基于支持细胞Toll样信号通路对不育症“久病入络”机制的研究

Testes subtle environment of idiopathic semen abnormalities have become one of the hotspot in recent years. It has been confirmed that the cytokine generated by Sertoli cell-mediated could induce second-injury to spermatogenic cells, and further to spermatogenesis. Deficiency of the kidney as the basic pathogenesis in Traditional Chinese Medicine(TCM),and mainly tonify the kidney as its treatment. Damage of blood-testis barrier induced disturbances of spermatogenic environmental will cause “qi and blood stasis” in seminiferous tubules, as TCM said “collaterals invaded and stasis by prolonged illness”. The therapy of invigorate the kidney and promote blood circulation has achieved satisfactory results. Based on this therapy we made Qiangjing pills, which has showed satisfactory results in clinical and significant improvement in sperm quality, recovery after epithelial damagement. We take sertoli cells and Toll-like signal pathway as the starting point, with Qiangjing pills interventing mice with weak spermatogenesis and Sertoli cells in vitro culture, analysing Sertoli cell pathology, the changes of cytokines, and Toll-like signal pathway MYD88, TRAF6 change, TAK1, NF-κB (AP1) by Western Blot, Qpcr and other methods, explore the mechanism of "collaterals invaded by prolonged illness", and revealing the regulative role of Chinese medicine in testes subtle environment.

睾丸生精微环境变化已成为特发性精液异常研究热点之一,已证实微环境中由支持细胞介导产生的细胞因子能对生精细胞形成二次损伤,从而进一步损伤生精功能。中医学以肾虚为基本病机,传统治疗以补肾为主,现代研究显示,血睾屏障破坏引起生精微环境紊乱导致生精小管内“气血瘀滞”,与“久病入络、久病成瘀”理论相一致,运用补肾活血法治疗本病亦取得满意疗效。以此法筛选出药物(强精片),前期临床效果满意,基础研究亦证明能明显改善精子质量,促进损伤上皮生理功能恢复。本研究以支持细胞及Toll样信号通路为切入点,用强精片对弱生精环境下小鼠及体外支持细胞培养株进行干预,用Western Blot、Qpcr等法,分析支持细胞病理、细胞因子改变及Toll样信号通路中MYD88、TRAF6、TAK1、NF-κB(AP1)等指标的变化,探讨不育症“久病入络”的理论机制,揭示补肾活血中药对睾丸生精微环境的调节作用 。

睾丸生精微环境变化已成为特发性精液异常研究热点之一,已证实微环境中由支持细胞介导产生的细胞因子能对生精细胞形成二次损伤,从而进一步损伤生精功能。中医学以肾虚为基本病机,传统治疗以补肾为主,现代研究显示,血睾屏障破坏引起生精微环境紊乱导致生精小管内“气血瘀滞”,与“久病入络、久病成瘀”理论相一致,运用补肾活血法治疗本病亦取得满意疗效。以此法筛选出药物(强精片),前期临床效果满意,基础研究亦证明能明显改善精子质量,促进损伤上皮生理功能恢复。本研究以支持细胞及Toll样信号通路为切入点,用强精片对弱生精环境下小鼠及体外支持细胞培养株进行干预,用Western Blot、Q-pcr等法,分析支持细胞病理、细胞因子改变及Toll样信号通路中MYD88、TRAF6、TAK1、NF-κB(AP1)等指标的变化。研究发现强精片能改善少弱精子症大鼠的生精功能，增加精子数量，提高睾丸组织HE染色MTBS评分，提高LH、T水平，降低FSH、E2水平，降低TLR4、MYD88表达、升高TRAF6mRNA表达。强精片对Sertoli细胞相关炎性细胞因子IL-1β、TNF-ɑ的分泌具有抑制作用。强精片可抑制TLR4、TRAF6基因的表达，减少TLR4、MyD88和TRAF6蛋白的含量，抑制NF-κB的磷酸化和核转位，其对TLR4/MyD88/TRAF6/NF-κB信号通路的干预方式存在多样性。强精片治疗少弱精子症的作用机理与改善生精微环境的炎性状态密切相关，其可通过调节睾丸Sertoli细胞TLR4/MyD88/TRAF6/NF-κB信号通路降低炎性细胞因子的转录水平，发挥改善炎性微环境作用。综上所述，本研究探讨了不育症“久病入络”的理论机制,揭示补肾活血中药对睾丸生精微环境的调节作用。