformational epitopes in gH/gL and neutralized both fibroblast and epithelial cell entry. That these antibodies were strain specific for neutralizing fibroblast but not epithelial cell entry suggests that polymorphisms external to certain gH/gL epitopes may influence antibody neutralization during fibroblast but not epithelial cell entry. These findings may have implications for elucidating the mechanisms of CMV entry, spread, and antibody evasion and may assist in determining which antibodies may be most efficacious following active immunization or passive administration.IMPORTANCE Cytomegalovirus (CMV) is a significant cause of birth defects among newborns infected in utero and morbidity and mortality in transplant and AIDS patients. Monoclonal antibodies

  14. Effectiveness of basic fibroblast growth factor for pediatric hand burns.

    Science.gov (United States)

    Hayashida, Kenji; Fujioka, Masaki; Morooka, Shin; Saijo, Hiroto; Akita, Sadanori

    2016-11-01

    Pediatric hand deep dermal and deep burns may lead to serious hand deformity with functional impairment and result in an esthetically unfavorable outcome. Since there is no guideline regarding the use of growth factors for pediatric hand burns, we sought to investigate the effectiveness of an angiogenic and regenerative growth factor, basic fibroblast growth factor (bFGF). Consecutive series of second degree or third degree palmer burns at less than 3 years of age seen from January 2010 to June 2014 were included for evaluation at 6 months post-wound healing. The bFGF treatment started from just after injury and continued up to 21 days. Each patient had their scars scored using the Vancouver Scar Scale (VSS) at 6 months after wound healing. There were 34 children with 49 acute palmar burns. The mean healing period was 13.5 ± 4.3 days (7-44 days) and 43 wounds healed within 21 days. There was no need of additional surgery in the 43 wounds, healed within 21 days. In comparison to the wounds for which healing took more than 21 days, the wounds that healed within 21 days demonstrated significantly better pigmentation, pliability, and height according to the VSS (p pediatric palmer burns. Copyright © 2016 Tissue Viability Society. Published by Elsevier Ltd. All rights reserved.

  15. Characterization of human fibroblastic reticular cells as potential immunotherapeutic tools.

    Science.gov (United States)

    Valencia, Jaris; Jiménez, Eva; Martínez, Víctor G; Del Amo, Beatriz G; Hidalgo, Laura; Entrena, Ana; Fernández-Sevilla, Lidia M; Del Río, Francisco; Varas, Alberto; Vicente, Ángeles; Sacedón, Rosa

    2017-05-01

    Fibroblastic reticular cells (FRCs) are essential players during adaptive immune responses not only as a structural support for the encounter of antigen-presenting cells and naive T lymphocytes but also as a source of modulatory signals. However, little is known about this cell population in humans. To address the phenotypical and functional analysis of human FRCs here we established splenic (SP) and mesenteric lymph node (LN) CD45(-)CD31(-)CD90(+)podoplanin(+) myofibroblastic cell cultures. They shared the phenotypical characteristics distinctive of FRCs, including the expression of immunomodulatory factors and peripheral tissue antigens. Nevertheless, human FRCs also showed particular features, some differing from mouse FRCs, like the lack of nitric oxide synthase (NOS2) expression after interferon (IFN)γstimulation. Interestingly, SP-FRCs expressed higher levels of interleukin (IL)-6, BMP4, CCL2, CXCL12 and Notch molecules, and strongly adapted their functional profile to lipopolysaccharide (LPS), polyinosinic:polycytidylic acid (Poly I:C) and IFNγ stimulation. In contrast, we found higher expression of transforming growth factor (TGF)β and Activin A in LN-FRCs that barely responded via Toll-Like Receptor (TLR)3 and constitutively expressed retinaldehyde dehydrogenase 1 enzyme, absent in SP-FRCs. This study reveals human FRCs can be valuable models to increase our knowledge about the physiology of human secondary lymphoid organs in health and disease and to explore the therapeutic options of FRCs. Copyright © 2017 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

  16. Mercury-induced micronuclei in skin fibroblasts of beluga whales

    Energy Technology Data Exchange (ETDEWEB)

    Gauthier, J.M.; Dubeau, H.; Rassart, E. [Univ. du Quebec, Montreal, Quebec (Canada). Dept. des Sciences Biologiques

    1998-12-01

    Beluga whales (Delphinapterus leucas) inhabiting the St. Lawrence estuary are highly contaminated with environmental pollutants and have a high incidence of cancer. Environmental contaminants may be partly responsible for the high incidence of cancer observed in this population. DNA damage plays an important role in the development of cancer. The micronuclei assay was used to test the genotoxic potential of mercury compounds in skin fibroblasts of an Arctic beluga whale. Both mercuric chloride (Hg) and methylmercury (MeHg) induced a highly significant dose-response increase of micronucleated cells. Statistically significant increases in micronucleated cells were observed for 0.5, 5, and 20 {micro}g/ml Hg and 0.05, 0.5, and 2 {micro}g/ml MeHg when compared to control cultures. Concentrations of 0.5, 5, and 20 {micro}g/ml Hg induced a two-, three- and fourfold increase of micronucleated cells, respectively. Treatment with MeHg was one order of magnitude more potent in inducing micronuclei and in inhibiting cell proliferation than Hg. Although results of this in vitro study do not imply that mercury compounds are involved in the etiology of cancer in St. Lawrence beluga whales, significant increases in micronuclei frequency were found at low concentrations of MeHg that are believed to be comparable to concentrations present in certain whales of this population.

  17. Enhanced G2 chromatid radiosensitivity in dyskeratosis congenita fibroblasts

    Energy Technology Data Exchange (ETDEWEB)

    DeBauche, D.M.; Pai, G.S.; Stanley, W.S. (Medical Univ. of South Carolina, Charleston (USA))

    1990-02-01

    Dyskeratosis congenita (DC) is an inherited disorder characterized by reticular pigmentation of the skin, dystrophic nails, mucosal leukoplakia, and a predisposition to cancer in early adult life. In the majority of cases, DC is an X-linked recessive trait. However, one or more autosomal form(s) of DC may exist. Although excessive spontaneous chromatid breakage has been reported in DC, it is not a consistent cytological marker for this disorder. We examined the frequency and specificity of X-irradiation-induced G2 chromatid breakage in fibroblasts from three unrelated DC patients (two males and one female). Metaphase cells from DC patients had significantly more chromatid breaks (16-18-fold and 17-26-fold at 50 and 100 rad X-irradiation, respectively) and chromatid gaps (10-12-fold and 6-7-fold at 50 and 100 rad, respectively) than those from two different controls. Analysis of banded chromosomes revealed a nonrandom distribution of chromatid aberrations in DC but not in controls, a distribution corresponding to some of the known breakpoints for cancer-specific rearrangements, constitutive fragile sites, and/or loci for cellular proto-oncogenes. The significance of this finding for cancer predisposition in DC patients is uncertain, but the increased susceptibility of X-irradiation-induced chromatid breakage may serve as a cellular marker of diagnostic value.

  18. Experimental Myocardial Infarction Upregulates Circulating Fibroblast Growth Factor‐23

    Science.gov (United States)

    Andrukhova, Olena; Slavic, Svetlana; Odörfer, Kathrin I; Erben, Reinhold G

    2015-01-01

    ABSTRACT Myocardial infarction (MI) is a major cause of death worldwide. Epidemiological studies have linked vitamin D deficiency to MI incidence. Because fibroblast growth factor‐23 (FGF23) is a master regulator of vitamin D hormone production and has been shown to be associated with cardiac hypertrophy per se, we explored the hypothesis that FGF23 may be a previously unrecognized pathophysiological factor causally linked to progression of cardiac dysfunction post‐MI. Here, we show that circulating intact Fgf23 was profoundly elevated, whereas serum vitamin D hormone levels were suppressed, after induction of experimental MI in rat and mouse models, independent of changes in serum soluble Klotho or serum parathyroid hormone. Both skeletal and cardiac expression of Fgf23 was increased after MI. Although the molecular link between the cardiac lesion and circulating Fgf23 concentrations remains to be identified, our study has uncovered a novel heart–bone–kidney axis that may have important clinical implications and may inaugurate the new field of cardio‐osteology. © 2015 The Authors. Journal of Bone and Mineral Research published by Wiley Periodicals, Inc. on behalf of American Society for Bone and Mineral Research (ASBMR). PMID:25858796

  19. Fibroblast Growth Factor 21 Mediates Glycemic Regulation by Hepatic JNK

    Directory of Open Access Journals (Sweden)

    Santiago Vernia

    2016-03-01

    Full Text Available The cJun NH2-terminal kinase (JNK-signaling pathway is implicated in metabolic syndrome, including dysregulated blood glucose concentration and insulin resistance. Fibroblast growth factor 21 (FGF21 is a target of the hepatic JNK-signaling pathway and may contribute to the regulation of glycemia. To test the role of FGF21, we established mice with selective ablation of the Fgf21 gene in hepatocytes. FGF21 deficiency in the liver caused marked loss of FGF21 protein circulating in the blood. Moreover, the protective effects of hepatic JNK deficiency to suppress metabolic syndrome in high-fat diet-fed mice were not observed in mice with hepatocyte-specific FGF21 deficiency, including reduced blood glucose concentration and reduced intolerance to glucose and insulin. Furthermore, we show that JNK contributes to the regulation of hepatic FGF21 expression during fasting/feeding cycles. These data demonstrate that the hepatokine FGF21 is a key mediator of JNK-regulated metabolic syndrome.

  20. Fibroblast functionality on novel Ti-30Ta nanotube array

    Energy Technology Data Exchange (ETDEWEB)

    Capellato, Patricia, E-mail: pat_capellato@yahoo.com.br [Department of Materials, Faculty of Engineering Guaratingueta, Sao Paulo State University-UNESP, Av. Ariberto Pereira da Cunha, 333, Pedregulho, CEP 12516-410, Guaratingueta, SP (Brazil); Smith, Barbara S. [School of Biomedical Engineering, Colorado State University, Fort Collins CO 80523 (United States); Popat, Ketul C. [School of Biomedical Engineering, Colorado State University, Fort Collins CO 80523 (United States); Department of Mechanical Engineering, Colorado State University, Fort Collins, CO 80523 (United States); Claro, Ana P.R. Alves [Department of Materials, Faculty of Engineering Guaratingueta, Sao Paulo State University-UNESP, Av. Ariberto Pereira da Cunha, 333, Pedregulho, CEP 12516-410, Guaratingueta, SP (Brazil)

    2012-10-01

    In this study, the mechanical substrate and topographical surface properties of anodized Ti-30Ta alloy were investigated using scanning electron microscopy (SEM), energy-dispersive spectroscopy (EDS) and contact angle measurement. The anodization process was performed in an electrolyte solution containing HF (48%) and H{sub 2}SO{sub 4} (98%) in the volumetric ratios 1:9 with the addition of 5% dimethyl sulfoxide (DMSO) at 15 V, 25 V and 35 V for 20 and 40 min, producing a nanotube architecture when anodized at 35 V for 40 min. Human dermal fibroblasts (HDF, neonatal) were utilized to evaluate the biocompatibility of Ti-30Ta nanotubes and Ti-30Ta alloy after 1 and 3 days of culture. Cellular adhesion, proliferation, viability, cytoskeletal organization and morphology were investigated using fluorescence microscope imaging, biochemical assay and SEM imaging respectively. The results presented identify altered material properties and improved cellular interaction on Ti-30Ta nanotubes as compared to Ti-30 Ta alloy. - Highlights: Black-Right-Pointing-Pointer The surface was modified by anodization, biomimetic treatment and ion bean etching. Black-Right-Pointing-Pointer SEM, EDS and contact angle measurements were used to characterize the surface. Black-Right-Pointing-Pointer Group 5 the most hydrophobic. Black-Right-Pointing-Pointer Group 4 the most hydrophilic. Black-Right-Pointing-Pointer Group 3 and 4 are the more indicated for biomedical application.

  1. Therapeutic Targeting of Fibroblast Growth Factor Receptors in Gastric Cancer

    Directory of Open Access Journals (Sweden)

    Mikito Inokuchi

    2015-01-01

    Full Text Available Chemotherapy has become the global standard treatment for patients with metastatic or unresectable gastric cancer (GC, although outcomes remain unfavorable. Many molecular-targeted therapies inhibiting signaling pathways of various tyrosine kinase receptors have been developed, and monoclonal antibodies targeting human epidermal growth factor receptor 2 (HER2 have become standard therapy for HER2-positive GC. An inhibitor of vascular endothelial growth factor receptor 2 or MET has also produced promising results in patients with GC. Fibroblast growth factor receptors (FGFR play key roles in tumor growth via activated signaling pathways in GC. Genomic amplification of FGFR2 leads to the aberrant activation found in GC tumors and is related to survival in patients with GC. This review discusses the clinical relevance of FGFR in GC and examines FGFR as a potential therapeutic target in patients with GC. Preclinical studies in animal models suggest that multitargeted tyrosine kinase inhibitors (TKIs, including FGFR inhibitor, suppress tumor cell proliferation and delay tumor progression. Several TKIs are now being evaluated in clinical trials as treatment for metastatic or unresectable GC harboring FGFR2 amplification.

  2. MicroRNA-mediated myostatin silencing in caprine fetal fibroblasts.

    Directory of Open Access Journals (Sweden)

    Bushuai Zhong

    Full Text Available Myostatin functions as a negative regulator of skeletal muscle growth by suppressing proliferation and differentiation of myoblasts. Dysfunction of the myostatin gene, either due to natural mutation or genetic manipulations such as knockout or knockdown, has been reported to increase muscle mass in mammalian species. RNA interference (RNAi mediated by microRNAs (miRNAs is a promising method for gene knockdown studies. In the present study, transient and stable silencing of the myostatin gene in caprine fetal fibroblasts (CFF was evaluated using the two most effective constructs selected from four different miRNA expression constructs screened in 293FT cells. Using these two miRNA constructs, we achieved up to 84% silencing of myostatin mRNA in transiently transfected CFF cells and up to 31% silencing in stably transfected CFF cells. Moreover, off-target effects due to induction of interferon (IFN response genes, such as interferon beta (IFN-β and 2'-5'-oligoadenylate synthetase 2 (OAS2, were markedly fewer in stably transfected CFF cells than in transiently transfected cells. Stable expression of anti-myostatin miRNA with minimal induction of interferon shows great promise for increasing muscle mass in transgenic goats.

  3. Selective conversion of fibroblasts into peripheral sensory neurons.

    Science.gov (United States)

    Blanchard, Joel W; Eade, Kevin T; Szűcs, Attila; Lo Sardo, Valentina; Tsunemoto, Rachel K; Williams, Daniel; Sanna, Pietro Paolo; Baldwin, Kristin K

    2015-01-01

    Humans and mice detect pain, itch, temperature, pressure, stretch and limb position via signaling from peripheral sensory neurons. These neurons are divided into three functional classes (nociceptors/pruritoceptors, mechanoreceptors and proprioceptors) that are distinguished by their selective expression of TrkA, TrkB or TrkC receptors, respectively. We found that transiently coexpressing Brn3a with either Ngn1 or Ngn2 selectively reprogrammed human and mouse fibroblasts to acquire key properties of these three classes of sensory neurons. These induced sensory neurons (iSNs) were electrically active, exhibited distinct sensory neuron morphologies and matched the characteristic gene expression patterns of endogenous sensory neurons, including selective expression of Trk receptors. In addition, we found that calcium-imaging assays could identify subsets of iSNs that selectively responded to diverse ligands known to activate itch- and pain-sensing neurons. These results offer a simple and rapid means for producing genetically diverse human sensory neurons suitable for drug screening and mechanistic studies.

  4. Treatment of Skin Avulsion Injuries with Basic Fibroblast Growth Factor

    Directory of Open Access Journals (Sweden)

    Hajime Matsumine, MD, PhD

    2015-04-01

    Full Text Available Summary: This report describes favorable outcomes in 9 patients with skin avulsion injuries of the extremities who underwent full-thickness skin grafting and basic fibroblast growth factor (bFGF application. Following removal of contaminated subcutaneous fat tissue on the inside of skin, the avulsed skin was processed into a full-thickness skin graft, with as much of the skin used as possible irrespective of damage. Several drainage holes (5–10 mm in diameter were made on the graft for drainage from the graft bed and to prevent seroma and hematoma formation. Genetically recombinant human bFGF was sprayed at a dose of 1 μg/cm2 onto the graft bed, which was then covered with the graft and sutured. Pressure immobilization with ointment gauzes and elastic bandages was administered for 1 week postoperatively, and the surface of the skin grafts that did not take was scraped away, preserving the revascularized dermal component on the debrided raw surface as much as possible. bFGF was sprayed again onto the debrided surface to promote epithelialization. Wound closure was achieved in all cases with conservative therapy. The surgical procedure was effective in preventing postoperative ulcer formation and scar contracture and resulted in wound healing with the formation of good-quality, flexible scars.

  5. Fascia implantation with fibroblast growth factor on vocal fold paralysis.

    Science.gov (United States)

    Nagai, Hiromi; Nishiyama, Koichiro; Seino, Yutomo; Kimura, Yu; Tabata, Yasuhiko; Okamoto, Makito

    2013-01-01

    The purpose of this prospective study was to determine the effect of autologous transplantation of fascia into the vocal fold (ATFV) with controlled release of basic fibroblast growth factor (bFGF) on unilateral vocal fold paralysis (UVFP) in a rat model. Unilateral recurrent laryngeal nerve (RLN) section was performed on 15 rats. Ten rats received an autologous fascia implant and gelatin hydrogel with or without bFGF (1 μg) to their larynxes (fascia only, "fascia group"; bFGF + fascia, "fascia + bFGF group"), while the rest underwent RLN transection ("RLN section group"). Four months later, evaluation of the laryngeal glottal gap and histological analysis were performed. The glottal gap was significantly reduced in the fascia + bFGF group, and fat volume increased significantly relative to the RLN section. The volume of the remaining fascia in the bFGF + fascia group was significantly greater than that of the fascia group. ATFV with controlled release of bFGF may compensate for diminished laryngeal volume in UVFP by reducing resorption of the implanted fascia and increasing fat volume. Our findings suggest that this modality may represent an attractive option for treating UVFP. Copyright © 2013 Elsevier Inc. All rights reserved.

  6. Morphometric Approach to Pulp Fibroblast Development in Tooth Germ

    Directory of Open Access Journals (Sweden)

    Irina-Draga Căruntu

    2014-01-01

    Full Text Available This paper builds a morphometric framework for the analysis of dental pulp fibroblast evolution during tooth development. We investigated 15 tooth germs (cases organized, by histological criteria, in three groups corresponding to cap, early bell, and late bell stages, respectively. Each group comprised five cases. The morphometric description used the following parameters: area A, perimeter P—automatically extracted by a color segmentation technique, and form factor (FF—calculated as 4πA/P2. The designed framework operated at inter- and intragroup levels. The intergroup analysis quantified the differences between groups, in the sense of a relative distance (RD adequately defined by mean-value scaling. We showed that the stage of early bell is approximately 5 times closer to late bell than to cap. The quantification procedure required concomitant information about A, P parameters (as P versus A dependences, or FF values, whereas the procedure failed for A or P separately used. The intragroup analysis quantified the similarity of the cases belonging to the same stage. We proved that, unlike the intergroup tests, the individual exploitation of all three descriptors A, P, and FF is effective, yielding highly compatible results. Within any group, most cases presented RDs less than 10% from the group mean value, regardless of the descriptor type.

  7. Fibroblast growth factor signaling in mammalian tooth development.

    Science.gov (United States)

    Li, Chun-Ying; Prochazka, Jan; Goodwin, Alice F; Klein, Ophir D

    2014-01-01

    In this review, we discuss the central role of fibroblast growth factor (FGF) signaling in mammalian tooth development. The FGF family consists of 22 members, most of which bind to four different receptor tyrosine kinases, which in turn signal through a cascade of intracellular proteins. This signaling regulates a number of cellular processes, including proliferation, differentiation, cell adhesion and cell mobility. FGF signaling first becomes important in the presumptive dental epithelium at the initiation stage of tooth development, and subsequently, it controls the invagination of the dental epithelium into the underlying mesenchyme. Later, FGFs are critical in tooth shape formation and differentiation of ameloblasts and odontoblasts, as well as in the development and homeostasis of the stem cell niche that fuels the continuously growing mouse incisor. In addition, FGF signaling is critical in human teeth, as mutations in genes encoding FGF ligands or receptors result in several congenital syndromes characterized by alterations in tooth number, morphology or enamel structure. The parallel roles of FGF signaling in mouse and human tooth development demonstrate the conserved importance of FGF signaling in mammalian odontogenesis.

  8. Clonal analysis of stable chromosome rearrangements in Bloom's syndrome fibroblasts.

    Science.gov (United States)

    Hoehn, H; Salk, D

    1984-04-01

    In situ cytogenetic analysis was performed on colonies derived from single cells of cultured skin fibroblast-like strains from two patients with Bloom's syndrome (GM 1492 and GM 2520). Metaphases in all of the colonies displayed structural chromosome rearrangements. Among 212 metaphases from 24 colonies of GM 1492, only 16% were pseudodiploid, and there was a high incidence of de novo rearrangements within individual colonies. There were two "families" of 16 and five colonies, respectively, each containing identical or related aneusomies, and these could be arranged into pedigrees showing clonal evolution. The heterochromatic region of chromosome #1 and the telomeric regions of chromosome arms 2q, 3q, 4p, and 11q were most frequently involved in the rearrangements. In contrast, strain GM 2520 showed less intraclonal variation, was primarily pseudodiploid, and displayed only three clonal types, one of which had extensive subclonal variation (19 of 24 clones). A remarkable finding in GM 2520 was that, in some clones, extra copies of specific chromosome segments were present as translocations. These results caution against the use of strain GM 1492 as a prototype Bloom's syndrome strain for cell biological studies.

  9. Study of cultured fibroblasts in vivo using NMR

    Energy Technology Data Exchange (ETDEWEB)

    Karczmar, G.S.

    1984-01-01

    The goal of this thesis was to study the compartmentation of phosphorylated glycolytic intermediates in intact Chicken Embryo Fibroblasts (CEFs) using /sup 31/P NMR at 109 MHz. Because glycolysis is regulated differently in normal and virally transformed CEFs, NMR experiments were performed on both types of cells. A technique for maintaining functional cells at high densities in an NMR magnet is described. Signals were detected from cytoplasmic inorganic phosphate (P/sub i/), ATP, NAD, NADH, phosphorylcholine and phosphorylethanolamine. The effect of external glucose on cytoplasmic pools of phosphates was studied. However, experiments with /sup 32/P labelled P/sub i/ showed that as the concentration of glucose in the medium was increased, the amount of phosphate sequestered in the cells increased. They conclude that there is a pool of P/sub i/ which is not detected by high resolution of NMR and that the size of this pool increases as the rate of glycolysis increases. These effects were found only in cultured cells; the data for transformed and normal cells were similar. Longitudinal relaxation times of intracellular phosphates in normal, transformed, and primary CEFs were measured.

  10. Study of cultured fibroblasts in vivo using NMR

    Energy Technology Data Exchange (ETDEWEB)

    Karczmar, G.S.

    1984-08-01

    The goal was to study the compartmentation of phosphorylated glycolytic intermediates in intact Chicken Embryo Fibroblasts (CEFs) using /sup 31/P NMR at 109 MHz. A technique for maintaining functional cells at high densities in an NMR magnet is described. Signals were detected from cytoplasmic inorganic phosphate (P/sub i/), ATP, NAD, NADH, phosphorylcholine and phosphorylethanolamine. The effect of external glucose on cytoplasmic pools of phosphates was studied. When cells were perfused with glucose-free medium the rate of glycolysis decreased, the amplitudes of the ATP resonances decreased, and the P/sub i/ intensity increased. The quantity of NMR-detectable P/sub i/ produced was significantly greater than the quantity of NMR-detectable ATP which was lost. Experiments with /sup 32/P labeled P/sub i/ showed that as the concentration of glucose in the medium was increase, the amount of phosphate sequestered in the cells increased. We conclude that there is a pool of P/sub i/ which is not detected by high resolution NMR and that the size of this pool increases as the rate of glycolysis increase. Longtitudinal relaxation times of intracellular phosphates in normal, transformed, and primary CEFs were measured. The results demonstrate that relaxation times of phosphates are sensitive to structural and metabolic changes which occur when cells are grown in culture. 59 references. 31 figures.

  11. The role of fibroblast growth factor 21 in atherosclerosis.

    Science.gov (United States)

    Kokkinos, John; Tang, Shudi; Rye, Kerry-Anne; Ong, Kwok Leung

    2017-02-01

    The metabolic properties of the endocrine fibroblast growth factor 21 (FGF21) have been extensively studied in the past decade. Previous studies have demonstrated the lipid-lowering, anti-inflammatory and anti-oxidant properties of FGF21. FGF21 is mainly secreted in the liver and adipose tissue in response to a range of physiological and pathological stimuli. In animal and in vitro studies, FGF21 has been shown to improve lipid profiles and inhibit key processes in the pathogenesis of atherosclerosis. It exerts its effects on the cardiovascular system via adiponectin dependent and independent mechanisms. However, the signalling pathways by which FGF21 exerts its effects on endothelial cells remains unknown and needs to be further investigated. The elevation of circulating FGF21 levels in cardiovascular disease has also raised questions as to whether FGF21 can be used as a biomarker to predict subclinical atherosclerosis and cardiovascular events. Recent findings from population studies must be validated in independent cohorts before FGF21 can be used as a biomarker in the clinical setting. The anti-atherosclerotic effects of FGF21 have been investigated in two recent clinical trials, where treatment with an FGF21 analog significantly improved the cardiometabolic profile in obese patients with type 2 diabetes. This review will evaluate recent advances that suggest there may be a role for FGF21 in atherosclerosis. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  12. Basal Cell Carcinoma in Gorlin's Patients: a Matter of Fibroblasts-Led Protumoral Microenvironment?

    Science.gov (United States)

    Gache, Yannick; Brellier, Florence; Rouanet, Sophie; Al-Qaraghuli, Sahar; Goncalves-Maia, Maria; Burty-Valin, Elodie; Barnay, Stéphanie; Scarzello, Sabine; Ruat, Martial; Sevenet, Nicolas; Avril, Marie-Françoise; Magnaldo, Thierry

    2015-01-01

    Basal cell carcinoma (BCC) is the commonest tumor in human. About 70% sporadic BCCs bear somatic mutations in the PATCHED1 tumor suppressor gene which encodes the receptor for the Sonic Hedgehog morphogen (SHH). PATCHED1 germinal mutations are associated with the dominant Nevoid Basal Cell Carcinoma Syndrome (NBCCS), a major hallmark of which is a high susceptibility to BCCs. Although the vast majority of sporadic BCCs arises exclusively in sun exposed skin areas, 40 to 50% BCCs from NBCCS patients develop in non photo-exposed skin. Since overwhelming evidences indicate that microenvironment may both be modified by- and influence the- epithelial tumor, we hypothesized that NBCCS fibroblasts could contribute to BCCs in NBCCS patients, notably those developing in non photo-exposed skin areas. The functional impact of NBCCS fibroblasts was then assessed in organotypic skin cultures with control keratinocytes. Onset of epidermal differentiation was delayed in the presence of primary NBCCS fibroblasts. Unexpectedly, keratinocyte proliferation was severely reduced and showed high levels of nuclear P53 in both organotypic skin cultures and in fibroblast-led conditioning experiments. However, in spite of increased levels of senescence associated β-galactosidase activity in keratinocytes cultured in the presence of medium conditioned by NBCCS fibroblasts, we failed to observe activation of P16 and P21 and then of bona fide features of senescence. Constitutive extinction of P53 in WT keratinocytes resulted in an invasive phenotype in the presence of NBCCS fibroblasts. Finally, we found that expression of SHH was limited to fibroblasts but was dependent on the presence of keratinocytes. Inhibition of SHH binding resulted in improved epidermal morphogenesis. Altogether, these data suggest that the repertoire of diffusible factors (including SHH) expressed by primary NBCCS fibroblasts generate a stress affecting keratinocytes behavior and epidermal homeostasis. Our findings

  13. Combined effects of interleukin-1? and cyclic stretching on metalloproteinase expression in corneal fibroblasts in vitro

    OpenAIRE

    Feng, Pengfei; Li, Xiaona; Chen, Weiyi; Liu, Chengxing; Rong, Shuo; Wang, Xiaojun; Du, Genlai

    2016-01-01

    Background Corneal tensile strain increases if the cornea becomes thin or if intraocular pressure increases. However, the effects of mechanical stress on extracellular matrix (ECM) remodelling in the corneal repair process and the corneal anomalies are unknown. Methods In this study, the combined effects of interleukin-1? (IL-1?) on matrix metalloproteinases (MMPs) in corneal fibroblasts under cyclic stretching were investigated in vitro. Cultured rabbit corneal fibroblasts were subjected to ...

  14. Fibroblast growth factor 1 ameliorates diabetic nephropathy by an anti-inflammatory mechanism.

    Science.gov (United States)

    Liang, Guang; Song, Lintao; Chen, Zilu; Qian, Yuanyuan; Xie, Junjun; Zhao, Longwei; Lin, Qian; Zhu, Guanghui; Tan, Yi; Li, Xiaokun; Mohammadi, Moosa; Huang, Zhifeng

    2018-01-01

    Inflammation plays a central role in the etiology of diabetic nephropathy, a global health issue. We observed a significant reduction in the renal expression of fibroblast growth factor 1, a known mitogen and insulin sensitizer, in patients with diabetic nephropathy and in mouse models implying that fibroblast growth factor 1 possesses beneficial anti-inflammatory and renoprotective activities in vivo. To test this possibility, we investigated the effects of chronic intraperitoneal administration of fibroblast growth factor 1 into both the streptozotocin-induced type 1 diabetes and db/db type 2 diabetes models. Indeed, recombinant fibroblast growth factor 1 significantly suppressed renal inflammation (i.e., cytokines, macrophage infiltration), glomerular and tubular damage, and renal dysfunction in both type 1 and type 2 diabetes mice. Fibroblast growth factor 1 was able to correct the elevated blood glucose levels in type 2 but not in type 1 diabetic mice, suggesting that the anti-inflammatory effect of fibroblast growth factor 1 was independent of its glucose-lowering activity. The mechanistic study demonstrated that fibroblast growth factor 1-mediated inhibition of the renal inflammation in vivo was accompanied by attenuation of the nuclear factor κB and c-Jun N-terminal kinase signaling pathways, further validated in vitro using cultured glomerular mesangial cells and podocytes. Thus, fibroblast growth factor 1 holds great promise for developing new treatments for diabetic nephropathy through countering inflammatory signaling cascades in injured renal tissue. Copyright © 2017 International Society of Nephrology. Published by Elsevier Inc. All rights reserved.

  15. Functional deficiency of fibroblasts heterozygous for Bloom syndrome as specific manifestation of the primary defect.

    OpenAIRE

    Bartram, C R; Rüdiger, H W; Schmidt-Preuss, U; Passarge, E

    1981-01-01

    The effect on the rate of sister chromatid exchanges (SCEs) in Bloom syndrome fibroblasts by cocultivation with Fanconi anemia and xeroderma pigmentosum fibroblasts and with Bloom syndrome heterozygotes was studied. Cells of Fanconi anemia and xeroderma origin reduced the rate of SCEs in Bloom cells by about 45%-50%, just as control cells do. In contrast, heterozygous Bloom cells reduced the rate of SCEs by only 16%-28%. In absolute figures, Fanconi cells reduced the mean rate of SCE in Bloom...

  16. Evaluation of cytotoxicity and gelatinases activity in 3T3 fibroblast cell by root repair materials

    OpenAIRE

    Varol Basak; Tuna Elif Bahar; Karsli Emine; Kasimoglu Yelda; Koruyucu Mine; Seymen Figen; Nurten Rustem

    2016-01-01

    The aim of this study was to investigate the effects of calcium silicate-based products on cytotoxicity in the 3T3 fibroblast and gelatinolytic activity of matrix metalloproteinases (MMPs). 3T3 fibroblasts were incubated directly with Ortho Mineral trioxide aggregate (MTA), BioAggregate, Biodentine, MTA Plus, MTA Angelus and MTA Cerkamed for 24 hours and seven days. The cytotoxicity was determined using an MTT assay. Supernatants were collected to determine MMP-2 and MMP-9. Data were analysed...

  17. Granulocyte macrophage colony stimulating factor (GM-CSF biological actions on human dermal fibroblasts

    Directory of Open Access Journals (Sweden)

    S Montagnani

    2009-12-01

    Full Text Available Fibroblasts are involved in all pathologies characterized by increased ExtraCellularMatrix synthesis, from wound healing to fibrosis. Granulocyte Macrophage-Colony Stimulating Factor (GM-CSF is a cytokine isolated as an hemopoietic growth factor but recently indicated as a differentiative agent on endothelial cells. In this work we demonstrated the expression of the receptor for GM-CSF (GMCSFR on human normal skin fibroblasts from healthy subjects (NFPC and on a human normal fibroblast cell line (NHDF and we try to investigate the biological effects of this cytokine. Human normal fibroblasts were cultured with different doses of GM-CSF to study the effects of this factor on GMCSFR expression, on cell proliferation and adhesion structures. In addition we studied the production of some Extra-Cellular Matrix (ECM components such as Fibronectin, Tenascin and Collagen I. The growth rate of fibroblasts from healthy donors (NFPC is not augmented by GM-CSF stimulation in spite of increased expression of the GM-CSFR. On the contrary, the proliferation of normal human dermal fibroblasts (NHDF cell line seems more influenced by high concentration of GM-CSF in the culture medium. The adhesion structures and the ECM components appear variously influenced by GM-CSF treatment as compared to fibroblasts cultured in basal condition, but newly only NHDF cells are really induced to increase their synthesis activity. We suggest that the in vitro treatment with GM-CSF can shift human normal fibroblasts towards a more differentiated state, due or accompanied by an increased expression of GM-CSFR and that such “differentiation” is an important event induced by such cytokine.

  18. Breast Cancer-Associated Fibroblasts: Where We Are and Where We Need to Go

    OpenAIRE

    Buchsbaum, Rachel J.; Oh, Sun Young

    2016-01-01

    Cancers are heterogeneous tissues comprised of multiple components, including tumor cells and microenvironment cells. The tumor microenvironment has a critical role in tumor progression. The tumor microenvironment is comprised of various cell types, including fibroblasts, macrophages and immune cells, as well as extracellular matrix and various cytokines and growth factors. Fibroblasts are the predominant cell type in the tumor microenvironment. However, neither the derivation of tissue-speci...

  19. Proliferative and inductive effects of Cyclosporine a on gingival fibroblast of child and adult

    Directory of Open Access Journals (Sweden)

    Bahareh Nazemi Salman

    2013-01-01

    Conclusions: The mechanism of a CsA-induced fibroblast overgrowth may converge on the steps involving fibroblast proliferation and cytokine network including IL-6, IL-8, IL-1β, TGF-β1, and PGE 2 , in both adults and pediatrics. As the prevalence and intensity of drug-induced gingival overgrowth is more serious in the pediatrics. As group than in adults, we suggest that more studies be conducted on the pediatric group.

  20. The fibroblast Tiam1-osteopontin pathway modulates breast cancer invasion and metastasis.

    Science.gov (United States)

    Xu, Kun; Tian, Xuejun; Oh, Sun Y; Movassaghi, Mohammad; Naber, Stephen P; Kuperwasser, Charlotte; Buchsbaum, Rachel J

    2016-01-28

    The tumor microenvironment has complex effects in cancer pathophysiology that are not fully understood. Most cancer therapies are directed against malignant cells specifically, leaving pro-malignant signals from the microenvironment unaddressed. Defining specific mechanisms by which the tumor microenvironment contributes to breast cancer metastasis may lead to new therapeutic approaches against advanced breast cancer. We use a novel method for manipulating three-dimensional mixed cell co-cultures, along with studies in mouse xenograft models of human breast cancer and a histologic study of human breast cancer samples, to investigate how breast cancer-associated fibroblasts affect the malignant behaviors of breast cancer cells. Altering fibroblast Tiam1 expression induces changes in invasion, migration, epithelial-mesenchymal transition, and cancer stem cell characteristics in associated breast cancer cells. These changes are both dependent on fibroblast secretion of osteopontin and also long-lasting even after cancer cell dissociation from the fibroblasts, indicating a novel Tiam1-osteopontin pathway in breast cancer-associated fibroblasts. Notably, inhibition of fibroblast osteopontin with low doses of a novel small molecule prevents lung metastasis in a mouse model of human breast cancer metastasis. Moreover, fibroblast expression patterns of Tiam1 and osteopontin in human breast cancers show converse changes correlating with invasion, supporting the hypothesis that this pathway in tumor-associated fibroblasts regulates breast cancer invasiveness in human disease and is thus clinically relevant. These findings suggest a new therapeutic paradigm for preventing breast cancer metastasis. Pro-malignant signals from the tumor microenvironment with long-lasting effects on associated cancer cells may perpetuate the metastatic potential of developing cancers. Inhibition of these microenvironment signals represents a new therapeutic strategy against cancer metastasis that

  1. Malignant transformation of human fibroblasts caused by expression of a transfected T24 HRAS oncogene.

    OpenAIRE

    Hurlin, P J; Maher, V M; McCormick, J J

    1989-01-01

    We showed previously that diploid human fibroblasts that express a transfected HRAS oncogene from the human bladder carcinoma cell line T24 exhibit several characteristics of transformed cells but do not acquire an infinite life-span and are not tumorigenic. To extend these studies of the T24 HRAS in human cells, we have utilized an infinite life-span, but otherwise phenotypically normal, human fibroblast cell strain, MSU-1.1, developed in this laboratory after transfection of diploid fibrobl...

  2. Evidence of two distinct functionally specialized fibroblast lineages in breast stroma

    DEFF Research Database (Denmark)

    Morsing, Mikkel; Klitgaard, Marie Christine; Jafari Kermani, Abbas

    2016-01-01

    breast. We here demonstrate the impact on epithelium of two lineages of breast stromal fibroblasts, one of which accumulates in the TDLU while the other resides outside the TDLU in the interlobular stroma. Methods The two lineages are prospectively isolated by fluorescence activated cell sorting (FACS...... remain phenotypically stable and functionally distinct in culture. Lobular fibroblasts have properties in common with bone marrow derived mesenchymal stem cells and they specifically convey growth and branching morphogenesis of epithelial progenitors. Conclusions Two distinct functionally specialized...

  3. Acquired Localized Cutis Laxa due to Increased Elastin Turnover

    DEFF Research Database (Denmark)

    Nygaard, Rie Harboe; Maynard, Scott; Schjerling, Peter

    2016-01-01

    obtained skin biopsies from affected and unaffected skin areas of the patient and analyzed these with microscopy, polymerase chain reaction, western blotting and cell culture experiments. Skin from the affected area lacked elastin fibers in electron microscopy but had higher mRNA expression of elastin....... In conclusion, we report a case of acquired localized cutis laxa with a lack of elastic fibers in the skin of the patient's upper body. The lack of elastic fibers in the affected skin was combined with increased mRNA expression and protein levels of elastin. These findings indicate that elastin synthesis...... and total RNA. Levels of an apparent tropoelastin degradation product were higher in the affected area. Fibroblast cultures from the affected area were able to produce elastin and showed higher proliferation and survival after oxidative and UVB stress compared to fibroblasts from the unaffected area...

  4. Capparis spinosa protects against oxidative stress in systemic sclerosis dermal fibroblasts.

    Science.gov (United States)

    Cao, Yue-Lan; Li, Xin; Zheng, Min

    2010-07-01

    High reactive oxygen species (ROS), Ha-Ras, and active ERK1/2 in fibroblasts play an essential role in the pathogenesis of systemic sclerosis (SSc). The present study was carried out to evaluate the effects of the ethanol extract from fruits of Capparis Spinosa L. (ECS) on oxidative stress and ROS-ERK1/2-Ha-Ras signal loop in SSc dermal fibroblasts in vitro. Cultured dermal fibroblasts from three SSc patients and three normal controls were treated with ECS by different concentration (10, 50, 100 microg/ml). ECS significantly reduced the production of O2(-), H2O2, and ROS in SSc fibroblasts in a dose-dependent manner. ECS effectively minimized the loss of cell viability and apoptosis induced by H2O2 in normal and SSc fibroblasts. Furthermore, the protective effect of ECS on SSc fibroblasts was more significant than on normal ones. ECS decreased the expression of P-ERK1/2 and Ha-Ras in a dose-dependent manner. In conclusion, ECS exhibits a notable activity in protecting against oxidative stress and interrupting of ROS-ERK1/2-Ha-Ras signal loop in SSc, suggesting its potential protective effects against skin sclerosis.

  5. Platelet-Derived Growth Factor-BB Enhances Adipogenesis in Orbital Fibroblasts.

    Science.gov (United States)

    Virakul, Sita; Dalm, Virgil A S H; Paridaens, Dion; van den Bosch, Willem A; Mulder, Monique T; Hirankarn, Nattiya; van Hagen, P Martin; Dik, Willem A

    2015-08-01

    Platelet-derived growth factor (PDGF)-BB has been identified as important factor in pathogenesis of Graves' ophthalmopathy (GO). It stimulates proliferation, cytokine, and hyaluronan production, and thyrotropin receptor expression by orbital fibroblasts. Therefore, the PDGF-pathway has been proposed as a target for pharmacological intervention in GO. However, increased adipogenesis is another major pathological characteristic of GO and it is unknown whether this is affected by PDGF-BB. The aim of this study was to investigate the effect of PDGF-BB on adipocyte differentiation by orbital fibroblasts. Orbital fibroblasts from five healthy controls and nine GO patients were collected. Adipogenesis was induced by culturing orbital fibroblasts in differentiation medium, either in the presence or absence of PDGF-BB. Adipogenesis was determined by Oil-Red-O staining, triglyceride measurement, and peroxisome proliferator-activated receptor (PPAR)-γ mRNA expression. Platelet-derived growth factor-BB significantly enhanced adipocyte differentiation by orbital fibroblasts (Oil-Red-O staining [P BB on adipogenesis was independent of autocrine IL-6 signaling as it was not abrogated by IL-6-receptor-α neutralizing antibody. The clinically applicable tyrosine kinase inhibitor dasatinib and tyrphostin AG1296, which both block PDGF receptor tyrosine kinase activity, inhibited PDGF-BB-enhanced adipogenesis (P BB enhances adipogenesis in orbital fibroblasts, and, thus, may contribute to adipose tissue expansion in GO. Therefore, the PDGF-signaling cascade may represent a target of therapy to interfere with adipogenesis in GO.

  6. Gamma-ray-enhanced reactivation of irradiated adenovirus in Xeroderma pigmentosum and Cockayne syndrome fibroblasts

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    Jeeves, W.P.; Rainbow, A.J.

    1983-06-01

    A ..gamma..-ray-enhanced reactivation (..gamma..RER) of uv-irradiated as well as of ..gamma..-irradiated human adenovirus type 2 (Ad 2) was detected following infection of normal, Xeroderma pigmentosum (XP), and Cockayne syndrome (CS) fibroblasts that had been preirradiated with ..gamma.. rays. Gamma-irradiated or nonirradiated fibroblasts were infected with either nonirradiated or irradiated Ad 2, and 48 hr after infection cells were examined for the presence of viral structural antigens (Vag) using immunofluorescent staining. Results obtained using seven different normal fibroblast strains showed that irradiation of host monolayers with 1 krad immediately prior to infection resulted in a ..gamma..RER factor +-SE of 4.5 +- 1.6 for uv-irradiated virus and 4.3 +- 1.3 for ..gamma..-irradiated virus. CS fibroblasts, as well as excision repair-deficient XP fibroblasts from complementation groups A and D, were all found to be capable of expressing ..gamma..RER of irradiated Ad 2. XP variant cells expressed lower levels of ..gamma..RER compared to most normal strains, suggesting a possible role for cellular postreplication repair in the mechanism responsible for ER in human cells. An excision-deficient XP fibroblast strain belonging to complementation group A, but derived from a patient afflicted with the severe De Sanctis-Cacchione form of XP, although proficient in ..gamma..RER of ..gamma..-irradiated Ad 2, yielded barely detectable levels of ..gamma..RER for uv-irradiated Ad 2.

  7. Cellular Mechanics of Primary Human Cervical Fibroblasts: Influence of Progesterone and a Pro-inflammatory Cytokine.

    Science.gov (United States)

    Shukla, Vasudha; Barnhouse, Victoria; Ackerman, William E; Summerfield, Taryn L; Powell, Heather M; Leight, Jennifer L; Kniss, Douglas A; Ghadiali, Samir N

    2018-01-01

    The leading cause of neonatal mortality, pre-term birth, is often caused by pre-mature ripening/opening of the uterine cervix. Although cervical fibroblasts play an important role in modulating the cervix's extracellular matrix (ECM) and mechanical properties, it is not known how hormones, i.e., progesterone, and pro-inflammatory insults alter fibroblast mechanics, fibroblast-ECM interactions and the resulting changes in tissue mechanics. Here we investigate how progesterone and a pro-inflammatory cytokine, IL-1β, alter the biomechanical properties of human cervical fibroblasts and the fibroblast-ECM interactions that govern tissue-scale mechanics. Primary human fibroblasts were isolated from non-pregnant cervix and treated with estrogen/progesterone, IL-1β or both. The resulting changes in ECM gene expression, matrix remodeling, traction force generation, cell-ECM adhesion and tissue contractility were monitored. Results indicate that IL-1β induces a significant reduction in traction force and ECM adhesion independent of pre-treatment with progesterone. These cell level effects altered tissue-scale mechanics where IL-1β inhibited the contraction of a collagen gel over 6 days. Interestingly, progesterone treatment alone did not modulate traction forces or gel contraction but did result in a dramatic increase in cell-ECM adhesion. Therefore, the protective effect of progesterone may be due to altered adhesion dynamics as opposed to altered ECM remodeling.

  8. [Effects of extracts of Dragon's blood on fibroblast proliferation and extracellular matrix hyaluronic acid].

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    Li, Dan; Hui, Rui; Hu, Yongwu; Han, Yan; Guo, Shuzhong

    2015-01-01

    To investigate the effects of Dragon' s blood extract on proliferation and secret extracellular matrix function of fibroblasts in vitro. Dragon' s blood was extracted by chloroform, acetoacetic ester, alcohol. Human fibroblast were cultured in vitro in media containing gradient dilutions of Dragon' s blood extracts (0.002, 0.02, 0.2, 2, 20 mg/ml) , which was followed by cell proliferation assessed with MTT assay on 0, 12, 24, 36, 48, 60, 72 h. Under the optimal concentration, the cell growth curves were drawn and the flow cytometry (FCM) was used to determine the changes of cell cycle. On 0, 12, 24, 36, 48, 60, 72 h, the concentration of hyaluronic acid in the supernatant of fibroblast culture was measured by radioimmunoassay. 0.2-2 mg/ml Dragon' s blood extracts enhanced the proliferation of fibroblasts in a dose-dependent manner. 2 mg/ml was the optimal dilution of Dragon's blood extract, and it increased the ratio of S cells in cell cycle [(25.80 ± 3.10)%] than control group [(7.50 ± 0.70)%, P < 0.01]. From 12 h to 72 h, in 2 mg/ml Dragon's blood group, concentration of Hyaluronic acid secreted by fibroblasts gradually increased, but were less than control (P < 0.01). Dragon's blood acetoacetic ester extract improved the proliferation of cultured human fibroblasts in vitro, might be beneficial to promote wound healing.

  9. A cell culture model using rat coronary artery adventitial fibroblasts to measure collagen production

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    Meszaros Gary

    2007-05-01

    Full Text Available Abstract Background We have developed a rat cell model for studying collagen type I production in coronary artery adventitial fibroblasts. Increased deposition of adventitial collagen type I leads to stiffening of the blood vessel, increased blood pressure, arteriosclerosis and coronary heart disease. Although the source and mechanism of collagen deposition is yet unknown, the adventitia appears to play a significant role. To demonstrate the application of our cell model, cultured adventitial fibroblasts were treated with sex hormones and the effect on collagen production measured. Methods Hearts (10–12 weeks were harvested and the left anterior descending coronary artery (LAD was isolated and removed. Tissue explants were cultured and cells (passages 2–4 were confirmed as fibroblasts using immunohistochemistry. Optimal conditions were determined for cell tissue harvest, timing, proliferation and culture conditions. Fibroblasts were exposed to 10-7 M testosterone or 10-7 M estrogen for 24 hours and either immunostained for collagen type I or subjected to ELISA. Results Results showed increased collagen staining in fibroblasts treated with testosterone compared to control and decreased staining with estrogen. ELISA results showed that testosterone increased collagen I by 20% whereas estrogen decreased collagen I by 15%. Conclusion Data demonstrates the usefulness of our cell model in studying the specific role of the adventitia apart from other blood vessel tissue in rat coronary arteries. Results suggest opposite effects of testosterone and estrogen on collagen synthesis in the rat coronary artery adventitial fibroblasts.

  10. Optimal Viscosity and Particle Shape of Hyaluronic Acid Filler as a Scaffold for Human Fibroblasts.

    Science.gov (United States)

    Kim, Deok-Yeol; Namgoong, Sik; Han, Seung-Kyu; Won, Chang-Hoon; Jeong, Seong-Ho; Dhong, Eun-Sang; Kim, Woo-Kyung

    2015-07-01

    The authors previously reported that cultured human fibroblasts suspended in a hyaluronic acid filler can produce human dermal matrices with extended in vivo stability in animal and clinical studies. The present study was undertaken to determine the optimal viscosity and particle shape of hyaluronic acid filler as a scaffold for cultured human dermal fibroblasts to enhance the maximal viability of injected cells. The fibroblasts were suspended in either 1 of 3 hyaluronic acid viscosities at 2 different particle shapes. The viscosities used in this study were low (600,000-800,000 centipoises), moderate (2,000,000-4,000,000 centipoises), and high (8,000,000-12,000,000 centipoises). The particle shape was evaluated by testing round and irregular shapes. The fibroblast mixed bioimplants were injected into the back of individual athymic nude mice. The levels of type I collagen were measured using fluorescent-activated cell sorting (FACS) and immunohistochemical staining at 16 weeks after the injections. Results of FACS demonstrated that the mean cell ratio with human collagens in the moderate viscosity group was greater than those of control, low, and high viscosity groups. An immunohistochemical study showed similar results. The moderate viscosity group demonstrated the highest positive staining of human collagens. However, there were no significant differences between groups of irregular and round shape particles. A hyaluronic acid bioimplant with moderate viscosity is superior to that with low or high viscosity in the viability for human fibroblasts. However, the particle shape does not influence the viability of the fibroblasts.

  11. Antimicrobial peptide KSL-W promotes gingival fibroblast healing properties in vitro.

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    Park, Hyun-Jin; Salem, Mabrouka; Semlali, Abdelhabib; Leung, Kai P; Rouabhia, Mahmoud

    2017-07-01

    We investigated the effect of synthetic antimicrobial decapeptide KSL-W (KKVVFWVKFK) on normal human gingival fibroblast growth, migration, collagen gel contraction, and α-smooth muscle actin protein expression. Results show that in addition to promoting fibroblast adhesion by increasing F-actin production, peptide KSL-W promoted cell growth by increasing the S and G2/M cell cycle phases, and enhanced the secretion of metalloproteinase (MMP)-1 and MMP-2 by upregulating MMP inhibitors, such as tissue inhibitors of metalloproteinase (TIMP)-1 and TIMP-2 in fibroblasts. An in vitro wound healing assay confirmed that peptide KSL-W promoted fibroblast migration and contraction of a collagen gel matrix. We also demonstrated a high expression of α-smooth muscle actin by gingival fibroblasts being exposed to KSL-W. This work shows that peptide KSL-W enhances gingival fibroblast growth, migration, and metalloproteinase secretion, and the expression of α-smooth muscle actin, thus promoting wound healing. Copyright © 2017 Elsevier Inc. All rights reserved.

  12. Hepatocyte growth factor improves direct reprogramming of fibroblasts towards endothelial progenitor cells via ETV2 transduction

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    Phuc Van Pham

    2016-09-01

    Full Text Available Human fibroblasts can be differentiated into endothelial progenitor cells by direct reprogramming via ETV-2 transfection. Previously, we have shown that the efficacy of direct reprogramming can be enhanced by hypoxia treatment. In this study, we aim to investigate whether the efficacy of direct reprogramming of fibroblasts into EPCs via Ets variant gene 2 (ETV2 transfection can be increased with hepatocyte growth factor (HGF treatment. Foreskin-derived fibroblasts were cultured in standard medium (DMEM/F12 supplemented with fetal bovine serum. They were then transduced with a viral vector expressing ETV2 in culture medium supplemented with HGF. The transduced fibroblasts were cultured in endothelial cell medium supplemented with HGF for 28 days. The efficacy of direct reprogramming was evaluated based on expression of CD31 and VEGFR2 markers by transduced cells. Phenotypic and functional characterization of induced EPCs were also confirmed by expression of particular genes and in vitro angiogenesis assays. Our results showed that HGF significantly increased the efficacy of direct reprogramming of fibroblasts towards EPCs via ETV2 transcription factors; efficiency increased from 5.41+/-1.51% for ETV2 transduction alone to 12.31+/-2.15% for ETV2 transduction combined with HGF treatment. These findings suggest the rationale for combined use of ETV2 and HGF in direct in vitro reprogramming of fibroblasts into EPCs. [Biomed Res Ther 2016; 3(9.000: 836-843

  13. Expression of TGF-β3 in Isolated Fibroblasts from Foreskin

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    Mahnaz Mahmoudi Rad

    2015-05-01

    Full Text Available Background: The multifunctional transforming growth factor beta (TGF-β is a glycoprotein that exists in three isoforms. TGF-β3 expression increases in fetal wound healing and reduces fibronectin and collagen I and III deposition, and also improves the architecture of the neodermis which is a combination of blood vessels and connective tissue during wound healing. Fibroblasts are key cells in the wound healing process. TGF-β3 plays a critical role in scar-free wound healing and fibroblast actions in the wound healing process. The aim of this study was to express the TGF-β3 gene (tgf-b3 in human foreskin fibroblasts (HFF’s. Methods: We obtained HFF’s from a newborn and a primary fibroblast culture was prepared. The cells were transfected with TGF-β3-pCMV6-XL5 plasmid DNA by both lipofection and electroporation. Expression of TGF-β3 was measured by enzyme-linked immunosorbent assay (ELISA. Results: The highest TGF-β3 expression (8.3-fold greater than control was obtained by lipofection after 72 hours using 3 μl of transfection reagent. Expression was 1.4-fold greater than control by electroporation. Conclusions: In this study, we successfully increased TGF-β3 expression in primary fibroblast cells. In the future, grafting these transfected fibroblasts onto wounds can help the healing process without scarring.

  14. Pentadecapeptide BPC 157 Enhances the Growth Hormone Receptor Expression in Tendon Fibroblasts

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    Chung-Hsun Chang

    2014-11-01

    Full Text Available BPC 157, a pentadecapeptide derived from human gastric juice, has been demonstrated to promote the healing of different tissues, including skin, muscle, bone, ligament and tendon in many animal studies. However, the underlying mechanism has not been fully clarified. The present study aimed to explore the effect of BPC 157 on tendon fibroblasts isolated from Achilles tendon of male Sprague-Dawley rat. From the result of cDNA microarray analysis, growth hormone receptor was revealed as one of the most abundantly up-regulated genes in tendon fibroblasts by BPC 157. BPC 157 dose- and time-dependently increased the expression of growth hormone receptor in tendon fibroblasts at both the mRNA and protein levels as measured by RT/real-time PCR and Western blot, respectively. The addition of growth hormone to BPC 157-treated tendon fibroblasts dose- and time-dependently increased the cell proliferation as determined by MTT assay and PCNA expression by RT/real-time PCR. Janus kinase 2, the downstream signal pathway of growth hormone receptor, was activated time-dependently by stimulating the BPC 157-treated tendon fibroblasts with growth hormone. In conclusion, the BPC 157-induced increase of growth hormone receptor in tendon fibroblasts may potentiate the proliferation-promoting effect of growth hormone and contribute to the healing of tendon.

  15. Oleic, Linoleic and Linolenic Acids Increase ROS Production by Fibroblasts via NADPH Oxidase Activation

    Science.gov (United States)

    Hatanaka, Elaine; Dermargos, Alexandre; Hirata, Aparecida Emiko; Vinolo, Marco Aurélio Ramirez; Carpinelli, Angelo Rafael; Newsholme, Philip; Armelin, Hugo Aguirre; Curi, Rui

    2013-01-01

    The effect of oleic, linoleic and γ-linolenic acids on ROS production by 3T3 Swiss and Rat 1 fibroblasts was investigated. Using lucigenin-amplified chemiluminescence, a dose-dependent increase in extracellular superoxide levels was observed during the treatment of fibroblasts with oleic, linoleic and γ-linolenic acids. ROS production was dependent on the addition of β-NADH or NADPH to the medium. Diphenyleneiodonium inhibited the effect of oleic, linoleic and γ-linolenic acids on fibroblast superoxide release by 79%, 92% and 82%, respectively. Increased levels of p47phox phosphorylation due to fatty acid treatment were detected by Western blotting analyses of fibroblast proteins. Increased p47phox mRNA expression was observed using real-time PCR. The rank order for the fatty acid stimulation of the fibroblast oxidative burst was as follows: γ-linolenic > linoleic > oleic. In conclusion, oleic, linoleic and γ-linolenic acids stimulated ROS production via activation of the NADPH oxidase enzyme complex in fibroblasts. PMID:23579616

  16. Influence of caffeine and hyaluronic acid on collagen biosynthesis in human skin fibroblasts.

    Science.gov (United States)

    Donejko, Magdalena; Przylipiak, Andrzej; Rysiak, Edyta; Głuszuk, Katarzyna; Surażyński, Arkadiusz

    2014-01-01

    The aim of this study was to evaluate the effect of caffeine on collagen biosynthesis in human skin fibroblasts and the influence of hyaluronic acid (HA) on this process. Collagen, [(3)H]-thymidine incorporation, and prolidase activity were measured in confluent human skin fibroblast cultures that had been treated with 1, 2, and 5 mM caffeine and with caffeine and 500 μg/mL HA. Western immunoblot analysis was performed to evaluate expression of β1-integrin receptor, insulin-like growth factor receptor phospho-Akt protein and mitogen-activated protein kinase (phospho-extracellular signal-regulated kinase). Caffeine inhibited collagen biosynthesis in a dose-dependent manner. The mechanism of this process was found at the level of prolidase activity. Caffeine significantly inhibited the enzyme activity. The addition of HA had no effect on collagen biosynthesis or prolidase activity in fibroblasts incubated with caffeine. Caffeine also had an inhibitory effect on DNA biosynthesis. HA, however, did not have any significant effect on this process. The inhibition of the expression of β1-integrin and insulin-like growth factor receptor in fibroblasts incubated with the caffeine indicates a possible mechanism of inhibition of collagen biosynthesis. Caffeine reduces collagen synthesis in human cultured skin fibroblasts. HA did not have any significant protective effect on this process. This is the first study to our knowledge that reports caffeine-induced inhibition of collagen synthesis in human skin fibroblasts.

  17. Pentadecapeptide BPC 157 enhances the growth hormone receptor expression in tendon fibroblasts.

    Science.gov (United States)

    Chang, Chung-Hsun; Tsai, Wen-Chung; Hsu, Ya-Hui; Pang, Jong-Hwei Su

    2014-11-19

    BPC 157, a pentadecapeptide derived from human gastric juice, has been demonstrated to promote the healing of different tissues, including skin, muscle, bone, ligament and tendon in many animal studies. However, the underlying mechanism has not been fully clarified. The present study aimed to explore the effect of BPC 157 on tendon fibroblasts isolated from Achilles tendon of male Sprague-Dawley rat. From the result of cDNA microarray analysis, growth hormone receptor was revealed as one of the most abundantly up-regulated genes in tendon fibroblasts by BPC 157. BPC 157 dose- and time-dependently increased the expression of growth hormone receptor in tendon fibroblasts at both the mRNA and protein levels as measured by RT/real-time PCR and Western blot, respectively. The addition of growth hormone to BPC 157-treated tendon fibroblasts dose- and time-dependently increased the cell proliferation as determined by MTT assay and PCNA expression by RT/real-time PCR. Janus kinase 2, the downstream signal pathway of growth hormone receptor, was activated time-dependently by stimulating the BPC 157-treated tendon fibroblasts with growth hormone. In conclusion, the BPC 157-induced increase of growth hormone receptor in tendon fibroblasts may potentiate the proliferation-promoting effect of growth hormone and contribute to the healing of tendon.

  18. The extracellular matrix modulates fibroblast phenotype and function in the infarcted myocardium

    Science.gov (United States)

    Dobaczewski, Marcin; de Haan, Judith J; Frangogiannis, Nikolaos G

    2012-01-01

    Cardiac fibroblasts are key cellular effectors of cardiac repair; their phenotype and function are modulated by interactions with extracellular matrix proteins. This review manuscript discusses the effects of the extracellular matrix on the inflammatory and reparative properties of fibroblasts in the infarcted myocardium. Early generation of matrix fragments in the infarct induces a pro-inflammatory and matrix-degrading fibroblast phenotype. Formation of a fibrin/fibronectin-rich provisional matrix serves as a conduit for migration of fibroblasts into the infarcted area. Induction of ED-A fibronectin and non-fibrillar collagens may contribute to myofibroblast transdifferentiation. Upregulation of matricellular proteins promotes transduction of growth factor and cytokine-mediated signals. As the scar matures, matrix cross-linking, clearance of matricellular proteins and reduced growth factor signaling cause deactivation and apoptosis of reparative infarct fibroblasts. Understanding the effects of matrix components on infarct fibroblasts may guide the design of peptides that reproduce, or inhibit, specific matricellular functions, attenuating adverse remodeling. PMID:22956156

  19. Effects of chlorhexidine, essential oils and herbal medicines (Salvia, Chamomile, Calendula) on human fibroblast in vitro

    Science.gov (United States)

    Urbaniak, Paulina; Szkaradkiewicz, Anna; Jankun, Jerzy; Kotwicka, Malgorzata

    2016-01-01

    Antiseptic rinses have been successfully used in inflammatory states of the gums and oral cavity mucosa. Antibacterial effects of chlorhexidine, essential oils and some herbs are well documented. Reaction of host tissue to these substances has much poorer documentation. The aim of the study was to analyse the influence of chlorhexidine (CHX), essential oil (EO: thymol, 0.064%; eucalyptol, 0.092%; methyl salicylate, 0.060%; menthol, 0.042%) mouth rinses and salvia, chamomile and calendula brews on fibroblast biology in vitro. The human fibroblast CCD16 line cells were cultured in incubation media which contained the examined substances. After 24 and 48 hours, the cell morphology, relative growth and apoptosis were evaluated. Exposure of fibroblasts to CHX, EO or salvia caused various changes in cell morphology. Cells cultured for 48 hours with CHX revealed a noticeably elongated shape of while cells cultured in high EO concentration or with salvia were considerably smaller and contracted with fewer projections. Chlorhexidine, EO and salvia reduced the fibroblast proliferation rate and stimulated cell death. Both reactions to EO were dose dependent. Cells exposure to chamomile or calendula brews did not change morphology or proliferation of fibroblasts. The results of this in vitro study showed that in contrast to chamomile and calendula, the brews of EO, CHX or salvia had a negative influence on fibroblast biology. PMID:27536196

  20. Gene expression signature of fibroblast serum response predicts human cancer progression: similarities between tumors and wounds.

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    Howard Y Chang

    2004-02-01

    Full Text Available Cancer invasion and metastasis have been likened to wound healing gone awry. Despite parallels in cellular behavior between cancer progression and wound healing, the molecular relationships between these two processes and their prognostic implications are unclear. In this study, based on gene expression profiles of fibroblasts from ten anatomic sites, we identify a stereotyped gene expression program in response to serum exposure that appears to reflect the multifaceted role of fibroblasts in wound healing. The genes comprising this fibroblast common serum response are coordinately regulated in many human tumors, allowing us to identify tumors with gene expression signatures suggestive of active wounds. Genes induced in the fibroblast serum-response program are expressed in tumors by the tumor cells themselves, by tumor-associated fibroblasts, or both. The molecular features that define this wound-like phenotype are evident at an early clinical stage, persist during treatment, and predict increased risk of metastasis and death in breast, lung, and gastric carcinomas. Thus, the transcriptional signature of the response of fibroblasts to serum provides a possible link between cancer progression and wound healing, as well as a powerful predictor of the clinical course in several common carcinomas.

  1. Therapeutic potential of gingival fibroblasts for cutaneous radiation syndrome: comparison to bone marrow-mesenchymal stem cell grafts.

    Science.gov (United States)

    Linard, Christine; Tissedre, Frederique; Busson, Elodie; Holler, Valerie; Leclerc, Thomas; Strup-Perrot, Carine; Couty, Ludovic; L'homme, Bruno; Benderitter, Marc; Lafont, Antoine; Lataillade, Jean Jacques; Coulomb, Bernard

    2015-05-15

    Mesenchymal stem cell (MSC) therapy has recently been investigated as a potential treatment for cutaneous radiation burns. We tested the hypothesis that injection of local gingival fibroblasts (GFs) would promote healing of radiation burn lesions and compared results with those for MSC transplantation. Human clinical- grade GFs or bone marrow-derived MSCs were intradermally injected into mice 21 days after local leg irradiation. Immunostaining and real-time PCR analysis were used to assess the effects of each treatment on extracellular matrix remodeling and inflammation in skin on days 28 and 50 postirradiation. GFs induced the early development of thick, fully regenerated epidermis, skin appendages, and hair follicles, earlier than MSCs did. The acceleration of wound healing by GFs involved rearrangement of the deposited collagen, modification of the Col/MMP/TIMP balance, and modulation of the expression and localization of tenascin-C and of the expression of growth factors (VEGF, EGF, and FGF7). As MSC treatment did, GF injection decreased the irradiation-induced inflammatory response and switched the differentiation of macrophages toward an M2-like phenotype, characterized by CD163(+) macrophage infiltration and strong expression of arginase-1. These findings indicate that GFs are an attractive target for regenerative medicine, for easier to collect, can grow in culture, and promote cutaneous wound healing in irradiation burn lesions.

  2. Key regulatory role of dermal fibroblasts in pigmentation as demonstrated using a reconstructed skin model: impact of photo-aging.

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    Christine Duval

    Full Text Available To study cutaneous pigmentation in a physiological context, we have previously developed a functional pigmented reconstructed skin model composed of a melanocyte-containing epidermis grown on a dermal equivalent comprising living fibroblasts. The present studies, using the same model, aimed to demonstrate that dermal fibroblasts influence skin pigmentation up to the macroscopic level. The proof of principle was performed with pigmented skins differing only in the fibroblast component. First, the in vitro system was reconstructed with or without fibroblasts in order to test the global influence of the presence of this cell type. We then assessed the impact of the origin of the fibroblast strain on the degree of pigmentation using fetal versus adult fibroblasts. In both experiments, impressive variation in skin pigmentation at the macroscopic level was observed and confirmed by quantitative parameters related to skin color, melanin content and melanocyte numbers. These data confirmed the responsiveness of the model and demonstrated that dermal fibroblasts do indeed impact the degree of skin pigmentation. We then hypothesized that a physiological state associated with pigmentary alterations such as photo-aging could be linked to dermal fibroblasts modifications that accumulate over time. Pigmentation of skin reconstructed using young unexposed fibroblasts (n = 3 was compared to that of tissues containing natural photo-aged fibroblasts (n = 3 which express a senescent phenotype. A stimulation of pigmentation in the presence of the natural photo-aged fibroblasts was revealed by a significant increase in the skin color (decrease in Luminance and an increase in both epidermal melanin content and melanogenic gene expression, thus confirming our hypothesis. Altogether, these data demonstrate that the level of pigmentation of the skin model is influenced by dermal fibroblasts and that natural photo-aged fibroblasts can contribute to the

  3. Aspirin suppresses cardiac fibroblast proliferation and collagen formation through downregulation of angiotensin type 1 receptor transcription

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    Wang, Xianwei, E-mail: XWang2@UAMS.edu; Lu, Jingjun; Khaidakov, Magomed; Mitra, Sona; Ding, Zufeng; Raina, Sameer; Goyal, Tanu; Mehta, Jawahar L., E-mail: MehtaJL@UAMS.edu

    2012-03-15

    Aspirin (acetyl salicylic acid, ASA) is a common drug used for its analgesic and antipyretic effects. Recent studies show that ASA not only blocks cyclooxygenase, but also inhibits NADPH oxidase and resultant reactive oxygen species (ROS) generation, a pathway that underlies pathogenesis of several ailments, including hypertension and tissue remodeling after injury. In these disease states, angiotensin II (Ang II) activates NADPH oxidase via its type 1 receptor (AT1R) and leads to fibroblast growth and collagen synthesis. In this study, we examined if ASA would inhibit NADPH oxidase activation, upregulation of AT1R transcription, and subsequent collagen generation in mouse cardiac fibroblasts challenged with Ang II. Mouse heart fibroblasts were isolated and treated with Ang II with or without ASA. As expected, Ang II induced AT1R expression, and stimulated cardiac fibroblast growth and collagen synthesis. The AT1R blocker losartan attenuated these effects of Ang II. Similarly to losartan, ASA, and its SA moiety suppressed Ang II-mediated AT1R transcription and fibroblast proliferation as well as expression of collagens and MMPs. ASA also suppressed the expression of NADPH oxidase subunits (p22{sup phox}, p47{sup phox}, p67{sup phox}, NOX2 and NOX4) and ROS generation. ASA did not affect total NF-κB p65, but inhibited its phosphorylation and activation. These observations suggest that ASA inhibits Ang II-induced NADPH oxidase expression, NF-κB activation and AT1R transcription in cardiac fibroblasts, and fibroblast proliferation and collagen expression. The critical role of NADPH oxidase activity in stimulation of AT1R transcription became apparent in experiments where ASA also inhibited AT1R transcription in cardiac fibroblasts challenged with H{sub 2}O{sub 2}. Since SA had similar effect as ASA on AT1R expression, we suggest that ASA's effect is mediated by its SA moiety. -- Highlights: ► Aspirin in therapeutic concentrations decreases mouse cardiac

  4. Identification of colonic fibroblast secretomes reveals secretory factors regulating colon cancer cell proliferation.

    Science.gov (United States)

    Chen, Sun-Xia; Xu, Xiao-En; Wang, Xiao-Qing; Cui, Shu-Jian; Xu, Lei-Lei; Jiang, Ying-Hua; Zhang, Yang; Yan, Hai-Bo; Zhang, Qian; Qiao, Jie; Yang, Peng-Yuan; Liu, Feng

    2014-10-14

    Stromal microenvironment influences tumor cell proliferation and migration. Fibroblasts represent the most abundant stromal constituents. Here, we established two pairs of normal fibroblast (NF) and cancer-associated fibroblast (CAF) cultures from colorectal adenocarcinoma tissues and the normal counterparts. The NFs and CAFs were stained positive for typical fibroblast markers and inhibited colon cancer (CC) cell proliferation in in vitro cocultures and in xenograft mouse models. The fibroblast conditioned media were analyzed using LC-MS and 227 proteins were identified at a false discovery rate of 1.3%, including 131 putative secretory and 20 plasma membrane proteins. These proteins were enriched for functional categories of extracellular matrix, adhesion, cell motion, inflammatory response, redox homeostasis and peptidase inhibitor. Secreted protein acidic and rich in cysteine, transgelin, follistatin-related protein 1 (FSTL1) and decorin was abundant in the fibroblast secretome as confirmed by Western blot. Silencing of FSTL1 and transgelin in colonic fibroblast cell line CCD-18Co induced an accelerated proliferation of CC cells in cocultures. Exogenous FSTL1 attenuates CC cell proliferation in a negative fashion. FSTL1 was upregulated in CC patient plasma and cancerous tissues but had no implication in prognosis. Our results provided novel insights into the molecular signatures and modulatory role of CC associated fibroblasts. In this study, a label-free LC-MS was performed to analyze the secretomes of two paired primary fibroblasts, which were isolated from fresh surgical specimen of colorectal adenocarcinoma and adjacent normal colonic tissues and exhibited negative modulatory activity for colon cancer cell growth in in vitro cocultures and in vivo xenograph mouse models. Follistatin-related protein 1 was further revealed to be one of the stroma-derived factors of potential suppression role for colon cancer cell proliferation. Our results provide novel

  5. Intrinsic innate immunity fails to control herpes simplex virus and vesicular stomatitis virus replication in sensory neurons and fibroblasts.

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    Rosato, Pamela C; Leib, David A

    2014-09-01

    Herpes simplex virus 1 (HSV-1) establishes lifelong latent infections in the sensory neurons of the trigeminal ganglia (TG), wherein it retains the capacity to reactivate. The interferon (IFN)-driven antiviral response is critical for the control of HSV-1 acute replication. We therefore sought to further investigate this response in TG neurons cultured from adult mice deficient in a variety of IFN signaling components. Parallel experiments were also performed in fibroblasts isolated concurrently. We showed that HSV-1 replication was comparable in wild-type (WT) and IFN signaling-deficient neurons and fibroblasts. Unexpectedly, a similar pattern was observed for the IFN-sensitive vesicular stomatitis virus (VSV). Despite these findings, TG neurons responded to IFN-β pretreatment with STAT1 nuclear localization and restricted replication of both VSV and an HSV-1 strain deficient in γ34.5, while wild-type HSV-1 replication was unaffected. This was in contrast to fibroblasts in which all viruses were restricted by the addition of IFN-β. Taken together, these data show that adult TG neurons can mount an effective antiviral response only if provided with an exogenous source of IFN-β, and HSV-1 combats this response through γ34.5. These results further our understanding of the antiviral response of neurons and highlight the importance of paracrine IFN-β signaling in establishing an antiviral state. Herpes simplex virus 1 (HSV-1) is a ubiquitous virus that establishes a lifelong latent infection in neurons. Reactivation from latency can cause cold sores, blindness, and death from encephalitis. Humans with deficiencies in innate immunity have significant problems controlling HSV infections. In this study, we therefore sought to elucidate the role of neuronal innate immunity in the control of viral infection. Using neurons isolated from mice, we found that the intrinsic capacity of neurons to restrict virus replication was unaffected by the presence or absence of

  6. Autologous glioma cell vaccine admixed with interleukin-4 gene transfected fibroblasts in the treatment of patients with malignant gliomas

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    Torres-Trejo Alejandro

    2007-12-01

    Full Text Available Abstract Background The prognosis for malignant gliomas remains dismal. We addressed the safety, feasibility and preliminary clinical activity of the vaccinations using autologous glioma cells and interleukin (IL-4 gene transfected fibroblasts. Methods In University of Pittsburgh Cancer Institute (UPCI protocol 95-033, adult participants with recurrent glioblastoma multiforme (GBM or anaplastic astrocytoma (AA received gross total resection (GTR of the recurrent tumors, followed by two vaccinations with autologous fibroblasts retrovirally transfected with TFG-IL4-Neo-TK vector admixed with irradiated autologous glioma cells. In UPCI 99-111, adult participants with newly diagnosed GBM or AA, following GTR and radiation therapy, received two intradermal vaccinations with the TFG-IL4-Neo-TK-transfected fibroblasts admixed with type-1 dendritic cells (DC loaded with autologous tumor lysate. The participants were evaluated for occurrence of adverse events, immune response, and clinical response by radiological imaging. Results and Discussion In UPCI 95-033, only 2 of 6 participants received the vaccinations. Four other participants were withdrawn from the trial because of tumor progression prior to production of the cellular vaccine. However, both participants who received two vaccinations demonstrated encouraging immunological and clinical responses. Biopsies from the local vaccine sites from one participant displayed IL-4 dose-dependent infiltration of CD4+ as well as CD8+ T cells. Interferon (IFN-γ Enzyme-Linked Immuno-SPOT (ELISPOT assay in another human leukocyte antigen (HLA-A2+ participant demonstrated systemic T-cell responses against an HLA-A2-restricted glioma-associated antigen (GAA epitope EphA2883–891. Moreover, both participants demonstrated clinical and radiological improvement with no evidence of allergic encephalitis, although both participants eventually succumbed with the tumor recurrence. In 99-111, 5 of 6 enrolled participants

  7. Adipose Tissue-Derived Stromal Cells Inhibit TGF-beta 1-Induced Differentiation of Human Dermal Fibroblasts and Keloid Scar-Derived Fibroblasts in a Paracrine Fashion

    NARCIS (Netherlands)

    Spiekman, Maroesjka; Przybyt, Ewa; Plantinga, Josee A.; Gibbs, Susan; van der Lei, Berend; Harmsen, Martin C.

    2014-01-01

    Background: Adipose tissue-derived stromal cells augment wound healing and skin regeneration. It is unknown whether and how they can also influence dermal scarring. The authors hypothesized that adipose tissue-derived stromal cells inhibit adverse differentiation of dermal fibroblasts induced by the

  8. Effects of basic fibroblast growth factor on density and morphology of fibroblasts grown on root surfaces with or without conditioning with tetracycline or EDTA.

    Science.gov (United States)

    Silvério, Karina G; Martinez, Aurora E T; Rossa, Carlos

    2007-09-01

    A study was conducted to evaluate in vitro the effect of root surface conditioning with basic fibroblast growth factor (b-FGF) on morphology and proliferation of fibroblasts. Three experimental groups were used: non-treated, and treated with 50 microg or 125 microg b-FGF/ml. The dentin samples in each group were divided into subgroups according to the chemical treatment received before application of b-FGF: none, or conditioned with tetracycline-HCl or EDTA. After contact with b-FGF for 5 min, the samples were incubated for 24 h with 1 ml of culture medium containing 1 x 10(5) cells/ml plus 1 ml of culture medium alone. The samples were then subjected to routine preparation for SEM, and random fields were photographed. Three calibrated and blind examiners performed the assessment of morphology and density according to two index systems. Classification and regression trees indicated that the root surfaces treated with 125 microg b-FGF and previously conditioned with tetracycline-HCl or EDTA presented a morphology more suggestive of cellular adhesion and viability (P = 0.004). The density of fibroblasts on samples previously conditioned with EDTA, regardless of treatment with b-FGF, was significantly higher than in the other groups (P < 0.001). The present findings suggest that topical application of b-FGF has a positive influence on both the density and morphology of fibroblasts.

  9. Monomethylarsonous acid inhibited endogenous cholesterol biosynthesis in human skin fibroblasts

    Energy Technology Data Exchange (ETDEWEB)

    Guo, Lei [Environmental Toxicology Graduate Program, University of California, Riverside, CA 92521-0403 (United States); Xiao, Yongsheng [Department of Chemistry, University of California, Riverside, CA 92521-0403 (United States); Wang, Yinsheng, E-mail: yinsheng.wang@ucr.edu [Environmental Toxicology Graduate Program, University of California, Riverside, CA 92521-0403 (United States); Department of Chemistry, University of California, Riverside, CA 92521-0403 (United States)

    2014-05-15

    Human exposure to arsenic in drinking water is a widespread public health concern, and such exposure is known to be associated with many human diseases. The detailed molecular mechanisms about how arsenic species contribute to the adverse human health effects, however, remain incompletely understood. Monomethylarsonous acid [MMA(III)] is a highly toxic and stable metabolite of inorganic arsenic. To exploit the mechanisms through which MMA(III) exerts its cytotoxic effect, we adopted a quantitative proteomic approach, by coupling stable isotope labeling by amino acids in cell culture (SILAC) with LC-MS/MS analysis, to examine the variation in the entire proteome of GM00637 human skin fibroblasts following acute MMA(III) exposure. Among the ∼ 6500 unique proteins quantified, ∼ 300 displayed significant changes in expression after exposure with 2 μM MMA(III) for 24 h. Subsequent analysis revealed the perturbation of de novo cholesterol biosynthesis, selenoprotein synthesis and Nrf2 pathways evoked by MMA(III) exposure. Particularly, MMA(III) treatment resulted in considerable down-regulation of several enzymes involved in cholesterol biosynthesis. In addition, real-time PCR analysis showed reduced mRNA levels of select genes in this pathway. Furthermore, MMA(III) exposure contributed to a distinct decline in cellular cholesterol content and significant growth inhibition of multiple cell lines, both of which could be restored by supplementation of cholesterol to the culture media. Collectively, the present study demonstrated that the cytotoxicity of MMA(III) may arise, at least in part, from the down-regulation of cholesterol biosynthesis enzymes and the resultant decrease of cellular cholesterol content. - Highlights: • MMA(III)-induced perturbation of the entire proteome of GM00637 cells is studied. • Quantitative proteomic approach revealed alterations of multiple cellular pathways. • MMA(III) inhibits de novo cholesterol biosynthesis. • MMA

  10. Fluid Flow Mechanotransduction in Vascular Smooth Muscle Cells and Fibroblasts

    Science.gov (United States)

    Shi, Zhong-Dong; Tarbell, John M.

    2011-01-01

    Understanding how vascular wall endothelial cells (ECs), smooth muscle cells (SMCs), and fibroblasts (FBs) sense and transduce the stimuli of hemodynamic forces (shear stress, cyclic strain, and hydrostatic pressure) into intracellular biochemical signals is critical to prevent vascular disease development and progression. ECs lining the vessel lumen directly sense alterations in blood flow shear stress and then communicate with medial SMCs and adventitial FBs to regulate vessel function and disease. Shear stress mechanotransduction in ECs has been extensively studied and reviewed. In the case of endothelial damage, blood flow shear stress may directly act on the superficial layer of SMCs and transmural interstitial flow may be elevated on medial SMCs and adventitial FBs. Therefore, it is also important to investigate direct shear effects on vascular SMCs as well as FBs. The work published in the last two decades has shown that shear stress and interstitial flow have significant influences on vascular SMCs and FBs. This review summarizes work that considered direct shear effects on SMCs and FBs and provides the first comprehensive overview of the underlying mechanisms that modulate SMC secretion, alignment, contraction, proliferation, apoptosis, differentiation, and migration in response to 2-dimensional (2D) laminar, pulsatile, and oscillating flow shear stresses and 3D interstitial flow. A mechanistic model of flow sensing by SMCs is also provided to elucidate possible mechanotransduction pathways through surface glycocalyx, integrins, membrane receptors, ion channels, and primary cilia. Understanding flow-mediated mechanotransduction in SMCs and FBs and the interplay with ECs should be helpful in exploring strategies to prevent flow-initiated atherosclerosis and neointima formation and has implications in vascular tissue engineering. PMID:21479754

  11. Myxoinflammatory fibroblastic sarcoma: spectrum of disease and imaging presentation

    Energy Technology Data Exchange (ETDEWEB)

    Gaetke-Udager, Kara; Yablon, Corrie M.; Morag, Yoav [University of Michigan Health System, Department of Radiology, Ann Arbor, MI (United States); Lucas, David R. [University of Michigan Health System, Department of Pathology, Ann Arbor, MI (United States)

    2016-03-15

    To describe the imaging findings of a series of myxoinflammatory fibroblastic sarcomas (MFSs) from our institution, including a case of dedifferentiated MFS and two cases with areas of high-grade tumor, in addition to typical cases of low-grade tumor. To correlate the imaging findings with the pathologic features of these tumors. IRB approval was obtained. Retrospective search of the pathology database at our institution from 2000 to 2015 identified seven cases of MFS with available imaging. Imaging, pathology, and clinical data were reviewed. Unlike the majority of well-differentiated tumors in our series (four cases), one tumor showed dedifferentiation and two cases had areas of high-grade tumor. The dedifferentiated tumor showed peripheral post-contrast enhancement. One case with a substantial high-grade component showed osseous destruction and peripheral enhancement in the high-grade area, while the low-grade component enhanced diffusely. The second case had a small high-grade area and showed diffuse enhancement. All three of these cases had non-acral locations and lacked association with a tendon. The four cases of low-grade MFS demonstrated diffuse enhancement, were located in the distal extremities, and were associated with a tendon. The imaging findings of dedifferentiated and high-grade MFS differ from the more typical low-grade tumors in that they have nonenhancing areas, a non-acral location, lack association with a tendon, and may involve bone. The radiologist should be aware that MFS represents a spectrum that includes low-grade tumors, tumors with high-grade areas, and tumors with dedifferentiation and that this spectrum presents with differing imaging features. (orig.)

  12. Effects of Calendula officinalis on human gingival fibroblasts.

    Science.gov (United States)

    Saini, Pragtipal; Al-Shibani, Nouf; Sun, Jun; Zhang, Weiping; Song, Fengyu; Gregson, Karen S; Windsor, L Jack

    2012-04-01

    Calendula officinalis is commonly called the marigold. It is a staple topical remedy in homeopathic medicine. It is rich in quercetin, carotenoids, lutein, lycopene, rutin, ubiquinone, xanthophylls, and other anti-oxidants. It has anti-inflammatory properties. Quercetin, one of the active components in Calendula, has been shown to inhibit recombinant human matrix metalloproteinase (MMP) activity and decrease the expression of tumor necrosis factor-α, interleukin-1β (IL), IL-6 and IL-8 in phorbol 12-myristate 13-acetate and calcium ionophore-stimulated human mast cells. To examine the effects of Calendula on human gingival fibroblast (HGF) mediated collagen degradation and MMP activity. Lactate dehydrogenate assays were performed to determine the non-toxic concentrations of Calendula, doxycycline and quercetin. Cell-mediated collagen degradation assays were performed to examine the inhibitory effect on cell-mediated collagen degradation. Gelatin zymography was performed to examine their effects on MMP-2 activity. The experiments were repeated three times and ANOVA used for statistical analyses. Calendula at 2-3% completely inhibited the MMP-2 activity in the zymograms. Doxycycline inhibited HGF-mediated collagen degradation at 0.005, 0.01, 0.02 and 0.05%, and MMP-2 activity completely at 0.05%. Quercetin inhibited HGF-mediated collagen degradation at 0.005, 0.01 and 0.02%, and MMP-2 activity in a dose-dependent manner. Calendula inhibited HGF-mediated collagen degradation and MMP-2 activity more than the same correlated concentration of pure quercetin. Calendula inhibits HGF-mediated collagen degradation and MMP-2 activity more than the corresponding concentration of quercetin. This may be attributed to additional components in Calendula other than quercetin. Published by Elsevier Ltd.

  13. Modulation of the Senescence-Associated Inflammatory Phenotype in Human Fibroblasts by Olive Phenols

    Directory of Open Access Journals (Sweden)

    Beatrice Menicacci

    2017-10-01

    Full Text Available Senescent cells display an increase in the secretion of growth factors, inflammatory cytokines and proteolytic enzymes, termed the “senescence-associated-secretory-phenotype” (SASP, playing a major role in many age-related diseases. The phenolic compounds present in extra-virgin olive oil are inhibitors of oxidative damage and have been reported to play a protective role in inflammation-related diseases. Particularly, hydroxytyrosol and oleuropein are the most abundant and more extensively studied. Pre-senescent human lung (MRC5 and neonatal human dermal (NHDF fibroblasts were used as cellular model to evaluate the effect of chronic (4–6 weeks treatment with 1 μM hydroxytyrosol (HT or 10 μM oleuropein aglycone (OLE on senescence/inflammation markers. Both phenols were effective in reducing β-galactosidase-positive cell number and p16 protein expression. In addition, senescence/inflammation markers such as IL-6 and metalloprotease secretion, and Ciclooxigenase type 2 (COX-2 and α-smooth-actin levels were reduced by phenol treatments. In NHDF, COX-2 expression, Nuclear Factor κ-light-chain-enhancer of activated B cells (NFκB protein level and nuclear localization were augmented with culture senescence and decreased by OLE and HT treatment. Furthermore, the inflammatory effect of Tumor Necrosis Factor α (TNFα exposure was almost completely abolished in OLE- and HT-pre-treated NHDF. Thus, the modulation of the senescence-associated inflammatory phenotype might be an important mechanism underlying the beneficial effects of olive oil phenols.

  14. Thyroid hormone receptor regulates most genes independently of fibroblast growth factor 21 in liver.

    Science.gov (United States)

    Zhang, Aijun; Sieglaff, Douglas H; York, Jean Philippe; Suh, Ji Ho; Ayers, Stephen D; Winnier, Glenn E; Kharitonenkov, Alexei; Pin, Christopher; Zhang, Pumin; Webb, Paul; Xia, Xuefeng

    2015-03-01

    Thyroid hormone (TH) acts through specific receptors (TRs), which are conditional transcription factors, to induce fibroblast growth factor 21 (FGF21), a peptide hormone that is usually induced by fasting and that influences lipid and carbohydrate metabolism via local hepatic and systemic endocrine effects. While TH and FGF21 display overlapping actions when administered, including reductions in serum lipids, according to the current models these hormones act independently in vivo. In this study, we examined mechanisms of regulation of FGF21 expression by TH and tested the possibility that FGF21 is required for induction of hepatic TH-responsive genes. We confirm that active TH (triiodothyronine (T3)) and the TRβ-selective thyromimetic GC1 increase FGF21 transcript and peptide levels in mouse liver and that this effect requires TRβ. T3 also induces FGF21 in cultured hepatocytes and this effect involves direct actions of TRβ1, which binds a TRE within intron 2 of FGF21. Gene expression profiles of WT and Fgf21-knockout mice are very similar, indicating that FGF21 is dispensable for the majority of hepatic T3 gene responses. A small subset of genes displays diminished T3 response in the absence of FGF21. However, most of these are not obviously directly involved in T3-dependent hepatic metabolic processes. Consistent with these results, T3-dependent effects on serum cholesterol are maintained in the Fgf21(-/-) background and we observe no effect of the Fgf21-knockout background on serum triglycerides and glucose. Our findings indicate that T3 regulates the genes involved in classical hepatic metabolic responses independently of FGF21. © 2015 Society for Endocrinology.

  15. Mitochondrial reactive oxygen species mediate the lipopolysaccharide-induced pro-inflammatory response in human gingival fibroblasts

    Energy Technology Data Exchange (ETDEWEB)

    Li, Xue; Wang, Xiaoxuan [Department of Periodontology, Peking University School and Hospital of Stomatology, National Engineering Laboratory for Digital and Material Technology of Stomatology, Beijing Key Laboratory of Digital Stomatology, 22 Zhongguancun Avenue South, Haidian District, Beijing 100081 (China); Zheng, Ming, E-mail: zhengm@bjmu.edu.cn [Department of Physiology and Pathophysiology, Peking University Health Science Center, 38 Xueyuan Road, Haidian District, Beijing 100191 (China); Luan, Qing Xian, E-mail: kqluanqx@126.com [Department of Periodontology, Peking University School and Hospital of Stomatology, National Engineering Laboratory for Digital and Material Technology of Stomatology, Beijing Key Laboratory of Digital Stomatology, 22 Zhongguancun Avenue South, Haidian District, Beijing 100081 (China)

    2016-09-10

    Although periodontal diseases are initiated by bacteria that colonize the tooth surface and gingival sulcus, the host response is believed to play an essential role in the breakdown of connective tissue and bone. Mitochondrial reactive oxygen species (mtROS) have been proposed to regulate the activation of the inflammatory response by the innate immune system. However, the role of mtROS in modulating the response of human gingival fibroblasts (HGFs) to immune stimulation by lipopolysaccharides (LPS) has yet to be fully elucidated. Here, we showed that LPS from Porphyromonas gingivalis stimulated HGFs to increase mtROS production, which could be inhibited by treatment with a mitochondrial-targeted exogenous antioxidant (mito-TEMPO) or transfection with manganese superoxide dismutase (MnSOD). A time-course study revealed that an increase in the concentration of mtROS preceded the expression of inflammatory cytokines in HGFs. Mito-TEMPO treatment or MnSOD transfection also significantly prevented the LPS-induced increase of interleukin (IL)-1β, IL-6, and tumor necrosis factor-α. Furthermore, suppressing LPS-induced mtROS generation inhibited the activation of p38, c-Jun N-terminal kinase, and inhibitor of nuclear factor-κB kinase, as well as the nuclear localization of nuclear factor-κB. These results demonstrate that mtROS generation is a key signaling event in the LPS-induced pro-inflammatory response of HGFs. - Highlights: • Inflammation is thought to promote pathogenic changes in periodontitis. • We investigated mtROS as a regulator of inflammation in gingival fibroblasts. • Targeted antioxidants were used to inhibit mtROS production after LPS challenge. • Inhibiting mtROS generation suppressed the secretion of pro-inflammatory cytokines. • JNK, p38, IKK, and NF-κB were shown to act as transducers of mtROS signaling.

  16. Expression pattern and regulation of genes differ between fibroblasts of adhesion and normal human peritoneum

    Directory of Open Access Journals (Sweden)

    Saed Ghassan M

    2005-01-01

    Full Text Available Abstract Background Injury to the peritoneum during surgery is followed by a healing process that frequently results in the attachment of adjacent organs by a fibrous mass, referred commonly as adhesions. Because injuries to the peritoneum during surgery are inevitable, it is imperative that we understand the mechanisms of adhesion formation to prevent its occurrence. This requires thorough understanding of the molecular sequence that results in the attachment of injured peritoneum and the development of fibrous tissue. Recent data show that fibroblasts from the injured peritoneum may play a critical role in the formation of adhesion tissues. Therefore, identifying changes in gene expression pattern in the peritoneal fibroblasts during the process may provide clues to the mechanisms by which adhesion develop. Methods In this study, we compared expression patterns of larger number of genes in the fibroblasts isolated from adhesion and normal human peritoneum using gene filters. Contributions of TGF-beta1 and hypoxia in the altered expression of specific genes were also examined using a semiquantitative RT-PCR technique. Results Results show that several genes are differentially expressed between fibroblasts of normal and adhesion peritoneum and that the peritoneal fibroblast may acquire a different phenotype during adhesion formation. Genes that are differentially expressed between normal and adhesion fibroblasts encode molecules involved in cell adhesion, proliferation, differentiation, migration and factors regulating cytokines, transcription, translation and protein/vesicle trafficking. Conclusions Our data substantiate that adhesion formation is a multigenic phenomenon and not all changes in gene expression pattern between normal and adhesion fibroblasts are the function of TGF-beta1 and hypoxia that are known to influence adhesion formation. Analysis of the gene expression data in the perspective of known functions of genes connote to

  17. Complement C3a Mobilizes Dental Pulp Stem Cells and Specifically Guides Pulp Fibroblast Recruitment.

    Science.gov (United States)

    Rufas, Pierre; Jeanneau, Charlotte; Rombouts, Charlotte; Laurent, Patrick; About, Imad

    2016-09-01

    Complement activation is considered a major mechanism in innate immunity. Although it is mainly involved in initiating inflammation, recent data reported its involvement in other processes such as tissue regeneration. In the dental pulp, complement C5a fragment has been shown to be involved in the recruitment of dental pulp stem cells (DPSCs). This study sought to investigate the possible role of C3a, another complement fragment, in the early steps of dentin-pulp regeneration. Expression of C3a receptor (C3aR) was investigated by immunofluorescence and reverse transcriptase polymerase chain reaction on cultured pulp fibroblasts, STRO-1-sorted DPSCs, as well as on human tooth sections in vivo. The effect of C3a on proliferation of both DPSCs and pulp fibroblasts was investigated by MTT assay. Cell migration under a C3a gradient was investigated by using microfluidic chemotaxis chambers. C3aR was expressed in vivo as well as in cultured pulp fibroblasts co-expressing fibroblast surface protein and in DPSCs co-expressing STRO-1. Addition of recombinant C3a induced a significant proliferation of both cell types. When subjected to a C3a gradient, DPSCs were mobilized but not specifically recruited, whereas pulp fibroblasts were specifically recruited following a C3a gradient. These results provide the first demonstration of C3aR expression in the dental pulp and demonstrate that C3a is involved in increasing DPSCs and fibroblast proliferation, in mobilizing DPSCs, and in specifically guiding fibroblast recruitment. This provides an additional link to the tight correlation between inflammation and tissue regeneration. Copyright © 2016. Published by Elsevier Inc.

  18. Reciprocal responses of fibroblasts and melanocytes to α-MSH depending on MC1R polymorphisms.

    Science.gov (United States)

    Stanisz, Hedwig; Seifert, Markus; Tilgen, Wolfgang; Vogt, Thomas; Rass, Knuth

    2011-10-01

    The melanocortin 1-receptor (MC1R) exhibits several variants in form of single nucleotide polymorphisms (SNPs) that are known to differentially regulate melanocyte function. However, whether and how MC1R polymorphisms also affect fibroblast function has not been investigated so far.Therefore we measured intracellular cyclic adenosine monophosphate (cAMP) concentrations and cellular proliferation upon stimulation with alpha-melanocyte stimulating hormone (α-MSH) in eight different human fibroblast and melanocyte cell lines with wild type and different MC1R SNPs.We found that fibroblasts, as well as melanocytes, show differences in MC1R function depending on the MC1R genotype. MC1R stimulation with α-MSH in wild type (MC1R(wt)) melanocytes results in an increase of intracellular cAMP and cellular proliferation. In contrast, MC1R(wt) fibroblasts react with a decrease of intracellular cAMP and proliferation. In MC1R polymorphic fibroblasts (R163Q, R151C and V60L) both effects are significantly alleviated. Similar, but inverse effects could be found in MC1R polymorphic melanocytes (R142H and V92M) with a significantly lower cAMP increase and proliferation rate compared to MC1R(wt) melanocytes.Our results indicate that the MC1R displays reciprocal growth responses in melanocytes and fibroblasts, depending on the MC1R genotype. Thus, the MC1R seems to be not solely important for the skin pigmentary system, but also for the fibroblast function, and might influence different processes of the dermal compartment like wound healing, fibrosis and keloid formation.

  19. Lipopolysaccharide promotes lipid accumulation in human adventitial fibroblasts via TLR4-NF-κB pathway

    Directory of Open Access Journals (Sweden)

    Wang Jun

    2012-10-01

    Full Text Available Abstract Background Atherosclerosis is a chronic degenerative disease of the arteries and is thought to be one of the most common causes of death globally. In recent years, the functions of adventitial fibroblasts in the development of atherosclerosis and tissue repair have gained increased interests. LPS can increase the morbidity and mortality of atherosclerosis-associated cardiovascular disease. Although LPS increases neointimal via TLR4 activation has been reported, how LPS augments atherogenesis through acting on adventitial fibroblasts is still unknown. Here we explored lipid deposition within adventitial fibroblasts mediated by lipopolysaccharide (LPS to imitate inflammatory conditions. Results In our study, LPS enhanced lipid deposition by the up-regulated expression of adipose differentiation-related protein (ADRP as the silencing of ADRP abrogated lipid deposition in LPS-activated adventitial fibroblasts. In addition, pre-treatment with anti-Toll-like receptor 4 (TLR4 antibody diminished the LPS-induced lipid deposition and ADRP expression. Moreover, LPS induced translocation of nuclear factor-κB (NF-κB, which could markedly up-regulate lipid deposition as pre-treatment with the NF-κB inhibitor, PDTC, significantly reduced lipid droplets. In addition, the lowering lipid accumulation was accompanied with the decreased ADRP expression. Furthermore, LPS-induced adventitial fibroblasts secreted more monocyte chemoattractant protein (MCP-1, compared with transforming growth factor-β1 (TGF-β1. Conclusions Taken together, these results suggest that LPS promotes lipid accumulation via the up-regulation of ADRP expression through TLR4 activated downstream of NF-κB in adventitial fibroblasts. Increased levels of MCP-1 released from LPS-activated adventitial fibroblasts and lipid accumulation may accelerate monocytes recruitment and lipid-laden macrophage foam cells formation. Here, our study provides a new explanation as to how bacterial

  20. Influence of caffeine and hyaluronic acid on collagen biosynthesis in human skin fibroblasts

    Directory of Open Access Journals (Sweden)

    Donejko M

    2014-10-01

    Full Text Available Magdalena Donejko,1 Andrzej Przylipiak,1 Edyta Rysiak,2 Katarzyna Głuszuk,2 Arkadiusz Surażyński2 1Department of Esthetic Medicine, 2Department of Medicinal Chemistry, Faculty of Pharmacy, Medical University of Białystok, Białystok, Poland Aim: The aim of this study was to evaluate the effect of caffeine on collagen biosynthesis in human skin fibroblasts and the influence of hyaluronic acid (HA on this process. Materials and methods: Collagen, [3H]-thymidine incorporation, and prolidase activity were measured in confluent human skin fibroblast cultures that had been treated with 1, 2, and 5 mM caffeine and with caffeine and 500 µg/mL HA. Western immunoblot analysis was performed to evaluate expression of ß1-integrin receptor, insulin-like growth factor receptor phospho-Akt protein and mitogen-activated protein kinase (phospho-extracellular signal-regulated kinase. Results: Caffeine inhibited collagen biosynthesis in a dose-dependent manner. The mechanism of this process was found at the level of prolidase activity. Caffeine significantly inhibited the enzyme activity. The addition of HA had no effect on collagen biosynthesis or prolidase activity in fibroblasts incubated with caffeine. Caffeine also had an inhibitory effect on DNA biosynthesis. HA, however, did not have any significant effect on this process. The inhibition of the expression of ß1-integrin and insulin-like growth factor receptor in fibroblasts incubated with the caffeine indicates a possible mechanism of inhibition of collagen biosynthesis. Conclusion: Caffeine reduces collagen synthesis in human cultured skin fibroblasts. HA did not have any significant protective effect on this process. This is the first study to our knowledge that reports caffeine-induced inhibition of collagen synthesis in human skin fibroblasts. Keywords: collagen, caffeine, hyaluronic acid, fibroblast

  1. Glucosylceramide and glucosylsphingosine metabolism in cultured fibroblasts deficient in acid beta-glucosidase activity.

    Science.gov (United States)

    Sasagasako, N; Kobayashi, T; Yamaguchi, Y; Shinnoh, N; Goto, I

    1994-01-01

    The metabolism of glucosylceramide (GlcCer) and glucosylsphingosine (GlcSph) was studied using cultured fibroblasts deficient in acid beta-glucosidase activity. In fibroblasts from patients with Gaucher's disease, in vitro beta-glucosidase activities were 2.7-11.7% and 4.8-13.6% of control values when 4-methylumbelliferyl beta-D-glucoside and GlcSph were used as substrates, respectively. In spite of the enzyme deficiency, GlcCer and GlcSph, the natural substrates of the deficient enzyme, did not accumulate in the cells. When normal fibroblasts were incubated with conduritol B epoxide (CBE), a specific inhibitor of acid beta-glucosidase, the in vitro enzyme activities decreased dose-dependently (2.2-2.4% of control values at 50 microM CBE), and GlcCer and GlcSph accumulated in the cells at concentrations of CBE higher than 50 microM. To investigate the intracellular metabolism of GlcCer and GlcSph, either radioactive GlcCer or GlcSph was loaded onto cultured fibroblasts. In fibroblasts treated with a high dose of CBE (1 mM), the degradation of GlcCer and GlcSph was retarded (5-21% on day 7; normal range, 81-99%), while in fibroblasts from patients with Gaucher's disease, both the pattern and rate of the degradation of the lipids (83-97% on day 7) were almost the same as those seen in the control cells. These results indicate that in Gaucher's disease fibroblasts the intracellular metabolism of GlcCer and GlcSph is normal in spite of the deficiency in beta-glucosidase activity.(ABSTRACT TRUNCATED AT 250 WORDS)

  2. The decrease of fibroblasts and fibroblast growth factor-2 expressions as a result of X-ray irradiation on the tooth extraction socket in Rattus novergicus

    Directory of Open Access Journals (Sweden)

    Fatma Yasmin Mahdani

    2015-06-01

    Full Text Available Background: Wound healing involves cellular, molecular, physiological, and biochemical processes as responses to tissue damage. For instance, when a failure during tooth extraction occurs, radiographic examination, X-rays, is required. X-rays as an enforcer diagnosis can damage DNA chain, resulting in cell death and inhibition of wound healing process. Purpose: This research aims to analyze fibroblasts cell number and fibroblast growth factor-2 (FGF-2 expressions during wound healing process after tooth extraction as a result of X-ray irradiation. Methods: There were three research groups, each consisting of ten rats. Incisor tooth extraction was performed on the left lower jaw, and then X-ray examination was conducted at certain irradiation doses, namely 0 mSv, 0.08 mSv, and 0.16 mSv. Those animals were sacrificed on day 3, and on day 7 after the extraction, histopathology and immunohistochemistry examinations were conducted to determine fibroblast cell number and FGF-2 expressions. Data obtained were then analyzed by oneway ANOVA and Tukey HSD tests. Results: The number of fibroblasts decreased significantly in the group with the irradiation dose of 0.16 mSv applied on day 7 after the extraction (p <0.05. Similarly, the number of FGF-2 expressions decreased significantly in the group with the irradiation dose of 0.16 mSv applied on days 3 and 7 after the extraction (p <0.05. Conclusion: X-ray irradiation at a dose of 0.16 mSv can inhibit the healing process of tooth extraction wound due to the decreasing of fibroblasts cell number and FGF-2 expressions.

  3. Quantum Locality?

    Energy Technology Data Exchange (ETDEWEB)

    Stapp, Henry

    2011-11-10

    vagaries that he cites do not upset the proof in question. It is show here in detail why the precise statement of this theorem justifies the specified application of CQT. It is also shown, in response to his challenge, why a putative proof of locality that he has proposed is not valid.

  4. MRC-5 fibroblast-conditioned medium influences multiple pathways regulating invasion, migration, proliferation, and apoptosis in hepatocellular carcinoma

    OpenAIRE

    Ding, Songming; Chen, Guoliang; Zhang, Wu; Xing, Chunyang; Xu, Xiao; Xie, Haiyang; Lu, Aili; Chen, Kangjie; Guo, Haijun; Ren, Zhigang; Zheng, Shusen; Zhou, Lin

    2015-01-01

    Background Carcinoma associated fibroblasts (CAFs), an important component of tumor microenvironment, are capable of enhancing tumor cells invasion and migration through initiation of epithelial?mesenchymal transition (EMT). MRC-5 fibroblasts are one of the CAFs expressing alpha-smooth muscle actin. It is ascertained that medium conditioned by MRC-5 fibroblasts stimulate motility and invasion of breast cancer cells. However, its role in hepatocellular carcinoma (HCC) is less clear. The aim of...

  5. Coxsackievirus B3 induces the formation of autophagosomes in cardiac fibroblasts both in vitro and in vivo

    Energy Technology Data Exchange (ETDEWEB)

    Zhai, Xia, E-mail: zhai_xia_cool@126.com [Department of Microbiology and Wu Lien-Teh Institute, Harbin Medical University, 157 Baojian Road, Harbin 150081 (China); Qin, Ying, E-mail: qinyinggaofeng@163.com [Department of Microbiology and Wu Lien-Teh Institute, Harbin Medical University, 157 Baojian Road, Harbin 150081 (China); Chen, Yang, E-mail: cy_hmu@126.com [Department of Microbiology and Wu Lien-Teh Institute, Harbin Medical University, 157 Baojian Road, Harbin 150081 (China); Lin, Lexun, E-mail: linlexun@163.com [Department of Microbiology and Wu Lien-Teh Institute, Harbin Medical University, 157 Baojian Road, Harbin 150081 (China); Wang, Tianying, E-mail: wangty0929@163.com [Department of Microbiology and Wu Lien-Teh Institute, Harbin Medical University, 157 Baojian Road, Harbin 150081 (China); Zhong, Xiaoyan, E-mail: littlerock712@163.com [Department of Microbiology and Wu Lien-Teh Institute, Harbin Medical University, 157 Baojian Road, Harbin 150081 (China); Wu, Xiaoyu, E-mail: xiaoyu_wu2006@163.com [Department of Cardiology, The First Hospital of Harbin Medical University, 23 Youzheng Street, Harbin 150001 (China); Chen, Sijia, E-mail: chensj0802@163.com [Department of Microbiology and Wu Lien-Teh Institute, Harbin Medical University, 157 Baojian Road, Harbin 150081 (China); Li, Jing, E-mail: jing070822@163.com [Center of Electron Microscopy, Harbin Medical University, 157 Baojian Road, Harbin 150081 (China); Wang, Yan, E-mail: wangyan@hrbmu.edu.cn [Department of Microbiology and Wu Lien-Teh Institute, Harbin Medical University, 157 Baojian Road, Harbin 150081 (China); Zhang, Fengmin, E-mail: fengminzhang@ems.hrbmu.edu.cn [Department of Microbiology and Wu Lien-Teh Institute, Harbin Medical University, 157 Baojian Road, Harbin 150081 (China); Zhao, Wenran, E-mail: zhaowenran2002@aliyun.com [Department of Cell Biology, Harbin Medical University, 157 Baojian Road, Harbin 150081 (China); and others

    2016-12-10

    Coxsackievirus group B (CVB) is one of the common pathogens that cause myocarditis and cardiomyopathy. Evidence has shown that CVB replication in cardiomyocytes is responsible for the damage and loss of cardiac muscle and the dysfunction of the heart. However, it remains largely undefined how CVB would directly impact cardiac fibroblasts, the most abundant cells in human heart. In this study, cardiac fibroblasts were isolated from Balb/c mice and infected with CVB type 3 (CVB3). Increased double-membraned, autophagosome-like vesicles in the CVB3-infected cardiac fibroblasts were observed with electron microscope. Punctate distribution of LC3 and increased level of LC3-II were also detected in the infected cardiac fibroblasts. Furthermore, we observed that the expression of pro-inflammatory cytokines, IL-6 and TNF-α, was increased in the CVB3-infected cardiac fibroblasts, while suppressed autophagy by 3-MA and Atg7-siRNA inhibited cytokine expression. Consistent with the in vitro findings, increased formation of autophagosomes was observed in the cardiac fibroblasts of Balb/c mice infected with CVB3. In conclusion, our data demonstrated that cardiac fibroblasts respond to CVB3 infection with the formation of autophagosomes and the release of the pro-inflammatory cytokines. These results suggest that the autophagic response of cardiac fibroblasts may play a role in the pathogenesis of myocarditis caused by CVB3 infection. - Highlights: • CVB3 replication induced autophagosome assembly in primary cardiac fibroblasts. • Both IL-6 and TNF-α in cardiac fibroblasts infected by CVB3 were increased. • IL-6 and TNF-α were reduced in cardiac fibroblasts when autophagy was inhibited. • Autophagosome assembly in cardiac fibroblasts of CVB-infected mice was increased.

  6. On-chip constructive cell-network study (I): contribution of cardiac fibroblasts to cardiomyocyte beating synchronization and community effect.

    Science.gov (United States)

    Kaneko, Tomoyuki; Nomura, Fumimasa; Yasuda, Kenji

    2011-05-23

    To clarify the role of cardiac fibroblasts in beating synchronization, we have made simple lined-up cardiomyocyte-fibroblast network model in an on-chip single-cell-based cultivation system. The synchronization phenomenon of two cardiomyocyte networks connected by fibroblasts showed (1) propagation velocity of electrophysiological signals decreased a magnitude depending on the increasing number of fibroblasts, not the lengths of fibroblasts; (2) fluctuation of interbeat intervals of the synchronized two cardiomyocyte network connected by fibroblasts did not always decreased, and was opposite from homogeneous cardiomyocyte networks; and (3) the synchronized cardiomyocytes connected by fibroblasts sometimes loses their synchronized condition and recovered to synchronized condition, in which the length of asynchronized period was shorter less than 30 beats and was independent to their cultivation time, whereas the length of synchronized period increased according to cultivation time. The results indicated that fibroblasts can connect cardiomyocytes electrically but do not significantly enhance and contribute to beating interval stability and synchronization. This might also mean that an increase in the number of fibroblasts in heart tissue reduces the cardiomyocyte 'community effect', which enhances synchronization and stability of their beating rhythms.

  7. bFGF regulates PI3-kinase-Rac1-JNK pathway and promotes fibroblast migration in wound healing.

    Directory of Open Access Journals (Sweden)

    Shigeyuki Kanazawa

    Full Text Available Fibroblast proliferation and migration play important roles in wound healing. bFGF is known to promote both fibroblast proliferation and migration during the process of wound healing. However, the signal transduction of bFGF-induced fibroblast migration is still unclear, because bFGF can affect both proliferation and migration. Herein, we investigated the effect of bFGF on fibroblast migration regardless of its effect on fibroblast proliferation. We noticed involvement of the small GTPases of the Rho family, PI3-kinase, and JNK. bFGF activated RhoA, Rac1, PI3-kinase, and JNK in cultured fibroblasts. Inhibition of RhoA did not block bFGF-induced fibroblast migration, whereas inhibition of Rac1, PI3-kinase, or JNK blocked the fibroblast migration significantly. PI3-kinase-inhibited cells down-regulated the activities of Rac1 and JNK, and Rac1-inhibited cells down-regulated JNK activity, suggesting that PI3-kinase is upstream of Rac1 and that JNK is downstream of Rac1. Thus, we concluded that PI3-kinase, Rac1, and JNK were essential for bFGF-induced fibroblast migration, which is a novel pathway of bFGF-induced cell migration.

  8. Synthesis and biological activity of M6-P and M6-P analogs on fibroblast and keratinocyte proliferation.

    Science.gov (United States)

    Clavel, Caroline; Barragan-Montero, Véronique; Garric, Xavier; Molès, Jean-Pierre; Montero, Jean-Louis

    2005-09-01

    A new synthetic route to obtain the carboxylate analog of mannose 6-phosphate (M6-P) is presented. The effects of the M6-P, the carboxylate and two other analogs (the phosphonate and the alpha,beta ethylenic carboxylate) on the proliferation of human keratinocytes and dermal fibroblasts as well as on the proliferation of a murine fibroblast cell line, 3T3-J2 are tested. We observed that M6-P is a potent inhibitor of proliferation of both fibroblasts and keratinocytes. Among its analogs, the phosphonate showed a similar effect on human dermal fibroblasts but not on keratinocytes.

  9. miR-29a-3p attenuates hypoxic pulmonary hypertension by inhibiting pulmonary adventitial fibroblast activation.

    Science.gov (United States)

    Luo, Ying; Dong, Hai-Ying; Zhang, Bo; Feng, Zhao; Liu, Yi; Gao, Yu-Qi; Dong, Ming-Qing; Li, Zhi-Chao

    2015-02-01

    Activation of pulmonary adventitial fibroblasts plays a key role in the pulmonary vascular remodeling in hypoxic pulmonary hypertension. Previous studies showed that miRNAs participated in the regulation of fibroblast activation. This study explored the role of miR-29 in the activation of pulmonary adventitial fibroblasts and the therapeutic potential in hypoxic pulmonary hypertension. We found that hypoxia-induced pulmonary adventitial fibroblasts activation was accompanied with a drastic decrease of miR-29a-3p expression. Knockdown of hypoxia-inducible factor-1 α or Smad3 reversed the hypoxia-induced decrease of miR-29-3p in cultured pulmonary adventitial fibroblasts. In vitro, miR-29a-3p mimic inhibited the hypoxia-induced proliferation, migration, and secretion of pulmonary adventitial fibroblasts, suppressed the hypoxia-induced expression of α-smooth muscle actin and extracellular matrix collagen in pulmonary adventitial fibroblasts; however, miR-29a-3p inhibitor mimicked the effect of hypoxia on the activation of pulmonary adventitial fibroblasts. Further studies revealed that preventative or therapeutic administration of miR-29a-3p significantly decreased pulmonary artery pressure and right ventricle hypertrophy index and ameliorated pulmonary vascular remodeling in hypoxic pulmonary hypertension rats. These findings suggest that miR-29a-3p regulates the activation and phenotype of pulmonary adventitial fibroblasts in hypoxia and has preventative and therapeutic potential in hypoxic pulmonary hypertension. © 2014 American Heart Association, Inc.

  10. Antifibrotic effects of curcumin are associated with overexpression of cathepsins K and L in bleomycin treated mice and human fibroblasts

    National Research Council Canada - National Science Library

    Zhang, Dongwei; Huang, Chuangfang; Yang, Changfu; Liu, Renzuo J; Wang, Jifeng; Niu, Jianzhao; Brömme, Dieter

    2011-01-01

    .... Curcumin, a polyphenol antioxidant from the spice tumeric, has been shown to effectively counteract fibroblast proliferation and reducing inflammation and fibrotic progression in animal models...

  11. Tubule-Derived Wnts Are Required for Fibroblast Activation and Kidney Fibrosis.

    Science.gov (United States)

    Zhou, Dong; Fu, Haiyan; Zhang, Lu; Zhang, Ke; Min, Yali; Xiao, Liangxiang; Lin, Lin; Bastacky, Sheldon I; Liu, Youhua

    2017-08-01

    Cell-cell communication via Wnt ligands is necessary in regulating embryonic development and has been implicated in CKD. Because Wnt ligands are ubiquitously expressed, the exact cellular source of the Wnts involved in CKD remains undefined. To address this issue, we generated two conditional knockout mouse lines in which Wntless (Wls), a dedicated cargo receptor that is obligatory for Wnt secretion, was selectively ablated in tubular epithelial cells or interstitial fibroblasts. Blockade of Wnt secretion by genetic deletion of Wls in renal tubules markedly inhibited myofibroblast activation and reduced renal fibrosis after unilateral ureteral obstruction. This effect associated with decreased activation of β-catenin and downstream gene expression and preserved tubular epithelial integrity. In contrast, fibroblast-specific deletion of Wls exhibited little effect on the severity of renal fibrosis after obstructive or ischemia-reperfusion injury. In vitro, incubation of normal rat kidney fibroblasts with tubule-derived Wnts promoted fibroblast proliferation and activation. Furthermore, compared with kidney specimens from patients without CKD, biopsy specimens from patients with CKD also displayed increased expression of multiple Wnt proteins, predominantly in renal tubular epithelium. These results illustrate that tubule-derived Wnts have an essential role in promoting fibroblast activation and kidney fibrosis via epithelial-mesenchymal communication. Copyright © 2017 by the American Society of Nephrology.

  12. Brimonidine reduces TGF-beta-induced extracellular matrix synthesis in human Tenon's fibroblasts.

    Science.gov (United States)

    Hong, Samin; Han, Sueng-Han; Kim, Chan Yun; Kim, Kang Yoon; Song, Yoo Kyung; Seong, Gong Je

    2015-05-28

    Brimonidine is a highly selective α2 adrenergic agonist that has been widely used in anti-glaucoma eyedrops. The aim of this study was to investigate its putative anti-fibrotic role in the fibrosis caused by activated Tenon's fibroblasts. Primary cultured human Tenon's fibroblasts were exposed to 2.0 ng/mL of transforming growth factor-β1 (TGF-β1) for up to 48 h. In the presence of various concentrations of brimonidine (from 0.0 to 10.0 μM), the expression levels of fibronectin, collagen types I and III, and β-actin were determined by Western immunoblots. The expression of phosphorylated SMAD2/3 (p-SMAD2/3) was then evaluated using immunofluorescence. TGF-β1 significantly increased the synthesis of fibronectin and collagens in human Tenon's fibroblasts; however brimonidine treatment distinctly attenuated the TGF-β1-induced production of extracellular matrix (ECM) proteins. TGF-β1 also changed the cellular morphology to be plump, while brimonidine treatment returned the cells to a spindle shape, similar to control fibroblasts. Regarding p-SMAD2/3, brimonidine treatment did not show any apparent changes in its expression. Our data revealed that brimonidine reduces TGF-β-induced ECM synthesis in human Tenon's fibroblasts in vitro. This finding implies that topical administration of brimonidine may be helpful in reducing the fibrosis caused by the long-term use of topical anti-glaucoma medications.

  13. The prognostic significance of cancer-associated fibroblasts in esophageal squamous cell carcinoma.

    Directory of Open Access Journals (Sweden)

    Sang Yun Ha

    Full Text Available BACKGROUND: Cancer-associated fibroblasts (CAF are activated fibroblasts in the cancer stroma and play an important role in cancer progression. Some reports have indicated the correlation between the expression of CAF markers and adverse prognosis in several cancers. However, no reports have studied CAF phenotype and its clinical relevance in esophageal squamous cell carcinoma (ESCC. METHODS: We investigated CAF phenotype of ESCC based on histology and immunohistochemical expressions of five CAF markers such as fibroblast activation protein (FAP, smooth muscle actin (SMA, fibroblast-specific protein-1 (FSP1, platelet-derived growth factor receptor (PDGFRα, and PDGFRβ in 116 ESCC tissue samples. Besides, we also examined the correlation of the CAF phenotype with clinical relevance as well as other cancer-microenvironment related factors. RESULTS: Histologically immature CAF phenotype was correlated with poor prognosis (p<0.001 and associated with increased microvessel density, increased tumor associated macrophages, and epithelial to mesenchymal transition. CAF markers were characteristically expressed in stromal fibroblast close to tumor cells and the expression pattern of 5 CAF markers was highly heterogeneous in every individual cases. Of five CAF markers, SMA, FSP1, and PDGFRα were unfavorable prognostic indicators of ESCC. The number of positive CAF markers was greater in ESCC with immature CAFs than in those with mature ones. CONCLUSIONS: Our results demonstrate that histologic classification of CAF phenotype is a reliable and significant prognostic predictor in ESCC. CAF markers have the potential to be diagnostic and therapeutic targets in ESCC.

  14. MicroRNA-22 increases senescence and activates cardiac fibroblasts in the aging heart.

    Science.gov (United States)

    Jazbutyte, Virginija; Fiedler, Jan; Kneitz, Susanne; Galuppo, Paolo; Just, Annette; Holzmann, Angelika; Bauersachs, Johann; Thum, Thomas

    2013-06-01

    MicroRNAs (miRs) are small non- coding RNA molecules controlling a plethora of biological processes such as development, cellular survival and senescence. We here determined miRs differentially regulated during cardiac postnatal development and aging. Cardiac function, morphology and miR expression profiles were determined in neonatal, 4 weeks, 6 months and 19 months old normotensive male healthy C57/Bl6N mice. MiR-22 was most prominently upregulated during cardiac aging. Cardiac expression of its bioinformatically predicted target mimecan (osteoglycin, OGN) was gradually decreased with advanced age. Luciferase reporter assays validated mimecan as a bona fide miR-22 target. Both, miR-22 and its target mimecan were co- expressed in cardiac fibroblasts and smooth muscle cells. Functionally, miR-22 overexpression induced cellular senescence and promoted migratory activity of cardiac fibroblasts. Small interference RNA-mediated silencing of mimecan in cardiac fibroblasts mimicked the miR-22-mediated effects. Rescue experiments revealed that the effects of miR-22 on cardiac fibroblasts were only partially mediated by mimecan. In conclusion, miR-22 upregulation in the aging heart contributed at least partly to accelerated cardiac fibroblast senescence and increased migratory activity. Our results suggest an involvement of miR-22 in age-associated cardiac changes, such as cardiac fibrosis.

  15. Anti-fibrotic effects of nintedanib in lung fibroblasts derived from patients with idiopathic pulmonary fibrosis.

    Science.gov (United States)

    Hostettler, Katrin E; Zhong, Jun; Papakonstantinou, Eleni; Karakiulakis, George; Tamm, Michael; Seidel, Petra; Sun, Qingzhu; Mandal, Jyotshna; Lardinois, Didier; Lambers, Christopher; Roth, Michael

    2014-12-12

    Idiopathic pulmonary fibrosis (IPF) is a progressive lung disease with poor prognosis. The kinase inhibitor nintedanib specific for vascular endothelial growth factor receptor (VEGFR), platelet-derived growth factor receptor (PDGFR) and fibroblast growth factor receptor (FGFR) significantly reduced the rate of decline of forced vital capacity versus placebo. To determine the in vitro effect of nintedanib on primary human lung fibroblasts. Fibroblasts were isolated from lungs of IPF patients and from non-fibrotic controls. We assessed the effect of VEGF, PDGF-BB and basic FGF (bFGF) ± nintedanib on: (i) expression/activation of VEGFR, PDGFR, and FGFR, (ii) cell proliferation, secretion of (iii) matrix metalloproteinases (MMP), (iv) tissue inhibitor of metalloproteinase (TIMP), and (v) collagen. IPF fibroblasts expressed higher levels of PDGFR and FGFR than controls. PDGF-BB, bFGF, and VEGF caused a pro-proliferative effect which was prevented by nintedanib. Nintedanib enhanced the expression of pro-MMP-2, and inhibited the expression of TIMP-2. Transforming growth factor-beta-induced secretion of collagens was inhibited by nintedanib. Our data demonstrate a significant anti-fibrotic effect of nintedanib in IPF fibroblasts. This effect consists of the drug's anti-proliferative capacity, and on its effect on the extracellular matrix, the degradation of which seems to be enhanced.

  16. Directly reprogramming fibroblasts into adipogenic, neurogenic and hepatogenic differentiation lineages by defined factors

    Science.gov (United States)

    Wu, Wei; Jin, Yu-Qing; Gao, Zhen

    2017-01-01

    The reprogramming of adult cells into pluripotent cells or directly into alternative adult cell types represents a great potential technology for regenerative medicine. In the present study, the potential of key developmental adipogenic, neurogenic and hepatogenic regulators to reprogram human fibroblasts into adipocytes, neurocytes and hepatocytes was investigated. The results demonstrated that direct reprogramming of octamer-binding transcription factor 4 (Oct4) and CCAAT-enhancer-binding protein (C/EBP)β activated C/EBPα and peroxisome proliferator-activated receptor-γ expression, inducing the conversion of fibroblasts into adipocytes. Similarly, direct reprogramming of the transcription factors sex determining region-box 2, trans-acting T-cell specific transcription factor (GATA-3) and neurogenic differentiation 1 in fibroblasts may induce neurogenic differentiation through hemagglutinating virus of Japan envelope (HVJ-E) transfection. Moreover, hepatogenic differentiation was induced by combining the direct reprogramming of Oct4, GATA-3, hepatocyte nuclear factor 1 homeobox α and forkhead box protein A2 in fibroblasts. These results demonstrate that specific transcription factors and reprogramming factors are able to directly reprogram fibroblasts into adipogenic, neurogenic and hepatogenic differentiation lineages by HVJ-E transfection. PMID:28587331

  17. Distinct function of estrogen receptor α in smooth muscle and fibroblast cells in prostate development.

    Science.gov (United States)

    Vitkus, Spencer; Yeh, Chiuan-Ren; Lin, Hsiu-Hsia; Hsu, Iawen; Yu, Jiangzhou; Chen, Ming; Yeh, Shuyuan

    2013-01-01

    Estrogen signaling, through estrogen receptor (ER)α, has been shown to cause hypertrophy in the prostate. Our recent report has shown that epithelial ERα knockout (KO) will not affect the normal prostate development or homeostasis. However, it remains unclear whether ERα in different types of stromal cells has distinct roles in prostate development. This study proposed to elucidate how KO of ERα in the stromal smooth muscle or fibroblast cells may interrupt cross talk between prostate stromal and epithelial cells. Smooth muscle ERαKO (smERαKO) mice showed decreased glandular infolding with the proximal area