Biomedical subjects

Tannia Gracia

Publications and source records attributed to Tannia Gracia.

4 recordsLinked to original sources

PubMed2006

The H295R system for evaluation of endocrine-disrupting effects.

The present studies were undertaken to evaluate the utility of the H295R system as an in vitro assay to assess the potential of chemicals to modulate steroidogenesis. The effects of four model chemicals on the expression of ten steroidogenic genes and on the production of three steroid hormones were examined. Exposures with individual model chemicals as well as binary mixtures were conducted. Although the responses reflect the known mode of action of the various compounds, the results show that designating a chemical as "specific inducer or inhibitor" is unwise. Not all changes in the mixture exposures could be predicted based on results from individual chemical exposures. Hormone production was not always directly related to gene expression. The H295R system integrates the effects of direct-acting hormone agonists and antagonists as well as chemicals affecting signal transduction pathways for steroid production and provides data on both gene expression and hormone secretion which makes this cell line a valuable tool to examine effects of chemicals on steroidogenesis.

Aminoglutethimide↗

PubMed2006

Alteration of steroidogenesis in H295R cells by organic sediment contaminants and relationships to other endocrine disrupting effects.

A novel bioassay with the human adrenocortical carcinoma cell line H295R can be used to screen for endocrine disrupting chemicals that affect the expression of genes important in steroidogenesis. This assay was employed to study the effects of organic contaminants associated with the freshwater pond sediments collected in the Ostrava-Karvina region, Czech Republic. The modulation of ten major genes involved in the synthesis of steroid hormones (CYP11A, CYP11B2, CYP17, CYP19, 17betaHSD1, 17betaHSD4, CYP21, 3betaHSD2, HMGR, StAR) after exposure of H295R cells to sediment extracts was investigated using quantitative real-time polymerase chain reaction (PCR). Crude sediment extracts, containing high concentrations of polycyclic aromatic hydrocarbons (PAHs) and moderate amounts of polychlorinated biphenyls (PCBs) and organochlorine pesticides (OCPs) significantly stimulated expression of the CYP11B2 gene (up to 10-fold induction), and suppressed expression of 3betaHSD2 and CYP21 genes. A similar pattern was observed with the extracts after treatment with concentrated sulfuric acid to remove labile chemicals (including PAHs) leaving only persistent PCBs, OCPs and potentially PCDD/Fs. Comparison of the results with other mechanistically based bioassays (arylhydrocarbon receptor, AhR, mediated responses in H4IIE-luc cells, and estrogen receptor mediated effects in MVLN cells) revealed significant endocrine disrupting potencies of organic contaminants present in the sediments (most likely antiestrogenicity). Pronounced effects were observed particularly in sediment extracts from the Pilnok Pond which harbors an unusual intersexual population of the narrow-cawed crayfish Pontastacus leptodactylus (Decapoda, Crustacea). This pilot study provided the first experimental evidence of the wider application of the H295R bioassay for screening complex environmental samples, and the results support the hypothesis of chemical-induced endocrine disruption in intersexual crayfish.

Cell Line, Tumor↗

PubMed2005

Quantitative RT-PCR methods for evaluating toxicant-induced effects on steroidogenesis using the H295R cell line.

Gene expression profiles show considerable promise for the evaluation of the toxic potential of environmental contaminants. For example, any alterations in the pathways of steroid synthesis or breakdown have the potential to Cause endocrine disruption. Therefore monitoring these pathways can provide information relative to a chemical's ability to impact endocrine function. One approach to monitoring these pathways has been to use a human adrenocortical carcinoma cell line (H295R) that expresses all the key enzymes necessary for steroidogenesis. In this study we have further developed these methods using accurate and specific quantification methods utilizing molecular beacon-based quantitative RT-PCR (Q-RT-PCR). The assay system was used to analyze the expression patterns of 11 steroidogenic genes in H295R cells. The expression of gene transcripts was measured using a real-time PCR system and quantified based on both a standard curve method using a dilution series of RNA standards and a comparative Ct method. To validate the optimized method, cells were exposed to specific and nonspecific model compounds (inducers and inhibitors of various steroidogenic enzymes) for gene expression profiling. Similar gene expression profiles were exhibited by cells treated with chemicals acting through common mechanisms of action. Overall, our findings demonstrated that the present assay can facilitate the development of compound-specific response profiles, and will provide a sensitive and integrative screen for the effects of chemicals on steroidogenesis.

Cell Line, Tumor↗

PubMed2004

Assessment of the effects of chemicals on the expression of ten steroidogenic genes in the H295R cell line using real-time PCR.

The potential for a variety of environmental contaminants to disturb endocrine function in wildlife and humans has been of recent concern. While much effort is being focused on the assessment of effects mediated through steroid hormone receptor-based mechanisms, there are potentially several other mechanisms that could lead to endocrine disruption. Recent studies have demonstrated that a variety of xenobiotics can alter the gene expression or activity of enzymes involved in steroidogenesis. By altering the production or catalytic activity of steroidogenic or steroid-catabolizing enzymes, these chemicals have the potential to alter the steroid balance in organisms. To assess the potential of chemicals to alter steroidogenesis, an assay system was developed using a human adrenocortical carcinoma cell line, the H295R cell line, which retains the ability to synthesize most of the important steroidogenic enzymes. Methods were developed, optimized, and validated to measure the expression of 10 genes involved in steroidogenesis by the use of real-time quantitative reverse transcriptase PCR. The effects of several model chemicals known to alter steroid metabolism, both inducers and inhibitors, were assessed. Similar expression patterns were observed for chemicals acting through common mechanisms of action. Time-course studies demonstrated distinct time-dependent expression profiles for chemicals able to modulate steroid metabolism. The assay, which allows simultaneous analysis of the expression of numerous steroidogenic enzymes, would be useful as a sensitive and integrative screen for the many effects of chemicals on steroidogenesis.

Adrenal Cortex↗