Family Planning Research Institute, Center for Reproductive Medicine, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China.
Shenzhen Key Laboratory of Fertility Regulation, Center of Assisted Reproduction and Embryology, The University of Hong Kong - ShenZhen Hospital, Guangdong, China.
Am J Reprod Immunol. 2019 Feb;81(2):e13079. doi: 10.1111/aji.13079. Epub 2019 Jan 8.
This study aims to determine the expression and localization of programmed cell death 1 (PD-1) and programmed cell death 1 ligand 1 (PD-L1) in the testes of mice at different developmental stages.
By means of RT-qPCR, Western blot and immunofluorescence, the expression and localization of PD-1 and PD-L1 were detected in the testicular tissues of mice at different postnatal times: P7, P14, P21, P28, P35, and adulthood. Meanwhile, the level of soluble PD-L1 (sPD-L1) was evaluated by ELISA in the testicular interstitial fluid (IF) of the adult mice, culture supernatants of TM4 cell lines (Sertoli cells lines), and primary Sertoli cells at P14.
Pd-1 mRNA levels were unexpectedly low. From P7 to P21, there was limited PD-1 protein detected while PD-1 was evident at P28 and afterward at significantly higher levels than at P14 and P21 (P < 0.05). Despite being found in the interstitial area at P7, P14, and P21, PD-1 was also detected in the germ cells of the seminiferous tubules after P28. Pd-l1 mRNA exhibited age-related changes, peaking at P21, while PD-L1 protein was constitutively expressed at any stage, specifically localized in the nucleus of Sertoli cells. Moreover, the level of sPD-L1 in IF was significantly higher than that in the culture supernatants of both TM4 and primary Sertoli cells at P14.
PD-1 and PD-L1 were present in the testicular tissue of adult mice. The expression and localization of PD-1 fluctuated with age, and PD-1 was mainly localized to advanced germ cells, suggesting that it may play a role in spermiogenesis. PD-L1 was constitutively expressed in the nucleus of Sertoli cells, which could secrete sPD-L1 into the testicular interstitial space and thus may be involved in testicular immune privilege.
本研究旨在确定程序性细胞死亡 1（PD-1）和程序性细胞死亡配体 1（PD-L1）在不同发育阶段小鼠睾丸中的表达和定位。
通过 RT-qPCR、Western blot 和免疫荧光法，检测不同出生后时间（P7、P14、P21、P28、P35 和成年期）小鼠睾丸组织中 PD-1 和 PD-L1 的表达和定位。同时，通过 ELISA 评估成年小鼠睾丸间质液（IF）、TM4 细胞系（支持细胞系）培养上清液和 P14 期原代支持细胞中可溶性 PD-L1（sPD-L1）的水平。
Pd-1 mRNA 水平出人意料地低。从 P7 到 P21，检测到有限的 PD-1 蛋白，而 PD-1 在 P28 后明显更高水平（P<0.05）。尽管在 P7、P14 和 P21 的间质区域发现 PD-1，但在 P28 后的生精小管生殖细胞中也检测到 PD-1。Pd-l1 mRNA 呈年龄相关性变化，在 P21 时达到峰值，而 PD-L1 蛋白在任何阶段均持续表达，特异性定位于支持细胞的核内。此外，IF 中的 sPD-L1 水平明显高于 P14 期 TM4 和原代支持细胞培养上清液中的水平。
PD-1 和 PD-L1 存在于成年小鼠的睾丸组织中。PD-1 的表达和定位随年龄波动，PD-1 主要定位于晚期生殖细胞，提示其可能在精子发生中发挥作用。PD-L1 在支持细胞的核内持续表达，可将 sPD-L1 分泌到睾丸间质空间，因此可能参与睾丸免疫豁免。
