Approximately 10% of essential hypertension cases is attributed to primary aldosteronism (PA), where inappropriately high levels of aldosterone are secreted. Almost half of these PA cases result from aldosterone-producing adenoma. microRNAs are single-stranded, short non-coding RNAs that negatively regulate gene expression post-transcriptionally. In a previous study, we showed that microRNAs directly modulate CYP11B2 (aldosterone synthase) gene expression and aldosterone levels. We also compared microRNA profiles of non-diseased adrenal tissue with those of aldosterone-producing adenoma (APA; each n=4). Now we have examined microRNA profiles from the widely-used adrenocortical carcinoma cell line, H295R, in both its basal state and following stimulation of aldosterone production for 24 hours with either 100 nM angiotensin II (AngII), 1 mM dibutyryl cyclic AMP (dbcAMP) or 20 mM potassium chloride (KCl; n=3 per cell group). The microRNA profiles of all cells and tissues were generated by microarray. CYP11B2 upregulation in stimulated H295R cells was confirmed by qRT-PCR.
We found five microRNAs to be consistently downregulated in APA relative to NA and in all stimulated H295R cells relative to basal. This is suggestive of common mechanisms underlying the abnormal secretion of aldosterone observed in APA as well as the normal physiological stimulation of aldosterone production in response to regulatory molecules. Furthermore, it implicates these microRNAs in the regulation of such mechanisms. Initial bioinformatic analysis was performed using Ingenuity Pathway Analysis (IPA) software in order to predict relevant mRNAs likely to be targeted by one or more of these 5 microRNAs. A predicted target genes of particular relevance to steroidogenesis is HMGCR, which encodes the rate-limiting enzyme for cholesterol biosynthesis targeted by statin treatment. As regulation of cholesterol supply is increasingly recognised as a determining factor in steroid output future study will focus on this interaction.
In conclusion, we suggest that these consistently altered microRNAs are likely to be involved in aldosterone regulation and have presented relevant putative targets, which are worthy of further investigation.