Role Of Sertoli Cell Tight Junctions In The Pathogenesis Of Male Reproductive Dysfunction Induced By Oxidative Stress In Mice

Objective:Oxidative stress, induced by many conditions or events, plays a key role in the pathogenesis of male infertility. This investigation was:(1) To investigate role of Clauaidinll on the spermatozoal dysfunction induced by oxidative stress;(2) To investigate the relative roles of Claudin-3, Claudin-11, key sertoli cell factors in the establishment of the blood-testis barrier (BTB), and recovery of spermatogenesis from irradiation, thus to explain the block of spermatogonial differentiation and abnormal appearance of endocrine on the recovery of spermatogonesis from irradiation.Methods:(1)48male Kunming mice were randomly allocated to a sham control group and three oxidative stress groups. Mice in three oxidative groups were restrained with anesthesia and irradiated to the lower abdominal and scrotal areas with2Gy,6Gy or lOGy60Co y-ray respectively. Animals were enthanized at p.i (post irradiation)6weeks, recorded each weight, then harvested testes and obtained3ml blood of each mouse. Histology was observe through sections with HE staining; serum FSH levels were detected by enzyme-linked immunosorbent assay; semiquantitative real time PCR was performed to determine the mRNA levels of claudin-11, a key factor in establishment of the hemotesticular barrier, in each groups.(2) The animals composed of42KW mice were divided into seven groups randomly:Grop A, sham controlled; the lower abnominal and scrotal area of the mice were irradiated with single dose of2,6,10,6,6,6Gy60Co gamma ray after anaesthetizd in Group B, C, D, E, F, G.17β-estradiol(E2) treatments were initiated immediately for mice in Group E, F, G after irradiation at the dose of1,2,4μg/d via hypodermic injection for4weeks. Animals were enthanized at p.i (post irradiation)6weeks. The tubule differentiation index (TDI) was computed in the testis sections of each groups. Semiquantitative real time PCR was performed to determine the mRNA levels of InhibinβB, claudin-3and claudin-11in each groups. Claudin-11protein’s profile was generated by western blot.Results:(1) Compared with sham control, testis weight and tubule differentiation index was reduced in three irradiated groups (all P<0.05). Testes under oxidative stress, induced by irradiation, revealed presence of diminished seminifeous tubules along with disorganization of the spermato genic cells in histology. Serum FSH was elevated in6Gy irradiated mice (all P<0.05), more significantly after10Gy irradiation, FSH levels increaced to1.9fold. Along with reproductive dysfunction, Claudinll transcription levels in testes tended towards increasing, Claudinll mRNA concentration in testes with6Gy or1OGy irradiation was markedly higher than that of sham control(all P<0.05).(2) The TDI was markedly reduced as the dose of irradiation increasing (rs=-0.977, P<0.05). Compared with sham controls, the expression level of InhibinβB mRNA was decreased in testes exposed to lOGy gamma ray (t=6.906, P<0.017), indicating that Sertoli cells’function was impacted by irradiation, whereas in group C and D, which was exposed to6and10Gy gamma ray respactively, claudin-3, claudin-11mRNA and claudin-11protein levels was significantly higher than sham controls (all P<0.017). Estrogen (E2) enhances recovery of spermatogonesis from irradiation. Testes weight was improved in group F and group G (all P<0.017), treatment with E2at2μg/d increased the TDI from35%to near62%. Overexpression of claudin-3, claudin-11trended down words, claudin-11’s transcription and protein levels of group F were similar to that of sham controlled(t=-2.497, P>0.013; t=-2.256, P>0.013). Expression of claudin-3and claudin-11negatively correlated with TDI during the recovery of spermatogonesis from irradiation (rs=-0.884, P<0.05; rs=-0.758, P<0.05).Conclusion:Oxidative stress elevated expression of Claudin-11, which delayed cyclical restitution of hemotesticular barrier; increased serum FSH implied sertoli cells’ nutrition and secretion funcion were also damaged despite resistance to oxidative stress. These factors were associated with the pathogenesis of male reproductive dysfunction induced by oxidative stress.Through estrogen elevation or testosterone suppression, TJs’compositions, as claudin-3, claudin-11, were moderated to normal, BTB’s restructuring restored, leading spermatogonia through from basal compartment to adluminal compartment, where meiosis and spermiogenesis occur. Elevated TJs impair recovery of spermatogenesis from oxidative stress.