Internal Medicine 4 (L.L., B.C., A.G.C., T.M.S., M.K., S.F., G.S., G.P.R.) and Surgical Pathology and Cytopathology Unit (A.F.), Department of Medicine, Department of Medicine, Section of Human Anatomy and Physiology (A.S.B.), Department of Molecular Medicine, University of Padua, 35128 Padua, Italy; Max-Delbrück-Center for Molecular Medicine (M.B.), 13092 Berlin, Germany; and Department of Molecular and Integrative Physiology (W.E.R.), University of Michigan, Ann Arbor, Michigan 48109.
J Clin Endocrinol Metab. 2014 Apr;99(4):E674-82. doi: 10.1210/jc.2013-2900. Epub 2013 Nov 27.
The molecular mechanisms of primary aldosteronism, a common cause of human hypertension, are unknown, but alterations of K(+) channels can play a key role.
The objective of the study was to investigate the following: 1) the expression of the Twik-related acid-sensitive K(+) channels (TASK) in aldosterone producing adenomas (APAs); 2) the role of TASK-2 in aldosterone synthesis; and 3) the determinants of TASK-2-blunted expression in APAs.
We analyzed the transcriptome and the microRNA profiles of 32 consecutive APAs and investigated the protein expression and localization of TASK-2 in APA and adrenocortical cell lines (H295R and HAC15) using immunoblotting and confocal microscopy. The functional effect of TASK-2 blunted activity caused by a dominant-negative mutation on steroidogenic enzymes, and aldosterone production was also assessed. TASK-2 regulation by selected microRNA was studied by a luciferase assay.
TASK-2 was consistently less expressed at the transcript and protein levels in APAs than in the normal human adrenal cortex. H295R cell transfection with a TASK-2 dominant-negative mutant construct significantly increased the aldosterone production by 153% and the gene expression of aldosterone synthase (CYP11B2, gene expression fold change 3.1 vs control, P < .05) and the steroidogenic acute regulatory protein (gene expression fold change 1.8 vs control, P < .05). Two microRNAs, hsa-miR-23 and hsa-miR-34, were found to decrease the TASK-2 expression by binding to the 3' untranslated region of the TASK-2 gene.
The TASK-2 channel lower expression represents a hallmark of APA and is associated with a higher expression of hsa-miR-23 and hsa-miR-34. The ensuing blunted TASK-2 activity increased the production of aldosterone in vitro and the expression of steroidogenic acute regulatory protein and CYP11B2. Hence, the lower expression of TASK-2 channel in APA cells can explain high aldosterone secretion in human primary aldosteronism despite the suppression of angiotensin II, hypertension, and hypokalemia.
原发性醛固酮增多症是人类高血压的常见病因，其分子机制尚不清楚，但 K+通道的改变可能起关键作用。
本研究旨在探讨以下问题：1）醛固酮分泌瘤（APA）中 Twik 相关酸敏感钾通道（TASK）的表达；2）TASK-2 在醛固酮合成中的作用；3）APA 中 TASK-2 表达减弱的决定因素。
我们分析了 32 例连续 APA 的转录组和 microRNA 谱，并使用免疫印迹和共聚焦显微镜研究 TASK-2 在 APA 和肾上腺皮质细胞系（H295R 和 HAC15）中的蛋白表达和定位。还评估了显性负突变引起的 TASK-2 活性减弱对类固醇生成酶和醛固酮产生的功能影响。通过荧光素酶测定研究了 TASK-2 受选定 microRNA 的调节。
TASK-2 在 APA 中的转录和蛋白水平均明显低于正常人肾上腺皮质。用 TASK-2 显性负突变体构建体转染 H295R 细胞，可使醛固酮产生增加 153%，醛固酮合酶（CYP11B2，基因表达倍数变化 3.1 与对照相比，P <.05）和类固醇急性调节蛋白（基因表达倍数变化 1.8 与对照相比，P <.05）的基因表达增加。发现 2 种 microRNA，hsa-miR-23 和 hsa-miR-34，通过结合 TASK-2 基因的 3'非翻译区来降低 TASK-2 的表达。
TASK-2 通道的低表达代表 APA 的一个标志，与 hsa-miR-23 和 hsa-miR-34 的表达较高相关。随之而来的 TASK-2 活性减弱增加了体外醛固酮的产生以及类固醇急性调节蛋白和 CYP11B2 的表达。因此，APA 细胞中 TASK-2 通道的低表达可以解释尽管抑制血管紧张素 II、高血压和低血钾，但人类原发性醛固酮增多症仍有高醛固酮分泌。
