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Sample records for human fen1 expression

  1. Genomic and protein expression analysis reveals Flap endonuclease 1 (FEN1) as a key biomarker in breast and ovarian cancer

    Science.gov (United States)

    Abdel-Fatah, Tarek MA; Russell, Roslin; Albarakati, Nada; Maloney, David J; Dorjsuren, Dorjbal; Rueda, Oscar M; Moseley, Paul; Mohan, Vivek; Sun, Hongmao; Abbotts, Rachel; Mukherjee, Abhik; Agarwal, Devika; Illuzzi, Jennifer L.; Jadhav, Ajit; Simeonov, Anton; Ball, Graham; Chan, Stephen; Caldas, Carlos; Ellis, Ian O; Wilson, David M; Madhusudan, Srinivasan

    2015-01-01

    FEN1 has key roles in Okazaki fragment maturation during replication, long patch base excision repair, rescue of stalled replication forks, maintenance of telomere stability and apoptosis. FEN1 may be dysregulated in breast and ovarian cancers and have clinicopathological significance in patients. We comprehensively investigated FEN1 mRNA expression in multiple cohorts of breast cancer [training set (128), test set (249), external validation (1952)]. FEN1 protein expression was evaluated in 568 oestrogen receptor (ER) negative breast cancers, 894 ER positive breast cancers and 156 ovarian epithelial cancers. FEN1 mRNA overexpression was highly significantly associated with high grade (p=4.89 × 10−57), high mitotic index (p=5.25 × 10−28), pleomorphism (p=6.31 × 10−19), ER negative (p=9.02 × 10−35), PR negative (p=9.24 × 10−24), triple negative phenotype (p=6.67 × 10−21), PAM50.Her2 (p=5.19 × 10−13), PAM50.Basal (p=2.7 × 10−41), PAM50.LumB (p=1.56 × 10−26), integrative molecular cluster 1 (intClust.1) (p=7.47 × 10−12), intClust.5 (p=4.05 × 10−12) and intClust. 10 (p=7.59 × 10−38) breast cancers. FEN1 mRNA overexpression is associated with poor breast cancer specific survival in univariate (p= 4.4 × 10−16) and multivariate analysis (p= 9.19 × 10−7). At the protein level, in ER positive tumours, FEN1 overexpression remains significantly linked to high grade, high mitotic index and pleomorphism (ps<0.01). In ER negative tumours, high FEN1 is significantly associated with pleomorphism, tumour type, lymphovascular invasion, triple negative phenotype, EGFR and HER2 expression (ps<0.05). In ER positive as well as in ER negative tumours, FEN1 protein overexpression is associated with poor survival in univariate and multivariate analysis (ps<0.01). In ovarian epithelial cancers, similarly, FEN1 overexpression is associated with high grade, high stage and poor survival (ps<0.05). We conclude that FEN1 is a promising biomarker in breast

  2. Genomic and protein expression analysis reveals flap endonuclease 1 (FEN1) as a key biomarker in breast and ovarian cancer.

    Science.gov (United States)

    Abdel-Fatah, Tarek M A; Russell, Roslin; Albarakati, Nada; Maloney, David J; Dorjsuren, Dorjbal; Rueda, Oscar M; Moseley, Paul; Mohan, Vivek; Sun, Hongmao; Abbotts, Rachel; Mukherjee, Abhik; Agarwal, Devika; Illuzzi, Jennifer L; Jadhav, Ajit; Simeonov, Anton; Ball, Graham; Chan, Stephen; Caldas, Carlos; Ellis, Ian O; Wilson, David M; Madhusudan, Srinivasan

    2014-10-01

    FEN1 has key roles in Okazaki fragment maturation during replication, long patch base excision repair, rescue of stalled replication forks, maintenance of telomere stability and apoptosis. FEN1 may be dysregulated in breast and ovarian cancers and have clinicopathological significance in patients. We comprehensively investigated FEN1 mRNA expression in multiple cohorts of breast cancer [training set (128), test set (249), external validation (1952)]. FEN1 protein expression was evaluated in 568 oestrogen receptor (ER) negative breast cancers, 894 ER positive breast cancers and 156 ovarian epithelial cancers. FEN1 mRNA overexpression was highly significantly associated with high grade (p = 4.89 × 10(-57)), high mitotic index (p = 5.25 × 10(-28)), pleomorphism (p = 6.31 × 10(-19)), ER negative (p = 9.02 × 10(-35)), PR negative (p = 9.24 × 10(-24)), triple negative phenotype (p = 6.67 × 10(-21)), PAM50.Her2 (p = 5.19 × 10(-13)), PAM50. Basal (p = 2.7 × 10(-41)), PAM50.LumB (p = 1.56 × 10(-26)), integrative molecular cluster 1 (intClust.1) (p = 7.47 × 10(-12)), intClust.5 (p = 4.05 × 10(-12)) and intClust. 10 (p = 7.59 × 10(-38)) breast cancers. FEN1 mRNA overexpression is associated with poor breast cancer specific survival in univariate (p = 4.4 × 10(-16)) and multivariate analysis (p = 9.19 × 10(-7)). At the protein level, in ER positive tumours, FEN1 overexpression remains significantly linked to high grade, high mitotic index and pleomorphism (ps cancers, similarly, FEN1 overexpression is associated with high grade, high stage and poor survival (ps breast and ovarian epithelial cancer.

  3. Studies with the human cohesin establishment factor, ChlR1. Association of ChlR1 with Ctf18-RFC and Fen1.

    Science.gov (United States)

    Farina, Andrea; Shin, Jae-Ho; Kim, Do-Hyung; Bermudez, Vladimir P; Kelman, Zvi; Seo, Yeon-Soo; Hurwitz, Jerard

    2008-07-25

    Human ChlR1 (hChlR1), a member of the DEAD/DEAH subfamily of helicases, was shown to interact with components of the cohesin complex and play a role in sister chromatid cohesion. In order to study the biochemical and biological properties of hChlR1, we purified the protein from 293 cells and demonstrated that hChlR1 possesses DNA-dependent ATPase and helicase activities. This helicase translocates on single-stranded DNA in the 5' to 3' direction in the presence of ATP and, to a lesser extent, dATP. Its unwinding activity requires a 5'-singlestranded region for helicase loading, since flush-ended duplex structures do not support unwinding. The helicase activity of hChlR1 is capable of displacing duplex regions up to 100 bp, which can be extended to 500 bp by RPA or the cohesion establishment factor, the Ctf18-RFC (replication factor C) complex. We show that hChlR1 interacts with the hCtf18-RFC complex, human proliferating cell nuclear antigen, and hFen1. The interactions between Fen1 and hChlR1 stimulate the flap endonuclease activity of Fen1. Selective depletion of either hChlR1 or Fen1 by targeted small interfering RNA treatment results in the precocious separation of sister chromatids. These findings are consistent with a role of hChlR1 in the establishment of sister chromatid cohesion and suggest that its action may contribute to lagging strand processing events important in cohesion.

  4. A cryptic targeting signal creates a mitochondrial FEN1 isoform with tailed R-Loop binding properties.

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    Lawrence Kazak

    Full Text Available A growing number of DNA transacting proteins is found in the nucleus and in mitochondria, including the DNA repair and replication protein Flap endonuclease 1, FEN1. Here we show a truncated FEN1 isoform is generated by alternative translation initiation, exposing a mitochondrial targeting signal. The shortened form of FEN1, which we term FENMIT, localizes to mitochondria, based on import into isolated organelles, immunocytochemistry and subcellular fractionation. In vitro FENMIT binds to flap structures containing a 5' RNA flap, and prefers such substrates to single-stranded RNA. FENMIT can also bind to R-loops, and to a lesser extent to D-loops. Exposing human cells to ethidium bromide results in the generation of RNA/DNA hybrids near the origin of mitochondrial DNA replication. FENMIT is recruited to the DNA under these conditions, and is released by RNase treatment. Moreover, high levels of recombinant FENMIT expression inhibit mtDNA replication, following ethidium bromide treatment. These findings suggest FENMIT interacts with RNA/DNA hybrids in mitochondrial DNA, such as those found at the origin of replication.

  5. The Influence of FEN-1 Gene on Cell Cycle and Genetic Stability%FEN-1基因对细胞周期和遗传稳定性的影响

    Institute of Scientific and Technical Information of China (English)

    石斌山; 余应年; 蔡朱男; 沈炳辉

    2001-01-01

    FEN-1 is essential in the cell replication, repair and in themaintenance of cellular genetic stability. In this report, it was verfied that FEN-1 antisense mRNA fragment was expressed in the cell line FL-FEN-1-, constructed in our lab, blocking FEN-1 gene expression. It was found by the flow cytometer analysis that the cell cycle of FL-FEN-1- cells was delayed in the S-phase DNA synthesis process and arrested in G1 phase. In a mutation assay, based on the shuttle-plasmid pZ189, the spontaneous mutation frequency of SupF tRNA gene in the plasmid in the FL-FEN-1-cells was 19.1×104, while it was 2.9×104 and 3.0×104 in the control cells FL and FL-M, respectively. Further study showed that nontargeted mutation frequency of the FL-FEN-1- cell induced by MNNG was almost the same as the control, indicating that the mutants derived from the block of FEN- 1 gene and the nontargeted mutants may be formed through different passways. The FL-FEN-1 cells exhibit increased sensitivity to alkylating agent MNNG.%结构特异性核酸酶FEN-1在DNA复制和修复,以及维持细胞遗传稳定性方面都起着重要作用。采用RT-PCR方法,证实了FEN-1基因阻断细胞(FL-FEN-1-)通过FEN-1反义mRNA片段的表达,阻断了FEN-1基因。应用流式细胞仪对FL-FEN-1-细胞进行细胞周期分析,发现FL-FEN-1-细胞进入S期DNA合成的过程被延缓,并发生了G1期的阻滞。在由穿梭质粒pZ189介导的突变试验中,质粒在FL-FEN-1-细胞中复制后,其SupFtRNA基因的自发突变频率为19.1×104,而对照细胞FL和FL-M细胞分别为2.9×104和3.0×104,但FLFEN-1-细胞经甲基硝基亚硝基胍(MNNG)诱发后的非定标突变频率与对照细胞并无明显差异,这表明由FEN-1阻断造成的突变频率升高与细胞在受低浓度MNNG攻击后发生的非定标突变可能通过不同的途径形成。FL-FEN-1-细胞对MNNG的敏感性高于对照细胞。

  6. Fen1 mutations that specifically disrupt its interaction with PCNA cause aneuploidy-associated cancer

    Institute of Scientific and Technical Information of China (English)

    Li Zheng; Sankar Mitra; Qin Huang; Kemp H Kernstine; Gerd P Pfeifer; Binghui Shen; Huifang Dai; Muralidhar L Hegde; Mian Zhou; Zhigang Guo; Xiwei Wu; Jun WU; Lei Su; Xueyan Zhong

    2011-01-01

    DNA replication and repair are critical processes for all living organisms to ensure faithful duplication and transmission of genetic information. Flap endonuclease 1 (Feni), a structure-specific nuclease, plays an important role in multiple DNA metabolic pathways and maintenance of genome stability. Human FEN1 mutations that impair its exonuclease activity have been linked to cancer development. FEN1 interacts with multiple proteins, including proliferation cell nuclear antigen (PCNA), to form various functional complexes. Interactions with these proteins are considered to be the key molecular mechanisms mediating FEN1's key biological functions. The current challenge is to experimentally demonstrate the biological consequence of a specific interaction without compromising other functions of a desired protein. To address this issue, we established a mutant mouse model harboring a FEN1 point mutation (F343A/F344A, FFAA), which specifically abolishes the FEN1/PCNA interaction. We show that the FFAA mutation causes defects in RNA primer removal and long-patch base excision repair, even in the heterozygous state, resulting in numerous DNA breaks. These breaks activate the G2/M checkpoint protein, Chk1, and induce neartetraploid aneuploidy, commonly observed in human cancer, consequently elevating the transformation frequency. Consistent with this, inhibition of aneupioidy formation by a Chk1 inhibitor significantly suppressed the cellular transformation. WT/FFAA FEN1 mutant mice develop aneuploidy-associated cancer at a high frequency. Thus, this study establishes an exemplary case for investigating the biological significance of protein-protein interactions by knock-in of a point mutation rather than knock-out of a whole gene.

  7. The Overexpression of FEN1 and RAD54B May Act as Independent Prognostic Factors of Lung Adenocarcinoma.

    Directory of Open Access Journals (Sweden)

    Jau-Chung Hwang

    Full Text Available Synthetic lethality arises when a combination of mutations in two or more genes leads to cell death. However, the prognostic role of concordant overexpression of synthetic lethality genes in protein level rather than a combination of mutations is not clear. In this study, we explore the prognostic role of combined overexpression of paired genes in lung adenocarcinoma. We used immunohistochemical staining to investigate 24 paired genes in 93 lung adenocarcinoma patients and Kaplan-Meier analysis and Cox proportional hazards models to evaluate their prognostic roles. Among 24 paired genes, only FEN1 (Flap endonuclease 1 and RAD54B (RAD54 homolog B were overexpressed in lung adenocarcinoma patients with poor prognosis. Patients with expression of both FEN1 and RAD54B were prone to have advanced nodal involvement and significantly poor prognosis (HR = 2.35, P = 0.0230. These results suggest that intensive follow up and targeted therapy might improve clinical outcome for patients who show expression of both FEN1 and RAD54B.

  8. Role of FEN1 S187 phosphorylation in counteracting oxygen-induced stress and regulating postnatal heart development.

    Science.gov (United States)

    Zhou, Lina; Dai, Huifang; Wu, Jian; Zhou, Mian; Yuan, Hua; Du, Juan; Yang, Lu; Wu, Xiwei; Xu, Hong; Hua, Yuejin; Xu, Jian; Zheng, Li; Shen, Binghui

    2017-01-01

    Flap endonuclease 1 (FEN1) phosphorylation is proposed to regulate the action of FEN1 in DNA repair as well as Okazaki fragment maturation. However, the biologic significance of FEN1 phosphorylation in response to DNA damage remains unknown. Here, we report an in vivo role for FEN1 phosphorylation, using a mouse line carrying S187A FEN1, which abolishes FEN1 phosphorylation. Although S187A mouse embryonic fibroblast cells showed normal proliferation under low oxygen levels (2%), the mutant cells accumulated oxidative DNA damage, activated DNA damage checkpoints, and showed G1-phase arrest at atmospheric oxygen levels (21%). This suggests an essential role for FEN1 phosphorylation in repairing oxygen-induced DNA damage and maintaining proper cell cycle progression. Consistently, the mutant cardiomyocytes showed G1-phase arrest due to activation of the p53-mediated DNA damage response at the neonatal stage, which reduces the proliferation potential of the cardiomyocytes and impairs heart development. Nearly 50% of newborns with the S187A mutant died in the first week due to failure to undergo the peroxisome proliferator-activated receptor signaling-dependent switch from glycolysis to fatty acid oxidation. The adult mutant mice developed dilated hearts and showed significantly shorter life spans. Altogether, our results reveal an important role of FEN1 phosphorylation to counteract oxygen-induced stress in the heart during the fetal-to-neonatal transition.-Zhou, L., Dai, H., Wu, J., Zhou, M., Yuan, H., Du, J., Yang, L., Wu, X., Xu, H., Hua, Y., Xu, J., Zheng, L., Shen, B. Role of FEN1 S187 phosphorylation in counteracting oxygen-induced stress and regulating postnatal heart development. © FASEB.

  9. Human Lacrimal Gland Gene Expression

    Science.gov (United States)

    Aakalu, Vinay Kumar; Parameswaran, Sowmya; Maienschein-Cline, Mark; Bahroos, Neil; Shah, Dhara; Ali, Marwan; Krishnakumar, Subramanian

    2017-01-01

    Background The study of human lacrimal gland biology and development is limited. Lacrimal gland tissue is damaged or poorly functional in a number of disease states including dry eye disease. Development of cell based therapies for lacrimal gland diseases requires a better understanding of the gene expression and signaling pathways in lacrimal gland. Differential gene expression analysis between lacrimal gland and other embryologically similar tissues may be helpful in furthering our understanding of lacrimal gland development. Methods We performed global gene expression analysis of human lacrimal gland tissue using Affymetrix ® gene expression arrays. Primary data from our laboratory was compared with datasets available in the NLM GEO database for other surface ectodermal tissues including salivary gland, skin, conjunctiva and corneal epithelium. Results The analysis revealed statistically significant difference in the gene expression of lacrimal gland tissue compared to other ectodermal tissues. The lacrimal gland specific, cell surface secretory protein encoding genes and critical signaling pathways which distinguish lacrimal gland from other ectodermal tissues are described. Conclusions Differential gene expression in human lacrimal gland compared with other ectodermal tissue types revealed interesting patterns which may serve as the basis for future studies in directed differentiation among other areas. PMID:28081151

  10. Human Neuroepithelial Cells Express NMDA Receptors

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    Cappell B

    2003-11-01

    Full Text Available Abstract L-glutamate, an excitatory neurotransmitter, binds to both ionotropic and metabotropic glutamate receptors. In certain parts of the brain the BBB contains two normally impermeable barriers: 1 cerebral endothelial barrier and 2 cerebral epithelial barrier. Human cerebral endothelial cells express NMDA receptors; however, to date, human cerebral epithelial cells (neuroepithelial cells have not been shown to express NMDA receptor message or protein. In this study, human hypothalamic sections were examined for NMDA receptors (NMDAR expression via immunohistochemistry and murine neuroepithelial cell line (V1 were examined for NMDAR via RT-PCR and Western analysis. We found that human cerebral epithelium express protein and cultured mouse neuroepithelial cells express both mRNA and protein for the NMDA receptor. These findings may have important consequences for neuroepithelial responses during excitotoxicity and in disease.

  11. Critical role for CaFEN1 and CaFEN12 of Candida albicans in cell wall integrity and biofilm formation

    Science.gov (United States)

    Alfatah, Md.; Bari, Vinay K.; Nahar, Anubhav S.; Bijlani, Swati; Ganesan, K.

    2017-01-01

    Sphingolipids are involved in several cellular functions, including maintenance of cell wall integrity. To gain insight into the role of individual genes of sphingolipid biosynthetic pathway, we have screened Saccharomyces cerevisiae strains deleted in these genes for sensitivity to cell wall perturbing agents calcofluor white and congo red. Only deletants of FEN1 and SUR4 genes were found to be sensitive to both these agents. Candida albicans strains deleted in their orthologs, CaFEN1 and CaFEN12, respectively, also showed comparable phenotypes, and a strain deleted for both these genes was extremely sensitive to cell wall perturbing agents. Deletion of these genes was reported earlier to sensitise cells to amphotericin B (AmB), which is a polyene drug that kills the cells mainly by binding and sequestering ergosterol from the plasma membrane. Here we show that their AmB sensitivity is likely due to their cell wall defect. Further, we show that double deletant of C. albicans is defective in hyphae formation as well as biofilm development. Together this study reveals that deletion of FEN1 and SUR4 orthologs of C. albicans leads to impaired cell wall integrity and biofilm formation, which in turn sensitise cells to AmB. PMID:28079132

  12. Expression of immunoreactive urocortin in human tissue

    Institute of Scientific and Technical Information of China (English)

    GU Qing; Vicki L Clifton; CUI Ying; HUI Ning; ZHOU Xiao-ning; HE Qian; HAN Qing-feng; SHA Jin-yan; Roger Smith

    2001-01-01

    To localize where urocortin is expressed in human tissue in an attempt to study its physiological functions. Methods: Expression of immunoreactive urocortin in different human tissue was examined using a specific urocortin antibody and the immunoperoxidase staining method. Results: Immunoreactive urocortin was observed in the anterior pituitary cells, decidual stromal cells, syncytiotrophoblasts, amnion epithelium, the vascular smooth muscles of myometrium, fallopian tube and small intestine. Conclusion: The study indicates that urocortin is expressed in some specific areas of human tissue. The data are consistent with the hypothesis that urocortin is produced locally as an endocrine factor, which may act as a neural regulator and a regulator of local blood flow.

  13. Neuropharmacology of Human Appetite Expression

    Science.gov (United States)

    Halford, Jason C. G.; Harrold, Joanne A.

    2008-01-01

    The regulation of appetite relies on the integration of numerous episodic (meal) and tonic (energy storage) generated signals in energy regulatory centres within the central nervous system (CNS). These centers provide the pharmacological potential to modify human appetite (hunger and satiety) to increase or decrease caloric intake, or to normalize…

  14. Construction and expression of recombined human AFP eukaryotic expression vector

    Institute of Scientific and Technical Information of China (English)

    Li-Wang Zhang; Yang-Lin Pan; Stephen M Festein; Jun Ren; Liang Zhang; Hong-Mei Zhang; Bin Jin; Bo-Rong Pan; Xiao-Ming Si; Yan-Jun Zhang; Zhong-Hua Wang

    2003-01-01

    AIM: To construct a recombined human AFP eukaryotic expression vector for the purpose of gene therapy and target therapy of hepatocellular carcinoma (HCC).METHODS: The full length AFP-cDNA of prokaryotic vector was digested, and subcloned to the multi-clony sites of the eukaryotic vector. The constructed vector was confirmed by enzymes digestion and electrophoresis, and the product expressed was detected by electrochemiluminescence and immunofluorescence methods.RESULTS: The full length AFP-cDNA successfully cloned to the eukaryotic vector through electrophoresis, 0.9723 IU/ml AFP antigen was detected in the supernatant of AFPCHO by electrochemiluminescence method. Compared with the control groups, the differences were significant (P<0.05).AFP antigen molecule was observed in the plasma of AFPCHO by immunofluorescence staining.CONCLUSION: The recombined human AFP eukaryotic expression vector can express in CHO cell line. It provides experimental data for gene therapy and target therapy of hepatocellular carcinoma.

  15. Expression of polarity genes in human cancer.

    Science.gov (United States)

    Lin, Wan-Hsin; Asmann, Yan W; Anastasiadis, Panos Z

    2015-01-01

    Polarity protein complexes are crucial for epithelial apical-basal polarity and directed cell migration. Since alterations of these processes are common in cancer, polarity proteins have been proposed to function as tumor suppressors or oncogenic promoters. Here, we review the current understanding of polarity protein functions in epithelial homeostasis, as well as tumor formation and progression. As most previous studies focused on the function of single polarity proteins in simplified model systems, we used a genomics approach to systematically examine and identify the expression profiles of polarity genes in human cancer. The expression profiles of polarity genes were distinct in different human tissues and classified cancer types. Additionally, polarity expression profiles correlated with disease progression and aggressiveness, as well as with identified cancer types, where specific polarity genes were commonly altered. In the case of Scribble, gene expression analysis indicated its common amplification and upregulation in human cancer, suggesting a tumor promoting function.

  16. Clusterin expression and human testicular seminoma.

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    Tang, Min; Li, Jie; Liu, Bianjiang; Song, Ninghong; Wang, Zengjun; Yin, Changjun

    2013-10-01

    Clusterin expression has a positive correlation with the occurrence and progression of various types of tumors from different genetic backgrounds. Clusterin overexpression may protect tumor cells from apoptosis and damage caused by autoimmunity or anti-tumor therapy. Using immunohistochemisty, one previous study showed that clusterin protein expression is downregulated in human testicular seminoma, which is highly sensitive to radiotherapy and chemotherapy. We thus postulate that clusterin expression in human testicular seminoma differs from clusterin expression in other tumors. It may be the cause of the treatable characteristics of testicular seminoma. In the present preliminary study, we detected the abundance of clusterin mRNA in human testicular seminoma and normal testis. The results showed decreased clusterin expression in seminoma at the gene transcription level. Our primary data and summarized previous literature suggest that the downregulation of clusterin expression may be the cause of the high sensitivity of testicular seminoma to radiotherapy and chemotherapy. It may be that the scarcity of clusterin leaves tumor cells with insufficient protection from treatment. This is the first study to focus on the relationship between clusterin expression and human testicular cancer. Copyright © 2013 Elsevier Ltd. All rights reserved.

  17. IL-21 Receptor Expression in Human Tendinopathy

    Science.gov (United States)

    Campbell, Abigail L.; Smith, Nicola C.; Reilly, James H.; Kerr, Shauna C.; Leach, William J.; Fazzi, Umberto G.; Rooney, Brian P.; Murrell, George A. C.; Millar, Neal L.

    2014-01-01

    The pathogenetic mechanisms underlying tendinopathy remain unclear, with much debate as to whether inflammation or degradation has the prominent role. Increasing evidence points toward an early inflammatory infiltrate and associated inflammatory cytokine production in human and animal models of tendon disease. The IL-21/IL-21R axis is a proinflammatory cytokine complex that has been associated with chronic inflammatory diseases including rheumatoid arthritis and inflammatory bowel disease. This project aimed to investigate the role and expression of the cytokine/receptor pair IL-21/IL-21R in human tendinopathy. We found significantly elevated expression of IL-21 receptor message and protein in human tendon samples but found no convincing evidence of the presence of IL-21 at message or protein level. The level of expression of IL-21R message/protein in human tenocytes was significantly upregulated by proinflammatory cytokines (TNFα/IL-1β) in vitro. These findings demonstrate that IL-21R is present in early human tendinopathy mainly expressed by tenocytes and macrophages. Despite a lack of IL-21 expression, these data again suggest that early tendinopathy has an inflammatory/cytokine phenotype, which may provide novel translational targets in the treatment of tendinopathy. PMID:24757284

  18. IL-21 Receptor Expression in Human Tendinopathy

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    Abigail L. Campbell

    2014-01-01

    Full Text Available The pathogenetic mechanisms underlying tendinopathy remain unclear, with much debate as to whether inflammation or degradation has the prominent role. Increasing evidence points toward an early inflammatory infiltrate and associated inflammatory cytokine production in human and animal models of tendon disease. The IL-21/IL-21R axis is a proinflammatory cytokine complex that has been associated with chronic inflammatory diseases including rheumatoid arthritis and inflammatory bowel disease. This project aimed to investigate the role and expression of the cytokine/receptor pair IL-21/IL-21R in human tendinopathy. We found significantly elevated expression of IL-21 receptor message and protein in human tendon samples but found no convincing evidence of the presence of IL-21 at message or protein level. The level of expression of IL-21R message/protein in human tenocytes was significantly upregulated by proinflammatory cytokines (TNFα/IL-1β in vitro. These findings demonstrate that IL-21R is present in early human tendinopathy mainly expressed by tenocytes and macrophages. Despite a lack of IL-21 expression, these data again suggest that early tendinopathy has an inflammatory/cytokine phenotype, which may provide novel translational targets in the treatment of tendinopathy.

  19. Gene Expression in the Human Endolymphatic Sac

    DEFF Research Database (Denmark)

    Møller, Martin Nue; Kirkeby, Svend; Vikeså, Jonas

    2015-01-01

    OBJECTIVES/HYPOTHESIS: The purpose of the present study is to explore, demonstrate, and describe the expression of genes related to the solute carrier (SLC) molecules of ion transporters in the human endolymphatic sac. STUDY DESIGN: cDNA microarrays and immunohistochemistry were used for analyses...... of fresh human endolymphatic sac tissue samples. METHODS: Twelve tissue samples of the human endolymphatic sac were obtained during translabyrinthine surgery for vestibular schwannoma. Microarray technology was used to investigate tissue sample expression of solute carrier family genes, using adjacent dura...... mater as control. Immunohistochemistry was used for verification of translation of selected genes, as well as localization of the specific protein within the sac. RESULTS: An extensive representation of the SLC family genes were upregulated in the human endolymphatic sac, including SLC26a4 Pendrin, SLC4...

  20. Expression of RHOGTPase regulators in human myometrium

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    Morrison John J

    2008-01-01

    Full Text Available Abstract Background RHOGTPases play a significant role in modulating myometrial contractility in uterine smooth muscle. They are regulated by at least three families of proteins, RHO guanine nucleotide exchange factors (RHOGEFs, RHOGTPase-activating proteins (RHOGAPs and RHO guanine nucleotide inhibitors (RHOGDIs. RHOGEFs activate RHOGTPases from the inactive GDP-bound to the active GTP-bound form. RHOGAPs deactivate RHOGTPases by accelerating the intrinsic GTPase activity of the RHOGTPases, converting them from the active to the inactive form. RHOGDIs bind to GDP-bound RHOGTPases and sequester them in the cytosol, thereby inhibiting their activity. Ezrin-Radixin-Moesin (ERM proteins regulate the cortical actin cytoskeleton, and an ERM protein, moesin (MSN, is activated by and can also activate RHOGTPases. Methods We therefore investigated the expression of various RHOGEFs, RHOGAPs, a RHOGDI and MSN in human myometrium, by semi-quantitative reverse transcription PCR, real-time fluorescence RT-PCR, western blotting and immunofluorescence microscopy. Expression of these molecules was also examined in myometrial smooth muscle cells. Results ARHGEF1, ARHGEF11, ARHGEF12, ARHGAP5, ARHGAP24, ARHGDIA and MSN mRNA and protein expression was confirmed in human myometrium at term pregnancy, at labour and in the non-pregnant state. Furthermore, their expression was detected in myometrial smooth muscle cells. It was determined that ARHGAP24 mRNA expression significantly increased at labour in comparison to the non-labour state. Conclusion This study demonstrated for the first time the expression of the RHOGTPase regulators ARHGEF1, ARHGEF11, ARHGEF12, ARHGAP5, ARHGAP24, ARHGDIA and MSN in human myometrium, at term pregnancy, at labour, in the non-pregnant state and also in myometrial smooth muscle cells. ARHGAP24 mRNA expression significantly increased at labour in comparison to the non-labouring state. Further investigation of these molecules may enable us

  1. Hypoxia induces apelin expression in human adipocytes.

    Science.gov (United States)

    Geiger, K; Muendlein, A; Stark, N; Saely, C H; Wabitsch, M; Fraunberger, P; Drexel, H

    2011-06-01

    Adipokines play a central role in the development of diseases associated with insulin resistance and obesity. Hypoxia in adipose tissue leads to a dysregulation of the expression of adipokines. The effect of hypoxia on the more recently identified adipokine apelin in human adipocytes is unclear. Therefore, we aimed at investigating the role of hypoxia on the expression of the adipokine apelin. Differentiated human Simpson-Golabi-Behmel syndrome (SGBS) adipocytes were cultured under hypoxic conditions for varying time periods. A modular incubator chamber was used to create a hypoxic tissue culture environment (defined as 1% O(2), 94% N, and 5% CO(2)). In addition, hypoxic conditions were mimicked by using CoCl(2). The effect of hypoxia on the expression of the investigated adipokines was measured by real-time PCR and the secretion of apelin was quantified by ELISA. Induction of hypoxia significantly induced mRNA expression of leptin and apelin in differentiated SGBS adipocytes compared with the normoxic control condition. Expression of adiponectin was significantly decreased by hypoxia. In addition, the amount of secreted apelin protein in response to hypoxia was elevated compared to untreated cells. Furthermore, we could demonstrate that the observed hypoxia-induced induction of apelin mRNA expression is in the first phase dependent on HIF-1α. In our study, we could demonstrate for the first time that apelin expression and secretion by human adipocytes are strongly induced under hypoxic conditions and that the early response on hypoxia with apelin induction is dependent on HIF-1α. © Georg Thieme Verlag KG Stuttgart · New York.

  2. Regulation of gene expression in human tendinopathy

    Science.gov (United States)

    2011-01-01

    Background Chronic tendon injuries, also known as tendinopathies, are common among professional and recreational athletes. These injuries result in a significant amount of morbidity and health care expenditure, yet little is known about the molecular mechanisms leading to tendinopathy. Methods We have used histological evaluation and molecular profiling to determine gene expression changes in 23 human patients undergoing surgical procedures for the treatment of chronic tendinopathy. Results Diseased tendons exhibit altered extracellular matrix, fiber disorientation, increased cellular content and vasculature, and the absence of inflammatory cells. Global gene expression profiling identified 983 transcripts with significantly different expression patterns in the diseased tendons. Global pathway analysis further suggested altered expression of extracellular matrix proteins and the lack of an appreciable inflammatory response. Conclusions Identification of the pathways and genes that are differentially regulated in tendinopathy samples will contribute to our understanding of the disease and the development of novel therapeutics. PMID:21539748

  3. Human plasma cells express granzyme B.

    Science.gov (United States)

    Xu, Wei; Narayanan, Priya; Kang, Ning; Clayton, Sandra; Ohne, Yoichiro; Shi, Peiqing; Herve, Marie-Cecile; Balderas, Robert; Picard, Capucine; Casanova, Jean-Laurent; Gorvel, Jean-Pierre; Oh, Sangkon; Pascual, Virginia; Banchereau, Jacques

    2014-01-01

    While studying the plasma cell (PC) compartment in human tonsils, we identified that immunoglobulin kappa or lambda chain-expressing PCs are the main cells expressing granzyme B (GrzB). In vitro studies revealed that activated B cells differentiated into GrzB-expressing PCs when co-cultured with macrophages and follicular helper T cells. This effect could be reproduced on combined stimulation of IL-15 (produced by macrophages) and IL-21 (produced by T follicular helper cells) in a STAT3-dependent manner. Whereas IL-21 triggers the transcription of mRNA of GrzB, IL-15 synergizes the translation of GrzB proteins. The precise role of GrzB in PC biology remains to be understood and studies in mice will not help as their PCs do not express GrzB.

  4. Fibulin-5 expression in the human placenta.

    Science.gov (United States)

    Gauster, Martin; Berghold, Veronika M; Moser, Gerit; Orendi, Kristina; Siwetz, Monika; Huppertz, Berthold

    2011-02-01

    Fibulin-5 is a secreted extracellular matrix glycoprotein and displays a diverse panel of biological functions, which can be segregated into elastogenic as well as extra-elastogenic functions. While elastogenic functions of fibulin-5 include essential roles in early steps of elastic fibre assembly, extra-elastogenic functions are widespread. Depending on the cell type used, fibulin-5 mediates cell adherence via a subset of integrins, antagonizes angiogenesis and inhibits migration as well as proliferation of endothelial and smooth muscle cells. In this study, we focused on the spatiotemporal expression of fibulin-5 in the human placenta. With progressing gestation, placental fibulin-5 expression increased from first trimester towards term. At term, placental fibulin-5 mRNA expression is lower when compared with other well-vascularized organs such as lung, kidney, heart, uterus and testis. In first trimester, placenta immunohistochemistry localized fibulin-5 in villous cytotrophoblasts and extravillous cytotrophoblasts of the proximal cell column. In term placenta, fibulin-5 was detected in the endothelial basement membrane and adventitia-like regions of vessels in the chorionic plate and stem villi. Cell culture experiments with the villous trophoblast-derived cell line BeWo showed that fibulin-5 expression was downregulated during functional differentiation and intercellular fusion. Moreover, cultivation of BeWo cells under low oxygen conditions impaired intercellular fusion and upregulated fibulin-5 expression. The spatiotemporal shift from the trophoblast compartment in first trimester to the villous vasculature at term suggests a dual role of fibulin-5 in human placental development.

  5. Circadian Kisspeptin expression in human term placenta.

    Science.gov (United States)

    de Pedro, M A; Morán, J; Díaz, I; Murias, L; Fernández-Plaza, C; González, C; Díaz, E

    2015-11-01

    Kisspeptin is an essential gatekeeper of reproductive function. During pregnancy high circulating levels of kisspeptin have been described, however the clear role of this neuropeptide in pregnancy remains unknown. We tested the existence of rhythmic kisspeptin expression in human full-term placenta from healthy pregnant women at six different time points during the day. The data obtained by Western blotting were fitted to a mathematical model (Fourier series), demonstrating, for the first time, the existence of a circadian rhythm in placental kisspeptin expression.

  6. The human endolymphatic sac expresses natriuretic peptides

    DEFF Research Database (Denmark)

    Møller, Martin Nue; Kirkeby, Svend; Vikeså, Jonas

    2017-01-01

    in intracranial pressure homeostasis. However, no direct evidence of such capacity exists. This study aims to explore and identify the hypothesized endocrine capacity of the human ES. STUDY DESIGN: DNA microarrays and immunohistochemistry were used for analyses of fresh human ES tissue samples. METHODS: Twelve...... vasopressin receptors and aquaporin-2 channels in the inner ear via OXT expression. We hypothesize that the ES is likely to regulate inner ear endolymphatic homeostasis, possibly through secretion of several peptides, but it may also influence systemic and/or intracranial blood pressure through direct...

  7. An Evolutionarily Conserved Synthetic Lethal Interaction Network Identifies FEN1 as a Broad-Spectrum Target for Anticancer Therapeutic Development

    OpenAIRE

    Derek M. van Pel; Barrett, Irene J.; Yoko Shimizu; Sajesh, Babu V.; Brent J Guppy; Tom Pfeifer; McManus, Kirk J.; Philip Hieter

    2013-01-01

    Harnessing genetic differences between cancerous and noncancerous cells offers a strategy for the development of new therapies. Extrapolating from yeast genetic interaction data, we used cultured human cells and siRNA to construct and evaluate a synthetic lethal interaction network comprised of chromosome instability (CIN) genes that are frequently mutated in colorectal cancer. A small number of genes in this network were found to have synthetic lethal interactions with a large number of canc...

  8. Genes of periodontopathogens expressed during human disease.

    Science.gov (United States)

    Song, Yo-Han; Kozarov, Emil V; Walters, Sheila M; Cao, Sam Linsen; Handfield, Martin; Hillman, Jeffrey D; Progulske-Fox, Ann

    2002-12-01

    Since many bacterial genes are environmentally regulated, the screening for virulence-associated factors using classical genetic and molecular biology approaches can be biased under laboratory growth conditions of a given pathogen, because the required conditions for expression of many virulence factors may not occur during in vitro growth. Thus, technologies have been developed during the past several years to identify genes that are expressed during disease using animal models of human disease. However, animal models are not always truly representative of human disease, and with many pathogens, there is no appropriate animal model. A new technology, in vivo-induced antigen technology (IVIAT) was thus engineered and tested in our laboratory to screen for genes of pathogenic organisms induced specifically in humans, without the use of animal or artificial models of infection. This technology uses pooled sera from patients to probe for genes expressed exclusively in vivo (or ivi, in vivo-induced genes). IVIAT was originally designed for the study of Actinobacillus actinomycetemcomitans pathogenesis, but we have now extended it to other oral pathogens including Porphyromonas gingivalis. One hundred seventy-one thousand (171,000) clones from P. gingivalis strain W83 were screened and 144 were confirmed positive. Over 300,000 A. actinomycetemcomitans clones were probed, and 116 were confirmed positive using a quantitative blot assay. MAT has proven useful in identifying previously unknown in vivo-induced genes that are likely involved in virulence and are thus excellent candidates for use in diagnostic : and therapeutic strategies, including vaccine design.

  9. A Comparison of Artificial Subtle Expressions with Human-like Expressions on Expressing Confidence Level

    Science.gov (United States)

    Komatsu, Takanori; Kobayashi, Kazuki; Yamada, Seiji; Funakoshi, Kotaro; Nakano, Mikio

    Expressing the confidence level of a system's suggestions by using speech sounds is an important cue to users of the system for perceiving how likely it is for the suggestions to be correct. We assume that expressing confidence levels by using human-like expressions would cause users to have a poorer impression of the systems than if artificial subtle expressions (ASEs) were used when the quality of the presented information does not match the expressed confidence level. We confirmed that this assumption was correct by conducting a psychological experiment.

  10. Human basophils express interleukin-4 receptors

    Energy Technology Data Exchange (ETDEWEB)

    Valent, P.; Besemer, J.; Kishi, K.; Di Padova, F.; Geissler, K.; Lechner, K.; Bettelheim, P. (Univ. of Vienna (Austria))

    1990-11-01

    Interleukin-4 (IL-4), a multipotential lymphokine reputed to play an important role in the regulation of immune responses, interacts with a variety of hemopoietic target cells through specific cell surface membrane receptors. The present study was designed to investigate whether human basophils express IL-4 binding sites. For this purpose, basophils were enriched to homogeneity (93% and 98% purity, respectively) from the peripheral blood of two chronic granulocytic leukemia (CGL) donors using a cocktail of monoclonal antibodies (MoAbs) and complement. Purified basophils bound 125I-radiolabeled recombinant human (rh) IL-4 in a specific manner. Quantitative binding studies and Scatchard plot analysis revealed the presence of a single class of high affinity IL-4 binding sites (280 +/- 40 sites per cell in donor 1 and 640 +/- 45 sites per cell in donor 2) with an apparent dissociation constant, kd, of 7.12 x 10(-11) +/- 2.29 x 10(-11) and 9.55 +/- 3.5 x 10(-11) mol/L, respectively. KU812-F, a human basophil precursor cell line, was found to express a single class of 810 to 1,500 high affinity IL-4 binding sites with a kd of 2.63 to 5.54 x 10(-10) mol/L. No change in the numbers or binding constants of IL-4 receptors was found after exposure of KU812-F cells to rhIL-3 (a potent activator of basophils) for 60 minutes. No effect of rhIL-4 on 3H-thymidine uptake, release or synthesis of histamine, or expression of basophil differentiation antigens (Bsp-1, CD11b, CD25, CD40, CD54) on primary human CGL basophils or KU812-F cells was observed.

  11. Brown Fat Expresses Adiponectin in Humans

    Directory of Open Access Journals (Sweden)

    Gianluca Iacobellis

    2013-01-01

    Full Text Available The presence of brown adipose tissue (BAT in humans is unclear. Pheochromocytomas (PHEO are rare tumors of neuroectodermal origin which occur in 0.1-0.2% of patients with hypertension. We sought to evaluate the presence and activity of BAT surrounding adrenal PHEO in a well-studied sample of 11 patients who were diagnosed with PHEO and then underwent adrenalectomy. Areas of white fat (WAT and BAT surrounding PHEO were obtained by Laser Capture Microdissection for analysis of uncoupling protein (UCP-1 and adiponectin mRNA expression. Adiponectin and UCP-1 mRNA levels were significantly higher in BAT than in WAT (0.62 versus 0.15 and 362.4 versus 22.1, resp., for both. Adiponectin mRNA levels significantly correlated with urinary metanephrines (, , vanilly mandelic acid (VMA (, , and serum adiponectin levels (, . Serum adiponectin levels significantly decreased ( μg/mL versus  μg/mL, after adrenalectomy in PHEO subjects. This study provides the following findings: (1 BAT surrounding PHEO expresses adiponectin and UCP-1 mRNA, (2 expression of adiponectin mRNA is significantly higher in BAT than in WAT surrounding PHEO, and (3 catecholamines and serum adiponectin levels significantly correlate with BAT UCP-1 and adiponectin mRNA.

  12. GLUT-3 expression in human skeletal muscle

    Science.gov (United States)

    Stuart, C. A.; Wen, G.; Peng, B. H.; Popov, V. L.; Hudnall, S. D.; Campbell, G. A.

    2000-01-01

    Muscle biopsy homogenates contain GLUT-3 mRNA and protein. Before these studies, it was unclear where GLUT-3 was located in muscle tissue. In situ hybridization using a midmolecule probe demonstrated GLUT-3 within all muscle fibers. Fluorescent-tagged antibody reacting with affinity-purified antibody directed at the carboxy-terminus demonstrated GLUT-3 protein in all fibers. Slow-twitch muscle fibers, identified by NADH-tetrazolium reductase staining, possessed more GLUT-3 protein than fast-twitch fibers. Electron microscopy using affinity-purified primary antibody and gold particle-tagged second antibody showed that the majority of GLUT-3 was in association with triads and transverse tubules inside the fiber. Strong GLUT-3 signals were seen in association with the few nerves that traversed muscle sections. Electron microscopic evaluation of human peripheral nerve demonstrated GLUT-3 within the axon, with many of the particles related to mitochondria. GLUT-3 protein was found in myelin but not in Schwann cells. GLUT-1 protein was not present in nerve cells, axons, myelin, or Schwann cells but was seen at the surface of the peripheral nerve in the perineurium. These studies demonstrated that GLUT-3 mRNA and protein are expressed throughout normal human skeletal muscle, but the protein is predominantly found in the triads of slow-twitch muscle fibers.

  13. The pharmacology of human appetite expression.

    Science.gov (United States)

    Halford, Jason C G; Cooper, Gillian D; Dovey, Terence M

    2004-04-01

    The discovery of the adiposity signal leptin a decade ago revolutionised our understanding of the hypothalamic mechanisms underpinning the central control of ingestive behaviour. Subsequently, the structure and function of various hypothalamic peptide systems (Neuropeptide Y (NPY), Orexins, Melanocortins, Cocaine and Amphetamine Regulating Transcript (CART), Galanin/Galanin Like Peptides (GALP) and endocannabinoids) have been characterised in detail in rodent models. The therapeutic benefit of targeting these systems remains to be discovered. More is becoming known about the pharmacological potential of peripheral, meal-induced, episodic endogenous peptides. Hormones such as Cholecystokinin (CCK), Gastrin Releasing Peptides (GRP), Glucagon-Like Peptide I (GLP-1) Enterostatin, Amylin, Peptide YY (PYY) and Ghrelin are released prior to, during and/or after a meal, controlling intake and subjective feelings of appetite (hunger and satiety). In addition, there is an expanding body of literature detailing the effects of a wide variety of drugs on human appetite and food intake. Some of these drugs act upon CNS monoamine systems such as Serotonin (5-HT). Dopamine (DA) and Noradrenaline (NA), have long been implicated in appetite regulation. Detailed examination of both the effect of agonising endogenous gut peptide systems and the effect of various monoaminergic drugs on the expression of human appetite can provide a greater understanding of mechanisms underpinning normal appetite regulation. However, such an understanding must be based on knowledge of the effect of the treatment on meal size, eating rate, meal pattern, food choice and the subjective experience of appetite flux (hunger and satiety), and notjust food intake.

  14. Positive selection on gene expression in the human brain

    DEFF Research Database (Denmark)

    Khaitovich, Philipp; Tang, Kun; Franz, Henriette

    2006-01-01

    Recent work has shown that the expression levels of genes transcribed in the brains of humans and chimpanzees have changed less than those of genes transcribed in other tissues [1] . However, when gene expression changes are mapped onto the evolutionary lineage in which they occurred, the brain...... shows more changes than other tissues in the human lineage compared to the chimpanzee lineage [1] , [2] and [3] . There are two possible explanations for this: either positive selection drove more gene expression changes to fixation in the human brain than in the chimpanzee brain, or genes expressed...... in the brain experienced less purifying selection in humans than in chimpanzees, i.e. gene expression in the human brain is functionally less constrained. The first scenario would be supported if genes that changed their expression in the brain in the human lineage showed more selective sweeps than other genes...

  15. Human RECQL5beta stimulates flap endonuclease 1

    DEFF Research Database (Denmark)

    Speina, Elzbieta; Dawut, Lale; Hedayati, Mohammad

    2010-01-01

    Human RECQL5 is a member of the RecQ helicase family which is implicated in genome maintenance. Five human members of the family have been identified; three of them, BLM, WRN and RECQL4 are associated with elevated cancer risk. RECQL1 and RECQL5 have not been linked to any human disorder yet; cells...... dramatically stimulates the rate of FEN1 cleavage of flap DNA substrates. Moreover, we show that RECQL5beta and FEN1 interact physically and co-localize in the nucleus in response to DNA damage. Our findings, together with the previous literature on WRN, BLM and RECQL4's stimulation of FEN1, suggests...

  16. EXPRESSION OF Fas LIGAND IN HUMAN COLON CANCER CELL LINES

    Institute of Scientific and Technical Information of China (English)

    张建军; 丁尔迅; 王强; 陈学云; 付志仁

    2001-01-01

    To investigate the expression of Fas ligand in human colon carcinoma cell lines. Methods: A total of six human colon cancer cell lines were examined for the expression of Fas ligand mRNA and cell surface protein by using RT-PCR and flow cytometry respectively. Results: The results showed that Fas ligand mRNA was expressed in all of the six cancer cell lines and Fas ligand cell surface protein was expressed in part of them. Conclusion: These data suggest that Fas ligand was expressed, at least in part, in human colon cancer cell lines and might facilitate to escape from immune surveillance of the host.

  17. Expression and immunohistochemical localization of leptin in human periapical granulomas

    OpenAIRE

    Martín González, Jenifer; Carmona Fernández, Antonio; Pérez Pérez, Antonio; Sánchez Jiménez, Flora; Sánchez-Margalet, Víctor; Segura-Egea, Juan J.

    2015-01-01

    Background Leptin, initially described as an adipocyte-derived hormone to regulate weight control, is expressed in normal and inflamed human dental pulp, being up-regulated during pulp experimental inflammation. Leptin receptor (LER) has been identified in human periapical granulomas. The aim of this study was to analyze and characterize the expression of leptin in human periapical granulomas. Material and Methods Fifteen periapical inflammatory lesions were obtained from extracted human teet...

  18. Cyclooxygenase-2 expression in the normal human eye and its expression pattern in selected eye tumours

    DEFF Research Database (Denmark)

    Wang, Jinmei; Wu, Yazhen; Heegaard, Steffen;

    2011-01-01

    using antibodies against COX-2 was performed on paraffin sections of normal human eyes and selected eye tumours arising from cells expressing COX-2. Results: Cyclooxygenase-2 expression was found in various structures of the normal eye. Abundant expression was seen in the cornea, iris, ciliary body......Purpose: Cyclooxygenase-2 (COX-2) is an enzyme involved in neoplastic processes. The purpose of the present study is to investigate COX-2 expression in the normal human eye and the expression pattern in selected eye tumours involving COX-2 expressing cells. Methods: Immunohistochemical staining...... and retina. The COX-2 expression was less in tumours deriving from the ciliary epithelium and also in retinoblastoma. Conclusion: Cyclooxygenase-2 is constitutively expressed in normal human eyes. The expression of COX-2 is much lower in selected eye tumours involving COX-2 expressing cells....

  19. Cyclooxygenase-2 expression in the normal human eye and its expression pattern in selected eye tumours

    DEFF Research Database (Denmark)

    Wang, Jinmei; Wu, Yazhen; Heegaard, Steffen;

    2011-01-01

    and retina. The COX-2 expression was less in tumours deriving from the ciliary epithelium and also in retinoblastoma. Conclusion: Cyclooxygenase-2 is constitutively expressed in normal human eyes. The expression of COX-2 is much lower in selected eye tumours involving COX-2 expressing cells....

  20. Expression and analysis of the glycosylation properties of recombinant human erythropoietin expressed in Pichia pastoris

    OpenAIRE

    Ser Huy Teh; Mun Yik Fong; Zulqarnain Mohamed

    2011-01-01

    The Pichia pastoris expression system was used to produce recombinant human erythropoietin, a protein synthesized by the adult kidney and responsible for the regulation of red blood cell production. The entire recombinant human erythropoietin (rhEPO) gene was constructed using the Splicing by Overlap Extension by PCR (SOE-PCR) technique, cloned and expressed through the secretory pathway of the Pichia expression system. Recombinant erythropoietin was successfully expressed in P. pastoris. The...

  1. CONSTRUCTION AND EXPRESSION OF A HUMAN-MOUSE CHIMERIC ANTIBODY AGAINST HUMAN BLADDER CANCER

    Institute of Scientific and Technical Information of China (English)

    白银; 王琰; 周丽君; 俞莉章

    2001-01-01

    To construct and express a human-mouse chimeric antibody against human bladder cancer. Method: The variable region genes of anti-human bladder cancer monoclonal antibody BDI-1 were cloned by RT-PCR. A human-mouse chimeric antibody expression vector was constructed and transfected into CHO cells. The chimeric antibody against bladder cancer was expressed and characterized. Result: Eukaryotic expression vector of the chimeric antibody against human bladder carcinoma was successfully constructed, and was expressed in eukaryotic cells; the expressed chimeric antibody ch-BDI showed same specificity as its parent McAb against human bladder cancer cells. Conclusion: The constructed chimeric antibody was expressed successfully in eukaryotic cells, and the chimeric antibody had desired affinity against human bladder cancer cells.

  2. Endothelium specific matrilysin (MMP-7) expression in human cancers

    NARCIS (Netherlands)

    Sier, C.F.M.; Hawinkels, L.J.A.C.; Zijlmans, H.J.M.A.A.; Zuidwijk, K.; Jonge de; Muller, E.S.M.; Ferreira, V.; Hanemaaijer, R.; Mulder-Stapel, A.A.; Kenter, G.G.; Verspaget, H.W.; Gorter, A.

    2008-01-01

    Over-expression of matrilysin (MMP-7) is predominantly associated with epithelial (pre)malignant cells. In the present study MMP-7 expression is also found in endothelial cells in various human cancer types. Endothelial MMP-7 was associated with CD34 and/or CD105 expression. These immunohistochemica

  3. Human thymic epithelial cells express functional HLA-DP molecules

    DEFF Research Database (Denmark)

    Jørgensen, A; Röpke, C; Nielsen, M

    1996-01-01

    T lymphocytes, we examined whether human thymic epithelial cells (TEC) expressed HLA-DP molecules. We present evidence that TEC obtained from short time culture express low but significant levels of HLA-DP molecules. The expression of HLA-DP molecules was comparable to or higher than the expression...... of HLA-DP allospecific primed lymphocyte typing (PLT) CD4 T cell lines. IFN-gamma treatment strongly upregulated the HLA-DP allospecific PLT responses whereas other PLT responses remained largely unchanged. In conclusion, these data indicate that human thymus epithelial cells express significant levels...

  4. Glucose transporter expression in human skeletal muscle fibers

    DEFF Research Database (Denmark)

    Gaster, M; Handberg, A; Beck-Nielsen, H

    2000-01-01

    amplification (TSA) technique to detect the localization of glucose transporter expression in human skeletal muscle. We found expression of GLUT-1, GLUT-3, and GLUT-4 in developing human muscle fibers showing a distinct expression pattern. 1) GLUT-1 is expressed in human skeletal muscle cells during gestation......, but its expression is markedly reduced around birth and is further reduced to undetectable levels within the first year of life; 2) GLUT-3 protein expression appears at 18 wk of gestation and disappears after birth; and 3) GLUT-4 protein is diffusely expressed in muscle cells throughout gestation, whereas...... after birth, the characteristic subcellular localization is as seen in adult muscle fibers. Our results show that GLUT-1, GLUT-3, and GLUT-4 seem to be of importance during muscle fiber growth and development. GLUT-5 protein was undetectable in fetal and adult skeletal muscle fibers. In adult muscle...

  5. Expression of haemopexin receptors by cultured human cytotrophoblast

    NARCIS (Netherlands)

    H.P. van Dijk (Hans); M.J. Kroos; J.S. Starreveld; H.G. van Eijk (Henk); S.P. Tang; D.X. Song

    1995-01-01

    textabstractThe expression of cell-surface haemopexin (Hx) receptors on human cytotrophoblasts was assessed by using four different Hx species purified from plasma: human Hx isolated by wheatgerm-affinity chromatography, human Hx isolated by haem-agarose-affinity

  6. [Human angiogenin: expression, purification, biological assay].

    Science.gov (United States)

    Yang, H; Zhang, Y Q; Yan, Z; Han, W; Yao, L B; Su, C Z

    2001-01-01

    Angiogenin cDNA was obtained by RT-PCR, and cloned into the fusion expression vector pRSETB. The recombinant Angiogenin protein was fused with His6 at its N-terminal and expressed as inclusion body. The expression level was about 10% of the total bacteria protein. After dissolved in 8 mol/L urea, the recombinant protein was purified by Ni2(+)-NTA chelating resin, according to the high affinity of His6 with Ni2+. The biological assay indicated that purified rhANG could induced the new blood vessel formation of CAM and degraded tRNA in vitro.

  7. Genes Expressed in Human Tumor Endothelium

    Science.gov (United States)

    St. Croix, Brad; Rago, Carlo; Velculescu, Victor; Traverso, Giovanni; Romans, Katharine E.; Montgomery, Elizabeth; Lal, Anita; Riggins, Gregory J.; Lengauer, Christoph; Vogelstein, Bert; Kinzler, Kenneth W.

    2000-08-01

    To gain a molecular understanding of tumor angiogenesis, we compared gene expression patterns of endothelial cells derived from blood vessels of normal and malignant colorectal tissues. Of over 170 transcripts predominantly expressed in the endothelium, 79 were differentially expressed, including 46 that were specifically elevated in tumor-associated endothelium. Several of these genes encode extracellular matrix proteins, but most are of unknown function. Most of these tumor endothelial markers were expressed in a wide range of tumor types, as well as in normal vessels associated with wound healing and corpus luteum formation. These studies demonstrate that tumor and normal endothelium are distinct at the molecular level, a finding that may have significant implications for the development of anti-angiogenic therapies.

  8. Rho GTPase expression in human myeloid cells.

    Directory of Open Access Journals (Sweden)

    Suzanne F G van Helden

    Full Text Available Myeloid cells are critical for innate immunity and the initiation of adaptive immunity. Strict regulation of the adhesive and migratory behavior is essential for proper functioning of these cells. Rho GTPases are important regulators of adhesion and migration; however, it is unknown which Rho GTPases are expressed in different myeloid cells. Here, we use a qPCR-based approach to investigate Rho GTPase expression in myeloid cells.We found that the mRNAs encoding Cdc42, RhoQ, Rac1, Rac2, RhoA and RhoC are the most abundant. In addition, RhoG, RhoB, RhoF and RhoV are expressed at low levels or only in specific cell types. More differentiated cells along the monocyte-lineage display lower levels of Cdc42 and RhoV, while RhoC mRNA is more abundant. In addition, the Rho GTPase expression profile changes during dendritic cell maturation with Rac1 being upregulated and Rac2 downregulated. Finally, GM-CSF stimulation, during macrophage and osteoclast differentiation, leads to high expression of Rac2, while M-CSF induces high levels of RhoA, showing that these cytokines induce a distinct pattern. Our data uncover cell type specific modulation of the Rho GTPase expression profile in hematopoietic stem cells and in more differentiated cells of the myeloid lineage.

  9. Expression and effects of human telomerase RNA in testicular tumor

    Institute of Scientific and Technical Information of China (English)

    叶哲伟; 陈晓春; 杨述华; 杨秀萍; 曾汉青; 谷龙杰; 鲁功成

    2004-01-01

    @@ Human telomerase RNA (hTR) plays an important role in determining repeated telomere sequence and the expression of an antisense telomerase RNA that leads to telomere shortening and cell death.1 Using highly sensitive in situ nucleic acid hybridisation, we investigated the expression of hTR in human testicular tumours and located its cellular expression. Our study may help in elucidating the role of hTR in human testicular tumours, finding a highly sensitive diagnostic method and a target for gene therapy of testicular tumours.

  10. Human Empathy, Personality and Experience Affect the Emotion Ratings of Dog and Human Facial Expressions

    OpenAIRE

    Kujala, Miiamaaria V.; Somppi, Sanni; Jokela, Markus; Vainio, Outi; Parkkonen, Lauri

    2017-01-01

    Facial expressions are important for humans in communicating emotions to the conspecifics and enhancing interpersonal understanding. Many muscles producing facial expressions in humans are also found in domestic dogs, but little is known about how humans perceive dog facial expressions, and which psychological factors influence people’s perceptions. Here, we asked 34 observers to rate the valence, arousal, and the six basic emotions (happiness, sadness, surprise, disgust, fear, and anger/aggr...

  11. Expression of histamine receptors in the human endolymphatic sac

    DEFF Research Database (Denmark)

    Møller, M Nue; Kirkeby, S; Vikeså, J.

    2016-01-01

    in 2012. This leaves betahistine (Betaserc) as the only drug for potential prevention of the incapacitating attacks of dizziness, tinnitus and hearing loss. However, the histamine receptors targeted by betahistine have never been demonstrated in the human ES. Accordingly, this study aims to investigate...... the expression of histamine receptors of the human ES epithelium and sub-epithelial stroma. Following sampling of human endolymphatic sac tissue during translabyrinthine surgery, the expression of histamine receptor genes was determined by cDNA microarray analysis. Results were subsequently verified by immuno......-histochemistry. The combined results of microarrays and immuno-histochemistry showed expression of the histamine receptor HRH1 in the epithelial lining of the ES, whereas HRH3 was expressed exclusively in the sub-epithelial capillary network. Receptors HRH2 and -4 were not expressed. The present data provide the first direct...

  12. Expression and characterization of recombinant human serum ...

    African Journals Online (AJOL)

    ajl yemi

    2011-11-14

    Nov 14, 2011 ... 1The Key Laboratory of Industrial Biotechnology, Ministry of Education, Jiangnan University, Wuxi ... might play a role in a broad range of molecular and ... The resulting recombinant expression vector pPIC9K/ HSA-CP was.

  13. Melanopsin expressing human retinal ganglion cells

    DEFF Research Database (Denmark)

    Hannibal, Jens; Christensen, Anders Tolstrup; Heegaard, Steffen

    2017-01-01

    Intrinsically photosensitive retinal ganglion cells (ipRGCs) expressing the photopigment melanopsin belong to a heterogenic population of RGCs which regulate the circadian clock, masking behavior, melatonin suppression, the pupillary light reflex and sleep/wake cycles. The different functions seem...

  14. Expression of connexins in human preimplantation embryos in vitro

    Directory of Open Access Journals (Sweden)

    Leese Henry J

    2004-06-01

    Full Text Available Abstract Intercellular communication via gap junctions is required to coordinate developmental processes in the mammalian embryo. We have investigated if the connexin (Cx isoforms known to form gap junctions in rodent preimplantation embryos are also expressed in human embryos, with the aim of identifying species differences in communication patterns in early development. Using a combination of polyA PCR and immunocytochemistry we have assessed the expression of Cx26, Cx31, Cx32, Cx40, Cx43 and Cx45 which are thought to be important in early rodent embryos. The results demonstrate that Cx31 and Cx43 are the main connexin isoforms expressed in human preimplantation embryos and that these isoforms are co-expressed in the blastocyst. Cx45 protein is expressed in the blastocyst but the protein may be translated from a generally low level of transcripts: which could only be detected in the PN to 4-cell embryos. Interestingly, Cx40, which is expressed by the extravillous trophoblast in the early human placenta, was not found to be expressed in the blastocyst trophectoderm from which this tissue develops. All of the connexin isoforms in human preimplantation embryos are also found in rodents pointing to a common regulation of these connexins in development of rodent and human early embryos and perhaps other species.

  15. Gene profile identifies zinc transporters differentially expressed in normal human organs and human pancreatic cancer.

    Science.gov (United States)

    Yang, J; Zhang, Y; Cui, X; Yao, W; Yu, X; Cen, P; Hodges, S E; Fisher, W E; Brunicardi, F C; Chen, C; Yao, Q; Li, M

    2013-03-01

    Deregulated expression of zinc transporters was linked to several cancers. However, the detailed expression profile of all human zinc transporters in normal human organs and in human cancer, especially in pancreatic cancer is not available. The objectives of this study are to investigate the complete expression patterns of 14 ZIP and 10 ZnT transporters in a large number of normal human organs and in human pancreatic cancer tissues and cell lines. We examined the expression patterns of ZIP and ZnT transporters in 22 different human organs and tissues, 11 pairs of clinical human pancreatic cancer specimens and surrounding normal/benign tissues, as well as 10 established human pancreatic cancer cell lines plus normal human pancreatic ductal epithelium (HPDE) cells, using real time RT-PCR and immunohistochemistry. The results indicate that human zinc transporters have tissue specific expression patterns, and may play different roles in different organs or tissues. Almost all the ZIPs except for ZIP4, and most ZnTs were down-regulated in human pancreatic cancer tissues compared to the surrounding benign tissues. The expression patterns of individual ZIPs and ZnTs are similar among different pancreatic cancer lines. Those results and our previous studies suggest that ZIP4 is the only zinc transporter that is significantly up-regulated in human pancreatic cancer and might be the major zinc transporter that plays an important role in pancreatic cancer growth. ZIP4 might serve as a novel molecular target for pancreatic cancer diagnosis and therapy.

  16. Cytokeratin expression in human fetal tongue and buccal mucosa

    Indian Academy of Sciences (India)

    M M Vaidya; Sharda S Sawant; Anita M Borges; N K Naresh; Manda C Purandare; A N Bhisey

    2000-09-01

    Expression of cytokeratins (CK), a subset of intermediate filament (IF) proteins in epithelia, is developmentally regulated. CK expression may also change after malignant transformation. Our earlier studies on CK expression in human oral tumours and pre-cancerous lesions have shown specific changes in CK expression. We analysed CK expression in human tongue and buccal mucosa (BM) in fetuses in the embryonic age group of 16 to 27 weeks using biochemical and immunohistochemical techniques to find out whether there is any similarity in CK expression in human oral squamous cell carcinomas (SCC) and fetal oral tissues. CK 1, 8 and 18 were detected in a majority of samples using both techniques. Our earlier studies had shown aberrant expression of CK 1 and 18 in many of the oral SCC and leukoplakias. Studies by immunohistochemistry showed that these different CK antigens were expressed in different cell layers. CK 1(2) were present in the stratified epithelial layers whereas CK 8 and 18 were restricted to glandular epithelium. Till 27 weeks of gestation, both tongue and BM expressed CK 1, 8 and 18 along with CK 6 and 16. Thus, fetal tissues showed some similarities in CK pattern with their respective SCC.

  17. Mucin gene expression in human middle ear epithelium.

    Science.gov (United States)

    Kerschner, Joseph Edward

    2007-09-01

    To investigate the expression of recently identified human mucin genes in human middle ear epithelial (MEE) specimens from in vivo middle ear (ME) tissue and to compare this mucin gene expression with mucin gene expression in an immortalized cell culture in vitro source of human MEE. Human MEE was harvested as in vivo specimens, and human MEE cell cultures were established for in vitro experimentation. RNA was extracted from MEE and primers designed for reverse-transcription polymerase chain reaction to assess for mucin gene MUC1, MUC2, MUC3, MUC4, MUC5AC, MUC5B, MUC6, MUC7, MUC8, MUC9, MUC11, MUC12, MUC13, MUC15, MUC16, MUC18, MUC19, and MUC20 expression. Mucin gene expression in the in vivo and in vitro ME tissue was compared against tissues with known expression of the mucin genes in question. Mucin genes MUC1, MUC2, MUC3, MUC4, MUC5AC, MUC5B, MUC7, MUC8, MUC9, MUC11, MUC13, MUC15, MUC16, MUC18, MUC19, and MUC20 were identified and expressed in both the in vivo and in vitro samples of MEE. Mucin genes MUC6, MUC12, and MUC17 were not identified in either tissue samples. Many of the mucin genes that have been recently identified are expressed in human MEE. These genes are expressed in a similar manner in both in vivo and in vitro models. Understanding the mechanisms in which these genes regulate the physiology and pathophysiology of MEE will provide a more thorough understanding of the molecular mechanics of the MEE and disease conditions such as otitis media.

  18. Expressive Writing: Enhancing the Emotional Intelligence of Human Services Majors

    Science.gov (United States)

    Castillo, Yuleinys; Fischer, Jerome M.

    2017-01-01

    The skills and tasks in the human services field are highly connected to emotional intelligence abilities. The purpose of the study was to investigate the effect of an expressive writing program involving human service students in an undergraduate rehabilitation services course. The program was developed to enhance their emotional intelligence.…

  19. Decorin gene expression and its regulation in human keratinocytes

    Energy Technology Data Exchange (ETDEWEB)

    Velez-DelValle, Cristina; Marsch-Moreno, Meytha; Castro-Munozledo, Federico [Department of Cell Biology, Centro de Investigacion y de Estudios Avanzados del IPN, Apdo. Postal 14-740, Mexico D.F. 07000 (Mexico); Kuri-Harcuch, Walid, E-mail: walidkuri@gmail.com [Department of Cell Biology, Centro de Investigacion y de Estudios Avanzados del IPN, Apdo. Postal 14-740, Mexico D.F. 07000 (Mexico)

    2011-07-22

    Highlights: {yields} We showed that cultured human diploid epidermal keratinocytes express and synthesize decorin. {yields} Decorin is found intracytoplasmic in suprabasal cells of cultures and in human epidermis. {yields} Decorin mRNA expression in cHEK is regulated by pro-inflammatory and proliferative cytokines. {yields} Decorin immunostaining of psoriatic lesions showed a lower intensity and altered intracytoplasmic arrangements. -- Abstract: In various cell types, including cancer cells, decorin is involved in regulation of cell attachment, migration and proliferation. In skin, decorin is seen in dermis, but not in keratinocytes. We show that decorin gene (DCN) is expressed in the cultured keratinocytes, and the protein is found in the cytoplasm of differentiating keratinocytes and in suprabasal layers of human epidermis. RT-PCR experiments showed that DCN expression is regulated by pro-inflammatory and proliferative cytokines. Our data suggest that decorin should play a significant role in keratinocyte terminal differentiation, cutaneous homeostasis and dermatological diseases.

  20. Expression Divergence of Tandemly Arrayed Genes in Human and Mouse

    Directory of Open Access Journals (Sweden)

    Valia Shoja

    2007-01-01

    Full Text Available Tandemly arrayed genes (TAGs account for about one third of the duplicated genes in eukaryotic genomes, yet there has not been any systematic study of their gene expression patterns. Taking advantage of recently published large-scale microarray data sets, we studied the expression divergence of 361 two-member TAGs in human and 212 two-member TAGs in mouse and examined the effect of sequence divergence, gene orientation, and chromosomal proximity on the divergence of TAG expression patterns. Our results show that there is a weak negative correlation between sequence divergence of TAG members and their expression similarity. There is also a weak negative correlation between chromosomal proximity of TAG members and their expression similarity. We did not detect any significant relationship between gene orientation and expression similarity. We also found that downstream TAG members do not show significantly narrower expression breadth than upstream members, contrary to what we predict based on TAG expression divergence hypothesis that we propose. Finally, we show that both chromosomal proximity and expression correlation in TAGs do not differ significantly from their neighboring non-TAG gene pairs, suggesting that tandem duplication is unlikely to be the cause for the higher-than-random expression association between neighboring genes on a chromosome in human and mouse.

  1. Neuroglobin and Cytoglobin expression in the human brain

    DEFF Research Database (Denmark)

    Hundahl, Christian Ansgar; Kelsen, Jesper; Hay-Schmidt, Anders

    2013-01-01

    expressed and up-regulated following stroke in the human brain. The present study aimed at confirming our previous observations in rodents using two post-mortem human brains. The anatomical localization of Neuroglobin and Cytoglobin in the human brain is much like what has been described for the rodent...... and Cytoglobin in the cerebral cortex, while no expression in the cerebellar cortex was detectable. We provide a neuroanatomical indication for a different role of Neuroglobin and Cytoglobin in the human brain.......Neuroglobin and Cytoglobin are new members of the heme-globin family. Both globins are primarily expressed in neurons of the brain and retina. Neuroglobin and Cytoglobin have been suggested as novel therapeutic targets in various neurodegenerative diseases based on their oxygen binding and cell...

  2. Downregulation of clusterin expression in human testicular seminoma.

    Science.gov (United States)

    Liu, Bianjiang; Han, Min Tang Zhijian; Zhang, Jiexiu; Lu, Pei; Li, Jie; Song, Ninghong; Wang, Zengjun; Yin, Changjun; Zhang, Wei

    2013-01-01

    Clusterin, a heterodimeric glycoprotein of approximately 80 kDa, exists extensively in human body fluids. The abnormal expression of clusterin is closely related to the occurrence, progression, and prognosis of tumors. Up to now, few studies have focused on clusterin in human testicular cancer. This study describes an extensive exploration of the presence and expression of clusterin in testicular seminoma. Tumor tissues and normal testis tissues were collected from 13 patients with testicular seminoma and 16 patients undergoing surgical castration for prostate cancer. Real-time polymerase chain reaction (PCR) was performed to detect the expression difference of clusterin mRNA between testicular seminoma and normal testis. Western blot and immunohistochemical analysis were performed to detect the presence and expression difference of clusterin protein between two groups. Real-time PCR showed the expression of clusterin mRNA in testicular seminoma to be significantly lower than in normal testis (only 13% relative quantification). Western blot analysis indicated marked reductions in the expression of clusterin protein in testicular seminoma. Similar results were observed upon immunohistochemical analysis. In testicular seminoma and normal testis, clusterin exists in its heterodimeric secretory isoform. Clusterin expression is significantly lower in testicular seminoma than in normal testis. This is the first comprehensive study of the presence and expression of clusterin in human testicular cancer. © 2013 S. Karger AG, Basel

  3. Downregulation of Clusterin Expression in Human Testicular Seminoma

    Directory of Open Access Journals (Sweden)

    Bianjiang Liu

    2013-11-01

    Full Text Available Background: Clusterin, a heterodimeric glycoprotein of approximately 80 kDa, exists extensively in human body fluids. The abnormal expression of clusterin is closely related to the occurrence, progression, and prognosis of tumors. Up to now, few studies have focused on clusterin in human testicular cancer. This study describes an extensive exploration of the presence and expression of clusterin in testicular seminoma. Methods: Tumor tissues and normal testis tissues were collected from 13 patients with testicular seminoma and 16 patients undergoing surgical castration for prostate cancer. Real-time polymerase chain reaction (PCR was performed to detect the expression difference of clusterin mRNA between testicular seminoma and normal testis. Western blot and immunohistochemical analysis were performed to detect the presence and expression difference of clusterin protein between two groups. Results: Real-time PCR showed the expression of clusterin mRNA in testicular seminoma to be significantly lower than in normal testis (only 13% relative quantification. Western blot analysis indicated marked reductions in the expression of clusterin protein in testicular seminoma. Similar results were observed upon immunohistochemical analysis. Conclusion: In testicular seminoma and normal testis, clusterin exists in its heterodimeric secretory isoform. Clusterin expression is significantly lower in testicular seminoma than in normal testis. This is the first comprehensive study of the presence and expression of clusterin in human testicular cancer.

  4. Regenerating human muscle fibres express GLUT3 protein

    DEFF Research Database (Denmark)

    Gaster, M; Beck-Nielsen, H; Schrøder, H D

    2002-01-01

    The presence of the GLUT3 glucose transporter protein in human muscle cells is a matter of debate. The present study was designed to establish whether GLUT3 is expressed in mature human skeletal muscle fibres and, if so, whether its expression changes under different conditions, such as metabolic...... stress (obesity, obese non-insulin-dependent diabetes mellitus), hypertrophy (training), de- and reinnervation (amyotrophic lateral sclerosis) or regeneration (polymyositis). We used an immunohistochemical approach to detect and localise GLUT3. GLUT3 immunoreactivity was not detectable in adult skeletal...... muscle fibres, nor did metabolic stress, training or de- and re-innervation induce GLUT3 expression, while a few GLUT3 expressing fibres were seen in some cases of polymyositis. In contrast, GLUT4 was expressed in all investigated muscle fibres. GLUT3 immunoreactivity was found in perineural...

  5. HCMV Infection Depress NGF Expression in Human Glioma Cells

    Institute of Scientific and Technical Information of China (English)

    Hai-tao WANG; Bin WANG; Zhi-jun LIU; Zhi-qiang BAI; Ling LI; Dong-meng QIAN; Zhi-yong YAN; Xu-xia SONG

    2009-01-01

    Human cytomegalovirus (HCMV) is the most common cause of congenital infection, resulting in birth defects such as microcephaly. In this study, RT-PCR and Western Blotting were performed to quantify the regulation of endogenic nerve growth factor expression in neuroglia cells by HCMV infection. The results showed that basal, endogenous NGF expression in U251 was unchanged during early HCMV infection. NGF expression is strongly down-regulated during the latent phase of infection. These results suggest that HCMV can depress the NGF expression in U251 cells.

  6. Human placental trophoblasts express the immunosuppressive cytokine IL-35.

    Science.gov (United States)

    Mao, Haiting; Gao, Wenjuan; Ma, Chao; Sun, Jintang; Liu, Jia; Shao, Qianqian; Song, Bingfeng; Qu, Xun

    2013-07-01

    Studies of maternal-fetal tolerance focus on defining mechanisms for establishment of immunological privilege within the uterus during pregnancy. Fetal trophoblasts play a key role in maternal tolerance, in part through cytokines production. As a novel inhibitory cytokine, IL-35 is produced by Foxp3(+) regulatory T cells (Tregs) and mediates maximal suppression of Tregs. The purpose of the study is to analyze the expression of IL-35 in first-trimester human placental trophoblasts. IL-35 expression was detected at both protein and mRNA levels by immunohistochemical staining and quantitative real-time PCR method, respectively and secretion of IL-35 was measured by ELISA assay. Our results demonstrated that human trophoblasts constitutively expressed IL-35. Ebi3 and p35 (two subunits of IL-35) mRNA was shown to be co-expressed in trophoblast cells. Moreover, large amounts of secreted IL-35 were detected in the supernatants of trophoblast cells. But we did not detect the constitutive expression of IL-35 in decidual stromal cells. Our findings confirmed for the first time that first-trimester human trophoblast cells expressed and secreted IL-35, which might contribute to their suppressive capacity to maternal immune cells. Therefore, IL-35 may be an important factor of the cytokine network regulating local immune responses during human pregnancy.

  7. Human Empathy, Personality and Experience Affect the Emotion Ratings of Dog and Human Facial Expressions.

    Science.gov (United States)

    Kujala, Miiamaaria V; Somppi, Sanni; Jokela, Markus; Vainio, Outi; Parkkonen, Lauri

    2017-01-01

    Facial expressions are important for humans in communicating emotions to the conspecifics and enhancing interpersonal understanding. Many muscles producing facial expressions in humans are also found in domestic dogs, but little is known about how humans perceive dog facial expressions, and which psychological factors influence people's perceptions. Here, we asked 34 observers to rate the valence, arousal, and the six basic emotions (happiness, sadness, surprise, disgust, fear, and anger/aggressiveness) from images of human and dog faces with Pleasant, Neutral and Threatening expressions. We investigated how the subjects' personality (the Big Five Inventory), empathy (Interpersonal Reactivity Index) and experience of dog behavior affect the ratings of dog and human faces. Ratings of both species followed similar general patterns: human subjects classified dog facial expressions from pleasant to threatening very similarly to human facial expressions. Subjects with higher emotional empathy evaluated Threatening faces of both species as more negative in valence and higher in anger/aggressiveness. More empathetic subjects also rated the happiness of Pleasant humans but not dogs higher, and they were quicker in their valence judgments of Pleasant human, Threatening human and Threatening dog faces. Experience with dogs correlated positively with ratings of Pleasant and Neutral dog faces. Personality also had a minor effect on the ratings of Pleasant and Neutral faces in both species. The results imply that humans perceive human and dog facial expression in a similar manner, and the perception of both species is influenced by psychological factors of the evaluators. Especially empathy affects both the speed and intensity of rating dogs' emotional facial expressions.

  8. A novel SGLT is expressed in the human kidney.

    Science.gov (United States)

    Kothinti, Rajendra K; Blodgett, Amy B; North, Paula E; Roman, Richard J; Tabatabai, Niloofar M

    2012-09-05

    Selective inhibitors of sodium-glucose cotransporter 2 (SGLT2)-mediated reabsorption of glucose in the proximal tubule of the kidney are being developed for the treatment of diabetes. SGLT2 shares high degree of homology with SGLT3; however, very little is known about the expression and functional role of SGLT3 in the human kidney. Indeed, the SGLT2 inhibitors that are currently in clinical trials might affect the expression and/or the activity of SGLT3. Therefore, the present study examined the expression of SGLT3 mRNA and protein in human kidney and in a human proximal tubule HK-2 cell line. The results indicated that human SGLT3 (hSGLT3) message and protein are expressed both in vivo and in vitro. We also studied the activity of hSGLT3 protein following its over-expression in mammalian kidney-derived COS-7 cells and in HK-2 cells treated with the imino sugar deoxynojirimycin (DNJ), a potent agonist of hSGLT3. Over-expression of hSGLT3 in COS-7 cells increased intracellular sodium concentration by 3-fold without affecting glucose transport. Activation of hSGLT3 with DNJ (50μM) increased sodium uptake in HK-2 cells by 5.5 fold and this effect could be completely blocked with SGLT inhibitor phlorizin (50μM). These results suggest that SGLT3 is expressed in human proximal tubular cells where it serves as a novel sodium transporter. Up-regulation of the expression of SGLT3 in the proximal tubule in diabetic patients may contribute to the elevated sodium transport in this segment of the nephron that has been postulated to promote hyperfiltration and renal injury.

  9. FasL EXPRESSION IN HUMAN COLON CARCINOMAS

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    Objective:The Fas and Fas ligand (FasL) play an important role in maintaining immune privilege on malignant tumors. In present study, we investigated the expression of FasL in SW480 and LS174 human colon carcinoma cell lines and twenty primary colon carcinoma specimens. Methods: The expression of FasL in human colon carcinoma cell lines and primary colon carcinomas specimens was detected by immunohistochemistry and Reverse Transcription-PCR (RT-PCR). Results: We found that all of detected human colon carcinoma cell lines and primary colon carcinoma specimens constitutively expressed FasL at the mRNA and protein level. However, the expression of FasL was not found in normal colon epithelial cells. Conclusion: The expression of FasL may occur during malignant transformation from normal colon epithelial cells to colon carcinoma cells. Our results suggest that tumor cells kill cytotoxic T lymphocytes (CTLS) and natural killer (NK) cells by expression of FasL. It may be a new mechanism for tumor cells to escape the host's immune surveillance. The expression of FasL may contribute to the formation of colon carcinomas.

  10. Cloning and expression of human colon mast cell carboxypeptidase

    Institute of Scientific and Technical Information of China (English)

    Zhang-Quan Chen; Shao-Heng He

    2004-01-01

    AIM: To clone and express the human colon mast cell METHODS: Total RNA was extracted from colon tissue, and the cDNA encoding human colon mast cell carboxypeptidase was amplified by reverse-transcription PCR (RT-PCR). The product cDNA was subcloned into the prokaryotic expression vector pMAL-c2x and eukaryotic expression vector pPIC9K to conrtruct prokaryotic expression vector pMAL/human MC-CP (hMC-CP) and eukaryotic pPIC9K/hMC-CP. The recombinant fusion protein expressed in E.coli was induced with IPTG and purified by amylose affinity chromatography. After digestion with factor Xa, recombinant hMC-CP was purified by heparin agarose chromatography. The recombinant hMC-CP expressed in Pichia pastoris (P.pastoris) was induced with methanol and analyzed by SDS-PAGE, Western blot, N-terminal amino acid RESULTS: The cDNA encoding the human colon mast cell carboxypeptidase was cloned, which had five nucleotide variations compared with skin MC-CP cDNA. The recombinant hMC-CP protein expressed in E.coli was purified with amylose affinity chromatography and heparin agarose chromatogphy.SDS-PAGE and Western blot analysis showed that the recombinant protein expressed by E. coli had a molecular weight of 36 kDa and reacted to the anti-native hMC-CP monoclonal antibody (CA5). The N-terminal amino acid sequence confirmed further the product was hMC-CP. E. coli generated hMC-CP showed a very low level of enzymatic activity, but P. pastoris produced hMC-CP had a relatively high enzymatic activity towards a synthetic substrate hippuryl-L-phenylalanine.carboxypeptidase can be successfully cloned and expressed in E.coli and P. pastoris, which will contribute greatly to the fonctional study on hMC-CP.

  11. Modulation of Progesterone Receptor Isoform Expression in Pregnant Human Myometrium

    Directory of Open Access Journals (Sweden)

    Marina Ilicic

    2017-01-01

    Full Text Available Background. Regulation of myometrial progesterone receptor (PR expression is an unresolved issue central to understanding the mechanism of functional progesterone withdrawal and initiation of labor in women. Objectives. To determine whether pregnant human myometrium undergoes culture-induced changes in PR isoform expression ex situ and, further, to determine if conditions approaching the in vivo environment stabilise PR isoform expression in culture. Methods. Term nonlaboring human myometrial tissues were cultured under specific conditions: serum supplementation, steroids, stretch, cAMP, PMA, PGF2α, NF-κB inhibitors, or TSA. Following 48 h culture, PR-T, PR-A, and PR-B mRNA levels were determined using qRT-PCR. PR-A/PR-B ratios were calculated. Results. PR-T and PR-A expression and the PR-A/PR-B ratio significantly increased in culture. Steroids prevented the culture-induced increase in PR-T and PR-A expression. Stretch blocked the effects of steroids on PR-T and PR-A expression. PMA further increased the PR-A/PR-B ratio, while TSA blocked culture-induced increases of PR-A expression and the PR-A/PR-B ratio. Conclusion. Human myometrial tissue in culture undergoes changes in PR gene expression consistent with transition toward a laboring phenotype. TSA maintained the nonlaboring PR isoform expression pattern. This suggests that preserving histone and/or nonhistone protein acetylation is critical for maintaining the progesterone dependent quiescent phenotype of human myometrium in culture.

  12. Modulation of Progesterone Receptor Isoform Expression in Pregnant Human Myometrium

    Science.gov (United States)

    2017-01-01

    Background. Regulation of myometrial progesterone receptor (PR) expression is an unresolved issue central to understanding the mechanism of functional progesterone withdrawal and initiation of labor in women. Objectives. To determine whether pregnant human myometrium undergoes culture-induced changes in PR isoform expression ex situ and, further, to determine if conditions approaching the in vivo environment stabilise PR isoform expression in culture. Methods. Term nonlaboring human myometrial tissues were cultured under specific conditions: serum supplementation, steroids, stretch, cAMP, PMA, PGF2α, NF-κB inhibitors, or TSA. Following 48 h culture, PR-T, PR-A, and PR-B mRNA levels were determined using qRT-PCR. PR-A/PR-B ratios were calculated. Results. PR-T and PR-A expression and the PR-A/PR-B ratio significantly increased in culture. Steroids prevented the culture-induced increase in PR-T and PR-A expression. Stretch blocked the effects of steroids on PR-T and PR-A expression. PMA further increased the PR-A/PR-B ratio, while TSA blocked culture-induced increases of PR-A expression and the PR-A/PR-B ratio. Conclusion. Human myometrial tissue in culture undergoes changes in PR gene expression consistent with transition toward a laboring phenotype. TSA maintained the nonlaboring PR isoform expression pattern. This suggests that preserving histone and/or nonhistone protein acetylation is critical for maintaining the progesterone dependent quiescent phenotype of human myometrium in culture. PMID:28540297

  13. Rescue and expression of human immunoglobulin genes to generate functional human monoclonal antibodies.

    Science.gov (United States)

    Lewis, A P; Parry, N; Peakman, T C; Crowe, J S

    1992-07-01

    Human monoclonal antibody production has been hampered for many years by the instability of cell lines and low levels of expression of the antibodies. We describe here the rescue of human immunoglobulin genes utilizing micro-mRNA preparation from a small number of human hybridoma cells and conventional cDNA cloning. This allows cloning and immediate high-level expression from full-length human heavy and light chain cDNA molecules and provides a mechanism to rescue whole human monoclonal antibodies of proven efficacy.

  14. Expression of human adenosine deaminase in murine hematopoietic cells.

    Science.gov (United States)

    Belmont, J W; MacGregor, G R; Wager-Smith, K; Fletcher, F A; Moore, K A; Hawkins, D; Villalon, D; Chang, S M; Caskey, C T

    1988-01-01

    Multiple replication-defective retrovirus vectors were tested for their ability to transfer and express human adenosine deaminase in vitro and in vivo in a mouse bone marrow transplantation model. High-titer virus production was obtained from vectors by using both a retrovirus long terminal repeat promoter and internal transcriptional units with human c-fos and herpes virus thymidine kinase promoters. After infection of primary murine bone marrow with one of these vectors, human adenosine deaminase was detected in 60 to 85% of spleen colony-forming units and in the blood of 14 of 14 syngeneic marrow transplant recipients. This system offers the opportunity to assess methods for increasing efficiency of gene transfer, for regulation of expression of foreign genes in hematopoietic progenitors, and for long-term measurement of the stability of expression in these cells. Images PMID:3072474

  15. Monoallelic expression of the human FOXP2 speech gene.

    Science.gov (United States)

    Adegbola, Abidemi A; Cox, Gerald F; Bradshaw, Elizabeth M; Hafler, David A; Gimelbrant, Alexander; Chess, Andrew

    2015-06-02

    The recent descriptions of widespread random monoallelic expression (RMAE) of genes distributed throughout the autosomal genome indicate that there are more genes subject to RMAE on autosomes than the number of genes on the X chromosome where X-inactivation dictates RMAE of X-linked genes. Several of the autosomal genes that undergo RMAE have independently been implicated in human Mendelian disorders. Thus, parsing the relationship between allele-specific expression of these genes and disease is of interest. Mutations in the human forkhead box P2 gene, FOXP2, cause developmental verbal dyspraxia with profound speech and language deficits. Here, we show that the human FOXP2 gene undergoes RMAE. Studying an individual with developmental verbal dyspraxia, we identify a deletion 3 Mb away from the FOXP2 gene, which impacts FOXP2 gene expression in cis. Together these data suggest the intriguing possibility that RMAE impacts the haploinsufficiency phenotypes observed for FOXP2 mutations.

  16. CHARACTERIZATION OF THE OLFACTORY RECEPTORS EXPRESSED IN HUMAN SPERMATOZOA

    Directory of Open Access Journals (Sweden)

    Caroline eFlegel

    2016-01-01

    Full Text Available The detection of external cues is fundamental for human spermatozoa to locate the oocyte in the female reproductive tract. This task requires a specific chemoreceptor repertoire that is expressed on the surface of human spermatozoa, which is not fully identified to date. Olfactory receptors (ORs are candidate molecules and have been attributed to be involved in sperm chemotaxis and chemokinesis, indicating an important role in mammalian spermatozoa. An increasing importance has been suggested for spermatozoal RNA, which led us to investigate the expression of all 387 OR genes. This study provides the first comprehensive analysis of OR transcripts in human spermatozoa of several individuals by RNA-Seq. We detected 91 different transcripts in the spermatozoa samples that could be aligned to annotated OR genes. Using stranded mRNA-Seq, we detected a class of these putative OR transcripts in an antisense orientation, indicating a different function, rather than coding for a functional OR protein. Nevertheless, we were able to detect OR proteins in various compartments of human spermatozoa, indicating distinct functions in human sperm. A panel of various OR ligands induced Ca2+ signals in human spermatozoa, which could be inhibited by mibefradil. This study indicated that a variety of ORs are expressed at the mRNA and protein level in human spermatozoa and demonstrates that ORs are involved in the physiological processes.

  17. Four zona pellucida glycoproteins are expressed in the human.

    Science.gov (United States)

    Lefièvre, L; Conner, S J; Salpekar, A; Olufowobi, O; Ashton, P; Pavlovic, B; Lenton, W; Afnan, M; Brewis, I A; Monk, M; Hughes, D C; Barratt, C L R

    2004-07-01

    The zona pellucida (ZP) is an extracellular glycoprotein matrix which surrounds all mammalian oocytes. Recent data have shown the presence of four human zona genes (ZP1, ZP2, ZP3 and ZPB). The aim of the study was to determine if all four ZP proteins are expressed and present in the human. cDNA derived from human oocytes were used to amplify by PCR the four ZP genes. In addition, isolated native human ZP were heat-solubilized, trypsin-digested and subjected to tandem mass spectrometry (MS/MS). All four genes were expressed and the respective proteins present in the human ZP. Moreover, a bioinformatics approach showed that the mouse ZPB gene, although present, is likely to encode a non-functional protein. Four ZP genes are expressed in human oocytes (ZP1, ZP2, ZP3 and ZPB) and preliminary data show that the four corresponding ZP proteins are present in the human ZP. Therefore, this is a fundamental difference with the mouse model

  18. Genetic control of human brain transcript expression in Alzheimer disease.

    Science.gov (United States)

    Webster, Jennifer A; Gibbs, J Raphael; Clarke, Jennifer; Ray, Monika; Zhang, Weixiong; Holmans, Peter; Rohrer, Kristen; Zhao, Alice; Marlowe, Lauren; Kaleem, Mona; McCorquodale, Donald S; Cuello, Cindy; Leung, Doris; Bryden, Leslie; Nath, Priti; Zismann, Victoria L; Joshipura, Keta; Huentelman, Matthew J; Hu-Lince, Diane; Coon, Keith D; Craig, David W; Pearson, John V; Heward, Christopher B; Reiman, Eric M; Stephan, Dietrich; Hardy, John; Myers, Amanda J

    2009-04-01

    We recently surveyed the relationship between the human brain transcriptome and genome in a series of neuropathologically normal postmortem samples. We have now analyzed additional samples with a confirmed pathologic diagnosis of late-onset Alzheimer disease (LOAD; final n = 188 controls, 176 cases). Nine percent of the cortical transcripts that we analyzed had expression profiles correlated with their genotypes in the combined cohort, and approximately 5% of transcripts had SNP-transcript relationships that could distinguish LOAD samples. Two of these transcripts have been previously implicated in LOAD candidate-gene SNP-expression screens. This study shows how the relationship between common inherited genetic variants and brain transcript expression can be used in the study of human brain disorders. We suggest that studying the transcriptome as a quantitative endo-phenotype has greater power for discovering risk SNPs influencing expression than the use of discrete diagnostic categories such as presence or absence of disease.

  19. Gene expression profiling of human erythroid progenitors by micro-serial analysis of gene expression.

    Science.gov (United States)

    Fujishima, Naohito; Hirokawa, Makoto; Aiba, Namiko; Ichikawa, Yoshikazu; Fujishima, Masumi; Komatsuda, Atsushi; Suzuki, Yoshiko; Kawabata, Yoshinari; Miura, Ikuo; Sawada, Ken-ichi

    2004-10-01

    We compared the expression profiles of highly purified human CD34+ cells and erythroid progenitor cells by micro-serial analysis of gene expression (microSAGE). Human CD34+ cells were purified from granulocyte colony-stimulating factor-mobilized blood stem cells, and erythroid progenitors were obtained by cultivating these cells in the presence of stem cell factor, interleukin 3, and erythropoietin. Our 10,202 SAGE tags allowed us to identify 1354 different transcripts appearing more than once. Erythroid progenitor cells showed increased expression of LRBA, EEF1A1, HSPCA, PILRB, RANBP1, NACA, and SMURF. Overexpression of HSPCA was confirmed by real-time polymerase chain reaction analysis. MicroSAGE revealed an unexpected preferential expression of several genes in erythroid progenitor cells in addition to the known functional genes, including hemoglobins. Our results provide reference data for future studies of gene expression in various hematopoietic disorders, including myelodysplastic syndrome and leukemia.

  20. Expression and analysis of the glycosylation properties of recombinant human erythropoietin expressed in Pichia pastoris

    Directory of Open Access Journals (Sweden)

    Ser Huy Teh

    2011-01-01

    Full Text Available The Pichia pastoris expression system was used to produce recombinant human erythropoietin, a protein synthesized by the adult kidney and responsible for the regulation of red blood cell production. The entire recombinant human erythropoietin (rhEPO gene was constructed using the Splicing by Overlap Extension by PCR (SOE-PCR technique, cloned and expressed through the secretory pathway of the Pichia expression system. Recombinant erythropoietin was successfully expressed in P. pastoris. The estimated molecular mass of the expressed protein ranged from 32 kDa to 75 kDa, with the variation in size being attributed to the presence of rhEPO glycosylation analogs. A crude functional analysis of the soluble proteins showed that all of the forms were active in vivo.

  1. Cyclooxygenase-2 Expression in Hamster and Human Pancreatic Neoplasia

    Directory of Open Access Journals (Sweden)

    Pamela L. Crowell

    2006-06-01

    Full Text Available Cyclooxygenase-2 (COX-2 has been implicated in the development of gastrointestinal malignancies. The aim of the present study was to determine COX-2 expression/activity throughout stages of experimental and human pancreatic neoplasia. COX-2 immunohistochemistry was performed in pancreata of hamsters subjected to the carcinogen N-nitrosobis-(2-oxopropylamine (BOP and in human pancreatic tumors. COX-2 activity was determined by prostaglandin E2 assay in tumor versus matched normal pancreatic tissues. The activity of the COX inhibitor sulindac was tested in the PC-1 hamster pancreatic cancer model. COX-2 expression was elevated in all pancreatic intraepithelial neoplasias (PanINs and adenocarcinomas. In BOP-treated hamsters, there were significant progressive elevations in COX-2 expression throughout pancreatic tumorigenesis. In human samples, peak COX-2 expression occurred in PanIN2 lesions and remained moderately elevated in PanIN3 and adenocarcinoma tissues. COX-2 activity was significantly elevated in hamster and human pancreatic cancers compared to pair-matched normal pancreas. Furthermore, hamster pancreatic tumor engraftment/formation in the PC-1 hamster pancreatic cancer model was reduced 4.9-fold by oral administration of sulindac. Increased COX-2 expression is an early event in pancreatic carcinogeneses. The BOP-induced hamster carcinogenesis model is a representative model used to study the role of COX-2 in well-differentiated pancreatic tumorigenesis. COX inhibitors may have a role in preventing tumor engraftment/formation.

  2. Expression profiling of insulin action in human myotubes

    DEFF Research Database (Denmark)

    Hansen, Lars; Gaster, Michael; Oakeley, Edward J

    2004-01-01

    ), 0.5, 1, 2, 4, 8, and 24 h, mRNA contents were analyzed in human myotubes for each time point using Affymetrix DNA chip technology. Insulin treatment induced an inflammatory and pro-angiogenic response in the myotubes, with expression of early response factors followed by inflammatory chemokines...... of diabetic skeletal muscle. We conclude, (i) that insulin induces a time-dependent inflammatory and pro-angiogenic transcriptional response in cultured human myotubes, (ii) that myotubes in vitro retain a gene expression pattern specific for type 2 diabetes and sharing five genes with that of type 2 diabetic...

  3. Converging Strategies in Expression of Human Complex Retroviruses

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    Ilaria Cavallari

    2011-08-01

    Full Text Available The discovery of human retroviruses in the early 1980s revealed the existence of viral-encoded non-structural genes that were not evident in previously described animal retroviruses. Based on the absence or presence of these additional genes retroviruses were classified as ‘simple’ and ‘complex’, respectively. Expression of most of these extra genes is achieved through the generation of alternatively spliced mRNAs. The present review summarizes the genetic organization and expression strategies of human complex retroviruses and highlights the converging mechanisms controlling their life cycles.

  4. Automated discovery of functional generality of human gene expression programs.

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    Georg K Gerber

    2007-08-01

    Full Text Available An important research problem in computational biology is the identification of expression programs, sets of co-expressed genes orchestrating normal or pathological processes, and the characterization of the functional breadth of these programs. The use of human expression data compendia for discovery of such programs presents several challenges including cellular inhomogeneity within samples, genetic and environmental variation across samples, uncertainty in the numbers of programs and sample populations, and temporal behavior. We developed GeneProgram, a new unsupervised computational framework based on Hierarchical Dirichlet Processes that addresses each of the above challenges. GeneProgram uses expression data to simultaneously organize tissues into groups and genes into overlapping programs with consistent temporal behavior, to produce maps of expression programs, which are sorted by generality scores that exploit the automatically learned groupings. Using synthetic and real gene expression data, we showed that GeneProgram outperformed several popular expression analysis methods. We applied GeneProgram to a compendium of 62 short time-series gene expression datasets exploring the responses of human cells to infectious agents and immune-modulating molecules. GeneProgram produced a map of 104 expression programs, a substantial number of which were significantly enriched for genes involved in key signaling pathways and/or bound by NF-kappaB transcription factors in genome-wide experiments. Further, GeneProgram discovered expression programs that appear to implicate surprising signaling pathways or receptor types in the response to infection, including Wnt signaling and neurotransmitter receptors. We believe the discovered map of expression programs involved in the response to infection will be useful for guiding future biological experiments; genes from programs with low generality scores might serve as new drug targets that exhibit minimal

  5. Importance of the brow in facial expressiveness during human communication.

    Science.gov (United States)

    Neely, John Gail; Lisker, Paul; Drapekin, Jesse

    2014-03-01

    The objective of this study was to evaluate laterality and upper/lower face dominance of expressiveness during prescribed speech using a unique validated image subtraction system capable of sensitive and reliable measurement of facial surface deformation. Observations and experiments of central control of facial expressions during speech and social utterances in humans and animals suggest that the right mouth moves more than the left during nonemotional speech. However, proficient lip readers seem to attend to the whole face to interpret meaning from expressed facial cues, also implicating a horizontal (upper face-lower face) axis. Prospective experimental design. Experimental maneuver: recited speech. image-subtraction strength-duration curve amplitude. Thirty normal human adults were evaluated during memorized nonemotional recitation of 2 short sentences. Facial movements were assessed using a video-image subtractions system capable of simultaneously measuring upper and lower specific areas of each hemiface. The results demonstrate both axes influence facial expressiveness in human communication; however, the horizontal axis (upper versus lower face) would appear dominant, especially during what would appear to be spontaneous breakthrough unplanned expressiveness. These data are congruent with the concept that the left cerebral hemisphere has control over nonemotionally stimulated speech; however, the multisynaptic brainstem extrapyramidal pathways may override hemiface laterality and preferentially take control of the upper face. Additionally, these data demonstrate the importance of the often-ignored brow in facial expressiveness. Experimental study. EBM levels not applicable.

  6. Expression of osteopontin in human endometrium throughout menstrual cycle

    Institute of Scientific and Technical Information of China (English)

    Ma Cai-hong; Chen Gui-an; Han Jin-song; Qi Yong-fen

    2005-01-01

    Objective:To analyze mRNA and protein expression of osteopontin(OPN) in human endometrium throughout menstrual cycle.Methods:Immunohistochemical method was used to determine the level and the location of OPN in normal cycling endometrium of 50 women. Western Blot analysis was also used to detect the existence of OPN at proliferative and secretory phases in six samples. The levels of OPN mRNA in 18 samples were measured by RT-PCR. Results:The OPN mainly expressed in human endometrial glandular epithelium but not in stromal cells. The expression was highest at mid and late secretory phases and menstruous phase as well. No expression was found in early and mid proliferative endometria. Two subtypes of OPN proteins (40 kDa-75 kDa) were detected by Western blot analysis in homogenized endometria at secretory phase.The level of OPN mRNA in secretory phase endometrium was significantly higher than that in proliferative phase. Conclusions:OPN was only found in endometrial glandular epithelial cells, and the OPN and its mRNA showed a pronounced cycle-dependent expression in human endometrium, higher expression at mid and late secretory phases and menstruous phase. It's probably involved in embryonic implantation and endometrial shedding.

  7. Cloning and expression of special F protein from human liver

    Institute of Scientific and Technical Information of China (English)

    Shu-Ye Liu; Xin-Da Yu; Chun-Juan Song; Wei Lu; Jian-Dong Zhang; Xin-Rong Shi; Ying Duan; Ju Zhang

    2007-01-01

    AIM:To clone human liver special F protein and to express it in a prokaryotic system.METHODS:Total RNA was isolated from human liver tissue and first-strand cDNA was reverse transcribed using the PCR reverse primer. Following this,cDNA of the F protein was ligated into the clone vector pUCm-T. The segment of F protein's cDNA was subcloned into the expression vector pET-15b and transformed into E coli BL21 (DEB) pLyss. Isopropy-β-D-thiogalactoside (IPTG) was then used to induce expression of the target protein.RESULTS:The cDNA clone of human liver special F protein (1134bp) was successfully produced,with the cDNA sequence being published in Gene-bank:DQ188836. We confirmed the expression of F protein by Western blot with a molecular weight of 43 kDa. The expressed protein accounted for 40% of the total protein extracted.CONCLUSION:F protein expresses cDNA clone in a proKaryotic system,which offers a relatively simple way of producing sufficient quantities of F protein and contributes to understanding the principal biological functions of this protein.

  8. Translational regulation of human p53 gene expression.

    OpenAIRE

    Fu, L.; Minden, M D; Benchimol, S

    1996-01-01

    In blast cells obtained from patients with acute myelogenous leukemia, p53 mRNA was present in all the samples examined while the expression of p53 protein was variable from patient to patient. Mutations in the p53 gene are infrequent in this disease and, hence, variable protein expression in the majority of the samples cannot be accounted for by mutation. In this study, we examined the regulation of p53 gene expression in human leukemic blasts and characterized the p53 transcripts in these c...

  9. Notch receptor expression in human brain arteriovenous malformations.

    Science.gov (United States)

    Hill-Felberg, Sandra; Wu, Hope Hueizhi; Toms, Steven A; Dehdashti, Amir R

    2015-08-01

    The roles of the Notch pathway proteins in normal adult vascular physiology and the pathogenesis of brain arteriovenous malformations are not well-understood. Notch 1 and 4 have been detected in human and mutant mice vascular malformations respectively. Although mutations in the human Notch 3 gene caused a genetic form of vascular stroke and dementia, its role in arteriovenous malformations development has been unknown. In this study, we performed immunohistochemistry screening on tissue microarrays containing eight surgically resected human brain arteriovenous malformations and 10 control surgical epilepsy samples. The tissue microarrays were evaluated for Notch 1-4 expression. We have found that compared to normal brain vascular tissue Notch-3 was dramatically increased in brain arteriovenous malformations. Similarly, Notch 4 labelling was also increased in vascular malformations and was confirmed by western blot analysis. Notch 2 was not detectable in any of the human vessels analysed. Using both immunohistochemistry on microarrays and western blot analysis, we have found that Notch-1 expression was detectable in control vessels, and discovered a significant decrease of Notch 1 expression in vascular malformations. We have demonstrated that Notch 3 and 4, and not Notch 1, were highly increased in human arteriovenous malformations. Our findings suggested that Notch 4, and more importantly, Notch 3, may play a role in the development and pathobiology of human arteriovenous malformations.

  10. Gene expression of manganese superoxide dismutase in human glioma cells

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    Novi S. Hardiany

    2010-02-01

    Full Text Available Aim This study analyze the MnSOD gene expression as endogenous antioxidant in human glioma cells compared with leucocyte cells as control.Methods MnSOD gene expression of 20 glioma patients was analyzed by measuring the relative expression of mRNA and enzyme activity of MnSOD in brain and leucocyte cells. The relative expression of mRNA MnSOD was determined by using quantitative Real Time RT-PCR and the enzyme activity of MnSOD using biochemical kit assay (xantine oxidase inhibition. Statistic analysis for mRNA and enzyme activity of MnSOD was performed using Kruskal Wallis test.Results mRNA of MnSOD in glioma cells of 70% sample was 0.015–0.627 lower, 10% was 1.002-1.059 and 20% was 1.409-6.915 higher than in leucocyte cells. Also the specific activity of MnSOD enzyme in glioma cells of 80% sample showed 0,064-0,506 lower and 20% sample was 1.249-2.718 higher than in leucocyte cells.Conclusion MnSOD gene expression in human glioma cells are significantly lower than its expression in leucocytes cells. (Med J Indones 2010; 19:21-5Keywords : MnSOD, glioma, gene expression

  11. Expression of aquaporin isoforms during human and mouse tooth development.

    Science.gov (United States)

    Felszeghy, S; Módis, L; Németh, P; Nagy, G; Zelles, T; Agre, P; Laurikkala, J; Fejerskov, O; Thesleff, I; Nielsen, S

    2004-04-01

    Previously, we described the development of hyaluronan (HA) deposition in human tooth germ tissues that are consistent with water transport in different stages of tooth development. The aquaporins (AQP) constitute a family of membrane water channels that are expressed in many organs. However, there are no data available about the expression pattern of aquaporin water channels in dental structures. In the present study we have characterised the expression of six different aquaporin isoforms (AQP1-5, AQP-9) in developing human and mouse tooth germs by immunohistochemistry using isoform specific antibodies. In the "bell stage" AQP1 was expressed in endothelial cells of small vessels whereas no other structures of the tooth primordial were labeled. AQP2, AQP3 and AQP9 immunoreactivity was not observed in tooth germs, whereas strong AQP4 and AQP5 expression was observed in dental lamina, inner enamel epithelium, stratum intermedium, stellate reticulum and the outer enamel epithelium. Oral epithelium also exhibited AQP4 and AQP5 immunolabeling. During development of the matrices of the dental hard tissues AQP4 and AQP5 immunostaining was observed in the odontoblasts and their processes, as well as in the secretory ameloblast and their apical processes. Immunolabeling controls were negative. In conclusion, AQP4 and AQP5 are expressed in tooth germ tissues in early development in cells that previously have been shown to express HA and/or CD44, indicating that AQP water channels may play a role for ECM hydration during tooth development.

  12. Expression and regulation of nampt in human islets.

    Directory of Open Access Journals (Sweden)

    Karen Kover

    Full Text Available Nicotinamide phosphoribosyltransferase (Nampt is a rate-limiting enzyme in the mammalian NAD+ biosynthesis of a salvage pathway and exists in 2 known forms, intracellular Nampt (iNampt and a secreted form, extracellular Nampt (eNampt. eNampt can generate an intermediate product, nicotinamide mononucleotide (NMN, which has been reported to support insulin secretion in pancreatic islets. Nampt has been reported to be expressed in the pancreas but islet specific expression has not been adequately defined. The aim of this study was to characterize Nampt expression, secretion and regulation by glucose in human islets. Gene and protein expression of Nampt was assessed in human pancreatic tissue and isolated islets by qRT-PCR and immunofluorescence/confocal imaging respectively. Variable amounts of Nampt mRNA were detected in pancreatic tissue and isolated islets. Immunofluorescence staining for Nampt was found in the exocrine and endocrine tissue of fetal pancreas. However, in adulthood, Nampt expression was localized predominantly in beta cells. Isolated human islets secreted increasing amounts of eNampt in response to high glucose (20 mM in a static glucose-stimulated insulin secretion assay (GSIS. In addition to an increase in eNampt secretion, exposure to 20 mM glucose also increased Nampt mRNA levels but not protein content. The secretion of eNampt was attenuated by the addition of membrane depolarization inhibitors, diazoxide and nifedipine. Islet-secreted eNampt showed enzymatic activity in a reaction with increasing production of NAD+/NADH over time. In summary, we show that Nampt is expressed in both exocrine and endocrine tissue early in life but in adulthood expression is localized to endocrine tissue. Enzymatically active eNampt is secreted by human islets, is regulated by glucose and requires membrane depolarization.

  13. Expression profile of frizzled receptors in human medulloblastomas.

    Science.gov (United States)

    Salsano, Ettore; Paterra, Rosina; Figus, Miriam; Menghi, Francesca; Maderna, Emanuela; Pollo, Bianca; Solero, Carlo Lazzaro; Massimi, Luca; Finocchiaro, Gaetano

    2012-01-01

    Secreted WNT proteins signal through ten receptors of the frizzled (FZD) family. Because of the relevance of the WNT/β-catenin (CTNNB1) signaling pathway in medulloblastomas (MBs), we investigated the expression of all ten members of the FZD gene family (FZD1-10) in 17 human MBs, four MB cell lines and in normal human cerebellum, using real-time PCR. We found that FZD2 transcript was over-expressed in all MBs and MB cell lines. Western blot analysis confirmed the expression of FZD2 at the protein level. Moreover, the levels of FZD2 transcript were found to correlate with those of ASPM transcript, a marker of mitosis essential for mitotic spindle function. Accordingly, ASPM mRNA was expressed at a very low level in the adult, post-mitotic, human cerebellum, at higher levels in fetal cerebellum and at highest levels in MB tissues and cell lines. Unlike FZD2, the other FZDs were overexpressed (e.g., FZD1, FZD3 and FZD8) or underexpressed (e.g., FZD7, FZD9 and FZD10) in a case-restricted manner. Interestingly, we did not find any nuclear immuno-reactivity to CTNNB1 in four MBs over-expressing both FZD2 and other FZD receptors, confirming the lack of nuclear CTNNB1 staining in the presence of increased FZD expression, as in other tumor types. Overall, our results indicate that altered expression of FZD2 might be associated with a proliferative status, thus playing a role in the biology of human MBs, and possibly of cerebellar progenitors from which these malignancies arise.

  14. Gene expression profiles of the developing human retina

    Institute of Scientific and Technical Information of China (English)

    WANG Feng; LI Huiming; LIU Wenwen; XU Ping; HU Gengxi; CHENG Yidong; JIA Libin; HUANG Qian

    2004-01-01

    Retina is a multilayer and highly specialized tissue important in converting light into neural signals. In humans, the critical period for the formation of complex multiplayer structure takes place during embryogenesis between 12 and 28 weeks. The morphologic changes during retinal development in humans have been studied but little is known about the molecular events essential for the formation of the retina. To gain further insights into this process, cDNA microarrays containing 16361 human gene probes were used to measure the gene expression levels in retinas. Of the 16361 genes, 68.7%, 71.4% and 69.7% showed positive hybridization with cDNAs made from 12-16 week fetal, 22-26 week fetal and adult retinas. A total of 814 genes showed a minimum of 3-fold changes between the lowest and highest expression levels among three time points and among them, 106 genes had expression levels with the hybridization intensity above 100 at one or more time points. The clustering analysis suggested that the majority of differentially expressed genes were down-regulated during the retinal development. The differentially expressed genes were further classified according to functions of known genes, and were ranked in decreasing order according to frequency: development, differentiation, signal transduction, protein synthesis and translation, metabolism, DNA binding and transcription, DNA synthesis-repair-recombination, immuno-response, ion channel- transport, cell receptor, cytoskeleton, cell cycle, pro-oncogene, stress and apoptosis related genes. Among these 106 differentially expressed genes, 60 are already present in NEI retina cDNA or EST Databank but the remaining 46 genes are absent and thus identified as "function unknown". To validate gene expression data from the microarray, real-time RT-PCR was performed for 46 "function unknown" genes and 6 known retina specific expression genes, and β-actin was used as internal control. Twenty-seven of these genes showed very similar

  15. Analysis of LINE-1 expression in human pluripotent cells.

    Science.gov (United States)

    Muñoz-Lopez, Martin; Garcia-Cañadas, Marta; Macia, Angela; Morell, Santiago; Garcia-Perez, Jose L

    2012-01-01

    Half of the human genome is composed of repeated DNA, and some types are mobile within our genome (transposons and retrotransposons). Despite their abundance, only a small fraction of them are currently active in our genome (Long Interspersed Element-1 (LINE-1), Alu, and SVA elements). LINE-1 or L1 elements are a family of active non-LTR retrotransposons, the ongoing mobilization of which still impacts our genome. As selfish DNA elements, L1 activity is more prominent in early human development, where new insertions would be transmitted to the progeny. Here, we describe the conventional methods aimed to determine the expression level of LINE-1 elements in pluripotent human cells.

  16. Proteoglycan and collagen expression during human air conducting system development

    Directory of Open Access Journals (Sweden)

    C. Godoy-Guzmán

    2012-06-01

    Full Text Available The lung is formed from a bud that grows and divides in a dichotomous way. A bud is a new growth center which is determined by epithelial-mesenchymal interactions where proteins of the extracellular matrix (ECM might be involved. To understand this protein participation during human lung development, we examined the expression and distribution of proteoglycans in relation to the different types of collagens during the period in which the air conducting system is installed. Using light microscopy and immunohistochemistry we evaluate the expression of collagens (I, III and VI and proteoglycans (decorin, biglycan and lumican between 8 to 10 weeks post fertilization and 11 to 14 weeks of gestational age of human embryo lungs. We show that decorin, lumican and all the collagen types investigated were expressed at the epithelium-mesenchymal interface, forming a sleeve around the bronchiolar ducts. In addition, biglycan was expressed in both the endothelial cells and the smooth muscle of the blood vessels. Thus, the similar distribution pattern of collagen and proteoglycans in the early developmental stages of the human lung may be closely related to the process of dichotomous division of the bronchial tree. This study provides a new insight concerning the participation of collagens and proteoglycans in the epithelial-mesenchymal interface during the period in which the air conducting system is installed in the human fetal lung.

  17. Gene expression in the aging human brain: an overview.

    Science.gov (United States)

    Mohan, Adith; Mather, Karen A; Thalamuthu, Anbupalam; Baune, Bernhard T; Sachdev, Perminder S

    2016-03-01

    The review aims to provide a summary of recent developments in the study of gene expression in the aging human brain. Profiling differentially expressed genes or 'transcripts' in the human brain over the course of normal aging has provided valuable insights into the biological pathways that appear activated or suppressed in late life. Genes mediating neuroinflammation and immune system activation in particular, show significant age-related upregulation creating a state of vulnerability to neurodegenerative and neuropsychiatric disease in the aging brain. Cellular ionic dyshomeostasis and age-related decline in a host of molecular influences on synaptic efficacy may underlie neurocognitive decline in later life. Critically, these investigations have also shed light on the mobilization of protective genetic responses within the aging human brain that help determine health and disease trajectories in older age. There is growing interest in the study of pre and posttranscriptional regulators of gene expression, and the role of noncoding RNAs in particular, as mediators of the phenotypic diversity that characterizes human brain aging. Gene expression studies in healthy brain aging offer an opportunity to unravel the intricately regulated cellular underpinnings of neurocognitive aging as well as disease risk and resiliency in late life. In doing so, new avenues for early intervention in age-related neurodegenerative disease could be investigated with potentially significant implications for the development of disease-modifying therapies.

  18. High expression of NPY receptors in the human testis.

    Science.gov (United States)

    Körner, Meike; Waser, Beatriche; Thalmann, George N; Reubii, Jean Claude

    2011-04-30

    NPY receptors represent novel molecular therapeutic targets in cancer and obesity. However, the extent of NPY receptor expression in normal human tissues is poorly investigated. Based on the role of NPY in reproductive functions, the NPY receptor expression was studied in 25 normal human testes and, additionally, 24 testicular tumors using NPY receptor autoradiography. In the normal testis, Leydig cells strongly expressed NPY receptor subtype Y2, and small arterial blood vessels Y1. Y2 receptors were found to be functional with agonist-stimulated [(35)S]GTPγS binding autoradiography. Full functional integrity of the NPY system was further suggested by the immunohistochemical detection of NPY peptide in nerve fibers directly adjacent to Leydig cells and arteries. Germ cell tumors expressed Y1 and Y2 on tumor cells in 33% and Y1 on intratumoral blood vessels in 50%. Based on its strong NPY receptor expression in Leydig cells and blood vessels, the normal human testis represents a potentially important physiological and pharmalogical NPY target.

  19. RANK and RANK ligand expression in primary human osteosarcoma

    Directory of Open Access Journals (Sweden)

    Daniel Branstetter

    2015-09-01

    Our results demonstrate RANKL expression was observed in the tumor element in 68% of human OS using IHC. However, the staining intensity was relatively low and only 37% (29/79 of samples exhibited≥10% RANKL positive tumor cells. RANK expression was not observed in OS tumor cells. In contrast, RANK expression was clearly observed in other cells within OS samples, including the myeloid osteoclast precursor compartment, osteoclasts and in giant osteoclast cells. The intensity and frequency of RANKL and RANK staining in OS samples were substantially less than that observed in GCTB samples. The observation that RANKL is expressed in OS cells themselves suggests that these tumors may mediate an osteoclastic response, and anti-RANKL therapy may potentially be protective against bone pathologies in OS. However, the absence of RANK expression in primary human OS cells suggests that any autocrine RANKL/RANK signaling in human OS tumor cells is not operative, and anti-RANKL therapy would not directly affect the tumor.

  20. Choline Transporters in Human Lung Adenocarcinoma: Expression and Functional Implications

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    Choline is an essential nutrient for cell survival and proliferation, however, the expression and function of choline transporters have not been well identified in cancer. In this study, we detected the mRNA and protein expression of organic cation transporter OCT3, carnitine/cation transporters OCTN 1 and OCTN2,and choline transporter-like protein CTL1 in human lung adenocarcinoma cell lines A549, H1299 and SPC-A-1.Their expression pattern was further confirmed in 25 human primary adenocarcinoma tissues. The choline uptake in these cell lines was significantly blocked by CTL1 inhibitor, but only partially inhibited by OCT or OCTN inhibitors. The efficacy of these inhibitors on cell proliferation is closely correlated with their abilities to block choline transport. Under the native expression of these transporters, the total choline uptake was notably blocked by specific PI3K/AKT inhibitors. These results describe the expression of choline transporters and their relevant function in cell proliferation of human lung adenocarcinoma, thus providing a potential"choline-starvation" strategy of cancer interference through targeting choline transporters, especially CTL1.

  1. FcgammaRIIb expression on human germinal center B lymphocytes.

    Science.gov (United States)

    Macardle, Peter J; Mardell, Carolyn; Bailey, Sheree; Wheatland, Loretta; Ho, Alice; Jessup, Claire; Roberton, Donal M; Zola, Heddy

    2002-12-01

    IgG antibody can specifically suppress the antibody response to antigen. This has been explained by the hypothesis that signaling through the B cell antigen receptor is negatively modulated by the co-ligation of immunoglobulin with the receptor for IgG, FcgammaRIIb. We hypothesized that inhibitory signaling through FcgammaRIIb would be counter-productive in germinal center cells undergoing selection by affinity maturation, since these cells are thought to receive a survival/proliferative signal by interacting with antigen displayed on follicular dendritic cells. We have identified and characterized a population of B lymphocytes with low/negative FcgammaRIIb expression that are present in human tonsil. Phenotypically these cells correspond to germinal center B cells and comprise both centroblast and centrocyte populations. In examining expression at the molecular level we determined that these B cells do not express detectable mRNA for FcgammaRIIb. We examined several culture conditions to induce expression of FcgammaRIIb on germinal center cells but could not determine conditions that altered expression. We then examined the functional consequence of cross-linking membrane immunoglobulin and the receptor for IgG on human B lymphocytes. Our results cast some doubt on the value of anti-IgG as a model for antigen-antibody complexes in studying human B cell regulation.

  2. Polymorphic cis- and trans-regulation of human gene expression.

    Directory of Open Access Journals (Sweden)

    Vivian G Cheung

    Full Text Available Expression levels of human genes vary extensively among individuals. This variation facilitates analyses of expression levels as quantitative phenotypes in genetic studies where the entire genome can be scanned for regulators without prior knowledge of the regulatory mechanisms, thus enabling the identification of unknown regulatory relationships. Here, we carried out such genetic analyses with a large sample size and identified cis- and trans-acting polymorphic regulators for about 1,000 human genes. We validated the cis-acting regulators by demonstrating differential allelic expression with sequencing of transcriptomes (RNA-Seq and the trans-regulators by gene knockdown, metabolic assays, and chromosome conformation capture analysis. The majority of the regulators act in trans to the target (regulated genes. Most of these trans-regulators were not known to play a role in gene expression regulation. The identification of these regulators enabled the characterization of polymorphic regulation of human gene expression at a resolution that was unattainable in the past.

  3. Investigation of G72 (DAOA expression in the human brain

    Directory of Open Access Journals (Sweden)

    Hirsch Steven

    2008-12-01

    Full Text Available Abstract Background Polymorphisms at the G72/G30 locus on chromosome 13q have been associated with schizophrenia or bipolar disorder in more than ten independent studies. Even though the genetic findings are very robust, the physiological role of the predicted G72 protein has thus far not been resolved. Initial reports suggested G72 as an activator of D-amino acid oxidase (DAO, supporting the glutamate dysfunction hypothesis of schizophrenia. However, these findings have subsequently not been reproduced and reports of endogenous human G72 mRNA and protein expression are extremely limited. In order to better understand the function of this putative schizophrenia susceptibility gene, we attempted to demonstrate G72 mRNA and protein expression in relevant human brain regions. Methods The expression of G72 mRNA was studied by northern blotting and semi-quantitative SYBR-Green and Taqman RT-PCR. Protein expression in human tissue lysates was investigated by western blotting using two custom-made specific anti-G72 peptide antibodies. An in-depth in silico analysis of the G72/G30 locus was performed in order to try and identify motifs or regulatory elements that provide insight to G72 mRNA expression and transcript stability. Results Despite using highly sensitive techniques, we failed to identify significant levels of G72 mRNA in a variety of human tissues (e.g. adult brain, amygdala, caudate nucleus, fetal brain, spinal cord and testis human cell lines or schizophrenia/control post mortem BA10 samples. Furthermore, using western blotting in combination with sensitive detection methods, we were also unable to detect G72 protein in a number of human brain regions (including cerebellum and amygdala, spinal cord or testis. A detailed in silico analysis provides several lines of evidence that support the apparent low or absent expression of G72. Conclusion Our results suggest that native G72 protein is not normally present in the tissues that we analysed

  4. Heterologous expression of human H1 histones in yeast.

    Science.gov (United States)

    Albig, W; Runge, D M; Kratzmeier, M; Doenecke, D

    1998-09-18

    The complete set of seven human H1 histone subtype genes was heterologously expressed in yeast. Since Saccharomyces cerevisiae lacks standard histone H1 we could isolate each recombinantly expressed human H1 subtype in pure form without contamination by endogenous H I histones. For isolation of the H1 histones in this expression system no tagging was needed and the isoforms could be extracted with the authentic primary structure by a single extraction step with 5%(0.74 M) perchloric acid. The isolated H1 histone proteins were used to assign the subtype genes to the corresponding protein spots or peaks after two-dimensional gel electrophoresis and capillary zone electrophoresis, respectively. This allowed us to correlate transcriptional data with protein data, which was barely possible until now.

  5. Distribution of cellular HSV-1 receptor expression in human brain.

    Science.gov (United States)

    Lathe, Richard; Haas, Juergen G

    2016-12-15

    Herpes simplex virus type 1 (HSV-1) is a neurotropic virus linked to a range of acute and chronic neurological disorders affecting distinct regions of the brain. Unusually, HSV-1 entry into cells requires the interaction of viral proteins glycoprotein D (gD) and glycoprotein B (gB) with distinct cellular receptor proteins. Several different gD and gB receptors have been identified, including TNFRSF14/HVEM and PVRL1/nectin 1 as gD receptors and PILRA, MAG, and MYH9 as gB receptors. We investigated the expression of these receptor molecules in different areas of the adult and developing human brain using online transcriptome databases. Whereas all HSV-1 receptors showed distinct expression patterns in different brain areas, the Allan Brain Atlas (ABA) reported increased expression of both gD and gB receptors in the hippocampus. Specifically, for PVRL1, TNFRFS14, and MYH9, the differential z scores for hippocampal expression, a measure of relative levels of increased expression, rose to 2.9, 2.9, and 2.5, respectively, comparable to the z score for the archetypical hippocampus-enriched mineralocorticoid receptor (NR3C2, z = 3.1). These data were confirmed at the Human Brain Transcriptome (HBT) database, but HBT data indicate that MAG expression is also enriched in hippocampus. The HBT database allowed the developmental pattern of expression to be investigated; we report that all HSV1 receptors markedly increase in expression levels between gestation and the postnatal/adult periods. These results suggest that differential receptor expression levels of several HSV-1 gD and gB receptors in the adult hippocampus are likely to underlie the susceptibility of this brain region to HSV-1 infection.

  6. Trefoil factor-3 expression in human colon cancer liver metastasis.

    Science.gov (United States)

    Babyatsky, Mark; Lin, Jing; Yio, Xianyang; Chen, Anli; Zhang, Jie-yu; Zheng, Yan; Twyman, Christina; Bao, Xiuliang; Schwartz, Myron; Thung, Swan; Lawrence Werther, J; Itzkowitz, Steven

    2009-01-01

    Deaths from colorectal cancer are often due to liver metastasis. Trefoil factor-3 (TFF3) is expressed by normal intestinal epithelial cells and its expression is maintained throughout the colon adenoma-carcinoma sequence. Our previous work demonstrated a correlation between TFF3 expression and metastatic potential in an animal model of colon cancer. The aim of this study was to determine whether TFF3 is expressed in human colon cancer liver metastasis (CCLM) and whether inhibiting TFF3 expression in colon cancer cells would alter their invasive potential in vitro. Human CCLMs were analyzed at the mRNA and protein level for TFF3 expression. Two highly metastatic rat colon cancer cell lines that either natively express TFF3 (LN cells) or were transfected with TFF3 (LPCRI-2 cells), were treated with two rat TFF3 siRNA constructs (si78 and si365), and analyzed in an in vitro invasion assay. At the mRNA and protein level, TFF3 was expressed in 17/17 (100%) CCLMs and 10/11 (91%) primary colon cancers, but not in normal liver tissue. By real time PCR, TFF3 expression was markedly inhibited by both siRNA constructs in LN and LPCRI-2 cells. The si365 and si78 constructs inhibited invasion by 44% and 53%, respectively, in LN cells, and by 74% and 50%, respectively, in LPCRI-2 cells. These results provide further evidence that TFF3 contributes to the malignant behavior of colon cancer cells. These observations may have relevance for designing new diagnostic and treatment approaches to colorectal cancer.

  7. [Generation of transgenic mice expressing human lysozyme in mammary gland].

    Science.gov (United States)

    Yan, Hua; Li, Guo-cai; Sun, Huai-chang

    2005-10-01

    To evaluate the feasibility of generating animal mammary gland bioreactors expressing human lysozyme (hLYZ). The recombinant vector p205C3-hLYZ, as a result of connecting the hLYZ cDNA with the mammry gland expression vector p205C3, was used to generate transfer genic mice by microinjection. A total of 136 F0 mice were obtained, of which 7 (2 females and 5 males) and 4 (1 females and 3 males) were found to contain the transfer-gene by PCR and Southern blotting respectively. The results of Western blotting indicated that the expressed protein had the same molecular weight as that of normal hLYZ. From the F1 generation on, the mice mated only with their brothers or sisters and a colony of F7 transgenic mice was obtained. Among the offspring, the female transgenic mice maintained and expressed the transfer-gene stably with an expression level as high as 750 mg/L. The expressed protein had strong tissue specificity, and in addition to the mammary glands, some degree of ectropic expression in the spleens and intestines of the transgenic mice was confirmed by dot blotting assay. These data indicate that the mice mammary gland bioreactors expressing hLYZ have been successfully generated.

  8. Over-expression of cyclooxygenase-2 in human prostate adenocarcinoma.

    Science.gov (United States)

    Gupta, S; Srivastava, M; Ahmad, N; Bostwick, D G; Mukhtar, H

    2000-01-01

    Aberrant or increased expression of cyclooxygenase (COX)-2 has been implicated in the pathogenesis of many diseases including carcinogenesis. COX-2 has been shown to be over-expressed in some human cancers. Employing semi-quantitative reverse transcription-PCR, immunoblotting, and immunohistochemistry we assessed COX-2 expression in samples of pair-matched benign and cancer tissue obtained from the same prostate cancer patient. Mean levels of COX-2 mRNA were 3.4-fold higher in prostate cancer tissue (n = 12) compared with the paired benign tissue. The immunoblot analysis demonstrated that as compared to benign tissue COX-2 protein was over-expressed in 10 of 12 samples examined. Immunohistochemical analysis also verified COX-2 over-expression in cancer than in benign tissue. To our knowledge, this is the first in vivo study showing an over-expression of COX-2 in prostate cancer. These data suggest that COX-2 inhibitors may be useful for prevention or therapy of prostate cancer in humans.

  9. TP53 mutations, expression and interaction networks in human cancers.

    Science.gov (United States)

    Wang, Xiaosheng; Sun, Qingrong

    2017-01-03

    Although the associations of p53 dysfunction, p53 interaction networks and oncogenesis have been widely explored, a systematic analysis of TP53 mutations and its related interaction networks in various types of human cancers is lacking. Our study explored the associations of TP53 mutations, gene expression, clinical outcomes, and TP53 interaction networks across 33 cancer types using data from The Cancer Genome Atlas (TCGA). We show that TP53 is the most frequently mutated gene in a number of cancers, and its mutations appear to be early events in cancer initiation. We identified genes potentially repressed by p53, and genes whose expression correlates significantly with TP53 expression. These gene products may be especially important nodes in p53 interaction networks in human cancers. This study shows that while TP53-truncating mutations often result in decreased TP53 expression, other non-truncating TP53 mutations result in increased TP53 expression in some cancers. Survival analyses in a number of cancers show that patients with TP53 mutations are more likely to have worse prognoses than TP53-wildtype patients, and that elevated TP53 expression often leads to poor clinical outcomes. We identified a set of candidate synthetic lethal (SL) genes for TP53, and validated some of these SL interactions using data from the Cancer Cell Line Project. These predicted SL genes are promising candidates for experimental validation and the development of personalized therapeutics for patients with TP53-mutated cancers.

  10. Expression of human acidic fibroblast growth factor in Pichia pastoris

    Institute of Scientific and Technical Information of China (English)

    YU Ying; CAI Shaoxi; Harald G. WERIRICH; XIA Yuxian

    2003-01-01

    Pichia pastoris expression system is similar to that of the mammal cell in modification of expressed protein, including refolding and glycosylation. A human aFGF gene was cloned into the intracellular expression vector pPIC9K. The Pichia pastoriS KM71 strain was transformed with the recombined expression plasmid. Transgenic expression was observed after screening the transformants with G418. The expression and secretion of recombinant human aFGF (rhaFGF) into the culture medium were testified by ELISA assay. The yield peaked after two days of induction and was approximately 10 mg.L-1 in shake-flask fermentation medium. The recombinant proteins were purified by the combination of heparin-Sepharose affinity chromatography and gel filtration chromatography. Two proteins with relative molecular masses (Mr) of 17 000 and 35 000 were purified as a single band in SDS-PAGE, whose biological activities were determined by MTT assay. It is found that the protein with Mr of 1 7 000 is nonglycosylated haFGF, and that with Mr of 35 000 is glycosylated haFGF; and the latter has a lower biological activity than the former.

  11. On Expression Patterns and Developmental Origin of Human Brain Regions.

    Science.gov (United States)

    Kirsch, Lior; Chechik, Gal

    2016-08-01

    Anatomical substructures of the human brain have characteristic cell-types, connectivity and local circuitry, which are reflected in area-specific transcriptome signatures, but the principles governing area-specific transcription and their relation to brain development are still being studied. In adult rodents, areal transcriptome patterns agree with the embryonic origin of brain regions, but the processes and genes that preserve an embryonic signature in regional expression profiles were not quantified. Furthermore, it is not clear how embryonic-origin signatures of adult-brain expression interplay with changes in expression patterns during development. Here we first quantify which genes have regional expression-patterns related to the developmental origin of brain regions, using genome-wide mRNA expression from post-mortem adult human brains. We find that almost all human genes (92%) exhibit an expression pattern that agrees with developmental brain-region ontology, but that this agreement changes at multiple phases during development. Agreement is particularly strong in neuron-specific genes, but also in genes that are not spatially correlated with neuron-specific or glia-specific markers. Surprisingly, agreement is also stronger in early-evolved genes. We further find that pairs of similar genes having high agreement to developmental region ontology tend to be more strongly correlated or anti-correlated, and that the strength of spatial correlation changes more strongly in gene pairs with stronger embryonic signatures. These results suggest that transcription regulation of most genes in the adult human brain is spatially tuned in a way that changes through life, but in agreement with development-determined brain regions.

  12. INSL-3 is expressed in human hyperplastic and neoplastic thyrocytes.

    Science.gov (United States)

    Hombach-Klonisch, Sabine; Hoang-Vu, Cuong; Kehlen, Astrid; Hinze, Raoul; Holzhausen, Hans-Jürgen; Weber, Ekkehard; Fischer, Bernd; Dralle, Henning; Klonisch, Thomas

    2003-05-01

    The insulin-like hormone INSL-3, also named relaxin-like factor (RLF) or Leydig-derived insulin-like peptide (LEY-IL), is expressed in various reproductive tissues and is regarded a marker of differentiation in human testicular Leydig cells. Recently, we have identified differential expression of human INSL-3 in neoplastic Leydig cells and mammary epithelial cells suggesting an involvement of INSL-3 in tumor biology. Here we have investigated the expression of INSL-3 in human thyroid carcinoma cell lines and in the human thyroid gland which has been shown to express transcripts for the G protein coupled INSL-3 receptor LGR8. When we determined the expression of INSL-3 in eight human thyroid carcinoma cell lines, a novel INSL-3 splice variant containing a 95 bp out-of-frame insertion at the beginning of exon II of the INSL-3 gene was discovered. Treatment of the human anaplastic thyroid carcinoma cell line 8505C with diethylstilbestrol (DES) caused a significant dose-dependent transcriptional down-regulation of INSL-3 and a marked up-regulation of LGR8. Employing in situ hybridization to detect INSL-3 transcripts and specific rabbit antisera against the INSL-3 proteins, both INSL-3 isoforms were detected in patients with Graves' disease (n=10), follicular carcinomas (FTC; n=12), papillary carcinomas (PTC; n=9) and undifferentiated anaplastic carcinomas (UTC; n=15). By contrast, thyrocytes of all 15 benign goiter tissues studied were devoid of both INSL-3 isoforms, mRNA and protein. Our data indicate that INSL-3 hormone is up-regulated in hyperplastic and neoplastic human thyrocytes suggesting that the INSL-3 isoforms may serve as additional markers for hyperplastic and neoplastic human thyrocytes. In the anaplastic thyroid carcinoma cell line 8505C, the regulation of both INSL-3 and LGR8 by estrogen may be the first indication of a novel hormonally responsive, auto-/paracrine INSL-3 LGR8 ligand receptor system active in human thyroid carcinoma cells.

  13. Quality of human milk expressed in a human milk bank and at home.

    Science.gov (United States)

    Borges, Mayla S; Oliveira, Angela M de M; Hattori, Wallisen T; Abdallah, Vânia O S

    2017-08-30

    To evaluate the quality of the human milk expressed at home and at a human milk bank. This a retrospective, analytical, and observational study, performed by assessing titratable acidity records and the microbiological culture of 100 human milk samples expressed at home and at a human milk bank, in 2014. For the statistical analysis, generalized estimating equations (GEE) and the chi-squared test were used. When comparing the two sample groups, no significant difference was found, with 98% and 94% of the samples being approved among those collected at the milk bank and at home, respectively. No main interaction effect between local and titratable acidity records (p=0.285) was observed, and there was no statistically significant difference between the expected and observed values for the association between the collection place and the microbiological culture results (p=0.307). The quality of human milk expressed at home and at the milk bank are in agreement with the recommended standards, confirming that the expression of human milk at home is as safe as expression at the human milk bank, provided that the established hygiene, conservation, storage, and transport standards are followed. Copyright © 2017 Sociedade Brasileira de Pediatria. Published by Elsevier Editora Ltda. All rights reserved.

  14. LIN28A expression reduces sickling of cultured human erythrocytes.

    Science.gov (United States)

    de Vasconcellos, Jaira F; Fasano, Ross M; Lee, Y Terry; Kaushal, Megha; Byrnes, Colleen; Meier, Emily R; Anderson, Molly; Rabel, Antoinette; Braylan, Raul; Stroncek, David F; Miller, Jeffery L

    2014-01-01

    Induction of fetal hemoglobin (HbF) has therapeutic importance for patients with sickle cell disease (SCD) and the beta-thalassemias. It was recently reported that increased expression of LIN28 proteins or decreased expression of its target let-7 miRNAs enhances HbF levels in cultured primary human erythroblasts from adult healthy donors. Here LIN28A effects were studied further using erythrocytes cultured from peripheral blood progenitor cells of pediatric subjects with SCD. Transgenic expression of LIN28A was accomplished by lentiviral transduction in CD34(+) sickle cells cultivated ex vivo in serum-free medium. LIN28A over-expression (LIN28A-OE) increased HbF, reduced beta (sickle)-globin, and strongly suppressed all members of the let-7 family of miRNAs. LIN28A-OE did not affect erythroblast differentiation or prevent enucleation, but it significantly reduced or ameliorated the sickling morphologies of the enucleated erythrocytes.

  15. KiSS-1 expression in human breast cancer.

    Science.gov (United States)

    Martin, Tracey A; Watkins, Gareth; Jiang, Wen G

    2005-01-01

    The KiSS-1 gene encodes a 145 amino acid residue peptide that is further processed to a final peptide, metastin, a ligand to a G-coupled orphan receptor (OT7T175/AXOR12). KiSS-1 has been identified as a putative human metastasis suppressor gene in melanomas and in breast cancer cell lines. This study aimed to determine the expression and distribution of KiSS-1 and its receptor in human breast cancer tissues and to identify a possible link between expression levels and patient prognosis. Frozen sections from breast cancer primary tumours (matched tumour 124 and background 33) were immuno-stained with KiSS-1 antibody. RNA was reverse transcribed and analyzed by Q-PCR (standardized using beta-actin, and normalized with cytokeratin-19 levels). Levels of expression of KiSS-1 were higher in tumour compared to background tissues (3,124+/-1,262 vs 2,397+/-1,181) and significantly increased in node positive tumours compared to node negative (3,637+/-1,719 vs 2,653+/-1,994, P = 0.02). KiSS-1 expression was also increased with increasing grade and TNM status. There were no such trends with the KiSS-1 receptor. Expression of KiSS-1 was higher in patients who had died from breast cancer than those who had remained healthy (4,631+/-3,024 vs 2,280+/-1,403) whereas expression of the receptor was reduced (480+/-162 vs 195+/-134). Immunohistochemical staining showed increased expression of KiSS-1 in tumour sections. Insertion of the KiSS-1 gene into the human breast cancer cell line MDA-MB-231, resulted in cells that were significantly more motile and invasive in behaviour, with reduced adhesion to matrix, using respective assays. In conclusion, KiSS-1 expression is increased in human breast cancer, particularly in patients with aggressive tumours and with mortality. Over-expression of KiSS-1 in breast cancer cells result in more aggressive phenotype. Together, it suggests that KiSS-1 plays a role beyond the initial metastasis repressor in this cancer type.

  16. Neural decoding of expressive human movement from scalp electroencephalography (EEG

    Directory of Open Access Journals (Sweden)

    Zachery Ryan Hernandez

    2014-04-01

    Full Text Available Although efforts to characterize human movement through EEG have revealed neural activities unique to limb control that can be used to infer movement kinematics, it is still unknown the extent to which EEG can be used to discern the expressive qualities that influence such movements. In this study we used EEG and inertial sensors to record brain activity and movement of five skilled and certified Laban Movement Analysis (LMA dancers. Each dancer performed whole body functional movements of three Action types: movements devoid of expressive qualities ('Neutral', non-expressive movements while thinking about specific expressive qualities ('Think’, and enacted expressive movements ('Do'. The expressive movement qualities that were used in the 'Think' and 'Do' actions consisted of a sequence of eight Laban Efforts as defined by LMA - a notation system and language for describing, visualizing, interpreting and documenting all varieties of human movement. We used delta band (0.2 – 4 Hz EEG as input to a machine learning algorithm that computed locality-preserving Fisher’s discriminant analysis (LFDA for dimensionality reduction followed by Gaussian mixture models (GMMs to decode the type of Action. We also trained our LFDA-GMM models to classify all the possible combinations of Action Type and Laban Effort (giving a total of 17 classes. Classification accuracy rates were 59.4 ± 0.6% for Action Type and 88.2 ± 0.7% for Laban Effort Type. Ancillary analyses of the potential relations between the EEG and movement kinematics of the dancer's body, indicated that motion-related artifacts did not significantly influence our classification results. In summary, this research demonstrates that EEG has valuable information about the expressive qualities of movement. These results may have applications for advancing the understanding of the neural basis of expressive movements and for the development of neuroprosthetics to restore movements.

  17. Neural decoding of expressive human movement from scalp electroencephalography (EEG)

    Science.gov (United States)

    Cruz-Garza, Jesus G.; Hernandez, Zachery R.; Nepaul, Sargoon; Bradley, Karen K.; Contreras-Vidal, Jose L.

    2014-01-01

    Although efforts to characterize human movement through electroencephalography (EEG) have revealed neural activities unique to limb control that can be used to infer movement kinematics, it is still unknown the extent to which EEG can be used to discern the expressive qualities that influence such movements. In this study we used EEG and inertial sensors to record brain activity and movement of five skilled and certified Laban Movement Analysis (LMA) dancers. Each dancer performed whole body movements of three Action types: movements devoid of expressive qualities (“Neutral”), non-expressive movements while thinking about specific expressive qualities (“Think”), and enacted expressive movements (“Do”). The expressive movement qualities that were used in the “Think” and “Do” actions consisted of a sequence of eight Laban Effort qualities as defined by LMA—a notation system and language for describing, visualizing, interpreting and documenting all varieties of human movement. We used delta band (0.2–4 Hz) EEG as input to a machine learning algorithm that computed locality-preserving Fisher's discriminant analysis (LFDA) for dimensionality reduction followed by Gaussian mixture models (GMMs) to decode the type of Action. We also trained our LFDA-GMM models to classify all the possible combinations of Action Type and Laban Effort quality (giving a total of 17 classes). Classification accuracy rates were 59.4 ± 0.6% for Action Type and 88.2 ± 0.7% for Laban Effort quality Type. Ancillary analyses of the potential relations between the EEG and movement kinematics of the dancer's body, indicated that motion-related artifacts did not significantly influence our classification results. In summary, this research demonstrates that EEG has valuable information about the expressive qualities of movement. These results may have applications for advancing the understanding of the neural basis of expressive movements and for the development of

  18. D2-40/podoplanin expression in the human placenta.

    Science.gov (United States)

    Wang, Y; Sun, J; Gu, Y; Zhao, S; Groome, L J; Alexander, J S

    2011-01-01

    Placental tissue expresses many lymphatic markers. The current study was undertaken to examine if D2-40/podoplanin, a lymphatic endothelial marker, was expressed in the human placenta, and how it is altered developmentally and pathologically. We examined D2-40/podoplanin and VEGFR-3 expressions in placentas from normotensive pregnancies at different gestational ages and in placentas from women with clinically defined preeclampsia. D2-40 expression in systemic lymphatic vessel endothelium served as a positive control. Protein expression for D2-40, VEGFR-3, and β-actin was determined by Western blot in placentas from normotensive (n = 6) and preeclamptic (n = 5) pregnancies. Our results show that D2-40/podoplanin was strongly expressed in the placenta, mainly as a network plexus pattern in the villous stroma throughout gestation. CD31 was limited to villous core fetal vessel endothelium and VEGFR-3 was found in both villous core fetal vessel endothelium and trophoblasts. D2-40/podoplanin expression was significantly decreased, and VEGFR-3 significantly increased in preeclamptic placental tissues compared to normotensive placental controls. Placental villous stroma is a reticular-like structure, and the localization of D2-40 to the stroma suggests that a lymphatic-like conductive network may exist in the human placenta. D2-40/podoplanin is an O-linked sialoglycoprotein. Although little is known regarding biological functions of sialylated glycoproteins within the placenta, placental D2-40/podoplanin may support fetal vessel angiogenesis during placenta development and reduced D2-40/podoplanin expression in preeclamptic placenta may contribute to altered interstitial fluid homeostasis and impaired angiogenesis in this pregnancy disorder.

  19. Neural decoding of expressive human movement from scalp electroencephalography (EEG).

    Science.gov (United States)

    Cruz-Garza, Jesus G; Hernandez, Zachery R; Nepaul, Sargoon; Bradley, Karen K; Contreras-Vidal, Jose L

    2014-01-01

    Although efforts to characterize human movement through electroencephalography (EEG) have revealed neural activities unique to limb control that can be used to infer movement kinematics, it is still unknown the extent to which EEG can be used to discern the expressive qualities that influence such movements. In this study we used EEG and inertial sensors to record brain activity and movement of five skilled and certified Laban Movement Analysis (LMA) dancers. Each dancer performed whole body movements of three Action types: movements devoid of expressive qualities ("Neutral"), non-expressive movements while thinking about specific expressive qualities ("Think"), and enacted expressive movements ("Do"). The expressive movement qualities that were used in the "Think" and "Do" actions consisted of a sequence of eight Laban Effort qualities as defined by LMA-a notation system and language for describing, visualizing, interpreting and documenting all varieties of human movement. We used delta band (0.2-4 Hz) EEG as input to a machine learning algorithm that computed locality-preserving Fisher's discriminant analysis (LFDA) for dimensionality reduction followed by Gaussian mixture models (GMMs) to decode the type of Action. We also trained our LFDA-GMM models to classify all the possible combinations of Action Type and Laban Effort quality (giving a total of 17 classes). Classification accuracy rates were 59.4 ± 0.6% for Action Type and 88.2 ± 0.7% for Laban Effort quality Type. Ancillary analyses of the potential relations between the EEG and movement kinematics of the dancer's body, indicated that motion-related artifacts did not significantly influence our classification results. In summary, this research demonstrates that EEG has valuable information about the expressive qualities of movement. These results may have applications for advancing the understanding of the neural basis of expressive movements and for the development of neuroprosthetics to restore

  20. Proteins differentially expressed in human beta-cells-enriched pancreatic islet cultures and human insulinomas

    DEFF Research Database (Denmark)

    Terra, Letícia F; Teixeira, Priscila C; Wailemann, Rosangela A M

    2013-01-01

    In view of the great demand for human beta-cells for physiological and medical studies, we generated cell lines derived from human insulinomas which secrete insulin, C-peptide and express neuroendocrine and islet markers. In this study, we set out to characterize their proteomes, comparing them t...

  1. Altered gene expression in human placenta after suspected preterm labour.

    Science.gov (United States)

    Oros, D; Strunk, M; Breton, P; Paules, C; Benito, R; Moreno, E; Garcés, M; Godino, J; Schoorlemmer, J

    2017-07-01

    Suspected preterm labour occurs in around 9% of pregnancies. However, almost two-thirds of women admitted for threatened preterm labour ultimately deliver at term and are considered risk-free for fetal development. We examined placental and umbilical cord blood samples from preterm or term deliveries after threatened preterm labour as well as term deliveries without threatened preterm labour. We quantitatively analysed the mRNA expression of inflammatory markers (IL6, IFNγ, and TNFα) and modulators of angiogenesis (FGF2, PGF, VEGFA, VEGFB, and VEGFR1). A total of 132 deliveries were analysed. Preterm delivery and term delivery after suspected preterm labour groups showed similar increases in TNFα expression compared with the term delivery control group in umbilical cord blood samples. Placental samples from preterm and term deliveries after suspected preterm labour exhibited significantly increased expression of TNFα and IL6 and decreased expression of IFNγ. Suspected preterm labour was also associated with altered expression of angiogenic factors, although not all differences reached statistical significance. We found gene expression patterns indicative of inflammation in human placentas after suspected preterm labour regardless of whether the deliveries occurred preterm or at term. Similarly, a trend towards altered expression of angiogeneic factors was not limited to preterm birth. These findings suggest that the biological mechanisms underlying threatened preterm labour affect pregnancies independently of gestational age at birth. Copyright © 2017 Elsevier Ltd. All rights reserved.

  2. NFATc1 regulation of TRAIL expression in human intestinal cells.

    Directory of Open Access Journals (Sweden)

    Qingding Wang

    Full Text Available TNF-related apoptosis-inducing ligand (TRAIL; Apo2 has been shown to promote intestinal cell differentiation. Nuclear factor of activated T cells (NFAT participates in the regulation of a variety of cellular processes, including differentiation. Here, we examined the role of NFAT in the regulation of TRAIL in human intestinal cells. Treatment with a combination of phorbol 12-myristate 13-acetate (PMA plus the calcium ionophore A23187 (Io increased NFAT activation and TRAIL expression; pretreatment with the calcineurin inhibitor cyclosporine A (CsA, an antagonist of NFAT signaling, diminished NFAT activation and TRAIL induction. In addition, knockdown of NFATc1, NFATc2, NFATc3, and NFATc4 blocked PMA/Io increased TRAIL protein expression. Expression of NFATc1 activated TRAIL promoter activity and increased TRAIL mRNA and protein expression. Deletion of NFAT binding sites from the TRAIL promoter did not significantly abrogate NFATc1-increased TRAIL promoter activity, suggesting an indirect regulation of TRAIL expression by NFAT activation. Knockdown of NFATc1 increased Sp1 transcription factor binding to the TRAIL promoter and, importantly, inhibition of Sp1, by chemical inhibition or RNA interference, increased TRAIL expression. These studies identify a novel mechanism for TRAIL regulation by which activation of NFATc1 increases TRAIL expression through negative regulation of Sp1 binding to the TRAIL promoter.

  3. BMP-7 PROTEIN EXPRESSION IS DOWNREGULATED IN HUMAN DIABETIC NEPHROPATHY.

    Science.gov (United States)

    Ivanac-Janković, Renata; Ćorić, Marijana; Furić-Čunko, Vesna; Lovičić, Vesna; Bašić-Jukić, Nikolina; Kes, Petar

    2015-06-01

    Bone morphogenetic protein-7 (BMP-7) is expressed in all parts of the normal kidney parenchyma, being highest in the epithelium of proximal tubules. It protects kidney against acute and chronic injury, inflammation and fibrosis. Diabetic nephropathy is the leading cause of chronic kidney disease, and is characterized by decreased expression of BMP-7. The aim of our study was to analyze whether the expression of BMP-7 is significantly changed in advanced stages of human diabetic nephropathy. Immunohistochemical analysis of the expression of BMP-7 was performed on archival material of 30 patients that underwent renal biopsy and had confirmed diagnosis of diabetic nephropathy. Results showed that BMP-7 was differently expressed in the cytoplasm of epithelial cells of proximal tubules and podocytes among all stages of diabetic nephropathy. At early stages of diabetic nephropathy, BMP-7 was strongly positive in proximal tubules and podocytes, while low expression was recorded in the majority of samples at advanced stages. In conclusion, increased expression of BMP-7 at initial stages of diabetic nephropathy with subsequent decrease at advanced stage highlights the role of BMP-7 in the protection of kidney structure and function. Further investigations should be focused on disturbances of BMP-7 receptors and signaling pathways in patients with diabetic nephropathy.

  4. Characterisation of the expression of NMDA receptors in human astrocytes.

    Directory of Open Access Journals (Sweden)

    Ming-Chak Lee

    Full Text Available Astrocytes have long been perceived only as structural and supporting cells within the central nervous system (CNS. However, the discovery that these glial cells may potentially express receptors capable of responding to endogenous neurotransmitters has resulted in the need to reassess astrocytic physiology. The aim of the current study was to characterise the expression of NMDA receptors (NMDARs in primary human astrocytes, and investigate their response to physiological and excitotoxic concentrations of the known endogenous NMDAR agonists, glutamate and quinolinic acid (QUIN. Primary cultures of human astrocytes were used to examine expression of these receptors at the mRNA level using RT-PCR and qPCR, and at the protein level using immunocytochemistry. The functionality role of the receptors was assessed using intracellular calcium influx experiments and measuring extracellular lactate dehydrogenase (LDH activity in primary cultures of human astrocytes treated with glutamate and QUIN. We found that all seven currently known NMDAR subunits (NR1, NR2A, NR2B, NR2C, NR2D, NR3A and NR3B are expressed in astrocytes, but at different levels. Calcium influx studies revealed that both glutamate and QUIN could activate astrocytic NMDARs, which stimulates Ca2+ influx into the cell and can result in dysfunction and death of astrocytes. Our data also show that the NMDAR ion channel blockers, MK801, and memantine can attenuate glutamate and QUIN mediated cell excitotoxicity. This suggests that the mechanism of glutamate and QUIN gliotoxicity is at least partially mediated by excessive stimulation of NMDARs. The present study is the first to provide definitive evidence for the existence of functional NMDAR expression in human primary astrocytes. This discovery has significant implications for redefining the cellular interaction between glia and neurons in both physiological processes and pathological conditions.

  5. Hierarchical Recognition Scheme for Human Facial Expression Recognition Systems

    Directory of Open Access Journals (Sweden)

    Muhammad Hameed Siddiqi

    2013-12-01

    Full Text Available Over the last decade, human facial expressions recognition (FER has emerged as an important research area. Several factors make FER a challenging research problem. These include varying light conditions in training and test images; need for automatic and accurate face detection before feature extraction; and high similarity among different expressions that makes it difficult to distinguish these expressions with a high accuracy. This work implements a hierarchical linear discriminant analysis-based facial expressions recognition (HL-FER system to tackle these problems. Unlike the previous systems, the HL-FER uses a pre-processing step to eliminate light effects, incorporates a new automatic face detection scheme, employs methods to extract both global and local features, and utilizes a HL-FER to overcome the problem of high similarity among different expressions. Unlike most of the previous works that were evaluated using a single dataset, the performance of the HL-FER is assessed using three publicly available datasets under three different experimental settings: n-fold cross validation based on subjects for each dataset separately; n-fold cross validation rule based on datasets; and, finally, a last set of experiments to assess the effectiveness of each module of the HL-FER separately. Weighted average recognition accuracy of 98.7% across three different datasets, using three classifiers, indicates the success of employing the HL-FER for human FER.

  6. Human leucocytes in asthenozoospermic patients: endothelial nitric oxide synthase expression.

    Science.gov (United States)

    Buldreghini, E; Hamada, A; Macrì, M L; Amoroso, S; Boscaro, M; Lenzi, A; Agarwal, A; Balercia, G

    2014-12-01

    In a basic study at the Andrology Unit, Department of Clinical and Molecular Sciences, Polytechnic University of Marche, Ancona, Italy, we evaluated the pattern of mRNA endothelial nitric oxide synthase (eNOS) expression in human blood leucocytes isolated from normozoospermic fertile and asthenozoospermic infertile men to elucidate any pathogenic involvement in sperm cell motility. Forty infertile men with idiopathic asthenozoospermia and 45 normozoospermic fertile donors, age-matched, were included. Semen parameters were evaluated, and expression analysis of mRNA was performed in human leucocytes using reverse transcription polymerase chain reaction. Sperm volume, count, motility and morphology were determined, and eNOS expression and Western blotting analyses were performed. A positive correlation was observed between the concentrations of NO and the percentage of immotile spermatozoa. The mRNA of eNOS was more expressed in peripheral blood leucocytes isolated from asthenozoospermic infertile men versus those of fertile normozoospermic men (7.46 ± 0.38 versus 7.06 ± 0.56, P = 0.0355). A significant up-regulation of eNOS gene in peripheral blood leucocytes was 1.52-fold higher than that of fertile donors. It is concluded that eNOS expression and activity are enhanced in blood leucocytes in men with idiopathic asthenozoospermia.

  7. Expression of the endocannabinoid receptors in human fascial tissue

    Directory of Open Access Journals (Sweden)

    C. Fede

    2016-06-01

    Full Text Available Cannabinoid receptors have been localized in the central and peripheral nervous system as well as on cells of the immune system, but recent studies on animal tissue gave evidence for the presence of cannabinoid receptors in different types of tissues. Their presence was supposed also in myofascial tissue, suggesting that the endocannabinoid system may help resolve myofascial trigger points and relieve symptoms of fibromyalgia. However, until now the expression of CB1 (cannabinoid receptor 1 and CB2 (cannabinoid receptor 2 in fasciae has not yet been established. Small samples of fascia were collected from volunteers patients during orthopedic surgery. For each sample were done a cell isolation, immunohistochemical investigation (CB1 and CB2 antibodies and real time RT-PCR to detect the expression of CB1 and CB2. Both cannabinoid receptors are expressed in human fascia and in human fascial fibroblasts culture cells, although to a lesser extent than the control gene. We can assume that the expression of mRNA and protein of CB1 and CB2 receptors in fascial tissue are concentrated into the fibroblasts. This is the first demonstration that the fibroblasts of the muscular fasciae express CB1 and CB2. The presence of these receptors could help to provide a description of cannabinoid receptors distribution and to better explain the role of fasciae as pain generator and the efficacy of some fascial treatments. Indeed the endocannabinoid receptors of fascial fibroblasts can contribute to modulate the fascial fibrosis and inflammation.

  8. Promoter hypomethylation regulates CD133 expression in human gliomas

    Institute of Scientific and Technical Information of China (English)

    Kouichi Tabu; Ken Sasai; Taichi Kimura; Lei Wang; Eiko Aoyanagi; Shinji Kohsaka; Mishie Tanino; Hiroshi Nishihara; Shinya Tanaka

    2008-01-01

    Brain tumor-initiating cells (BTICs) have been enriched using antibodies against the cell surface protein CD133;however,the biological relevance and the regulatory mechanism of CD133 expression in human gliomas are not yet understood.In this study,we initially demonstrated that CD133 was overexpressed in high-grade human glioblastomas where CD133-positive cells were focally observed as a micro-cluster.In addition,CD133 transcripts with exon 1A,1B,or 1C were predominantly expressed in glioblastomas.To elucidate the mechanism regulating this aberrant expression of CD133,three proximal promoters (P1,P2,and P3) containing a CpG island were isolated.In U251MG and T98Gglioblastoma cells,the P1 region flanking exon 1A exhibited the highest activity among the three promoters,and this activity was significantly inactivated by in vitro methylation.After treatment with the demethylating agent 5-azacytidine and/or the histone deacetylase inhibitor valproic acid,the expression level of CD133 mRNA was significantly restored in glioma cells.Importantly,hypomethylation of CpG sites within the P1,P2,and P3 regions was observed by bisulfite sequencing in human glioblastoma tissues with abundant CD133 mRNA.Taken together,our results indicate that DNA hypomethylation is an important determinant of CD133 expression in glioblastomas,and this epigenetic event may be associated with the development of BTICs expressing CD133.

  9. Expression of retinoic acid receptors in human endometrial carcinoma.

    Science.gov (United States)

    Tanabe, Kojiro; Utsunomiya, Hiroki; Tamura, Mitsutoshi; Niikura, Hitoshi; Takano, Tadao; Yoshinaga, Kohsuke; Nagase, Satoru; Suzuki, Takashi; Ito, Kiyoshi; Matsumoto, Mitsuyo; Hayashi, Shin-ichi; Yaegashi, Nobuo

    2008-02-01

    The retinoids (vitamin A and its biologically active derivatives) are essential for the health and survival of the individual. Several studies have reported a strong rationale for the use of retinoids in cancer treatment and chemoprevention. It has been discovered that expression of retinoic acid receptor (RAR) beta is frequently silenced in epithelial carcinogenesis, which has led to the hypothesis that RAR beta could act as a tumor suppressor. However, the status of RAR beta in human endometrial carcinoma has not been examined. In the present study, we initially studied the effects of retinoic acid on cell proliferation and the expression of RAR alpha, RAR beta, and RAR gamma using AM580 (a RAR-specific agonist) in the Ishikawa endometrial cancer cell line. We also examined the expression of RAR in human eutopic endometrium (30 cases), endometrial hyperplasia (28 cases), and endometrial carcinoma (103 cases) using immunohistochemistry. Finally, we correlated these findings with the clinicopathological parameters. In vitro, cell growth was inhibited and RAR beta and RAR gamma mRNA was significantly induced by AM580, compared with vehicle controls, whereas RAR alpha mRNA was significantly attenuated by AM580, compared with vehicle. RAR beta was detected predominantly in endometrial hyperplasia, compared with endometrial carcinoma. No statistically significant correlation was obtained between the expression of any other RAR subtypes and clinicopathological parameters in human endometrial carcinoma. The results of our study demonstrate that AM580 inhibits cell growth and induces RAR beta mRNA expression in the Ishikawa cell line, and the expression level of RAR beta in endometrial carcinoma is significantly lower than that in endometrial hyperplasia. AM580 might therefore be considered as a potential treatment for endometrial carcinoma.

  10. Bacterial flagellin induces IL-6 expression in human basophils.

    Science.gov (United States)

    Jeon, Jun Ho; Ahn, Ki Bum; Kim, Sun Kyung; Im, Jintaek; Yun, Cheol-Heui; Han, Seung Hyun

    2015-05-01

    Binding of allergen to IgE on basophils positively affects allergic inflammation by releasing inflammatory mediators. Recently, basophils were shown to express pattern-recognition receptors, such as toll-like receptors (TLRs), for recognizing microbe-associated molecular patterns (MAMPs) that are independent of allergen-IgE binding. In this study, we investigated whether MAMP alone can induce IL-6 production in a human basophil cell line, KU812. Stimulation with flagellin in the absence of allergen-IgE association induced IL-6 expression in KU812 cells, while stimulation with lipoteichoic acid, peptidoglycan, or poly I:C did not under the same condition. Flagellin-induced IL-6 expression was also observed in human primary basophils. Flow cytometric analysis showed that KU812 cells expressed flagellin-recognizing TLR5 both on the cell surface and in the cytoplasm while TLR2 and TLR3 were observed only in the cytoplasm. We further demonstrated that although flagellin augmented the phosphorylation of mitogen-activated protein kinases including p38 kinase, ERK, and JNK, flagellin-induced IL-6 production was attenuated by inhibitors for p38 kinase and ERK, but not by JNK inhibitors. In addition, flagellin enhanced phosphorylation of signaling molecules including CREB, PKCδ, and AKT. The inhibitors for PKA and PKC also showed inhibitory effects. Interestingly, flagellin-induced IL-6 production was further enhanced by pretreatment with inhibitors for PI3K, implying that PI3K negatively affects the flagellin-induced IL-6 production. Furthermore, DNA binding activities of NF-κB, AP-1, and CREB, which play pivotal roles in the induction of IL-6 gene expression, were increased by flagellin. These results suggest that flagellin alone is sufficient to induce IL-6 gene expression via TLR5 signaling pathways in human basophils.

  11. Expression of Calbindin-d28k in Human Endometrium

    Institute of Scientific and Technical Information of China (English)

    Duo XU; Wei-jie ZHU

    2007-01-01

    Objective To investigate the expression of Calbindin-d28k (CaBP-d28k) in human endometrium.Methods Thirty-three samples of human normal endometrial tissues were divided into 6 groups: early proliferative stage (n =6), mid proliferative stage (n =5), late proliferative stage (n=5), early secretory stage (n=7), mid secretory stage (n=5) and late secretory stage (n=5). The expression and change of CaBP-d28k protein and gene were determined by immunohistochemistry and reverse transcription polymerase chain reaction methods.Results In endometrial samples, the expression of CaBP-d28k protein was mainly observed in the cytoplasm of luminal and glandular epithelium. In the menstrual cycle, the level of CaBP-d28k protein in the epithelium was the lowest during the early and mid proliferative stages, and was the highest during the mid secretory stage, then decreased in the late secretory stage (P<0.05). In the stroma, the expressed type of CaBP-d28k protein was the same as in the epithelium, but was lower than that in the epithelium(P<0. 05). The CaBP-d28k mRNA was at the lowest level in the early proliferative stage(P<0. 05), and significantly increased in the late proliferative, and early,mid secretory stages (P<0. 05).Conclusion Both CaBP-d28k protein and gene were expressed in human endometrium,and their expression had cyclic changes.

  12. Effect of Human Cytomegalovirus Infection on Nerve Growth Factor Expression in Human Glioma U251 Cells

    Institute of Scientific and Technical Information of China (English)

    HAI-TAO WANG; BIN WANG; ZHI-JUN LIU; ZHI-QIANG BAI; LING LI; HAI-YAN LIU; DONG-MENG QIAN; ZHI-YONG YAN; XU-XIA SONG

    2009-01-01

    Objectives To explore the change of endogenic nerve growth factor (NGF) expression in human glioma cells infected with human cytomegalovirus (HCMV). Methods U251 cells were cultured in RPMI 1640 culture medium and infected with HCMV AD169 strain in vitro to establish a cell model of viral infection. Morphologic changes of U251 cells were observed under inverted microscope before and after infection with HCMV. Expression of NGF gene and protein of cells was detected by RT-PCR and Western blotting before and after infection with HCMV. Results The cytopathic effects of HCMV-infected cells appeared on day 5 after infection. However, differential NGF expression was evident on day 7. NGF expression was decreased significantly in U251 cells on day 7 after infection in comparison with control group (P<0.05). Conclusion HCMV can down-regulate endogenous NGF levels in human glioma cell line U251.

  13. Sequence and expression analysis of gaps in human chromosome 20

    DEFF Research Database (Denmark)

    Minocherhomji, Sheroy; Seemann, Stefan; Mang, Yuan;

    2012-01-01

    The finished human genome-assemblies comprise several hundred un-sequenced euchromatic gaps, which may be rich in long polypurine/polypyrimidine stretches. Human chromosome 20 (chr 20) currently has three unfinished gaps remaining on its q-arm. All three gaps are within gene-dense regions and....../or overlap disease-associated loci, including the DLGAP4 locus. In this study, we sequenced ~99% of all three unfinished gaps on human chr 20, determined their complete genomic sizes and assessed epigenetic profiles using a combination of Sanger sequencing, mate pair paired-end high-throughput sequencing...... and chromatin, methylation and expression analyses. We found histone 3 trimethylated at Lysine 27 to be distributed across all three gaps in immortalized B-lymphocytes. In one gap, five novel CpG islands were predominantly hypermethylated in genomic DNA from peripheral blood lymphocytes and human cerebellum...

  14. Prognostic value of metastin expression in human pancreatic cancer

    OpenAIRE

    Kawaguchi Yoshiya; Masui Toshihiko; Koizumi Masayuki; Kida Atsushi; Ito Tatsuo; Katagiri Fumihiko; Doi Ryuichiro; Nagai Kazuyuki; Tomita Kenji; Oishi Shinya; Fujii Nobutaka; Uemoto Shinji

    2009-01-01

    Abstract Background KiSS-1 was identified as a metastasis-suppressing gene in melanoma cells. The KiSS-1 gene product (metastin) was isolated from human placenta as the ligand of GPR54, a G-protein-coupled receptor. The role of metastin and GPR54 in tumor progression is not fully understood. Methods We investigated the clinical significance of metastin and GPR54 expression in pancreatic cancer. We evaluated immunohistochemical expression of metastin and GPR54 in pancreatic ductal adenocarcino...

  15. Prognostic value of metastin expression in human pancreatic cancer

    OpenAIRE

    Nagai, Kazuyuki; Doi, Ryuichiro; Katagiri, Fumihiko; Ito, Tatsuo; Kida, Atsushi; Koizumi, Masayuki; Masui, Toshihiko; Kawaguchi, Yoshiya; Tomita, Kenji; Oishi, Shinya; Fujii, Nobutaka; Uemoto, Shinji

    2009-01-01

    Background KiSS-1 was identified as a metastasis-suppressing gene in melanoma cells. The KiSS-1 gene product (metastin) was isolated from human placenta as the ligand of GPR54, a G-protein-coupled receptor. The role of metastin and GPR54 in tumor progression is not fully understood. Methods We investigated the clinical significance of metastin and GPR54 expression in pancreatic cancer. We evaluated immunohistochemical expression of metastin and GPR54 in pancreatic ductal adenocarcinoma tissue...

  16. Prognostic value of metastin expression in human pancreatic cancer.

    OpenAIRE

    Nagai, Kazuyuki; Doi, Ryuichiro; Katagiri, Fumihiko; Ito, Tatsuo; Kida, Atsushi; Koizumi, Masayuki; Masui, Toshihiko; Kawaguchi, Yoshiya; Tomita, Kenji; Oishi, Shinya; Fujii, Nobutaka; Uemoto, Shinji

    2009-01-01

    [Background]KiSS-1 was identified as a metastasis-suppressing gene in melanoma cells. The KiSS-1 gene product (metastin) was isolated from human placenta as the ligand of GPR54, a G-protein-coupled receptor. The role of metastin and GPR54 in tumor progression is not fully understood. [Methods]We investigated the clinical significance of metastin and GPR54 expression in pancreatic cancer. We evaluated immunohistochemical expression of metastin and GPR54 in pancreatic ductal adenocarcinoma tiss...

  17. A novel SGLT is expressed in the human kidney

    OpenAIRE

    Kothinti, Rajendra K.; Blodgett, Amy B.; North, Paula E.; Roman, Richard J.; Tabatabai, Niloofar M.

    2012-01-01

    Selective inhibitors of sodium-glucose cotransporter 2 (SGLT2)-mediated reabsorption of glucose in the proximal tubule of the kidney are being developed for the treatment of diabetes. SGLT2 shares high degree of homology with SGLT3; however, very little is known about the expression and functional role of SGLT3 in the human kidney. Indeed, the SGLT2 inhibitors that are currently in clinical trials might affect the expression and/or the activity of SGLT3. Therefore, the present study examined ...

  18. Prognostic value of metastin expression in human pancreatic cancer

    Directory of Open Access Journals (Sweden)

    Kawaguchi Yoshiya

    2009-01-01

    Full Text Available Abstract Background KiSS-1 was identified as a metastasis-suppressing gene in melanoma cells. The KiSS-1 gene product (metastin was isolated from human placenta as the ligand of GPR54, a G-protein-coupled receptor. The role of metastin and GPR54 in tumor progression is not fully understood. Methods We investigated the clinical significance of metastin and GPR54 expression in pancreatic cancer. We evaluated immunohistochemical expression of metastin and GPR54 in pancreatic ductal adenocarcinoma tissues obtained from 53 consecutive patients who underwent resection between July 2003 and May 2007 at Kyoto University Hospital. In 23 consecutive patients, the plasma metastin level was measured before surgery by enzyme immunoassay. Results Strong immunohistochemical expression of metastin was detected in 13 tumors (24.5%, while strong expression of GPR54 was detected in 30 tumors (56.6%. Tumors that were negative for both metastin and GPR54 expression were significantly larger than tumors that were positive for either metastin or GPR54 (p = 0.047. Recurrence was less frequent in patients who had metastin-positive tumors compared with those who had metastin-negative tumors (38.5% versus 70.0%, p = 0.04. Strong expression of metastin and GPR54 was significantly correlated with longer survival (p = 0.02. Metastin expression by pancreatic cancer was an independent prognostic factor for longer survival (hazard ratio, 2.1; 95% confidence interval, 1.1–4.7; p = 0.03, and the patients with a high plasma metastin level (n = 6 did not die after surgical resection. Conclusion Strong expression of metastin and GPR54 by pancreatic cancer is associated with longer survival. Metastin expression is an independent prognostic factor for the survival of pancreatic cancer patients. The plasma metastin level could become a noninvasive prognostic factor for the assessment of pancreatic cancer.

  19. MicroRNA expression variability in human cervical tissues.

    Directory of Open Access Journals (Sweden)

    Patrícia M Pereira

    Full Text Available MicroRNAs (miRNAs are short (approximately 22 nt non-coding regulatory RNAs that control gene expression at the post-transcriptional level. Deregulation of miRNA expression has been discovered in a wide variety of tumours and it is now clear that they contribute to cancer development and progression. Cervical cancer is one of the most common cancers in women worldwide and there is a strong need for a non-invasive, fast and efficient method to diagnose the disease. We investigated miRNA expression profiles in cervical cancer using a microarray platform containing probes for mature miRNAs. We have evaluated miRNA expression profiles of a heterogeneous set of cervical tissues from 25 different patients. This set included 19 normal cervical tissues, 4 squamous cell carcinoma, 5 high-grade squamous intraepithelial lesion (HSIL and 9 low-grade squamous intraepithelial lesion (LSIL samples. We observed high variability in miRNA expression especially among normal cervical samples, which prevented us from obtaining a unique miRNA expression signature for this tumour type. However, deregulated miRNAs were identified in malignant and pre-malignant cervical tissues after tackling the high expression variability observed. We were also able to identify putative target genes of relevant candidate miRNAs. Our results show that miRNA expression shows natural variability among human samples, which complicates miRNA data profiling analysis. However, such expression noise can be filtered and does not prevent the identification of deregulated miRNAs that play a role in the malignant transformation of cervical squamous cells. Deregulated miRNAs highlight new candidate gene targets allowing for a better understanding of the molecular mechanism underlying the development of this tumour type.

  20. The structure and expression of the human neuroligin-3 gene.

    Science.gov (United States)

    Philibert, R A; Winfield, S L; Sandhu, H K; Martin, B M; Ginns, E I

    2000-04-04

    The neuroligins are a family of proteins that are thought to mediate cell to cell interactions between neurons. During the sequencing at an Xq13 locus associated with a mental retardation syndrome in some studies, we discovered a portion of the human orthologue of the rat neuroligin-3 gene. We now report the structure and the expression of that gene. The gene spans approximately 30kb and contains eight exons. Unlike the rat gene, it codes for at least two mRNAs and at least one of which is expressed outside the CNS. Interestingly, the putative promoter for the gene overlaps the last exon of the neighboring HOPA gene and is located less than 1kb from an OPA element in which a polymorphism associated with mental retardation is found. These findings suggest a possible role for the neuroligin gene in mental retardation and that the role of the gene in humans may differ from its role in rats.

  1. ANALYSES ON DIFFERENTIALLY EXPRESSED GENES ASSOCIATED WITH HUMAN BREAST CANCER

    Institute of Scientific and Technical Information of China (English)

    MENG Xu-li; DING Xiao-wen; XU Xiao-hong

    2006-01-01

    Objective: To investigate the molecular etiology of breast cancer by way of studying the differential expression and initial function of the related genes in the occurrence and development of breast cancer. Methods: Two hundred and eighty-eight human tumor related genes were chosen for preparation of the oligochips probe. mRNA was extracted from 16 breast cancer tissues and the corresponding normal breast tissues, and cDNA probe was prepared through reverse-transcription and hybridized with the gene chip. A laser focused fluorescent scanner was used to scan the chip. The different gene expressions were thereafter automatically compared and analyzed between the two sample groups. Cy3/Cy5>3.5 meant significant up-regulation. Cy3/Cy5<0.25 meant significant down-regulation. Results: The comparison between the breast cancer tissues and their corresponding normal tissues showed that 84 genes had differential expression in the Chip. Among the differently expressed genes, there were 4 genes with significant down-regulation and 6 with significant up-regulation. Compared with normal breast tissues, differentially expressed genes did partially exist in the breast cancer tissues. Conclusion: Changes in multi-gene expression regulations take place during the occurrence and development of breast cancer; and the research on related genes can help understanding the mechanism of tumor occurrence.

  2. Copper induces the expression of cholesterogenic genes in human macrophages.

    Science.gov (United States)

    Svensson, Per Arne; Englund, Mikael C O; Markström, Emilia; Ohlsson, Bertil G; Jernås, Margareta; Billig, Håkan; Torgerson, Jarl S; Wiklund, Olov; Carlsson, Lena M S; Carlsson, Björn

    2003-07-01

    Accumulation of lipids and cholesterol by macrophages and subsequent transformation into foam cells are key features in development of atherosclerosis. Serum copper concentrations have been shown to be associated with cardiovascular disease. However, the mechanism behind the proatherogenic effect of copper is not clear. We used DNA microarrays to define the changes in gene expression profile in response to copper exposure of human macrophages. Expression monitoring by DNA microarray revealed 91 genes that were regulated. Copper increased the expression of seven cholesterogenic genes (3-hydroxy-3-methylglutaryl coenzyme A (HMG CoA) synthase, IPP isomerase, squalene synthase, squalene epoxidase, methyl sterol oxidase, H105e3 mRNA and sterol-C5-desaturase) and low-density lipoprotein receptor (LDL-R), and decreased the expression of CD36 and lipid binding proteins. The expression of LDL-R and HMG CoA reductase was also investigated using real time PCR. The expression of both of these genes was increased after copper treatment of macrophages (Pmechanism for the association between copper and atherosclerosis. The effect of copper on cholesterogenic genes may also have implications for liver steatosis in early stages of Wilson's disease.

  3. Expression of human α1-proteinase inhibitor in Aspergillus niger

    Directory of Open Access Journals (Sweden)

    Punt Peter J

    2007-10-01

    Full Text Available Abstract Background Human α1-proteinase inhibitor (α1-PI, also known as antitrypsin, is the most abundant serine protease inhibitor (serpin in plasma. Its deficiency is associated with development of progressive, ultimately fatal emphysema. Currently in the United States, α1-PI is available for replacement therapy as an FDA licensed plasma-derived (pd product. However, the plasma source itself is limited; moreover, even with efficient viral inactivation steps used in manufacture of plasma products, the risk of contamination from emerging viruses may still exist. Therefore, recombinant α1-PI (r-α1-PI could provide an attractive alternative. Although r-α1-PI has been produced in several hosts, protein stability in vitro and rapid clearance from the circulation have been major issues, primarily due to absent or altered glycosylation. Results We have explored the possibility of expressing the gene for human α1-PI in the filamentous fungus Aspergillus niger (A. niger, a system reported to be capable of providing more "mammalian-like" glycosylation patterns to secretable proteins than commonly used yeast hosts. Our expression strategy was based on fusion of α1-PI with a strongly expressed, secreted leader protein (glucoamylase G2, separated by dibasic processing site (N-V-I-S-K-R that provides in vivo cleavage. SDS-PAGE, Western blot, ELISA, and α1-PI activity assays enabled us to select the transformant(s secreting a biologically active glycosylated r-α1-PI with yields of up to 12 mg/L. Matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS analysis further confirmed that molecular mass of the r-α1-PI was similar to that of the pd-α1-PI. In vitro stability of the r-α1-PI from A. niger was tested in comparison with pd-α1-PI reference and non-glycosylated human r-α1-PI from E. coli. Conclusion We examined the suitability of the filamentous fungus A. niger for the expression of the human gene for α1-PI, a medium size

  4. Transgenic rabbit that expresses a functional human lipoprotein (a)

    Science.gov (United States)

    Rouy, Didier; Duverger, Nicolas; Emmanuel, Florence; Denefle, Patrice; Houdebine, Louis-Marie; Viglietta, Celine; Rubin, Edward M.; Hughes, Steven D.

    2003-01-01

    A transgenic rabbit which has in its genomic DNA sequences that encode apolipoprotein (a) and apolipoprotein B polypeptides which are capable of combining to produce lipoprotein (a), a process for creating such a rabbit, and the use of the rabbit to identify compounds which are effective in the treatment of human diseases which are associated with, induced and/or exacerbated by Lp(a) expression.

  5. The Signature of Selection Mediated by Expression on Human Genes

    OpenAIRE

    Urrutia, Araxi O.; Hurst, Laurence D

    2003-01-01

    As the efficacy of natural selection is expected to be a function of population size, in humans it is usually presumed that selection is a weak force and hence that gene characteristics are mostly determined by stochastic forces. In contrast, in species with large population sizes, selection is expected to be a much more effective force. Evidence for this has come from examining how genic parameters vary with expression level, which appears to determine many of a gene's features, such as codo...

  6. Robust, synergistic regulation of human gene expression using TALE activators.

    Science.gov (United States)

    Maeder, Morgan L; Linder, Samantha J; Reyon, Deepak; Angstman, James F; Fu, Yanfang; Sander, Jeffry D; Joung, J Keith

    2013-03-01

    Artificial activators designed using transcription activator-like effector (TALE) technology have broad utility, but previous studies suggest that these monomeric proteins often exhibit low activities. Here we demonstrate that TALE activators can robustly function individually or in synergistic combinations to increase expression of endogenous human genes over wide dynamic ranges. These findings will encourage applications of TALE activators for research and therapy, and guide design of monomeric TALE-based fusion proteins.

  7. Identification of Haemophilus ducreyi genes expressed during human infection.

    Science.gov (United States)

    Bauer, Margaret E; Fortney, Kate R; Harrison, Alistair; Janowicz, Diane M; Munson, Robert S; Spinola, Stanley M

    2008-04-01

    To identify Haemophilus ducreyi transcripts that are expressed during human infection, we used selective capture of transcribed sequences (SCOTS) with RNA isolated from pustules obtained from three volunteers infected with H. ducreyi, and with RNA isolated from broth-grown bacteria used to infect volunteers. With SCOTS, competitive hybridization of tissue-derived and broth-derived sequences identifies genes that may be preferentially expressed in vivo. Among the three tissue specimens, we identified 531 genes expressed in vivo. Southern blot analysis of 60 genes from each tissue showed that 87 % of the identified genes hybridized better with cDNA derived from tissue specimens than with cDNA derived from broth-grown bacteria. RT-PCR on nine additional pustules confirmed in vivo expression of 10 of 11 selected genes in other volunteers. Of the 531 genes, 139 were identified in at least two volunteers. These 139 genes fell into several functional categories, including biosynthesis and metabolism, regulation, and cellular processes, such as transcription, translation, cell division, DNA replication and repair, and transport. Detection of genes involved in anaerobic and aerobic respiration indicated that H. ducreyi likely encounters both microenvironments within the pustule. Other genes detected suggest an increase in DNA damage and stress in vivo. Genes involved in virulence in other bacterial pathogens and 32 genes encoding hypothetical proteins were identified, and may represent novel virulence factors. We identified three genes, lspA1, lspA2 and tadA, known to be required for virulence in humans. This is the first study to broadly define transcripts expressed by H. ducreyi in humans.

  8. Human myometrial gene expression before and during parturition.

    Science.gov (United States)

    Havelock, Jon C; Keller, Patrick; Muleba, Ndaya; Mayhew, Bobbie A; Casey, Brian M; Rainey, William E; Word, R Ann

    2005-03-01

    Identification of temporal and spatial changes in myometrial gene expression during parturition may further the understanding of the coordinated regulation of myometrial contractions during parturition. The objective of this study was to compare the gene expression profiles of human fundal myometrium from pregnant women before and after the onset of labor using a functional genomics approach, and to further characterize the spatial and temporal expression patterns of three genes believed to be important in parturition. Fundal myometrial mRNA was isolated from five women in labor and five women not in labor, and analyzed using human UniGEM-V microarrays with 9182 cDNA elements. Real-time polymerase chain reaction using myometrial RNA from pregnant women in labor or not in labor was used to examine mRNA levels for three of the genes; namely, prostaglandin-endoperoxide synthase 2 (PTGS2), calgranulin B (S100A9), and oxytocin receptor (OXTR). The spatial expression pattern of these genes throughout the pregnant uterus before and after labor was also determined. Immunolocalization of cyclooxygenase-2 (also known as PTGS2) and S100A9 within the uterine cervix and myometrium were analyzed by immunohistochemistry. Few genes were differentially expressed in fundal myometrial tissues at term with the onset of labor. However, there appears to be a subset of genes important in the parturition cascade. The cellular properties of S100A9, its spatial localization, and dramatic increase in cervix and myometrium of women in labor suggest that this protein may be very important in the initiation or propagation of human labor.

  9. Transcriptional regulation of human thromboxane synthase gene expression

    Energy Technology Data Exchange (ETDEWEB)

    Lee, K.D.; Baek, S.J.; Fleischer, T [Univ. of Maryland Medical School, Baltimore, MD (United States)] [and others

    1994-09-01

    The human thromboxane synthase (TS) gene encodes a microsomal enzyme catalyzing the conversion of prostaglandin endoperoxide into thromboxane A{sub 2}(TxA{sub 2}), a potent inducer of vasoconstriction and platelet aggregation. A deficiency in platelet TS activity results in bleeding disorders, but the underlying molecular mechanism remains to be elucidated. Increased TxA{sub 2} has been associated with many pathophysiological conditions such as cardiovascular disease, pulmonary hypertension, pre-eclampsia, and thrombosis in sickle cell patients. Since the formation of TxA{sub 2} is dependent upon TS, the regulation of TS gene expression may presumably play a crucial role in vivo. Abrogation of the regulatory mechanism in TS gene expression might contribute, in part, to the above clinical manifestations. To gain insight into TS gene regulation, a 1.7 kb promoter of the human TS gene was cloned and sequenced. RNase protection assay and 5{prime} RACE protocols were used to map the transcription initiation site to nucleotide A, 30 bp downstream from a canonical TATA box. Several transcription factor binding sites, including AP-1, PU.1, and PEA3, were identified within this sequence. Transient expression studies in HL-60 cells transfected with constructs containing various lengths (0.2 to 5.5 kb) of the TS promoter/luciferase fusion gene indicated the presence of multiple repressor elements within the 5.5 kb TS promoter. However, a lineage-specific up-regulation of TS gene expression was observed in HL-60 cells induced by TPA to differentiate along the macrophage lineage. The increase in TS transcription was not detectable until 36 hr after addition of the inducer. These results suggest that expression of the human TS gene may be regulated by a mechanism involving repression and derepression of the TS promoter.

  10. Study of human dopamine sulfotransferases based on gene expression programming.

    Science.gov (United States)

    Si, Hongzong; Zhao, Jiangang; Cui, Lianhua; Lian, Ning; Feng, Hanlin; Duan, Yun-Bo; Hu, Zhide

    2011-09-01

    A quantitative model is developed to predict the Km of 47 human dopamine sulfotransferases by gene expression programming. Each kind of compound is represented by several calculated structural descriptors of moment of inertia A, average electrophilic reactivity index for a C atom, relative number of triple bonds, RNCG relative negative charge, HA-dependent HDSA-1, and HBCA H-bonding charged surface area. Eight fitness functions of the gene expression programming method are used to find the best nonlinear model. The best quantitative model with squared standard error and square of correlation coefficient are 0.096 and 0.91 for training data set, and 0.102 and 0.88 for test set, respectively. It is shown that the gene expression programming-predicted results with fitness function are in good agreement with experimental ones.

  11. Human Neural Cells Transiently Express Reelin during Olfactory Placode Development.

    Directory of Open Access Journals (Sweden)

    M Cristina Antal

    Full Text Available Reelin, an extracellular glycoprotein is essential for migration and correct positioning of neurons during development. Since the olfactory system is known as a source of various migrating neuronal cells, we studied Reelin expression in the two chemosensory olfactory systems, main and accessory, during early developmental stages of human foetuses/embryos from Carnegie Stage (CS 15 to gestational week (GW 14. From CS 15 to CS 18, but not at later stages, a transient expression of Reelin was detected first in the presumptive olfactory and then in the presumptive vomeronasal epithelium. During the same period, Reelin-positive cells detach from the olfactory/vomeronasal epithelium and migrate through the mesenchyme beneath the telencephalon. Dab 1, an adaptor protein of the Reelin pathway, was simultaneously expressed in the migratory mass from CS16 to CS17 and, at later stages, in the presumptive olfactory ensheathing cells. Possible involvements of Reelin and Dab 1 in the peripheral migrating stream are discussed.

  12. Expression of cyclooxygenase-2 in human esophageal squamous cell carcinomas

    Institute of Scientific and Technical Information of China (English)

    Jian-Gang Jiang; Dao-Wen Wang; Jiang-Bo Tang; Chun-Lian Chen; Bao-Xing Liu; Xiang-Ning Fu; Zhi-Hui Zhu; Wei Qu; Katherine Cianflone; Michael P. Waalkes

    2004-01-01

    AIM: To determine whether cyclooxygenase-2 (COX-2) was expressed in human esophageal squamous cell carcinoma.METHODS: Quantitative reverse transcription-polymerase chain reaction (RT-PCR), western blotting, immunohistochemistry and immunofluorescence were used to assess the expression level of COX-2 in esophageal tissue.RESULTS: COX-2 mRNA levels were increased by >80-fold in esophageal squamous cell carcinoma when compared to adjacent noncancerous tissue. COX-2 protein was present in 21 of 30 cases of esophageal squamous cell carcinoma tissues, but was undetectable in noncancerous tissue. Immunohistochemistry was performed to directly show expression of COX-2 in tumor tissue.CONCLUSION: These results suggest that COX-2 may be an important factor for esophageal cancer and inhibition of COX-2 may be helpful for prevention and possibly treatment of this cancer.

  13. A recombinant vaccinia virus expressing human carcinoembryonic antigen (CEA).

    Science.gov (United States)

    Kaufman, H; Schlom, J; Kantor, J

    1991-07-30

    Carcinoembryonic antigen (CEA) is a 180-kDa glycoprotein expressed on most gastrointestinal carcinomas. A 2.4-kb cDNA clone, containing the complete coding sequence, was isolated from a human colon tumor cell library and inserted into a vaccinia virus genome. This newly developed construct was characterized by Southern blotting, DNA hybridization studies, and polymerase chain reaction analysis. The CEA gene was stably integrated into the vaccinia virus thymidine kinase gene. The recombinant was efficiently replicated upon serial passages in cell cultures and in animals. The recombinant virus expresses on the surface of infected cells a protein product recognized by a monoclonal antibody (COL-I) directed against CEA. Immunization of mice with the vaccinia construct elicited a humoral immune response against CEA. Pilot studies also showed that administration of the recombinant CEA vaccinia construct was able to greatly reduce the growth in mice of a syngeneic murine colon adenocarcinoma which had been transduced with the human CEA gene. The use of this new recombinant CEA vaccinia construct may thus provide an approach in the specific active immunotherapy of human GI cancer and other CEA expressing carcinoma types.

  14. Mapping the genetic architecture of gene expression in human liver.

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    Eric E Schadt

    2008-05-01

    Full Text Available Genetic variants that are associated with common human diseases do not lead directly to disease, but instead act on intermediate, molecular phenotypes that in turn induce changes in higher-order disease traits. Therefore, identifying the molecular phenotypes that vary in response to changes in DNA and that also associate with changes in disease traits has the potential to provide the functional information required to not only identify and validate the susceptibility genes that are directly affected by changes in DNA, but also to understand the molecular networks in which such genes operate and how changes in these networks lead to changes in disease traits. Toward that end, we profiled more than 39,000 transcripts and we genotyped 782,476 unique single nucleotide polymorphisms (SNPs in more than 400 human liver samples to characterize the genetic architecture of gene expression in the human liver, a metabolically active tissue that is important in a number of common human diseases, including obesity, diabetes, and atherosclerosis. This genome-wide association study of gene expression resulted in the detection of more than 6,000 associations between SNP genotypes and liver gene expression traits, where many of the corresponding genes identified have already been implicated in a number of human diseases. The utility of these data for elucidating the causes of common human diseases is demonstrated by integrating them with genotypic and expression data from other human and mouse populations. This provides much-needed functional support for the candidate susceptibility genes being identified at a growing number of genetic loci that have been identified as key drivers of disease from genome-wide association studies of disease. By using an integrative genomics approach, we highlight how the gene RPS26 and not ERBB3 is supported by our data as the most likely susceptibility gene for a novel type 1 diabetes locus recently identified in a large

  15. Expression of 5-Lipoxygenase in human colorectal cancer

    Institute of Scientific and Technical Information of China (English)

    Labile Togba Soumaoro; Satoru Iida; Hiroyuki Uetake; Megumi Ishiguro; Yoko Takagi; Tetsuro Higuchi; Masamichi Yasuno; Masayuki Enomoto; Kenichi Sugihara

    2006-01-01

    AIM: To evaluate the 5-lipoxygenases (Loxs) expression level in human colorectal cancer specimens in order to determine its clinicopathologic significance in human tumorigenesis.METHODS: The relative quantity of 5-Lox mRNA in paired 91 colorectal tumor and adjacent normal mucosa samples was determined by real time quantitative PCR. Additionally, the expression of 5-Lox and cyclooxygenase (Cox)-2 proteins was also examined using immunohistochemical staining methods.RESULTS: There was a marked increase in 5-Lox mRNA levels in the tumor compared with paired normal mucosa samples (P < 0.0001). Sixty six (72.5%) tumors showed high 5-Lox mRNA levels. The positivity rate of 5-Lox and Cox-2 protein expression was 68.7% and 79.1%respectively. There was a significant association between tumoral 5-Lox mRNA level and tumor size (Rho = 0.392,P = 0.0002), depth or vessel invasion.CONCLUSION: These results suggest that 5-Lox is up-regulated in colorectal cancer and that inhibition of its expression might be valuable in the prevention and treatment of colorectal cancer.

  16. Expression of pemphigus-autoantigen desmoglein 1 in human thymus.

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    Mouquet, H; Berrih-Aknin, S; Bismuth, J; Joly, P; Gilbert, D; Tron, F

    2008-05-01

    Desmoglein (Dsg) 1 is a transmembrane glycoprotein of the desmosome allowing cell-cell adhesion between keratinocytes, whose expression is restricted to stratified squamous epithelia-like epidermis. Dsg1 is the target autoantigen of pathogenic autoantibodies produced by pemphigus foliaceus and 50% of pemphigus vulgaris patients in a Dsg1-specific T-cell-dependent pathway. Herewith, we show that mRNA of the DSG1 gene is present in normal human thymus and show by quantitative real-time polymerase chain reaction analysis that the expression of DSG1 transcript increases with age. Although immunoblot analysis on human thymus extracts using different anti-Dsg1 antibodies did not allow to detect the protein, we show by double-immunofluorescence assay that Dsg1 is expressed at protein level by CD19+ CD63+ cells located in the medulla. These data provide another illustration of the thymic expression of a tissue-specific autoantigen involved in an organ-specific autoimmune disease, which may participate in the tolerance acquisition and/or regulation of Dsg1-specific T cells.

  17. EXPRESSION AND SIGNIFICANCE OF ERK PROTEIN IN HUMAN BREAST CARCINOMA

    Institute of Scientific and Technical Information of China (English)

    张秀梅; 李柏林; 宋敏; 宋继谒

    2004-01-01

    Objective: To investigate the expression of ERK and p-ERK protein in human breast cancer and their corresponding tissue, to assess the significance of ERK signal pathway in tumorigenesis and progression of breast carcinoma. Methods: 40 breast cancer cases were used in S-P immunohistochemistry technique and Western Blot study. Results: The expression of ERK1, ERK2, and p- ERK protein levels increased remarkably in breast cancer tissues in comparison to normal tissues (P<0.01). The expression was upregulated by 1.32-, 1.53-and 4.27-fold, respectively. The overexpressions of ERK1, ERK2, and p- ERK proteins were obviously correlated with clinical stage of breast cancer. Protein levels of ERK and p-ERK were higher in stage III patients than in stage I and stage II patients (P<0.05). These proteins were strongly related with axillary lymph node metastasis of breast cancer, but not correlated with histopathological type and status of ER and PR of breast cancer. Expression of ERK1, and ERK2, protein showed a positive linear correlation. Conclusion: ERK signal transduction pathway is a key factor during human breast tumorigenesis and breast cancer progression.

  18. Modulation of HLA Expression in Human Cytomegalovirus Immune Evasion

    Institute of Scientific and Technical Information of China (English)

    Aifen Lin; Huihui Xu; Weihua Yan

    2007-01-01

    Human cytomegalovirus (hCMV) has evolved multiple mechanisms to escape the host immune recognition and innate or adaptive immune responses. Among them, hCMV has developed strategies to modulate the expression and/or function of human leukocyte antigens (HLAs), including by encoding series of infection stage-dependent hCMV proteins to detain and destroy the expression of HLA molecules on the surface of infected cells. This disturbs the antigen presentation and processing, by encoding MHC class Ⅰ homologues or selective up-regulation of particular HLA class Ⅰ molecules binding to NK cell inhibitory receptors, and by encoding specific ligand antagonists to interfere with NK cell activating receptors. Here we discussed the molecular mechanisms utilized by the hCMV to alter the formation, transportation and expression of HLA antigens on the infected cell surface. The knowledge about hCMV modulating HLA expression could benefit us to further understand the pathogenesis of viral diseases and may eventually develop novel effective immunotherapies to counteract viral infections and viral associated diseases.

  19. Comparative expression pathway analysis of human and canine mammary tumors

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    Marconato Laura

    2009-03-01

    Full Text Available Abstract Background Spontaneous tumors in dog have been demonstrated to share many features with their human counterparts, including relevant molecular targets, histological appearance, genetics, biological behavior and response to conventional treatments. Mammary tumors in dog therefore provide an attractive alternative to more classical mouse models, such as transgenics or xenografts, where the tumour is artificially induced. To assess the extent to which dog tumors represent clinically significant human phenotypes, we performed the first genome-wide comparative analysis of transcriptional changes occurring in mammary tumors of the two species, with particular focus on the molecular pathways involved. Results We analyzed human and dog gene expression data derived from both tumor and normal mammary samples. By analyzing the expression levels of about ten thousand dog/human orthologous genes we observed a significant overlap of genes deregulated in the mammary tumor samples, as compared to their normal counterparts. Pathway analysis of gene expression data revealed a great degree of similarity in the perturbation of many cancer-related pathways, including the 'PI3K/AKT', 'KRAS', 'PTEN', 'WNT-beta catenin' and 'MAPK cascade'. Moreover, we show that the transcriptional relationships between different gene signatures observed in human breast cancer are largely maintained in the canine model, suggesting a close interspecies similarity in the network of cancer signalling circuitries. Conclusion Our data confirm and further strengthen the value of the canine mammary cancer model and open up new perspectives for the evaluation of novel cancer therapeutics and the development of prognostic and diagnostic biomarkers to be used in clinical studies.

  20. EOTAXIN AND EOTAXIN-2 EXPRESSION IN HUMAN BRONCHIAL EPITHELIAL CELL

    Institute of Scientific and Technical Information of China (English)

    TANG Wei; DENG Wei-wu; Albert CHAN; Stanley CHIK; Adrain WU

    2005-01-01

    Objective To study the role of eotaxin and eotaxin-2 expression by Th2 cytokine and analyze their relationship in normal human bronchial epithelial cell line-BEAS-2B cell. Methods Levels of eotaxin mRNA and protein expression in the bronchial epithelial cell line BEAS-2B cell were determined with RT-PCR and ELISA. We also used RT-PCR to evaluate eotaxin-2 expression under the regulation of Th2 cytokine IL-4 and IL-13 as well as proinflammatory agent-TNFα. Results Eotaxin mRNA expression was the highest at the time point of 12h under the stimulation of TNF-α. While Th2 cytokine IL-4 and IL-13 had the amplification effect on the expression. Eotaxin protein was also elevated with the combination stimulation of proinflammatory agent TNF-α and IL-4 in dose and time dependent manner(P<0.01). These results were also seen when the cells were stimulated by TNF-α and IL-13. Eotaxin-2 mRNA expression was the highest at the time point of 8h. The expression evaluated by semi-quantitative RT-PCR also elevated under the co-stimulation of TNF-α and IL-4 or TNF-α and IL-13 and it should significantly correlate with Eotaxin(P<0.05). Conclusion This study demonstrated that Th2 cytokine like IL-4 and IL-13 enhances eotaxin and eotaxin-2 expression when co-stimulated with proinflammatory agent TNF-α. These results showed that Th2 cytokines existence is the strong evidence for bronchial epithelial cells taking part in the allergic inflammation especially in eosinophils recruitment.

  1. Midkine expression in 52 human meningiomas A correlation analysis

    Institute of Scientific and Technical Information of China (English)

    Xinjun Li; Xiangguo Xia

    2008-01-01

    BACKGROUND: Several studies have shown that midkine directly participates in tumor cell growth and invasion, as well as the regulation of angiogenesis.OBJECTIVE: To investigate midkine expression in meningioma tissue in relation to angiogenesis, invasion, peritumoral edema, and clinicopathology.DESIGN, TIME AND SETTING: The present clinical, case-controlled, neuropathological study was performed at the Laboratory of Molecular Organism, People's Hospital of Deyang City between May 2007 and April 2008.MATERIALS: Fifty-two meningioma tissues were classified by WHO tumor classification of the central nervous system, comprising 40 grade Ⅰ meningioma, five grade Ⅱ meningioma, and seven grade Ⅲ meningioma. Ten normal, human cerebral maters were selected from cerebral trauma patients.METHODS: Midkine protein expression and mean microvessel density were detected using immunohistochemical techniques. Simultaneously, all data were statistically analyzed.MAIN OUTCOME MEASURES: Midkine expression and microvessel density in meningiomas and normal cerebral maters.RESULTS: The positive midkine expression rate was 64% in the meningioma tissues. However, midkine expression was not detected in normal cerebral mater tissue. The mean microvessel density was 82.0±22.7 in the meningiomas, and 25.8±6.2 in the normal cerebral mater tissues. There was significant difference in midkine expression and mean microvessel density between meningioma tissues and human cerebral maters (P 0.05). However, it closely correlated with patient clinical condition, pathological grade, invasion, and peritumoral edema (r =0.378 5, 0.741 2, 0.651 8, 0.614 2, P < 0.05).CONCLUSION: Midkinc protein was overexpressed in meningiomas and correlated to tumor angiogenesis, invasion, peritumoral edema, and clinicopathology.

  2. Cytochrome P450 aromatase expression in human seminoma

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    Montanaro Daniela

    2005-12-01

    Full Text Available Abstract Background The enzyme cytochrome P450 aromatase, catalysing the conversion of androgens into estrogens, has been detected in normal human testicular cells suggesting a physiological role of local estrogen biosynthesis on spermatogenesis control. Estrogens, regulating cell growth and apoptosis, can also be involved in tumorigenesis process, but the possible link between estrogens and testicular neoplastic process is, up to now, scarcely known. This study examined aromatase expression in human seminoma, which is the most common germ cell tumour of the testis. Methods The tumour-bearing testes were obtained from 20 patients with classic seminoma undergoing to therapeutic orchidectomy. Paraffin embedded tissues were processed for immunohistochemistry using a mouse monoclonal antibody generated against human placental cytochrome P450 arom, as primary antibody, and a biotinylated goat-anti-mouse IgG, as secondary antibody. Furthermore, Western blot analysis of seminoma extracts was carried out. Results Intense P450 arom immunoreactivity was observed in the seminoma cells and Western blot analysis confirmed the immunodetection. A strong immunostaining was also detected in cells of intratubular germ cell neoplasia (IGCN, adjacent to seminoma. Conclusion The present study demonstrated, for the first time in human, aromatase expression in neoplastic cells of seminoma suggesting a relation between local estrogen biosynthesis and germ cell tumorigenesis. The P450 arom immunolocalization in the cells of IGCN, representing the common precursor of most germ cell tumors, seems to support these findings.

  3. Regulation of MYCN expression in human neuroblastoma cells

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    de Vries I

    2009-07-01

    Full Text Available Abstract Background Amplification of the MYCN gene in neuroblastoma (NB is associated with a poor prognosis. However, MYCN-amplification does not automatically result in higher expression of MYCN in children with NB. We hypothesized that the discrepancy between MYCN gene expression and prognosis in these children might be explained by the expression of either MYCN-opposite strand (MYCNOS or the shortened MYCN-isoform (ΔMYCN that was recently identified in fetal tissues. Both MYCNOS and ΔMYCN are potential inhibitors of MYCN either at the mRNA or at the protein level. Methods Expression of MYCN, MYCNOS and ΔMYCN was measured in human NB tissues of different stages. Transcript levels were quantified using a real-time reverse transcriptase polymerase chain reaction assay (QPCR. In addition, relative expression of these three transcripts was compared to the number of MYCN copies, which was determined by genomic real-time PCR (gQPCR. Results Both ΔMYCN and MYCNOS are expressed in all NBs examined. In NBs with MYCN-amplification, these transcripts are significantly higher expressed. The ratio of MYCN:ΔMYCN expression was identical in all tested NBs. This indicates that ΔMYCN and MYCN are co-regulated, which suggests that ΔMYCN is not a regulator of MYCN in NB. However, the ratio of MYCNOS:MYCN expression is directly correlated with NB disease stage (p = 0.007. In the more advanced NB stages and NBs with MYCN-amplification, relatively more MYCNOS is present as compared to MYCN. Expression of the antisense gene MYCNOS might be relevant to the progression of NB, potentially by directly inhibiting MYCN transcription by transcriptional interference at the DNA level. Conclusion The MYCNOS:MYCN-ratio in NBs is significantly correlated with both MYCN-amplification and NB-stage. Our data indicate that in NB, MYCN expression levels might be influenced by MYCNOS but not by ΔMYCN.

  4. Interference of CD95 expression on human lymphocytes.

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    Petanova, Jitka; Fucikova, Terezie; Bencko, Vladimir

    2002-02-01

    The study presents the exogenous influence of cadmium in comparison with zinc on the apoptosis of human lymphocytes by CD95 expression and its kinetic changes. The salts of both metals were used in final concentrations of 20 microM in cell cultures with whole blood. The duration of cultivation was 18 and 90 hours. The expression of surface antigens was evaluated by flow cytometry with monoclonal antibodies. In cultures of not stimulated cells we found in average 51.54% CD95 positive lymphocytes. The kinetic study of untreated cells showed elevation after 18 hours of cultivation and a very low expression after 90 hours. The CD95 expression on lymphocytes in cell culture with cadmium and zinc was lower after 18 hours of cultivation than in untreated cells. After 90 hours cultivation we found low levels of CD95 expression on cells treated with cadmium and a great individual variability in the number of positive cells upon the influence of zinc.

  5. Differentially Expressed Genes and Signature Pathways of Human Prostate Cancer.

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    Jennifer S Myers

    Full Text Available Genomic technologies including microarrays and next-generation sequencing have enabled the generation of molecular signatures of prostate cancer. Lists of differentially expressed genes between malignant and non-malignant states are thought to be fertile sources of putative prostate cancer biomarkers. However such lists of differentially expressed genes can be highly variable for multiple reasons. As such, looking at differential expression in the context of gene sets and pathways has been more robust. Using next-generation genome sequencing data from The Cancer Genome Atlas, differential gene expression between age- and stage- matched human prostate tumors and non-malignant samples was assessed and used to craft a pathway signature of prostate cancer. Up- and down-regulated genes were assigned to pathways composed of curated groups of related genes from multiple databases. The significance of these pathways was then evaluated according to the number of differentially expressed genes found in the pathway and their position within the pathway using Gene Set Enrichment Analysis and Signaling Pathway Impact Analysis. The "transforming growth factor-beta signaling" and "Ran regulation of mitotic spindle formation" pathways were strongly associated with prostate cancer. Several other significant pathways confirm reported findings from microarray data that suggest actin cytoskeleton regulation, cell cycle, mitogen-activated protein kinase signaling, and calcium signaling are also altered in prostate cancer. Thus we have demonstrated feasibility of pathway analysis and identified an underexplored area (Ran for investigation in prostate cancer pathogenesis.

  6. Expression analysis of Stat3 in human lung carcinoma

    Institute of Scientific and Technical Information of China (English)

    WANG Hong; HAN Yi-ping

    2002-01-01

    Objective: To analyze the relationship of Stat3 expression with clinical stages, tissue types, p53and proliferation cell nuclear antigen (PCNA) in human lung carcinoma, and to evaluate the role of Stat3 in the pathogenesis of lung carcinoma. Methods: Immunohistochemical method were used to detected Stat3,p53 and PCNA in different tissues of patients (n= 42) with lung carcinoma who accepted neither radiotherapy nor chemotherapy. Results: The positive rate of Stat3 was 81.0% in lung carcinoma and its expression level was related to the tissue type but not to T, N or the clinical stage. The expression level of Stat3 in non-small cell lung carcinoma (NSCLC) was higher than that in small cell lung carcinoma (SCLC). A positive correlation of the expression of Stat3 with that of p53 and PCNA was identified. Conclusion: The expression level of Stat3 is abnormal in lung carcinoma. Stat3 may be involved in the regulation of p53 gene in lung carcinoma cell, it may accelerate the proliferation of lung carcinoma cells and play an important role in the pathogenesis of lung carcinoma.

  7. Differential Mucin Expression by Respiratory Syncytial Virus and Human Metapneumovirus Infection in Human Epithelial Cells

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    Ma. Del Rocío Baños-Lara

    2015-01-01

    Full Text Available Mucins (MUC constitute an important component of the inflammatory and innate immune response. However, the expression of these molecules by respiratory viral infections is still largely unknown. Respiratory syncytial virus (RSV and human metapneumovirus (hMPV are two close-related paramyxoviruses that can cause severe low respiratory tract disease in infants and young children worldwide. Currently, there is not vaccine available for neither virus. In this work, we explored the differential expression of MUC by RSV and hMPV in human epithelial cells. Our data indicate that the MUC expression by RSV and hMPV differs significantly, as we observed a stronger induction of MUC8, MUC15, MUC20, MUC21, and MUC22 by RSV infection while the expression of MUC1, MUC2, and MUC5B was dominated by the infection with hMPV. These results may contribute to the different immune response induced by these two respiratory viruses.

  8. Identification of differentially expressed circular RNAs in human colorectal cancer.

    Science.gov (United States)

    Zhang, Peili; Zuo, Zhigui; Shang, Wenjing; Wu, Aihua; Bi, Ruichun; Wu, Jianbo; Li, Shaotang; Sun, Xuecheng; Jiang, Lei

    2017-03-01

    Circular RNA, a class of non-coding RNA, is a new group of RNAs and is related to tumorigenesis. Circular RNAs are suggested to be ideal candidate biomarkers with potential diagnostic and therapeutic implications. However, little is known about their expression in human colorectal cancer. In our study, differentially expressed circular RNAs were detected using circular RNA array in paired tumor and adjacent non-tumorous tissues from six colorectal cancer patients. Expression levels of selected circular RNAs (hsa_circRNA_103809 and hsa_circRNA_104700) were measured by real-time polymerase chain reaction in 170 paired colorectal cancer samples for validation. Statistical analyses were conducted to investigate the association between hsa_circRNA_103809 and hsa_circRNA_104700 expression levels and respective patient clinicopathological features. Receiver operating characteristic curve was constructed to evaluate the diagnostic values. Our results indicated that there were 125 downregulated and 76 upregulated circular RNAs in colorectal cancer tissues compared with normal tissues. We also first demonstrated that the expression levels of hsa_circRNA_103809 ( p colorectal cancer than in normal tissues. The expression level of hsa_circRNA_103809 was significantly correlated with lymph node metastasis ( p = 0.021) and tumor-node-metastasis stage ( p = 0.011), and the expression level of hsa_circRNA_104700 was significantly correlated with distal metastasis ( p = 0.036). The area under receiver operating characteristic curves of hsa_circRNA_103809 and hsa_circRNA_104700 were 0.699 ( p colorectal cancer and serve as potential biomarkers for the diagnosis of colorectal cancer.

  9. Cyclooxygenases in human and mouse skin and cultured human keratinocytes: association of COX-2 expression with human keratinocyte differentiation

    Science.gov (United States)

    Leong, J.; Hughes-Fulford, M.; Rakhlin, N.; Habib, A.; Maclouf, J.; Goldyne, M. E.

    1996-01-01

    Epidermal expression of the two isoforms of the prostaglandin H-generating cyclooxygenase (COX-1 and COX-2) was evaluated both by immunohistochemistry performed on human and mouse skin biopsy sections and by Western blotting of protein extracts from cultured human neonatal foreskin keratinocytes. In normal human skin, COX-1 immunostaining is observed throughout the epidermis whereas COX-2 immunostaining increases in the more differentiated, suprabasilar keratinocytes. Basal cell carcinomas express little if any COX-1 or COX-2 immunostaining whereas both isozymes are strongly expressed in squamous cell carcinomas deriving from a more differentiated layer of the epidermis. In human keratinocyte cultures, raising the extracellular calcium concentration, a recognized stimulus for keratinocyte differentiation, leads to an increased expression of both COX-2 protein and mRNA; expression of COX-1 protein, however, shows no significant alteration in response to calcium. Because of a recent report that failed to show COX-2 in normal mouse epidermis, we also looked for COX-1 and COX-2 immunostaining in sections of normal and acetone-treated mouse skin. In agreement with a previous report, some COX-1, but no COX-2, immunostaining is seen in normal murine epidermis. However, following acetone treatment, there is a marked increase in COX-1 expression as well as the appearance of significant COX-2 immunostaining in the basal layer. These data suggest that in human epidermis as well as in human keratinocyte cultures, the expression of COX-2 occurs as a part of normal keratinocyte differentiation whereas in murine epidermis, its constitutive expression is absent, but inducible as previously published.

  10. Endothelin-1 downregulates Mas receptor expression in human cardiomyocytes.

    Science.gov (United States)

    Chen, Zhiheng; Tang, Yamei; Yang, Zuocheng; Liu, Shaojun; Liu, Yong; Li, Yan; He, Wei

    2013-09-01

    Endothelin-1 (ET-1) and the renin-angiotensin system (RAS) are involved in the pathogenesis of cardiac dysfunction. The Mas receptor is a functional binding site for angiotensin (Ang)‑(1-7), which is now considered a critical component of the RAS and exerts cardioprotective effects. To the best of our knowledge, the present study aimed to examine, for the first time, the effects of ET-1 on Mas expression in cultured human cardiomyocytes. Human cardiomyocytes were treated with ET-1 at different concentrations (1, 5, 10, 20 and 30 nM) for varied time periods (0.5, 1.5, 3, 4.5 or 6 h) with or without the transcription inhibitor actinomycin D, endothelin A (ETA) receptor blocker BQ123 and ETB receptor blocker BQ788, or different kinase inhibitors. ET-1 decreased the Mas mRNA level in a statistically significant dose- and time-dependent manner within 4.5 h, which was reflected in the dose-dependent downregulation of Mas promoter activity, Mas protein levels and Ang-(1-7) binding on the cell membrane. Actinomycin D (1 mg/ml), BQ123 (1 µM), p38 mitogen-activated protein kinase (MAPK) siRNA and inhibitor PD169316 (25 µM), completely eliminated the inhibitory effects of ET-1 on Mas expression in human cardiomyocytes. In conclusion, the present study demonstrated that ET-1 downregulates Mas expression at the transcription level in human cardiomyocytes via the ETA receptor by a p38 MAPK‑dependent mechanism. This study provides novel insights into the function of ET-1 and the Ang‑(1-7)/Mas axis in cardiac pathophysiology.

  11. Transient Expression of Tetrameric Recombinant Human Butyrylcholinesterase in Nicotiana benthamiana

    Science.gov (United States)

    Alkanaimsh, Salem; Karuppanan, Kalimuthu; Guerrero, Andrés; Tu, Aye M.; Hashimoto, Bryce; Hwang, Min Sook; Phu, My L.; Arzola, Lucas; Lebrilla, Carlito B.; Dandekar, Abhaya M.; Falk, Bryce W.; Nandi, Somen; Rodriguez, Raymond L.; McDonald, Karen A.

    2016-01-01

    To optimize the expression, extraction and purification of plant-derived tetrameric recombinant human butyrylcholinesterase (prBChE), we describe the development and use of plant viral amplicon-based gene expression system; Tobacco Mosaic Virus (TMV) RNA-based overexpression vector (TRBO) to express enzymatically active FLAG-tagged plant made recombinant butyrylcholinesterase (rBChE) in Nicotiana benthamiana leaves using transient agroinfiltration. Two gene expression cassettes were designed to express the recombinant protein in either the ER or to the apoplastic compartment. Leaf homogenization was used to isolate ER-retained recombinant butyrylcholinesterase (prBChE-ER) while apoplast-targeted rBChE was isolated by either leaf homogenization (prBChE) or vacuum-extraction of apoplastic wash fluid (prBChE-AWF). rBChE from apoplast wash fluid had a higher specific activity but lower enzyme yield than leaf homogenate. To optimize the isolation and purification of total recombinant protein from leaf homogenates, an acidic extraction buffer was used. The acidic extraction buffer yielded >95% enzymatically active tetrameric rBChE as verified by Coomassie stained and native gel electrophoresis. Furthermore, when compared to human butyrylcholinesterase, the prBChE was found to be similar in terms of tetramerization and enzyme kinetics. The N-linked glycan profile of purified prBChE-ER was found to be mostly high mannose structures while the N-linked glycans on prBChE-AWF were primarily complex. The glycan profile of the prBChE leaf homogenates showed a mixture of high mannose, complex and paucimannose type N-glycans. These findings demonstrate the ability of plants to produce rBChE that is enzymatically active and whose oligomeric state is comparable to mammalian butyrylcholinesterase. The process of plant made rBChE tetramerization and strategies for improving its pharmacokinetics properties are also discussed. PMID:27379103

  12. Expression of Recombinant Human Coagulation Factor VII by the Lizard Leishmania Expression System

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    Sina Mirzaahmadi

    2011-01-01

    Full Text Available The variety of recombinant protein expression systems have been developed as a resource of FVII gene expression. In the current study, the authors used a novel protein expression system based on the Iranian Lizard Leishmania, a trypanosomatid protozoan as a host for expression of FVII. Plasmid containing cDNA encoding full-length human FVII was introduced into Lizard Leishmania and positive transfectants were analyzed by SDS-PAGE and Western blot analysis. Furthermore, biological activity of purified protein was detected by PT assay. The recombinant strain harboring a construct was analyzed for expression of FVII at the mRNA and protein level. Purified rFVII was obtained and in order to confirm the purified compound was in fact rFVII. Western blot analysis was carried out. Clotting time in PT assay was reduced about 30 seconds with the purified rFVII. In Conclusion, this study has demonstrated, for the first time, that Leishmania cells can be used as an expression system for producing recombinant FVII.

  13. YY1 positively regulates human UBIAD1 expression

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    Funahashi, Nobuaki, E-mail: nfunahashi@ri.ncgm.go.jp [Department of Hygienic Sciences, Kobe Pharmaceutical University, Kobe (Japan); Department of Metabolic Disorder, Diabetes Research Center, Research Institute, National Center for Global Health and Medicine, Tokyo (Japan); Hirota, Yoshihisa [Department of Hygienic Sciences, Kobe Pharmaceutical University, Kobe (Japan); Faculty of Pharmaceutical Sciences, Suzuka University of Medical Science, Suzuka (Japan); Nakagawa, Kimie; Sawada, Natumi; Watanabe, Masato [Department of Hygienic Sciences, Kobe Pharmaceutical University, Kobe (Japan); Suhara, Yoshitomo [Department of Bioscience and Engineering, Shibaura Institute of Technology, Saitama (Japan); Okano, Toshio [Department of Hygienic Sciences, Kobe Pharmaceutical University, Kobe (Japan)

    2015-05-01

    Vitamin K is involved in bone formation and blood coagulation. Natural vitamin K compounds are composed of the plant form phylloquinone (vitamin K{sub 1}) and a series of bacterial menaquionones (MK-n; vitamin K{sub 2}). Menadione (vitamin K{sub 3}) is an artificial vitamin K compound. MK-4 contains 4-isoprenyl as a side group in the 2-methyl-1,4-naphthoquinone common structure and has various bioactivities. UbiA prenyltransferase domain containing 1 (UBIAD1 or TERE1) is the menaquinone-4 biosynthetic enzyme. UBIAD1 transcript expression significantly decreases in patients with prostate carcinoma and overexpressing UBIAD1 inhibits proliferation of a tumour cell line. UBIAD1 mRNA expression is ubiquitous in mouse tissues, and higher UBIAD1 mRNA expression levels are detected in the brain, heart, kidneys and pancreas. Several functions of UBIAD1 have been reported; however, regulation of the human UBIAD1 gene has not been elucidated. Here we report cloning and characterisation of the human UBIAD1 promoter. A 5′ rapid amplification of cDNA ends analysis revealed that the main transcriptional start site was 306 nucleotides upstream of the translation initiation codon. Deletion and mutation analyses revealed the functional importance of the YY1 consensus motif. Electrophoretic gel mobility shift and chromatin immunoprecipitation assays demonstrated that YY1 binds the UBIAD1 promoter in vitro and in vivo. In addition, YY1 small interfering RNA decreased endogenous UBIAD1 mRNA expression and UBIAD1 conversion activity. These results suggest that YY1 up-regulates UBIAD1 expression and UBIAD1 conversion activity through the UBIAD1 promoter. - Highlights: • We cloned the human UBIAD1 promoter. • The functional importance of the YY1 motif was identified in the UBIAD1 promoter. • YY1 binds the UBIAD1 promoter in vitro and in vivo. • Knockdown of YY1 significantly decreased UBIAD1 expression. • YY1 up-regulates UBIAD1 conversion activity through the UBIAD1

  14. Characterization of Leukocyte Formin FMNL1 Expression in Human Tissues

    Science.gov (United States)

    Heuser, Vanina D.; Iljin, Kristiina; Kampf, Caroline; Uhlen, Mathias; Carpén, Olli

    2014-01-01

    Formins are cytoskeleton regulating proteins characterized by a common FH2 structural domain. As key players in the assembly of actin filaments, formins direct dynamic cytoskeletal processes that influence cell shape, movement and adhesion. The large number of formin genes, fifteen in the human, suggests distinct tasks and expression patterns for individual family members, in addition to overlapping functions. Several formins have been associated with invasive cell properties in experimental models, linking them to cancer biology. One example is FMNL1, which is considered to be a leukocyte formin and is known to be overexpressed in lymphomas. Studies on FMNL1 and many other formins have been hampered by a lack of research tools, especially antibodies suitable for staining paraffin-embedded formalin-fixed tissues. Here we characterize, using bioinformatics tools and a validated antibody, the expression pattern of FMNL1 in human tissues and study its subcellular distribution. Our results indicate that FMNL1 expression is not restricted to hematopoietic tissues and that neoexpression of FMNL1 can be seen in epithelial cancer. PMID:24700756

  15. Downregulation of sulfotransferase expression and activity in diseased human livers.

    Science.gov (United States)

    Yalcin, Emine B; More, Vijay; Neira, Karissa L; Lu, Zhenqiang James; Cherrington, Nathan J; Slitt, Angela L; King, Roberta S

    2013-09-01

    Sulfotransferase (SULT) function has been well studied in healthy human subjects by quantifying mRNA and protein expression and determining enzyme activity with probe substrates. However, it is not well known if sulfotransferase activity changes in metabolic and liver disease, such as diabetes, steatosis, or cirrhosis. Sulfotransferases have significant roles in the regulation of hormones and excretion of xenobiotics. In the present study of normal subjects with nonfatty livers and patients with steatosis, diabetic cirrhosis, and alcoholic cirrhosis, we sought to determine SULT1A1, SULT2A1, SULT1E1, and SULT1A3 activity and mRNA and protein expression in human liver tissue. In general, sulfotransferase activity decreased significantly with severity of liver disease from steatosis to cirrhosis. Specifically, SULT1A1 and SULT1A3 activities were lower in disease states relative to nonfatty tissues. Alcoholic cirrhotic tissues further contained lower SULT1A1 and 1A3 activities than those affected by either of the two other disease states. SULT2A1, on the other hand, was only reduced in alcoholic cirrhotic tissues. SULT1E1 was reduced both in diabetic cirrhosis and in alcoholic cirrhosis tissues, relative to nonfatty liver tissues. In conclusion, the reduced levels of sulfotransferase expression and activity in diseased versus nondiseased liver tissue may alter the metabolism and disposition of xenobiotics and affect homeostasis of endobiotic sulfotransferase substrates.

  16. Identification of differently expressed genes in human colorectal adenocarcinoma

    Institute of Scientific and Technical Information of China (English)

    Yao Chen; Yi-Zeng Zhang; Zong-Guang Zhou; Gang Wang; Zeng-Ni Yi

    2006-01-01

    AIM: To investigate the differently expressed genes in human colorectal adenocarcinoma.METHODS: The integrated approach for gene expression profiling that couples suppression subtractive hybridization, high-throughput cDNA array, sequencing,bioinformatics analysis, and reverse transcriptase realtime quantitative polymerase chain reaction (PCR)was carried out. A set of cDNA clones including 1260SSH inserts amplified by PCR was arrayed using robotic printing. The cDNA arrays were hybridized with florescent-labeled probes prepared from RNA of human colorectal adenocarcinoma (HCRAC) and normal colorectal tissues.RESULTS: A total of 86 genes were identified, 16 unknown genes and 70 known genes. The transcription factor Sox9 influencing cell differentiation was downregulated. At the same time, Heat shock protein 10 KDis downregulated and Calmoulin is up-regulated.CONCLUSION: Downregulation of heat shock protein 10 KD lost its inhibition of Ras, and then attenuated the Ras GTPase signaling pathway, increased cell proliferation and inhibited cell apoptosis. Down-regulated transcription factor Sox9 influences cell differentiation and cell-specific gene expression. Down-regulated Sox9 also decreases its binding to calmodulin, accumulates calmodulin as receptor-activated kinase and phosphorylase kinase due to the activation of PhK.

  17. Dogs can discriminate emotional expressions of human faces.

    Science.gov (United States)

    Müller, Corsin A; Schmitt, Kira; Barber, Anjuli L A; Huber, Ludwig

    2015-03-01

    The question of whether animals have emotions and respond to the emotional expressions of others has become a focus of research in the last decade [1-9]. However, to date, no study has convincingly shown that animals discriminate between emotional expressions of heterospecifics, excluding the possibility that they respond to simple cues. Here, we show that dogs use the emotion of a heterospecific as a discriminative cue. After learning to discriminate between happy and angry human faces in 15 picture pairs, whereby for one group only the upper halves of the faces were shown and for the other group only the lower halves of the faces were shown, dogs were tested with four types of probe trials: (1) the same half of the faces as in the training but of novel faces, (2) the other half of the faces used in training, (3) the other half of novel faces, and (4) the left half of the faces used in training. We found that dogs for which the happy faces were rewarded learned the discrimination more quickly than dogs for which the angry faces were rewarded. This would be predicted if the dogs recognized an angry face as an aversive stimulus. Furthermore, the dogs performed significantly above chance level in all four probe conditions and thus transferred the training contingency to novel stimuli that shared with the training set only the emotional expression as a distinguishing feature. We conclude that the dogs used their memories of real emotional human faces to accomplish the discrimination task.

  18. Expression of Obesity Hormone Leptin in Human Colorectal Cancer

    Institute of Scientific and Technical Information of China (English)

    Jin-chun Cong; Xian-wei Dai; Ming-yang Shen; Jun-jiang Wang; Chun-sheng Chen; Hong Zhang; Lei Qiao

    2009-01-01

    Objective: The obesity hormone, leptin, has been found to participate in the development and proliferation of normal and malignant tissues. The aim of this study was to evaluate the role of leptin in human colorectal cancer.Methods: Serum leptin levels were measured via ABC-ELLSA in 30 colorectal cancers and 24 normal controls. Leptin concentration in colorectal cancer was analyzed in terms of selected clinicopathological features and some oncogenes.Results: The mean concentration of leptin was significantly higher for colorectal cancers(3.54±1.46 ng/ml) than normal controls(2.27±0.99 ng/ml), no gender difference was observed in this study. Leptin expression in poorly differentiated tumors was obviously lower than those in moderately and well differentiated tumors. There were no statistically significant correlations between leptin and the serum CEA and CA199 in colorectal cancers (P>0.05), and between leptin and the expressions of K-RAS, P53, APC, DCC genes in tumor tissues (P>0.05).Conclusion: Leptin is overexpressed in human colorectal cancer, which is related to the differentiation degrees of the tumor. There is no correlation between leptin expression and chages of oncogenes in colorectal cancers.

  19. Expression of human soluble TRAIL in Chlamydomonas reinhardtii chloroplast

    Institute of Scientific and Technical Information of China (English)

    YANG Zongqi; LI yinü; CHEN Feng; LI Dong; ZHANG Zhifang; LIU Yanxin; ZHENG Dexian; WANG Yong; SHEN Guifang

    2006-01-01

    Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) induces selectively apoptosis in various tumor cells and virus-infected cells, but rarely in normal cells. A chloroplast expression vector, p64TRAIL, containing the cDNA coding for the soluble TRAIL (sTRAIL), was constructed with clpP-trnL-petB-chlL-rpl23-rpl2 as Chlamydomonas reinhardtii plastid homologous recombinant fragments and spectinomycin-resistant aadA gene as a select marker. The plasmid p64TRAIL was transferred into the chloroplast genome of C. reinhardtii by the biolistic method. Three independently transformed lines were obtained by 100 mg/L spectinomycin selection. PCR amplification, Southern blot analysis of the sTRAIL coding region DNA and cultivation cells in the dark all showed that the exogenous DNA had been integrated into chloroplast genome of C. reinhardtii. Western blot analysis showed that human soluble TRAIL was expressed in C. reinhardtii chloroplast. The densitometric analysis of Western blot indicated that the expressed human sTRAIL protein in the chloroplasts of C. reinhardtii accounted for about 0.43%-0.67% of the total soluble proteins.These experimental results demonstrated the possibility of using transgenic chloroplasts of green alga as bioreactors for production of biopharmaceuticals.

  20. Expression of epidermal growth factor receptors in human brain tumors.

    Science.gov (United States)

    Libermann, T A; Razon, N; Bartal, A D; Yarden, Y; Schlessinger, J; Soreq, H

    1984-02-01

    The expression of receptors for epidermal growth factor (EGF-R) was determined in 29 samples of brain tumors from 22 patients. Primary gliogenous tumors, of various degrees of cancer, five meningiomas, and two neuroblastomas were examined. Tissue samples were frozen in liquid nitrogen immediately after the operation and stored at -70 degrees until use. Cerebral tissue samples from 11 patients who died from diseases not related to the central nervous system served as controls. Immunoprecipitation of functional EGF-R-kinase complexes revealed high levels of EGF-R in all of the brain tumors of nonneuronal origin that were examined. The level of EGF-R varied between tumors from different patients and also between specimens prelevated from different areas of the same tumor. In contrast, the levels of EGF-R from control specimens were invariably low. The biochemical properties of EGF-R in brain tumor specimens were found to be indistinguishable from those of the well-characterized EGF-R from the A-431 cell line, derived from human epidermoid carcinomas. Human brain EGF-R displays a molecular weight of 170,000 by polyacrylamide-sodium dodecyl sulfate gel electrophoresis. It is phosphorylated mainly in tyrosine residues and shows a 2-dimensional phosphopeptide map similar to that obtained with the phosphorylated EGF-R from membranes of A-431 cells. Our observations suggest that induction of EGF-R expression may accompany the malignant transformation of human brain cells of nonneuronal origin.

  1. CCM2 expression during prenatal development and adult human neocortex.

    Science.gov (United States)

    Tanriover, Gamze; Sozen, Berna; Gunel, Murat; Demir, Necdet

    2011-08-01

    Cerebral cavernous malformation (CCM) is one of the most common types of vascular malformations of the central nervous system, affecting nearly one in 200 people. CCM lesions are characterized by grossly dilated vascular channels lined by a single layer of endothelium. Genetic linkage analyses have mapped three CCM loci to CCM1, CCM2 and CCM3. All three causative genes have now been identified allowing new insights into CCM pathophysiology. We focused on the CCM2 protein that might take place in blood vessel formation; we report here the expression patterns of CCM2 in prenatal development and adult human neocortex by means of immunohistochemistry and Western blot analysis. CCM2 was obviously detected in vascular endothelium and neuroglial precursor cells during development and also observed in arterial endothelium, neurons, some of the glial cells in adult neocortex. The expression patterns suggest that it could be one of the arterial markers whether this is a cause or a consequence of an altered vascular identity. CCM2 might play a role during vasculogenesis and angiogenesis during human brain development. Furthermore, with this study, CCM2 have been described for the first time in developing human neocortex.

  2. Muscle Gene Expression Patterns in Human Rotator Cuff Pathology

    Science.gov (United States)

    Choo, Alexander; McCarthy, Meagan; Pichika, Rajeswari; Sato, Eugene J.; Lieber, Richard L.; Schenk, Simon; Lane, John G.; Ward, Samuel R.

    2014-01-01

    Background: Rotator cuff pathology is a common source of shoulder pain with variable etiology and pathoanatomical characteristics. Pathological processes of fatty infiltration, muscle atrophy, and fibrosis have all been invoked as causes for poor outcomes after rotator cuff tear repair. The aims of this study were to measure the expression of key genes associated with adipogenesis, myogenesis, and fibrosis in human rotator cuff muscle after injury and to compare the expression among groups of patients with varied severities of rotator cuff pathology. Methods: Biopsies of the supraspinatus muscle were obtained arthroscopically from twenty-seven patients in the following operative groups: bursitis (n = 10), tendinopathy (n = 7), full-thickness rotator cuff tear (n = 8), and massive rotator cuff tear (n = 2). Quantitative polymerase chain reaction (qPCR) was performed to characterize gene expression pathways involved in myogenesis, adipogenesis, and fibrosis. Results: Patients with a massive tear demonstrated downregulation of the fibrogenic, adipogenic, and myogenic genes, indicating that the muscle was not in a state of active change and may have difficulty responding to stimuli. Patients with a full-thickness tear showed upregulation of fibrotic and adipogenic genes; at the tissue level, these correspond to the pathologies most detrimental to outcomes of surgical repair. Patients with bursitis or tendinopathy still expressed myogenic genes, indicating that the muscle may be attempting to accommodate the mechanical deficiencies induced by the tendon tear. Conclusions: Gene expression in human rotator cuff muscles varied according to tendon injury severity. Patients with bursitis and tendinopathy appeared to be expressing pro-myogenic genes, whereas patients with a full-thickness tear were expressing genes associated with fatty atrophy and fibrosis. In contrast, patients with a massive tear appeared to have downregulation of all gene programs except inhibition of

  3. Expression of Neuropeptide Y in Human Pituitary Adenoma

    Institute of Scientific and Technical Information of China (English)

    Laizhao Chen; Jingjian Ma; Anchao Zheng; Honggang Zheng

    2006-01-01

    OBJECTIVE Neuropeptid e Y (NPY) acts as a neuroendocrine modulator in the anterior pituitary, and NPY mRNA and NPY-immunoreactivity have been detected in normal human anterior pituitaries. However, only a few studies of NPY expression in human pituitary adenomas have been published. Our study was conducted to determine whether or not adenomatous cells express NPY, to investigate the relationship between NPY expression and the subtypes of pituitary adenoma and to explore the clinical significance of NPY.METHODS The study included tissues from 58 patients with pituitary adenomas who underwent surgery because of their clinical diagnosis.Using a highly specific anti-NPY polyclonal antibody, immunohistochemical analysis was performed on the surgically removed pituitary adenomas. Six fresh specimens also were examined using immuno-electron microscopy. NPY was labeled with colloidal gold in order to study the distribution of NPY at the subcellular level.RESULTS The NPY expression level was significantly different among subgroups of pituitary adenomas (P<0.05). NPY was immuno-detected in 58.6% of all adenomas, in 91.7% of gonadotrophic adenomas and in 14.3% of prolactinomas. NPY expression was slightly lower in invasive pituitary adenomas compared to noninvasive adenomas, but the difference was not significant (t=1.81, P>0.05). Of particular interest was the finding that vascular endothelial cells showed positive NPY expression in some pituitary adenomas. Parts of strongly positive tumor cells were seen in channels formed without endothelial cells, but which contained some red blood cells in a formation similar to so-called vasculogenic mimicry. Immuno-electron microscopy demonstrated that 4 of the 6 fresh specimens displayed positive NPY staining with a high density of gold particles located mainly in the secretory granulas. In addition, gold particles were sparsely detected in the rough endoplasmic reticulum and cell matrix.CONCLUSION NPY exists in pituitary adenomas

  4. Developmental gene expression profiles of the human pathogen Schistosoma japonicum

    Science.gov (United States)

    Gobert, Geoffrey N; Moertel, Luke; Brindley, Paul J; McManus, Donald P

    2009-01-01

    Background The schistosome blood flukes are complex trematodes and cause a chronic parasitic disease of significant public health importance worldwide, schistosomiasis. Their life cycle is characterised by distinct parasitic and free-living phases involving mammalian and snail hosts and freshwater. Microarray analysis was used to profile developmental gene expression in the Asian species, Schistosoma japonicum. Total RNAs were isolated from the three distinct environmental phases of the lifecycle – aquatic/snail (eggs, miracidia, sporocysts, cercariae), juvenile (lung schistosomula and paired but pre-egg laying adults) and adult (paired, mature males and egg-producing females, both examined separately). Advanced analyses including ANOVA, principal component analysis, and hierarchal clustering provided a global synopsis of gene expression relationships among the different developmental stages of the schistosome parasite. Results Gene expression profiles were linked to the major environmental settings through which the developmental stages of the fluke have to adapt during the course of its life cycle. Gene ontologies of the differentially expressed genes revealed a wide range of functions and processes. In addition, stage-specific, differentially expressed genes were identified that were involved in numerous biological pathways and functions including calcium signalling, sphingolipid metabolism and parasite defence. Conclusion The findings provide a comprehensive database of gene expression in an important human pathogen, including transcriptional changes in genes involved in evasion of the host immune response, nutrient acquisition, energy production, calcium signalling, sphingolipid metabolism, egg production and tegumental function during development. This resource should help facilitate the identification and prioritization of new anti-schistosome drug and vaccine targets for the control of schistosomiasis. PMID:19320991

  5. Developmental gene expression profiles of the human pathogen Schistosoma japonicum

    Directory of Open Access Journals (Sweden)

    McManus Donald P

    2009-03-01

    Full Text Available Abstract Background The schistosome blood flukes are complex trematodes and cause a chronic parasitic disease of significant public health importance worldwide, schistosomiasis. Their life cycle is characterised by distinct parasitic and free-living phases involving mammalian and snail hosts and freshwater. Microarray analysis was used to profile developmental gene expression in the Asian species, Schistosoma japonicum. Total RNAs were isolated from the three distinct environmental phases of the lifecycle – aquatic/snail (eggs, miracidia, sporocysts, cercariae, juvenile (lung schistosomula and paired but pre-egg laying adults and adult (paired, mature males and egg-producing females, both examined separately. Advanced analyses including ANOVA, principal component analysis, and hierarchal clustering provided a global synopsis of gene expression relationships among the different developmental stages of the schistosome parasite. Results Gene expression profiles were linked to the major environmental settings through which the developmental stages of the fluke have to adapt during the course of its life cycle. Gene ontologies of the differentially expressed genes revealed a wide range of functions and processes. In addition, stage-specific, differentially expressed genes were identified that were involved in numerous biological pathways and functions including calcium signalling, sphingolipid metabolism and parasite defence. Conclusion The findings provide a comprehensive database of gene expression in an important human pathogen, including transcriptional changes in genes involved in evasion of the host immune response, nutrient acquisition, energy production, calcium signalling, sphingolipid metabolism, egg production and tegumental function during development. This resource should help facilitate the identification and prioritization of new anti-schistosome drug and vaccine targets for the control of schistosomiasis.

  6. Clinicopathological significance of PTPN12 expression in human breast cancer

    Energy Technology Data Exchange (ETDEWEB)

    Yuan, Xunyi [Breast Disease Diagnosis and Treatment Centre, Affiliated Hospital of Medical College, Qingdao University, Qingdao Shandong Province (China); Yuan, Zhentao [Department of Anesthesiology, Shengli Oilfield Central Hospital, Dongying Shandong Province (China); Jiang, Dandan; Li, Funian [Breast Disease Diagnosis and Treatment Centre, Affiliated Hospital of Medical College, Qingdao University, Qingdao Shandong Province (China)

    2012-10-15

    Protein tyrosine phosphatase non-receptor type 12 (PTPN12) is a recently identified tumor suppressor gene (TSG) that is frequently compromised in human triple-negative breast cancer. In the present study, we investigated the expression of PTPN12 protein by patients with breast cancer in a Chinese population and the relationship between PTPN12 expression levels and patient clinicopathological features and prognosis. Additionally, we explored the underlying down-regulation mechanism from the perspective of an epigenetic alteration. We examined PTPN12 mRNA expression in five breast cancer cell lines using semi-quantitative reverse-transcription PCR, and detected PTPN12 protein expression using immunohistochemistry in 150 primary invasive breast cancer cases and paired adjacent non-tumor tissues. Methylation-specific PCR was performed to analyze the promoter CpG island methylation status of PTPN12. PTPN12 was significantly down-regulated in breast cancer cases (48/150) compared to adjacent noncancerous tissues (17/150; P < 0.05). Furthermore, low expression of PTPN12 showed a significant positive correlation with tumor size (P = 0.047), lymph node metastasis (P = 0.001), distant metastasis (P = 0.009), histological grade (P = 0.012), and survival time (P = 0.019). Additionally, promoter CpG island hypermethylation occurs more frequently in breast cancer cases and breast cancer cell lines with low PTPN12 expression. Our findings suggest that PTPN12 is potentially a methylation-silenced TSG for breast cancer that may play an important role in breast carcinogenesis and could potentially serve as an independent prognostic factor for invasive breast cancer patients.

  7. LIN28A expression reduces sickling of cultured human erythrocytes.

    Directory of Open Access Journals (Sweden)

    Jaira F de Vasconcellos

    Full Text Available Induction of fetal hemoglobin (HbF has therapeutic importance for patients with sickle cell disease (SCD and the beta-thalassemias. It was recently reported that increased expression of LIN28 proteins or decreased expression of its target let-7 miRNAs enhances HbF levels in cultured primary human erythroblasts from adult healthy donors. Here LIN28A effects were studied further using erythrocytes cultured from peripheral blood progenitor cells of pediatric subjects with SCD. Transgenic expression of LIN28A was accomplished by lentiviral transduction in CD34(+ sickle cells cultivated ex vivo in serum-free medium. LIN28A over-expression (LIN28A-OE increased HbF, reduced beta (sickle-globin, and strongly suppressed all members of the let-7 family of miRNAs. LIN28A-OE did not affect erythroblast differentiation or prevent enucleation, but it significantly reduced or ameliorated the sickling morphologies of the enucleated erythrocytes.

  8. Expression of Formyl-peptide Receptors in Human Lung Carcinoma.

    Science.gov (United States)

    Cattaneo, Fabio; Guerra, Germano; Parisi, Melania; Lucariello, Angela; De Luca, Antonio; De Rosa, Nicolina; Mazzarella, Gennaro; Bianco, Andrea; Ammendola, Rosario

    2015-05-01

    Formyl-peptide receptors (FPRs) are expressed in several tissues and cell types. The identification of markers involved in cell growth may further allow for molecular profiling of lung cancer. We investigated the possible role of FPRs as molecular markers in several types of lung carcinomas which is the main cause of cancer death worldwide. Tumor tissue samples were collected from six patients affected by lung cancer. Biopsies were analyzed for expression of FPR isoforms both in tumoral and peritumoral tissue by real-time polymerase chain reaction (PCR), western blot and immunofluorescence. Real-time PCR, western blot and immunofluorescence analyses showed that FPR expression is lower in types of human lung cancer tissues when compared to the surrounding peritumoral tissues. The study of the mechanistic basis for the control of FPR expression in normal peritumoral versus tumoral tissues could provide the basis for new diagnostic and therapeutic interventions. Copyright© 2015 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.

  9. Progesterone Upregulates Gene Expression in Normal Human Thyroid Follicular Cells

    Directory of Open Access Journals (Sweden)

    Ana Paula Santin Bertoni

    2015-01-01

    Full Text Available Thyroid cancer and thyroid nodules are more prevalent in women than men, so female sex hormones may have an etiological role in these conditions. There are no data about direct effects of progesterone on thyroid cells, so the aim of the present study was to evaluate progesterone effects in the sodium-iodide symporter NIS, thyroglobulin TG, thyroperoxidase TPO, and KI-67 genes expression, in normal thyroid follicular cells, derived from human tissue. NIS, TG, TPO, and KI-67 mRNA expression increased significantly after TSH 20 μUI/mL, respectively: 2.08 times, P<0.0001; 2.39 times, P=0.01; 1.58 times, P=0.0003; and 1.87 times, P<0.0001. In thyroid cells treated with 20 μUI/mL TSH plus 10 nM progesterone, RNA expression of NIS, TG, and KI-67 genes increased, respectively: 1.78 times, P<0.0001; 1.75 times, P=0.037; and 1.95 times, P<0.0001, and TPO mRNA expression also increased, though not significantly (1.77 times, P=0.069. These effects were abolished by mifepristone, an antagonist of progesterone receptor, suggesting that genes involved in thyroid cell function and proliferation are upregulated by progesterone. This work provides evidence that progesterone has a direct effect on thyroid cells, upregulating genes involved in thyroid function and growth.

  10. Correlation between Gene Expression and Osteoarthritis Progression in Human

    Directory of Open Access Journals (Sweden)

    Leilei Zhong

    2016-07-01

    Full Text Available Osteoarthritis (OA is a multifactorial disease characterized by gradual degradation of joint cartilage. This study aimed to quantify major pathogenetic factors during OA progression in human cartilage. Cartilage specimens were isolated from OA patients and scored 0–5 according to the Osteoarthritis Research Society International (OARSI guidelines. Protein and gene expressions were measured by immunohistochemistry and qPCR, respectively. Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL assays were used to detect apoptotic cells. Cartilage degeneration in OA is a gradual progress accompanied with gradual loss of collagen type II and a gradual decrease in mRNA expression of SOX9, ACAN and COL2A1. Expression of WNT antagonists DKK1 and FRZB was lost, while hypertrophic markers (RUNX2, COL10A1 and IHH increased during OA progression. Moreover, DKK1 and FRZB negatively correlated with OA grading, while RUNX2 and IHH showed a significantly positive correlation with OA grading. The number of apoptotic cells was increased with the severity of OA. Taken together, our results suggested that genetic profiling of the gene expression could be used as markers for staging OA at the molecular level. This helps to understand the molecular pathology of OA and may lead to the development of therapies based on OA stage.

  11. Expression of epidermal growth factor receptors in human endometrial carcinoma

    DEFF Research Database (Denmark)

    Nyholm, H C; Nielsen, Anette Lynge; Ottesen, B

    1993-01-01

    Little data exist on the expression of epidermal growth factor receptors (EGF-Rs) in human endometrial cancer. EGF-R status was studied in 65 patients with endometrial carcinomas and in 26 women with nonmalignant postmenopausal endometria, either inactive/atrophic endometrium or adenomatous...... hyperplasia. EGF-R was identified on frozen tissue sections by means of an indirect immunoperoxidase technique with a monoclonal antibody against the external domain of the EGF-R. Seventy-one percent of the carcinomas expressed positive EGF-R immunoreactivity. In general, staining was most prominent....../inactive endometria and seven of 13 (54%) endometria with adenomatous hyperplasia were EGF-R positive, with an immunostaining pattern rather similar to that of the carcinomas....

  12. Exercise-induced metallothionein expression in human skeletal muscle fibres

    DEFF Research Database (Denmark)

    Penkowa, Milena; Keller, Pernille; Keller, Charlotte;

    2005-01-01

    in both type I and II muscle fibres. This is the first report demonstrating that MT-I + II are significantly induced in human skeletal muscle fibres following exercise. As MT-I + II are antioxidant factors that protect various tissues during pathological conditions, the MT-I + II increases post exercise......Exercise induces free oxygen radicals that cause oxidative stress, and metallothioneins (MTs) are increased in states of oxidative stress and possess anti-apoptotic effects. We therefore studied expression of the antioxidant factors metallothionein I and II (MT-I + II) in muscle biopsies obtained...... in response to 3 h of bicycle exercise performed by healthy men and in resting controls. Both MT-I + II proteins and MT-II mRNA expression increased significantly in both type I and II muscle fibres after exercise. Moreover, 24 h after exercise the levels of MT-II mRNA and MT-I + II proteins were still highly...

  13. DMPD: LPS induction of gene expression in human monocytes. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 11257452 LPS induction of gene expression in human monocytes. Guha M, Mackman N. Ce...ll Signal. 2001 Feb;13(2):85-94. (.png) (.svg) (.html) (.csml) Show LPS induction of gene expression in human... monocytes. PubmedID 11257452 Title LPS induction of gene expression in human monocytes. Authors Guha M, Ma

  14. Expression of the human sodium/iodide symporter (hNIS) in xenotransplanted human thyroid carcinoma

    NARCIS (Netherlands)

    Smit, J.W.A.; Schröder - van der Elst, J.P.; Karperien, M.; Que, I.; Romijn, J.A.; Heide, van der D.

    2001-01-01

    The uptake of iodide in thyroid epithelial cells is mediated by the sodium/iodide symporter (NIS). The uptake of iodide is of vital importance for thyroid physiology and is a prerequisite for radioiodine therapy in thyroid cancer. Loss of iodide uptake due to diminished expression of the human NIS (

  15. Efficient expression and purification of biologically active human cystatin proteins.

    Science.gov (United States)

    Chauhan, Sakshi; Tomar, Raghuvir S

    2016-02-01

    Cystatins are reversible cysteine protease inhibitor proteins. They are known to play important roles in controlling cathepsins, neurodegenerative disease, and in immune system regulation. Production of recombinant cystatin proteins is important for biochemical and function characterization. In this study, we cloned and expressed human stefin A, stefin B and cystatin C in Escherichia coli. Human stefin A, stefin B and cystatin C were purified from soluble fraction. For cystatin C, we used various chaperone plasmids to make cystatin C soluble, as it is reported to localize in inclusion bodies. Trigger factor, GroES-GroEL, DnaK-DnaJ-GrpE chaperones lead to the presence of cystatin C in the soluble fraction. Immobilized metal affinity chromatography, glutathione sepharose and anion exchange chromatography techniques were employed for efficient purification of these proteins. Their biological activities were tested by inhibition assays against cathepsin L and H3 protease.

  16. The antibody preparation and expression of human Pescadillo

    Institute of Scientific and Technical Information of China (English)

    ZHANG Hao; NING Kang; ZHU JianHua; LI JieZhi; HUANG CuiFen; LIU AiJun; YE QiNong; LI JiePing; WANG XiaoHui; SUN Yan; YUAN Bin; YANG ZhiHong; JIANG YanChao; ZENG Min; DING LiHua

    2007-01-01

    To explore the biological roles of human Pescadillo and investigate its potential effect on tumorigene sis, the eDNA of Pescadillo was fused with that of GST. After purification and elution, the purified GST-Pescadillo fusion protein was obtained, and the antibody against the fusion protein was generated.Endogenous Pescadillo protein was observed to be remarkably induced by estrogen. It was mainly distributed in the tissues such as breast, ovary and intestine, all of which contain proliferating cells,and was also detected in many cell lines of human cancer: renal carcinoma, hepatoma, ovarian cancer,colon carcinoma, and breast cancer. The expression level of Pescadillo was increased significantly in breast cancer tissues compared with their paired margin tissues. Taken together, these data suggest that Pescadillo may play important roles in the initiation and development of cancer and may be a potential target in cancer diagnosis and therapy.

  17. Astaxanthin reduces matrix metalloproteinase expression in human chondrocytes.

    Science.gov (United States)

    Chen, Wei-Ping; Xiong, Yan; Shi, Yong-Xiang; Hu, Peng-Fei; Bao, Jia-Peng; Wu, Li-Dong

    2014-03-01

    Astaxanthin is a red carotenoid pigment which exerts multiple biological activities. However, little is known about the effects of astaxanthin on matrix metalloproteinases (MMPs) in OA. The present study investigated the effects of astaxanthin on MMPs in human chondrocytes. Human chondrocytes were pretreated with astaxanthin at 1, 10 or 50μM, then, cells were stimulated with IL-1β (10ng/ml) for 24h. MMP-1, MMP-3 and MMP-13 were observed. We found that astaxanthin reduced the expression of MMP-1, MMP-3 and MMP-13 as well as the phosphorylation of two mitogen-activated protein kinases (MAPK) (p38 and ERK1/2) in IL-1β-stimulated chondrocytes. Astaxanthin also blocked the IκB-α degradation. These results suggest that astaxanthin may be beneficial in the treatment of OA.

  18. The antibody preparation and expression of human Pescadillo

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    To explore the biological roles of human Pescadillo and investigate its potential effect on tumorigene- sis, the cDNA of Pescadillo was fused with that of GST. After purification and elution, the purified GST-Pescadillo fusion protein was obtained, and the antibody against the fusion protein was generated. Endogenous Pescadillo protein was observed to be remarkably induced by estrogen. It was mainly distributed in the tissues such as breast, ovary and intestine, all of which contain proliferating cells, and was also detected in many cell lines of human cancer: renal carcinoma, hepatoma, ovarian cancer, colon carcinoma, and breast cancer. The expression level of Pescadillo was increased significantly in breast cancer tissues compared with their paired margin tissues. Taken together, these data suggest that Pescadillo may play important roles in the initiation and development of cancer and may be a po- tential target in cancer diagnosis and therapy.

  19. Genome-Wide Associations of Gene Expression Variation in Humans.

    Directory of Open Access Journals (Sweden)

    2005-12-01

    Full Text Available The exploration of quantitative variation in human populations has become one of the major priorities for medical genetics. The successful identification of variants that contribute to complex traits is highly dependent on reliable assays and genetic maps. We have performed a genome-wide quantitative trait analysis of 630 genes in 60 unrelated Utah residents with ancestry from Northern and Western Europe using the publicly available phase I data of the International HapMap project. The genes are located in regions of the human genome with elevated functional annotation and disease interest including the ENCODE regions spanning 1% of the genome, Chromosome 21 and Chromosome 20q12-13.2. We apply three different methods of multiple test correction, including Bonferroni, false discovery rate, and permutations. For the 374 expressed genes, we find many regions with statistically significant association of single nucleotide polymorphisms (SNPs with expression variation in lymphoblastoid cell lines after correcting for multiple tests. Based on our analyses, the signal proximal (cis- to the genes of interest is more abundant and more stable than distal and trans across statistical methodologies. Our results suggest that regulatory polymorphism is widespread in the human genome and show that the 5-kb (phase I HapMap has sufficient density to enable linkage disequilibrium mapping in humans. Such studies will significantly enhance our ability to annotate the non-coding part of the genome and interpret functional variation. In addition, we demonstrate that the HapMap cell lines themselves may serve as a useful resource for quantitative measurements at the cellular level.

  20. Genome-wide associations of gene expression variation in humans.

    Directory of Open Access Journals (Sweden)

    Barbara E Stranger

    2005-12-01

    Full Text Available The exploration of quantitative variation in human populations has become one of the major priorities for medical genetics. The successful identification of variants that contribute to complex traits is highly dependent on reliable assays and genetic maps. We have performed a genome-wide quantitative trait analysis of 630 genes in 60 unrelated Utah residents with ancestry from Northern and Western Europe using the publicly available phase I data of the International HapMap project. The genes are located in regions of the human genome with elevated functional annotation and disease interest including the ENCODE regions spanning 1% of the genome, Chromosome 21 and Chromosome 20q12-13.2. We apply three different methods of multiple test correction, including Bonferroni, false discovery rate, and permutations. For the 374 expressed genes, we find many regions with statistically significant association of single nucleotide polymorphisms (SNPs with expression variation in lymphoblastoid cell lines after correcting for multiple tests. Based on our analyses, the signal proximal (cis- to the genes of interest is more abundant and more stable than distal and trans across statistical methodologies. Our results suggest that regulatory polymorphism is widespread in the human genome and show that the 5-kb (phase I HapMap has sufficient density to enable linkage disequilibrium mapping in humans. Such studies will significantly enhance our ability to annotate the non-coding part of the genome and interpret functional variation. In addition, we demonstrate that the HapMap cell lines themselves may serve as a useful resource for quantitative measurements at the cellular level.

  1. Preferential gene expression in quiescent human lung fibroblasts.

    Science.gov (United States)

    Coppock, D L; Kopman, C; Scandalis, S; Gilleran, S

    1993-06-01

    The exit from the proliferative cell cycle into a reversible quiescence (G0) is an active process that is not yet well understood at the molecular level. Investigation of G0-specific gene expression is an important step in studying the mechanism regulating the entrance to quiescence. Using the human embryo lung fibroblast (WI38) as a model system, we have isolated complementary DNA clones that are expressed at a higher level in quiescent cells than in logarithmically growing cells. We have identified complementary DNAs from eight genes including collagen alpha 1(VI), collagen alpha 1(III), decorin, complement C1r, collagen alpha 1(I), collagen alpha 2(I), and two novel genes, Q6 and Q10. We have named this class of quiescence-inducible genes quiescins. Expression of these genes was induced just as proliferation slowed, as indicated by the level of histone H2B mRNA, [3H]-thymidine incorporation, and cell number. The level of expression of the novel genes, Q6 and Q10, increased at the same time as the other genes. Q6 has two mRNAs of 3 and 4 kb, whereas Q10 mRNA is about 1.0 kb. The expression of the quiescins was not induced by blocking the cell cycle in S phase with aphidicolin or in G1 with lovastatin. However, the genes were highly induced by trypsinization or scraping of the cells during logarithmic growth. This induction was not blocked by inhibitors of RNA synthesis. The expression of decorin and Q6 was very low in SV40-transformed cells (VA13) either in logarithmic growth or at high density, whereas the gene Q10 was expressed more highly in VA13 than in WI38 cells. The finding that expression of some components of the extracellular matrix is induced as cells enter G0 suggests that they may have a role in both the induction and the maintenance of the quiescent state. The quiescins will serve as molecular markers for the investigation of mechanisms that regulate the onset of quiescence.

  2. Regulation of human genome expression and RNA splicing by human papillomavirus 16 E2 protein.

    Science.gov (United States)

    Gauson, Elaine J; Windle, Brad; Donaldson, Mary M; Caffarel, Maria M; Dornan, Edward S; Coleman, Nicholas; Herzyk, Pawel; Henderson, Scott C; Wang, Xu; Morgan, Iain M

    2014-11-01

    Human papillomavirus 16 (HPV16) is causative in human cancer. The E2 protein regulates transcription from and replication of the viral genome; the role of E2 in regulating the host genome has been less well studied. We have expressed HPV16 E2 (E2) stably in U2OS cells; these cells tolerate E2 expression well and gene expression analysis identified 74 genes showing differential expression specific to E2. Analysis of published gene expression data sets during cervical cancer progression identified 20 of the genes as being altered in a similar direction as the E2 specific genes. In addition, E2 altered the splicing of many genes implicated in cancer and cell motility. The E2 expressing cells showed no alteration in cell growth but were altered in cell motility, consistent with the E2 induced altered splicing predicted to affect this cellular function. The results present a model system for investigating E2 regulation of the host genome.

  3. Selenoprotein P mRNA expression in human hepatic tissues

    Institute of Scientific and Technical Information of China (English)

    Chun-Li Li; Ke-Jun Nan; Tao Tian; Chen-Guang Sui; Yan-Fang Liu

    2007-01-01

    AIM: To investigate the expression of Selenoprotein P mRNA (SePmRNA) in tissues of normal liver, liver cirrhosis and hepatocellular carcinoma (HCC), and its relationship with HCC occurrence and development.METHODS: The expression of SePmRNA in tissues of normal liver, liver cirrhosis and HCC were detected by in situ hybridization using a cDNA probe.RESULTS: The enzyme digesting products of pBluescript-Human Selenoprotein P were evaluated by electrophoresis.The positive expression of SePmRNA was found in the tissues of normal liver,liver cirrhosis and HCC.The expression of SeP mRNA was found in hepatic interstitial substance,especially in endothelial cells and lymphocytes of vasculature.The positive rate of SePmRNA in normal liver tissue was 84.6% (11/13) and the positive signals appeared in the nucleus and cytoplasm,mostly in the nucleolus,and the staining granules were larger in the nucleolus and around the nucleus.The positive rate of SePmRNA in liver cirrhosis tissue was 45.O% (9/20) and the positive signals were mainly in the nucleolus and cytoplasm,being less around the nucleus and inner nucleus than that in normal liver tissue. The positive rate of SePmRNA in HCC tissue was 30.0% (9/30) and the positive signals were in the cytoplasm, but less in the nucleus.CONCLUSION: SePmRNA expression in the tissues of normal liver and HCC is significantly different (84.6% vs 30.0%, P = 0.003), suggesting that SeP might play a role in the occurrence and development of HCC.

  4. APOBEC3B expression in human leptomeninges and meningiomas

    Science.gov (United States)

    Johnson, Mahlon D.; Reeder, Jay E.; O'Connell, Mary

    2016-01-01

    Nucleic acid-editing enzymes of the apolipoprotein B mRNA-editing enzyme (APOBEC) family have been associated with somatic mutation in cancer. However, the role of APOBEC catalytic subunit 3B (APOBEC3B) editing in the pathogenesis of base substitutions in meningiomas is unknown. In the present study, the expression of APOBEC3B was examined by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and western blot analyses in five fetal and one adult human leptomeninges and 38 meningiomas. Genomic DNA was sequenced using the Illumina Tru-Seq Cancer Panel. Three meningioma primary cultures were also established and treated with cerebrospinal fluid form patients without neurological disease or platelet-derived growth factor-BB (PDGF-BB), prior to evaluation of APOBEC3B expression. By western blotting, APOBEC3B was revealed to be present in 100% of the fetal leptomeninges, and in 88% of World Health Organization grade I, 100% of grade II and 83% of grade III meningiomas tested, but was not different between grades. RT-qPCR revealed no difference in the mRNA expression of APOBEC3B between grades. Sequencing revealed no elevated levels of the C>T mutations that are characteristic of APOBEC3B editing of genomic DNA. Treatment with cerebrospinal fluid and PDGF-BB had no effect on APOBEC3B protein expression in the leptomeningeal or meningioma cells. These findings suggest that the mutations associated with increased APOBEC3B expression may not be central to the pathogenesis of meningiomas. PMID:28101245

  5. Connective Tissue Growth Factor Expression in Human Bronchial Epithelial Cells

    Institute of Scientific and Technical Information of China (English)

    Amrita DOSANJH

    2006-01-01

    Connective tissue growth factor (CTGF) is a cysteine-rich protein that promotes extracellular matrix deposition. CTGF is selectively induced by transforming growth factor β and des-Arg kallidin in lung fibroblasts and increases steady-state mRNA levels of α type I collagen, 5α-integrin and fibronectin in fibroblasts. Bronchial epithelial cells have been proposed to functionally interact with lung fibroblasts. We therefore investigated if bronchial epithelial cells are able to synthesize CTGF. Human bronchial epithelial cells were grown to subconfluence in standard growth media. Proliferating cells grown in small airway growth media were harvested following starvation for up to 24 h. Expression of CTGF transcripts was measured by PCR. Immunocytochemistry was also completed using a commercially available antibody.The cells expressed readily detectable CTGF transcripts. Starvation of these cells resulted in a quantitative decline of CTGF transcripts. Direct sequencing of the PCR product identified human CTGF. Immunocytochemistry confirmed intracellular CTGF in the cells and none in negative control cells. We conclude that bronchial epithelial cells could be a novel source of CTGF. Bronchial epithelial cell-derived CTGF could thus directly influence the deposition of collagen in certain fibrotic lung diseases.

  6. Bordetella pertussis modulates human macrophage defense gene expression.

    Science.gov (United States)

    Valdez, Hugo Alberto; Oviedo, Juan Marcos; Gorgojo, Juan Pablo; Lamberti, Yanina; Rodriguez, Maria Eugenia

    2016-08-01

    Bordetella pertussis, the etiological agent of whooping cough, still causes outbreaks. We recently found evidence that B. pertussis can survive and even replicate inside human macrophages, indicating that this host cell might serve as a niche for persistence. In this work, we examined the interaction of B. pertussis with a human monocyte cell line (THP-1) that differentiates into macrophages in culture in order to investigate the host cell response to the infection and the mechanisms that promote that intracellular survival. To that end, we investigated the expression profile of a selected number of genes involved in cellular bactericidal activity and the inflammatory response during the early and late phases of infection. The bactericidal and inflammatory response of infected macrophages was progressively downregulated, while the number of THP-1 cells heavily loaded with live bacteria increased over time postinfection. Two of the main toxins of B. pertussis, pertussis toxin (Ptx) and adenylate cyclase (CyaA), were found to be involved in manipulating the host cell response. Therefore, failure to express either toxin proved detrimental to the development of intracellular infections by those bacteria. Taken together, these results support the relevance of host defense gene manipulation to the outcome of the interaction between B. pertussis and macrophages.

  7. Adult human liver mesenchymal progenitor cells express phenylalanine hydroxylase.

    Science.gov (United States)

    Baruteau, Julien; Nyabi, Omar; Najimi, Mustapha; Fauvart, Maarten; Sokal, Etienne

    2014-09-01

    Phenylketonuria (PKU) is one of the most prevalent inherited metabolic diseases and is accountable for a severe encephalopathy by progressive intoxication of the brain by phenylalanine. This results from an ineffective L-phenylalanine hydroxylase enzyme (PAH) due to a mutated phenylalanine hydroxylase (PAH) gene. Neonatal screening programs allow an early dietetic treatment with restrictive phenylalanine intake. This diet prevents most of the neuropsychological disabilities but remains challenging for lifelong compliance. Adult-derived human liver progenitor cells (ADHLPC) are a pool of precursors that can differentiate into hepatocytes. We aim to study PAH expression and PAH activity in a differenciated ADHLPC. ADHLPC were isolated from human hepatocyte primary culture of two different donors and differenciated under specific culture conditions. We demonstrated the high expression of PAH and a large increase of PAH activity in differenciated LPC. The age of the donor, the cellular viability after liver digestion and cryopreservation affects PAH activity. ADHLPC might therefore be considered as a suitable source for cell therapy in PKU.

  8. Gene expression profiles of single human mature oocytes in relation to age

    DEFF Research Database (Denmark)

    Grøndahl, M L; Andersen, Claus Yding; Bogstad, J

    2010-01-01

    The development competence of human oocytes declines with increasing age. The objective of this study was to investigate the effect of age on gene expression profile in mature human oocytes.......The development competence of human oocytes declines with increasing age. The objective of this study was to investigate the effect of age on gene expression profile in mature human oocytes....

  9. Expression and biochemical characterization of recombinant human epididymis protein 4.

    Science.gov (United States)

    Hua, Ling; Liu, Yunhui; Zhen, Shuai; Wan, Deyou; Cao, Jiyue; Gao, Xin

    2014-10-01

    Whey acidic proteins (WAP) belong to a large gene family of antibacterial peptides that perform critical immune system functions. The function of human epididymis protein 4 (HE4), a 124-amino acid long polypeptide that has two whey acidic protein four-disulfide core (WFDC) domains, is not well studied. Here, a fusion gene encoding the HE4 protein fused to an IgG1 Fc domain was constructed. The recombinant HE4 protein was expressed as a secretory protein in Pichia pastoris and mammalian HEK293-F cells and was subsequently purified. Our data suggested that the HE4 protein produced by these two expression systems bound to both gram-negative and gram-positive bacteria, but demonstrated slightly inhibitory activity towards the growth of Staphylococcus aureus. Moreover, HE4 exhibited proteinase inhibitory activity towards trypsin, elastase, matrix metallopeptidase 9, and the secretory proteinases from Bacillus subtilis. The effects of glycosylation on the biochemical characterization of HE4 were also investigated. LC-ESI-MS glycosylation analysis showed that the high-mannose glycosylated form of HE4 expressed by P. pastoris has lower biological activity when compared to its complex-glycosylated form produced from HEK293-F cells. The implications of this are discussed, which may be provide theoretical basis for its important role in the development of cancer and innate immune system.

  10. Expression of DNA-dependent protein kinase in human granulocytes

    Institute of Scientific and Technical Information of China (English)

    Annahita SALLMYR; Anna MILLER; Aida GABDOULKHAKOVA; Valentina SAFRONOVA; Gunnel HENRIKSSON; Anders BREDBERG

    2004-01-01

    Human polymorphonuclear leukocytes (PMN) have been reported to completely lack of DNA-dependent protein kinase (DNA-PK) which is composed of Ku protein and the catalytic subunit DNA-PKcs, needed for nonhomologous end-joining (NHEJ) of DNA double-strand breaks. Promyelocytic HL-60 cells express a variant form of Ku resulting in enhanced radiation sensitivity. This raises the question if low efficiency of NHEJ, instrumental for the cellular repair of oxidative damage, is a normal characteristic of myeloid differentiation. Here we confirmed the complete lack of DNAPK in P MN protein extracts, and the expression of the truncated Ku86 variant form in HL-60. However, this degradation of DNA-PK was shown to be due to a DNA-PK-degrading protease in PMN and HL-60. In addition, by using a protease-resistant whole cell assay, both Ku86 and DNA-PKcs could be demonstrated in PMN, suggesting the previously reported absence in PMN of DNA-PK to be an artefact. The levels of Ku86 and DNA-PKcs were much reduced in PMN, as compared with that of the lymphocytes, whereas HL-60 displayed a markedly elevated DNA-PK concentration.In conclusion, our findings provide evidence of reduced, not depleted expression of DNA-PK during the mature stages of myeloid differentiation.

  11. Expression and purification of recombinant human hemangiopoietin in Escherichia coli

    Institute of Scientific and Technical Information of China (English)

    Ren Qian; Ma Fengxia; Chen Zhong; Lu Shihong; Han Zhibo; Liu Yongiun; Xu Bin; Zhang Xiangyu; Han Zhongchao

    2008-01-01

    Objective: To express the soluble recombinant hemangiopoietin protein in E. coli BL21(DE3). Methods: Using human fetal live cDNA as a template, a partial cDNA fragment of HAPO coding N-terminal region was subcloned into plasmids pTrc99, pQE60 and pET32c to construct different recombinant prokaryotic expression systems. After selecting,the soluble rhHAPO fusion protein was expressed stably in E. coli BL21 (DE3) by vector pET32c-HAPO and further isolated by nickelnitrilotriacetic acid (NTA) affinity chromatography. After cleavage with enterokinase, the rhHAPO protein was applied to Fast Flow SP sepharose column. Results: The rhHAPO protein had a purity of more than 95% and a good bioactivity based on the cell adhesion assay in ECV304 cells. Conclusion: We have established a protein engineering system to produce rhHAPO which may provide the possibility for clinical application.

  12. Regulated expression of erythropoietin by two human hepatoma cell lines

    Energy Technology Data Exchange (ETDEWEB)

    Goldberg, M.A.; Glass, G.A.; Cunningham, J.M.; Bunn, H.F.

    1987-11-01

    The development of a cell culture system that produces erythropoietin (Epo) in a regulated manner has been the focus of much effort. The authors have screened multiple renal and hepatic cell lines for either constitutive or regulated expression of Epo. Only the human hepatoma cell lines, Hep3B and HepG2, made significant amounts of Epo as measured both by radioimmunoassay and in vitro bioassay (as much as 330 milliunits per 10/sup 6/ cells in 24 hr). The constitutive production of Epo increased dramatically as a function of cell density in both cell lines. At cell densities < 3.3 x 10/sup 5/ cells per cm/sup 2/, there was little constitutive release of Epo in the medium. With Hep3B cells grown at low cell densities, a mean 18-fold increase in Epo expression was seen in response to hypoxia and a 6-fold increase was observed in response to incubation in medium containing 50 ..mu..M cobalt(II) chloride. At similar low cell densities, Epo production in HepG2 cells could be enhanced an average of about 3-fold by stimulation with either hypoxia or cobalt(II) chloride. Upon such stimulation, both cell lines demonstrated markedly elevated levels of Epo mRNA. Hence, both Hep3B and HepG2 cell lines provide an excellent in vitro system in which to study the physiological regulation of Epo expression.

  13. Impact of Statins on Gene Expression in Human Lung Tissues.

    Directory of Open Access Journals (Sweden)

    Jérôme Lane

    Full Text Available Statins are 3-hydroxy-3-methylglutaryl-coenzyme A reductase inhibitors that alter the synthesis of cholesterol. Some studies have shown a significant association of statins with improved respiratory health outcomes of patients with asthma, chronic obstructive pulmonary disease and lung cancer. Here we hypothesize that statins impact gene expression in human lungs and may reveal the pleiotropic effects of statins that are taking place directly in lung tissues. Human lung tissues were obtained from patients who underwent lung resection or transplantation. Gene expression was measured on a custom Affymetrix array in a discovery cohort (n = 408 and two replication sets (n = 341 and 282. Gene expression was evaluated by linear regression between statin users and non-users, adjusting for age, gender, smoking status, and other covariables. The results of each cohort were combined in a meta-analysis and biological pathways were studied using Gene Set Enrichment Analysis. The discovery set included 141 statin users. The lung mRNA expression levels of eighteen and three genes were up-regulated and down-regulated in statin users (FDR < 0.05, respectively. Twelve of the up-regulated genes were replicated in the first replication set, but none in the second (p-value < 0.05. Combining the discovery and replication sets into a meta-analysis improved the significance of the 12 up-regulated genes, which includes genes encoding enzymes and membrane proteins involved in cholesterol biosynthesis. Canonical biological pathways altered by statins in the lung include cholesterol, steroid, and terpenoid backbone biosynthesis. No genes encoding inflammatory, proteases, pro-fibrotic or growth factors were altered by statins, suggesting that the direct effect of statin in the lung do not go beyond its antilipidemic action. Although more studies are needed with specific lung cell types and different classes and doses of statins, the improved health outcomes and survival

  14. Aberrant phenotypes of transgenic mice expressing dimeric human erythropoietin

    Directory of Open Access Journals (Sweden)

    Yun Seong-Jo

    2012-01-01

    Full Text Available Abstract Background Dimeric human erythropoietin (dHuEPO peptides are reported to exhibit significantly higher biological activity than the monomeric form of recombinant EPO. The objective of this study was to produce transgenic (tg mice expressing dHuEPO and to investigate the characteristics of these mice. Methods A dHuEPO-expressing vector under the control of the goat beta-casein promoter, which produced a dimer of human EPO molecules linked by a 2-amino acid peptide linker (Asp-Ile, was constructed and injected into 1-cell fertilized embryos by microinjection. Mice were screened using genomic DNA samples obtained from tail biopsies. Blood samples were obtained by heart puncture using heparinized tubes, and hematologic parameters were assessed. Using the microarray analysis tool, we analyzed differences in gene expression in the spleens of tg and control mice. Results A high rate of spontaneous abortion or death of the offspring was observed in the recipients of dHuEPO embryos. We obtained 3 founder lines (#4, #11, and #47 of tg mice expressing the dHuEPO gene. However, only one founder line showed stable germline integration and transmission, subsequently establishing the only transgenic line (#11. We obtained 2 F1 mice and 3 F2 mice from line #11. The dHuEPO protein could not be obtained because of repeated spontaneous abortions in the tg mice. Tg mice exhibited symptoms such as short lifespan and abnormal blood composition. The red blood cell count, white blood cell count, and hematocrit levels in the tg mice were remarkably higher than those in the control mice. The spleens of the tg mice (F1 and F2 females were 11- and -21-fold larger than those of the control mice. Microarray analysis revealed 2,672 spleen-derived candidate genes; more genes were downregulated than upregulated (849/764. Reverse transcriptase-polymerase chain reaction (RT-PCR and quantitative real-time PCR (qRT-PCR were used for validating the results of the microarray

  15. Cloning, expression and location of RNase9 in human epididymis

    Directory of Open Access Journals (Sweden)

    Lin YQ

    2008-11-01

    Full Text Available Abstract Background Mammalian spermatozoa become fully motile and fertile during transit through the luminal fluid of the epididymis. At least 200 proteins are present in the epididymal lumen, but the potential roles of these luminal proteins in male fertility are unknown. Investigation of the function of these proteins will elucidate the mechanism of sperm maturation, and also provide new drug targets for male contraception. We cloned RNase9 from a human epididymis cDNA library for characterization and analysis of its functions. Findings It was predicted that human RNase9 gene was located on chromosome 14q11.2 and encoded a 205 amino acids protein with a signal peptide of 26 amino acids at the N-terminus. The protein had eight conserved cysteine residues characteristic of the RNase A family members and several potential post-translational modification sites. At the transcriptional level, RNase9 was expressed in a wide variety of tissues, and the expression was higher in men than in boys. RNase9 was localized to the post-equatorial region of the sperms' head. Immunofluorescence staining showed that RNase9 protein was present mostly in the epithelium of the epididymal tubule. Recombinant RNase9 had no ribonuclease activity. In addition, RNase9 had no detectable effect on sperm motility and fertilization as demonstrated by blocking spermatozoa with anti-RNase9 polyclonal serum. Conclusion RNase9 is expressed in a wide variety of tissues. It is located on the post-equatorial region of the sperm head and the epithelium of epididymal tubule. Although RNase9 belongs to the RNase A family, it has no ribonuclease activity.

  16. Human adipose tissue expresses intrinsic circadian rhythm in insulin sensitivity.

    Science.gov (United States)

    Carrasco-Benso, Maria P; Rivero-Gutierrez, Belen; Lopez-Minguez, Jesus; Anzola, Andrea; Diez-Noguera, Antoni; Madrid, Juan A; Lujan, Juan A; Martínez-Augustin, Olga; Scheer, Frank A J L; Garaulet, Marta

    2016-09-01

    In humans, insulin sensitivity varies according to time of day, with decreased values in the evening and at night. Mechanisms responsible for the diurnal variation in insulin sensitivity are unclear. We investigated whether human adipose tissue (AT) expresses intrinsic circadian rhythms in insulin sensitivity that could contribute to this phenomenon. Subcutaneous and visceral AT biopsies were obtained from extremely obese participants (body mass index, 41.8 ± 6.3 kg/m(2); 46 ± 11 y) during gastric-bypass surgery. To assess the rhythm in insulin signaling, AKT phosphorylation was determined every 4 h over 24 h in vitro in response to different insulin concentrations (0, 1, 10, and 100 nM). Data revealed that subcutaneous AT exhibited robust circadian rhythms in insulin signaling (P Insulin sensitivity reached its maximum (acrophase) around noon, being 54% higher than during midnight (P = 0.009). The amplitude of the rhythm was positively correlated with in vivo sleep duration (r = 0.53; P = 0.023) and negatively correlated with in vivo bedtime (r = -0.54; P = 0.020). No circadian rhythms were detected in visceral AT (P = 0.643). Here, we demonstrate the relevance of the time of the day for how sensitive AT is to the effects of insulin. Subcutaneous AT shows an endogenous circadian rhythm in insulin sensitivity that could provide an underlying mechanism for the daily rhythm in systemic insulin sensitivity.-Carrasco-Benso, M. P., Rivero-Gutierrez, B., Lopez-Minguez, J., Anzola, A., Diez-Noguera, A., Madrid, J. A., Lujan, J. A., Martínez-Augustin, O., Scheer, F. A. J. L., Garaulet, M. Human adipose tissue expresses intrinsic circadian rhythm in insulin sensitivity. © FASEB.

  17. Recombinant Functional Human Lactoferrin Expressed in Baculovirus System

    Institute of Scientific and Technical Information of China (English)

    Tao LIU; Yao-Zhou ZHANG; Xiang-Fu WU

    2006-01-01

    Human lactoferrin (hLf) is a multifunctional iron-binding glycoprotein. In this study, we amplified hLfcDNA by reverse transcription-polymerase chain reaction from normal human mammary gland.The nucleotide sequence of the hLf was identical to the known hLf. We constructed a recombinant virus,vBm-hLf, harboring the hLfgene and exploited the BmN cells as host to produce recombinant human lactoferrin(rhLf). It was found that a recombinant protein with a molecular mass of approximately 78 kDa was expressed.Approximately 13.5 μg rhLf was purified from 1-2× 105 BmN cells infected by vBm-hLf and the rhLf proved to be biologically active. This method established in our study will pave the way for efficient production of rhLf for further application of this protein in the future.

  18. Sperm protein 17 is expressed in human nervous system tumours

    Directory of Open Access Journals (Sweden)

    Frezza Eldo E

    2006-01-01

    Full Text Available Abstract Background Human sperm protein 17 (Sp17 is a highly conserved protein that was originally isolated from a rabbit epididymal sperm membrane and testis membrane pellet. It has recently been included in the cancer/testis (CT antigen family, and shown to be expressed in multiple myeloma and ovarian cancer. We investigated its immunolocalisation in specimens of nervous system (NS malignancies, in order to establish its usefulness as a target for tumour-vaccine strategies. Methods The expression of Sp17 was assessed by means of a standardised immunohistochemical procedure [(mAb/antigen MF1/Sp17] in formalin-fixed and paraffin embedded surgical specimens of NS malignancies, including 28 neuroectodermal primary tumours (6 astrocytomas, 16 glioblastoma multiforme, 5 oligodendrogliomas, and 1 ependymoma, 25 meningeal tumours, and five peripheral nerve sheath tumours (4 schwannomas, and 1 neurofibroma,. Results A number of neuroectodermal (21% and meningeal tumours (4% were found heterogeneously immunopositive for Sp17. None of the peripheral nerve sheath tumours was immunopositive for Sp17. The expression pattern was heterogeneous in all of the positive samples, and did not correlate with the degree of malignancy. Conclusion The frequency of expression and non-uniform cell distribution of Sp17 suggest that it cannot be used as a unique immunotherapeutic target in NS cancer. However, our results do show the immunolocalisation of Sp17 in a proportion of NS tumour cells, but not in their non-pathological counterparts. The emerging complex function of Sp17 makes further studies necessary to clarify the link between it and immunopositive cells.

  19. Dissecting cis regulation of gene expression in human metabolic tissues.

    Directory of Open Access Journals (Sweden)

    Radu Dobrin

    Full Text Available Complex diseases such as obesity and type II diabetes can result from a failure in multiple organ systems including the central nervous system and tissues involved in partitioning and disposal of nutrients. Studying the genetics of gene expression in tissues that are involved in the development of these diseases can provide insights into how these tissues interact within the context of disease. Expression quantitative trait locus (eQTL studies identify mRNA expression changes linked to proximal genetic signals (cis eQTLs that have been shown to affect disease. Given the high impact of recent eQTL studies, it is important to understand what role sample size and environment plays in identification of cis eQTLs. Here we show in a genotyped obese human population that the number of cis eQTLs obey precise scaling laws as a function of sample size in three profiled tissues, i.e. omental adipose, subcutaneous adipose and liver. Also, we show that genes (or transcripts with cis eQTL associations detected in a small population are detected at approximately 90% rate in the largest population available for our study, indicating that genes with strong cis acting regulatory elements can be identified with relatively high confidence in smaller populations. However, by increasing the sample size we allow for better detection of weaker and more distantly located cis-regulatory elements. Yet, we determined that the number of tissue specific cis eQTLs saturates in a modestly sized cohort while the number of cis eQTLs common to all tissues fails to reach a maximum value. Understanding the power laws that govern the number and specificity of eQTLs detected in different tissues, will allow a better utilization of genetics of gene expression to inform the molecular mechanism underlying complex disease traits.

  20. Expression cartography of human tissues using self organizing maps

    Science.gov (United States)

    2011-01-01

    Background Parallel high-throughput microarray and sequencing experiments produce vast quantities of multidimensional data which must be arranged and analyzed in a concerted way. One approach to addressing this challenge is the machine learning technique known as self organizing maps (SOMs). SOMs enable a parallel sample- and gene-centered view of genomic data combined with strong visualization and second-level analysis capabilities. The paper aims at bridging the gap between the potency of SOM-machine learning to reduce dimension of high-dimensional data on one hand and practical applications with special emphasis on gene expression analysis on the other hand. Results The method was applied to generate a SOM characterizing the whole genome expression profiles of 67 healthy human tissues selected from ten tissue categories (adipose, endocrine, homeostasis, digestion, exocrine, epithelium, sexual reproduction, muscle, immune system and nervous tissues). SOM mapping reduces the dimension of expression data from ten of thousands of genes to a few thousand metagenes, each representing a minicluster of co-regulated single genes. Tissue-specific and common properties shared between groups of tissues emerge as a handful of localized spots in the tissue maps collecting groups of co-regulated and co-expressed metagenes. The functional context of the spots was discovered using overrepresentation analysis with respect to pre-defined gene sets of known functional impact. We found that tissue related spots typically contain enriched populations of genes related to specific molecular processes in the respective tissue. Analysis techniques normally used at the gene-level such as two-way hierarchical clustering are better represented and provide better signal-to-noise ratios if applied to the metagenes. Metagene-based clustering analyses aggregate the tissues broadly into three clusters containing nervous, immune system and the remaining tissues. Conclusions The SOM technique

  1. Expression cartography of human tissues using self organizing maps

    Directory of Open Access Journals (Sweden)

    Löffler Markus

    2011-07-01

    Full Text Available Abstract Background Parallel high-throughput microarray and sequencing experiments produce vast quantities of multidimensional data which must be arranged and analyzed in a concerted way. One approach to addressing this challenge is the machine learning technique known as self organizing maps (SOMs. SOMs enable a parallel sample- and gene-centered view of genomic data combined with strong visualization and second-level analysis capabilities. The paper aims at bridging the gap between the potency of SOM-machine learning to reduce dimension of high-dimensional data on one hand and practical applications with special emphasis on gene expression analysis on the other hand. Results The method was applied to generate a SOM characterizing the whole genome expression profiles of 67 healthy human tissues selected from ten tissue categories (adipose, endocrine, homeostasis, digestion, exocrine, epithelium, sexual reproduction, muscle, immune system and nervous tissues. SOM mapping reduces the dimension of expression data from ten of thousands of genes to a few thousand metagenes, each representing a minicluster of co-regulated single genes. Tissue-specific and common properties shared between groups of tissues emerge as a handful of localized spots in the tissue maps collecting groups of co-regulated and co-expressed metagenes. The functional context of the spots was discovered using overrepresentation analysis with respect to pre-defined gene sets of known functional impact. We found that tissue related spots typically contain enriched populations of genes related to specific molecular processes in the respective tissue. Analysis techniques normally used at the gene-level such as two-way hierarchical clustering are better represented and provide better signal-to-noise ratios if applied to the metagenes. Metagene-based clustering analyses aggregate the tissues broadly into three clusters containing nervous, immune system and the remaining tissues

  2. Expression cartography of human tissues using self organizing maps.

    Science.gov (United States)

    Wirth, Henry; Löffler, Markus; von Bergen, Martin; Binder, Hans

    2011-07-27

    Parallel high-throughput microarray and sequencing experiments produce vast quantities of multidimensional data which must be arranged and analyzed in a concerted way. One approach to addressing this challenge is the machine learning technique known as self organizing maps (SOMs). SOMs enable a parallel sample- and gene-centered view of genomic data combined with strong visualization and second-level analysis capabilities. The paper aims at bridging the gap between the potency of SOM-machine learning to reduce dimension of high-dimensional data on one hand and practical applications with special emphasis on gene expression analysis on the other hand. The method was applied to generate a SOM characterizing the whole genome expression profiles of 67 healthy human tissues selected from ten tissue categories (adipose, endocrine, homeostasis, digestion, exocrine, epithelium, sexual reproduction, muscle, immune system and nervous tissues). SOM mapping reduces the dimension of expression data from ten of thousands of genes to a few thousand metagenes, each representing a minicluster of co-regulated single genes. Tissue-specific and common properties shared between groups of tissues emerge as a handful of localized spots in the tissue maps collecting groups of co-regulated and co-expressed metagenes. The functional context of the spots was discovered using overrepresentation analysis with respect to pre-defined gene sets of known functional impact. We found that tissue related spots typically contain enriched populations of genes related to specific molecular processes in the respective tissue. Analysis techniques normally used at the gene-level such as two-way hierarchical clustering are better represented and provide better signal-to-noise ratios if applied to the metagenes. Metagene-based clustering analyses aggregate the tissues broadly into three clusters containing nervous, immune system and the remaining tissues. The SOM technique provides a more intuitive and

  3. Human immunodeficiency virus receptor and coreceptor expression on human uterine epithelial cells: regulation of expression during the menstrual cycle and implications for human immunodeficiency virus infection.

    Science.gov (United States)

    Yeaman, Grant R; Howell, Alexandra L; Weldon, Sally; Demian, Douglas J; Collins, Jane E; O'Connell, Denise M; Asin, Susana N; Wira, Charles R; Fanger, Michael W

    2003-05-01

    Human immunodeficiency virus-1 (HIV-1) is primarily a sexually transmitted disease. Identification of cell populations within the female reproductive tract that are initially infected, and the events involved in transmission of infection to other cells, remain to be established. In this report, we evaluated expression of HIV receptors and coreceptors on epithelial cells in the uterus and found they express several receptors critical for HIV infection including CD4, CXCR4, CCR5 and galactosylceramide (GalC). Moreover, expression of these receptors varied during the menstrual cycle. Expression of CD4 and CCR5 on uterine epithelial cells is high throughout the proliferative phase of the menstrual cycle when blood levels of oestradiol are high. In contrast, CXCR4 expression increased gradually throughout the proliferative phase. During the secretory phase of the cycle when both oestradiol and progesterone are elevated, CD4 and CCR5 expression decreased whereas CXCR4 expression remained elevated. Expression of GalC on endometrial glands is higher during the secretory phase than during the proliferative phase of the menstrual cycle. Because epithelial cells line the female reproductive tract and express HIV receptors and coreceptors, it is likely that they are one of the first cell types to become infected. The hormonal regulation of HIV receptor expression may affect a woman's susceptibility to HIV infection during her menstrual cycle. Moreover, selective coreceptor expression could account for the preferential transmission of R5-HIV-1 strains to women. In addition, these studies provide evidence that the uterus, and potentially the entire upper reproductive tract, are important sites for the initial events involved in HIV infection.

  4. Expression of Human Immunodeficiency Virus Type 1 Neutralizing Antibody Fragments Using Human Vaginal Lactobacillus

    Science.gov (United States)

    Marcobal, Angela; Liu, Xiaowen; Zhang, Wenlei; Dimitrov, Antony S.; Jia, Letong; Lee, Peter P.; Fouts, Timothy R.; Parks, Thomas P.

    2016-01-01

    Abstract Eradication of human immunodeficiency virus type 1 (HIV-1) by vaccination with epitopes that produce broadly neutralizing antibodies is the ultimate goal for HIV prevention. However, generating appropriate immune responses has proven difficult. Expression of broadly neutralizing antibodies by vaginal colonizing lactobacilli provides an approach to passively target these antibodies to the mucosa. We tested the feasibility of expressing single-chain and single-domain antibodies (dAbs) in Lactobacillus to be used as a topical microbicide/live biotherapeutic. Lactobacilli provide an excellent platform to express anti-HIV proteins. Broadly neutralizing antibodies have been identified against epitopes on the HIV-1 envelope and have been made into active antibody fragments. We tested single-chain variable fragment m9 and dAb-m36 and its derivative m36.4 as prototype antibodies. We cloned and expressed the antibody fragments m9, m36, and m36.4 in Lactobacillus jensenii-1153 and tested the expression levels and functionality. We made a recombinant L. jensenii 1153-1128 that expresses dAb-m36.4. All antibody fragments m9, m36, and m36.4 were expressed by lactobacilli. However, we noted the smaller m36/m36.4 were expressed to higher levels, ≥3 μg/ml. All L. jensenii-expressed antibody fragments bound to gp120/CD4 complex; Lactobacillus-produced m36.4 inhibited HIV-1BaL in a neutralization assay. Using a TZM-bl assay, we characterized the breadth of neutralization of the m36.4. Delivery of dAbs by Lactobacillus could provide passive transfer of these antibodies to the mucosa and longevity at the site of HIV-1 transmission. PMID:26950606

  5. Microarray data integration for genome-wide analysis of human tissue-selective gene expression

    OpenAIRE

    Wang, Liangjiang; Srivastava, Anand K; Schwartz, Charles E

    2010-01-01

    Background Microarray gene expression data are accumulating in public databases. The expression profiles contain valuable information for understanding human gene expression patterns. However, the effective use of public microarray data requires integrating the expression profiles from heterogeneous sources. Results In this study, we have compiled a compendium of microarray expression profiles of various human tissue samples. The microarray raw data generated in different research laboratorie...

  6. Construction and Expression of Eukaryotic Expression Vector and Plasmid Expressing siRNA of Human Protection of Telomeres 1

    Institute of Scientific and Technical Information of China (English)

    Di-Nan HUANG; Ying-Hua JIANG; Hou GAN

    2005-01-01

    @@ 1 Introduction The POT1 (protection of telomeres 1 ) protein binds the single-stranded overhang at the ends of chromosomes in diverse eukaryocytes. It is essential for chromosome end-protection in the fission yeast Schizosaccharomyces pombe, and it is involved in regulation of telomere lengthin human cells. Human POT1 had been identified in 2001year. Its amino terminal is highly conservative in eukaryocytes. Since Pot1 can bind internal loops and directly adjacent DNA-binding sites, it is likely to fully coat and protect both G-strand overhangs and the displaced G strand of a T-loop. It also participates in the regulation of telomere maintenance by telomerase or of chromosome by other enzymes. But its biological function and mechanism are still unknown. hPOT eukaryotic expression vector(pcDNA3- hPOT1 ) and 3 kinds of siRNA expression vectors(pmU6-shDNA1, pmU6-shDNA2 and pmU6-shDNA3)had been constructed. And there were overexpression and RNA silence in HeLa cells.

  7. Human fetal liver stromal cells expressing erythropoietin promote hematopoietic development from human embryonic stem cells.

    Science.gov (United States)

    Yang, Chao; Ji, Lei; Yue, Wen; Shi, Shuang-Shuang; Wang, Ruo-Yong; Li, Yan-Hua; Xie, Xiao-Yan; Xi, Jia-Fei; He, Li-Juan; Nan, Xue; Pei, Xue-Tao

    2012-02-01

    Blood cells transfusion and hematopoietic stem cells (HSCs) transplantation are important methods for cell therapy. They are widely used in the treatment of incurable hematological disorder, infectious diseases, genetic diseases, and immunologic deficiency. However, their availability is limited by quantity, capacity of proliferation and the risk of blood transfusion complications. Recently, human embryonic stem cells (hESCs) have been shown to be an alternative resource for the generation of hematopoietic cells. In the current study, we describe a novel method for the efficient production of hematopoietic cells from hESCs. The stable human fetal liver stromal cell lines (hFLSCs) expressing erythropoietin (EPO) were established using the lentiviral system. We observed that the supernatant from the EPO transfected hFLSCs could induce the hESCs differentiation into hematopoietic cells, especially erythroid cells. They not only expressed fetal and embryonic globins but also expressed the adult-globin chain on further maturation. In addition, these hESCs-derived erythroid cells possess oxygen-transporting capacity, which indicated hESCs could generate terminally mature progenies. This should be useful for ultimately developing an animal-free culture system to generate large numbers of erythroid cells from hESCs and provide an experimental model to study early human erythropoiesis.

  8. Prokaryotic expression, purification and characterization of human cyclooxygenase-2.

    Science.gov (United States)

    Liao, Xiangzhi; Wang, Wenhan; Fan, Chuanxi; Yang, Ning; Zhao, Jialiang; Zhang, Ying; Gao, Ruijuan; Shen, Guannan; Xia, Simin; Li, Guiying

    2017-07-01

    Cyclooxygenase-2 (COX-2) is a key enzyme which catalyzes the conversion of arachidonic acid (AA) into prostaglandins (PGs). It plays an important role in pathophysiological processes, such as tumorigenesis, angiogenesis, inflammation and tumor cell drug resistance. Therefore, COX-2 has been viewed as an important target for cancer therapy. The preparation of COX-2 protein is an important initial step for the subsequent development of COX-2 inhibitors. In this study, we report a strategy to heterologously express truncated human COX-2 (trCOX-2) in Escherichia coli (E. coli) BL21(DE3) host cells. Following denaturation, purification and renaturation, we successfully obtained enzymatically active trCOX-2 containing 257 residues of the C-terminus. Homology modeling and molecular docking analyses revealed that trCOX-2 retained the predicted 3D catalytic domain structure and AA could still bind to its hydrophobic groove. Western blot analysis and ELISA indicated that the trCOX-2 still retained its characteristic antigenicity and binding activity, while COX assays revealed that trCOX-2 maintained its enzyme activity. On the whole, in this study, we provided a novel method to isolate trCOX-2 possessing AA binding and catalytic activities. This study thus lays a foundation to facilitate further investigations of COX-2 and offers a valuable method with which to achieve the prokaryotic expression of a eukaryotic membrane protein.

  9. Syndecan expressions in the human amnion and chorionic plate

    Directory of Open Access Journals (Sweden)

    T. Lorenzi

    2010-10-01

    Full Text Available The syndecan family consists of four distinct membrane glycoproteins in mammals. Syndecans control cell proliferation, differentiation, adhesion and migration through participation in cell-cell interactions, anchorage of cells to the extracellular environment, and modulation of multiple growth factors. Therefore, syndecans may play a pivotal role in the regulation of cell behaviour depending on the cellular microenvironment. Here, we demonstrate that syndecan-1, syndecan-2 and syndecan-4 are expressed in fetal membrane tissue with different immunolocalizations. Syndecan-1 is expressed in the amniotic epithelium, localizing at basolateral cell surfaces. Syndecan-2 and syndecan-4, in contrast, are mostly localized in intracellular compartments, in the extravillous cytotrophoblastic cells and in some fibroblasts of the chorionic plate as well as in the amniotic epithelial cells. In the latter, syndecan-4 is mainly localized in the apical part of the cells. Our results strongly suggest a key role of syndecan-1, syndecan-2 and syndecan-4 in the determination of structural and functional characteristics of human amnion and chorionic plate. Since the solute exchanges between fetus and mother take place in fetal membranes, our data suggest that syndecans are important players in the placenta for the establishment of the fetal-maternal inter-communication.

  10. Syndecan expressions in the human amnion and chorionic plate.

    Science.gov (United States)

    Lorenzi, T; Turi, A; Crescimanno, C; Morroni, M; Castellucci, M; David, G; Tranquilli, A L; Marzioni, D

    2010-10-27

    The syndecan family consists of four distinct membrane glycoproteins in mammals. Syndecans control cell proliferation, differentiation, adhesion and migration through participation in cell-cell interactions, anchorage of cells to the extracellular environment, and modulation of multiple growth factors. Therefore, syndecans may play a pivotal role in the regulation of cell behaviour depending on the cellular microenvironment. Here, we demonstrate that syndecan-1, syndecan-2 and syndecan-4 are expressed in fetal membrane tissue with different immunolocalizations. Syndecan-1 is expressed in the amniotic epithelium, localizing at basolateral cell surfaces. Syndecan-2 and syndecan-4, in contrast, are mostly localized in intracellular compartments, in the extravillous cytotrophoblastic cells and in some fibroblasts of the chorionic plate as well as in the amniotic epithelial cells. In the latter, syndecan-4 is mainly localized in the apical part of the cells. Our results strongly suggest a key role of syndecan-1, syndecan-2 and syndecan-4 in the determination of structural and functional characteristics of human amnion and chorionic plate. Since the solute exchanges between fetus and mother take place in fetal membranes, our data suggest that syndecans are important players in the placenta for the establishment of the fetal-maternal inter-communication.

  11. Gene expression profiling gut microbiota in different races of humans

    Science.gov (United States)

    Chen, Lei; Zhang, Yu-Hang; Huang, Tao; Cai, Yu-Dong

    2016-03-01

    The gut microbiome is shaped and modified by the polymorphisms of microorganisms in the intestinal tract. Its composition shows strong individual specificity and may play a crucial role in the human digestive system and metabolism. Several factors can affect the composition of the gut microbiome, such as eating habits, living environment, and antibiotic usage. Thus, various races are characterized by different gut microbiome characteristics. In this present study, we studied the gut microbiomes of three different races, including individuals of Asian, European and American races. The gut microbiome and the expression levels of gut microbiome genes were analyzed in these individuals. Advanced feature selection methods (minimum redundancy maximum relevance and incremental feature selection) and four machine-learning algorithms (random forest, nearest neighbor algorithm, sequential minimal optimization, Dagging) were employed to capture key differentially expressed genes. As a result, sequential minimal optimization was found to yield the best performance using the 454 genes, which could effectively distinguish the gut microbiomes of different races. Our analyses of extracted genes support the widely accepted hypotheses that eating habits, living environments and metabolic levels in different races can influence the characteristics of the gut microbiome.

  12. Antigenic assessment of a recombinant human CD90 protein expressed in prokaryotic expression system.

    Science.gov (United States)

    Yousefi-Rad, Narges; Shokrgozar, Mohammad Ali; Behdani, Mahdi; Moradi-Kalbolandi, Shima; Motamedi-Rad, Mahdieh; Habibi-Anbouhi, Mahdi

    2015-12-01

    Cluster of Differentiation 90 (CD90, Thy-1) has been proposed as one of the most important biomarkers in several cancer cells including cancer stem cells (CSCs). CD90 is considered as a potential normal stem cell and CSCs biomarker and also has been identified in lung cancer stem cells, hepatocellular carcinoma cells and high-grade gliomas. Using eukaryotic host systems involves complex procedures and frequently results in low protein yields. The expression of recombinant proteins in Escherichia coli is comparatively easier than eukaryotic host cells. The potential of large scale production of recombinant protein has made this system an economic production platform. In this study we expressed the extra-membrane domain of human CD90 (exCD90) antigen (Gln15-Cys130) in E. coli expression host cells. The epitope integrity of purified recombinant antigen was confirmed by antibody-antigen interaction using 5E10 anti-CD90 monoclonal antibody and binding study through ELISA and florescent staining of CD90(+) cells in a flow cytometry experiment. Copyright © 2015 Elsevier Inc. All rights reserved.

  13. Impaired Pten expression in human malignant peripheral nerve sheath tumours.

    Directory of Open Access Journals (Sweden)

    Maren Bradtmöller

    Full Text Available Malignant peripheral nerve sheath tumours (MPNST are aggressive sarcomas that develop in about 10% of patients with the genetic disease neurofibromatosis type 1 (NF1. Molecular alterations contributing to MPNST formation have only partially been resolved. Here we examined the role of Pten, a key regulator of the Pi3k/Akt/mTOR pathway, in human MPNST and benign neurofibromas. Immunohistochemistry showed that Pten expression was significantly lower in MPNST (n=16 than in neurofibromas (n=16 and normal nervous tissue. To elucidate potential mechanisms for Pten down-regulation or Akt/mTOR activation in MPNST we performed further experiments. Mutation analysis revealed absence of somatic mutations in PTEN (n=31 and PIK3CA (n=38. However, we found frequent PTEN promotor methylation in primary MPNST (11/26 and MPNST cell lines (7/8 but not in benign nerve sheath tumours. PTEN methylation was significantly associated with early metastasis. Moreover, we detected an inverse correlation of Pten-regulating miR-21 and Pten protein levels in MPNST cell lines. The examination of NF1-/- and NF1+/+Schwann cells and fibroblasts showed that Pten expression is not regulated by NF1. To determine the significance of Pten status for treatment with the mTOR inhibitor rapamycin we treated 5 MPNST cell lines with rapamycin. All cell lines were sensitive to rapamycin without a significant correlation to Pten levels. When rapamycin was combined with simvastatin a synergistic anti-proliferative effect was achieved. Taken together we show frequent loss/reduction of Pten expression in MPNST and provide evidence for the involvement of multiple Pten regulating mechanisms.

  14. Microbiota diversity and gene expression dynamics in human oral biofilms.

    Science.gov (United States)

    Benítez-Páez, Alfonso; Belda-Ferre, Pedro; Simón-Soro, Aurea; Mira, Alex

    2014-04-27

    Micro-organisms inhabiting teeth surfaces grow on biofilms where a specific and complex succession of bacteria has been described by co-aggregation tests and DNA-based studies. Although the composition of oral biofilms is well established, the active portion of the bacterial community and the patterns of gene expression in vivo have not been studied. Using RNA-sequencing technologies, we present the first metatranscriptomic study of human dental plaque, performed by two different approaches: (1) A short-reads, high-coverage approach by Illumina sequencing to characterize the gene activity repertoire of the microbial community during biofilm development; (2) A long-reads, lower-coverage approach by pyrosequencing to determine the taxonomic identity of the active microbiome before and after a meal ingestion. The high-coverage approach allowed us to analyze over 398 million reads, revealing that microbial communities are individual-specific and no bacterial species was detected as key player at any time during biofilm formation. We could identify some gene expression patterns characteristic for early and mature oral biofilms. The transcriptomic profile of several adhesion genes was confirmed through qPCR by measuring expression of fimbriae-associated genes. In addition to the specific set of gene functions overexpressed in early and mature oral biofilms, as detected through the short-reads dataset, the long-reads approach detected specific changes when comparing the metatranscriptome of the same individual before and after a meal, which can narrow down the list of organisms responsible for acid production and therefore potentially involved in dental caries. The bacteria changing activity during biofilm formation and after meal ingestion were person-specific. Interestingly, some individuals showed extreme homeostasis with virtually no changes in the active bacterial population after food ingestion, suggesting the presence of a microbial community which could be

  15. HMGA1-pseudogene expression is induced in human pituitary tumors

    Science.gov (United States)

    Esposito, Francesco; De Martino, Marco; D'Angelo, Daniela; Mussnich, Paula; Raverot, Gerald; Jaffrain-Rea, Marie-Lise; Fraggetta, Filippo; Trouillas, Jacqueline; Fusco, Alfredo

    2015-01-01

    Numerous studies have established that High Mobility Group A (HMGA) proteins play a pivotal role on the onset of human pituitary tumors. They are overexpressed in pituitary tumors, and, consistently, transgenic mice overexpressing either the Hmga1 or the Hmga2 gene develop pituitary tumors. In contrast with HMGA2, HMGA1 overexpression is not related to any rearrangement or amplification of the HMGA1 locus in these tumors. We have recently identified 2 HMGA1 pseudogenes, HMGA1P6 and HMGA1P7, acting as competitive endogenous RNA decoys for HMGA1 and other cancer related genes. Here, we show that HMGA1 pseudogene expression significantly correlates with HMGA1 mRNA levels in growth hormone and nonfunctioning pituitary adenomas likely inhibiting the repression of HMGA1 through microRNAs action. According to our functional studies, these HMGA1 pseudogenes enhance the proliferation and migration of the mouse pituitary tumor cell line, at least in part, through their upregulation. Our results point out that the overexpression of HMGA1P6 and HMGA1P7 could contribute to increase HMGA1 levels in human pituitary tumors, and then to pituitary tumorigenesis. PMID:25894544

  16. Human recombinant soluble guanylyl cyclase: expression, purification, and regulation

    Science.gov (United States)

    Lee, Y. C.; Martin, E.; Murad, F.

    2000-01-01

    The alpha1- and beta1-subunits of human soluble guanylate cyclase (sGC) were coexpressed in the Sf9 cells/baculovirus system. In addition to the native enzyme, constructs with hexahistidine tag at the amino and carboxyl termini of each subunit were coexpressed. This permitted the rapid and efficient purification of active recombinant enzyme on a nickel-affinity column. The enzyme has one heme per heterodimer and was readily activated with the NO donor sodium nitroprusside or 3-(5'-hydroxymethyl-2'furyl)-1-benzyl-indazole (YC-1). Sodium nitroprusside and YC-1 treatment potentiated each other in combination and demonstrated a remarkable 2,200-fold stimulation of the human recombinant sGC. The effects were inhibited with 1H-(1,2, 4)oxadiazole(4,3-a)quinoxalin-1one (ODQ). The kinetics of the recombinant enzyme with respect to GTP was examined. The products of the reaction, cGMP and pyrophosphate, inhibited the enzyme. The extent of inhibition by cGMP depended on the activation state of the enzyme, whereas inhibition by pyrophosphate was not affected by the enzyme state. Both reaction products displayed independent binding and cooperativity with respect to enzyme inhibition. The expression of large quantities of active enzyme will facilitate structural characterization of the protein.

  17. Expression, purification and bioactivity of human augmenter of liver regeneration

    Institute of Scientific and Technical Information of China (English)

    Yang-De Zhang; Jian Zhou; Jin-Feng Zhao; Jian Peng; Xiao-Dong Liu; Xin-Sheng Liu; Ze-Ming Jia

    2006-01-01

    AIM: To construct the expression vectors for prokaryotic and eukaryotic human augmenter of liver regeneration (hALR) and to study their biological activity.METHODS: hALRcDNA clone was obtained from plasmid pGEM-T-hALR, and cDNA was subcloned into the prokatyotic expression vector pGEX-4T-2.The recombinant vector and pGEX-4T-2hALR were identified by enzyme digestion and DNA sequencing and transformed into E coli JM109. The positively selected clone was induced by the expression of GST-hALR fusion protein with IPTG, then the fusion protein was purified by glutathine s-transferase (GST) sepharose 4B affinity chromatography, cleaved by thrombin and the hALR monomer was obtained and detected by measuring H thymidine incorporation.RESULTS: The product of PCR from plasmid pGEM-ThALR was examined by 1.5% sepharose electrophoresis.The specific strap was coincident with the theoretical one. The sequence was accurate and pGEX-4T-hALP digested by enzymes was coincident with the theoretical one. The sequence was accurate and the fragment was inserted in the positive direction. The recombinant vector was transformed into E coli JM109. SDS-PAGE proved that the induced expressive fusion protein showed a single band with a molecular weight of 41 kDa. The product was purified and cleaved. The molecular weights of GST and hALR were 26 kDa, 15 kDa respectively. The recombinant fusion protein accounted for 31% of the total soluble protein of bacterial lysate. HALR added to the culture medium of adult rat hepatocytes in primary culture and HepG2 cell line could significantly enhance the rate of DNA synthesis compared to the relevant control groups (P < 0.01).CONCLUSION: Purified hALR has the ability to stimulate DNA synthesis of adult rat hepatocytes in primary culture and HepG2 cells in vitro, and can provide evidence for its clinical application.

  18. Cloning of human brevican cDNA and expression of its mRNA in human glioma

    Institute of Scientific and Technical Information of China (English)

    韩唏; 董艳; 由振东; 何成; 卢亦成

    2003-01-01

    Objective:To clone the cDNA of human brevican secreting isoform and to investigate its mRNA expression in human glioma.Methods:The full-length cDNA of human brevican secreted isoform was cloned from a human ahaplastic astrocytoma by RT-PCR,and the expression of human brevican mRNA in 22 cases of human glioma and 13 cases of non-glial brain tumors were investigated by in situ hybridization.Results:The cDNA which including the whole open reading frame of human brevican secreted isoform was obtained.In situ hybridization showed that brevican positive cells were present in all of the 22 cases of gliomas(100%),whereas none were found in the 13 cases of non-glial and metastasis brain tumors examined.Conclusion:The results suggest that brevican mRNA is highly and specifically expressed in human glioma.

  19. hTERT protein expression is independent of clinicopathological parameters and c-Myc protein expression in human breast cancer

    Directory of Open Access Journals (Sweden)

    Meligonis G

    2005-01-01

    Full Text Available Abstract Background Telomerase is a ribonucleoprotein enzyme that synthesises telomeres after cell division and maintains chromosomal length and stability thus leading to cellular immortalisation. The hTERT (human telomerase reverse transcriptase subunit seems to be the rate-limiting determinant of telomerase and knowledge of factors controlling hTERT transcription may be useful in therapeutic strategies. The hTERT promoter contains binding sites for c-Myc and there is some experimental and in vitro evidence that c-Myc may increase hTERT expression. We previously reported no correlation between c-Myc mRNA expression and hTERT mRNA or telomerase activity in human breast cancer. This study aims to examine the correlation between hTERT expression as determined by immunohistochemistry and c-Myc expression, lymph node status, and tumour size and grade in human breast cancer. Materials and methods The immunohistochemical expression of hTERT and c-Myc was investigated in 38 malignant breast tumours. The expression of hTERT was then correlated with the lymph node status, c-Myc expression and other clinicopathological parameters of the tumours. Results hTERT expression was positive in 27 (71% of the 38 tumours. 15 (79% of 19 node positive tumours were hTERT positive compared with 11 (63% of 19 node negative tumours. The expression was higher in node positive tumours but this failed to reach statistical significance (p = 0.388. There was no significant association with tumour size, tumour grade or c-Myc expression. However, hTERT expression correlated positively with patients' age (correlation coefficient = 0.415, p = 0.0097. Conclusion hTERT protein expression is independent of lymph node status, tumour size and grade and c-Myc protein expression in human breast cancer

  20. Effect of 5'-flanking sequence deletions on expression of the human insulin gene in transgenic mice

    DEFF Research Database (Denmark)

    Fromont-Racine, M; Bucchini, D; Madsen, O;

    1990-01-01

    Expression of the human insulin gene was examined in transgenic mouse lines carrying the gene with various lengths of DNA sequences 5' to the transcription start site (+1). Expression of the transgene was demonstrated by 1) the presence of human C-peptide in urine, 2) the presence of specific tra...... of the transgene was observed in cell types other than beta-islet cells.......Expression of the human insulin gene was examined in transgenic mouse lines carrying the gene with various lengths of DNA sequences 5' to the transcription start site (+1). Expression of the transgene was demonstrated by 1) the presence of human C-peptide in urine, 2) the presence of specific......, and -168 allowed correct initiation of the transcripts and cell specificity of expression, while quantitative expression gradually decreased. Deletion to -58 completely abolished the expression of the gene. The amount of human product that in mice harboring the longest fragment contributes up to 50...

  1. Expression of basal cell keratins in human prostate cancer metastases and cell lines.

    NARCIS (Netherlands)

    Leenders, G.J.L.H. van; Aalders, M.W.; Hulsbergen-van de Kaa, C.A.; Ruiter, D.J.; Schalken, J.A.

    2001-01-01

    Within normal human prostate epithelium, basal and luminal cells can be discriminated by their expression of keratins (K). While basal cells express K5/14, luminal cells show expression of K8/18 and an intermediate cell population can be identified by co-expression of K5/18. Prostate cancer is predo

  2. Expression of muscarinic binding sites in primary human brain tumors.

    Science.gov (United States)

    Gurwitz, D; Razon, N; Sokolovsky, M; Soreq, H

    1984-05-01

    The expression of muscarinic binding sites was examined in a collection of primary brain tumors of different cellular origins and various degrees of dedifferentiation, as compared to control specimens. Eleven gliogenous tumors were examined, all of which contained substantial amounts of muscarinic binding sites. Most of the other tumor types examined did not display detectable binding of [3H]N-methyl-4-piperidyl benzilate ([3H]4NMPB). Scatchard analysis indicated the existence of homogeneous antagonist sites in both normal forebrain and glioblastoma multiforme, with Kd values of 1.2 nM and 0.9 nM, respectively. The density of muscarinic binding sites varied between tumors from different patients, and also between specimens prelevated from different areas of the same tumor. This variability, as well as the average density of binding sites, appeared to be larger in highly malignant tumors than in less malignant ones. In contrast, the density of muscarinic receptors from control specimens was invariably high, but within the same order of magnitude. To test whether the muscarinic binding activity in the brain tumors is correlated to other cholinoceptive properties, cholinesterase activity was also examined. Individual data for density of [3H]4NMPB binding sites were then plotted against corresponding values of cholinesterase activity. The pattern of distribution of these values was clearly different in tumor specimens, when compared to that observed in samples derived from non-malignant brain. Our observations indicate that human brain cells of gliogenous origin are capable of expressing muscarinic binding sites, and that, if a correlation exists between muscarinic receptors and cholinesterase levels in gliogenous tumors, it differs from that of non-malignant brain tissue.

  3. Expression of stem cell markers in the human fetal kidney.

    Directory of Open Access Journals (Sweden)

    Sally Metsuyanim

    Full Text Available In the human fetal kidney (HFK self-renewing stem cells residing in the metanephric mesenchyme (MM/blastema are induced to form all cell types of the nephron till 34(th week of gestation. Definition of useful markers is crucial for the identification of HFK stem cells. Because wilms' tumor, a pediatric renal cancer, initiates from retention of renal stem cells, we hypothesized that surface antigens previously up-regulated in microarrays of both HFK and blastema-enriched stem-like wilms' tumor xenografts (NCAM, ACVRIIB, DLK1/PREF, GPR39, FZD7, FZD2, NTRK2 are likely to be relevant markers. Comprehensive profiling of these putative and of additional stem cell markers (CD34, CD133, c-Kit, CD90, CD105, CD24 in mid-gestation HFK was performed using immunostaining and FACS in conjunction with EpCAM, an epithelial surface marker that is absent from the MM and increases along nephron differentiation and hence can be separated into negative, dim or bright fractions. No marker was specifically localized to the MM. Nevertheless, FZD7 and NTRK2 were preferentially localized to the MM and emerging tubules (50% of HFK cells and predominantly co-express EpCAM(bright, indicating they are mostly markers of differentiation. Furthermore, localization of NCAM exclusively in the MM and in its nephron progenitor derivatives but also in stroma and the expression pattern of significantly elevated renal stem/progenitor genes Six2, Wt1, Cited1, and Sall1 in NCAM(+EpCAM(- and to a lesser extent in NCAM(+EpCAM(+ fractions confirmed regional identity of cells and assisted us in pinpointing the presence of subpopulations that are putative MM-derived progenitor cells (NCAM(+EpCAM(+FZD7(+, MM stem cells (NCAM(+EpCAM(-FZD7(+ or both (NCAM(+FZD7(+. These results and concepts provide a framework for developing cell selection strategies for human renal cell-based therapies.

  4. Expression of leptin and leptin receptor isoforms in the human stomach

    OpenAIRE

    Mix, H.; Widjaja, A; Jandl, O.; Cornberg, M; Kaul, A; Goke, M; Beil, W.; Kuske, M.; Brabant, G; Manns, M; Wagner, S.

    2000-01-01

    BACKGROUND—Leptin is an important regulator of food intake and energy expenditure. Initially it was thought to be expressed exclusively in and secreted by adipocytes. Recently, leptin expression was also noted in other tissues, including rat gastric mucosa. Information on leptin and leptin receptor expression in the human stomach is lacking.
AIM—To investigate expression of leptin and its corresponding receptors in human gastric epithelial cells.
METHODS—Fundic and antral gastric mucosal biop...

  5. Expression of leptin and leptin receptor isoforms in the human stomach

    OpenAIRE

    Mix, H.; Widjaja, A; Jandl, O.; Cornberg, M.; Kaul, A.; GOKE, M; Beil, W; Kuske, M.; Brabant, G; Manns, M; Wagner, S.

    2000-01-01

    BACKGROUND—Leptin is an important regulator of food intake and energy expenditure. Initially it was thought to be expressed exclusively in and secreted by adipocytes. Recently, leptin expression was also noted in other tissues, including rat gastric mucosa. Information on leptin and leptin receptor expression in the human stomach is lacking.
AIM—To investigate expression of leptin and its corresponding receptors in human gastric epithelial cells.
METHODS—Fundic and antral gastric mucosal biop...

  6. Expression and alternative splicing pattern of human telomerase reverse transcriptase in human lung cancer cells.

    Science.gov (United States)

    Fujiwara, Masachika; Kamma, Hiroshi; Wu, Wenwen; Hamasaki, Makoto; Kaneko, Setsuko; Horiguchi, Hisashi; Matsui-Horiguchi, Miwa; Satoh, Hiroaki

    2004-04-01

    Telomerase activity is generally considered to be necessary for cancer cells to avoid senescence. The expression of human telomerase reverse transcriptase (hTERT) is believed to be a rate-limiting step in telomerase activation. Recently, it has been proposed that the alternative splicing of hTERT is also involved in regulation of telomerase activity. However, the regulatory mechanism of telomerase in cancer cells has not been thoroughly investigated. To clarify it in lung cancer cells, we measured the expression of the hTERT transcript, analyzed its alternative splicing by RT-PCR, and compared it with telomerase activity and telomere length. The expression of the hTERT transcript was positively correlated with telomerase activity in lung cancer cells. Cancer cells with high telomerase activity contained 4 splicing variants of hTERT, and the full-length variant was 31.3-54.2% of the total transcripts. Cells of the TKB-20 cell line, which has extremely low telomerase activity, showed a different splicing pattern of hTERT in addition to low expression. The functional full-length variant was scarcely detected in TKB-20 cells, suggesting that the telomerase activity was repressed by alternative splicing of hTERT. Telomere length was not necessarily correlated with telomerase activity or hTERT expression in lung cancer cells. Cells of the TKB-4 cell line that also showed relatively low telomerase activity (as TKB-20 cells) had long telomeres. In conclusion, hTERT expression is regulated at both the transcriptional and post-transcriptional levels in lung cancer cells, and the alternative splicing of hTERT is involved in the control of telomerase activity.

  7. Establishing the baseline level of repetitive element expression in the human cortex

    National Research Council Canada - National Science Library

    Tyekucheva, Svitlana; Yolken, Robert H; McCombie, W Richard; Parla, Jennifer; Kramer, Melissa; Wheelan, Sarah J; Sabunciyan, Sarven

    2011-01-01

    .... Hence, we performed whole transcriptome sequencing to investigate the expression of repetitive elements in human frontal cortex using postmortem tissue obtained from the Stanley Medical Research Institute...

  8. Turning Avatar into Realistic Human Expression Using Linear and Bilinear Interpolations

    Science.gov (United States)

    Hazim Alkawaz, Mohammed; Mohamad, Dzulkifli; Rehman, Amjad; Basori, Ahmad Hoirul

    2014-06-01

    The facial animation in term of 3D facial data has accurate research support of the laser scan and advance 3D tools for complex facial model production. However, the approach still lacks facial expression based on emotional condition. Though, facial skin colour is required to offers an effect of facial expression improvement, closely related to the human emotion. This paper presents innovative techniques for facial animation transformation using the facial skin colour based on linear interpolation and bilinear interpolation. The generated expressions are almost same to the genuine human expression and also enhance the facial expression of the virtual human.

  9. Effect of 5'-flanking sequence deletions on expression of the human insulin gene in transgenic mice

    DEFF Research Database (Denmark)

    Fromont-Racine, M; Bucchini, D; Madsen, O

    1990-01-01

    Expression of the human insulin gene was examined in transgenic mouse lines carrying the gene with various lengths of DNA sequences 5' to the transcription start site (+1). Expression of the transgene was demonstrated by 1) the presence of human C-peptide in urine, 2) the presence of specific......, and -168 allowed correct initiation of the transcripts and cell specificity of expression, while quantitative expression gradually decreased. Deletion to -58 completely abolished the expression of the gene. The amount of human product that in mice harboring the longest fragment contributes up to 50...... of the transgene was observed in cell types other than beta-islet cells....

  10. Intramammary expression and therapeutic effect of a human lysozyme-expressing vector for treating bovine mastitis

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    To develop a gene therapy strategy for treating bovine mastitis, a new mammary-specific vector containing human lysozyme (hLYZ) cDNA and kanamycin resistance gene was constructed for intramammary expression and clinical studies. After one time acupuncture or intracisternal infusion of healthy cows with 400 μg of the p215C3LYZ vector, over 2.0 μg/ml of rhLYZ could be detected by enzymatic assay for about 3 weeks in the milk samples. Western blotting showed that rhLYZ secreted into milk samples from the vector-injected cows had molecular weight similar to that of the natural hLYZ in human colostrums.Twenty days after the primary injection, the quarters were re-injected with the same vector by quarter acupuncture and even higher concentrations of rhLYZ could be detected. Indirect competitive ELISA of milk samples showed that the vector injection did not induce detectable humoral immune response against hLYZ. Clinical studies showed that twice acupuncture of quarters with the p215C3LYZ vector had overt therapeutic effect on clinical and subclinical mastitis previously treated with antibiotics, including disappearance of clinical symptoms and relatively high microbiological cure rates. These data provide a solid rationale for using the vector to develop gene therapy for treating bovine mastitis.

  11. Factors affecting the gene expression of in vitro cultured human preimplantation embryos

    NARCIS (Netherlands)

    Mantikou, E.; Jonker, M.J.; Wong, K.M.; van Montfoort, A.P.A.; de Jong, M.; Breit, T.M.; Repping, S.; Mastenbroek, S.

    2016-01-01

    STUDY QUESTION: What is the relative effect of common environmental and biological factors on transcriptome changes during human preimplantation development? SUMMARY ANSWER: Developmental stage and maternal age had a larger effect on the global gene expression profile of human preimplantation

  12. Effects of human papillomavirus (HPV type 16 oncoproteins on the expression of involucrin in human keratinocytes

    Directory of Open Access Journals (Sweden)

    Gyöngyösi Eszter

    2012-02-01

    Full Text Available Abstract Background The human papillomavirus (HPV life cycle is closely linked to keratinocyte differentiation. Oncogenic HPV infection has been shown to hamper the normal differentiation of keratinocytes; however, the underlying mechanisms responsible for this phenomenon are yet to be clarified. Here, we aimed to study the effects of HPV16 E6 and E7 oncogenes on the expression of involucrin (IVL, an established marker of keratinocyte differentiation, in human foreskin keratinocyte (HFK cells. Results The differentiation of HFK cells by serum and high calcium significantly increased both the mRNA and the protein levels of IVL. The E6 and E7 oncoproteins of HPV16 together caused strong down-regulation of IVL mRNA and protein both in proliferating and in differentiating HFK cells. To study the effects of HPV oncogenes on the IVL promoter, we made transient transfection assays and luciferase tests and found that HPV 16 E6 but not E7 repressed IVL promoter activity in proliferating HFK cells. The inhibitory effect of HPV 16 E6 on the human IVL promoter could be localised to the proximal regulatory region (PRR of the gene. Conclusions These results suggest that the down-regulation of IVL promoter activity by HPV 16 E6 significantly contribute to the inhibition of endogenous IVL expression by the HPV 16 oncoproteins. In contrast, the down-regulation of endogenous IVL expression by HPV16 E7 is probably not caused by a direct and specific effect of E7 on the IVL promoter.

  13. Measuring Escherichia coli Gene Expression during Human Urinary Tract Infections

    Science.gov (United States)

    Mobley, Harry L. T.

    2016-01-01

    Extraintestinal Escherichia coli (E. coli) evolved by acquisition of pathogenicity islands, phage, plasmids, and DNA segments by horizontal gene transfer. Strains are heterogeneous but virulent uropathogenic isolates more often have specific fimbriae, toxins, and iron receptors than commensal strains. One may ask whether it is the virulence factors alone that are required to establish infection. While these virulence factors clearly contribute strongly to pathogenesis, bacteria must survive by metabolizing nutrients available to them. By constructing mutants in all major metabolic pathways and co-challenging mice transurethrally with each mutant and the wild type strain, we identified which major metabolic pathways are required to infect the urinary tract. We must also ask what else is E. coli doing in vivo? To answer this question, we examined the transcriptome of E. coli CFT073 in the murine model of urinary tract infection (UTI) as well as for E. coli strains collected and analyzed directly from the urine of patients attending either a urology clinic or a university health clinic for symptoms of UTI. Using microarrays and RNA-seq, we measured in vivo gene expression for these uropathogenic E. coli strains, identifying genes upregulated during murine and human UTI. Our findings allow us to propose a new definition of bacterial virulence. PMID:26784237

  14. Modified expression of surface glyconjugates in stored human platelets

    Energy Technology Data Exchange (ETDEWEB)

    Dhar, A.; Ganguly, P.

    1987-05-01

    Platelets are anucleated cells which play an important part in blood coagulation and thrombosis. These cells may be stored in the blood bank for only 4/5 days. In order to improve the storage of platelets, it is essential to first understand the changes in these cells due to storage. In this work, human platelets were stored in autologous plasma at 4/sup 0/ or 22/sup 0/ and their surface changes were monitored with three lectins - wheat germ afflutinin (WGA), concanavalin A (Con A) and lentil lectin (LL). Blood was drawn from healthy donors and platelet rich plasma (PRP) was collected by slow speed centrifugation. Platelets stored at either temperature for different times showed increased sensitivity to agglutination by WGA after 34-48 hrs. Lectins, Con A and LL, which were not agglutinating to fresh platelets readily caused agglutination after 48-72 hrs. The platelets stored for 25 hrs or longer period were insensitive to thrombin but showed enhanced aggregation with WGA. Labelling of surface glycoconjugates of stored platelets with /sup 3/H-boro-hydride revealed progressive loss of a glycoprotein of Mr 150,000 (GPIb infinity) together with the appearance of components of Mr 69,000; Mr 60,000; Mr 25,000. New high molecular weight glycoproteins were also detected only in stored platelets. The author studies clearly indicate that modification or altered expression of platelets surface glycoproteins may be one factor of storage related dysfunction of platelets.

  15. Expression of inducible nitric oxide in human lung epithelial cells.

    Science.gov (United States)

    Robbins, R A; Barnes, P J; Springall, D R; Warren, J B; Kwon, O J; Buttery, L D; Wilson, A J; Geller, D A; Polak, J M

    1994-08-30

    Nitric oxide (NO) is increased in the exhaled air of subjects with several airway disorders. To determine if cytokines could stimulate epithelial cells accounting for the increased NO, the capacity of the proinflammatory cytokines (cytomix: tumor necrosis factor-alpha, interleukin-1 beta, and interferon-gamma) to increase inducible nitric oxide synthase (iNOS) was investigated in A549 and primary cultures of human bronchial epithelial cells. Cytomix induced a time-dependent increase in nitrite levels in culture supernatant fluids (p < 0.05). Increased numbers of cells stained for iNOS and increased iNOS mRNA was detected in the cytokine-stimulated cells compared to control (p < 0.05). Dexamethasone diminished the cytokine-induced increase in nitrite, iNOS by immunocytochemistry, and iNOS mRNA. These data demonstrate that cytokines, such as those released by mononuclear cells, can induce lung epithelial iNOS expression and NO release, and that this is attenuated by dexamethasone.

  16. Iron acquisition functions expressed by the human pathogen Acinetobacter baumannii.

    Science.gov (United States)

    Zimbler, Daniel L; Penwell, William F; Gaddy, Jennifer A; Menke, Sharon M; Tomaras, Andrew P; Connerly, Pamela L; Actis, Luis A

    2009-02-01

    Acinetobacter baumannii is a gram-negative bacterium that causes serious infections in compromised patients. More recently, it has emerged as the causative agent of severe infections in military personnel wounded in Iraq and Afghanistan. This pathogen grows under a wide range of conditions including iron-limiting conditions imposed by natural and synthetic iron chelators. Initial studies using the type strain 19606 showed that the iron proficiency of this pathogen depends on the expression of the acinetobactin-mediated iron acquisition system. More recently, we have observed that hemin but not human hemoglobin serves as an iron source when 19606 isogenic derivatives affected in acinetobactin transport and biosynthesis were cultured under iron-limiting conditions. This finding is in agreement with the observation that the genome of the strain 17978 has a gene cluster coding for putative hemin-acquisition functions, which include genes coding for putative hemin utilization functions and a TonBExbBD energy transducing system. This system restored enterobactin biosynthesis in an E. coli ExbBD deficient strain but not when introduced into a TonB mutant. PCR and Southern blot analyses showed that this hemin-utilization gene cluster is also present in the 19606 strain. Analysis of the 17978 genome also showed that this strain harbors genes required for acinetobactin synthesis and transport as well as a gene cluster that could code for additional iron acquisition functions. This hypothesis is in agreement with the fact that the inactivation of the basD acinetobactin biosynthetic gene did not affect the growth of A. baumannii 17978 cells under iron-chelated conditions. Interestingly, this second iron uptake gene cluster is flanked by perfect inverted repeats and includes transposase genes that are expressed transcriptionally. Also interesting is the observation that this additional cluster could not be detected in the type strain 19606, an observation that suggests some

  17. Cholinergic regulation of VIP gene expression in human neuroblastoma cells

    DEFF Research Database (Denmark)

    Kristensen, Bo; Georg, Birgitte; Fahrenkrug, Jan

    1997-01-01

    Vasoactive intestinal polypeptide, muscarinic receptor, neuroblastoma cell, mRNA, gene expression, peptide processing......Vasoactive intestinal polypeptide, muscarinic receptor, neuroblastoma cell, mRNA, gene expression, peptide processing...

  18. MicroRNA expression and regulation in human, chimpanzee, and macaque brains.

    Directory of Open Access Journals (Sweden)

    Hai Yang Hu

    2011-10-01

    Full Text Available Among other factors, changes in gene expression on the human evolutionary lineage have been suggested to play an important role in the establishment of human-specific phenotypes. However, the molecular mechanisms underlying these expression changes are largely unknown. Here, we have explored the role of microRNA (miRNA in the regulation of gene expression divergence among adult humans, chimpanzees, and rhesus macaques, in two brain regions: prefrontal cortex and cerebellum. Using a combination of high-throughput sequencing, miRNA microarrays, and Q-PCR, we have shown that up to 11% of the 325 expressed miRNA diverged significantly between humans and chimpanzees and up to 31% between humans and macaques. Measuring mRNA and protein expression in human and chimpanzee brains, we found a significant inverse relationship between the miRNA and the target genes expression divergence, explaining 2%-4% of mRNA and 4%-6% of protein expression differences. Notably, miRNA showing human-specific expression localize in neurons and target genes that are involved in neural functions. Enrichment in neural functions, as well as miRNA-driven regulation on the human evolutionary lineage, was further confirmed by experimental validation of predicted miRNA targets in two neuroblastoma cell lines. Finally, we identified a signature of positive selection in the upstream region of one of the five miRNA with human-specific expression, miR-34c-5p. This suggests that miR-34c-5p expression change took place after the split of the human and the Neanderthal lineages and had adaptive significance. Taken together these results indicate that changes in miRNA expression might have contributed to evolution of human cognitive functions.

  19. Astrocyte cultures derived from human brain tissue express angiotensinogen mRNA

    Energy Technology Data Exchange (ETDEWEB)

    Milsted, A.; Barna, B.P.; Ransohoff, R.M.; Brosnihan, K.B.; Ferrario, C.M. (Cleveland Clinic Foundation, OH (USA))

    1990-08-01

    The authors have identified human cultured cell lines that are useful for studying angiotensinogen gene expression and its regulation in the central nervous system. A model cell system of human central nervous system origin expressing angiotensinogen has not previously been available. Expression of angiotensinogen mRNA appears to be a basal property of noninduced human astrocytes, since astrocytic cell lines derived from human glioblastomas or nonneoplastic human brain tissue invariably produced angiotensinogen mRNA. In situ hybridization histochemistry revealed that angiotensinogen mRNA production was not limited to a subpopulation of astrocytes because >99% of cells in these cultures contained angiotensinogen mRNA. These cell lines will be useful in studies of the molecular mechanisms controlling angiotensin synthesis and the role of biologically active angiotensin in the human brain by allowing the authors to examine regulation of expression of the renin-angiotensin system in human astrocyte cultures.

  20. Gene expression analysis in human breast cancer associated blood vessels.

    Directory of Open Access Journals (Sweden)

    Dylan T Jones

    Full Text Available Angiogenesis is essential for solid tumour growth, whilst the molecular profiles of tumour blood vessels have been reported to be different between cancer types. Although presently available anti-angiogenic strategies are providing some promise for the treatment of some cancers it is perhaps not surprisingly that, none of the anti-angiogenic agents available work on all tumours. Thus, the discovery of novel anti-angiogenic targets, relevant to individual cancer types, is required. Using Affymetrix microarray analysis of laser-captured, CD31-positive blood vessels we have identified 63 genes that are upregulated significantly (5-72 fold in angiogenic blood vessels associated with human invasive ductal carcinoma (IDC of the breast as compared with blood vessels in normal human breast. We tested the angiogenic capacity of a subset of these genes. Genes were selected based on either their known cellular functions, their enriched expression in endothelial cells and/or their sensitivity to anti-VEGF treatment; all features implicating their involvement in angiogenesis. For example, RRM2, a ribonucleotide reductase involved in DNA synthesis, was upregulated 32-fold in IDC-associated blood vessels; ATF1, a nuclear activating transcription factor involved in cellular growth and survival was upregulated 23-fold in IDC-associated blood vessels and HEX-B, a hexosaminidase involved in the breakdown of GM2 gangliosides, was upregulated 8-fold in IDC-associated blood vessels. Furthermore, in silico analysis confirmed that AFT1 and HEX-B also were enriched in endothelial cells when compared with non-endothelial cells. None of these genes have been reported previously to be involved in neovascularisation. However, our data establish that siRNA depletion of Rrm2, Atf1 or Hex-B had significant anti-angiogenic effects in VEGF-stimulated ex vivo mouse aortic ring assays. Overall, our results provide proof-of-principle that our approach can identify a cohort of

  1. Differences in gene expression profiles between human preimplantation embryos cultured in two different IVF culture media

    NARCIS (Netherlands)

    Kleijkers, S.H.M.; Eijssen, L.M.T.; Coonen, E.; Derhaag, J.G.; Mantikou, E.; Jonker, M.J.; Mastenbroek, S.; Repping, S.; Evers, J.L.H.; Dumoulin, J.C.M.; van Montfoort, A.P.A.

    2015-01-01

    STUDY QUESTION: Is gene expression in human preimplantation embryos affected by the medium used for embryo culture in vitro during an IVF treatment? SUMMARY ANSWER: Six days of in vitro culture of human preimplantation embryos resulted in medium-dependent differences in expression level of genes inv

  2. HER2 and COX2 expression in human prostate cancer.

    Science.gov (United States)

    Edwards, J; Mukherjee, R; Munro, A F; Wells, A C; Almushatat, A; Bartlett, J M S

    2004-01-01

    COX2 and HER2 expression are associated with a poor prognosis in prostate cancer and HER2 has been linked to COX2 expression in colorectal cancer. The association between COX2 and HER2 expression was investigated in 117 patients with prostate cancer (89) or Benign prostatic hyperplasia (BPH) (28). Tissue was analysed for HER2 amplification by fluorescent in situ hybridisation, and HER2 and COX2 protein expression by immunohistochemistry (IHC). All tumours analysed expressed COX2 at a significantly higher level than BPH tissue (P=0.041). Only low levels of HER2 gene amplification (8%, 7/89) and HER2 protein expression (12%, 11/89) were observed. HER2 protein expression was rarely observed and did not correlate with HER2 amplification or COX2 expression. Although HER2 does not drive COX2 expression in prostate cancer, this study identified high levels of COX2 expressed in locally advanced prostate cancer, suggesting COX2 could be a potential therapeutic target. COX2 inhibitors are currently being used in clinical trials for the treatment of other tumour types.

  3. A Human "eFP" Browser for Generating Gene Expression Anatograms.

    Science.gov (United States)

    Patel, Rohan V; Hamanishi, Erin T; Provart, Nicholas J

    2016-01-01

    Transcriptomic studies help to further our understanding of gene function. Human transcriptomic studies tend to focus on a particular subset of tissue types or a particular disease state; however, it is possible to collate into a compendium multiple studies that have been profiled using the same expression analysis platform to provide an overview of gene expression levels in many different tissues or under different conditions. In order to increase the knowledge and understanding we gain from such studies, intuitive visualization of gene expression data in such a compendium can be useful. The Human eFP ("electronic Fluorescent Pictograph") Browser presented here is a tool for intuitive visualization of large human gene expression data sets on pictographic representations of the human body as gene expression "anatograms". Pictographic representations for new data sets may be generated easily. The Human eFP Browser can also serve as a portal to other gene-specific information through link-outs to various online resources.

  4. Gene cloning of human soluble CD14 and its expression in eucaryotic cells

    Institute of Scientific and Technical Information of China (English)

    荫俊; 白洁; 王威; 宋伟; 王忠泽

    2002-01-01

    To express human soluble CD14 (sCD14) in eukaryotic cells. Methods: Human sCD14 cDNA was amplified from U937 cells with RT-PCR method. The recombinant expression plasmid pEF1/HisC/sCD14348aa was constructed and the expression in COS-7 cells was carried out using liposome transfection method. The yield was examined with scanning map identification. The expressed product was purified by immuno-affinity chromatography. Results: Sequence analysis demonstrated that the amplified gene sequence and those reported by documents were completely identical. sCD14 was expressed with high-yield. The expressed product was purified to above 90%. Recombinant sCD14, specifically combinable with endotoxins, had a natural biological activity. Conclusions: Human sCD14 was expressed in COS-7 cells, which laid a foundation for further study.

  5. The human protooncogene product p33pim is expressed during fetal hematopoiesis and in diverse leukemias.

    OpenAIRE

    Amson, R; Sigaux, F; Przedborski, S; Flandrin, G; Givol, D; Telerman, A

    1989-01-01

    We measured the human pim-1 protooncogene (PIM) expression during fetal development and in hematopoietic malignancies. Our data indicate that during human fetal hematopoiesis the 33-kDa pim product, p33pim, is highly expressed in the liver and spleen. In contrast, at the adult stage it is only slightly expressed in circulating granulocytes. Out of 70 hematopoietic malignancies analyzed, 51 patients and 19 cell lines, p33pim was overexpressed in approximately 30% of the samples, particularly i...

  6. Serial Analysis of Gene Expression: Applications in Human Studies

    OpenAIRE

    2004-01-01

    Serial analysis of gene expression (SAGE) is a powerful tool, which provides quantitative and comprehensive expression profile of genes in a given cell population. It works by isolating short fragments of genetic information from the expressed genes that are present in the cell being studied. These short sequences, called SAGE tags, are linked together for efficient sequencing. The frequency of each SAGE tag in the cloned multimers directly reflects the transcript abundance. Therefore, SAGE r...

  7. [VEGF gene expression in transfected human multipotent stromal cells].

    Science.gov (United States)

    Smirnikhina, S A; Lavrov, A V; Bochkov, N P

    2011-01-01

    Dynamics of VEGF gene expression in transfected multipotent stromal cells from adipose tissue was examined using electroporation and lipofection. Differences in the potency and dynamics of plasmid elimination (up to day 9) between cell cultures were observed. All cultures were divided into fast and slow plasmid-eliminating ones. Interculture differences in VEGF expression were detected. The possibility of a 5-6-fold increase of VEGF expression was shown. There were no differences in transfection potency, plasmid elimination dynamics, and VEGF expression after transfection by both nonviral methods.

  8. Expression of human apolipoprotein B and assembly of lipoprotein(a) in transgenic mice

    Energy Technology Data Exchange (ETDEWEB)

    Callow, M.J.; Stoltzfus, L.J.; Rubin, E.M. [Lawrence Berkeley Lab., CA (United States); Lawn, R.M. [Stanford Univ., CA (United States)

    1994-03-15

    The atherogenic macromolecule lipoprotein(a) [Lp(a)] has resisted in vivo analyses partly because it is found in a limited number of experimental animals. Although transgenic mice expressing human apolipoprotein (a) [apo(a)] have previously been described, they failed to assemble Lp(a) particles because of the inability of human apo(a) to associate with mouse apolipoprotein B (apoB). The authors isolated a 90-kilobase P1 phagemid containing the human apoB gene and with this DNA generated 13 lines of transgenic mice of which 11 expressed human apoB. The human apoB transcript was expressed and edited in the liver of the transgenic mice. Plasma concentrations of human apoB, as well as low density lipoprotein (LDL), were related to transgene copy number; the transgenic line with the most copies of human apoB had a >4-fold increase in LDL cholesterol compared with nontransgenics and a lipoprotein profile similar to that of humans. When human apoB and apo(a) transgenic mice were bred together, plasma apo(a) in mice expressing both human proteins was tightly associated with lipoproteins in the LDL density region. These studies demonstrate the successful expression of human apoB and the efficient assembly of Lp(a) in mice.

  9. Human BCAS3 expression in embryonic stem cells and vascular precursors suggests a role in human embryogenesis and tumor angiogenesis.

    Directory of Open Access Journals (Sweden)

    Kavitha Siva

    Full Text Available Cancer is often associated with multiple and progressive genetic alterations in genes that are important for normal development. BCAS3 (Breast Cancer Amplified Sequence 3 is a gene of unknown function on human chromosome 17q23, a region associated with breakpoints of several neoplasms. The normal expression pattern of BCAS3 has not been studied, though it is implicated in breast cancer progression. Rudhira, a murine WD40 domain protein that is 98% identical to BCAS3 is expressed in embryonic stem (ES cells, erythropoiesis and angiogenesis. This suggests that BCAS3 expression also may not be restricted to mammary tissue and may have important roles in other normal as well as malignant tissues. We show that BCAS3 is also expressed in human ES cells and during their differentiation into blood vascular precursors. We find that BCAS3 is aberrantly expressed in malignant human brain lesions. In glioblastoma, hemangiopericytoma and brain abscess we note high levels of BCAS3 expression in tumor cells and some blood vessels. BCAS3 may be associated with multiple cancerous and rapidly proliferating cells and hence the expression, function and regulation of this gene merits further investigation. We suggest that BCAS3 is mis-expressed in brain tumors and could serve as a human ES cell and tumor marker.

  10. High-level expression of a chemically synthesized gene for human interferon-gamma using a prokaryotic expression vector.

    OpenAIRE

    1984-01-01

    A chemically synthesized gene for human interferon-gamma has been cloned into a prokaryotic expression vector under the regulation of a synthetic constitutive transcriptional-translational control unit that contains a strong bacteriophage T5 early promoter and a strong ribosome-binding site. Cells harboring the recombinant plasmid express high levels (4 X 10(9) units per liter of culture) of antiviral activity specific for interferon-gamma. Analysis of total cell lysates on NaDodSO4/polyacryl...

  11. Heat shock protein 27 expression in the human testis showing normal and abnormal spermatogenesis.

    Science.gov (United States)

    Adly, Mohamed A; Assaf, Hanan A; Hussein, Mahmoud Rezk A

    2008-10-01

    Heat shock proteins (HSPs) are molecular chaperones involved in protein folding, assembly and transport, and which play critical roles in the regulation of cell growth, survival and differentiation. We set out to test the hypothesis that HSP27 protein is expressed in the human testes and its expression varies with the state of spermatogenesis. HSP27 expression was examined in 30 human testicular biopsy specimens (normal spermatogenesis, maturation arrest and Sertoli cell only syndrome, 10 cases each) using immunofluorescent methods. The biopsies were obtained from patients undergoing investigations for infertility. The seminiferous epithelium of the human testes showing normal spermatogenesis had a cell type-specific expression of HSP27. HSP27 expression was strong in the cytoplasm of the Sertoli cells, spermatogonia, and Leydig cells. Alternatively, the expression was moderate in the spermatocytes, weak in the spermatids and absent in the spermatozoa. In testes showing maturation arrest, HSP27 expression was strong in the Sertoli cells, weak in the spermatogonia, and spermatocytes. It was absent in the spermatids and Leydig cells. In Sertoli cell only syndrome, HSP27 expression was strong in the Sertoli cells and absent in the Leydig cells. We report for the first time the expression patterns of HSP27 in the human testes and show differential expression during normal spermatogenesis, indicating a possible role in this process. The altered expression of this protein in testes showing abnormal spermatogenesis may be related to the pathogenesis of male infertility.

  12. Heterologously expressed bacterial and human multidrug resistance proteins confer cadmium resistance to Escherichia coli

    NARCIS (Netherlands)

    Achard-Joris, M; van Saparoea, HBV; Driessen, AJM; Bourdineaud, JP; Bourdineaud, Jean-Paul

    2005-01-01

    The human MDR1 gene is induced by cadmium exposure although no resistance to this metal is observed in human cells overexpressing hMDR1. To access the role of MDR proteins in cadmium resistance, human MDR1, Lactococcus lactis lmrA, and Oenococcus oeni omrA were expressed in an Escherichia coli tolC

  13. CXCL12/SDF-1 over-expression in human insulinomas and its biological relevance

    DEFF Research Database (Denmark)

    Ilhan, Aysegul; Nabokikh, Anastasiya; Maj, Magdalena

    2009-01-01

    This study was performed on the basis of previously obtained investigative gene array data concerning the over-expression of CXCL12/SDF-1 in human insulinomas versus human pancreatic islet preparations. The presence of CXCL12/SDF-1 was studied by RT-qPCR in human insulinomas (n=8) versus pancreatic...

  14. FcRn expression, ligands binding properties and its regulation in human immune cells and hepatocytes

    OpenAIRE

    2007-01-01

    ABSTRACT Expression and diverse functions of MHC class I related neonatal Fc receptor in different tissues is continually reported. To contribute to the understanding of how the receptor functions according to cell type, we investigated the expression and ligands binding properties of FcRn in human immune cells and hepatocytes. Here, we report that heterodimeric FcRn is expressed in these cells as evidenced by RT-PCR, Western immunoblottting and flow cytometry. The receptor expression i...

  15. Cardiac protein kinases: the cardiomyocyte kinome and differential kinase expression in human failing hearts

    OpenAIRE

    Fuller, Stephen J.; Osborne, Sally A.; Leonard, Sam J.; Hardyman, Michelle A.; Vaniotis, George; Allen, Bruce G.; Sugden, Peter H.; Clerk, Angela

    2015-01-01

    Aims. Protein kinases are potential therapeutic targets for heart failure, but most studies of cardiac protein kinases derive from other systems, an approach that fails to account for specific kinases expressed in the heart and the contractile cardiomyocytes. We aimed to define the cardiomyocyte kinome (i.e. the protein kinases expressed in cardiomyocytes) and identify kinases with altered expression in human failing hearts. Methods and Results. Expression profiling (Affymetrix microarrays) d...

  16. Determinants of ABH expression on human blood platelets.

    Science.gov (United States)

    Cooling, Laura L W; Kelly, Kathleen; Barton, James; Hwang, Debbie; Koerner, Theodore A W; Olson, John D

    2005-04-15

    Platelets express ABH antigens, which can adversely effect platelet transfusion recovery and survival in ABH-incompatible recipients. To date, there has been no large, comprehensive study comparing specific donor factors with ABH expression on platelet membranes and glycoconjugates. We studied ABH expression in 166 group A apheresis platelet donors by flow cytometry, Western blotting, and thin layer chromatography relative to donor age, sex, A1/A2 subgroup, and Lewis phenotype. Overall, A antigen on platelet membranes, glycoproteins, and glycosphingolipids was linked to an A1 red blood cell (RBC) phenotype. Among A1 donors, platelet ABH varied significantly between donors (0%-87%). Intradonor variability, however, was minimal, suggesting that platelet ABH expression is a stable, donor-specific characteristic, with 5% of A1 donors typing as either ABH high- or low-expressers. Group A2 donors, in contrast, possessed a Bombay-like phenotype, lacking both A and H antigens. Unlike RBCs, ABH expression on platelets may be determined primarily by H-glycosyltransferase (FUT1) activity. Identification of A2 and A1 low expressers may increase the availability and selection of crossmatched and HLA-matched platelets. Platelets from group A2 may also be a superior product for patients undergoing A/O major mismatch allogeneic progenitor cell transplantation.

  17. Effects of Aging and Anatomic Location on Gene Expression in Human Retina

    Directory of Open Access Journals (Sweden)

    Hui eCai

    2012-05-01

    Full Text Available Objective: To determine the effects of age and topographic location on gene expression in human neural retina.Methods: Macular and peripheral neural retina RNA were isolated from human donor eyes for DNA microarray and quantitative RT-PCR analyses.Results: Total RNA integrity from human donors was preserved. Hierarchical clustering analysis demonstrates that the gene expression profiles of young, old, macula and peripheral retina cluster into four distinct groups. Genes which are highly expressed in macular, peripheral, young or old retina were identified, including inhibitors of Wnt Signaling Pathway (DKK1, FZD10 and SFRP2 which are preferably expressed in the periphery. Conclusions: The transcriptome of the human retina is affected by age and topographic location. Wnt pathway inhibitors in the periphery may maintain peripheral retinal cells in an undifferentiated state. Understanding the effects of age and topographic location on gene expression may lead to the development of new therapeutic interventions for age-related eye diseases.

  18. Expression of hSef in various human tissues and cell lines

    Institute of Scientific and Technical Information of China (English)

    Huang Guanrong; Xiong Shiqin; Zhao Qiuhui; Wang Yinyin; Reng Fangli; Ye Xiongjun; Chang Zhijie

    2006-01-01

    Sef is a transmembrane protein inhibiting FGF signaling.To determine the correlation of Sef with human diseases,Sef expression patterns were observed in cell lines and human cancer tissues.Western blot using anti-hSef antibodies showed that hSef,when expressed in Cos7 cells gave a molecular mass of 100 KD as compared with 80 KD in an in vitro translation assay suggesting occurrence of glycosylation at the potential N-linked glycosylation sites in the extracellular domain.Northern blot showed that hSef was mainly expressed in human kidney and testis.RT-PCR analysis showed a widely spread expression pattern in several cell lines.Immunohistochemical analysis revealed ahigh expression level of hSef in kidney,testis,and the corresponding carcinoma tissues.Results demonstrated that Sef might be up-regulated in the cancer tissues suggesting a possible role of Sef in pathophysiology of human diseases.

  19. Expression of cytosolic 5' nucleotidase does not correlate with expression of oxidative metabolism marker: myoglobine in human skeletal muscles

    Institute of Scientific and Technical Information of China (English)

    Katarzyna Lechward; Kinga Tkacz-Stachowska

    2009-01-01

    Our previous studies had shown that eytosolic 5'nucleotidase-I (eN-I) is expressed in several tissues in pigeons, including brain and several different skeletal muscles. We observed that cN-I mRNA levels varied among different pigeon muscles. Initial quantification of the differences revealed that ~5-10 times more of cN-I transcript was present in red, oxidative muscles (breast muscle and gastrocnemius) than in white ones,composed of glycolytic fibers (biceps brachii). We had found this observation very intriguing and decided to compare human skeletal muscles distribution of cN-I with the type of oxygen metabolism. Our screen involved 60 samples of several human muscles and we assayed the correlation between the amount of tran-scripts of cN-I and myoglobine, which we took as a measure of oxidative-slow twitch fibers. Our question was whether in humans, cN-I presence in skeletal muscles was related to their fiber composition. If that was the case, then cN-I expression could serve as a tool to assess the percentage of oxidative fibers in any given human muscle sample, where myogiobine expression could not be readily measured. After quantification of expression of both genes, we concluded that there was no correlation between expression of cN-I and fiber type. Therefore, contrary to the pigeon muscles, cN-Idid not reflect the ratio of oxidative fibers to the total mass of the muscle sample in humans. That difference indicated that there were certain mechanisms that dif-ferentially regulated the expression of cN-I in muscle tissues of mammals and lower vertebrates.

  20. Transgenic silkworms expressing human insulin receptors for evaluation of therapeutically active insulin receptor agonists.

    Science.gov (United States)

    Matsumoto, Yasuhiko; Ishii, Masaki; Ishii, Kenichi; Miyaguchi, Wataru; Horie, Ryo; Inagaki, Yoshinori; Hamamoto, Hiroshi; Tatematsu, Ken-ichiro; Uchino, Keiro; Tamura, Toshiki; Sezutsu, Hideki; Sekimizu, Kazuhisa

    2014-12-12

    We established a transgenic silkworm strain expressing the human insulin receptor (hIR) using the GAL4/UAS system. Administration of human insulin to transgenic silkworms expressing hIR decreased hemolymph sugar levels and facilitated Akt phosphorylation in the fat body. The decrease in hemolymph sugar levels induced by injection of human insulin in the transgenic silkworms expressing hIR was blocked by co-injection of wortmannin, a phosphoinositide 3-kinase inhibitor. Administration of bovine insulin, an hIR ligand, also effectively decreased sugar levels in the transgenic silkworms. These findings indicate that functional hIRs that respond to human insulin were successfully induced in the transgenic silkworms. We propose that the humanized silkworm expressing hIR is useful for in vivo evaluation of the therapeutic activities of insulin receptor agonists. Copyright © 2014 Elsevier Inc. All rights reserved.

  1. Human pancreatic triglyceride lipase expressed in yeast cells: purification and characterization.

    Science.gov (United States)

    Yang, Y; Lowe, M E

    1998-06-01

    A cDNA clone encoding human pancreatic triglyceride lipase was cloned into a yeast expression vector so that the yeast PHO1 signal peptide replaced the native signal peptide. Pichia pastoris cells were transfected with the vector, and clones expressing human pancreatic triglyceride lipase were isolated. Recombinant human pancreatic lipase was expressed in broth cultures and was purified from the medium by DEAE blue Sepharose and hydroxyapatite chromatography. The highly purified lipase had specific activities for various triglyceride substrates identical to those of tissue-purified human pancreatic triglyceride lipase; it was inhibited by bile salts, required colipase for activity, and demonstrated interfacial activation. This expression system is suitable for the rapid, efficient production of human pancreatic triglyceride lipase in amounts adequate for biophysical studies.

  2. Activated human mast cells induce LOX-1-specific scavenger receptor expression in human monocyte-derived macrophages.

    Directory of Open Access Journals (Sweden)

    Mervi Alanne-Kinnunen

    Full Text Available Activated mast cells in atherosclerotic lesions degranulate and release bioactive compounds capable of regulating atherogenesis. Here we examined the ability of activated human primary mast cells to regulate the expression of the major scavenger receptors in cultured human primary monocyte-derived macrophages (HMDMs.Components released by immunologically activated human primary mast cells induced a transient expression of lectin-like oxidized LDL receptor (LOX-1 mRNA in HMDMs, while the expression of two other scavenger receptors, MSR1 and CD36, remained unaffected. The LOX-1-inducing secretory components were identified as histamine, tumor necrosis factor alpha (TNF-α, and transforming growth factor beta (TGF-β1, which exhibited a synergistic effect on LOX-1 mRNA expression. Histamine induced a transient expression of LOX-1 protein. Mast cell -induced increase in LOX-1 expression was not associated with increased uptake of oxidized LDL by the macrophages.Mast cell-derived histamine, TNF-α, and TGF-β1 act in concert to induce a transient increase in LOX-1 expression in human primary monocyte-derived macrophages. The LOX-1-inducing activity potentially endows mast cells a hitherto unrecognized role in the regulation of innate immune reactions in atherogenesis.

  3. How is mRNA expression predictive for protein expression? A correlation study on human circulating monocytes

    Institute of Scientific and Technical Information of China (English)

    Yanfang Guo; Yuan Chen; Hui Jiang; Lijun Tan; Jingyun Xie; Xuezhen Zhu; Songping Liang; Hongwen Deng; Peng Xiao; Shufeng Lei; Feiyan Deng; Gary Guishan Xiao; Yaozhong Liu; Xiangding Chen; Liming Li; Shan Wu

    2008-01-01

    A key assumption in studying mRNA expression is that it is informative in the prediction of protein expression. However,only limited studies have explored the mRNA-protein expression correlation in yeast or human tissues and the results have been relatively inconsistent. We carried out correlation analyses on mRNA-protein expressions in freshly isolated human circulating monocytes from 30 unrelated women. The expressed proteins for 71 genes were quantified and identified by 2-D electrophoresis coupled with mass spectrometry. The corresponding mRNA expressions were quantified by Affymetrix gene chips. Significant correlation (r=0.235, P<0.0001) was observed for the whole dataset including all studied genes and all samples. The correlations varied in different biological categories of gene ontology. For example, the highest correlation was achieved for genes of the extracellular region in terms of cellular component (r=0.643, P<0.0001) and the lowest correlation was obtained for genes of regulation (r=0.099, P=0.213) in terms of biological process. In the genome, half of the samples showed significant positive correlation for the 71 genes and significant correlation was found between the average mRNA and the average protein expression levels in all samples (r=0.296, P<0.01). However, at the study group level, only five studied genes had significant positive correlation across all the samples. Our results showed an overall positive correlation between mRNA and protein expression levels.However, the moderate and varied correlations suggest that mRNA expression might be sometimes useful, but certainly far from perfect, in predicting protein expression levels.

  4. Highly expressed genes in human high grade gliomas: immunohistochemical analysis of data from the Human Protein Atlas

    Directory of Open Access Journals (Sweden)

    Michael A. Meyer

    2014-06-01

    Full Text Available Gene expression within human glioblastomas were analyzed from data on 20,083 genes entered into the on-line Human Protein Atlas. In selecting genes that are strongly expressed within normal human brain tissue, 58 genes were identified from a search of the 20,083 entries that were rated as showing 90% or greater intensity of expression within normal brain tissues. Of these 58, a subset of 48 genes was identified that not only had expression data for human glioblastomas but also for the human glioblastoma cell line U-251. Four of these 48 selected genes were found to be strongly expressed within the cytoplasm when assessed by both histologic sampling of high grade glioma patient cases as well as U-251 glioblastoma cell line immunofluoresence analysis. These four human genes are: AGBL2 (ATP/GTP binding protein-like 2, BLOC1S6 (biogenesis of lysosomal organelles complex-1, subunit 6, MAP1A (microtubule-associated protein 1A and ZSWIM5 (zinc finger, SWIM-type containing 5, also known as KIAA1511. Further research is advocated to investigate the role of ZSWIM5 and AGBL2 in glioma cell biology.

  5. Recognition of Expressions on Human Face using AI Techniques

    Directory of Open Access Journals (Sweden)

    Arpita Nagpal

    2011-08-01

    Full Text Available Facial expressions convey non-verbal cues, which play animportant role in interpersonal relations. Facial expressionsrecognition technology helps in designing an intelligent humancomputer interfaces. This paper discusses a three phase techniquefor the facial expression recognition of the Indian faces. In thefirst phase the faces are tracked using Haar classifier in the livevideos of Indian student’s community. In the second phase 38facial feature points are detected using Active Appearance Model(AAM technique. In the last step the support vector machine(SVM is used to classify four primary facial expression.Integrating these broader techniques and obtaining a reasonablygood performance is a very big challenge. The performance ofthe proposed facial expressions recognizer is 82.7%.

  6. Expression of human interferon gamma in Brassica napus seeds

    African Journals Online (AJOL)

    TUOYO

    2010-08-09

    Aug 9, 2010 ... Accepted 8 June, 2010. Expressions of heterologous proteins in suitable plant tissues and targeting it into subcellular ... in plant and animal cells (Fischer et al., 2000). ..... Comparison of dot blot results in transgenic and wild-.

  7. Increased cortical expression of two synaptogenic thrombospondins in human brain evolution.

    Science.gov (United States)

    Cáceres, Mario; Suwyn, Carolyn; Maddox, Marcelia; Thomas, James W; Preuss, Todd M

    2007-10-01

    Thrombospondins are extracellular-matrix glycoproteins implicated in the control of synaptogenesis and neurite growth. Previous microarray studies suggested that one gene of this family, thrombospondin 4 (THBS4), was upregulated during human brain evolution. Using independent techniques to examine thrombospondin expression patterns in adult brain samples, we report approximately 6-fold and approximately 2-fold greater expression of THBS4 and THBS2 messenger RNA (mRNA), respectively, in human cerebral cortex compared with chimpanzees and macaques, with corresponding differences in protein levels. In humans and chimpanzees, thrombospondin expression differences were observed in the forebrain (cortex and caudate), whereas the cerebellum and most nonbrain tissues exhibited similar levels of the 2 mRNAs. Histological examination revealed THBS4 mRNA and protein expression in numerous pyramidal and glial cells in the 3 species but humans also exhibited very prominent immunostaining of the synapse-rich cortical neuropil. In humans, additionally, THBS4 antibodies labeled beta-amyloid containing plaques in Alzheimer's cases and some control cases. This is the first detailed characterization of gene-expression changes in human evolution that involve specific brain regions, including portions of cerebral cortex. Increased expression of thrombospondins in human brain evolution could result in changes in synaptic organization and plasticity, and contribute to the distinctive cognitive abilities of humans, as well as to our unique vulnerability to neurodegenerative disease.

  8. Diversity of human and mouse homeobox gene expression in development and adult tissues.

    Science.gov (United States)

    Dunwell, Thomas L; Holland, Peter W H

    2016-11-03

    Homeobox genes encode a diverse set of transcription factors implicated in a vast range of biological processes including, but not limited to, embryonic cell fate specification and patterning. Although numerous studies report expression of particular sets of homeobox genes, a systematic analysis of the tissue specificity of homeobox genes is lacking. Here we analyse publicly-available transcriptome data from human and mouse developmental stages, and adult human tissues, to identify groups of homeobox genes with similar expression patterns. We calculate expression profiles for 242 human and 278 mouse homeobox loci across a combination of 59 human and 12 mouse adult tissues, early and late developmental stages. This revealed 20 human homeobox genes with widespread expression, primarily from the TALE, CERS and ZF classes. Most homeobox genes, however, have greater tissue-specificity, allowing us to compile homeobox gene expression lists for neural tissues, immune tissues, reproductive and developmental samples, and for numerous organ systems. In mouse development, we propose four distinct phases of homeobox gene expression: oocyte to zygote; 2-cell; 4-cell to blastocyst; early to mid post-implantation. The final phase change is marked by expression of ANTP class genes. We also use these data to compare expression specificity between evolutionarily-based gene classes, revealing that ANTP, PRD, LIM and POU homeobox gene classes have highest tissue specificity while HNF, TALE, CUT and CERS are most widely expressed. The homeobox genes comprise a large superclass and their expression patterns are correspondingly diverse, although in a broad sense related to an evolutionarily-based classification. The ubiquitous expression of some genes suggests roles in general cellular processes; in contrast, most human homeobox genes have greater tissue specificity and we compile useful homeobox datasets for particular tissues, organs and developmental stages. The identification of a

  9. Gene expression profile analysis of human intervertebral disc degeneration

    OpenAIRE

    Kai Chen; Dajiang Wu; Xiaodong Zhu; Haijian Ni; Xianzhao Wei; Ningfang Mao; Yang Xie; Yunfei Niu; Ming Li

    2013-01-01

    In this study, we used microarray analysis to investigate the biogenesis and progression of intervertebral disc degeneration. The gene expression profiles of 37 disc tissue samples obtained from patients with herniated discs and degenerative disc disease collected by the National Cancer Institute Cooperative Tissue Network were analyzed. Differentially expressed genes between more and less degenerated discs were identified by significant analysis of microarray. A total of 555 genes were signi...

  10. Gene expression and functional annotation of the human and mouse choroid plexus epithelium.

    Directory of Open Access Journals (Sweden)

    Sarah F Janssen

    Full Text Available BACKGROUND: The choroid plexus epithelium (CPE is a lobed neuro-epithelial structure that forms the outer blood-brain barrier. The CPE protrudes into the brain ventricles and produces the cerebrospinal fluid (CSF, which is crucial for brain homeostasis. Malfunction of the CPE is possibly implicated in disorders like Alzheimer disease, hydrocephalus or glaucoma. To study human genetic diseases and potential new therapies, mouse models are widely used. This requires a detailed knowledge of similarities and differences in gene expression and functional annotation between the species. The aim of this study is to analyze and compare gene expression and functional annotation of healthy human and mouse CPE. METHODS: We performed 44k Agilent microarray hybridizations with RNA derived from laser dissected healthy human and mouse CPE cells. We functionally annotated and compared the gene expression data of human and mouse CPE using the knowledge database Ingenuity. We searched for common and species specific gene expression patterns and function between human and mouse CPE. We also made a comparison with previously published CPE human and mouse gene expression data. RESULTS: Overall, the human and mouse CPE transcriptomes are very similar. Their major functionalities included epithelial junctions, transport, energy production, neuro-endocrine signaling, as well as immunological, neurological and hematological functions and disorders. The mouse CPE presented two additional functions not found in the human CPE: carbohydrate metabolism and a more extensive list of (neural developmental functions. We found three genes specifically expressed in the mouse CPE compared to human CPE, being ACE, PON1 and TRIM3 and no human specifically expressed CPE genes compared to mouse CPE. CONCLUSION: Human and mouse CPE transcriptomes are very similar, and display many common functionalities. Nonetheless, we also identified a few genes and pathways which suggest that the CPE

  11. Gene expression analysis of precision-cut human liver slices indicate stable expression of hepatotoxicity related genes

    NARCIS (Netherlands)

    Elferink, Maria; Olinga, Peter; van Leeuwen, E. M.; Bauerschmidt, S.; Polman, J.; Schoonen, W. G.; Heisterkamp, S. H.; Groothuis, Genoveva

    2011-01-01

    In the process of drug development it is of high importance to test the safety of new drugs with predictive value for human toxicity. A promising approach of toxicity testing is based on changes in the gene expression profile of the liver. Toxicity screening based on animal liver cells cannot be dir

  12. Gene expression analysis of precision cut human liver slices indicate stable expression of ADME-Tox related genes

    NARCIS (Netherlands)

    Elferink, M.; Olinga, P.; Van Leeuwen, E.; Bauerschmidt, S.; Polman, J.; Schoonen, W.; Heisterkamp, S.; Groothuis, G.

    2010-01-01

    In the process of drug development it is of high importance to test the safety of new drugs with predictive value for human toxicity. Currently, for toxicity studies toxicogenomic analysis of changes in gene expression profile of the liver is increasingly applied. Toxicity screening based on animal

  13. Gene expression analysis of precision-cut human liver slices indicates stable expression of ADME-Tox related genes

    NARCIS (Netherlands)

    Elferink, M. G. L.; Olinga, P.; van Leeuwen, E. M.; Bauerschmidt, S.; Polman, J.; Schoonen, W. G.; Heisterkamp, S. H.; Groothuis, G. M. M.

    2011-01-01

    In the process of drug development it is of high importance to test the safety of new drugs with predictive value for human toxicity. A promising approach of toxicity testing is based on shifts in gene expression profiling of the liver. Toxicity screening based on animal liver cells cannot be direct

  14. Differential expression of sphingolipids in P-glycoprotein or multidrug resistance-related protein 1 expressing human neuroblastoma cell lines

    NARCIS (Netherlands)

    Dijkhuis, AJ; Douwes, J; Kamps, W; Sietsma, H; Kok, JW

    2003-01-01

    The sphingolipid composition and multidrug resistance status of three human neuroblastoma cell lines were established. SK-N-FI cells displayed high expression and functional (efflux) activity of P-glycoprotein, while multidrug resistance-related protein 1 was relatively abundant and most active in S

  15. Differential expression of sphingolipids in P-glycoprotein or multidrug resistance-related protein 1 expressing human neuroblastoma cell lines

    NARCIS (Netherlands)

    Dijkhuis, AJ; Douwes, J; Kamps, W; Sietsma, H; Kok, JW

    2003-01-01

    The sphingolipid composition and multidrug resistance status of three human neuroblastoma cell lines were established. SK-N-FI cells displayed high expression and functional (efflux) activity of P-glycoprotein, while multidrug resistance-related protein 1 was relatively abundant and most active in

  16. Human cancer cells express Slug-based epithelial-mesenchymal transition gene expression signature obtained in vivo

    Directory of Open Access Journals (Sweden)

    Anastassiou Dimitris

    2011-12-01

    Full Text Available Abstract Background The biological mechanisms underlying cancer cell motility and invasiveness remain unclear, although it has been hypothesized that they involve some type of epithelial-mesenchymal transition (EMT. Methods We used xenograft models of human cancer cells in immunocompromised mice, profiling the harvested tumors separately with species-specific probes and computationally analyzing the results. Results Here we show that human cancer cells express in vivo a precise multi-cancer invasion-associated gene expression signature that prominently includes many EMT markers, among them the transcription factor Slug, fibronectin, and α-SMA. We found that human, but not mouse, cells express the signature and Slug is the only upregulated EMT-inducing transcription factor. The signature is also present in samples from many publicly available cancer gene expression datasets, suggesting that it is produced by the cancer cells themselves in multiple cancer types, including nonepithelial cancers such as neuroblastoma. Furthermore, we found that the presence of the signature in human xenografted cells was associated with a downregulation of adipocyte markers in the mouse tissue adjacent to the invasive tumor, suggesting that the signature is triggered by contextual microenvironmental interactions when the cancer cells encounter adipocytes, as previously reported. Conclusions The known, precise and consistent gene composition of this cancer mesenchymal transition signature, particularly when combined with simultaneous analysis of the adjacent microenvironment, provides unique opportunities for shedding light on the underlying mechanisms of cancer invasiveness as well as identifying potential diagnostic markers and targets for metastasis-inhibiting therapeutics.

  17. Re-evaluation of the prolactin receptor expression in human breast cancer

    DEFF Research Database (Denmark)

    Galsgaard, Elisabeth Douglas; Rasmussen, Birgitte Bruun; Folkesson, Charlotta Grånäs;

    2009-01-01

    and decidual cells in tissue sections of human placenta. Screening of 160 mammary adenocarcinomas demonstrated significant immunoreactivity in only four tumours, indicating that PRLR is generally not strongly upregulated in human breast cancer. However, even a very low level of PRLR expression was found......The pituitary hormone PRL is involved in tumorigenesis in rodents and humans. PRL promotes proliferation, survival and migration of cancer cells acting via the PRL receptor (PRLR). Aiming to perform a large-scale immunohistochemical (IHC) screening of human mammary carcinomas for PRLR expression...... specificity for PRLR and to rather recognise a PRLR-associated protein. The mAb U5 raised against the rat PRLR did not cross-react with the human receptor. Only one mAb, 1A2B1, was found useful for detection of PRLR in IHC applications. This antibody recognised PRLR expressed in human breast cancer cell lines...

  18. A Progenitor Cell Expressing Transcription Factor RORγt Generates All Human Innate Lymphoid Cell Subsets.

    Science.gov (United States)

    Scoville, Steven D; Mundy-Bosse, Bethany L; Zhang, Michael H; Chen, Li; Zhang, Xiaoli; Keller, Karen A; Hughes, Tiffany; Chen, Luxi; Cheng, Stephanie; Bergin, Stephen M; Mao, Hsiaoyin C; McClory, Susan; Yu, Jianhua; Carson, William E; Caligiuri, Michael A; Freud, Aharon G

    2016-05-17

    The current model of murine innate lymphoid cell (ILC) development holds that mouse ILCs are derived downstream of the common lymphoid progenitor through lineage-restricted progenitors. However, corresponding lineage-restricted progenitors in humans have yet to be discovered. Here we identified a progenitor population in human secondary lymphoid tissues (SLTs) that expressed the transcription factor RORγt and was unique in its ability to generate all known ILC subsets, including natural killer (NK) cells, but not other leukocyte populations. In contrast to murine fate-mapping data, which indicate that only ILC3s express Rorγt, these human progenitor cells as well as human peripheral blood NK cells and all mature ILC populations expressed RORγt. Thus, all human ILCs can be generated through an RORγt(+) developmental pathway from a common progenitor in SLTs. These findings help establish the developmental signals and pathways involved in human ILC development.

  19. Lineage-specific expression of bestrophin-2 and bestrophin-4 in human intestinal epithelial cells

    DEFF Research Database (Denmark)

    Ito, Go; Okamoto, Ryuichi; Murano, Tatsuro

    2013-01-01

    Intestinal epithelial cells (IECs) regulate the absorption and secretion of anions, such as HCO3(-) or Cl(-). Bestrophin genes represent a newly identified group of calcium-activated Cl(-) channels (CaCCs). Studies have suggested that, among the four human bestrophin-family genes, bestrophin-2...... (BEST2) and bestrophin-4 (BEST4) might be expressed within the intestinal tissue. Consistently, a study showed that BEST2 is expressed by human colonic goblet cells. However, their precise expression pattern along the gastrointestinal tract, or the lineage specificity of the cells expressing these genes...

  20. Survivin expression in canine epidermis and in canine and human cutaneous squamous cell carcinomas.

    Science.gov (United States)

    Bongiovanni, Laura; Colombi, Isabella; Fortunato, Carmine; Della Salda, Leonardo

    2009-10-01

    Survivin, a member of the inhibitor of apoptosis protein (IAP) family, is ubiquitously expressed during tissue development, undetectable in most normal tissues, but re-expressed in most cancers, including skin malignancies. Expression of survivin was evaluated retrospectively in 19 canine cutaneous squamous cell carcinomas (SCCs; one in situ; 16 well differentiated; one invasive, one lymph node metastasis) and 19 well differentiated SCCs from human beings. Seven specimens of normal canine skin were included. Immunohistochemical expression of full-length survivin was determined using a commercially available antibody. In addition, apoptotic rate [Terminal deoxynucleotidyl Transferase Biotin-dUTP Nick End Labelling index (TUNEL) index] and mitotic index (MI), counting mitoses in 10 high power fields (HPF), were determined. Scattered survivin positive nuclei were identified in the epidermal basal cell layer of normal canine skin. Nuclear survivin expression was identified in 18 of 19 human and in all canine SCCs, mainly along the base of the tumour cell population. Cytoplasmic survivin expression was rarely observed in human SCCs and in 84.2% of canine SCCs. The TUNEL index ranged from 0.1 to 2.6 in human beings and from 7.5 to 69.4 in dogs, while MIs ranged from 0 to 4 in human beings and dogs. No correlation was found between survivin expression and apoptotic or mitotic rates. Canine and human tumours showed similar nuclear survivin expression, indicating similar functions of the molecule. We demonstrated survivin expression in normal adult canine epidermis. Increased nuclear survivin expression in pre-neoplastic and neoplastic lesions demonstrates a possible association of survivin with development of SCCs in human beings and dogs.

  1. EXPRESSION OF FLIP IN HUMAN COLON CARCINOMAS:A NEW MECHANISM OF IMMUNE EVASION

    Institute of Scientific and Technical Information of China (English)

    XING Bao-cai; S. Wimmenauer; EH. Farthmann

    2005-01-01

    Objective: It has been proposed that Fas ligand (FasL) may play an important role in immune escape of tumors and FLIP is an important mediator of Fas/FasL pathway. In this study, the expression of FLIP was determined in human colon carcinoma cell lines and tissue to investigate the new mechanism of immune evasion of human colon carcinomas. Methods:RT-PCR and immunohistochemistry (IHC) were performed to investigate the expression of FLIP in human colon carcinoma cell lines SW480, LS174 and twenty human primary colon carcinoma specimens. Results: It was shown that SW480 cells,LS174 cells and primary colon carcinoma specimen constitutively expressed FLIP at the mRNA and protein level. The expression of FLIP was not found in the epithelial cells of normal colon mucosa. Conclusion: FLIP was expressed in human primary colon carcinoma specimens but not in the normal counterpart. It suggested that the expression of FLIP may occur during the malignant transformation from normal colon epithelial cells to colon carcinoma cells. Tumor cells might obtain the ability to resist the Fas-mediated apoptosis by expressing FLIP. The expression of FLIP might contribute to the formation of colon carcinomas.

  2. Ki-67 Expression in Human Tumors Measured by Flow Cytometry

    Science.gov (United States)

    1990-01-01

    proliferation. One of the first proliferation antigens to be studied was the transferrin receptor (TfR). Proliferating normal and tumor cells require iron and...obtained by incubation in NP-40. When the antibody was used to stain frozen sections of human tonsil , the chromosomes were stained. The antibody was...proliferation. When applied to frozen sections of human tonsil , the antibody appeared to be reactive with a mitotic spindle-associated protein. It bound

  3. Human population-specific gene expression and transcriptional network modification with polymorphic transposable elements.

    Science.gov (United States)

    Wang, Lu; Rishishwar, Lavanya; Mariño-Ramírez, Leonardo; Jordan, I King

    2016-12-19

    Transposable element (TE) derived sequences are known to contribute to the regulation of the human genome. The majority of known TE-derived regulatory sequences correspond to relatively ancient insertions, which are fixed across human populations. The extent to which human genetic variation caused by recent TE activity leads to regulatory polymorphisms among populations has yet to be thoroughly explored. In this study, we searched for associations between polymorphic TE (polyTE) loci and human gene expression levels using an expression quantitative trait loci (eQTL) approach. We compared locus-specific polyTE insertion genotypes to B cell gene expression levels among 445 individuals from 5 human populations. Numerous human polyTE loci correspond to both cis and trans eQTL, and their regulatory effects are directly related to cell type-specific function in the immune system. PolyTE loci are associated with differences in expression between European and African population groups, and a single polyTE loci is indirectly associated with the expression of numerous genes via the regulation of the B cell-specific transcription factor PAX5 The polyTE-gene expression associations we found indicate that human TE genetic variation can have important phenotypic consequences. Our results reveal that TE-eQTL are involved in population-specific gene regulation as well as transcriptional network modification.

  4. Human population-specific gene expression and transcriptional network modification with polymorphic transposable elements

    Science.gov (United States)

    Wang, Lu; Mariño-Ramírez, Leonardo

    2017-01-01

    Abstract Transposable element (TE) derived sequences are known to contribute to the regulation of the human genome. The majority of known TE-derived regulatory sequences correspond to relatively ancient insertions, which are fixed across human populations. The extent to which human genetic variation caused by recent TE activity leads to regulatory polymorphisms among populations has yet to be thoroughly explored. In this study, we searched for associations between polymorphic TE (polyTE) loci and human gene expression levels using an expression quantitative trait loci (eQTL) approach. We compared locus-specific polyTE insertion genotypes to B cell gene expression levels among 445 individuals from 5 human populations. Numerous human polyTE loci correspond to both cis and trans eQTL, and their regulatory effects are directly related to cell type-specific function in the immune system. PolyTE loci are associated with differences in expression between European and African population groups, and a single polyTE loci is indirectly associated with the expression of numerous genes via the regulation of the B cell-specific transcription factor PAX5. The polyTE-gene expression associations we found indicate that human TE genetic variation can have important phenotypic consequences. Our results reveal that TE-eQTL are involved in population-specific gene regulation as well as transcriptional network modification. PMID:27998931

  5. Yes-associated protein 1 is widely expressed in human brain tumors and promotes glioblastoma growth.

    Science.gov (United States)

    Orr, Brent A; Bai, Haibo; Odia, Yazmin; Jain, Deepali; Anders, Robert A; Eberhart, Charles G

    2011-07-01

    The hippo pathway and its downstream mediator yes-associated protein 1 (YAP1) regulate mammalian organ size in part through modulating progenitor cell numbers. YAP1 has also been implicated as an oncogene in multiple human cancers. Currently, little is known about the expression of YAP1 either in normal human brain tissue or in central nervous system neoplasms. We used immunohistochemistry to evaluate nuclear YAP1 expression in the fetal and normal adult human brains and in 264 brain tumors. YAP1 was expressed in fetal and adult brain regions known to harbor neural progenitor cells, but there was little YAP1 immunoreactivity in the adult cerebral cortex. YAP1 protein was also readily detected in the nuclei of human brain tumors. In medulloblastoma, the expression varied between histologic subtypes and was most prominent in nodular/desmoplastic tumors. In gliomas, it was frequently expressed in infiltrating astrocytomas and oligodendrogliomas but rarely in pilocytic astrocytomas. Using a loss-of-function approach, we show that YAP1 promoted growth of glioblastoma cell lines in vitro. High levels of YAP1 messenger RNA expression were associated with aggressive molecular subsets of glioblastoma and with a nonsignificant trend toward reduced mean survival in human astrocytoma patients. These findings suggest that YAP1 may play an important role in normal human brain development and that it could represent a new target in human brain tumors.

  6. Improved methodology for the affinity isolation of human protein complexes expressed at near endogenous levels

    DEFF Research Database (Denmark)

    Domanski, Michal; Molloy, Kelly; Jiang, Hua;

    2012-01-01

    An efficient and reliable procedure for the capture of affinity-tagged proteins and associated complexes from human cell lines is reported. Through multiple optimizations, high yield and low background affinity-purifications are achieved from modest quantities of human cells expressing endogenous......-level tagged proteins. Isolations of triple-FLAG and GFP-tagged fusion proteins involved in RNA metabolism are presented.......An efficient and reliable procedure for the capture of affinity-tagged proteins and associated complexes from human cell lines is reported. Through multiple optimizations, high yield and low background affinity-purifications are achieved from modest quantities of human cells expressing endogenous...

  7. Androgen-Dependent Regulation of Human MUC1 Mucin Expression

    Directory of Open Access Journals (Sweden)

    Stephen Mitchell

    2002-01-01

    Full Text Available MUC1 mucin is transcriptionally regulated by estrogen, progesterone, and glucocorticoids. Our objective was to determine whether androgen receptor. (20AR activation regulates expression of MUC1. The following breast and prostatic cell lines were phenotyped and grouped according to AR and MUC1protein expression: 1 AR+MUCi + [DAR17+19. (20AR transfectants of DU-145, ZR-75-1, MDA-MB-453, and T47D]; 2 AR-MUCi+ [DZeoi. (20AR- vector control, DU-145, BT20, MDA-MB231, and MCF7]; 3 AIR +MUCi -. (20LNCaP and LNCaP-r. Cell proliferation was determined using the MTT assay in the presence of synthetic androgen R1881, 0.1 pM to 1 µM. Cell surface MUC1expression was determined by flow cytometry in the presence or absence of oestradiol, medroxy progesterone acetate or R1881, with and without 4 hydroxy-flutamide. (204-OH, a nonsteroidal AR antagonist. The functional significance of MUC1expression was investigated with a cell-cell aggregation assay. Only AR+ MUC1 + cell lines showed a significant increase in MUC1expression with AR activation. (20P. (20range =.01 to .0001, reversed in the presence of 4-OHF. Cell proliferation was unaffected. Increased expression of MUC1was associated with a significant. (20P. (20range =.002 to .001 reduction in cell-cell adhesion. To our knowledge, this is the first description of androgen-dependent regulation of MUC1mucin. This is also functionally associated with decreased cell-cell adhesion, a recognised feature of progressive malignancy. These findings have important implications for physiological and pathological processes.

  8. Expression of estrogen receptor β in human colorectal cancer

    Institute of Scientific and Technical Information of China (English)

    Li-Qun Xie; Jie-Ping Yu; He-Sheng Luo

    2004-01-01

    AIM: To determine the expression of estrogen receptor (ER)β in Chinese colorectal carcinoma (CRC) patients.METHODS: Erβ expression in CRC was investigated by immunohistochemical staining of formalin-fixed, paraffin-embedded tissue sections from 40 CRCs, 10 colonic adenomas,and 10 normal colon mucosa biopsies. The percentage of positive cells was recorded, mRNA expression of Erα and Erβ in 12 CRC tissues and paired normal colon tissues were detected by RT-PCR.RESULTS: Positive ER immunoreactivity was present in part of normal epithelium of biopsy (2/10), adenomas (3/10),and the sections of CRC tissue, most of them were nuclear positive. In CRCs, nuclear Erβ immunoreactivity was detected in over 10% of the cancer cells in 57.5% of the cases and was always associated with cytoplasmic immunoreactivity.There was no statistical significance between Erβ positive and negative groups in regard to depth of invasion and nodal metastases. Of the 12 CRC tissues and paired normal colon tissues, the expression rate of Erα mRNA in CRC tissue and corresponding normal colon tissue was 25% and 16.6%,respectively. Erβ mRNA was expressed in 83.3% CRC tissue and 91.7% paired normal colon tissue, respectively. Therewas no significant difference in Erβ mRNA level between CRC tissues and paired normal colon tissues.CONCLUSION: A large number of CRCs are positive for Erβ, which can also be detected in normal colonic epithelia.There is a different localization of Erβ immunoreactivity among normal colon mucosae, adenomas and CRCs. Erαand Erβ mRNA can be detected both in CRC tissue and in corresponding normal colon tissue. A post-transcriptional mechanism may account for the decrease of Erβ protein expression in CRC tissues.

  9. EXPRESSION OF GST-π GENE IN HUMAN ESOPHAGEAL CARCINOGENESIS

    Institute of Scientific and Technical Information of China (English)

    FU Baojin; ZHANG Yunhan; WANG Yaohe; GAO Dongling; FU Shuli; WEN Xiaogang; ZHANG Sanshen; WANG Jiang

    1999-01-01

    Objective: To investigate the possible role of GSTπ in esophageal carcinogenesis. Methods: GST-πexpression at mRNA level was studied by in situ hybridization (ISH) and at protein level by immunohistochemistry (IHC). GST-π expression in normal epithelial cells (NC) of the esophagus,hyperplastic cells (HC), dysplastic cells (DC) from grade Ⅰ to Ⅲ, carcinoma in situ (CIS) and all the cells in squamous cell carcinomas (SCC) were examined in the same esophageal cancer specimens (n=48) which provided a model reflecting the process of esophageal carcinogenesis. Results: The positive rate of IHC staining was 87. 5% for NC, 95.3% for HC, 55.9% for DC (grade Ⅰ: 73.9%, grade Ⅱ: 47.4%, grade Ⅲ: 41.2%),36.4% for CIS and 45.8% for SCC. The positive rate of GST-π mRNA expression was 81.2% for NC, 94.4% for HC, 61.9% for DC (grade Ⅰ: 76.5%, grade Ⅱ: 61.5%,grade Ⅲ: 41.7%), 44.4% for CIS and 83.3% for grade ⅠSCC, 30.0% for grade Ⅱ SCC and 0% for grade ⅢSCC. There was no statistically significant difference in GST-π expression at the mRNA and the protein level.Conclusion: There is a decreasing tendency of GST-πexpression from dysplasia to CIS and SCC. The decrease in GST-π expression is an early event in esophageal carcinogenesis.

  10. Regulation of fibrinogen receptor expression on human platelets

    Energy Technology Data Exchange (ETDEWEB)

    Shattil, S.J.; Motulsky, H.J.; Insel, P.A.; Brass, L.F.

    1986-03-01

    Platelet aggregation requires the binding of fibrinogen to specific receptors on the plasma membrane glycoprotein IIb-IIIa complex. Although the IIb-IIIa complex is identifiable on the surface of resting platelets, the fibrinogen receptor is expressed only after platelet activation. The authors have developed a monoclonal anti-IIb-IIIa antibody (PAC-1) that binds only to stimulated platelets and only in the presence of Ca. In order to better understand the steps leading to platelet aggregation, the authors used radiolabeled PAC-1 and fibrinogen to examine the effect of the ..cap alpha../sub 2/-adrenergic agonist, epinephrine, on the expression and function of the fibrinogen receptor. The addition of epinephrine to unstirred platelets caused and immediate increase in PAC-1 and fibrinogen binding that was associated with platelet aggregation once the platelets were stirred. Even after prolonged incubation of the platelets with epinephrine, fibrinogen receptor expression could be reversed by adding EGTA, PGl/sub 2/, or the ..cap alpha../sub 2/-adrenergic antagonist, phentolamine. When unstirred platelets were exposed to epinephrine for more than 10 min, the extent of aggregation caused by subsequent stirring was decreased by 70%. Surprisingly, these desensitized platelets bound PAC-1 and fibrinogen normally, indicating that the loss of aggregation was not due to a decrease in fibrinogen receptor expression or function. These studies demonstrate that: (1) fibrinogen receptor expression is dependent on extracellular CA; (2) induction of the fibrinogen receptor by epinephrine requires the continued presence of the agonist; and (3) prolonged stimulation of the platelet by epinephrine can lead to a reduced aggregation response by a mechanism that does not involve a loss of either fibrinogen recepor expression or fibrinogen binding.

  11. Large Scale Gene Expression Meta-Analysis Reveals Tissue-Specific, Sex-Biased Gene Expression in Humans

    Science.gov (United States)

    Mayne, Benjamin T.; Bianco-Miotto, Tina; Buckberry, Sam; Breen, James; Clifton, Vicki; Shoubridge, Cheryl; Roberts, Claire T.

    2016-01-01

    The severity and prevalence of many diseases are known to differ between the sexes. Organ specific sex-biased gene expression may underpin these and other sexually dimorphic traits. To further our understanding of sex differences in transcriptional regulation, we performed meta-analyses of sex biased gene expression in multiple human tissues. We analyzed 22 publicly available human gene expression microarray data sets including over 2500 samples from 15 different tissues and 9 different organs. Briefly, by using an inverse-variance method we determined the effect size difference of gene expression between males and females. We found the greatest sex differences in gene expression in the brain, specifically in the anterior cingulate cortex, (1818 genes), followed by the heart (375 genes), kidney (224 genes), colon (218 genes), and thyroid (163 genes). More interestingly, we found different parts of the brain with varying numbers and identity of sex-biased genes, indicating that specific cortical regions may influence sexually dimorphic traits. The majority of sex-biased genes in other tissues such as the bladder, liver, lungs, and pancreas were on the sex chromosomes or involved in sex hormone production. On average in each tissue, 32% of autosomal genes that were expressed in a sex-biased fashion contained androgen or estrogen hormone response elements. Interestingly, across all tissues, we found approximately two-thirds of autosomal genes that were sex-biased were not under direct influence of sex hormones. To our knowledge this is the largest analysis of sex-biased gene expression in human tissues to date. We identified many sex-biased genes that were not under the direct influence of sex chromosome genes or sex hormones. These may provide targets for future development of sex-specific treatments for diseases.

  12. Large scale gene expression meta-analysis reveals tissue-specific, sex-biased gene expression in humans

    Directory of Open Access Journals (Sweden)

    Benjamin Mayne

    2016-10-01

    Full Text Available The severity and prevalence of many diseases are known to differ between the sexes. Organ specific sex-biased gene expression may underpin these and other sexually dimorphic traits. To further our understanding of sex differences in transcriptional regulation, we performed meta-analyses of sex biased gene expression in multiple human tissues. We analysed 22 publicly available human gene expression microarray data sets including over 2500 samples from 15 different tissues and 9 different organs. Briefly, by using an inverse-variance method we determined the effect size difference of gene expression between males and females. We found the greatest sex differences in gene expression in the brain, specifically in the anterior cingulate cortex, (1818 genes, followed by the heart (375 genes, kidney (224 genes, colon (218 genes and thyroid (163 genes. More interestingly, we found different parts of the brain with varying numbers and identity of sex-biased genes, indicating that specific cortical regions may influence sexually dimorphic traits. The majority of sex-biased genes in other tissues such as the bladder, liver, lungs and pancreas were on the sex chromosomes or involved in sex hormone production. On average in each tissue, 32% of autosomal genes that were expressed in a sex-biased fashion contained androgen or estrogen hormone response elements. Interestingly, across all tissues, we found approximately two-thirds of autosomal genes that were sex-biased were not under direct influence of sex hormones. To our knowledge this is the largest analysis of sex-biased gene expression in human tissues to date. We identified many sex-biased genes that were not under the direct influence of sex chromosome genes or sex hormones. These may provide targets for future development of sex-specific treatments for diseases.

  13. Identification and validation of suitable endogenous reference genes for gene expression studies in human peripheral blood

    Directory of Open Access Journals (Sweden)

    Turner Renee J

    2009-08-01

    Full Text Available Abstract Background Gene expression studies require appropriate normalization methods. One such method uses stably expressed reference genes. Since suitable reference genes appear to be unique for each tissue, we have identified an optimal set of the most stably expressed genes in human blood that can be used for normalization. Methods Whole-genome Affymetrix Human 2.0 Plus arrays were examined from 526 samples of males and females ages 2 to 78, including control subjects and patients with Tourette syndrome, stroke, migraine, muscular dystrophy, and autism. The top 100 most stably expressed genes with a broad range of expression levels were identified. To validate the best candidate genes, we performed quantitative RT-PCR on a subset of 10 genes (TRAP1, DECR1, FPGS, FARP1, MAPRE2, PEX16, GINS2, CRY2, CSNK1G2 and A4GALT, 4 commonly employed reference genes (GAPDH, ACTB, B2M and HMBS and PPIB, previously reported to be stably expressed in blood. Expression stability and ranking analysis were performed using GeNorm and NormFinder algorithms. Results Reference genes were ranked based on their expression stability and the minimum number of genes needed for nomalization as calculated using GeNorm showed that the fewest, most stably expressed genes needed for acurate normalization in RNA expression studies of human whole blood is a combination of TRAP1, FPGS, DECR1 and PPIB. We confirmed the ranking of the best candidate control genes by using an alternative algorithm (NormFinder. Conclusion The reference genes identified in this study are stably expressed in whole blood of humans of both genders with multiple disease conditions and ages 2 to 78. Importantly, they also have different functions within cells and thus should be expressed independently of each other. These genes should be useful as normalization genes for microarray and RT-PCR whole blood studies of human physiology, metabolism and disease.

  14. Expression

    Directory of Open Access Journals (Sweden)

    Wang-Xia Wang

    2014-02-01

    Full Text Available The miR-15/107 family comprises a group of 10 paralogous microRNAs (miRNAs, sharing a 5′ AGCAGC sequence. These miRNAs have overlapping targets. In order to characterize the expression of miR-15/107 family miRNAs, we employed customized TaqMan Low-Density micro-fluid PCR-array to investigate the expression of miR-15/107 family members, and other selected miRNAs, in 11 human tissues obtained at autopsy including the cerebral cortex, frontal cortex, primary visual cortex, thalamus, heart, lung, liver, kidney, spleen, stomach and skeletal muscle. miR-103, miR-195 and miR-497 were expressed at similar levels across various tissues, whereas miR-107 is enriched in brain samples. We also examined the expression patterns of evolutionarily conserved miR-15/107 miRNAs in three distinct primary rat brain cell preparations (enriched for cortical neurons, astrocytes and microglia, respectively. In primary cultures of rat brain cells, several members of the miR-15/107 family are enriched in neurons compared to other cell types in the central nervous system (CNS. In addition to mature miRNAs, we also examined the expression of precursors (pri-miRNAs. Our data suggested a generally poor correlation between the expression of mature miRNAs and their precursors. In summary, we provide a detailed study of the tissue and cell type-specific expression profile of this highly expressed and phylogenetically conserved family of miRNA genes.

  15. Malaria infected mosquitoes express enhanced attraction to human odor.

    Directory of Open Access Journals (Sweden)

    Renate C Smallegange

    Full Text Available There is much evidence that some pathogens manipulate the behaviour of their mosquito hosts to enhance pathogen transmission. However, it is unknown whether this phenomenon exists in the interaction of Anopheles gambiae sensu stricto with the malaria parasite, Plasmodium falciparum--one of the most important interactions in the context of humanity, with malaria causing over 200 million human cases and over 770 thousand deaths each year. Here we demonstrate, for the first time, that infection with P. falciparum causes alterations in behavioural responses to host-derived olfactory stimuli in host-seeking female An. gambiae s.s. mosquitoes. In behavioural experiments we showed that P. falciparum-infected An. gambiae mosquitoes were significantly more attracted to human odors than uninfected mosquitoes. Both P. falciparum-infected and uninfected mosquitoes landed significantly more on a substrate emanating human skin odor compared to a clean substrate. However, significantly more infected mosquitoes landed and probed on a substrate emanating human skin odor than uninfected mosquitoes. This is the first demonstration of a change of An. gambiae behaviour in response to olfactory stimuli caused by infection with P. falciparum. The results of our study provide vital information that could be used to provide better predictions of how malaria is transmitted from human being to human being by An. gambiae s.s. females. Additionally, it highlights the urgent need to investigate this interaction further to determine the olfactory mechanisms that underlie the differential behavioural responses. In doing so, new attractive compounds could be identified which could be used to develop improved mosquito traps for surveillance or trapping programmes that may even specifically target P. falciparum-infected An. gambiae s.s. females.

  16. Expression from second-generation feline immunodeficiency virus vectors is impaired in human hematopoietic cells.

    Science.gov (United States)

    Price, Mary A; Case, Scott S; Carbonaro, Denise A; Yu, Xiao-Jin; Petersen, Denise; Sabo, Kathleen M; Curran, Michael A; Engel, Barbara C; Margarian, Hovanes; Abkowitz, Janis L; Nolan, Garry P; Kohn, Donald B; Crooks, Gay M

    2002-11-01

    Vectors based on the feline immunodeficiency virus (FIV) have been developed as an alternative to those based on another lentivirus, human immunodeficiency virus-1 (HIV-1), because of theoretical safety advantages. We compared the efficiency of gene transfer and expression in human and feline hematopoietic progenitors using second-generation HIV-1 and FIV-based vectors. Vector pairs were tested using either human cytomegalovirus or murine phospho-glycerate kinase (PGK) internal promoters and were pseudotyped with the vesicular stomatitis virus G protein (VSV-G). Vector proviral copy numbers were similar in human and feline hematopoietic primary cells and cell lines transduced by HIV-1 or FIV vectors, demonstrating that both vectors are able to transfer genes efficiently to these cell types. HIV-1 vectors were well expressed in human primary hematopoietic cells and cell lines. However, transgene expression from FIV vectors was almost undetectable in human hematopoietic cells. In contrast, the FIV vector was expressed well in primary hematopoietic feline cells and human non-hematopoietic cells, demonstrating that low transgene expression from the FIV vector is a phenomenon specific to human hematopoietic cells. Northern blot analysis demonstrated decreased vector transcript levels in human CEM cells transduced with FIV relative to cells transduced with HIV-1, despite high vector copy numbers. No evidence of vector transcript instability was seen in studies of transduced CEM cells treated with actinomycin D. We conclude that FIV vectors can transfer genes into human hematopoietic cells as effectively as HIV-1 vectors, but that unknown elements in the current FIV backbone inhibit expression from FIV vectors in human hematopoietic cells.

  17. Expression of vascular endothelial growth factor and its two receptors in normal human endometrium

    Institute of Scientific and Technical Information of China (English)

    王海燕; 陈贵安

    2003-01-01

    Objectives: We try to demonstrate the expression of vascular endothelial growthfactor (VEGF) and its receptors, flt-1 and KDR, in normal human emdometrium duringthe menstrual cycle.Methods: Immunohistochemical method was used to observe the expression ofVEGF and its two receptors in emdometrium throughout the normal menstrual cyclemeanwhile the isoforms of VEGF were also detected by Western blot analysis. The en-dothelial cells of micro-vessels were marked with Ⅷ factor antibody.Results: VEGF and its receptors existed in endometrial glandular, stromal and vas-cular endothelial cells of human endometrium. Their expressions were higher in the mid-secretory phase of menstrual cycle and highest at menstruation. VEGF121 and VEGF165were the predominant isoforms in normal human endometrium.Conclusion: The expression of VEGF and its two receptors showed cycle-dependentin human endometrium, probably involved in embryonic implantation and endometrialproliferation and differentiation.

  18. Global gene expression profiling of asymptomatic bacteriuria Escherichia coli during biofilm growth in human urine

    DEFF Research Database (Denmark)

    Hancock, Viktoria; Klemm, Per

    2007-01-01

    asymptomatic bacteriuria (ABU) E. coli strains 83972 and VR50 are significantly better biofilm formers in their natural growth medium, human urine, than the two uropathogenic E. coli isolates CFT073 and 536. We used DNA microarrays to monitor the expression profile during biofilm growth in urine of the two ABU...... strains 83972 and VR50. Significant differences in expression levels were seen between the biofilm expression profiles of the two strains with the corresponding planktonic expression profiles in morpholinepropanesulfonic acid minimal laboratory medium and human urine; 417 and 355 genes were up- and down...... versions of 83972 and VR50; all mutants showed reduced biofilm formation in urine by 18 to 43% compared with the wild type (P profile of strain 83972 in the human urinary tract partially overlaps with the biofilm expression profile....

  19. EXPRESSION OF IL-13Ra2 GENE IN HUMAN BRAIN TUMORS

    Institute of Scientific and Technical Information of China (English)

    WU An-hua; TIE Xin-xin; WANG Yun-jie; YANG Guo-rui

    2005-01-01

    Objective: To investigate the expression of IL-13Ra2 gene in brain tumors. Methods: Seventy-nine human brain tumors were obtained from the department of Neurosurgery of China Medical University. Human IL-13Ra2 expression was evaluated by reverse transcriptase polymerase chain reaction and immunohistochemical analysis. Results: IL-13Ra2 gene was highly expressed in glioblastoma, medulloblastoma, malignant meningioma and benign meningioma. Conclusion:Human IL-13Ra2 gene is expressed in brain tumors in addition to gliomas, and our result indicates that the IL-13Ra2 gene promoter based gene therapy method can be used to treat brain tumors in addition to gliomas. Further studies involving larger numbers of samples are necessary to fully understand the expression profile of IL-13Ra2 gene in the brain tumors.

  20. Cloning, expression, and chromosome mapping of human galectin-7

    DEFF Research Database (Denmark)

    Madsen, Peder; Rasmussen, H H; Flint, T

    1995-01-01

    comigrated with IEF 17 as determined by two-dimensional (two-dimensional gel electrophoresis) analysis of proteins expressed by transiently transfected COS-1 cells, and bound lactose. Alignment of the amino acid sequences with other members of the family showed that the amino acids central to the beta...

  1. SREBP inhibits VEGF expression in human smooth muscle cells.

    Science.gov (United States)

    Motoyama, Koka; Fukumoto, Shinya; Koyama, Hidenori; Emoto, Masanori; Shimano, Hitoshi; Maemura, Koji; Nishizawa, Yoshiki

    2006-03-31

    Sterol regulatory element-binding proteins (SREBPs) are transcription factors that regulate expression of genes encoding enzymes for lipid biosynthesis. SREBPs are activated by HMG-CoA reductase inhibitors (statins). Statins have been also reported to suppress vascular endothelial growth factor (VEGF) expression in vascular smooth muscle cells (VSMCs). Therefore, we hypothesized that SREBPs are involved in statin-mediated regulation of VEGF production in VSMCs. SREBP1 was robustly expressed, and was activated by atorvastatin in VSMCs, as demonstrated by increased levels of the mature nuclear form of SREBP1, and increased promoter activities of a reporter containing sterol regulatory elements by atorvastatin. Moreover, overexpression of SREBP1a dose-dependently suppressed VEGF promoter activity. Site-specific mutation or deletion of the proximal Sp1 sites reduced the inhibitory effects of SREBP1a on VEGF promoter activity. These data demonstrated that SREBP1, activated by atorvastatin, suppressed VEGF expression through the indirect interaction with the proximal tandem Sp1 sites in VSMCs.

  2. Gene structural analysis and expression of human renal dipeptidase

    Energy Technology Data Exchange (ETDEWEB)

    Satoh, Susumu; Ohtsuka, Kazuyuki; Keida, Yuriko; Kusunoki, Chihiro; Niwa, Mineo; Kohsaka, Masanobu (Fujisawa Pharmaceutical Company, Ltd., Osaka (Japan)); Konta, Yoshiyuki (Hirosaki Univ. (Japan))

    Human renal dipeptidase cDNA and genomic DNA were isolated from human kidney cDNA and genomic libraries, respectively. The human renal dipeptidase gene has a total length of approximately 6 kb and consists of ten exons and nine introns. The exons and cDNA each encode the 411 amino acid residues of the precursor protein, including 16 amino acid residues of signal sequence and a hydrophobic carboxyl terminal sequence for the attachment of a phosphatidylinositol glycan. Although the cDNA was slightly different from the cDNA reported by Adachi et al. (1990), the differences observed suggest, by comparison with human genomic DNA, that it may not represent an allelic variant but a cloning artifact. The recombinant human renal dipeptidase was produced on the surface of transfected L929 cells and had the same character as native renal dipeptidase. Northern blotting hybridization analysis showed that renal dipeptidase mRNA is only transcribed in kidney. 21 refs., 5 figs., 2 tabs.

  3. Expression of Neuropeptide Y and Its Relationship with Molecular and Morphological Changes in Human Pituitary Adenomas.

    Science.gov (United States)

    Jia, Ruichao; Li, Mu; Chang, Binge; Chen, Laichao; Ma, Jingjian

    2015-12-01

    The purpose of this study was to explore the role of neuropeptide Y (NPY) on molecular and histological changes in human pituitary adenomas. The localization of NPY and its expression at the protein, messenger RNA (mRNA), and receptor levels were investigated here in different subcategories of pituitary adenomas. Immunohistochemical staining was performed in all cases to assess expression of NPY. Reverse transcription-polymerase chain reaction (RT-PCR) was used to study the mRNA expression of NPY. NPY subcellular localization was observed using immunoelectron microscopy in cytoplasm, rough endoplasmic reticulum, and cell matrix in four of the six cases of pituitary adenoma. NPY protein expression was observed in 59.6% of 57 cases of pituitary adenoma and in 2 cases of pituitary hyperplasia. mRNA expression of NPY was observed in all 57 cases of pituitary adenoma and in 2 cases of pituitary hyperplasia. Significantly different levels of expression were observed across different subcategories of pituitary adenoma. mRNA expression of Y1R and Y2R was observed across all subcategories of pituitary adenomas, and a positive correlation was observed between NPY and Y2R. In conclusion, evidence is provided here for the expression of NPY and its receptors, Y1R and Y2R, in human pituitary adenoma, and the levels of expression were found to differ across different subcategories. Differences in expression of Y2R in human pituitary adenomas were found to have remarkable statistical significance.

  4. Human Vascular Endothelial Growth Factor cDNA Cloning and Expression in Osteoblasts

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    Human vascular endothelial growth factor (VEGF) cDNA was amplified by nested polymerase chain reaction method from the HL60 cells. Then a pCD-hVEGF165 recombinant plasmid was constructed. Rabbit osteoblasts were transfected with pCD-hVEGF165 plasmid by lipofectin mediated gene transfer. The transient expressive results were detected by immunohistochemical method. It was observed that the expression of human VEGF gene was detected 72 h after transfecting distinctly.

  5. Dissimilar processing of emotional facial expressions in human and monkey temporal cortex.

    Science.gov (United States)

    Zhu, Qi; Nelissen, Koen; Van den Stock, Jan; De Winter, François-Laurent; Pauwels, Karl; de Gelder, Beatrice; Vanduffel, Wim; Vandenbulcke, Mathieu

    2013-02-01

    Emotional facial expressions play an important role in social communication across primates. Despite major progress made in our understanding of categorical information processing such as for objects and faces, little is known, however, about how the primate brain evolved to process emotional cues. In this study, we used functional magnetic resonance imaging (fMRI) to compare the processing of emotional facial expressions between monkeys and humans. We used a 2×2×2 factorial design with species (human and monkey), expression (fear and chewing) and configuration (intact versus scrambled) as factors. At the whole brain level, neural responses to conspecific emotional expressions were anatomically confined to the superior temporal sulcus (STS) in humans. Within the human STS, we found functional subdivisions with a face-selective right posterior STS area that also responded to emotional expressions of other species and a more anterior area in the right middle STS that responded specifically to human emotions. Hence, we argue that the latter region does not show a mere emotion-dependent modulation of activity but is primarily driven by human emotional facial expressions. Conversely, in monkeys, emotional responses appeared in earlier visual cortex and outside face-selective regions in inferior temporal cortex that responded also to multiple visual categories. Within monkey IT, we also found areas that were more responsive to conspecific than to non-conspecific emotional expressions but these responses were not as specific as in human middle STS. Overall, our results indicate that human STS may have developed unique properties to deal with social cues such as emotional expressions.

  6. Genome-wide expression profiling of five mouse models identifies similarities and differences with human psoriasis.

    Directory of Open Access Journals (Sweden)

    William R Swindell

    Full Text Available Development of a suitable mouse model would facilitate the investigation of pathomechanisms underlying human psoriasis and would also assist in development of therapeutic treatments. However, while many psoriasis mouse models have been proposed, no single model recapitulates all features of the human disease, and standardized validation criteria for psoriasis mouse models have not been widely applied. In this study, whole-genome transcriptional profiling is used to compare gene expression patterns manifested by human psoriatic skin lesions with those that occur in five psoriasis mouse models (K5-Tie2, imiquimod, K14-AREG, K5-Stat3C and K5-TGFbeta1. While the cutaneous gene expression profiles associated with each mouse phenotype exhibited statistically significant similarity to the expression profile of psoriasis in humans, each model displayed distinctive sets of similarities and differences in comparison to human psoriasis. For all five models, correspondence to the human disease was strong with respect to genes involved in epidermal development and keratinization. Immune and inflammation-associated gene expression, in contrast, was more variable between models as compared to the human disease. These findings support the value of all five models as research tools, each with identifiable areas of convergence to and divergence from the human disease. Additionally, the approach used in this paper provides an objective and quantitative method for evaluation of proposed mouse models of psoriasis, which can be strategically applied in future studies to score strengths of mouse phenotypes relative to specific aspects of human psoriasis.

  7. Gene expression analysis of primary normal human hepatocytes infected with human hepatitis B virus

    Institute of Scientific and Technical Information of China (English)

    Hyun Mi Ryu; Sung Gyoo Park; Sung Su Yea; Won Hee Jang; Young-Il Yang; Guhung Jung

    2006-01-01

    AIM: To find the relationship between hepatitis B virus (HBV) and hepatocytes during the initial state of infection by cDNA microarray.METHODS: Primary normal human hepatocytes (PNHHs)were isolated and infected with HBV. From the PNHHs,RNA was isolated and inverted into complement DNA (cDNA) with Cy3- or Cy5- labeled dUTP for microarray analysis. The labeled cDNA was hybridized with microarray chip, including 4224 cDNAs. From the image of the microarray, expression profiles were produced and some of them were confirmed by RT-PCR, immunoblot analysis, and NF-κB luciferase reporter assay.RESULTS: From the cDNA microarray, we obtained 98differentially regulated genes. Of the 98 genes, 53 were up regulated and 45 down regulated. Interestingly, in the up regulated genes, we found the TNF signaling pathway-related genes: LT-α, TRAF2, and NIK. By using RT-PCR, we confirmed the up-regulation of these genes in HepG2, Huh7, and Chang liver cells, which were transfected with pHBV1.2x, a plasmid encoding all HBV messages. Moreover, these three genes participated in HBVmediated NF-κB activation.CONCLUSION: During the initial state of HBV infection,hepatocytes facilitate the activation of NF-κB through up regulation of LT-α, TRAF2, and NIK.

  8. TGF-β1 inhibits connexin-43 expression in cultured smooth muscle cells of human bladder

    Institute of Scientific and Technical Information of China (English)

    Chi Qiang; Zhou Fenghai; Wang Yangmin

    2009-01-01

    Objective: In this research, we studied the TGF-β1 effects on connexin-43 expression in cultured human bladder smooth muscle cells. Methods: Human bladder smooth muscle cells primary cultures, with bladder tissue obtained from patients undergoing cystectomy, were intervened by recombinant human TGF-β1. Connexin-43 expression in human bladder smooth muscle cells was then examined by Western blotting and immunocytochemistry. Results: Stimulation with TGF-β1 led to significant reduction of cormexin-43 immunoreactivity and coupling (P<0.0001). Connexin-43 protein expression was significantly downregnlated (P<0.05). Simultaneously, low phosphorylation species of connexin-43 were particularly affected. Conclusion: Our experiments demonstrated a significant downregulation of connexin-43 by TGF-β1 in cultured human bladder smooth muscle cells. These findings support the view that TGF-β1 is involved in the pathophysiology of urinary bladder dysfunction.

  9. In vitro cytokeratin expression profiling of human oral mucosa substitutes developed by tissue engineering.

    Science.gov (United States)

    Garzon, Ingrid; Serrato, Deyanira; Roda, Olga; Del Carmen Sanchez-Quevedo, Maria; Gonzales-Jaranay, Maximino; Moreu, Gerardo; Nieto-Aguilar, Renato; Alaminos, Miguel; Campos, Antonio

    2009-10-01

    In this work we performed a study of cytokeratin (CK) expression profiling on human artificial oral mucosa developed in vitro by tissue engineering at different stages of maturation (from immature to well-developed stages) at the protein and mRNA levels. Human artificial oral mucosa was generated in the laboratory using fibrin-agarose biomaterials. As controls, we used human native normal oral mucosa and embryonic oral tissues. Our results demonstrated that human embryonic oral tissues tended to express CK8 and CK19. In contrast, monolayered bioengineered oral mucosa did not show any CK expression by immunohistochemistry, whereas bilayered and multilayered artificial oral mucosa showed several markers of stratified epithelia, but did not express CK10. These results suggest that the CK expression pattern is strongly dependent on the maturation state of the artificial tissues and that the CK expression profile of our model of artificial oral mucosa was partially similar to that of the non-keratinized human adult oral mucosa. However, the expression of CK8 by the artificial oral mucosa suggests that these samples correspond to an early stage of development while kept in vitro.

  10. Expression of vitamin D receptor and cathelicidin in human corneal epithelium cells during fusarium solani infection.

    Science.gov (United States)

    Cong, Lin; Xia, Yi-Ping; Zhao, Gui-Qiu; Lin, Jing; Xu, Qiang; Hu, Li-Ting; Qu, Jian-Qiu; Peng, Xu-Dong

    2015-01-01

    To observe the expression of vitamin D receptor (VDR) in human specimen and immortalized human corneal epithelium cells (HCEC) when challenged with fusarium solani. Moreover, we decided to discover the pathway of VDR expression. Also, we would like to detect the expression of cathelicidin antimicrobial peptide (CAMP) in the downstream pathway of VDR. Immunohistochemistry was used to examine the VDR expression in HCEC from healthy and fungal keratitis patients. Real time quantitative polymerase chain reaction (qPCR) was performed to observe the messenger ribonucleic acid (mRNA) change of VDR when immortalized HCEC were challenged with fusarium solani for different hours. CAMP was detected at both mRNA and protein levels. We found out that the VDR expression in fusarium solani keratitis patients' specimen was much more than that in healthy people. The mRNA and protein expression of VDR increased when we stimulated HCEC with fusarium solani antigen (Pfusarium solani antigen stimulation (Pfusarium solani antigen.

  11. A micro-RNA expression signature for human NAFLD progression.

    Science.gov (United States)

    Guo, Yan; Xiong, Yanhua; Sheng, Quanghu; Zhao, Shilin; Wattacheril, Julia; Flynn, Charles Robb

    2016-10-01

    The spectrum of nonalcoholic fatty liver disease (NAFLD) describes disease conditions deteriorating from nonalcoholic fatty liver (NAFL) to nonalcoholic steatohepatitis (NASH) to cirrhosis (CIR) to hepatocellular carcinoma (HCC). From a molecular and biochemical perspective, our understanding of the etiology of this disease is limited by the broad spectrum of disease presentations, the lack of a thorough understanding of the factors contributing to disease susceptibility, and ethical concerns related to repeat sampling of the liver. To better understand the factors associated with disease progression, we investigated by next-generation RNA sequencing the altered expression of microRNAs (miRNAs) in liver biopsies of class III obese subjects (body mass index ≥40 kg/m(2)) biopsied at the time of elective bariatric surgery. Clinical characteristics and unbiased RNA expression profiles for 233 miRs, 313 transfer RNAs (tRNAs), and 392 miscellaneous small RNAs (snoRNAs, snRNAs, rRNAs) were compared among 36 liver biopsy specimens stratified by disease severity. The abundances of 3 miRNAs that were found to be differentially regulated (miR-301a-3p and miR-34a-5p increased and miR-375 decreased) with disease progression were validated by RT-PCR. No tRNAs or miscellaneous RNAs were found to be associated with disease severity. Similar patterns of increased miR-301a and decreased miR-375 expression were observed in 134 hepatocellular carcinoma (HCC) samples deposited in The Cancer Genome Atlas (TCGA). Our analytical results suggest that NAFLD severity is associated with a specific pattern of altered hepatic microRNA expression that may drive the hallmark of this disorder: altered lipid and carbohydrate metabolism. The three identified miRNAs can potentially be used as biomarkers to access the severity of NAFLD. The persistence of this miRNA expression pattern in an external validation cohort of HCC samples suggests that specific microRNA expression patterns may permit and

  12. Ectopic expression of telomerase enhances osteopontin and osteocalcin expression during osteogenic differentiation of human mesenchymal stem cells from elder donors

    Directory of Open Access Journals (Sweden)

    Machado CB

    2009-01-01

    Full Text Available Age related bone loss is one of the most prevalent diseases in the elder population. The osteoblasts are the effectors cells of bone formation and regeneration. With the aging the osteoblasts become senescent reducing their ability to produce bone. Cellular replicative senescence is triggered by telomers shortening. Telomerase elongate the telomers length and maintain the cell proliferative capacity. Here, we demonstrated that the expression of human telomerase reverse transcriptase mediated by an adenovirus vector increases the levels of osteopontin and osteocalcin mRNA during the in vitro osteogenic differentiation of elderly human mesenchymal stem cells. Bone marrow human mesenchymal stem cells were obtained from old donors (>65 years and induced to differentiate into osteoblasts for 14 days. The levels of mRNA of human telomerase reverse transcriptase, osteopontin and osteocalcin during the differentiation were assessed by semi-quantitative PCR before and during the differentiation on days 7 and 14. Infected cells showed 1.5 fold increase in telomerase expression. Also telomerized cells exhibit 1.5 fold increase in osteopontin and 0.5 fold increase in osteocalcin expression compared to primary osteoblasts isolated from the same donors. The transformed cells were not able to form tumours in NUDE mice.

  13. The Human Anatomic Gene Expression Library (H-ANGEL), the H-Inv integrative display of human gene expression across disparate technologies and platforms

    Science.gov (United States)

    Tanino, Motohiko; Debily, Marie-Anne; Tamura, Takuro; Hishiki, Teruyoshi; Ogasawara, Osamu; Murakawa, Katsuji; Kawamoto, Shoko; Itoh, Kouichi; Watanabe, Shinya; de Souza, Sandro José; Imbeaud, Sandrine; Graudens, Esther; Eveno, Eric; Hilton, Phillip; Sudo, Yukio; Kelso, Janet; Ikeo, Kazuho; Imanishi, Tadashi; Gojobori, Takashi; Auffray, Charles; Hide, Winston; Okubo, Kousaku

    2005-01-01

    The Human Anatomic Gene Expression Library (H-ANGEL) is a resource for information concerning the anatomical distribution and expression of human gene transcripts. The tool contains protein expression data from multiple platforms that has been associated with both manually annotated full-length cDNAs from H-InvDB and RefSeq sequences. Of the H-Inv predicted genes, 18 897 have associated expression data generated by at least one platform. H-ANGEL utilizes categorized mRNA expression data from both publicly available and proprietary sources. It incorporates data generated by three types of methods from seven different platforms. The data are provided to the user in the form of a web-based viewer with numerous query options. H-ANGEL is updated with each new release of cDNA and genome sequence build. In future editions, we will incorporate the capability for expression data updates from existing and new platforms. H-ANGEL is accessible at http://www.jbirc.aist.go.jp/hinv/h-angel/. PMID:15608263

  14. Increased expression of aquaporin-4 in human traumatic brain injury and brain tumors

    Institute of Scientific and Technical Information of China (English)

    HuaHu; Wei-PingZhang; LeiZhang; ZhongChen; Er-QingWei

    2004-01-01

    Aquaporin-4 (AQP4) is one of the aquaporins (AQPs), a water channel family. In the brain, AQP4 is expressed in astroeyte foot processes, and plays an important role in water homeostasis and in the formation of brain edema. In our study, AQP4 expression in human brain specimens from patients with traumatic brain injury or different brain tumors was detected

  15. Systematic analysis of gene expression patterns associated with postmortem interval in human tissues.

    Science.gov (United States)

    Zhu, Yizhang; Wang, Likun; Yin, Yuxin; Yang, Ence

    2017-07-14

    Postmortem mRNA degradation is considered to be the major concern in gene expression research utilizing human postmortem tissues. A key factor in this process is the postmortem interval (PMI), which is defined as the interval between death and sample collection. However, global patterns of postmortem mRNA degradation at individual gene levels across diverse human tissues remain largely unknown. In this study, we performed a systematic analysis of alteration of gene expression associated with PMI in human tissues. From the Genotype-Tissue Expression (GTEx) database, we evaluated gene expression levels of 2,016 high-quality postmortem samples from 316 donors of European descent, with PMI ranging from 1 to 27 hours. We found that PMI-related mRNA degradation is tissue-specific, gene-specific, and even genotype-dependent, thus drawing a more comprehensive picture of PMI-associated gene expression across diverse human tissues. Additionally, we also identified 266 differentially variable (DV) genes, such as DEFB4B and IFNG, whose expression is significantly dispersed between short PMI (S-PMI) and long PMI (L-PMI) groups. In summary, our analyses provide a comprehensive profile of PMI-associated gene expression, which will help interpret gene expression patterns in the evaluation of postmortem tissues.

  16. Endothelial lipase is highly expressed in macrophages in advanced human atherosclerotic lesions

    DEFF Research Database (Denmark)

    Bartels, Emil D; Nielsen, John E; Lindegaard, Marie Louise Skakkebæk;

    2007-01-01

    Endothelial lipase (EL) is expressed in endothelial cells, and affects plasma lipoprotein metabolism by hydrolyzing phospholipids in HDL. To determine the cellular expression of EL mRNA and protein in human atherosclerotic lesions, we performed in situ hybridization and immunohistochemical studies...

  17. GLUT12 expression in human placenta in first trimester and term

    NARCIS (Netherlands)

    Gude, NM; Stevenson, JL; Rogers, S; Best, JD; Kalionis, B; Erwich, JJHM; Timmer, A; King, RG

    2003-01-01

    The aim of this study was to characterize the expression of a novel glucose transporter protein GLUT12 in human placenta. GLUT12 mRNA expression was identified by RT-PCR in extracts from five normal term placentae and in extracts from cultured cells of the JAR, JEG-3 and HTR-8Svneo cell lines. In fu

  18. RNA interference inhibits expression of vascular endothelial growth factor (VEGF) in human retinal pigment epithelial cells

    Institute of Scientific and Technical Information of China (English)

    CAI Chun-mei; SUN Bao-chen; LIU Xu-yang; WANG Jin-jin; LI Jun-fa; HAN Song; WANG Ning-li; LU Qing-jun

    2005-01-01

    @@ Choroidal neovascularization (CNV), a major cause of vision loss, is the result of the increased vascular endothelial growth factor (VEGF) expression in human retinal pigment epithelial (RPE) cells. It is important to inhibit the expression of VEGF protein in RPE cells.

  19. Characterization of hematopoietic GATA transcription factor expression in mouse and human dendritic cells.

    Science.gov (United States)

    Scheenstra, Maaike R; Salunkhe, Vishal; De Cuyper, Iris M; Hoogenboezem, Mark; Li, Eveline; Kuijpers, Taco W; van den Berg, Timo K; Gutiérrez, Laura

    2015-12-01

    Dendritic cells (DCs) are key initiators and regulators of the immune response. The development of the DC lineage and their subsets requires an orchestrated regulation of their transcriptional program. Gata1, a transcription factor expressed in several hematopoietic cell lineages, has been recently reported to be required for mouse DC development and function. In humans, GATA1 is involved in the lineage separation between monocyte-derived DCs and Langerhans cells (LC) and loss of GATA1 results in differentiation arrest at the monocyte stage. The hematopoietic GATA factors (i.e. Gata1, Gata2, Gata3) are known to regulate each other's expression and to function consecutively throughout lineage commitment (so-called GATA switch). In humans, mutations in GATA2 are causative of MonoMAC disease, a human immunodeficiency syndrome characterized by loss of DCs, monocytes, B and NK cells. However, additional data on the expression of hematopoietic GATA factors in the DC lineage is missing. In this study, we have characterized the expression of hematopoietic GATA factors in murine and human DCs and their expression dynamics upon TLR stimulation. We found that all hematopoietic GATA factors are expressed in DCs, but identified species-specific differences in the relative expression of each GATA factor, and how their expression fluctuates upon stimulation.

  20. Comparative pharmacology of a new recombinant FSH expressed by a human cell Line

    DEFF Research Database (Denmark)

    Koechling, Wolfgang; Plaksin, Daniel; Croston, Glenn

    2017-01-01

    Recombinant FSH proteins are important therapeutic agents for the treatment of infertility, including follitropin alfa expressed in Chinese Hamster Ovary (CHO) cells and, more recently, follitropin delta expressed in the human cell line PER.C6. These recombinant FSH proteins have distinct...

  1. Expression of biologically active human interferon alpha 2 in aloe vera

    Science.gov (United States)

    We have developed a system for transgenic expression of proteins in Aloe Vera. Using this approach we have generated plants expressing the human gene interferon alpha 2, IFNa2. IFNa2 is a small secreted cytokine that plays a vital role in regulating the body’s immune response to viral infections a...

  2. Polymorphic GGC repeat differentially regulates human reelin gene expression levels.

    Science.gov (United States)

    Persico, A M; Levitt, P; Pimenta, A F

    2006-10-01

    The human gene encoding Reelin (RELN), a pivotal protein in neurodevelopment, includes a polymorphic GGC repeat in its 5' untranslated region (UTR). CHO cells transfected with constructs encompassing the RELN 5'UTR with 4-to-13 GGC repeats upstream of the luciferase reporter gene show declining luciferase activity with increasing GGC repeat number (P autism.

  3. Human intrinsic factor expressed in the plant Arabidopsis thaliana

    DEFF Research Database (Denmark)

    Fedosov, Sergey N; Laursen, Niels B; Nexø, Ebba

    2003-01-01

    Intrinsic factor (IF) is the gastric protein that promotes the intestinal uptake of vitamin B12. Gastric IF from animal sources is used in diagnostic tests and in vitamin pills. However, administration of animal IF to humans becomes disadvantageous because of possible pathogenic transmission...

  4. Identification of Human HK Genes and Gene Expression Regulation Study in Cancer from Transcriptomics Data Analysis

    Science.gov (United States)

    Zhang, Zhang; Liu, Jingxing; Wu, Jiayan; Yu, Jun

    2013-01-01

    The regulation of gene expression is essential for eukaryotes, as it drives the processes of cellular differentiation and morphogenesis, leading to the creation of different cell types in multicellular organisms. RNA-Sequencing (RNA-Seq) provides researchers with a powerful toolbox for characterization and quantification of transcriptome. Many different human tissue/cell transcriptome datasets coming from RNA-Seq technology are available on public data resource. The fundamental issue here is how to develop an effective analysis method to estimate expression pattern similarities between different tumor tissues and their corresponding normal tissues. We define the gene expression pattern from three directions: 1) expression breadth, which reflects gene expression on/off status, and mainly concerns ubiquitously expressed genes; 2) low/high or constant/variable expression genes, based on gene expression level and variation; and 3) the regulation of gene expression at the gene structure level. The cluster analysis indicates that gene expression pattern is higher related to physiological condition rather than tissue spatial distance. Two sets of human housekeeping (HK) genes are defined according to cell/tissue types, respectively. To characterize the gene expression pattern in gene expression level and variation, we firstly apply improved K-means algorithm and a gene expression variance model. We find that cancer-associated HK genes (a HK gene is specific in cancer group, while not in normal group) are expressed higher and more variable in cancer condition than in normal condition. Cancer-associated HK genes prefer to AT-rich genes, and they are enriched in cell cycle regulation related functions and constitute some cancer signatures. The expression of large genes is also avoided in cancer group. These studies will help us understand which cell type-specific patterns of gene expression differ among different cell types, and particularly for cancer. PMID:23382867

  5. Medical Photography: Documentation, Art, and the Expression of Human Emotions

    Directory of Open Access Journals (Sweden)

    Elisabeth Aberer

    2016-09-01

    Full Text Available Medical photography is the state of the art for the documentation of dermatological disease. Experienced photographers take pictures of the most typical skin lesions in order to assist the clinician in assessing disease morphology and activity. In this study, we present 6 individuals with a variety of dermatoses and the expression of the patients’ emotions. The patients were asked to show their diseased skin and to present typically involved areas in the respective disease. The feelings expressed by their body movements and positions are viewed and interpreted. The patients’ history will be reported retrospectively. The aim of the report is to show that the art of medical photography does not only document skin lesions but also the disease burden and the associated impairment of quality of life. Moreover, dermatologic photography is a sensitive intervention for patients viewed in the light of teaching and patient care.

  6. Medical Photography: Documentation, Art, and the Expression of Human Emotions.

    Science.gov (United States)

    Aberer, Elisabeth; Stieber, Werner; Homayoon, Donja; Fink-Puches, Regina; Lichen, Roland; Salmhofer, Wolfgang; Gruber-Wackernagel, Alexandra; Aberer, Werner

    2016-01-01

    Medical photography is the state of the art for the documentation of dermatological disease. Experienced photographers take pictures of the most typical skin lesions in order to assist the clinician in assessing disease morphology and activity. In this study, we present 6 individuals with a variety of dermatoses and the expression of the patients' emotions. The patients were asked to show their diseased skin and to present typically involved areas in the respective disease. The feelings expressed by their body movements and positions are viewed and interpreted. The patients' history will be reported retrospectively. The aim of the report is to show that the art of medical photography does not only document skin lesions but also the disease burden and the associated impairment of quality of life. Moreover, dermatologic photography is a sensitive intervention for patients viewed in the light of teaching and patient care.

  7. Expression of NGF, Trka and p75 in human cartilage

    Directory of Open Access Journals (Sweden)

    A Gigante

    2009-06-01

    Full Text Available Nerve growth factor (NGF exerts its action through two types of receptor: high-affinity tyrosine kinase A receptor (trkA and low-affinity p75 receptor. NGF has a neurotrophic role in central and peripheral nervous system development, but there is also clear evidence of its involvement in the developing skeleton. The aim of the present immunohistochemical study was to investigate the expression and distribution of NGF, trkA, and p75 in normal cartilaginous tissues from adult subjects: articular and meniscal cartilage of the knee, cartilage from the epiglottis, and intervertebral disc tissue. Detection of NGF mRNA was also performed by in situ hybridization. Immunoreaction for NGF and the two receptors in articular chondrocytes, chondrocyte-like cells of meniscus and annulus fibrosus, and chondrocytes of the epiglottis demonstrated that they are all expressed in hyaline, fibrous and elastic cartilaginous tissues, suggesting that they could be involved in cartilage physio-pathology.

  8. Low microRNA-199a expression in human amniotic epithelial cell feeder layers maintains human-induced pluripotent stem cell pluripotency via increased leukemia inhibitory factor expression

    Institute of Scientific and Technical Information of China (English)

    Te Liu; Qing Chen; Yongyi Huang; Qin Huang; Lizhen Jiang; Lihe Guo

    2012-01-01

    Human-induced pluripotent stem (iPS) cells share the same key properties as embryonic stem cells,and may be generated from patient- or disease-specific sources,which makes them attractive for personalized medicine,drug screens,or cellular therapy.Long-term cultivation and maintenance of normal iPS cells in an undifferentiated self-renewing state is a major challenge.Our previous studies have shown that human amniotic epithelial cells (HuAECs) could provide a good source of feeder cells for mouse and human embryonic stem cells,or spermatogonial stem cells,as they express endogenous leukemia inhibitory factor (LIF) at high levels.Here,we examined the effect of exogenous microRNA-199a regulation on endogenous LIF expression in HuAECs,and in torn on human iPS cell pluripotency.We found that HuAECs feeder cells transfected with microRNA-199a mutant expressed LIF at high levels,allowing iPS to maintain a high level of alkaline phosphatase activity in longterm culture and form teratomas in severe combined immunodeficient mice.The expression of stem cell markers was increased in iPS cultured on HuAECs feeder cells transfected with the microRNA-199a mutant,compared with iPS cultured on HuAECs transfected with microRNA-199a or mouse embryo fibroblasts.Taken together,these results suggested that LIF expression might be regulated by microRNA-199a,and LIF was a crucial component in feeder cells,and also was required for maintenance of human iPS cells in an undifferentiated,proliferative state capable of self-renewal.

  9. Expression and function of beta1 integrins on human eosinophils

    Directory of Open Access Journals (Sweden)

    Maria-Cristina Seminario

    1997-12-01

    Full Text Available Eosinophils preferentially accumulate at sites of chronic allergic diseases such as bronchial asthma. The mechanisms by which selective eosinophil migration occurs are not fully understood. However, interactions of cell-surface adhesion molecules on the eosinophil with molecular counterligands on endothelial and epithelial cells, and on extracellular matrix proteins, are likely to be critical during the recruitment process. One possible mechanism for selective eosinophil recruitment involves the alpha4beta 1 (VLA-4 integrin which is not expressed on neutrophils. Correlations have been found between infiltration of eosinophils and endothelial expression of VCAM-1, the ligand for VLA-4, in the lungs of asthmatic individuals as well as in late phase reactions in the lungs, nose and skin. Epithelial and endothelial cells respond to the Th2-type cytokines IL-4 and IL-13 with selective de novo expression of VCAM-1, consistent with the possible role of VCAM-1/VLA-4 interactions in eosinophil influx during allergic inflammation. Both beta 1 and beta 2 integrins on eosinophils exist in a state of partial activation. For example, eosinophils can be maximally activated for adhesion to VCAM-1 or fibronectin after exposure to beta 1 integrin-activating antibodies or divalent cations, conditions that do not necessarily affect the total cell surface expression of beta 1 integrins. In contrast, cytokines like IL-5 prevent beta 1 integrin activation while promoting beta 2 integrin function. Furthermore, ligation of integrins can regulate the effector functions of the cell. For example, eosinophil adhesion via beta 1 and/or beta 2 integrins has been shown to alter a variety of functional responses including degranulation and apoptosis. Thus, integrins appear to be important in mediating eosinophil migration and activation in allergic inflammation. Strategies that interfere with these processes may prove to be useful for treatment of allergic diseases.

  10. Decreased PCSK9 expression in human hepatocellular carcinoma

    OpenAIRE

    2015-01-01

    Background The management of hepatocellular carcinoma (HCC) is limited by the lack of adequate screening biomarkers and chemotherapy. In response, there has been much interest in tumor metabolism as a therapeutic target. PCSK9 stimulates internalization of the LDL-receptor, decreases cholesterol uptake into hepatocytes and affects liver regeneration. Thus, we investigated whether PCSK9 expression is altered in HCC, influencing its ability to harness cholesterol metabolism. Methods Thirty-nine...

  11. Detecting positive darwinian selection in brain-expressed genes during human evolution

    Institute of Scientific and Technical Information of China (English)

    QI XueBin; Alice A. LIN; Luca L. CAVALLI-SFORZA; WANG Jun; SU Bing; YANG Su; ZHENG HongKun; WANG YinQiu; LIAO ChengHong; LIU Ying; CHEN XiaoHua; SHI Hong; YU XiaoJing

    2007-01-01

    To understand the genetic basis that underlies the phenotypic divergence between human and nonhuman primates, we screened a total of 7176 protein-coding genes expressed in the human brain and compared them with the chimpanzee orthologs to identify genes that show evidence of rapid evolution in the human lineage. Our results showed that the nonsynonymous/synonymous substitution (Ka/Ks) ratio for genes expressed in the brain of human and chimpanzee is 0.3854, suggesting that the brain-expressed genes are under functional constraint. The X-linked human brain-expressed genes evolved more rapidly than autosomal ones. We further dissected the molecular evolutionary patterns of 34 candidate genes by sequencing representative primate species to identify lineage-specific adaptive evolution. Fifteen out of the 34 candidate genes showed evidence of positive Darwinian selection in human and/or chimpanzee lineages. These genes are predicted to play diverse functional roles in embryonic development, spermatogenesis and male fertility, signal transduction, sensory nociception, and neural function. This study together with others demonstrated the usefulness and power of phylogenetic comparison of multiple closely related species in detecting lineage-specific adaptive evolution, and the identification of the positively selected brain-expressed genes may add new knowledge to the understanding of molecular mechanism of human origin.

  12. Computational analysis of expression of human embryonic stem cell-associated signatures in tumors

    Directory of Open Access Journals (Sweden)

    Wang Xiaosheng

    2011-10-01

    Full Text Available Abstract Background The cancer stem cell model has been proposed based on the linkage between human embryonic stem cells and human cancer cells. However, the evidences supporting the cancer stem cell model remain to be collected. In this study, we extensively examined the expression of human embryonic stem cell-associated signatures including core genes, transcription factors, pathways and microRNAs in various cancers using the computational biology approach. Results We used the class comparison analysis and survival analysis algorithms to identify differentially expressed genes and their associated transcription factors, pathways and microRNAs among normal vs. tumor or good prognosis vs. poor prognosis phenotypes classes based on numerous human cancer gene expression data. We found that most of the human embryonic stem cell- associated signatures were frequently identified in the analysis, suggesting a strong linkage between human embryonic stem cells and cancer cells. Conclusions The present study revealed the close linkage between the human embryonic stem cell associated gene expression profiles and cancer-associated gene expression profiles, and therefore offered an indirect support for the cancer stem cell theory. However, many interest issues remain to be addressed further.

  13. Neurotrophin and Trk expression by cells of the human lamina cribrosa following oxygen-glucose deprivation

    OpenAIRE

    Clark Abbot F; Lambert Wendi S; Wordinger Robert J

    2004-01-01

    Abstract Background Ischemia within the optic nerve head (ONH) may contribute to retinal ganglion cell (RGC) loss in primary open angle glaucoma (POAG). Ischemia has been reported to increase neurotrophin and high affinity Trk receptor expression by CNS neurons and glial cells. We have previously demonstrated neurotrophin and Trk expression within the lamina cribrosa (LC) region of the ONH. To determine if ischemia alters neurotrophin and Trk protein expression in cells from the human LC, cul...

  14. Lineage-specific expression of bestrophin-2 and bestrophin-4 in human intestinal epithelial cells.

    Directory of Open Access Journals (Sweden)

    Go Ito

    Full Text Available Intestinal epithelial cells (IECs regulate the absorption and secretion of anions, such as HCO3(- or Cl(-. Bestrophin genes represent a newly identified group of calcium-activated Cl(- channels (CaCCs. Studies have suggested that, among the four human bestrophin-family genes, bestrophin-2 (BEST2 and bestrophin-4 (BEST4 might be expressed within the intestinal tissue. Consistently, a study showed that BEST2 is expressed by human colonic goblet cells. However, their precise expression pattern along the gastrointestinal tract, or the lineage specificity of the cells expressing these genes, remains largely unknown. Here, we show that BEST2 and BEST4 are expressed in vivo, each in a distinct, lineage-specific manner, in human IECs. While BEST2 was expressed exclusively in colonic goblet cells, BEST4 was expressed in the absorptive cells of both the small intestine and the colon. In addition, we found that BEST2 expression is significantly down-regulated in the active lesions of ulcerative colitis, where goblet cells were depleted, suggesting that BEST2 expression is restricted to goblet cells under both normal and pathologic conditions. Consistently, the induction of goblet cell differentiation by a Notch inhibitor, LY411575, significantly up-regulated the expression of not BEST4 but BEST2 in MUC2-positive HT-29 cells. Conversely, the induction of absorptive cell differentiation up-regulated the expression of BEST4 in villin-positive Caco-2 cells. In addition, we found that the up- or down-regulation of Notch activity leads to the preferential expression of either BEST4 or BEST2, respectively, in LS174T cells. These results collectively confirmed that BEST2 and BEST4 could be added to the lineage-specific genes of humans IECs due to their abilities to clearly identify goblet cells of colonic origin and a distinct subset of absorptive cells, respectively.

  15. Duchenne Muscular Dystrophy Gene Expression in Normal and Diseased Human Muscle

    Science.gov (United States)

    Oronzi Scott, M.; Sylvester, J. E.; Heiman-Patterson, T.; Shi, Y.-J.; Fieles, W.; Stedman, H.; Burghes, A.; Ray, P.; Worton, R.; Fischbeck, K. H.

    1988-03-01

    A probe for the 5' end of the Duchenne muscular dystrophy (DMD) gene was used to study expression of the gene in normal human muscle, myogenic cell cultures, and muscle from patients with DMD. Expression was found in RNA from normal fetal muscle, adult cardiac and skeletal muscle, and cultured muscle after myoblast fusion. In DMD muscle, expression of this portion of the gene was also revealed by in situ RNA hybridization, particularly in regenerating muscle fibers.

  16. Expression of thyroid hormone transporters in the human placenta and changes associated with intrauterine growth restriction.

    Science.gov (United States)

    Loubière, L S; Vasilopoulou, E; Bulmer, J N; Taylor, P M; Stieger, B; Verrey, F; McCabe, C J; Franklyn, J A; Kilby, M D; Chan, S-Y

    2010-04-01

    Thyroid hormones (TH) are important for the development of the human fetus and placenta from very early gestation. The transplacental passage of TH from mother to fetus and the supply of TH into trophoblasts require the expression of placental TH plasma membrane transporters. We describe the ontogeny of the TH transporters MCT8, MCT10, LAT1, LAT2, OATP1A2 and OATP4A1 in a large series (n = 110) of normal human placentae across gestation and describe their expression changes with intrauterine fetal growth restriction (IUGR n = 22). Quantitative RT-PCR revealed that all the mRNAs encoding TH transporters are expressed in human placenta from 6 weeks gestation and throughout pregnancy. MCT8, MCT10, OATP1A2 and LAT1 mRNA expression increased with gestation. OATP4A1 and CD98 (LATs obligatory associated protein) mRNA expression reached a nadir in mid-gestation before increasing towards term. LAT2 mRNA expression did not alter throughout gestation. Immunohistochemistry localised MCT10 and OATP1A2 to villous cytotrophoblasts and syncytiotrophoblasts, and extravillous trophoblasts while OATP4A1 was preferentially expressed in the villous syncytiotrophoblasts. Whilst MCT8 protein expression was increased, MCT10 mRNA expression was decreased in placentae from IUGR pregnancies delivered in the early 3rd trimester compared to age matched appropriately grown for gestational age controls. No significant change was found in the mRNA expression of the other transporters with IUGR. In conclusion, several TH transporters are present in the human placenta from early 1st trimester with varying patterns of expression throughout gestation. Their coordinated effects may regulate both transplacental TH passage and TH supply to trophoblasts, which are critical for the normal development of the fetus and placenta. Increased MCT8 and decreased MCT10 expression within placentae of pregnancies complicated by IUGR may contribute to aberrant development of the fetoplacental unit.

  17. Functional hemichannels formed by human connexin 26 expressed in bacteria.

    Science.gov (United States)

    Fiori, Mariana C; Krishnan, Srinivasan; Cortes, D Marien; Retamal, Mauricio A; Reuss, Luis; Altenberg, Guillermo A; Cuello, Luis G

    2015-03-18

    Gap-junction channels (GJCs) communicate the cytoplasm of adjacent cells and are formed by head-to-head association of two hemichannels (HCs), one from each of the neighbouring cells. GJCs mediate electrical and chemical communication between cells, whereas undocked HCs participate in paracrine signalling because of their permeability to molecules such as ATP. Sustained opening of HCs under pathological conditions results in water and solute fluxes that cannot be compensated by membrane transport and therefore lead to cell damage. Mutations of Cx26 (connexin 26) are the most frequent cause of genetic deafness and it is therefore important to understand the structure-function relationship of wild-type and deafness-associated mutants. Currently available connexin HC expression systems severely limit the pace of structural studies and there is no simple high-throughput HC functional assay. The Escherichia coli-based expression system presented in the present study yields milligram amounts of purified Cx26 HCs suitable for functional and structural studies. We also show evidence of functional activity of recombinant Cx26 HCs in intact bacteria using a new growth complementation assay. The E. coli-based expression system has high potential for structural studies and high-throughput functional screening of HCs.

  18. RAS AND p53 EXPRESSION IN HUMAN THYROID CARCINOMA

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    Objective: To investigate the possible interaction between the ras and p53 genes over-expression in thyroid carcinoma, and whether there is a correlation between the ras and p53 over-expression and clinicopathological criteria. Methods: Eighty patients with thyroid lesions were examined for expression of ras and p53 genes by the labeled streptavidin biotin peroxidase (LSAB) method. Of these patients, 54 were diagnosed (average age: 39.9± 15.9 years) with malignant lesions. Of those included in the study, 31 has papillary carcinoma, 13 had follicular carcinoma, 7 had medullary carcinoma, 3 had undifferentiated carcinoma and 19 were stratified to stage I, 28 to stage II, 2 to stage III and 5 to stage IV according to TNM staging system. Twenty-six benign nodular thyroid disorders were studied as control. Results: Positive immunostain results for ras and p53 genes were statistically significant between thyroid carcinomas and benign disorders (90.7% vs 23%, 55.5% vs 30.7%, P<0.05). Both p53 and ras overexpressions coexisted in 30 thyroid carcinomas, and of these, 3 died and 5 had recurrences within 4 years. Conclusions: Activation of ras gene and inactivation of p53 gene were cooperatively associated in thyroid tumorigenesis. The concurrent overexpressions of ras and p53 could result in a poor prognosis.

  19. COX-2 expression positively correlates with PD-L1 expression in human melanoma cells.

    Science.gov (United States)

    Botti, Gerardo; Fratangelo, Federica; Cerrone, Margherita; Liguori, Giuseppina; Cantile, Monica; Anniciello, Anna Maria; Scala, Stefania; D'Alterio, Crescenzo; Trimarco, Chiara; Ianaro, Angela; Cirino, Giuseppe; Caracò, Corrado; Colombino, Maria; Palmieri, Giuseppe; Pepe, Stefano; Ascierto, Paolo Antonio; Sabbatino, Francesco; Scognamiglio, Giosuè

    2017-02-23

    The resistance to PD-1/PD-L1 inhibitors for the treatment of melanoma have prompted investigators to implement novel clinical trials which combine immunotherapy with different treatment modalities. Moreover is also important to investigate the mechanisms which regulate the dynamic expression of PD-L1 on tumor cells and PD-1 on T cells in order to identify predictive biomarkers of response. COX-2 is currently investigated as a major player of tumor progression in several type of malignancies including melanoma. In the present study we investigated the potential relationship between COX-2 and PD-L1 expression in melanoma. Tumor samples obtained from primary melanoma lesions and not matched lymph node metastases were analyzed for both PD-L1 and COX-2 expression by IHC analysis. Status of BRAF and NRAS mutations was analyzed by sequencing and PCR. Co-localization of PD-L1 and COX-2 expression was analyzed by double fluorescence staining. Lastly the BRAF(V600E) A375 and NRAS(Q61R) SK-MEL-2 melanoma cell lines were used to evaluate the effect of COX-2 inhibition by celecoxib on expression of PD-L1 in vitro. BRAF(V600E/V600K) and NRAS(Q61R/Q61L) were detected in 57.8 and 8.9% of the metastatic lesions, and in 65.9 and 6.8% of the primary tumors, respectively. PD-L1 and COX-2 expression were heterogeneously expressed in both primary melanoma lesions and not matched lymph node metastases. A significantly lower number of PD-L1 negative lesions was found in primary tumors as compared to not matched metastatic lesions (P = 0.002). COX-2 expression significantly correlated with PD-L1 expression in both primary (P = 0.001) and not matched metastatic (P = 0.048) lesions. Furthermore, in melanoma tumors, cancer cells expressing a higher levels of COX-2 also co-expressed a higher level of PD-L1. Lastly, inhibition of COX-2 activity by celecoxib down-regulated the expression of PD-L1 in both BRAF(V600E) A375 and NRAS(Q61R) SK-MEL-2 melanoma cell lines. COX-2 expression

  20. Transcriptional and epigenetic regulation of KIAA1199 gene expression in human breast cancer.

    Directory of Open Access Journals (Sweden)

    Cem Kuscu

    Full Text Available Emerging evidence has demonstrated that upregulated expression of KIAA1199 in human cancer bodes for poor survival. The regulatory mechanism controlling KIAA1199 expression in cancer remains to be characterized. In the present study, we have isolated and characterized the human KIAA1199 promoter in terms of regulation of KIAA1199 gene expression. A 3.3 kb fragment of human genomic DNA containing the 5'-flanking sequence of the KIAA1199 gene possesses both suppressive and activating elements. Employing a deletion mutagenesis approach, a 1.4 kb proximal region was defined as the basic KIAA1199 promoter containing a TATA-box close to the transcription start site. A combination of 5'-primer extension study with 5'RACE DNA sequencing analysis revealed one major transcription start site that is utilized in the human KIAA1199 gene. Bioinformatics analysis suggested that the 1.4 kb KIAA1199 promoter contains putative activating regulatory elements, including activator protein-1(AP-1, Twist-1, and NF-κB sites. Sequential deletion and site-direct mutagenesis analysis demonstrated that the AP-1 and distal NF-κB sites are required for KIAA1199 gene expression. Further analyses using an electrophoretic mobility-shift assay and chromatin immunoprecipitation confirmed the requirement of these cis- and trans-acting elements in controlling KIAA1199 gene expression. Finally, we found that upregulated KIAA1199 expression in human breast cancer specimens correlated with hypomethylation of the regulatory region. Involvement of DNA methylation in regulation of KIAA1199 expression was recapitulated in human breast cancer cell lines. Taken together, our study unraveled the regulatory mechanisms controlling KIAA1199 gene expression in human cancer.

  1. Profiling of olfactory receptor gene expression in whole human olfactory mucosa.

    Directory of Open Access Journals (Sweden)

    Christophe Verbeurgt

    Full Text Available Olfactory perception is mediated by a large array of olfactory receptor genes. The human genome contains 851 olfactory receptor gene loci. More than 50% of the loci are annotated as nonfunctional due to frame-disrupting mutations. Furthermore haplotypic missense alleles can be nonfunctional resulting from substitution of key amino acids governing protein folding or interactions with signal transduction components. Beyond their role in odor recognition, functional olfactory receptors are also required for a proper targeting of olfactory neuron axons to their corresponding glomeruli in the olfactory bulb. Therefore, we anticipate that profiling of olfactory receptor gene expression in whole human olfactory mucosa and analysis in the human population of their expression should provide an opportunity to select the frequently expressed and potentially functional olfactory receptors in view of a systematic deorphanization. To address this issue, we designed a TaqMan Low Density Array (Applied Biosystems, containing probes for 356 predicted human olfactory receptor loci to investigate their expression in whole human olfactory mucosa tissues from 26 individuals (13 women, 13 men; aged from 39 to 81 years, with an average of 67±11 years for women and 63±12 years for men. Total RNA isolation, DNase treatment, RNA integrity evaluation and reverse transcription were performed for these 26 samples. Then 384 targeted genes (including endogenous control genes and reference genes specifically expressed in olfactory epithelium for normalization purpose were analyzed using the same real-time reverse transcription PCR platform. On average, the expression of 273 human olfactory receptor genes was observed in the 26 selected whole human olfactory mucosa analyzed, of which 90 were expressed in all 26 individuals. Most of the olfactory receptors deorphanized to date on the basis of sensitivity to known odorant molecules, which are described in the literature, were

  2. Factors affecting the gene expression of in vitro cultured human preimplantation embryos

    NARCIS (Netherlands)

    Mantikou, E.; Jonker, M.J.; Wong, K.M.; van Montfoort, A.P.A.; de Jong, M.; Breit, T.M.; Repping, S.; Mastenbroek, S.

    2016-01-01

    STUDY QUESTION: What is the relative effect of common environmental and biological factors on transcriptome changes during human preimplantation development? SUMMARY ANSWER: Developmental stage and maternal age had a larger effect on the global gene expression profile of human preimplantation embryo

  3. Subcellular localization and N-glycosylation of human ABCC6, expressed in MDCKII cells.

    NARCIS (Netherlands)

    Sinko, E; Ilias, A; Ujhelly, O; Homolya, L; Scheffer, G.L.; Bergen, AA; Sarkadi, B; Varadi, A

    2003-01-01

    Mutations in the gene coding for a human ABC transporter protein, ABCC6 (MRP6), are responsible for the development of pseudoxanthoma elasticum. Here, we demonstrate that human ABCC6, when expressed by retroviral transduction in polarized mammalian (MDCKII) cells, is exclusively localized to the bas

  4. Impact of cigarette smoke on the human and mouse lungs: a gene-expression comparison study.

    Directory of Open Access Journals (Sweden)

    Mathieu C Morissette

    Full Text Available Cigarette smoke is well known for its adverse effects on human health, especially on the lungs. Basic research is essential to identify the mechanisms involved in the development of cigarette smoke-related diseases, but translation of new findings from pre-clinical models to the clinic remains difficult. In the present study, we aimed at comparing the gene expression signature between the lungs of human smokers and mice exposed to cigarette smoke to identify the similarities and differences. Using human and mouse whole-genome gene expression arrays, changes in gene expression, signaling pathways and biological functions were assessed. We found that genes significantly modulated by cigarette smoke in humans were enriched for genes modulated by cigarette smoke in mice, suggesting a similar response of both species. Sixteen smoking-induced genes were in common between humans and mice including six newly reported to be modulated by cigarette smoke. In addition, we identified a new conserved pulmonary response to cigarette smoke in the induction of phospholipid metabolism/degradation pathways. Finally, the majority of biological functions modulated by cigarette smoke in humans were also affected in mice. Altogether, the present study provides information on similarities and differences in lung gene expression response to cigarette smoke that exist between human and mouse. Our results foster the idea that animal models should be used to study the involvement of pathways rather than single genes in human diseases.

  5. High glucose-induced oxidative stress increases transient receptor potential channel expression in human monocytes

    DEFF Research Database (Denmark)

    Wuensch, Tilo; Thilo, Florian; Krueger, Katharina;

    2010-01-01

    Transient receptor potential (TRP) channel-induced cation influx activates human monocytes, which play an important role in the pathogenesis of atherosclerosis. In the present study, we investigated the effects of high glucose-induced oxidative stress on TRP channel expression in human monocytes....

  6. Differential expression of glutamic acid decarboxylase in rat and human islets

    DEFF Research Database (Denmark)

    Petersen, J S; Russel, S; Marshall, M O;

    1993-01-01

    The GABA synthesizing enzyme GAD is a prominent islet cell autoantigen in type I diabetes. The two forms of GAD (GAD64 and GAD67) are encoded by different genes in both rats and humans. By in situ hybridization analysis of rat and human pancreases, expression of both genes was detected in rat isl...

  7. Dissecting Daily and Circadian Expression Rhythms of Clock-Controlled Genes in Human Blood

    NARCIS (Netherlands)

    K. Lech (Karolina); K. Ackermann (Katrin); V.L. Revell (Victoria); O.S.C.A.R. Lao; D.J. Skene (Debra); M.H. Kayser (Manfred)

    2016-01-01

    textabstractThe identification and investigation of novel clock-controlled genes (CCGs) has been conducted thus far mainly in model organisms such as nocturnal rodents, with limited information in humans. Here, we aimed to characterize daily and circadian expression rhythms of CCGs in human

  8. Endothelin converting-enzyme-1 mRNA expression in human cardiovascular disease

    NARCIS (Netherlands)

    Bohnemeier, H; Pinto, YM; Horkay, F; Toth, M; Juhasz-Nagy, A; Orzechowski, HD; Bohm, M; Paul, M

    1998-01-01

    Endothelin-1 converting-enzyme (ECE-1) cleaves the precursor, big-endothelin-1, to the active peptide endothelin-1. The aim of this study was to investigate whether ECE-1 mRNA expression is modified in human cardiovascular disease. Tissue samples from the left human atrium were analyzed for ECE-1 ex

  9. Expression and roles of Slit/Robo in human ovarian cancer.

    Science.gov (United States)

    Dai, Cai Feng; Jiang, Yi Zhou; Li, Yan; Wang, Kai; Liu, Pei Shu; Patankar, Manish S; Zheng, Jing

    2011-05-01

    The Slit glycoproteins and their Roundabout (Robo) receptors regulate migration and growth of many types of cells including human cancer cells. However, little is known about the expression and roles of Slit/Robo in human ovarian cancer. Herein, we examined the expression of Slit/Robo in human normal and malignant ovarian tissues and its potential participation in regulating migration and proliferation of human ovarian cancer cells using two ovarian cancer cell lines, OVCAR-3 and SKOV-3. We demonstrated that Slit2/3 and Robo1 were immunolocalized primarily in stromal cells in human normal ovaries and in cancer cells in many histotypes of ovarian cancer tissues. Protein expression of Slit2/3 and Robo1/4 was also identified in OVCAR-3 and SKOV-3 cells. However, recombinant human Slit2 did not significantly affect SKOV-3 cell migration, and OVCAR-3 and SKOV-3 cell proliferation. Slit2 also did not induce ERK1/2 and AKT1 phosphorylation in OVCAR-3 and SKOV-3 cells. The current findings indicate that three major members (Slit2/3 and Robo1) of Slit/Robo family are widely expressed in the human normal and malignant ovarian tissues and in OVCAR-3 and SKOV-3 cells. However, Slit/Robo signaling may not play an important role in regulating human ovarian cancer cell proliferation and migration.

  10. An RNA gene expressed during cortical development evolved rapidly in humans

    DEFF Research Database (Denmark)

    Pollard, Katherine S; Salama, Sofie R; Lambert, Nelle;

    2006-01-01

    in the developing human neocortex from 7 to 19 gestational weeks, a crucial period for cortical neuron specification and migration. HAR1F is co-expressed with reelin, a product of Cajal-Retzius neurons that is of fundamental importance in specifying the six-layer structure of the human cortex. HAR1 and the other...

  11. Expression of a humanized viral 2A-mediated lux operon efficiently generates autonomous bioluminescence in human cells.

    Directory of Open Access Journals (Sweden)

    Tingting Xu

    Full Text Available Expression of autonomous bioluminescence from human cells was previously reported to be impossible, suggesting that all bioluminescent-based mammalian reporter systems must therefore require application of a potentially influential chemical substrate. While this was disproven when the bacterial luciferase (lux cassette was demonstrated to function in a human cell, its expression required multiple genetic constructs, was functional in only a single cell type, and generated a significantly reduced signal compared to substrate-requiring systems. Here we investigate the use of a humanized, viral 2A-linked lux genetic architecture for the efficient introduction of an autobioluminescent phenotype across a variety of human cell lines.The lux cassette was codon optimized and assembled into a synthetic human expression operon using viral 2A elements as linker regions. Human kidney, breast cancer, and colorectal cancer cell lines were both transiently and stably transfected with the humanized operon and the resulting autobioluminescent phenotype was evaluated using common imaging instrumentation. Autobioluminescent cells were screened for cytotoxic effects resulting from lux expression and their utility as bioreporters was evaluated through the demonstration of repeated monitoring of single populations over a prolonged period using both a modified E-SCREEN assay for estrogen detection and a classical cytotoxic compound detection assay for the antibiotic Zeocin. Furthermore, the use of self-directed bioluminescent initiation in response to target detection was assessed to determine its amenability towards deployment as fully autonomous sensors. In all cases, bioluminescent measurements were supported with traditional genetic and transcriptomic evaluations.Our results demonstrate that the viral 2A-linked, humanized lux genetic architecture successfully produced autobioluminescent phenotypes in all cell lines tested without the induction of cytotoxicity

  12. GSTCD and INTS12 regulation and expression in the human lung.

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    Ma'en Obeidat

    Full Text Available Genome-Wide Association Study (GWAS meta-analyses have identified a strong association signal for lung function, which maps to a region on 4q24 containing two oppositely transcribed genes: glutathione S-transferase, C-terminal domain containing (GSTCD and integrator complex subunit 12 (INTS12. Both genes were found to be expressed in a range of human airway cell types. The promoter regions and transcription start sites were determined in mRNA from human lung and a novel splice variant was identified for each gene. We obtained the following evidence for GSTCD and INTS12 co-regulation and expression: (i correlated mRNA expression was observed both via Q-PCR and in a lung expression quantitative trait loci (eQTL study, (ii induction of both GSTCD and INTS12 mRNA expression in human airway smooth muscle cells was seen in response to TGFβ1, (iii a lung eQTL study revealed that both GSTCD and INTS12 mRNA levels positively correlate with percent predicted FEV1, and (iv FEV1 GWAS associated SNPs in 4q24 were found to act as an eQTL for INTS12 in a number of tissues. In fixed sections of human lung tissue, GSTCD protein expression was ubiquitous, whereas INTS12 expression was predominantly in epithelial cells and pneumocytes. During human fetal lung development, GSTCD protein expression was observed to be highest at the earlier pseudoglandular stage (10-12 weeks compared with the later canalicular stage (17-19 weeks, whereas INTS12 expression levels did not alter throughout these stages. Knowledge of the transcriptional and translational regulation and expression of GSTCD and INTS12 provides important insights into the potential role of these genes in determining lung function. Future work is warranted to fully define the functions of INTS12 and GSTCD.

  13. Identification of differentially expressed microRNAs in human male breast cancer

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    Schipper Elisa

    2010-03-01

    Full Text Available Abstract Background The discovery of small non-coding RNAs and the subsequent analysis of microRNA expression patterns in human cancer specimens have provided completely new insights into cancer biology. Genetic and epigenetic data indicate oncogenic or tumor suppressor function of these pleiotropic regulators. Therefore, many studies analyzed the expression and function of microRNA in human breast cancer, the most frequent malignancy in females. However, nothing is known so far about microRNA expression in male breast cancer, accounting for approximately 1% of all breast cancer cases. Methods The expression of 319 microRNAs was analyzed in 9 primary human male breast tumors and in epithelial cells from 15 male gynecomastia specimens using fluorescence-labeled bead technology. For identification of differentially expressed microRNAs data were analyzed by cluster analysis and selected statistical methods. Expression levels were validated for the most up- or down-regulated microRNAs in this training cohort using real-time PCR methodology as well as in an independent test cohort comprising 12 cases of human male breast cancer. Results Unsupervised cluster analysis separated very well male breast cancer samples and control specimens according to their microRNA expression pattern indicating cancer-specific alterations of microRNA expression in human male breast cancer. miR-21, miR519d, miR-183, miR-197, and miR-493-5p were identified as most prominently up-regulated, miR-145 and miR-497 as most prominently down-regulated in male breast cancer. Conclusions Male breast cancer displays several differentially expressed microRNAs. Not all of them are shared with breast cancer biopsies from female patients indicating male breast cancer specific alterations of microRNA expression.

  14. Recombinant expression of hydroxylated human collagen in Escherichia coli

    OpenAIRE

    Rutschmann, Christoph; Baumann, Stephan; Cabalzar, Jürg; Luther, Kelvin B.; Hennet, Thierry

    2014-01-01

    Collagen is the most abundant protein in the human body and thereby a structural protein of considerable biotechnological interest. The complex maturation process of collagen, including essential post-translational modifications such as prolyl and lysyl hydroxylation, has precluded large-scale production of recombinant collagen featuring the biophysical properties of endogenous collagen. The characterization of new prolyl and lysyl hydroxylase genes encoded by the giant virus mimivirus reveal...

  15. Similarity of GATA-3 Expression between Rat and Human Mammary Glands

    Science.gov (United States)

    Kinoshita, Yuichi; Yoshizawa, Katsuhiko; Emoto, Yuko; Yuki, Michiko; Yuri, Takashi; Shikata, Nobuaki; Tsubura, Airo

    2014-01-01

    The GATA family members are zinc finger transcription factors involved in cell differentiation and proliferation. In particular, GATA-3 is necessary for mammary gland maturation and is a useful marker in the characterization of mammary carcinoma in humans. The expression of GATA-3 protein in normal mammary glands, fibroadenomas and carcinomas was immunohistochemically compared in female rats and humans. In normal mammary glands of rats and humans, scattered luminal cells in the acini and whole ductal epithelial cells were positive for GATA-3 in the nuclei. No positive cells were detected in rat or human fibroadenomas. In rat and human mammary carcinomas, the nuclei of proliferating luminal-derived cancer cells expressed GATA-3. Therefore, GATA-3 protein is a candidate marker for mammary carcinoma in rats as well as humans. PMID:25352719

  16. Expression of the homeobox genes OTX2 and OTX1 in the early developing human brain

    DEFF Research Database (Denmark)

    Larsen, Karen B; Lutterodt, Melissa C; Møllgård, Kjeld

    2010-01-01

    protein was found in the subcommissural organ, pineal gland, and cerebellum. The early expression of OTX2 and OTX1 in proliferative cell layers of the human fetal brain supports the concept that these homeobox genes are important in neuronal cell development and differentiation: OTX1 primarily...... of young neurons of the deeper cortical layers. We have studied the spatial and temporal expression of the two homeobox genes OTX2 and OTX1 in human fetal brains from 7 to 14 weeks postconception by in situ hybridization and immunohistochemistry. OTX2 was expressed in the diencephalon, mesencephalon...... in the neocortex, and OTX2 in the archicortex, diencephalon, rostral brain stem, and cerebellum....

  17. Vitamin D receptor and vitamin D metabolizing enzymes are expressed in the human male reproductive tract

    DEFF Research Database (Denmark)

    Blomberg Jensen, Martin; Nielsen, John E; Jørgensen, Anne

    2010-01-01

    , since it is not solely dependent on VDR expression, but also on cellular uptake of circulating VD and presence and activity of VD metabolizing enzymes. Expression of VD metabolizing enzymes has not previously been investigated in human testis and male reproductive tract. Therefore, we performed......The vitamin D receptor (VDR) is expressed in human testis, and vitamin D (VD) has been suggested to affect survival and function of mature spermatozoa. Indeed, VDR knockout mice and VD deficient rats show decreased sperm counts and low fertility. However, the cellular response to VD is complex...

  18. Assessment of differential gene expression in human peripheral nerve injury

    Directory of Open Access Journals (Sweden)

    Sangameswaran Lakshmi

    2002-09-01

    Full Text Available Abstract Background Microarray technology is a powerful methodology for identifying differentially expressed genes. However, when thousands of genes in a microarray data set are evaluated simultaneously by fold changes and significance tests, the probability of detecting false positives rises sharply. In this first microarray study of brachial plexus injury, we applied and compared the performance of two recently proposed algorithms for tackling this multiple testing problem, Significance Analysis of Microarrays (SAM and Westfall and Young step down adjusted p values, as well as t-statistics and Welch statistics, in specifying differential gene expression under different biological states. Results Using SAM based on t statistics, we identified 73 significant genes, which fall into different functional categories, such as cytokines / neurotrophin, myelin function and signal transduction. Interestingly, all but one gene were down-regulated in the patients. Using Welch statistics in conjunction with SAM, we identified an additional set of up-regulated genes, several of which are engaged in transcription and translation regulation. In contrast, the Westfall and Young algorithm identified only one gene using a conventional significance level of 0.05. Conclusion In coping with multiple testing problems, Family-wise type I error rate (FWER and false discovery rate (FDR are different expressions of Type I error rates. The Westfall and Young algorithm controls FWER. In the context of this microarray study, it is, seemingly, too conservative. In contrast, SAM, by controlling FDR, provides a promising alternative. In this instance, genes selected by SAM were shown to be biologically meaningful.

  19. Differential gene expression in human granulosa cells from recombinant FSH versus human menopausal gonadotropin ovarian stimulation protocols

    Directory of Open Access Journals (Sweden)

    Bietz Mandi G

    2010-03-01

    Full Text Available Abstract Background The study was designed to test the hypothesis that granulosa cell (GC gene expression response differs between recombinant FSH and human menopausal gonadotropin (hMG stimulation regimens. Methods Females Results After exclusions, 1736 genes exhibited differential expression between groups. Over 400 were categorized as signal transduction genes, ~180 as transcriptional regulators, and ~175 as enzymes/metabolic genes. Expression of selected genes was confirmed by RT-PCR. Differentially expressed genes included A kinase anchor protein 11 (AKAP11, bone morphogenetic protein receptor II (BMPR2, epidermal growth factor (EGF, insulin-like growth factor binding protein (IGFBP-4, IGFBP-5, and hypoxia-inducible factor (HIF-1 alpha. Conclusions Results suggest that major differences exist in the mechanism by which pure FSH alone versus FSH/LH regulate gene expression in preovulatory GC that could impact oocyte maturity and developmental competence.

  20. Expression, purification and characterization of human Dopamine ß-monooxygenase

    DEFF Research Database (Denmark)

    Vendelboe, Trine Vammen

    This thesis deals with expression, purification and characterization of the copper containing enzyme dopamine ß-monooxygenase (DBM). DBM is an ascorbate dependent protein that requires Cu in the active site in order to be functional. DBM is made of four domains; An Nterminal DOMON domain, the two...... others, one of the reasons why these proteins are considered to follow the same mechanism. DBM converts dopamine (DA) into Norepinphrine (NE). Both substrate and product functions as neurotransmitters and the levels of these are involved in many different disorders such as depression and hypertension...

  1. Expression, purification and characterization of human Dopamine ß-monooxygenase

    DEFF Research Database (Denmark)

    Vendelboe, Trine Vammen

    This thesis deals with expression, purification and characterization of the copper containing enzyme dopamine ß-monooxygenase (DBM). DBM is an ascorbate dependent protein that requires Cu in the active site in order to be functional. DBM is made of four domains; An Nterminal DOMON domain, the two...... others, one of the reasons why these proteins are considered to follow the same mechanism. DBM converts dopamine (DA) into Norepinphrine (NE). Both substrate and product functions as neurotransmitters and the levels of these are involved in many different disorders such as depression and hypertension...

  2. Expression of serotonin receptors in human lower esophageal sphincter

    OpenAIRE

    Li, He-Fei; Liu, Jun-Feng; Zhang, Ke; Feng, Yong

    2014-01-01

    Serotonin (5-HT) is a neurotransmitter and vasoactive amine that is involved in the regulation of a large number of physiological functions. The wide variety of 5-HT-mediated functions is due to the existence of different classes of serotonergic receptors in the mammalian gastrointestinal tract and nervous system. The aim of this study was to explore the expression of multiple types of 5-HT receptor (5-HT1AR, 5-HT2AR, 5-HT3AR, 5-HT4R, 5-HT5AR, 5-HT6R and 5-HT7R) in sling and clasp fibers from...

  3. Posttranscriptional control of human gamma interferon gene expression in transfected mouse fibroblasts.

    OpenAIRE

    1986-01-01

    Human gamma interferon genomic DNA was introduced into NIH 3T3 fibroblasts by calcium phosphate precipitation and was not expressed in these cells at the cytoplasmic mRNA or protein level. Treatment of the transfected cells with cycloheximide (1 microgram/ml) induced the accumulation of cytoplasmic gamma interferon mRNA and biologically active human gamma interferon. Analysis of the nuclear enriched RNA from untreated cells indicated that human gamma interferon mRNA was present, suggesting th...

  4. Short Chain Fatty Acids (SCFA) Reprogram Gene Expression in Human Malignant Epithelial and Lymphoid Cells

    OpenAIRE

    Lidiia Astakhova; Mtakai Ngara; Olga Babich; Aleksandr Prosekov; Lyudmila Asyakina; Lyubov Dyshlyuk; Tore Midtvedt; Xiaoying Zhou; Ingemar Ernberg; Liudmila Matskova

    2016-01-01

    The effect of short chain fatty acids (SCFAs) on gene expression in human, malignant cell lines was investigated, with a focus on signaling pathways. The commensal microbial flora produce high levels of SCFAs with established physiologic effects in humans. The most abundant SCFA metabolite in the human microflora is n-butyric acid. It is well known to activate endogenous latent Epstein-Barr virus (EBV), that was used as a reference read out system and extended to EBV+ epithelial cancer cell l...

  5. Comparison of epigenetic mediator expression and function in mouse and human embryonic blastomeres

    OpenAIRE

    Chavez, Shawn L.; McElroy, Sohyun L.; Bossert, Nancy L.; De Jonge, Christopher J.; Rodriguez, Maria Vera; Denise E Leong; Behr, Barry; Westphal, Lynn M.; Reijo Pera, Renee A.

    2014-01-01

    A map of human embryo development that combines imaging, molecular, genetic and epigenetic data for comparisons to other species and across pathologies would be greatly beneficial for basic science and clinical applications. Here, we compared mRNA and protein expression of key mediators of DNA methylation and histone modifications between mouse and human embryos, embryos from fertile/infertile couples, and following growth factor supplementation. We observed that individual mouse and human em...

  6. Computational analysis of expression of human embryonic stem cell-associated signatures in tumors

    OpenAIRE

    Wang, Xiaosheng

    2011-01-01

    Background The cancer stem cell model has been proposed based on the linkage between human embryonic stem cells and human cancer cells. However, the evidences supporting the cancer stem cell model remain to be collected. In this study, we extensively examined the expression of human embryonic stem cell-associated signatures including core genes, transcription factors, pathways and microRNAs in various cancers using the computational biology approach. Results We used the class comparison analy...

  7. Computational analysis of expression of human embryonic stem cell-associated signatures in tumors

    OpenAIRE

    Wang Xiaosheng

    2011-01-01

    Abstract Background The cancer stem cell model has been proposed based on the linkage between human embryonic stem cells and human cancer cells. However, the evidences supporting the cancer stem cell model remain to be collected. In this study, we extensively examined the expression of human embryonic stem cell-associated signatures including core genes, transcription factors, pathways and microRNAs in various cancers using the computational biology approach. Results We used the class compari...

  8. Constitutively Expressed IFITM3 Protein in Human Endothelial Cells Poses an Early Infection Block to Human Influenza Viruses.

    Science.gov (United States)

    Sun, Xiangjie; Zeng, Hui; Kumar, Amrita; Belser, Jessica A; Maines, Taronna R; Tumpey, Terrence M

    2016-12-15

    A role for pulmonary endothelial cells in the orchestration of cytokine production and leukocyte recruitment during influenza virus infection, leading to severe lung damage, has been recently identified. As the mechanistic pathway for this ability is not fully known, we extended previous studies on influenza virus tropism in cultured human pulmonary endothelial cells. We found that a subset of avian influenza viruses, including potentially pandemic H5N1, H7N9, and H9N2 viruses, could infect human pulmonary endothelial cells (HULEC) with high efficiency compared to human H1N1 or H3N2 viruses. In HULEC, human influenza viruses were capable of binding to host cellular receptors, becoming internalized and initiating hemifusion but failing to uncoat the viral nucleocapsid and to replicate in host nuclei. Unlike numerous cell types, including epithelial cells, we found that pulmonary endothelial cells constitutively express a high level of the restriction protein IFITM3 in endosomal compartments. IFITM3 knockdown by small interfering RNA (siRNA) could partially rescue H1N1 virus infection in HULEC, suggesting IFITM3 proteins were involved in blocking human influenza virus infection in endothelial cells. In contrast, selected avian influenza viruses were able to escape IFITM3 restriction in endothelial cells, possibly by fusing in early endosomes at higher pH or by other, unknown mechanisms. Collectively, our study demonstrates that the human pulmonary endothelium possesses intrinsic immunity to human influenza viruses, in part due to the constitutive expression of IFITM3 proteins. Notably, certain avian influenza viruses have evolved to escape this restriction, possibly contributing to virus-induced pneumonia and severe lung disease in humans. Avian influenza viruses, including H5N1 and H7N9, have been associated with severe respiratory disease and fatal outcomes in humans. Although acute respiratory distress syndrome (ARDS) and progressive pulmonary endothelial damage

  9. MHC-unrestricted lysis of MUC1-expressing cells by human peripheral blood mononuclear cells.

    Science.gov (United States)

    Wright, Stephen E; Rewers-Felkins, Kathleen A; Quinlin, Imelda S; Fogler, William E; Phillips, Catherine A; Townsend, Mary; Robinson, William; Philip, Ramila

    2008-01-01

    Many human adenocarcinomas can be killed in vitro by targeted cytotoxic T-lymphocytes (CTL); however, major histocompatibility complex (MHC)-restrictions are typically required. The MUC1 antigen is common in many human adenocarcinomas, and is associated with a variable number of tandem repeats. It has been proposed that antigens with such repeated epitopes may be vulnerable to cytotoxic T-lymphocyte killing without MHC-restriction. Therefore, it is possible that MUC1-expressing malignant cells may be killed by targeted cytotoxic T-lymphocyte in the absence of MHC-restriction. In this study, a human MUC1-expressing murine mammary carcinoma cell line was used to determine if cytotoxic T-lymphocyte killing of MUC1-expressing adenocarcinoma cells requires MHC-restriction. Specifically, MUC1-stimulated human mononuclear cells (M1SMC) were observed to kill human MUC1-transfected, MUC1-expressing murine mammary carcinoma cells, but not the mock-transfected, non-MUC1-expressing murine mammary carcinoma cells. Furthermore, the killing was blocked by antibody to MUC1, indicating MUC1-specific killing. In conclusion, cytotoxic T-lymphocyte killing of MUC1-expressing adenocarcinoma cells can be MHC-unrestricted.

  10. LINE FUSION GENES: a database of LINE expression in human genes

    Directory of Open Access Journals (Sweden)

    Park Hong-Seog

    2006-06-01

    Full Text Available Abstract Background Long Interspersed Nuclear Elements (LINEs are the most abundant retrotransposons in humans. About 79% of human genes are estimated to contain at least one segment of LINE per transcription unit. Recent studies have shown that LINE elements can affect protein sequences, splicing patterns and expression of human genes. Description We have developed a database, LINE FUSION GENES, for elucidating LINE expression throughout the human gene database. We searched the 28,171 genes listed in the NCBI database for LINE elements and analyzed their structures and expression patterns. The results show that the mRNA sequences of 1,329 genes were affected by LINE expression. The LINE expression types were classified on the basis of LINEs in the 5' UTR, exon or 3' UTR sequences of the mRNAs. Our database provides further information, such as the tissue distribution and chromosomal location of the genes, and the domain structure that is changed by LINE integration. We have linked all the accession numbers to the NCBI data bank to provide mRNA sequences for subsequent users. Conclusion We believe that our work will interest genome scientists and might help them to gain insight into the implications of LINE expression for human evolution and disease. Availability http://www.primate.or.kr/line

  11. Human platelets express Toll-like receptor 3 and respond to poly I:C.

    Science.gov (United States)

    Anabel, Antonio-Santos; Eduardo, Pérez-Campos; Pedro Antonio, Hernández-Cruz; Carlos, Solórzano-Mata; Juana, Narváez-Morales; Honorio, Torres-Aguilar; Nicolás, Villegas-Sepúlveda; Sergio Roberto, Aguilar-Ruiz

    2014-12-01

    Platelets functions in hemostasis have been widely studied. Currently, growing evidence shows that platelets have also a role in the immune innate response. Recently, protein expression of Toll-like receptors (TLR's) 2, 4, 7, 8, and 9, and the presence of TLRs 1 and 6 mRNA in human platelets was described. Up to now the functionality of TLR-2, 4 and 9 in human platelets has been demonstrated. Due to the relevance of TLRs functions to PAMPS (pathogen-associated molecular patterns) recognizing, we evaluated the presence of TLR3 in human platelets founding low percentages of platelets expressing surface or intracellular TLR3 protein. The activation with thrombin induced an increase in the percentage of platelets expressing surface TLR3 and higher levels of TLR3 expression in the whole population. Human platelets responded to poly I:C by increasing [Ca(2+)]i, the percentages of cells expressing TLR4 and CD62P, and by releasing CXCL4 and IL-1β in comparison to unstimulated platelets. These results demonstrate that human platelets express TLR3 and are capable of responding to poly I:C, suggesting that these cells might influence the immune innate response when detecting viral dsRNA. Copyright © 2014 American Society for Histocompatibility and Immunogenetics. Published by Elsevier Inc. All rights reserved.

  12. Identification of CD146 Expression in Human and Mouse Preimplantation Embryo

    Institute of Scientific and Technical Information of China (English)

    Hong-bo WANG; Xuan DU; Ya-hui XU; Ze-hua WANG

    2008-01-01

    Objective To investigate whether CD146, a cell adhesion molecule, is expressed in mouse and human preimplantation blastocysts and to localize CD146 in the layer of trophectoderm(TE) and/or inner cell mass(ICM). Methods Human and mouse embryos were collected. Using reverse transcription polymerase chain reaction(RT-PCR), the expression of CD146 mRNA in blastocyst was evaluated in human and mouse embryos. Single embryo immunohistochemical staining was applicated in the examination of the expression of CD146 in protein level. The statistical significance of the data was analyzed using t-test. Results CD146 transcript was detected in all human and mouse preimplantation morula and blastocyst. The expression of CD146 was found to localize in human and mouse compacted morula stage embryos and the TE and ICM of the expanded blastocysts. Conclusion mRNA and protein of CD146 was expressed in preimplantation embryos,which may have a profound influence on early preimplantation development for the differentiation of the trophectoderm and the morphogenesis of the blastocyst.Furthermore, the expression of CD146 in blastocyst stage may be implicated in the assistance of embryo implantation.

  13. Augmentation of arginase Ⅱ expression in the human endometrial epithelium in the secretory phase.

    Science.gov (United States)

    Tajima, Makiko; Harada, Tatsuya; Ishikawa, Tomonori; Iwahara, Yuki; Kubota, Toshiro

    2012-12-03

    L-arginine is the common substrate for arginase and nitric oxide synthase (NOS). Arginase converts L-arginine to urea and L-ornithine. L-Ornithine is the principal precursor for the production of polyamines and L-proline, which are required for cell proliferation and collagen synthesis. Endothelial NOS is expressed in the human endometrial glandular epithelium, but the expression and physiological roles of arginase in the human endometrium are not clear. The objective of this study was to investigate the expression and distribution patterns of arginases Ⅰ (A-Ⅰ) and Ⅱ (A-Ⅱ) in the human endometrium by using immunohistochemistry, reverse transcription-polymerase chain reaction (RTPCR), and western blotting. A-Ⅰ and A-Ⅱ were detected by immunohistochemistry in human endometrial epithelial cells during the proliferative and secretory phases of the menstrual cycle. RT-PCR showed that A-Ⅰ and A-Ⅱ mRNA were expressed in human endometrial tissue. Western blotting analysis results showed the expression of A-Ⅱ protein. Immunohistochemistry and western blotting results showed that expression levels of A-Ⅱ were significantly higher in the secretory phase than in the proliferative phase. Increased A-Ⅱ levels in the secretory phase may be responsible for endometrial growth by increasing polyamines and proline products.

  14. Neuronal Fibers and Neurotransmitter Receptor Expression in the Human Endolymphatic Sac

    DEFF Research Database (Denmark)

    Møller, Martin Nue; Kirkeby, Svend; Vikeså, Jonas

    2017-01-01

    in intracranial pressure homeostasis. The anatomical location towards the sigmoid sinus would suggest a possible endo- and/or paracrine signaling. However, neuronal connections may also apply, but it remains very scarcely explored in the human ES. STUDY DESIGN: DNA micro-arrays and immunohistochemistry were used...... for analyses of fresh human ES tissue samples. METHODS: A total of 30 tissue samples from the human ES were obtained during translabyrinthine surgery for vestibular schwannoma. Microarray technology was used to investigate tissue sample gene expression, using adjacent dura mater as control. The expression...

  15. Concordance of gene expression in human protein complexes reveals tissue specificity and pathology

    DEFF Research Database (Denmark)

    Börnigen, Daniela; Pers, Tune Hannes; Thorrez, Lieven

    2013-01-01

    Disease-causing variants in human genes usually lead to phenotypes specific to only a few tissues. Here, we present a method for predicting tissue specificity based on quantitative deregulation of protein complexes. The underlying assumption is that the degree of coordinated expression among...... proteins in a complex within a given tissue may pinpoint tissues that will be affected by a mutation in the complex and coordinated expression may reveal the complex to be active in the tissue. We identified known disease genes and their protein complex partners in a high-quality human interactome. Each...... susceptibility gene's tissue involvement was ranked based on coordinated expression with its interaction partners in a non-disease global map of human tissue-specific expression. The approach demonstrated high overall area under the curve (0.78) and was very successfully benchmarked against a random model...

  16. Increased expression of aquaporin-4 in human traumatic brain injury and brain tumors

    Institute of Scientific and Technical Information of China (English)

    HU Hua; YAO Hong-tian; ZHANG Wei-ping; ZHANG LEI; DING Wei; ZHANG Shi-hong; CHEN Zhong; WEI Er-qing

    2005-01-01

    Objective: To characterize the expression of aquaporin-4 (AQP4), one of the aquaporins (AQPs), in human brain specimens from patients with traumatic brain injury or brain tumors. Methods: Nineteen human brain specimens were obtained from the patients with traumatic brain injury, brain tumors, benign meningioma or early stage hemorrhagic stroke. MRI or CT imaging was used to assess brain edema. Hematoxylin and eosin staining were used to evaluate cell damage. Immunohistochemistry was used to detect the AQP4 expression. Results: AQP4 expression was increased from 15h to at least 8 d after injury. AQP4immunoreactivity was strong around astrocytomas, ganglioglioma and metastatic adenocarcinoma. However, AQP4 immunoreactivity was only found in the centers of astrocytomas and ganglioglioma, but not in metastatic adenocarcinoma derived from lung.Conclusion: AQP4 expression increases in human brains after traumatic brain injury, within brain-derived tumors, and around brain tumors.

  17. Gene expression analysis uncovers novel hedgehog interacting protein (HHIP) effects in human bronchial epithelial cells.

    Science.gov (United States)

    Zhou, Xiaobo; Qiu, Weiliang; Sathirapongsasuti, J Fah; Cho, Michael H; Mancini, John D; Lao, Taotao; Thibault, Derek M; Litonjua, Augusto A; Bakke, Per S; Gulsvik, Amund; Lomas, David A; Beaty, Terri H; Hersh, Craig P; Anderson, Christopher; Geigenmuller, Ute; Raby, Benjamin A; Rennard, Stephen I; Perrella, Mark A; Choi, Augustine M K; Quackenbush, John; Silverman, Edwin K

    2013-05-01

    Hedgehog interacting protein (HHIP) was implicated in chronic obstructive pulmonary disease (COPD) by genome-wide association studies (GWAS). However, it remains unclear how HHIP contributes to COPD pathogenesis. To identify genes regulated by HHIP, we performed gene expression microarray analysis in a human bronchial epithelial cell line (Beas-2B) stably infected with HHIP shRNAs. HHIP silencing led to differential expression of 296 genes; enrichment for variants nominally associated with COPD was found. Eighteen of the differentially expressed genes were validated by real-time PCR in Beas-2B cells. Seven of 11 validated genes tested in human COPD and control lung tissues demonstrated significant gene expression differences. Functional annotation indicated enrichment for extracellular matrix and cell growth genes. Network modeling demonstrated that the extracellular matrix and cell proliferation genes influenced by HHIP tended to be interconnected. Thus, we identified potential HHIP targets in human bronchial epithelial cells that may contribute to COPD pathogenesis.

  18. Gene expression profiling in the human middle cerebral artery after cerebral ischemia

    DEFF Research Database (Denmark)

    Vikman, P; Edvinsson, L

    2006-01-01

    MCA samples distributing to the ischemic area, 7-10 days post-stroke. The gene expression was examined with real-time polymerase chain reaction (PCR) and microarray, proteins were studied with immunohistochemistry. We investigated genes previously shown to be upregulated in animal models of cerebral...... with microarray and seven genes chosen for further investigation with real-time PCR; ELK3, LY64, Metallothionin IG, POU3F4, Actin alpha2, RhoA and smoothelin. Six of these were regulated the same way when confirming array expression with real-time PCR. Gene expression studies in the human MCA leading......We have investigated the gene expression in human middle cerebral artery (MCA) after ischemia. Ischemic stroke affects the perfusion in the affected area and experimental cerebral ischemia results in upregulation of vasopressor receptors in the MCA leading to the ischemic area. We obtained human...

  19. Ornithine decarboxylase, mitogen-activated protein kinase and matrix metalloproteinase-2 expressions in human colon tumors

    Institute of Scientific and Technical Information of China (English)

    Takahiro Nemoto; Shunichiro Kubota; Hideyuki Ishida; Nobuo Murata; Daijo Hashimoto

    2005-01-01

    AIM: To investigate the expressions of omithine decarboxylase (ODC), MMP-2, and Erk, and their relationship in human colon tumors.METHODS: ODC activity, MMP-2 expression, and mitogenactivated protein (MAP) kinase activity (Erk phosphorylation) were determined in 58 surgically removed human colon tumors and their adjacent normal tissues, using [1-14C]-ornithine as a substrate, ELISA assay, and Western blotting, respectively.RESULTS: ODC activity, MMP-2 expression, and Erk phosphorylation were significantly elevated in colon tumors, compared to those in adjacent normal tissues. A significant correlation was observed between ODC activities and MMP-2 levels.CONCLUSION: This is the first report showing a significant correlation between ODC activities and MMP-2 levels in human colon tumors. As MMP-2 is involved in cancer invasion and metastasis, and colon cancer overexpresses ODC, suppression of ODC expression may be a rational approach to treat colon cancer which overexpresses ODC.

  20. Prokaryotic Expression and Biological Activity Analysis of Human Ar-resten Gene

    Institute of Scientific and Technical Information of China (English)

    SONG Zifang; ZHENG Qichang; LI Wei; XIONG Jun; SHANG Dan; SHU Xiaogang

    2005-01-01

    To express recombinant arresten in Escherichia coli (E. Coli) and investigate its biological activity, prokaryotic expression vector of human arresten gene was constructed by gene engineering. Human arresten gene was amplified from recombinant plasmid pGEMArr by polymerase chain reaction (PCR), and inserted into prokaryotic expression vector pRSET containing T7 promoter. Restriction analysis and DNA sequencing verified that the arresten gene was correctly cloned into the expression vector. The recombinant plasmid pRSETAt was subsequently transformed into E. Coli BL21 (DE3), and the target gene was expressed under induction of IPTG. SDS-PAGE analysis revealed that the recombinant protein with a molecular weight of 29 kD (1 kD=0. 992 1 ku) amounted to 29 % of the total bacterial proteins. After purification and renaturation, the recombinant protein could significantly suppress the proliferation of human umbilical vein endothelial cells (HUVECs). These results suggested that the expression of a biologically active form of human arresten in the pRSET expression system laid a foundation for further study on the mechanistic insight into arresten action on angiogenesis and the development of powerful anti-cancer drugs.

  1. [Expressions and significance of NDRG2 and Bcl-2 in human gastric cancer tissues].

    Science.gov (United States)

    Zhu, Ruixue; Shi, Yongquan; Zhang, Lianfeng

    2015-04-01

    To analyze the expressions of N-myc downstream regulated gene 2 (NDRG2) and B cell lymphoma/leukemia-2 (Bcl-2) in human gastric cancer in an attempt to explore their correlation and clinical significance. Immunohistochemical staining was used to detect the expression of NDRG2 and Bcl-2 in human gastric cancer, para-carcinoma tissues and normal tissues. The correlation between their expressions and clinicopathologic data were analyzed using statistical software in gastric cancer tissues. The tissue microarray consisting of 64 gastric cancer and 10 normal gastric tissues showed NDRG2 expression in gastric cancer tissues was significantly lower than that in normal tissues, whereas Bcl-2 expression in gastric cancer tissues was significantly higher than that in normal tissues. It was also indicated that NDRG2 was negatively correlated with Bcl-2 in gastric cancer tissues. NDRG2 and Bcl-2 were further analyzed in 206 gastric cancer and paired para-carcinoma tissues. It was displayed that the expression levels of NDRG2 and Bcl-2 in human gastric cancer were not associated with age and sex, but significantly associated with tumor differentiation, clinical stage and lymph node metastasis. There is a negative correlation between NDRG2 and Bcl-2 expressions in human gastric cancer, suggesting they might be synergistically involved in the development of gastric cancer.

  2. The roadmap of WT1 protein expression in the human fetal heart.

    Science.gov (United States)

    Duim, Sjoerd N; Smits, Anke M; Kruithof, Boudewijn P T; Goumans, Marie-José

    2016-01-01

    The transcription factor Wilms' Tumor-1 (WT1) is essential for cardiac development. Deletion of Wt1 in mice results in disturbed epicardial and myocardial formation and lack of cardiac vasculature, causing embryonic lethality. Little is known about the role of WT1 in the human fetal heart. Therefore, as a first step, we analyzed the expression pattern of WT1 protein during human cardiac development from week 4 till week 20. WT1 expression was apparent in epicardial, endothelial and endocardial cells in a spatiotemporal manner. The expression of WT1 follows a pattern starting at the epicardium and extending towards the lumen of the heart, with differences in timing and expression levels between the atria and ventricles. The expression of WT1 in cardiac arterial endothelial cells reduces in time, whereas WT1 expression in the endothelial cells of cardiac veins and capillaries remains present at all stages studied. This study provides for the first time a detailed description of the expression of WT1 protein during human cardiac development, which indicates an important role for WT1 also in human cardiogenesis. Copyright © 2015 Elsevier Ltd. All rights reserved.

  3. Cloning, Sequencing, and the Expression of the Elusive Sarcomeric TPM4α Isoform in Humans

    Directory of Open Access Journals (Sweden)

    Dipak K. Dube

    2016-01-01

    Full Text Available In mammals, tropomyosin is encoded by four known TPM genes (TPM1, TPM2, TPM3, and TPM4 each of which can generate a number of TPM isoforms via alternative splicing and/or using alternate promoters. In humans, the sarcomeric isoform(s of each of the TPM genes, except for the TPM4, have been known for a long time. Recently, on the basis of computational analyses of the human genome sequence, the predicted sequence of TPM4α has been posted in GenBank. We designed primer-pairs for RT-PCR and showed the expression of the transcripts of TPM4α and a novel isoform TPM4δ in human heart and skeletal muscle. qRT-PCR shows that the relative expression of TPM4α and TPM4δ is higher in human cardiac muscle. Western blot analyses using CH1 monoclonal antibodies show the absence of the expression of TPM4δ protein (~28 kDa in human heart muscle. 2D western blot analyses with the same antibody show the expression of at least nine distinct tropomyosin molecules with a mass ~32 kD and above in adult heart. By Mass spectrometry, we determined the amino acid sequences of the extracted proteins from these spots. Spot “G” reveals the putative expression of TPM4α along with TPM1α protein in human adult heart.

  4. Cloning, Sequencing, and the Expression of the Elusive Sarcomeric TPM4α Isoform in Humans

    Science.gov (United States)

    Abbott, Lynn; Alshiekh-Nasany, Ruham; Mitschow, Charles

    2016-01-01

    In mammals, tropomyosin is encoded by four known TPM genes (TPM1, TPM2, TPM3, and TPM4) each of which can generate a number of TPM isoforms via alternative splicing and/or using alternate promoters. In humans, the sarcomeric isoform(s) of each of the TPM genes, except for the TPM4, have been known for a long time. Recently, on the basis of computational analyses of the human genome sequence, the predicted sequence of TPM4α has been posted in GenBank. We designed primer-pairs for RT-PCR and showed the expression of the transcripts of TPM4α and a novel isoform TPM4δ in human heart and skeletal muscle. qRT-PCR shows that the relative expression of TPM4α and TPM4δ is higher in human cardiac muscle. Western blot analyses using CH1 monoclonal antibodies show the absence of the expression of TPM4δ protein (~28 kDa) in human heart muscle. 2D western blot analyses with the same antibody show the expression of at least nine distinct tropomyosin molecules with a mass ~32 kD and above in adult heart. By Mass spectrometry, we determined the amino acid sequences of the extracted proteins from these spots. Spot “G” reveals the putative expression of TPM4α along with TPM1α protein in human adult heart. PMID:27703814

  5. A humanized pattern of aromatase expression is associated with mammary hyperplasia in mice.

    Science.gov (United States)

    Zhao, Hong; Pearson, Elizabeth K; Brooks, David C; Coon, John S; Chen, Dong; Demura, Masashi; Zhang, Ming; Clevenger, Charles V; Xu, Xia; Veenstra, Timothy D; Chatterton, Robert T; DeMayo, Francesco J; Bulun, Serdar E

    2012-06-01

    Aromatase is essential for estrogen production and is the target of aromatase inhibitors, the most effective endocrine treatment for postmenopausal breast cancer. Peripheral tissues in women, including the breast, express aromatase via alternative promoters. Female mice lack the promoters that drive aromatase expression in peripheral tissues; thus, we generated a transgenic humanized aromatase (Arom(hum)) mouse line containing a single copy of the human aromatase gene to study the link between aromatase expression in mammary adipose tissue and breast pathology. Arom(hum) mice expressed human aromatase, driven by the proximal human promoters II and I.3 and the distal promoter I.4, in breast adipose fibroblasts and myoepithelial cells. Estrogen levels in the breast tissue of Arom(hum) mice were higher than in wild-type mice, whereas circulating levels were similar. Arom(hum) mice exhibited accelerated mammary duct elongation at puberty and an increased incidence of lobuloalveolar breast hyperplasia associated with increased signal transducer and activator of transcription-5 phosphorylation at 24 and 64 wk. Hyperplastic epithelial cells showed remarkably increased proliferative activity. Thus, we demonstrated that the human aromatase gene can be expressed via its native promoters in a wide variety of mouse tissues and in a distribution pattern nearly identical to that of humans. Locally increased tissue levels, but not circulating levels, of estrogen appeared to exert hyperplastic effects on the mammary gland. This novel mouse model will be valuable for developing tissue-specific aromatase inhibition strategies.

  6. EXPRESSION AND SWITCHING OF TH 1/TH2 TYPE CYTOKINES GENE IN HUMAN GLIOMAS

    Institute of Scientific and Technical Information of China (English)

    Yong-sheng Hu; Xin-gang Li; Qing-lin Zhang; Dong-hai Wang; Song-feng Gong

    2005-01-01

    Objective To study the expression and switching of Th1/Th2 cytokines gene in hman gliomas and its effects on occurring and developing of human gliomas.Methods Interleukin(IL)-2 and intefferon-γ represent Th1 type cytokines. IL-4, IL-6, IL-10, and IL-13 represent Th2 type cytokines. The gene expressions of Th1/Th2 cytokines in human glioma cells, glioma infiltrating lymphocytes,and glioma cell lines were detected by reverse transcription polymerase chain reaction (RT-PCR). The biological activity of cytokines in the supematant of glioma cell lines was assayed by enzyme-linked immunosorbent assay (ELISA)method.Results The total positive rates of Th1 and Th2 type cytokines gene in human glioma cells were 14.77% and 75%. The total positive rates of Thl and Th2 type cytokines gene in glioma infiltrating lymphocytes were 22.73% and 68.17%. There was obviously predominant expression of Th2 type cytokines in human glioma tissues, glioma infiltrating lymphocytes, and glioma cell lines. There was no unbalanced expression of Th1/Th2 cytokines in normal brain tissues.Conclusion There is a predominant expression of Th2 type cytokines in human glioma cells. The switching of Th1/Th2 cytokines gene may play an important role in the occurring and developing of human gliomas.

  7. Expression of SHH signaling molecules in the developing human primary dentition

    Science.gov (United States)

    2013-01-01

    Background Our current knowledge on tooth development derives primarily from studies in mice. Very little is known about gene expression and function during human odontogenesis. Sonic Hedgehog (SHH) signaling has been demonstrated to play crucial roles in the development of multiple organs in mice, including the tooth. However, if SHH signaling molecules are expressed and function in the developing human embryonic tooth remain unknown. Results We conducted microarray assay to reveal the expression profile of SHH signaling pathway molecules. We then used in situ hybridization to validate and reveal spatial and temporal expression patterns of a number of selected molecules, including SHH, PTC1, SMO, GLI1, GLI2, and GLI3, in the developing human embryonic tooth germs, and compared them with that in mice. We found that all these genes exhibit similar but slightly distinct expression patterns in the human and mouse tooth germ at the cap and bell stages. Conclusions Our results demonstrate the operation of active SHH signaling in the developing human tooth and suggest a conserved function of SHH signaling pathway during human odontogenesis. PMID:23566240

  8. Distribution of miRNA expression across human tissues.

    Science.gov (United States)

    Ludwig, Nicole; Leidinger, Petra; Becker, Kurt; Backes, Christina; Fehlmann, Tobias; Pallasch, Christian; Rheinheimer, Steffi; Meder, Benjamin; Stähler, Cord; Meese, Eckart; Keller, Andreas

    2016-05-05

    We present a human miRNA tissue atlas by determining the abundance of 1997 miRNAs in 61 tissue biopsies of different organs from two individuals collected post-mortem. One thousand three hundred sixty-four miRNAs were discovered in at least one tissue, 143 were present in each tissue. To define the distribution of miRNAs, we utilized a tissue specificity index (TSI). The majority of miRNAs (82.9%) fell in a middle TSI range i.e. were neither specific for single tissues (TSI > 0.85) nor housekeeping miRNAs (TSI tissues. Clustering of miRNA abundances revealed that tissues like several areas of the brain clustered together. Considering -3p and -5p mature forms we observed miR-150 with different tissue specificity. Analysis of additional lung and prostate biopsies indicated that inter-organism variability was significantly lower than inter-organ variability. Tissue-specific differences between the miRNA patterns appeared not to be significantly altered by storage as shown for heart and lung tissue. MiRNAs TSI values of human tissues were significantly (P = 10(-8)) correlated with those of rats; miRNAs that were highly abundant in certain human tissues were likewise abundant in according rat tissues. We implemented a web-based repository enabling scientists to access and browse the data (https://ccb-web.cs.uni-saarland.de/tissueatlas). © The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.

  9. Expression of ATP7B in normal human liver

    Directory of Open Access Journals (Sweden)

    D Fanni

    2009-06-01

    Full Text Available ATP7B is a copper transporting P-type ATPase, also known as Wilson disease protein, which plays a key role in copper distribution inside cells. Recent experimental data in cell culture have shown that ATP7B putatively serves a dual function in hepatocytes: when localized to the Golgi apparatus, it has a biosynthetic role, delivering copper atoms to apoceruloplasmin; when the hepatocytes are under copper stress, ATP7B translocates to the biliary pole to transport excess copper out of the cell and into the bile canaliculus for subsequent excretion from the body via the bile. The above data on ATP7B localization have been mainly obtained in tumor cell systems in vitro. The aim of the present work was to assess the presence and localization of the Wilson disease protein in the human liver. We tested immunoreactivity for ATP7B in 10 human liver biopsies, in which no significant pathological lesion was found using a polyclonal antiserum specific for ATP7B. In the normal liver, immunoreactivity for ATP7B was observed in hepatocytes and in biliary cells. In the hepatocytes, immunoreactivity for ATP7B was observed close to the plasma membrane, both at the sinusoidal and at the biliary pole. In the biliary cells, ATP7B was localized close to the cell membrane, mainly concentrated at the basal pole of the cells. The data suggest that, in human liver, ATP7B is localized to the plasma membrane of both hepatocytes and biliary epithelial cells.

  10. Expression of metastasis-associated protein 3 in human brain glioma related to tumor prognosis.

    Science.gov (United States)

    Shan, Shouqin; Hui, Guangyan; Hou, Fanggao; Shi, Hua; Zhou, Guoqing; Yan, Han; Wang, Lu; Liu, Jinfeng

    2015-10-01

    Glioma represents a disparate group of tumors characterized by high invasion ability, and therefore it is of clinical significance to identify molecular markers and therapeutic targets for better clinical management. Previously, metastasis-associated protein family (MTA) is considered to promote tumor cell invasion and metastasis of human malignancies. Recently, the newly identified MTA3 has been shown to play conflicting roles in human malignancies, while the expression pattern and potential clinical significance of MTA3 in human glioma have not been addressed yet. In the present study, we investigated the protein expression of MTA3 by immunohistochemistry assay and analyzed its association with glioma prognosis in 186 cases of patients. Results showed that MTA3 expression was decreased in glioma compared with that in normal brain (P human glioma and negatively associated with prognosis of patients, suggesting that MTA3 may play a tumor suppressor role in glioma.

  11. Expression of the human era cDNA in E.coli

    Institute of Scientific and Technical Information of China (English)

    WU Yuan-ming; CHEN Su-min; ZHANG Jun-jie; JI Zong-ling; LIU Hui-ping; CHEN Nan-chun

    2001-01-01

    To amplify human era (Hera) gene, then express it in E.coli. Methods: Human era gene, after amplified by PCR and identified by sequencing, was inserted into the expression vector pGEX4T3 in which exogenous gene was controlled by Ptac promoter. The recombinant plasmid pGEX-Hera was transformed into DH5 ( and induced with IPTG chemically. Results: The human era gene was amplified and the sequence was correct. When the bacteria with pGEX-Hera was induced, an anticipated 65 000 protein band appeared on SDS-PAGE gel and amounted to 23% of total bacterial protein. Conclusion: The human era gene has been successfully amplified and efficiently expressed in E.coli.

  12. Expression of survivin, a novel apoptosis inhibitor and cell cycle regulatory protein, in human gliomas

    Institute of Scientific and Technical Information of China (English)

    焦保华; 姚志刚; 耿少梅; 左书浩

    2004-01-01

    @@ Recently, a novel anti-apoptosis gene, named survivin,was identified as a structurally unique member of the inhibitor of apoptosis protein (lAP) family. The gene is located on chromosome 17q25. Survivin is a 16.5 kDa protein that is expressed in vivo in common human cancers, but not in normal adjacent tissue,1 during the G2/M phase of the cell cycle. Survivin expression is turned off during fetal development and not found in nonneoplastic adult human tissue, and it is turned on in most common human cancers. We investigated the expression of survivin in 50 patients with human gliomas, and determined its association with cell apoptosis and cell proliferation, and its impact on tumor progression and prognosis.

  13. Melanopsin-expressing retinal ganglion cells: implications for human diseases

    DEFF Research Database (Denmark)

    La Morgia, Chiara; Ross-Cisneros, Fred N; Hannibal, Jens

    2011-01-01

    interest on these cells, mainly focused on animal models. Only recently, a few studies have started to address the relevance of the mRGC system in humans and related diseases. We recently discovered that mRGCs resist neurodegeneration in two inherited mitochondrial disorders that cause blindness, i.......e. Leber hereditary optic neuropathy and dominant optic atrophy. The mechanism leading to mRGCs sparing in these blinding disorders, characterized by extensive and selective loss of RGCs, is currently unknown and under investigation. Other studies reported on mRGCs in glaucoma, on genetic variation...

  14. Effects of bright light exposure during daytime on peripheral clock gene expression in humans

    Science.gov (United States)

    Sato, Maki; Wakamura, Tomoko; Morita, Takeshi; Okamoto, Akihiko; Akashi, Makoto; Matsui, Takuya; Sato, Motohiko

    2016-12-01

    Light is the strongest synchronizer controlling circadian rhythms. The intensity and duration of light change throughout the year, thereby influencing body weight, food preferences, and melatonin secretion in humans and animals. Although the expression of clock genes has been examined using human samples, it currently remains unknown whether bright light during the daytime affects the expression of these genes in humans. Therefore, we herein investigated the effects of bright light exposure during the daytime on clock gene expression in the hair follicular and root cells of the human scalp. Seven healthy men (20.4 ± 2.2 years old; 172.3 ± 5.8 cm; 64.3 ± 8.5 kg; BMI 21.7 ± 3.1 kg/m2, mean ± SD) participated in this study. Subjects completed 3-day experimental sessions twice in 1 month during which they were exposed to bright and dim light conditions. The mRNA expression of Per1-3, Cry1-2, Rev-erb-α (Nr1d1), Rev-erb-β (Nr1d2), and Dec1 was analyzed using branched DNA probes. No significant changes were observed in the expression of Per1, Per2, Per3, Cry1, Cry2, Rev-erb-α (Nr1d1), or Dec1 following exposure to bright light conditions. However, the expression of Rev-erb-β (Nr1d2) tended to be stronger under bright light than dim light conditions. These results suggest that the bright light stimulus did not influence the expression of clock genes in humans. Long-lasting bright light exposure during the daytime may be required to change the expression of clock genes in humans.

  15. Expression profiles and function of Toll-like receptors in human corneal epithelia

    Institute of Scientific and Technical Information of China (English)

    WU Xin-yi; GAO Jian-lu; REN Mei-yu

    2007-01-01

    Background Toll-like receptors play an important role in the human immune system. This study was conducted to investigate the expression profiles and function of Toll-like receptor (TLR)1-9 in human corneal epithelium.Methods The expression of TLR1-9 mRNA in 20 human donor corneal epithelia samples abraded during photorefractive keratotomy (PRK) and cultivated telomerase-immortalized human corneal epithelial cells (THCEs) was examined by semi-quantitative reverse-transcriptase polymerase chain reaction (RT-PCR) analysis. Human peripheral blood mononuclear cells (PBMCs) were used as positive controls. The expression of the TLR2 and TLR4 proteins was detected by Western analysis. ELISA was used to detect IL-8 secretion from THCEs challenged with ligands for TLR3 and TLR4 with and without antibody blockade.Results The expression of TLR1-9 at the mRNA level was detected in the epithelia of 20 patients and in THCE.Significant differences among individuals were observed. One patient was found to lack of the expression of TLR3, 4, 6 and 8, whereas another did not express TLR5. The expression of TLR2 and TLR4 protein was detected in human corneal epithelial cells. As THCE cells express TLR1-9, cells were challenged with lipopolysaccharides (LPS) and poly I:C to determine whether TLR4 and TLR3 were functional. The results showed that secretion of IL-8 by cells stimulated with LPS and Poly I:C was 7 to 10 fold greater than secretion by unchallenged cells. Blocking TLR4 with an anti-TLR4 antibody significantly inhibited the LPS-induced IL-8 production by THCE (P<0.05).Conclusion Human corneal epithelial cells express multiple TLRs and are able to recognize LPS and poly I:C. Different expression profiles among individuals suggest that differences in the susceptibilities and sensitivities to bacterial and viral infection in human populations relate to different patterns of TLR expression.

  16. WISP-1 positively regulates angiogenesis by controlling VEGF-A expression in human osteosarcoma.

    Science.gov (United States)

    Tsai, Hsiao-Chi; Tzeng, Huey-En; Huang, Chun-Yin; Huang, Yuan-Li; Tsai, Chun-Hao; Wang, Shih-Wei; Wang, Po-Chuan; Chang, An-Chen; Fong, Yi-Chin; Tang, Chih-Hsin

    2017-04-13

    In recent years, much research has focused on the role of angiogenesis in osteosarcoma, which occurs predominantly in adolescents and young adults. The vascular endothelial growth factor-A (VEGF-A) pathway is the key regulator of angiogenesis and in osteosarcoma. VEGF-A expression has been recognized as a prognostic marker in angiogenesis. Aberrant WNT1-inducible signaling pathway protein-1 (WISP-1) expression is associated with various cancers. However, the function of WISP-1 in osteosarcoma angiogenesis is poorly understood. We demonstrate a positive correlation between WISP-1 and VEGF-A expression in human osteosarcoma. Moreover, we show that WISP-1 promotes VEGF-A expression in human osteosarcoma cells, subsequently inducing human endothelial progenitor cell (EPC) migration and tube formation. The focal adhesion kinase (FAK), Jun amino-terminal kinase (JNK), and hypoxia-inducible factor (HIF)-1α signaling pathways were activated after WISP-1 stimulation, while FAK, JNK, and HIF-1α inhibitors or small interfering RNA (siRNA) abolished WISP-1-induced VEGF-A expression and angiogenesis. In vitro and in vivo studies revealed down-regulation of microRNA-381 (miR-381) in WISP-1-induced VEGF-A expression and angiogenesis. Our findings reveal that WISP-1 enhances VEGF-A expression and angiogenesis through the FAK/JNK/HIF-1α signaling pathways, as well as via down-regulation of miR-381 expression. WISP-1 may be a promising target in osteosarcoma angiogenesis.

  17. Genomic and expression array profiling of chromosome 20q amplicon in human colon cancer cells

    Directory of Open Access Journals (Sweden)

    Carter Jennifer

    2005-01-01

    Full Text Available Background: Gain of the q arm of chromosome 20 in human colorectal cancer has been associated with poorer survival time and has been reported to increase in frequency from adenomas to metastasis. The increasing frequency of chromosome 20q amplification during colorectal cancer progression and the presence of this amplification in carcinomas of other tissue origin has lead us to hypothesize that 20q11-13 harbors one or more genes which, when over expressed promote tumor invasion and metastasis. Aims: Generate genomic and expression profiles of the 20q amplicon in human cancer cell lines in order to identify genes with increased copy number and expression. Materials and Methods: Utilizing genomic sequencing clones and amplification mapping data from our lab and other previous studies, BAC/ PAC tiling paths spanning the 20q amplicon and genomic microarrays were generated. Array-CGH on the custom array with human cancer cell line DNAs was performed to generate genomic profiles of the amplicon. Expression array analysis with RNA from these cell lines using commercial oligo microarrays generated expression profiles of the amplicon. The data were then combined in order to identify genes with increased copy number and expression. Results: Over expressed genes in regions of increased copy number were identified and a list of potential novel genetic tumor markers was assembled based on biological functions of these genes Conclusions: Performing high-resolution genomic microarray profiling in conjunction with expression analysis is an effective approach to identify potential tumor markers.

  18. RAGE Expression in Human T Cells: A Link between Environmental Factors and Adaptive Immune Responses

    Science.gov (United States)

    Akirav, Eitan M.; Preston-Hurlburt, Paula; Garyu, Justin; Henegariu, Octavian; Clynes, Raphael; Schmidt, Ann Marie; Herold, Kevan C.

    2012-01-01

    The Receptor for Advanced Glycation Endproducts (RAGE) is a scavenger ligand that binds glycated endproducts as well as molecules released during cell death such as S100b and HMGB1. RAGE is expressed on antigen presenting cells where it may participate in activation of innate immune responses but its role in adaptive human immune responses has not been described. We have found that RAGE is expressed intracellularly in human T cells following TCR activation but constitutively on T cells from patients with diabetes. The levels of RAGE on T cells from patients with diabetes are not related to the level of glucose control. It co-localizes to the endosomes. Its expression increases in activated T cells from healthy control subjects but bystander cells also express RAGE after stimulation of the antigen specific T cells. RAGE ligands enhance RAGE expression. In patients with T1D, the level of RAGE expression decreases with T cell activation. RAGE+ T cells express higher levels of IL-17A, CD107a, and IL-5 than RAGE− cells from the same individual with T1D. Our studies have identified the expression of RAGE on adaptive immune cells and a role for this receptor and its ligands in modulating human immune responses. PMID:22509345

  19. Discoidin domain receptor 1: isoform expression and potential functions in cirrhotic human liver.

    Science.gov (United States)

    Song, Sunmi; Shackel, Nicholas A; Wang, Xin M; Ajami, Katerina; McCaughan, Geoffrey W; Gorrell, Mark D

    2011-03-01

    Discoidin domain receptor 1 (DDR1) is a receptor tyrosine kinase that binds and is activated by collagens. Transcriptional profiling of cirrhosis in human liver using a DNA array and quantitative PCR detected elevated mRNA expression of DDR1 compared with that in nondiseased liver. The present study characterized DDR1 expression in cirrhotic and nondiseased human liver and examined the cellular effects of DDR1 expression. mRNA expression of all five isoforms of DDR1 was detected in human liver, whereas DDR1a demonstrated differential expression in liver with hepatitis C virus and primary biliary cirrhosis compared with nondiseased liver. In addition, immunoblot analysis detected shed fragments of DDR1 more readily in cirrhotic liver than in nondiseased liver. Inasmuch as DDR1 is subject to protease-mediated cleavage after prolonged interaction with collagen, this differential expression may indicate more intense activation of DDR1 protein in cirrhotic compared with nondiseased liver. In situ hybridization and immunofluorescence localized intense DDR1 mRNA and protein expression to epithelial cells including hepatocytes at the portal-parenchymal interface and the luminal aspect of the biliary epithelium. Overexpression of DDR1a altered hepatocyte behavior including increased adhesion and less migration on extracelular matrix substrates. DDR1a regulated extracellular expression of matrix metalloproteinases 1 and 2. These data elucidate DDR1 function pertinent to cirrhosis and indicate the importance of epithelial cell-collagen interactions in chronic liver injury.

  20. Expression and functions of long noncoding RNAs during human T helper cell differentiation.

    Science.gov (United States)

    Spurlock, Charles F; Tossberg, John T; Guo, Yan; Collier, Sarah P; Crooke, Philip S; Aune, Thomas M

    2015-04-23

    Long noncoding RNAs (lncRNAs) regulate an array of biological processes in cells and organ systems. Less is known about their expression and function in lymphocyte lineages. Here we have identified >2000 lncRNAs expressed in human T-cell cultures and those that display a TH lineage-specific pattern of expression and are intragenic or adjacent to TH lineage-specific genes encoding proteins with immunologic functions. One lncRNA cluster selectively expressed by the effector TH2 lineage consists of four alternatively spliced transcripts that regulate the expression of TH2 cytokines, IL-4, IL-5 and IL-13. Genes encoding this lncRNA cluster in humans overlap the RAD50 gene and thus are contiguous with the previously described TH2 locus control region (LCR) in the mouse. Given its genomic synteny with the TH2-LCR, we refer to this lncRNA cluster as TH2-LCR lncRNA.

  1. Expression of insulin signalling components in the sensory epithelium of the human saccule

    DEFF Research Database (Denmark)

    Degerman, Eva; Rauch, Uwe; Lindberg, Sven

    2013-01-01

    Several studies have demonstrated a link between diabetes and the dysfunction of the inner ear. Few studies, however, have reported the signalling mechanisms involved in metabolic control in human inner ear cells. Knowledge of the expression and role of the insulin receptor and downstream...... signalling components in the inner ear is sparce. Our immunohistochemistry approach has shown that the insulin receptor, insulin receptor substrate 1 (IRS1), protein kinase B (PKB) and insulin-sensitive glucose transporter (GLUT4) are expressed in the sensory epithelium of the human saccule, which also...... exhibits expression of a calcium-sensitive cAMP/cGMP phosphodiesterase 1C (PDE1C) and the vasopressin type 2 receptor. IRS1 and PDE1C are selectively expressed in sensory epithelial hair cells, whereas the other components are expressed in sensory epithelial supporting cells or in both cell types...

  2. In vitro expression and analysis of the 826 human G protein-coupled receptors

    Directory of Open Access Journals (Sweden)

    Xuechen Lv

    2016-04-01

    Full Text Available ABSTRACT G protein-coupled receptors (GPCRs are involved in all human physiological systems where they are responsible for transducing extracellular signals into cells. GPCRs signal in response to a diverse array of stimuli including light, hormones, and lipids, where these signals affect downstream cascades to impact both health and disease states. Yet, despite their importance as therapeutic targets, detailed molecular structures of only 30 GPCRs have been determined to date. A key challenge to their structure determination is adequate protein expression. Here we report the quantification of protein expression in an insect cell expression system for all 826 human GPCRs using two different fusion constructs. Expression characteristics are analyzed in aggregate and among each of the five distinct subfamilies. These data can be used to identify trends related to GPCR expression between different fusion constructs and between different GPCR families, and to prioritize lead candidates for future structure determination feasibility.

  3. Mosquito Passage Dramatically Changes var Gene Expression in Controlled Human Plasmodium falciparum Infections.

    Science.gov (United States)

    Bachmann, Anna; Petter, Michaela; Krumkamp, Ralf; Esen, Meral; Held, Jana; Scholz, Judith A M; Li, Tao; Sim, B Kim Lee; Hoffman, Stephen L; Kremsner, Peter G; Mordmüller, Benjamin; Duffy, Michael F; Tannich, Egbert

    2016-04-01

    Virulence of the most deadly malaria parasite Plasmodium falciparum is linked to the variant surface antigen PfEMP1, which is encoded by about 60 var genes per parasite genome. Although the expression of particular variants has been associated with different clinical outcomes, little is known about var gene expression at the onset of infection. By analyzing controlled human malaria infections via quantitative real-time PCR, we show that parasite populations from 18 volunteers expressed virtually identical transcript patterns that were dominated by the subtelomeric var gene group B and, to a lesser extent, group A. Furthermore, major changes in composition and frequency of var gene transcripts were detected between the parental parasite culture that was used to infect mosquitoes and Plasmodia recovered from infected volunteers, suggesting that P. falciparum resets its var gene expression during mosquito passage and starts with the broad expression of a specific subset of var genes when entering the human blood phase.

  4. Tissue-specific mRNA expression profiles of human solute carrier 35 transporters.

    Science.gov (United States)

    Nishimura, Masuhiro; Suzuki, Satoshi; Satoh, Tetsuo; Naito, Shinsaku

    2009-01-01

    Pairs of forward and reverse primers and TaqMan probes specific to each of 23 human solute carrier 35 (SLC35) transporters were prepared. The mRNA expression level of each target transporter was analyzed in total RNA from single and pooled specimens of adult human tissues (adipose tissue, adrenal gland, bladder, bone marrow, brain, cerebellum, colon, heart, kidney, liver, lung, mammary gland, ovary, pancreas, peripheral leukocytes, placenta, prostate, retina, salivary gland, skeletal muscle, small intestine, smooth muscle, spinal cord, spleen, stomach, testis, thymus, thyroid gland, tonsil, trachea, and uterus), from pooled specimens of fetal human tissues (brain, heart, kidney, liver, spleen, and thymus), and from three human cell lines (HeLa cell line ATCC#: CCL-2, human cell line Hep G2, and human breast carcinoma cell line MDA-435) by real-time reverse transcription PCR using an Applied Biosystems 7500 Fast Real-Time PCR System. The mRNA expression of SLC35As, SLC35Bs, SLC35Cs, SLC35D1, SLC35D2, SLC35Es, and SLC35F5 was found to be ubiquitous in both adult and fetal tissues. SLC35D3 mRNA was expressed at the highest levels in the adult retina. SLC35F1 mRNA was expressed at high levels in the adult and fetal brain. SLC35F2 mRNA was expressed at the highest levels in the adult salivary gland. Both SLC35F3 and SLC35F4 mRNAs were expressed at the highest levels in the adult cerebellum. Further, individual differences in the mRNA expression levels of human SLC35 transporters in the liver were also evaluated. Our newly determined expression profiles were used to study the gene expression in 31 adult human tissues, 6 fetal human tissues, and 3 cell lines, and tissues with high transcriptional activity for human SLC35 transporters were identified. These results are expected to be valuable for research concerning the clinical diagnosis of disease.

  5. MOLECULAR CLONING AND HETEROLOGOUS EXPRESSION OF HUMAN INTERFERON ALPHA2b GENE

    Directory of Open Access Journals (Sweden)

    I. Made Artika

    2013-01-01

    Full Text Available Human alpha Interferons (hIFNα have been shown to have antiviral, antiproliferative and immunomodulatory activities. The human interferon alpha2b (hIFNα2b, is one of the human interferon alpha2 sub variants, naturally synthesized as a polypeptide of 188 amino acid residues, the first 23 residues of which represents a signal peptide. In the present study, the hIFNα2b gene was expressed after being fused with Glutathione S-Transferase (GST gene. The hIFNα2b gene was amplified from human genomic DNA by using a pair of specific primers, cloned into an Escherichia coli expression vector and expressed in E. coli cells under the direction of the tac promoter. The expressed protein was purified using a one-step affinity chromatography column containing immobilized gluthatione-bound resin. The purified protein was shown to react specifically with anti-human-interferon-alpha antibody, confirming that the protein was the human interferon alpha molecule. This strategy has the potential to be used as an alternative mean for production of pure human interferon α proteins for therapeutic purposes and for further studies on their molecular characterization and mechanism of action.

  6. Evaluation of reference genes for gene expression studies in human brown adipose tissue.

    Science.gov (United States)

    Taube, Magdalena; Andersson-Assarsson, Johanna C; Lindberg, Kristin; Pereira, Maria J; Gäbel, Markus; Svensson, Maria K; Eriksson, Jan W; Svensson, Per-Arne

    2015-01-01

    Human brown adipose tissue (BAT) has during the last 5 year been subjected to an increasing research interest, due to its putative function as a target for future obesity treatments. The most commonly used method for molecular studies of human BAT is the quantitative polymerase chain reaction (qPCR). This method requires normalization to a reference gene (genes with uniform expression under different experimental conditions, e.g. similar expression levels between human BAT and WAT), but so far no evaluation of reference genes for human BAT has been performed. Two different microarray datasets with samples containing human BAT were used to search for genes with low variability in expression levels. Seven genes (FAM96B, GNB1, GNB2, HUWE1, PSMB2, RING1 and TPT1) identified by microarray analysis, and 8 commonly used reference genes (18S, B2M, GAPDH, LRP10, PPIA, RPLP0, UBC, and YWHAZ) were selected and further analyzed by quantitative PCR in both BAT containing perirenal adipose tissue and subcutaneous adipose tissue. Results were analyzed using 2 different algorithms (Normfinder and geNorm). Most of the commonly used reference genes displayed acceptably low variability (geNorm M-values genes identified by microarray displayed an even lower variability (M-values genes for qPCR analysis of human BAT and we recommend that they are included in future gene expression studies of human BAT.

  7. Expression of adrenomedullin in rats after spinal cord injury and intervention effect of recombinant human erythropoietin

    Science.gov (United States)

    Zhao, Liang; Jing, Yu; Qu, Lin; Meng, Xiangwei; Cao, Yang; Tan, Huibing

    2016-01-01

    The expression of adrenomedullin (ADM) in injured tissue of rat spinal cord was observed and the effect of recombinant human erythropoietin was analyzed. A total of 45 Sprague-Dawley rats were selected and divided into 3 equal groups including, a sham-operation group in which rats received an excision of vertebral plate; a spinal cord injury model group and a recombinant human erythropoietin group in which rats with spinal cord injury received a caudal vein injection of 300 units recombinant human erythropoietin after injury. Hematoxylin and eosin staining was performed to observe the spinal cord injury conditions. Immunohistochemical staining was performed to observe the expression of ADM. Pathologic changes in the group of recombinant human erythropoietin at various times were significantly less severe than those in the group of spinal cord injury model. The expression of ADM was increased particularly in the group of recombinant human erythropoietin (P<0.01). The improved Tarlov scores of the group of spinal cord injury model and the group of recombinant human erythropoietin were lower than those of the sham-operation group at 3, 6 and 9 days (P<0.01). Thus, the recombinant human erythropoietin is capable of alleviating the secondary injury of spinal cord. One of the mechanisms may be achieved by promoting the increase of ADM expression. PMID:28101163

  8. Expression of the cytochrome P450 epoxygenase CYP2J2 in human monocytic leukocytes.

    Science.gov (United States)

    Nakayama, Kaeko; Nitto, Takeaki; Inoue, Teruo; Node, Koichi

    2008-08-29

    CYP2J2 is one of the cytochrome P450 epoxygenases involved in the metabolism of arachidonic acid. CYP2J2 has been identified in several tissues, especially cardiovascular tissues. CYP2J2 has cardiovascular effects, as epoxyeicosatrienoic acid, one of its metabolites, has anti-inflammatory and vasodilative activities. We investigated the expression of CYP2J2 in human leukocytes using reverse transcription-polymerase chain reaction, immunoblotting and immunostaining. Human monocytic cells, but not human neutrophils, exhibited constitutive expression of CYP2J2. Furthermore, the expression of CYP2J2 mRNA increased when the human monocytic cell line THP-1 cells and human monocytes were stimulated with phorbol 12-myristate 13-acetate and macrophage-colony stimulating factor in combination with granulocyte/macrophage-colony stimulating factor, respectively. These results suggest that expression of CYP2J2 was up-regulated when human monocytes differentiated into macrophages and that human monocytic cells and macrophages have a pathway to metabolize arachidonic acid using CYP epoxygenases.

  9. Transgenic chickens expressing human urokinase-type plasminogen activator.

    Science.gov (United States)

    Lee, Sung Ho; Gupta, Mukesh Kumar; Ho, Young Tae; Kim, Teoan; Lee, Hoon Taek

    2013-09-01

    Urokinase-type plasminogen activator is a serine protease that is clinically used in humans for the treatment of thrombolytic disorders and vascular diseases such as acute ischemic stroke and acute peripheral arterial occlusion. This study explored the feasibility of using chickens as a bioreactor for producing human urokinase-type plasminogen activator (huPA). Recombinant huPA gene, under the control of a ubiquitous Rous sarcoma virus promoter, was injected into the subgerminal cavity of freshly laid chicken eggs at stage X using the replication-defective Moloney murine leukemia virus (MoMLV)-based retrovirus vectors encapsidated with VSV-G (vesicular stomatitis virus G) glycoprotein. A total of 38 chicks, out of 573 virus-injected eggs, hatched and contained the huPA gene in their various body parts. The mRNA transcript of the huPA gene was present in various organs, including blood and egg, and was germ-line transmitted to the next generation. The level of active huPA protein was 16-fold higher in the blood of the transgenic chicken than in the nontransgenic chicken (P pharming of the huPA drug but also be useful for studying huPA-induced bleeding and other disorders.

  10. mTOR expression in human testicular seminoma.

    Science.gov (United States)

    Yaba, A; Bozkurt, E R; Demir, N

    2016-08-01

    The mammalian target of rapamycin (TOR) has been implicated in the control of different stressors, growth factors, nutrients and hormones, participating in the control of key cellular functions. Controlling this many pathways poses mTOR signalling as a potential new target in new treatment strategies for multiple cancer types. mTOR components could potentially mislocated in tumour cells, which could lead to activation of signalling pathway that should not be active. Therefore, we aimed to show localisation of mTOR signal proteins in testicular seminoma. Tumoural testicular tissues were obtained from 10 patients with unilateral classic seminoma undergoing to therapeutic orchidectomy and compared with control human testicular tissues. Upon immunohistochemical evaluation, we detected mTOR and p-mTOR (serine 2448), P70S6K, p-P70S6K, PKCalpha and p-PKCalpha, CD36 and MAPLC3 proteins in the cytoplasm of Sertoli cells in the seminiferous tubules. We also showed cytoplasmic perinuclear staining in seminoma cells. This study demonstrated the interaction of mTOR signalling pathway and testicular seminoma by showing intense cytoplasmic mTOR pathway proteins immunoreactivity in the seminoma, for the first time in humans. Therefore, we suggested that mTOR signalling components could create new clinical targets for treatment of testicular seminoma patients and male infertility in the future. © 2015 Blackwell Verlag GmbH.

  11. Purinergic receptors expressed in human skeletal muscle fibres

    DEFF Research Database (Denmark)

    Bornø, A; Ploug, Thorkil; Bune, L T

    2012-01-01

    Purinergic receptors are present in most tissues and thought to be involved in various signalling pathways, including neural signalling, cell metabolism and local regulation of the microcirculation in skeletal muscles. The present study aims to determine the distribution and intracellular content...... of purinergic receptors in skeletal muscle fibres in patients with type 2 diabetes and age-matched controls. Muscle biopsies from vastus lateralis were obtained from six type 2 diabetic patients and seven age-matched controls. Purinergic receptors were analysed using light and confocal microscopy...... in immunolabelled transverse sections of muscle biopsies. The receptors P2Y(4), P2Y(11) and likely P2X(1) were present intracellularly or in the plasma membrane of muscle fibres and were thus selected for further detailed morphological analysis. P2X(1) receptors were expressed in intracellular vesicles...

  12. Over-expression of HOX-8, the human homologue of the mouse Hox-8 homeobox gene, in human tumors.

    Science.gov (United States)

    Suzuki, M; Tanaka, M; Iwase, T; Naito, Y; Sugimura, H; Kino, I

    1993-07-15

    A human ovarian yolk sac tumor cDNA library was screened for homeobox genes with an oligonucleotide probe under low stringent condition. Three homeobox genes were isolated, two of which were identified as HHO.c1 and HB24. The third was highly homologous with the mouse Hox-8 gene and was designated as HOX-8. Studies on RNAs from 25 human tumor tissues and cell lines showed that the profile of HOX-8 expression was different from those of HHO.c1 and HB24. The expression of HOX-8 was not detected in hematopoietic tumor cells, in which HHO.c1 and HB24 were highly expressed. HOX-8 was expressed at higher levels in a variety of tumors of epithelial origin than in their corresponding normal tissues more frequently than HHO.c1 and HB24. All three homeobox genes were highly expressed in a yolk sac tumor, an immature tumor of gonadal origin. These results suggest that HOX-8 plays a more important role in human tumors of epithelial origin than those of hematopoietic origin.

  13. Interferon-alpha induces transient suppressors of cytokine signalling expression in human T cells

    DEFF Research Database (Denmark)

    Brender, C; Nielsen, M; Röpke, C;

    2001-01-01

    The suppressors of cytokine signalling (SOCS) proteins comprise a newly identified family of negative feedback regulators of cytokine signalling. SOCS expression is differentially induced upon cytokine stimulation in different cell types. Here we show that interferon-alpha (IFNalpha) is a potent...... induction neither CIS, SOCS-1, nor SOCS-2 expression levels declined after 6 h. In conclusion, we provide the first evidence that IFNalpha induces SOCS expression in human T cells. Moreover, we show that IFNalpha and IL-2 induce distinct patterns of expression kinetics, suggesting that dynamic changes...

  14. Differential transcriptome expression in human nucleus accumbens as a function of loneliness.

    Science.gov (United States)

    Canli, T; Wen, R; Wang, X; Mikhailik, A; Yu, L; Fleischman, D; Wilson, R S; Bennett, D A

    2016-11-01

    Loneliness is associated with impaired mental and physical health. Studies of lonely individuals reported differential expression of inflammatory genes in peripheral leukocytes and diminished activation in brain reward regions such as nucleus accumbens, but could not address gene expression in the human brain. Here, we examined genome-wide RNA expression in post-mortem nucleus accumbens from donors (N=26) with known loneliness measures. Loneliness was associated with 1710 differentially expressed transcripts and genes from 1599 genes (DEGs; false discovery rate PMolecular Psychiatry advance online publication, 1 November 2016; doi:10.1038/mp.2016.186.

  15. Relationship between pancreatic vesicular monoamine transporter 2 (VMAT2) and insulin expression in human pancreas

    OpenAIRE

    Saisho, Yoshifumi; Harris, Paul E.; Butler, Alexandra E.; Galasso, Ryan; GURLO, TATYANA; Rizza, Robert A.; Butler, Peter C.

    2008-01-01

    Vesicular monoamine transporter 2 (VMAT2) is expressed in pancreatic beta cells and has recently been proposed as a target for measurement of beta cell mass in vivo. We questioned, (1) What proportion of beta cells express VMAT2? (2) Is VMAT2 expressed by other pancreatic endocrine or non-endocrine cells? (3) Is the relationship between VMAT2 and insulin expression disturbed in type 1 (T1DM) or type 2 diabetes (T2DM)? Human pancreas (7 non-diabetics, 5 T2DM, 10 T1DM) was immunostained for ins...

  16. Butyrate stimulates IL-32α expression in human intestinal epithelial cell lines

    Institute of Scientific and Technical Information of China (English)

    Ayako; Kobori; Shigeki; Bamba; Hirotsugu; Imaeda; Hiromitsu; Ban; Tomoyuki; Tsujikawa; Yasuharu; Saito; Yoshihide; Fujiyama; Akira; Andoh

    2010-01-01

    AIM: To investigate the effects of butyrate on interleukin (IL)-32α expression in epithelial cell lines. METHODS: The human intestinal epithelial cell lines HT-29, SW480, and T84 were used. Intracellular IL- 32α was determined by Western blotting analyses. IL- 32α mRNA expression was analyzed by real-time poly-merase chain reaction. RESULTS: Acetate and propionate had no effects on IL-32α mRNA expression. Butyrate significantly enhanced IL-32α expression in all cell lines. Butyrate also up-regulated IL-1β-i...

  17. Construction and Expression of Eukaryotic Expression Vector and Plasmid Expressing siRNA of Human Protection of Telomeres 1

    Institute of Scientific and Technical Information of China (English)

    2005-01-01

    1 IntroductionThe POT1 (protection of telomeres 1) protein binds the single-stranded overhang at the ends of chromosomes in diverse eukaryocytes. It is essential for chromosome end-protection in the fission yeast Schizosaccharomyces pombe, and it is involved in regulation of telomere length in human cells. Human POT1 had been identified in 2001 year. Its amino terminal is highly conservative in eukaryocytes. Since Pot1 can bind internal loops and directly adjacent DNA-binding sites, it is likely to fully co...

  18. Expression of fibroblast growth factor (FGF)-8 isoforms and FGF receptors in human ovarian tumors.

    Science.gov (United States)

    Valve, E; Martikainen, P; Seppänen, J; Oksjoki, S; Hinkka, S; Anttila, L; Grenman, S; Klemi, P; Härkönen, P

    2000-12-01

    FGF-8 is a mitogenic growth factor, which is widely expressed during embryonic development but only at a very low level in adult tissues. Alternative splicing of the human FGF-8 gene potentially allows coding for 4 protein isoforms (a, b, e, f), which differ in their transforming capacity. The FGF-8 isoforms preferentially activate the receptors FGFR1IIIc, FGFR2IIIc, FGFR3IIIc and FGFR4. FGF-8 is over-expressed in human breast and prostate cancers. Expression has also been found in RT-PCR studies of human ovarian and testicular cancers. The present study was undertaken to examine which FGF-8 isoforms are expressed in ovarian cancer and whether FGF-8 receptors are also expressed. Specimens from 5 normal human ovaries and 51 ovarian tumors (1 benign tumor, 8 borderline malignancies, 42 malignant tumors of different histopathological types) were studied by RT-PCR and immunohistochemistry. FGF-8 isoform b was expressed in all ovarian tumors and in all 7 ovarian-cancer cell lines studied. Isoform a was co-expressed in 9 malignant ovarian tumors. FGF-8 mRNA was not detected by RT-PCR of 3 normal ovary samples. Immunohistochemical staining localized FGF-8 protein to cancer cells. In general, the increased intensity of FGF-8 staining was associated with loss of differentiation within the tumors (Bowker's test, p = 0.37). FGF-8 staining of surface epithelium observed on 2 normal ovaries was very faint. RT-PCR showed that FGFR1IIIc, FGFR2IIIc and FGFR4 were the FGF-8 receptors expressed in normal ovaries and in ovarian tumors. FGF-8 receptor immunoreactivity was preferentially found in normal ovary surface epithelium and tumor cells but also in some stromal cells. Collectively, our results show that ovarian cancers of a wide variety of histological types expressing receptors for FGF-8 have acquired the capacity of expressing FGF-8. This suggests that FGF-8 has an important role in ovarian tumorigenesis.

  19. Expression of nitric oxide synthase in human gastric carcinoma and its relation to p53, PCNA

    Institute of Scientific and Technical Information of China (English)

    Yong-Zhong Wang; You-Qing Cao; Jian-Nong Wu; Miao Chen; Xiao-Ying Cha

    2005-01-01

    AIM: To investigate the expression of NOS in gastric carcinoma, and to explore the relationship between the expression of nitric oxide synthases (NOS) and p53, PCNA,pathological features and clinical staging of gastric cancer.METHODS: The activity of NOS protein was investigated in 85 samples of human gastric carcinoma and 25 samples of normal gastric mucosal tissue by biochemical assay. We then examined the expression of NOS, p53, PCNA in 85 samples of human gastric cancer was examined by immunohistochemistry, and NOS mRNA expression in 85 gastric cancer tissue specimens by in situ hybridization.RESULTS: Biochemical assay showed that the activity of NOS was significantly higher in gastric carcinoma than in normal gastric mucosal tissues (t = 0.4161, P<0.01).Immunohistochemistry revealed that endothelial nitric oxide synthase (eNOS) expressed in all samples of normal gastric mucosa, but only 6 cases of 85 gastric cancer specimens showed weak positive immunohistochemical reactions to eNOS (20%). Inducible nitric oxide synthase (iNOS) was expressed strongly in human gastric carcinoma (81.2%). In situ hybridization analysis showed that iNOS mRNA expression was significantly stronger than eNOS mRNA expression in gastric cancer tissue (x2 = 10.23, P<0.01). The expression of iNOS in gastric cancer was associated with differentiation, clinical stages or lymph node metastases (r= 0.3426, P<0.05). However,iNOS expression did not correlate with histological classifications and morphological types. The expression of iNOS was significantly correlated with p53 or PCNA expression (r = 0.3612, P<0.05). The expression of neuronal nitric oxide synthase (nNOS) was not examined by immunohistochemistry and in situ hybridization in gastric cancer specimens and normal gastric mucosa.CONCLUSION: In human gastric cancer, there is an enhanced expression of iNOS, but not of eNOS. NOS promotes the proliferation of tumor cells and plays an important role in gastric cancer spread

  20. Myosin heavy chain isoform expression in human extraocular muscles: longitudinal variation and patterns of expression in global and orbital layers.

    Science.gov (United States)

    Park, Kyung-Ah; Lim, Jeonghee; Sohn, Seongsoo; Oh, Sei Yeul

    2012-05-01

    We investigated the distribution of myosin heavy chain (MyHC) isoforms along the length of the global and orbital layers of human extraocular muscles (EOMs). Whole muscle tissue extracts of human EOMs were cross-sectioned consecutively and separated into orbital and global layers. The extracts from these layers were subjected to electrophoretic analysis, followed by quantification with scanning densitometry. MyHC isoforms displayed different distributions along the lengths of EOMs. In the orbital and global layers of all EOMs except for the superior oblique muscle, MyHCeom was enriched in the central regions. MyHCIIa and MyHCI were most abundant in the proximal and distal ends. A variation in MyHC isoform expression was apparent along the lengths of human EOMs. These results provide a basis for understanding the molecular mechanisms underlying the functional diversity of EOMs. Copyright © 2012 Wiley Periodicals, Inc.

  1. Expression of secreted human single-chain fragment variable antibody against human amyloid beta peptide in Pichia pastoris

    Institute of Scientific and Technical Information of China (English)

    Jiong Cai; Fang Li; Shizhen Wang

    2008-01-01

    BACKGROUND: Studies have shown that monoclonal or polyclonal antibody injections ofamyloid β peptide arc effective in removing amyloid β peptide overload in the brain.OBJECTIVE: Based on successful screening of a human single-chain fragment variable antibody specific to amyloid β peptide, this paper aimed to express recombinant human single-chain variable antibody against amyloid β peptide.DESIGN, TIME AND SETTING: A single sample experiment was performed at the Department of Nuclear Medicine, Peking Union Medical College Hospital, Chinese Academy of Medical Sciences and Peking Union Hospital (Beijing, China) from January to July 2006.MATERIALS: Human single-chain fragment variable antibody gene against amyloid β peptide was screened from a human phage-display antibody library.METHODS: Human single-chain fragment variable antibody gene was mutated to eliminate a BamHI restriction site and cloned into a Teasy plasmid for pT-seFvAβ construction, which was identified by PCR amplification and endonuclease digestion. Plasmid pT-scFvA β was cut by EcoRl and Notl endonucleases, and the antibody gene was cloned into pPIC9K plasmid to construct pPIC9K-scFvA β expression vector, which was confirmed by gene sequencing. Linearized pPICgK-scFvA β was used to transform a Pichia pastoris GS115 cell line, and the recombinant was induced by 0.5 % methanol to express human single-chain fragment variable antibody specific to amyloid β peptide.MAIN OUTCOME MEASURES: Protein electrophoresis was used to identify PCR products, gene sequencing was uscd to verify the pPIC9K-scFvA sequence, and SDS-PAGE was used to detect recombinant expression of human single-chain fragment variable antibody specific to amyloid β peptide in Pichia pastoris.RESULTS: Gene sequencing confirmed pPICgK-scFvA β orientation. Rccomhinants were obtained by lineadzed pPIC9K-scFvA β transformation. After induction with 0.5% methanol, the recombinant yeast cells secreted proteins of 33-ku size

  2. Thiocyanate supplementation decreases atherosclerotic plaque in mice expressing human myeloperoxidase

    DEFF Research Database (Denmark)

    Morgan, P E; Laura, R P; Maki, R A

    2015-01-01

    Elevated levels of the heme enzyme myeloperoxidase (MPO) are associated with adverse cardiovascular outcomes. MPO predominantly catalyzes formation of the oxidants hypochlorous acid (HOCl) from Cl(-), and hypothiocyanous acid (HOSCN) from SCN(-), with these anions acting as competitive substrates....... HOSCN is a less powerful and more specific oxidant than HOCl, and selectively targets thiols; such damage is largely reversible, unlike much HOCl-induced damage. We hypothesized that increased plasma SCN(-), and hence HOSCN formation instead of HOCl, may decrease artery wall damage. This was examined...... using high-fat fed atherosclerosis-prone LDLR(-/-) mice transgenic for human MPO, with and without SCN(-) (10 mM) added to drinking water. Serum samples, collected fortnightly, were analyzed for cholesterol, triglycerides, thiols, MPO, and SCN(-); study-long exposure was calculated by area under...

  3. Clinicopathological significance of p53 and mdm2 protein expression in human pancreatic cancer

    Institute of Scientific and Technical Information of China (English)

    Ming Dong; Gang Ma; Wei Tu; Ke-Jian Guo; Yu-Lin Tian; Yu-Ting Dong

    2005-01-01

    AIM: To study the clinicopathological significance of p53 and mdm2 protein expression in human pancreatic cancer. METHODS: To investigate the expression of p53 and mdm2 in pancreatic cancer by immunohistochemistry, and the relationships between the p53 and mdm2 protein expression and clinicopathological parameters in pancreatic cancer.RESULTS: The positive expression of p53 protein was found in 40 of 59 patients (67.8%) and that of mdm2 protein in 17 of 59 patients (28.8%). No obvious relationships were found between p53 as well as mdm2 expression and sex, tumor site, TNM staging and histological differentiation. p53 expression was increased in patients younger than 65 years old, while mdm2 had no relationship with age. The survival time of the patients with the positive expression of p53 and mdm2 proteins was obviously shorter than the other groups. CONCLUSION: Both p53 and mdm2 presented relatively high expression in human pancreatic cancer. The overexpression of p53 and mdm2 might reflect the malignant proliferation of pancreatic cancer and their co-expression might be helpful to evaluate the prognosis of the patients with pancreatic cancer.

  4. NLRP3 Inflammasome Expression and Signaling in Human Diabetic Wounds and in High Glucose Induced Macrophages

    Directory of Open Access Journals (Sweden)

    Xiaotian Zhang

    2017-01-01

    Full Text Available Introduction. To investigate the contribution and mechanism of NLRP3 inflammasome expression in human wounds in diabetes mellitus and in high glucose induced macrophages. Methods. In the present study, we compared the expression of NLRP3 inflammasome in debridement wound tissue from diabetic and nondiabetic patients. We also examined whether high glucose induces NLRP3 inflammasome expression in cultures THP-1-derived macrophages and the influence on IL-1β expression. Results. The expressions of NLRP3, caspase1, and IL-1β, at both the mRNA and protein level, were significantly higher in wounds of diabetic patients compared with nondiabetic wounds (P<0.05. High glucose induced a significant increase in NLRP3 inflammasome and IL-1β expression in THP-1-derived macrophages. M1 macrophage surface marker with CCR7 was significantly upregulated after high glucose stimulation. SiRNA-mediated silencing of NLRP3 expression downregulates the expression of IL-1β. Conclusion. The higher expression of NLRP3, caspase1, and secretion of IL-1β, signaling, and activation might contribute to the hyperinflammation in the human diabetic wound and in high glucose induced macrophages. It may be a novel target to treat the DM patients with chronic wound.

  5. Gene expression demonstrates an immunological capacity of the human endolymphatic sac

    DEFF Research Database (Denmark)

    Møller, Martin Nue; Kirkeby, Svend; Vikeså, Jonas

    2015-01-01

    OBJECTIVES/HYPOTHESIS: The purpose of the present study is to explore, demonstrate, and describe the expression of genes related to the innate immune system in the human endolymphatic sac. It is hypothesized that the endolymphatic sac has a significant immunological function in the human inner ear....... STUDY DESIGN: DNA microarrays and immunohistochemistry were used for analyses of fresh human endolymphatic-sac tissue samples. METHODS: Twelve tissue samples from the human endolymphatic sac were obtained during translabyrinthine surgery for vestibular schwannoma. Microarray technology was used...... was obtained. Multiple key elements of both the cellular and humoral innate immune system were expressed, including Toll-like receptors 4 and 7, as well as beta-defensin and lactoferrin. CONCLUSIONS: The present data provides the first direct evidence of an immunological capacity of the human endolymphatic sac...

  6. Transcriptional Factor PU.1 Regulates Decidual C1q Expression in Early Pregnancy in Human.

    Science.gov (United States)

    Madhukaran, Shanmuga Priyaa; Kishore, Uday; Jamil, Kaiser; Teo, Boon Heng Dennis; Choolani, Mahesh; Lu, Jinhua

    2015-01-01

    C1q is the first recognition subcomponent of the complement classical pathway, which in addition to being synthesized in the liver, is also expressed by macrophages and dendritic cells (DCs). Trophoblast invasion during early placentation results in accumulation of debris that triggers the complement system. Hence, both early and late components of the classical pathway are widely distributed in the placenta and decidua. In addition, C1q has recently been shown to significantly contribute to feto-maternal tolerance, trophoblast migration, and spiral artery remodeling, although the exact mechanism remains unknown. Pregnancy in mice, genetically deficient in C1q, mirrors symptoms similar to that of human preeclampsia. Thus, regulated complement activation has been proposed as an essential requirement for normal successful pregnancy. Little is known about the molecular pathways that regulate C1q expression in pregnancy. PU.1, an Ets-family transcription factor, is required for the development of hematopoietic myeloid lineage immune cells, and its expression is tissue-specific. Recently, PU.1 has been shown to regulate C1q gene expression in DCs and macrophages. Here, we have examined if PU.1 transcription factor regulates decidual C1q expression. We used immune-histochemical analysis, PCR, and immunostaining to localize and study the gene expression of PU.1 transcription factor in early human decidua. PU.1 was highly expressed at gene and protein level in early human decidual cells including trophoblast and stromal cells. Surprisingly, nuclear as well as cytoplasmic PU.1 expression was observed. Decidual cells with predominantly nuclear PU.1 expression had higher C1q expression. It is likely that nuclear and cytoplasmic PU.1 localization has a role to play in early pregnancy via regulating C1q expression in the decidua during implantation.

  7. Rate of evolution in brain-expressed genes in humans and other primates.

    Directory of Open Access Journals (Sweden)

    Hurng-Yi Wang

    2007-02-01

    Full Text Available Brain-expressed genes are known to evolve slowly in mammals. Nevertheless, since brains of higher primates have evolved rapidly, one might expect acceleration in DNA sequence evolution in their brain-expressed genes. In this study, we carried out full-length cDNA sequencing on the brain transcriptome of an Old World monkey (OWM and then conducted three-way comparisons among (i mouse, OWM, and human, and (ii OWM, chimpanzee, and human. Although brain-expressed genes indeed appear to evolve more rapidly in species with more advanced brains (apes > OWM > mouse, a similar lineage effect is observable for most other genes. The broad inclusion of genes in the reference set to represent the genomic average is therefore critical to this type of analysis. Calibrated against the genomic average, the rate of evolution among brain-expressed genes is probably lower (or at most equal in humans than in chimpanzee and OWM. Interestingly, the trend of slow evolution in coding sequence is no less pronounced among brain-specific genes, vis-à-vis brain-expressed genes in general. The human brain may thus differ from those of our close relatives in two opposite directions: (i faster evolution in gene expression, and (ii a likely slowdown in the evolution of protein sequences. Possible explanations and hypotheses are discussed.

  8. Rate of Evolution in Brain-Expressed Genes in Humans and Other Primates

    Science.gov (United States)

    Wang, Hurng-Yi; Chien, Huan-Chieh; Osada, Naoki; Hashimoto, Katsuyuki; Sugano, Sumio; Gojobori, Takashi; Chou, Chen-Kung; Tsai, Shih-Feng; Wu, Chung-I; Shen, C.-K. James

    2007-01-01

    Brain-expressed genes are known to evolve slowly in mammals. Nevertheless, since brains of higher primates have evolved rapidly, one might expect acceleration in DNA sequence evolution in their brain-expressed genes. In this study, we carried out full-length cDNA sequencing on the brain transcriptome of an Old World monkey (OWM) and then conducted three-way comparisons among (i) mouse, OWM, and human, and (ii) OWM, chimpanzee, and human. Although brain-expressed genes indeed appear to evolve more rapidly in species with more advanced brains (apes > OWM > mouse), a similar lineage effect is observable for most other genes. The broad inclusion of genes in the reference set to represent the genomic average is therefore critical to this type of analysis. Calibrated against the genomic average, the rate of evolution among brain-expressed genes is probably lower (or at most equal) in humans than in chimpanzee and OWM. Interestingly, the trend of slow evolution in coding sequence is no less pronounced among brain-specific genes, vis-à-vis brain-expressed genes in general. The human brain may thus differ from those of our close relatives in two opposite directions: (i) faster evolution in gene expression, and (ii) a likely slowdown in the evolution of protein sequences. Possible explanations and hypotheses are discussed. PMID:17194215

  9. Dopamine D2 receptor expression in the corticotroph cells of the human normal pituitary gland.

    Science.gov (United States)

    Pivonello, Rosario; Waaijers, Marlijn; Kros, Johan M; Pivonello, Claudia; de Angelis, Cristina; Cozzolino, Alessia; Colao, Annamaria; Lamberts, Steven W J; Hofland, Leo J

    2017-08-01

    The dopamine D2 receptor is the main dopamine receptor expressed in the human normal pituitary gland. The aim of the current study was to evaluate dopamine D2 receptor expression in the corticotroph cell populations of the anterior lobe and pars intermedia, as well as posterior lobe of the human normal pituitary gland by immunohistochemistry. Human normal pituitary gland samples obtained from routine autopsies were used for the study. In all cases, histology together with immunostaining for adrenocorticotropic hormone, melanocyte-stimulating hormone, prolactin, and neurofilaments were performed and compared to the immunostaining for D2 receptor. D2 receptor was heterogeneously expressed in the majority of the cell populations of the anterior and posterior lobe as well as in the area localized between the anterior and posterior lobe, and arbitrary defined as "intermediate zone". This zone, characterized by the presence of nerve fibers included the residual pars intermedia represented by the colloid-filled cysts lined by the remnant melanotroph cells strongly expressing D2 receptors, and clusters of corticotroph cells, belonging to the anterior lobe but localized within the cysts and adjacent to the posterior lobe, variably expressing D2 receptors. D2 dopamine receptor is expressed in the majority of the cell populations of the human normal pituitary gland, and particularly, in the different corticotroph cell populations localized in the anterior lobe and the intermediate zone of the pituitary gland.

  10. Modulation of cytokine expression in human macrophages by endocrine-disrupting chemical Bisphenol-A

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Yanzhen; Mei, Chenfang [State Key Laboratory of Applied Microbiology Southern China, Guangzhou 510070 (China); Guangdong Provincial Key Laboratory of Microbial Culture Collection and Application, Guangdong Institute of Microbiology, Guangzhou 510070 (China); Liu, Hao [Affiliated Cancer Hospital and Cancer Research Institute, Guangzhou Medical University, Guangzhou 510095 (China); Wang, Hongsheng [Department of Microbial and Biochemical Pharmacy, School of Pharmaceutical Sciences, Sun Yat-sen University, Guangzhou 510006 (China); Zeng, Guoqu; Lin, Jianhui [State Key Laboratory of Applied Microbiology Southern China, Guangzhou 510070 (China); Guangdong Provincial Key Laboratory of Microbial Culture Collection and Application, Guangdong Institute of Microbiology, Guangzhou 510070 (China); Xu, Meiying, E-mail: xumy@gdim.cn [State Key Laboratory of Applied Microbiology Southern China, Guangzhou 510070 (China); Guangdong Provincial Key Laboratory of Microbial Culture Collection and Application, Guangdong Institute of Microbiology, Guangzhou 510070 (China)

    2014-09-05

    Highlights: • Effects of BPA on the cytokines expression of human macrophages were investigated. • BPA increased pro-inflammation cytokines TNF-α and IL-6 production. • BPA decreased anti-inflammation IL-10 and TGF-β production. • ERα/β/ERK/NF-κB signaling involved in BPA-mediated cytokines expression. - Abstract: Exposure to environmental endocrine-disrupting chemical Bisphenol-A (BPA) is often associated with dysregulated immune homeostasis, but the mechanisms remain unclear. In the present study, the effects of BPA on the cytokines responses of human macrophages were investigated. Treatment with BPA increased pro-inflammation cytokines tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) production, but decreased anti-inflammation cytokines interleukin-10 (IL-10) and transforming growth factor-β (TGF-β) production in THP1 macrophages, as well as in primary human macrophages. BPA effected cytokines expression through estrogen receptor α/β (ERα/β)-dependent mechanism with the evidence of ERα/β antagonist reversed the expression of cytokines. We also identified that activation of extracellular regulated protein kinases (ERK)/nuclear factor κB (NF-κB) signal cascade marked the effects of BPA on cytokines expression. Our results indicated that BPA effected inflammatory responses of macrophages via modulating of cytokines expression, and provided a new insight into the link between exposure to BPA and human health.

  11. THE CONSTRUCTION AND EXPRESSION OF THE MURINE SCFV GENE IN E. COLI AGAINST HUMAN CERVICAL CANCER

    Institute of Scientific and Technical Information of China (English)

    Wang Ying; Chen Wei; Li Xu

    2006-01-01

    Objective To obtain the gene of murine Single chain Fv fragment (ScFv) against human cervical cancer and to express it in E. coli. Methods The variable region gene fragments of the heavy and light chains, which were amplified respectively using recombinant DNA techniques from CsA125 hybridoma cells, were spliced together through a flexible linker to ScFv against human cervical cancer. The ScFv genes were then cloned into expression vector pCANTAB 5E and expressed in E. coli HB2151 and TG1 respectively. The soluble ScFv were characterized by SDS PAGE and Western blot. The antigen-binding activities of the soluble and phage displayed ScFv were assayed by ELISA and cell immunohistochemical analysis. Results The expressed ScFv antibodies were soluble and phage displayed. The soluble ScFv secreted and expressed in E. coli HB2151 induced by IPTG were confirmed with SDS-PAGE, Western blot and ELISA. The specific binding capacity of the soluble and phage displayed ScFv to the surface associated antigen of human cervical cancer cell line was further confirmed with immunohistochemical studies. Conclusion The soluble and phage displayed ScFv expressed in E. coli against human cervical cancer showed high, specific affinity for the cervical cancer cell line surface associated antigen.

  12. A Human "eFP" Browser for Generating Gene Expression Anatograms.

    Directory of Open Access Journals (Sweden)

    Rohan V Patel

    Full Text Available Transcriptomic studies help to further our understanding of gene function. Human transcriptomic studies tend to focus on a particular subset of tissue types or a particular disease state; however, it is possible to collate into a compendium multiple studies that have been profiled using the same expression analysis platform to provide an overview of gene expression levels in many different tissues or under different conditions. In order to increase the knowledge and understanding we gain from such studies, intuitive visualization of gene expression data in such a compendium can be useful. The Human eFP ("electronic Fluorescent Pictograph" Browser presented here is a tool for intuitive visualization of large human gene expression data sets on pictographic representations of the human body as gene expression "anatograms". Pictographic representations for new data sets may be generated easily. The Human eFP Browser can also serve as a portal to other gene-specific information through link-outs to various online resources.

  13. Expression of SHH signaling pathway components in the developing human lung.

    Science.gov (United States)

    Zhang, Mingfeng; Wang, Hong; Teng, Hongqi; Shi, Jueping; Zhang, Yanding

    2010-10-01

    The Sonic hedgehog (Shh) cascade is crucial for the patterning of the early lung morphogenesis in mice, but its role in the developing human lung remains to be determined. In the present study, the expression patterns of SHH signaling pathway components, including SHH, PTCH1, SMO, GLI1, GLI2 and GLI3 were examined by in situ hybridization and immunohistochemistry, and compared with the equivalent patterns in mice. Our results showed that, as in mice, SHH was expressed in the epithelium of the developing human lung. However, SHH receptors (PTCH1 and SMO) and SHH signaling effectors (GLI1-3) were strongly detected in the human lung epithelium, but weakly in the mesenchyme, slightly different from their expressions in mice. Furthermore, the expression levels of SHH signaling pathway genes in human lung, but not that of GLI1, were subsequently downregulated at the canalicular stage evaluated by real-time PCR, coincident with a decline in the developing murine lung. In conclusion, in spite of slight differences, the considerable similarities of gene expression in human and mice suggest that conserved molecular networks regulate mammalian lung development.

  14. Gene expression profiles of human promyelocytic leukemia cell lines exposed to volatile organic compounds.

    Science.gov (United States)

    Sarma, Sailendra Nath; Kim, Youn-Jung; Ryu, Jae-Chun

    2010-05-27

    Benzene, toluene, o-xylene, ethylbenzene, trichloroethylene and dichloromethane are the most widely used volatile organic compounds (VOCs), and their toxic mechanisms are still undefined. This study analyzed the genome-wide expression profiles of human promyelocytic leukemia HL-60 cells exposed to VOCs using a 35-K whole human genome oligonucleotide microarray to ascertain potential biomarkers. Genes with a significantly increased expression levels (over 1.5-fold and p-values p53 signaling pathway, apoptosis, and natural killer cell-mediated cytotoxicity pathway. Functionally important immune response- and apoptosis-related genes were further validated by real-time RT-PCR. The results showed that IFIT1, IFIT2, IFIT3, USP18, INFGR2, PMAIP1, GADD45A, NFKBIA, TNFAIP3, and BIRC3 genes altered their expression profiles in a dose-dependent manner. Similar expressions profiles were also found in human erythromyeloblastoid leukemia K562 cells and in human leukemic monocyte lymphoma U937 cells. In conclusion, both gene expression profiles and gene ontology analysis have elucidated potential gene-based biomarkers and provided insights into the mechanism underlying the response of human leukemia cell lines to VOC exposure.

  15. Zinc at Sub-Cytotoxic Concentrations Induces Heme Oxygenase-1 Expression in Human Cancer Cells

    Directory of Open Access Journals (Sweden)

    Jing Xue

    2013-07-01

    Full Text Available Background/Aims: This study investigated the effects of zinc on heme oxygenase-1 (HO-1 expression in human cancer cells. Methods/Results: Zinc at sub-cytotoxic concentrations (50-100 μM induces HO-1 expression in the MDA-MB-231 (human breast cancer and A2780 (human ovarian cancer cell lines in a concentration- and time-dependent manner. The induction of HO-1 by zinc was detected after 4-6 hours of treatment, reached maximal level at 8 hours, and declined thereafter. Using a human HO-1 gene promoter reporter construct, we identified two antioxidant response elements (AREs that mediated the zinc-induced increase in HO-1 gene transcription, indicating that the nuclear factor (erythroid-derived 2-like 2 (Nrf2 signaling pathway is involved in this event. This assumption was supported by the observations that knockdown of Nrf2 expression compromised the zinc-induced increase in HO-1 gene transcription, and that zinc increased Nrf2 protein expression and the Nrf2 binding to the AREs. Additionally, we found that the zinc-induced HO-1 gene transcription can be enhanced by clioquinol, a zinc ionophore, and reversed by pretreatment with TPEN, a known zinc chelator, indicating that an increase in intracellular zinc levels is responsible for this induction. Conclusion: These findings demonstrate that zinc at sub-cytotoxic concentrations induces HO-1 expression in human cancer cells. The biological significance of this induction merits further investigation.

  16. Differential expression of CD150 (SLAM) family receptors by human hematopoietic stem and progenitor cells.

    Science.gov (United States)

    Sintes, Jordi; Romero, Xavier; Marin, Pedro; Terhorst, Cox; Engel, Pablo

    2008-09-01

    Human hematopoietic stem cell (HSC)-containing grafts are most commonly used to treat various blood diseases, including leukemias and autoimmune disorders. CD150 (SLAM) family receptors have recently been shown to be differentially expressed by mouse HSC and progenitor cells. Members of the CD150 family are key regulators of leukocyte activation and differentiation. The goal of the present study is to analyze the expression patterns of the CD150 receptors CD48, CD84, CD150 (SLAM), CD229 (Ly9), and CD244 (2B4) on the different sources of human hematopoietic stem and progenitor cells. Expression of CD150 receptors was analyzed on human mobilized peripheral blood CD133(+)-isolated cells and CD34(+) bone marrow (BM) and umbilical cord blood (CB) cells using multicolor flow cytometry. CD244 was present on most CD133(+)Lin(-)-mobilized cells and CD34(+)Lin(-) BM and CB cells, including virtually all CD38(-)Lin(-) primitive progenitor cells. CD48 had a restricted expression pattern on CD133(+)Lin(-)CD38(-) cells, while its levels were significantly higher in CD34(+)Lin(-) BM and CB cells. In addition, CD84 was present on a significant number of CD133(+)Lin(-) cells, but only on a small fraction of CD133(+)Lin(-)CD38(-) peripheral blood mobilized cells. In contrast, CD84 was expressed on practically all CD34(+)Lin(-) BM cells. No CD150 expression was observed in mobilized peripheral blood CD133(+)Lin(-) or CD34(+)Lin(-) BM and CB cells. Furthermore, only a small fraction of CD34(+)Lin(-) BM and CB cells expressed CD229. Our results show that CD150 family molecules are present on human hematopoietic stem and progenitor cells and that their expression patterns differ between humans and mice.

  17. Systematic analysis of a novel human renal glomerulus-enriched gene expression dataset.

    Directory of Open Access Journals (Sweden)

    Maja T Lindenmeyer

    Full Text Available Glomerular diseases account for the majority of cases with chronic renal failure. Several genes have been identified with key relevance for glomerular function. Quite a few of these genes show a specific or preferential mRNA expression in the renal glomerulus. To identify additional candidate genes involved in glomerular function in humans we generated a human renal glomerulus-enriched gene expression dataset (REGGED by comparing gene expression profiles from human glomeruli and tubulointerstitium obtained from six transplant living donors using Affymetrix HG-U133A arrays. This analysis resulted in 677 genes with prominent overrepresentation in the glomerulus. Genes with 'a priori' known prominent glomerular expression served for validation and were all found in the novel dataset (e.g. CDKN1, DAG1, DDN, EHD3, MYH9, NES, NPHS1, NPHS2, PDPN, PLA2R1, PLCE1, PODXL, PTPRO, SYNPO, TCF21, TJP1, WT1. The mRNA expression of several novel glomerulus-enriched genes in REGGED was validated by qRT-PCR. Gene ontology and pathway analysis identified biological processes previously not reported to be of relevance in glomeruli of healthy human adult kidneys including among others axon guidance. This finding was further validated by assessing the expression of the axon guidance molecules neuritin (NRN1 and roundabout receptor ROBO1 and -2. In diabetic nephropathy, a prevalent glomerulopathy, differential regulation of glomerular ROBO2 mRNA was found.In summary, novel transcripts with predominant expression in the human glomerulus could be identified using a comparative strategy on microdissected nephrons. A systematic analysis of this glomerulus-specific gene expression dataset allows the detection of target molecules and biological processes involved in glomerular biology and renal disease.

  18. Unusual patterns of immunoglobulin gene rearrangement and expression during human B cell ontogeny: human B cells can simultaneously express cell surface kappa and lambda light chains

    OpenAIRE

    1993-01-01

    Immunoglobulin gene rearrangement during mammalian B cell development generally follows an ordered progression, beginning with heavy (H) chain genes and proceeding through kappa and lambda light (L) chain genes. To determine whether the predicted kappa-->lambda hierarchy was occurring in vitro, we generated Epstein-Barr virus-transformed cell lines from cultures undergoing human pre-B cell differentiation. A total of 143 cell lines were established. 24 expressed cell surface mu/lambda by flow...

  19. Expression of neurotrimin in the normal and injured adult human spinal cord.

    Science.gov (United States)

    Grijalva, I; Li, X; Marcillo, A; Salzer, J L; Levi, A D

    2006-05-01

    Neurotrimin (Ntm) is a member of the family of neural cell adhesion molecules. Its expression pattern suggests that Ntm promotes axonal fasciculation, guides nerve fibers to specific targets and stabilizes synapses as it accumulates coincident with synaptogenesis. Strong labeling of Ntm was observed in motor and sensory areas of the postnatal rat cortex. It is not known whether Ntm is present in adult human spinal cord (SC). In the present study, a monoclonal antibody specific for Ntm (1B1), is applied to the first study of the expression of Ntm in normal and injured adult human SC. (1) To investigate the expression pattern of Ntm in adult normal human SC, and (2) to observe the changes of Ntm expression after SC injury and compare the differences between normal and injured adult human SC. Human SC tissue was obtained from necropsies of patients with (n=5) and without (n=4) SC injury. The 1B1 Ntm monoclonal antibody was used for immunohistochemical staining on paraffin embedded sections with an ABC kit. (1) In total, 12 slides were analyzed for each group from both cervical and thoracic levels. Motor neurons and Clarke's neurons and glial-like cells were mild to moderately positive in all uninjured SC specimens. (2) In injured SC, no staining was observed in the injury epicenter between two and three levels proximally and distally, but was detected five levels away. (3) In patients older than 67 years of age, Ntm-positive inclusions were present in the white matter of the SC with or without injury. (4) Some meningeal cells were strongly Ntm-positive, especially in the uninjured human SC. Ntm is expressed by motor and Clarke's neurons and glial cells in uninjured human SC. The downregulation of Ntm in the injured SC suggests that its expression is regulated by afferent input. Spinal Cord (2006) 44, 275-279. doi:10.1038/sj.sc.3101840; published online 20 September 2005.

  20. Secretory expression of human protein in the Yeast Pichia pastoris by controlled fermentor culture.

    Science.gov (United States)

    Murasugi, Akira

    2010-06-01

    The heterologous protein expression system of Pichia pastoris is now widely used for expression of many human proteins, because the efficiently expressed proteins will be correctly folded in Pichia pastoris cells and also efficiently secreted from the cells. Recombinant human serum albumin (rHSA) is efficiently secreted from Pichia pastoris. Nowadays, the expression of rHSA exceeds 10g in 1 L fermentor culture broth, and the protein is completely purified. Recombinant HSA expressed in Pichia pastoris was approved as a medicine by the authorities in 2007, and launched in 2008 in Japan. One of the insulin precursors (IP) was also successfully expressed in Pichia pastoris, and secreted up to 3.6g in 1 L medium using a multi-copy transformant. The insulin precursor could be efficiently converted to insulin, the final product, in vitro. Human growth hormone was also expressed in Pichia pastoris, and secreted up to 49 mg in 1 L medium. These proteins are also important for clinical applications. Midkine and pleiotrophin may be two of the candidates for clinical applications. Secretion signals, the copy number of an expression cassette in transformants, and culture conditions for fermentation were examined for efficient expression of these proteins in Pichia pastoris. The best signal was selected, and other factors were optimized. The amounts of native midkine and native pleiotrophin expressed were approximately 0.36g and 0.26g in 1 L medium, respectively. Expression of bile-salt stimulated lipase (BSSL) had been extremely low in the beginning of a fermentor culture experiment. However, approximately 1 g rBSSL in 1 L medium was finally expressed in a fermentor by unlimited feeding of glycerol for cell growth and optimization of other factors. BSSL from human milk and rBSSL from Pichia cells are glycosylated. The structure differences between these glycans are obvious. When humanization of Pichia glycans is established by genetic engineering, the Pichia pastoris expression

  1. Expression and Bioinformatics Analysis of SPACA4 in Human and Mice

    Institute of Scientific and Technical Information of China (English)

    Ai-fa TANG; Zhen-dong YU; Yao-ting GUI; Xin GUO; Xian-xin LI; Wei-xiang LIU; Hui ZHU; Zhi-ming CAI

    2008-01-01

    Objective To analyze the expression of SPACA4 in human and mice. Methods Testes cRNA samples from Balb/c mice of different postnatal days were performed with mouse affymetrix chip to screen the expression of SPACA4 in mice. Sub-quantitative RT-PCR and bioinformatic tools were used here to describe the expression profile of SPACA4 in mice and human. Results The results of gene chip analysis indicated that the expression of mSPACA4 began after d 35 of postnatal testis in mice. Sub-quantitative RT-PCR assay showed that SPACA4 gene expressed exclusively in mouse and human testis, and mouse mSPACA4 gene expressed after d 35 of postnatal testis that was consistency with the results of gene chip analysis. By bioinformatics analysis, mSPACA4 is located in cell membrane (34.8%) or plasma membrane (34.8%), the signal peptide cleavage site between position 19 and 20 amino acids, transmembrane region between 2-20 and 101-126 amino acids, respectively, on mSPACA4 protein. Conclusion mSPACA4 and hSPACA4 were testis-specific genes, and the expression of mSPACA4 begins after d 35 of postnatal testis in mice. SPACA4 is a candidate for targeting in a sperm-based contraceptive vaccine.

  2. SOX9 is expressed in normal stomach, intestinal metaplasia, and gastric carcinoma in humans.

    Science.gov (United States)

    Sashikawa Kimura, Miho; Mutoh, Hiroyuki; Sugano, Kentaro

    2011-11-01

    SOX9 is a marker for stem cells in the intestine and overexpression of SOX9 is found in some types of cancer. However, the expression of SOX9 in normal stomach, precancerous intestinal metaplasia, and gastric carcinoma has not yet been clarified. This study aimed to investigate SOX9 expression in the corpus and pyloric regions of the normal human stomach, premalignant intestinal metaplasia, and gastric carcinoma by using immunohistochemistry. We evaluated SOX9 expression in 46 clinical samples (early gastric well-differentiated adenocarcinoma including surrounding intestinal metaplasia) resected under esophagogastroduodenoscopy. A small amount of SOX9 was expressed in the neck/isthmus of the corpus region and SOX9 expression was predominantly restricted to the neck/isthmus of the pyloric region in normal human stomach. In the intestinal metaplastic mucosa, SOX9- and PCNA-positive cells were located at the base of the intestinal metaplastic mucosa. Almost all of the gastric carcinoma cells expressed SOX9. SOX9 is expressed in intestinal metaplasia and gastric carcinoma in humans.

  3. MHC class II expression in human basophils: induction and lack of functional significance.

    Directory of Open Access Journals (Sweden)

    Astrid L Voskamp

    Full Text Available The antigen-presenting abilities of basophils and their role in initiating a Th2 phenotype is a topic of current controversy. We aimed to determine whether human basophils can be induced to express MHC Class II and act as antigen presenting cells for T cell stimulation. Isolated human basophils were exposed to a panel of cytokines and TLR-ligands and assessed for MHC Class II expression. MHC Class II was expressed in up to 17% of isolated basophils following incubation with a combination of IL-3, IFN-γ and GM-CSF for 72 hours. Costimulatory molecules (CD80 and CD86 were expressed at very low levels after stimulation. Gene expression analysis of MHC Class II-positive basophils confirmed up-regulation of HLA-DR, HLA-DM, CD74 and Cathepsin S. However, MHC Class II expressing basophils were incapable of inducing antigen-specific T cell activation or proliferation. This is the first report of significant cytokine-induced MHC Class II up-regulation, at both RNA and protein level, in isolated human basophils. By testing stimulation with relevant T cell epitope peptide as well as whole antigen, the failure of MHC Class II expressing basophils to induce T cell response was shown not to be solely due to inefficient antigen uptake and/or processing.

  4. Expression of nucleophosmin in glandular epithelium of non-pregnant human endometrium during the menstrual cycle

    Institute of Scientific and Technical Information of China (English)

    KUANG Ye; XU Peng; WEN Hai-xia; KONG Xian-chao; GUAN Li-li; LI Pei-ling

    2011-01-01

    Background Nucleophosmin plays a critical role in embryonic development. This study aimed to examine the expression pattern of nucleophosmin in glandular epithelium of human endometrium during the menstrual cycle.Methods Endometrial tissues used for this study were obtained from 46 non-pregnant patients who underwent hysterectomy which had been performed to treat benign diseases. Nucleophosmin expression was assessed by in situ hybridization and immunohistochemistry.Results At the early-, mid- and late-proliferative phase, nucleophosmin mRNA was highly expressed in glandular epithelium of human endometrium. At the secretory phase, the expression of nucleophosmin mRNA was reduced in glandular epithelium in early-secretory phase, and the expression in mid- and late-secretory phases was not detected.Similarly, nucleophosmin protein was strongly expressed in endometrial glands throughout the proliferative phase, but was gradually reduced during secretory phase.Conclusion Nucleophosmin mRNA and protein are expressed in glandular epithelium of human endometrium throughout the menstrual cycle.

  5. Regulation of osteoprotegerin expression by Notch signaling in human oral squamous cell carcinoma cell line

    Institute of Scientific and Technical Information of China (English)

    Jeeranan Manokawinchoke; Thanaphum Osathanon; Prasit Pavasant

    2016-01-01

    Objective: To investigate the influence of Notch signaling on osteoprotegerin(OPG)expression in a human oral squamous cell carcinoma cell line.Methods: Activation of Notch signaling was performed by seeding cells on Jagged1 immobilized surfaces. In other experiments, a g-secretase inhibitor was added to the culture medium to inhibit intracellular Notch signaling. OPG m RNA and protein were determined by real-time PCR and ELISA, respectively. Finally, publicly available microarray database analysis was performed using connection up- or down-regulation expression analysis of microarrays software.Results: Jagged1-treatment of HSC-4 cells enhanced HES1 and HEY1 m RNA expression, confirming the intracellular activation of Notch signaling. OPG m RNA and protein levels were significantly suppressed upon Jagged1 treatment. Correspondingly, HSC-4 cells treated with a g-secretase inhibitor resulted in a significant reduction of HES1 and HEY1 m RNA levels, and a marked increase in OPG protein expression was observed.These results implied that Notch signaling regulated OPG expression in HSC-4 cells.However, Jagged1 did not alter OPG expression in another human oral squamous cell carcinoma cell line(HSC-5) or a human head and neck squamous cell carcinoma cell line(HN22).Conclusions: Notch signaling regulated OPG expression in an HSC-4 cell line and this mechanism could be cell line specific.

  6. Expression of interleukin-6 is downregulated by 17-(allylamino)-17-demethoxygeldanamycin in human prostatic carcinoma cells

    Institute of Scientific and Technical Information of China (English)

    Ke-hung TSUI; Tsui-hsia FENG; Wen-chi HSIEH; Phei-lang CHANG; Horng-heng JUANG

    2008-01-01

    Aim: Interleukin-6 (IL-6) is a pleiotropic cytokine that is associated with tumor metastasis and prostate cancer. We evaluated the mechanism and effect of 17-(allylamino)-17-demethoxygeldanamycin (17AAG), a novel inhibitor of heat shock protein 90 (Hsp90), on the IL-6 gene expression in human prostatic carcinoma (PC-3) cells. Methods: Quantitative IL-6 and IL-6 receptor (IL-6R) expressions were assessed using RT-PCR. The deregulation of 17AAG and phor-bol 12-myristate 13-acetate (PMA) on the IL-6 gene was determined by ELISA and transient gene expression assays using an IL-6 reporter vector. Results: Although the IL-6R is ubiquitously expressed by prostatic epithelium cells, the IL-6 expression is only found in advanced prostatic carcinoma cells, such as PC-3 and DU145. Further studies using RT-PCR indicated that 17AAG downregulated the gene expression of IL-6. ELISA and the transient gene expression assay revealed that 17AAG blocked the stimulation of PMA of IL-6 gene expression in PC-3 cells. The PMA-induced IL-6 gene expression is dependent on the NF-κB response element. However, the effect of 17AAG appears to be mediated via a region located at -149 to +8 bp upstream of the transcriptional starting site of the IL-6 gene, and might not be through the NF-κB signaling pathway. Conclusion: The present study reveals that IL-6 is transcriptionally downregulated in human prostatic carcinoma cells in response to 17AAG. This result suggests the presence of a novel Hsp90 mediation pathway that is involved in the deregulation on the transcription of the human IL-6 gene in human prostate cancer.

  7. Global gene expression profiling of individual human oocytes and embryos demonstrates heterogeneity in early development.

    Directory of Open Access Journals (Sweden)

    Lisa Shaw

    Full Text Available Early development in humans is characterised by low and variable embryonic viability, reflected in low fecundity and high rates of miscarriage, relative to other mammals. Data from assisted reproduction programmes provides additional evidence that this is largely mediated at the level of embryonic competence and is highly heterogeneous among embryos. Understanding the basis of this heterogeneity has important implications in a number of areas including: the regulation of early human development, disorders of pregnancy, assisted reproduction programmes, the long term health of children which may be programmed in early development, and the molecular basis of pluripotency in human stem cell populations. We have therefore investigated global gene expression profiles using polyAPCR amplification and microarray technology applied to individual human oocytes and 4-cell and blastocyst stage embryos. In order to explore the basis of any variability in detail, each developmental stage is replicated in triplicate. Our data show that although transcript profiles are highly stage-specific, within each stage they are relatively variable. We describe expression of a number of gene families and pathways including apoptosis, cell cycle and amino acid metabolism, which are variably expressed and may be reflective of embryonic developmental competence. Overall, our data suggest that heterogeneity in human embryo developmental competence is reflected in global transcript profiles, and that the vast majority of existing human embryo gene expression data based on pooled oocytes and embryos need to be reinterpreted.

  8. Long non-coding RNA expression during aging in the human subependymal zone

    Directory of Open Access Journals (Sweden)

    Guy eBarry

    2015-03-01

    Full Text Available The human subependymal zone (SEZ is debatably a source of newly born neurons throughout life and neurogenesis is a multi-step process requiring distinct transcripts during cell proliferation and early neuronal maturation, along with orchestrated changes in gene expression during cell state/fate transitions. Furthermore, it is becoming increasingly clear that the majority of our genome that results in production of non-protein coding RNAs plays vital roles in the evolution, development, adaptation and region-specific function of the human brain. We predicted that some transcripts expressed in the SEZ may be unique to this specialized brain region, and that a comprehensive transcriptomic analysis of this region would aid in defining expression changes during neuronal birth and growth in adult humans. Here, we used deep RNA sequencing of human SEZ tissue during adulthood and aging to characterize the transcriptional landscape with a particular emphasis on long non-coding RNAs (lncRNAs. The data shows predicted age-related changes in mRNAs encoding proliferation, progenitor and inflammatory proteins as well as a unique subset of lncRNAs that are highly expressed in the human SEZ, many of which have unknown functions. Our results suggest the existence of robust proliferative and neuronal differentiation potential in the adult human SEZ and lay the foundation for understanding the involvement of lncRNAs in postnatal neurogenesis and potentially associated neurodevelopmental diseases that emerge after birth.

  9. Expression of CD1d protein in human testis showing normal and abnormal spermatogenesis.

    Science.gov (United States)

    Adly, Mohamed A; Abdelwahed Hussein, Mahmoud-Rezk

    2011-05-01

    CD1d is a member of CD1 family of transmembrane glycoproteins, which represent antigen-presenting molecules. Immunofluorescent staining methods were utilized to examine expression pattern of CD1d in human testicular specimens. In testis showing normal spermatogenesis, a strong CD1d cytoplasmic expression was seen the Sertoli cells, spermatogonia, and Leydig cells. A moderate expression was observed in the spermatocytes. In testes showing maturation arrest, CD1d expression was strong in the Sertoli cells and weak in spermatogonia and spermatocytes compared to testis with normal spermatogenesis. In Sertoli cell only syndrome, CD1d expression was strong in the Sertoli and Leydig cells. This preliminary study displayed testicular infertility-related changes in CD1d expression. The ultrastructural changes associated with with normal and abnormal spermatogenesis are open for further investigations.

  10. Regulation of protease-activated receptor-1 expression in human buccal fibroblasts stimulated with arecoline.

    Science.gov (United States)

    Tsai, Chung-Hung; Lee, Shiuan-Shinn; Huang, Fu-Mei; Chang, Yu-Chao

    2013-09-01

    The purpose of this study was to compare the major thrombin receptor protease-activated receptor-1 (PAR-1) expression in normal human buccal mucosa and oral submucous fibrosis (OSF) specimens and further explore the potential mechanisms that may lead to induce PAR-1 expression. Thirty OSF and 10 normal buccal mucosa specimens were examined by immunohistochemistry. Buccal mucosal fibroblasts (BMFs) were challenged with arecoline by using Western blot analysis. N-acetyl-L-cysteine (NAC), LY294002, herbimycin A, NS-398, and PD98059 were added to find the possible regulatory mechanisms. PAR-1 expression was significantly higher in OSF specimens (p Arecoline was found to elevate PAR-1 expression in a dose-dependent and time-dependent manner (p arecoline-induced PAR-1 expression (p Arecoline-induced PAR-1 expression was downregulated by NAC, LY294002, herbimycin A, NS398, and PD98059. Copyright © 2012 Wiley Periodicals, Inc.

  11. Expression of hexokinase II and Glut-1 in untreated human breast cancer

    Energy Technology Data Exchange (ETDEWEB)

    Brown, Raya S.; Goodman, Tonya M.; Zasadny, Kenneth R.; Greenson, Joel K.; Wahl, Richard L. E-mail: rsbrown@umich.edu

    2002-05-01

    Expressions of HKII and Glut-1 were studied in untreated primary human breast cancers by immunohistochemistry. 79% of the breast cancers were HKII-positive and 61% were Glut-1-positive. Average positive malignant cell areas were 66{+-}41% for HKII and 29{+-}36% for Glut-1. HKII staining was cytoplasmic, suggesting mitochondrial localization with no variations in staining intensities. Glut-1 staining was heterogeneous, cytoplasmic and membranous and varied with histology and tumor stage. Cells expressing HKII did not always express Glut-1 and vice versa. Increased FDG-uptake appeared to be associated with increased Glut-1 expression (P=0.021), but not with HKII expression (p=0.6). FDG uptake in breast cancer tissue appears to be associated with the extent of immunodetectable expression of Glut-1, but not that of HKII, and FDG uptake may differ between individual tumors depending on tumor stage and histology.

  12. Regulation of bradykinin receptor gene expression in human lung fibroblasts.

    Science.gov (United States)

    Phagoo, S B; Yaqoob, M; Herrera-Martinez, E; McIntyre, P; Jones, C; Burgess, G M

    2000-06-01

    In WI-38 human fibroblasts, interleukin-1 beta and tumour necrosis factor-alpha (TNF-alpha) increased bradykinin B(1) receptor mRNA, which peaked between 2 and 4 h, remaining elevated for 20 h. Binding of the bradykinin B(1) receptor selective ligand [3H]des-Arg(10)-kallidin, also increased, peaking at 4 h and remaining elevated for 20 h. The B(max) value for [3H]des-Arg(10)-kallidin rose from 280+/-102 fmol/mg (n=3) to 701+/-147 fmol/mg (n=3), but the K(D) value remained unaltered (control, 1.04+/-0.33 nM (n=3); interleukin-1 beta, 0.88+/-0.41 nM (n=3)). The interleukin-1 beta-induced [3H]des-Arg(10)-kallidin binding sites were functional receptors, as bradykinin B(1) receptor agonist-induced responses increased in treated cells. Bradykinin B(2) receptor mRNA and [3H]bradykinin binding were upregulated by interleukin-1 beta, but not TNF-alpha. The effect of interleukin-1 beta on bradykinin B(2) receptors was smaller than for bradykinin B(1) receptors. Cycloheximide prevented interleukin-1 beta-mediated increases in B(1) and B(2) binding, but not mRNA suggesting that de novo synthesis of a transcriptional activator was unnecessary.

  13. Expression, purification and preliminary crystallographic studies of human ketohexokinase.

    Science.gov (United States)

    Kozak, M; Hayward, B; Borek, D; Bonthron, D T; Jaskólski, M

    2001-04-01

    Ketohexokinase (KHK; E.C. 2.7.1.3) catalyses the (reversible) phosphorylation of fructose to fructose-1-phosphate. KHK is the first enzyme in a specialized catabolic pathway metabolizing dietary fructose to the glycolytic intermediate glyceraldehyde-3-phosphate. Mutations inactivating KHK underlie the metabolic disorder essential fructosuria. The primary structure of KHK shows no significant homology to other mammalian hexokinases. It is most similar to prokaryotic ribokinases, but catalyses a distinct phosphorylation reaction. Recombinant human KHK has been crystallized in the orthorhombic form (space group P2(1)2(1)2 or P2(1)2(1)2(1)). Single crystals of this polymorph suitable for X-ray diffraction have been obtained by vapour diffusion using 2-propanol and MPD as precipitants (pH 7.5). The crystals have unit-cell parameters a = 93.4, b = 121.5, c = 108.4 A. Diffraction data were collected to 4.3 A resolution. The asymmetric unit contains four protein molecules.

  14. Expression profile of critical genes involved in FG