Hyperhomocysteinemia (HHcy) plays a salient role in male infertility. However, whether HHcy interferes with testosterone production remains inconclusive. Here, we reported a lower serum testosterone level in HHcy mice. Single-cell RNA sequencing revealed that genes related to testosterone biosynthesis, together with nuclear receptor subfamily 5 group A member 1 (Nr5a1), a key transcription factor for steroidogenic genes, were downregulated in the Leydig cells (LCs) of HHcy mice. Mechanistically, Hcy lowered trimethylation of histone H3 on lysine 4 (H3K4me3), which was bound on the promoter region of Nr5a1, resulting in downregulation of Nr5a1. Intriguingly, we identified an unknown cell cluster annotated as Macrophage-like Leydig cells (McLCs), expressing both LCs and macrophages markers. In HHcy mice, McLCs were shifted toward pro-inflammatory phenotype and thus promoted inflammatory response in LC. Betaine supplementation rescued the downregulation of NR5A1 and restored the serum testosterone level in HHcy mice. Overall, our study highlights an etiological role of HHcy in LCs dysfunction.

Graphical Abstract

中文翻译：

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高同型半胱氨酸血症通过损害间质细胞中睾酮的产生来降低血清睾酮浓度

高同型半胱氨酸血症（HHcy）在男性不育中起着重要作用。然而，HHcy 是否会干扰睾酮的产生仍无定论。在这里，我们报告了 HHcy 小鼠血清睾酮水平较低。单细胞 RNA 测序显示，与睾酮生物合成相关的基因以及核受体亚家族 5 A 组成员 1 ( Nr5a1 )（类固醇生成基因的关键转录因子）在 HHcy 小鼠的 Leydig 细胞 (LC) 中下调。从机制上讲，Hcy 降低了组蛋白 H3 在赖氨酸 4 (H3K4me3) 上的三甲基化，赖氨酸 4 (H3K4me3) 结合在Nr5a1的启动子区域，导致Nr5a1的下调。有趣的是，我们发现了一个未知的细胞簇，被注释为巨噬细胞样间质细胞（McLC），表达 LC 和巨噬细胞标记物。在 HHcy 小鼠中，McLC 转向促炎表型，从而促进 LC 的炎症反应。补充甜菜碱可以挽救 HHcy 小鼠中 NR5A1 的下调并恢复血清睾酮水平。总体而言，我们的研究强调了 HHcy 在 LC 功能障碍中的病因学作用。

图形概要