Zhou B, Zhang C, Deng H, Chen S, Chang Y, Yang Y, Fu G, Yuan D, Zhao H
Third-grade Pharmacological Laboratory on Traditional Chinese Medicine of State Administration of Traditional Chinese Medicine, China Three Gorges University, Yichang 443002, China.
College of Basic Medical Science, China Three Gorges University, Yichang 443002, China.
Nan Fang Yi Ke Da Xue Xue Bao. 2023 Jul 20;43(7):1145-1154. doi: 10.12122/j.issn.1673-4254.2023.07.11.
To investigate the protective effects of total saponins from (TSPJ) against high-fat dietinduced testicular Sertoli cell junction damage in mice.
Forty male C57BL/6J mice were randomized into normal diet group, high-fat diet group, and low-dose (25 mg/kg) and high-dose (75 mg/kg) TSPJ treatment groups (=10). The mice in the normal diet group were fed a normal diet, while the mice in the other groups were fed a high-fat diet. After TSPJ treatment via intragastric administration for 5 months, the testes and epididymis of the mice were collected for measurement of weight, testicular and epididymal indices and sperm parameters. HE staining was used for histological evaluation of the testicular tissues and measurement of seminiferous tubule diameter and seminiferous epithelium height. The expression levels of ZO-1, occludin, claudin11, N-cadherin, E-cadherin and β-catenin in Sertoli cells were detected with Western blot, and the localization and expression levels of ZO-1 and β-catenin in the testicular tissues were detected with immunofluorescence assay. The protein expressions of LC3B, p-AKT and p-mTOR in testicular Sertoli cells were detected using double immunofluorescence assay.
Treatment with TSPJ significantly improved high-fat diet-induced testicular dysfunction by reducing body weight ( < 0.001), increasing testicular and epididymal indices ( < 0.05), and improving sperm concentration and sperm viability ( < 0.05). TSPJ ameliorated testicular pathologies and increased seminiferous epithelium height of the mice with high-fat diet feeding ( < 0.05) without affecting the seminiferous tubule diameter. TSPJ significantly increased the expression levels of ZO-1, occludin, N-cadherin, E-cadherin and β-catenin ( < 0.05) but did not affect claudin11 expression in the testicular tissues. Immunofluorescence assay showed that TSPJ significantly increased ZO-1 and β-catenin expression in the testicular tissues ( < 0.001), downregulated LC3B expression and upregulated p-AKT and p-mTOR expressions in testicular Sertoli cells.
TSPJ alleviates high-fat diet-induced damages of testicular Sertoli cell junctions and spermatogenesis possibly by activating the AKT/mTOR signaling pathway and inhibiting autophagy of testicular Sertoli cells.
探讨绞股蓝总皂苷（TSPJ）对高脂饮食诱导的小鼠睾丸支持细胞连接损伤的保护作用。
将40只雄性C57BL/6J小鼠随机分为正常饮食组、高脂饮食组、低剂量（25 mg/kg）和高剂量（75 mg/kg）TSPJ治疗组（每组n = 10）。正常饮食组小鼠给予正常饮食，其他组小鼠给予高脂饮食。经灌胃给予TSPJ治疗5个月后，收集小鼠的睾丸和附睾，测量其重量、睾丸和附睾指数以及精子参数。采用苏木精-伊红（HE）染色对睾丸组织进行组织学评估，并测量生精小管直径和生精上皮高度。用蛋白质免疫印迹法检测支持细胞中紧密连接蛋白1（ZO-1）、闭合蛋白（occludin）、紧密连接蛋白11（claudin11）、N-钙黏蛋白、E-钙黏蛋白和β-连环蛋白的表达水平，用免疫荧光法检测睾丸组织中ZO-1和β-连环蛋白的定位及表达水平。采用双重免疫荧光法检测睾丸支持细胞中微管相关蛋白1轻链3β（LC3B）、磷酸化蛋白激酶B（p-AKT）和磷酸化哺乳动物雷帕霉素靶蛋白（p-mTOR）的蛋白表达。
TSPJ治疗可显著改善高脂饮食诱导的睾丸功能障碍，表现为降低体重（P < 0.001）、增加睾丸和附睾指数（P < 0.05）以及提高精子浓度和精子活力（P < 0.05）。TSPJ改善了高脂饮食喂养小鼠的睾丸病理状况并增加了生精上皮高度（P < 0.05），但不影响生精小管直径。TSPJ显著增加了睾丸组织中ZO-1、occludin、N-钙黏蛋白、E-钙黏蛋白和β-连环蛋白的表达水平（P < 0.05），但不影响claudin11的表达。免疫荧光分析显示，TSPJ显著增加了睾丸组织中ZO-1和β-连环蛋白的表达（P < 0.001），下调了睾丸支持细胞中LC3B的表达，并上调了p-AKT和p-mTOR的表达。
TSPJ可能通过激活AKT/mTOR信号通路并抑制睾丸支持细胞的自噬，减轻高脂饮食诱导的睾丸支持细胞连接损伤和精子发生损伤。
