H295R-cells as a model system for detection of toxic effects on the adrenal aldosterone synthesis pathways
Abstract: The adrenal gland is exposed to high concentrations of circulating xenobiotics due to its high rate of blood flow and it may also accumulate lipid soluble chemicals in its lipid rich tissue. These substances can affect the aldosterone synthesis in the glomerulosa cells by activation or suppression of the steroidogenic enzymes' gene expression, hence resulting in effects on blood pressure since this is the main action of aldosterone. The purpose of this project was to set up a model system for detection of xenobiotic effects on aldosterone synthesis due to changes in gene expression of involved enzymes and transporter proteins. The human adrenocarcionoma cell line H295R was subjected to angiotensin II (ang II) and potassium acetate (KAc), in order to establish which modulator that most efficiently differentiates the cells into glomerulosa like aldosterone secreting cells. Further the cells were tested for differences in differentiation due to various time intervals and for Ultroser SF (USF) supplementation in the medium. Leads effect on the involved genes in aldosterone formation was investigated, since lead is a known activator of aldosterone secretion. The results indicated that ang II was the best modulator for differentiating the cells, KAc had cytotoxic effects at higher concentrations. The results also indicated that USF supplementation in the medium had a rising effect on basal gene transcription levels of the steroidogenic enzymes, some genes differed as much as 70-fold in expression levels between the two medium types. Non-USF-supplemented medium was therefore chosen for exposure experiments, since the chemical effect hence became clearer. The lead exposures indicated that this substance did not affect the gene expression level of the investigated genes, except for small effects in medium without USF and ang II. Some other differences in gene expression were noted between the control and the samples, but they were very small even if they were statistically significant. The model will need more testing with other substances and the aldosterone level during chemical exposure will need to be determined. We have so far established that ang II is an efficient stimulator of glomerulosa like differentiation and that non-USF-supplemented medium is favorable in the experiments.
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