CNP/cGMP信号通路调节精子成熟和运动的分子机制研究

Epididymal sperm maturation is the key process for sperm to acquire its motility and fertilizability. Although C-type natriuretic peptide (CNP) is highly expressed in seminal plasma and male reproductive system, its role is still unclear. Applicant’s previous study showed that CNP could not only promote sperm motility and induce acrosome reaction through the signaling molecule cGMP, but also facillated sperm motility in epididymal head in vitro. Therefore, we hypothesize that CNP/cGMP could be a new signal transduction pathway of sperm maturation and motility. This project is divided into three parts: firstly, the expressions of CNP/NPR - B in the different parts of epididymis will be examied and the effects of CNP on sperm maturation will be assessed. Secondly, we will establish the conditional knockout mice model, then testify the role of the CNP/cGMP signal pathway in epididymal sperm maturation and motility by the application of knockout mice model and through a series of experiments like in vitro stimulation experiments. Finally, we will analyze the key downstream moleculars of CNP/cGMP signal pathway in epididymal sperm maturation by using gene chip and proteomics, thus illuminating the molecular mechanism of CNP in regulating sperm function. The aim of the project is to investigate the role and mechanism of CNP/cGMP in epididymal sperm maturation and motility, which will provide a new target for diagnosis and treatment of male infertility.

附睾精子成熟是精子获得运动能力及受精能力的关键。C型利钠肽（CNP）在精浆及雄性生殖系统中极高表达，但具体作用尚未明确。前期研究发现CNP体外不仅能通过cGMP提高精子活力及顶体反应率，还能促进附睾头部精子运动能力的获得。推测CNP/cGMP可能是促进附睾精子成熟及运动的新调节通路。该项目首先明确附睾不同部位CNP/NPR-B的表达情况及CNP对附睾精子成熟的影响，然后通过建立附睾条件性基因敲除小鼠模型和体外刺激等实验证实CNP/cGMP通路对附睾精子成熟及运动的影响；最后运用基因芯片及蛋白组学等方法分析CNP/cGMP信号通路在调节附睾精子成熟作用中的下游关键信号传导分子，从而阐明CNP调节精子成熟的分子机制。本课题旨在探讨CNP/cGMP对附睾精子成熟及运动的影响及其作用机制，为男性不育的诊治提供新的靶标和思路。

附睾精子成熟是精子获得运动能力及受精能力的关键。睾丸中产生的精子还不具备运动能力、精-卵识别和结合的能力，只有进入附睾后，经过附睾头、体、尾运行和附睾的贮存过程中， 精子才开始获得了运动能力、精-卵识别和结合能力。若附睾精子成熟出现障碍则会导致精子运动能力及受精能力障碍，最直接的体现是精子运动能力的下降，最终引发男性不育。因此，针对附睾精子成熟和运动的调控机制研究将有助于深入了解男性不育症的发病机制，并为临床诊断和治疗提供新的靶点及途径。.本研究发现：1）CNP可以通过cGMP通过影响精子钙内流及酪氨酸磷酸化，从而促进精子获能。2）CNP可以通过NPR-B/cGMP/PKG通路影响支持细胞内分泌功能,从而调节精子的发生. 3）CNP可通过提高精子活力来提高精卵受精率及后继的胚胎发育.同时也可通过直接作用于受精卵及胚胎来提高胚胎的质量.4）CNP/NPR-B在附睾组织高表达，尤其在头部表达更为明显，其表达受雄激素调控，并可通过cGMP通路促进附睾精子运动能力的获得及成熟, 用CNP处理附睾头部精子，可促进附睾精子运动能力的获得，增加精子上Bin1b、CD52mRNA、LDCH、Dnah17等表达，提示CNP能参与附睾精子成熟过程。5）在大肠肝菌诱导的附睾炎动物模型中，给予CNP可以明显减轻DEPC诱发的附睾炎症水肿程度及组织破坏程度，其机制是CNP可明显抑制附睾组织促炎因子分泌（IL-6、TNF-α、IL-1β），结果说明CNP可能是一个新的附睾炎关键免疫调节因子. 现已发表SCI文章6篇，在投SCI 二篇。.该课题找到新的附睾精子成熟和运动的调控通路，为男性不育诊疗的新方法,尤其是弱精子症的潜在治疗靶点。目前在不孕夫妇中男性因素约占50%，而造成男性不育的重要原因之一就是精子活力低下或无活力。因此CNP具有潜在的临床应用前景。