Pan Chengshuang, Fei Qianjin, Jin Jianyuan, Zheng Jiujia, Wu Didi, Li Honggang, Huang Xuefeng, Kong Xiangbin
Department of Reproductive Medical Center, The First Affiliated Hospital of Wenzhou Medical University, Wenzhou, 325000, Zhejiang, China.
Duolaimi Biotechnology (Wuhan) Co., Ltd, Wuhan, 430000, China.
In Vitro Cell Dev Biol Anim. 2023 Jan;59(1):63-75. doi: 10.1007/s11626-022-00745-z. Epub 2023 Jan 30.
Late-onset hypogonadism (LOH) is an age-related clinical and biological syndrome in which serum testosterone deficiency is an important characteristic and diagnostic indicator. In this study, we firstly analyzed the difference in the expression level of three miR-133 s (including miR-133a-3p, miR-133a-5p, and miR-133b-3p) in rat testis samples, blood samples from mice before and 1 wk after testis removal, and mouse TM3 cells. Secondly, the mimics and inhibitors corresponding to the three miR-133 s of mouse were transfected into TM3 cells separately to determine the correlation between the three miRNAs. Finally, using mouse TM3 cells to analyze the effect of miR-133b overexpression or inhibition on the proliferation and apoptosis of mouse testicular Leydig cells, the effect on genes related to testosterone synthesis, and the effect on the level of testosterone in the culture medium. We found that, compared with the testis tissue of newborn rats, miR-133a-5p was increased in adult rats, and miR-133a-3p and miR-133b-3p were decreased. In addition, 1 wk after the testis was removed, the expression levels of these three miRNAs in the blood of adult mice decreased. The correlation of the three miRNAs was summarized, and it was found that miR-133b-3p played an important role in it. In TM3 cells, overexpression of miR-133b-3p suppressed the proliferation and promotes apoptosis of cells, suppressed the expression level of most genes related to cell proliferation and testosterone synthesis, and the concentration of testosterone in the culture medium decreased while these phenomena can be reversed by the inhibition of miR-133b-3p expression. It was found that miR-133b-3p can regulate testosterone production in TM3 cells at least by targeting FSCN1. The above results suggest that miR-133b-3p plays an important role in regulating testosterone synthesis. These findings also provide new candidate diagnostic indicators for late-onset hypogonadism in men and provide new clues for the further study of pathogenesis.
迟发性性腺功能减退（LOH）是一种与年龄相关的临床和生物学综合征，其中血清睾酮缺乏是一个重要特征和诊断指标。在本研究中，我们首先分析了三种miR-133（包括miR-133a-3p、miR-133a-5p和miR-133b-3p）在大鼠睾丸样本、睾丸切除前及切除后1周的小鼠血液样本以及小鼠TM3细胞中的表达水平差异。其次，将小鼠的三种miR-133对应的模拟物和抑制剂分别转染到TM3细胞中，以确定这三种微小RNA之间的相关性。最后，利用小鼠TM3细胞分析miR-133b过表达或抑制对小鼠睾丸间质细胞增殖和凋亡的影响、对睾酮合成相关基因的影响以及对培养基中睾酮水平的影响。我们发现，与新生大鼠的睾丸组织相比，成年大鼠中miR-133a-5p升高，而miR-133a-3p和miR-133b-3p降低。此外，睾丸切除后1周，成年小鼠血液中这三种微小RNA的表达水平下降。总结了这三种微小RNA的相关性，发现miR-133b-3p在其中起重要作用。在TM3细胞中，miR-133b-3p的过表达抑制细胞增殖并促进细胞凋亡，抑制大多数与细胞增殖和睾酮合成相关基因的表达水平，培养基中睾酮浓度降低，而抑制miR-133b-3p的表达可逆转这些现象。研究发现，miR-133b-3p至少可通过靶向FSCN1来调节TM3细胞中的睾酮生成。上述结果表明，miR-133b-3p在调节睾酮合成中起重要作用。这些发现也为男性迟发性性腺功能减退提供了新的候选诊断指标，并为进一步研究发病机制提供了新线索。
