附睾钙稳态与载脂蛋白的相互作用对调控精子成熟微环境的功能研究

附睾微环境是精子成熟的重要场所，对精子功能和雄性生育力至关重要。附睾功能通过上皮细胞建立管腔微环境让精子成熟。进年来，特发性男性不育比例呈上升趋势，但人们对其潜在的致病因素却知之甚少。已有证据表明附睾功能与特发性男性不育息息相关。申请人之前的研究表明附睾上皮细胞形成了错综复杂的通讯网络为精子成熟维持适宜生理酸性低钙微环境，但其具体功能作用机制尚不完全清楚。申请人先前研究揭示钙稳态依赖性TRPV6与TMEM16A电偶联调控附睾尾部微环境，也发现了在附睾体部的维生素K2依赖GGCX羧基化MGP通路对管腔中低钙浓度起着重要作用，并发现羧基化的MGP与附睾特异表达的载脂蛋白互作，但其生理功能以及具体分子机制尚不完全清楚。本课题因此计划深入研究MGP结合蛋白复合体与与载脂LCN家族蛋白互作的功能作用，旨在阐明附睾低钙稳态微环境的分子机制及其对精子成熟的影响，并提供附睾精子成熟的新理论模型。

The production of high quality sperm with normal structure and function is the benchmark of male fertility. This is achieved by normal spermatogenesis in the testis and functional maturation in the epididymis. Epididymis establishes a unique luminal microenvironment for the immature testicular sperm to undergo a dynamic but tightly-controlled sequential changes, to let them become functionally competent – the ability of fertilizing an oocyte. For instance, the epididymal fluid is acidic and progressively low in calcium concentration down the epididymal tubule. Our previous studies have demonstrated an intriguing intercellular communication network underlying the barrier function of epididymal epithelium. Among these elaborate modulated microenvironmental factors, Ca2+ is known to play a critical role for spermatozoa to undergo fertilization activities such as motility and capacitation – the abilities of spermatozoa obtained after transiting through the epididymis. It has been reported that an aberrant high luminal Ca2+ level results in premature spermatozoa activation followed with reduced sperm motility and impaired fertility without affecting spermatogenesis, however, the underlying functional mechanism is still largely unclear. Recently, we have also reported that the epithelial calcium channel TRPV6 can electrically couple with the calcium-dependent chloride channel TMEM16A in principal cells of specific regions of epididymis. This study is not only consistent with the putative role of TRPV6 in calcium homeostatic regulation and male fertility, but also introduces a new TRPV6-TMEM16A coupled chloride-driven secretion mechanism in the epididymis, which is a pH-sensitive calcium-dependent and electrically regulatory process. In addition, we also identified the vitamin-K2 dependent (VK2D) GGCX carboxylation of MGP is an additional pathway in the caput and corpus regions for calcium homeostatic regulation where the TRPV6-TMEM16A coupled complex is absent. Interestingly, our results showed that carboxylated MGP binds with lipocalins (LCNs), whose expression is particularly high in the epididymis. The goal of study is therefore aimed to elucidate the functional mechanism in the interplay of carboxylated MGP and its interacting LCNs, particularly epididymal specific LCNs, in calcium homeostatic regulation and function in the epididymis. The ultimate goal of this study is to elucidate the mechanisms of epididymal luminal environment for sperm maturation under normal physiological conditions, and to provide sights to better understand the pathophysiology behind the more and more concerned “idiopathic” male infertility cases in the clinics.