Gene Expression Changes Related to Endocrine Function and Decline in Reproduction in Fathead Minnow (Pimephales promelas) after Dietary Methylmercury Exposure

BACKGROUND: Methylmercury (MeHg) is a known neurotoxic agent, but the mechanisms by which MeHg may act on reproductive pathways are relatively unknown. Several studies have indicated potential changes in hormone levels as well as declines in vertebrates with increasing dietary MeHg exposure. OBJECTIVES: The purpose of this study was to identify alterations in gene expression associated with MeHg exposure, specifically those associated with previously observed changes in reproduction and reproductive biomarkers. Fathead minnows, Pimephales promelas, were fed one of three diets that were similar to documented concentrations of MeHg in the diets of wild invertivorous and piscivorous fish. We used a commercial macroarray in conjunction with quantitative polymerase chain reaction to examine gene expression in fish in relation to exposure to these environmentally relevant doses of MeHg. RESULTS: Expression of genes commonly associated with endocrine disruption was altered with Hg exposure. Specifically, we observed a marked up-regulation in vitellogenin mRNA in individual Hg-exposed males and a significant decline in vitellogenin gene expression in female fish with increasing Hg concentrations. Other genes identified by the macroarray experiment included those associated with egg fertilization and development, sugar metabolism, apoptosis, and electron transport. We also observed differences in expression patterns between male and female fish not related to genes specifically associated with reproduction, indicating a potential physiological difference in the reaction of males and females to MeHg. CONCLUSION: Gene expression data may provide insight into the mechanisms by which MeHg affects reproduction in fish and indicate how MeHg differs in its effect from other heavy metals and endocrine-disrupting compounds

Evaluation of glyceraldehyde-3-phosphate, prolylpeptidyl isomerase A, and a set of stably expressed genes as reference mRNAs in urate crystal inflammation

ABSTRACT

Vascular endothelial growth factor A (VEGF-A) mRNA expression levels decrease after menopause in normal breast tissue but not in breast cancer lesions

We hypothesized that the regulation of microvascular functions and angiogenesis in breast tissue, a well known target of ovarian steroid action, is dependent on the hormonal exposure of the breast. Relative expression levels of VEGF-A (vascular endothelial growth factor A), a putative key regulator of angiogenesis in breast cancer, were analysed in the tumour and the adjacent non-neoplastic breast tissue of 19 breast cancer patients by quantitative reverse transcriptase polymerase chain reaction. In non-neoplastic breast specimens the expression levels of all detected VEGF-A-isoforms (189, 165, 121) were significantly higher in premenopausal compared to post-menopausal women (P = 0.02) and were inversely correlated with the patient's age (P = 0.006). In contrast, in cancerous tissues menopausal status had no influence on VEGF-A-expression levels. Benign and malignant tissues exhibited a similar expression pattern of VEGF-A-isoforms relative to each other. Thus, the regulation of the vasculature in normal breast tissue, as opposed to breast cancer tissue, appears to be hormonally dependent. Endogenous and therapeutically used hormonal steroids might, therefore, cause clinically relevant changes of the angiogenic phenotype of the human breast. © 1999 Cancer Research Campaig

In vivo Identification and Specificity assessment of mRNA markers of hypoxia in human and mouse tumors

<p>Abstract</p> <p>Background</p> <p>Tumor hypoxia is linked to poor prognosis, but identification and quantification of tissue hypoxia remains a challenge. The hypoxia-specificity of HIF-1α target genes in vivo has been questioned due to the confounding influence of other microenvironmental abnormalities known to affect gene expression (e.g., low pH). Here we describe a new technique that by exploiting intratumoral oxygenation heterogeneity allows us to identify and objectively rank the most robust mRNA hypoxia biomarkers.</p> <p>Methods</p> <p>Mice carrying human (FaDu_dd) or murine (SCCVII) tumors were injected with the PET hypoxia tracer FAZA. Four hours post-injection tumors were removed, frozen, and crushed into milligram-sized fragments, which were transferred individually to pre-weighed tubes containing RNAlater and then weighed. For each fragment radioactivity per tissue mass and expression patterns of selected mRNA biomarkers were analyzed and compared.</p> <p>Results</p> <p>In both tumour models, fragmentation into pieces weighing 10 to 60 mg resulted in tissue fragments with highly variable relative content of hypoxic cells as evidenced by an up to 13-fold variation in FAZA radioactivity per mass of tissue. Linear regression analysis comparing FAZA retention with patterns of gene expression in individual tissue fragments revealed that CA9, GLUT1 and LOX mRNA levels were equally and strongly correlated to hypoxic extent in FaDu_dd. The same link between hypoxia and gene expression profile was observed for CA9 and GLUT1, but not LOX, in SCCVII tumors. Apparent in vivo hypoxia-specificity for other putative molecular markers of tissue hypoxia was considerably weaker.</p> <p>Conclusions</p> <p>The portrayed technique allows multiple pairwise measurements of mRNA transcript levels and extent of hypoxia in individual tumors at a smallest possible volumetric scale which (by limiting averaging effects inherent to whole-tumor analysis) strengthen the conclusiveness on true hypoxia-specificity of candidate genes while limiting the required number of tumors. Among tested genes, our study identified CA9, GLUT1 and possibly LOX as highly specific biomarkers of tumor hypoxia in vivo.</p

Decellularized Matrix from Tumorigenic Human Mesenchymal Stem Cells Promotes Neovascularization with Galectin-1 Dependent Endothelial Interaction

BACKGROUND: Acquisition of a blood supply is fundamental for extensive tumor growth. We recently described vascular heterogeneity in tumours derived from cell clones of a human mesenchymal stem cell (hMSC) strain (hMSC-TERT20) immortalized by retroviral vector mediated human telomerase (hTERT) gene expression. Histological analysis showed that cells of the most vascularized tumorigenic clone, -BD11 had a pericyte-like alpha smooth muscle actin (ASMA+) and CD146+ positive phenotype. Upon serum withdrawal in culture, -BD11 cells formed cord-like structures mimicking capillary morphogenesis. In contrast, cells of the poorly tumorigenic clone, -BC8 did not stain for ASMA, tumours were less vascularized and serum withdrawal in culture led to cell death. By exploring the heterogeneity in hMSC-TERT20 clones we aimed to understand molecular mechanisms by which mesenchymal stem cells may promote neovascularization. METHODOLOGY/PRINCIPAL FINDINGS: Quantitative qRT-PCR analysis revealed similar mRNA levels for genes encoding the angiogenic cytokines VEGF and Angiopoietin-1 in both clones. However, clone-BD11 produced a denser extracellular matrix that supported stable ex vivo capillary morphogenesis of human endothelial cells and promoted in vivo neovascularization. Proteomic characterization of the -BD11 decellularized matrix identified 50 extracellular angiogenic proteins, including galectin-1. siRNA knock down of galectin-1 expression abrogated the ex vivo interaction between decellularized -BD11 matrix and endothelial cells. More stable shRNA knock down of galectin-1 expression did not prevent -BD11 tumorigenesis, but greatly reduced endothelial migration into -BD11 cell xenografts. CONCLUSIONS: Decellularized hMSC matrix had significant angiogenic potential with at least 50 angiogenic cell surface and extracellular proteins, implicated in attracting endothelial cells, their adhesion and activation to form tubular structures. hMSC -BD11 surface galectin-1 expression was required to bring about matrix-endothelial interactions and for xenografted hMSC -BD11 cells to optimally recruit host vasculature

Novel roles for GAPDH in cell death and carcinogenesis

Publicado en línea el 25 de septiembre de 2009Growing evidence points to the fact that glucose metabolism has a central role in carcinogenesis. Among the enzymes controlling this energy production pathway, glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is of particular interest. Initially identified as a glycolytic enzyme and considered as a housekeeping gene, this enzyme is actually tightly regulated and is involved in numerous cellular functions. Particularly intriguing are recent reports describing GAPDH as a regulator of cell death. However, its role in cell death is unclear; whereas some studies point toward a proapoptotic function, others describe a protective role and suggest its participation in tumor progression. In this study, we highlight recent findings and discuss potential mechanisms through which cells regulate GAPDH to fulfill its diverse functions to influence cell fate.This work was supported in part by l’Association pour la Recherche sur le Cancer, by l’Agence Nationnal de la Recherche, la Fondation de France, Plan Nacional I+D SAF2008-04974 and by grants from The U.S. National Institutes of Health. J-E.R. is a recipient of a contrat d’interface INSERM-CHU de Nice.Peer reviewe