睾丸炎中PK2通过NLRP3炎性小体通路促进IL-1β分泌影响生精功能的研究

Inflammation of male reproductive system is a common clinical condition affecting male fertility. Although testes have immune privileged mechanisms, microbial infections can still cause inflammation and result in impairment of testicular spermatogenesis. The uropathogenic E. coli (UPEC) is the most frequently isolated pathogen from patients with inflammatory diseases in the genital tract, which is a focus of clinical attention. Recent studies have found that the inflammatory cytokine IL-1β is secreted by testicular macrophages (TM) with immunosuppressive properties under inflammatory conditions, but the mechanism is not clear. PK2 protein plays an important role in the regulation of immune function and many physiological processes by regulating intracellular calcium concentration. Our previous study found that PK2 protein was significantly increased in TM of the UPEC orchitis rat model and involved in the regulation of NLRP3 inflammasome pathway, which is closely related to the production of IL-1β. We conclude that: PK2 protein induced TM intracellular calcium concentration increased and NLRP3 pathway activated to generate a great quantity of IL-1β under inflammation condition, resulting in damage to testicular spermatogenesis. To verify the conclusion, the UPEC orchitis rat model was established to detect the relation among PK2 , IL-1β production and impairment of spermatogenesis by activating NLRP3 pathway in TM. The knockout experiments will be used to explore the role of PK2 protein in the regulation of NLRP3 pathway through calcium channel in vitro. The completion of this project not only confirms the mechanism of PK2 protein in the innate immunity of the testis, but also provides a target for the treatment of orchitis.

生殖系统炎症是临床影响男性生育的常见病症，尽管睾丸具有免疫豁免机制，但微生物感染仍可引起炎症，以尿路致病性大肠杆菌UPEC感染最为常见。我们前期在UPEC睾丸炎模型中发现睾丸巨噬细胞在低免疫条件下分泌过量炎性因子IL-1β，但机制并不明确。而PK2蛋白在睾丸巨噬细胞中显著升高，并参与调控NLRP3炎性小体通路，与IL-1β产生相关。已知PK2调控细胞内钙离子浓度，且钙离子信号参与激活NLRP3炎性小体通路。因此我们提出假设：睾丸炎中睾丸巨噬细胞PK2蛋白动员细胞内钙离子浓度升高，调控NLRP3通路中Caspase-1活性产生过量IL-1β，对睾丸生精功能造成损伤。为证明此，我们拟在UPEC睾丸炎模型中，证实PK2通过激活NLRP3通路促进IL-1β产生并造成生精功能损伤，并在原代分离培养的睾丸巨噬细胞中探索PK2蛋白通过钙离子通路调控上述炎性反应的分子机制。本项目完成为治疗睾丸炎提供靶点。

睾丸炎是影响男性生育的常见病症，为了解相关免疫调控机制，寻找治疗靶点，研究根据课题假设开展相关实验。在构建成功的UPEC睾丸炎模型中，发现UPEC可入侵至睾丸间质，促进PK2在巨噬细胞内的表达及向细胞外释放。PK2的上调通过激活NLRP3炎性小体通路促进睾丸巨噬细胞分泌IL-1β，IL-1β的浓度升高可抑制睾酮合成相关的酶P450scc和P450c17的表达，从而降低睾酮水平。进一步发现UPEC感染后，PK2可能从睾丸巨噬细胞内释放，通过促进钙离子内流来激活NLRP3炎性小体通路，从而对精子活力、总数以及血清睾酮的水平产生负面影响，然而CaSR抑制剂可以在一定程度上缓解这些炎性损伤。本项目的明确PK2在睾丸炎中的作用，为治疗睾丸炎提供了潜在靶点。