Hao Jianheng, Ren Jia, Chang Boya, Xu Huichao, Wang Haijun, Ji Laixi
College of Acupuncture and Massage, Chengdu University of Traditional Chinese Medicine, Chengdu, 610075, China.
The Second Clinical College, Shanxi University of Traditional Chinese Medicine, Jinzhong, 030619, China.
Heliyon. 2024 Aug 22;10(17):e36664. doi: 10.1016/j.heliyon.2024.e36664. eCollection 2024 Sep 15.
Acupuncture is an integral component of complementary and alternative medicine that has been reported to enhance sperm motility, improve semen quality, and consequently augment male fertility. However, the precise mechanisms of action and the underlying molecular pathways remain unclear. In the present study, we aimed to elucidate the potential mechanisms through which acupuncture improves reproductive function in a mouse model of cyclophosphamide-induced asthenozoospermia. We collected sperm from the epididymis for semen analysis, collected serum to determine gonadotropin and oxidative stress marker levels, conducted histological examination of testicular tissue using hematoxylin and eosin (HE) and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining, and observed mitochondrial morphology using transmission electron microscopy (TEM). We also assessed oxidative stress levels and total iron content in testicular tissue and validated the proteomic and transcriptomic analysis results of testicular tissue using real-time reverse transcription-quantitative polymerase chain reaction (RT-qPCR), protein imprinting analysis, and immunohistochemistry (IHC). Our results indicate that acupuncture enhances sperm quality in asthenozoospermic mice; increases serum testosterone (T), follicle-stimulating hormone (FSH), and luteinizing hormone (LH) levels; and attenuates oxidative damage, iron accumulation, and mitochondrial injury in mouse testicular tissues. Through protein and transcriptomic analyses, we identified 21 key genes, of which cytochrome -245 heavy chain (CYBB), glutathione peroxidase 4 (GPX4), acyl-CoA synthetase long-chain family member 1 (ACSL1), and ferritin mitochondria (FTMT) were closely associated with ferroptosis. RT-qPCR, protein imprinting, and immunofluorescence (IF) analyses collectively indicated that acupuncture reduced ACSL1 and CYBB expression, and increased GPX4 and FTMT expression. Overall, the ferroptosis pathway associated with ACSL1/CYBB/FTMT/GPX4 represents a potential strategy through which acupuncture can improve the reproductive function in asthenozoospermic mice.
针灸是补充和替代医学的一个重要组成部分，据报道它能增强精子活力、改善精液质量，从而提高男性生育能力。然而，其确切的作用机制和潜在的分子途径仍不清楚。在本研究中，我们旨在阐明针灸改善环磷酰胺诱导的弱精子症小鼠模型生殖功能的潜在机制。我们从附睾收集精子进行精液分析，收集血清以测定促性腺激素和氧化应激标志物水平，使用苏木精和伊红（HE）以及末端脱氧核苷酸转移酶dUTP缺口末端标记（TUNEL）染色对睾丸组织进行组织学检查，并使用透射电子显微镜（TEM）观察线粒体形态。我们还评估了睾丸组织中的氧化应激水平和总铁含量，并使用实时逆转录定量聚合酶链反应（RT-qPCR）、蛋白质印迹分析和免疫组织化学（IHC）验证了睾丸组织的蛋白质组学和转录组学分析结果。我们的结果表明，针灸可提高弱精子症小鼠的精子质量；增加血清睾酮（T）、促卵泡激素（FSH）和促黄体生成素（LH）水平；并减轻小鼠睾丸组织中的氧化损伤、铁积累和线粒体损伤。通过蛋白质和转录组分析，我们鉴定出21个关键基因，其中细胞色素-245重链（CYBB）、谷胱甘肽过氧化物酶4（GPX4）、酰基辅酶A合成酶长链家族成员1（ACSL1）和线粒体铁蛋白（FTMT）与铁死亡密切相关。RT-qPCR、蛋白质印迹和免疫荧光（IF）分析共同表明，针灸降低了ACSL1和CYBB的表达，并增加了GPX4和FTMT的表达。总体而言，与ACSL1/CYBB/FTMT/GPX4相关的铁死亡途径代表了针灸改善弱精子症小鼠生殖功能的一种潜在策略。
