From the Clinical Pharmacology Unit, Department of Medicine (C.M., S.G., J.Z.) and Medical Research Council Cancer Unit (T.I.J.), University of Cambridge, United Kingdom; Human Research Tissue Bank, Cambridge University Hospitals NHS Foundation Trust, Addenbrooke's Hospital, United Kingdom (W.Z.); Metabolic Research Laboratories, Wellcome Trust-MRC Institute of Metabolic Science, Cambridge, United Kingdom (M.G.); and the Centre for Clinical Pharmacology, William Harvey Research Institute, Barts and the London School of Medicine and Dentistry, Queen Mary University of London, United Kingdom (J.Z., M.J.B.).
Hypertension. 2017 Aug;70(2):357-364. doi: 10.1161/HYPERTENSIONAHA.117.09231. Epub 2017 Jun 5.
Heterogeneity among aldosterone-producing adenomas (APAs) has been highlighted by the discovery of somatic mutations. mutations predominate in large zona fasciculata (ZF)-like APAs; mutations in , , and are more likely to be found in small zona glomerulosa (ZG)-like APAs. Microarray comparison of mutant versus wild-type APAs revealed significant differences in transcriptomes. , encoding a neurofilament subunit which is a D1R (dopamine D1 receptor)-interacting protein, was 4-fold upregulated in ZG-like versus ZF-like APAs and 14-fold more highly expressed in normal ZG versus ZF. Immunohistochemistry confirmed selective expression of NEFM (neurofilament medium) polypeptide in ZG and in ZG-like APAs. Silencing in adrenocortical H295R cells increased basal aldosterone secretion and cell proliferation; silencing also amplified aldosterone stimulation by the D1R agonist, fenoldopam, and inhibition by the D1R antagonist, SCH23390. NEFM coimmunoprecipitated with D1R, and its expression was stimulated by fenoldopam. Immunohistochemistry for D1R was mainly intracellular in ZG-like APAs but membranous in ZF-like APAs. Aldosterone secretion in response to fenoldopam in primary cells from ZF-like APAs was higher than in cells from ZG-like APAs. Transfection of mutant KCNJ5 caused a large reduction in NEFM expression in H295R cells. We conclude that NEFM is a negative regulator of aldosterone production and cell proliferation, in part by facilitating D1R internalization from the plasma membrane. Downregulation of NEFM in ZF-like APAs may contribute to a D1R/D2R imbalance underlying variable pharmacological responses to dopaminergic drugs among patients with APAs. Finally, taken together, our data point to the possibility that ZF-like APAs are in fact ZG in origin.
醛固酮瘤（APAs）中的异质性已被发现的体细胞突变所强调。在大束状带（ZF）样 APA 中，突变占主导地位；在小球状带（ZG）样 APA 中，更可能发现 、 和 突变。对 突变型与野生型 APA 的微阵列比较显示转录组存在显著差异。编码神经丝亚单位的 ，是多巴胺 D1 受体（D1R）相互作用蛋白，在 ZG 样 APA 中比 ZF 样 APA 上调 4 倍，在正常 ZG 中比 ZF 上调 14 倍。免疫组织化学证实 NEFM（神经丝中间）多肽在 ZG 和 ZG 样 APA 中选择性表达。在肾上腺皮质 H295R 细胞中沉默 ，增加基础醛固酮分泌和细胞增殖；沉默还放大了 D1R 激动剂芬氟拉明和 D1R 拮抗剂 SCH23390 对醛固酮的刺激作用。NEFM 与 D1R 共免疫沉淀，其表达受芬氟拉明刺激。在 ZG 样 APA 中，D1R 的免疫组织化学主要为细胞内，而在 ZF 样 APA 中主要为膜性。来自 ZF 样 APA 的原代细胞对芬氟拉明的反应性醛固酮分泌高于来自 ZG 样 APA 的细胞。在 H295R 细胞中，转染突变型 KCNJ5 导致 NEFM 表达大量减少。我们得出结论，NEFM 是醛固酮产生和细胞增殖的负调节剂，部分通过促进 D1R 从质膜内化。在 ZF 样 APA 中下调 NEFM 可能导致 D1R/D2R 失衡，这是 APA 患者对多巴胺能药物的药理反应不同的原因之一。最后，综上所述，我们的数据表明，ZF 样 APA 实际上可能起源于 ZG。
