Department of General Medicine, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama 700-8558, Japan.
Endocr J. 2021 Feb 28;68(2):243-250. doi: 10.1507/endocrj.EJ20-0359. Epub 2020 Oct 6.
Functional interactions between the levels of clock gene expression and adrenal steroidogenesis were studied in human adrenocortical H295R cells. Fluctuations of Bmal1, Clock, Per2 and Cry1 mRNA levels were found in H295R cells treated with forskolin (FSK) in a serum-free condition. The changes of clock gene expression levels were diverged, with Clock mRNA level being significantly higher than Cry1 and Per2 mRNA levels after 12-h stimulation with FSK. After FSK induction, mRNA levels of StAR and CYP11B2 were highest at 12 hours and CYP17 mRNA level reached a peak at 6 hours, but HSD3B1 mRNA level was transiently decreased at 3 hours. The expression levels of Clock mRNA showed a significant positive correlation with StAR among the interrelationships between mRNA levels of key steroidogenic factors and clock genes. Knockdown of Clock gene by siRNA led to a significant reduction of FSK-induced expression of StAR and CYP17 after 12-h treatment with FSK. BMP-6 and activin, which modulate adrenal steroidogenesis, had inhibitory effects on Clock mRNA expression, whereas treatment with follistatin, a binding protein of activin, increased Clock mRNA levels in the presence of FSK, suggesting an endogenous function of activin in regulation of Clock mRNA expression. Collectively, the results indicated that changes of Clock mRNA expression, being upregulated by FSK and suppressed by BMP-6 and activin, were tightly linked to StAR expression by human adrenocortical cells.
研究了生物钟基因表达水平与肾上腺类固醇生成之间的功能相互作用在人类肾上腺皮质 H295R 细胞中。在无血清条件下用 forskolin (FSK) 处理 H295R 细胞时，发现 Bmal1、Clock、Per2 和 Cry1 mRNA 水平波动。时钟基因表达水平的变化存在差异，FSK 刺激 12 小时后，Clock mRNA 水平明显高于 Cry1 和 Per2 mRNA 水平。在 FSK 诱导后，StAR 和 CYP11B2 的 mRNA 水平在 12 小时时最高，CYP17 mRNA 水平在 6 小时时达到峰值，但 HSD3B1 mRNA 水平在 3 小时时短暂下降。Clock mRNA 表达水平与关键类固醇生成因子和时钟基因之间的 mRNA 水平的相互关系中与 StAR 呈显著正相关。用 siRNA 敲低 Clock 基因后，用 FSK 处理 12 小时后，FSK 诱导的 StAR 和 CYP17 的表达显著减少。调节肾上腺类固醇生成的 BMP-6 和激活素对 Clock mRNA 表达有抑制作用，而激活素的结合蛋白 follistatin 的处理在 FSK 存在的情况下增加了 Clock mRNA 水平，提示激活素在调节 Clock mRNA 表达中的内源性作用。总之，这些结果表明，Clock mRNA 表达的变化被 FSK 上调，被 BMP-6 和激活素抑制，与人类肾上腺皮质细胞的 StAR 表达密切相关。
