| Objective: To study on the influence of ureaplasma urealyticum(UU) on spermatogenic cells and the role of antibiotic therapeutic intervention on UU infection.These could provide insight into exploring the mechanisms of UU infection to cause male infertility and giving the new way for Clinical treatment to male infertility.Method:(1) The clean male SD rats were employed for the present study and randomly divided into four groups:the control group, the 7-day group, the 14-day group and the treatment group. Except of the control group, each rat in the other three groups was injected one time with 0.6 ml 1 × 107CCU/ml UU through punctio vesicae. In the same way, each rat in the control group was injected with the same volume of UU liquid medium. Each rat in the treatment group was intragastrically administered with minocycline at the seventh day after injection. 7 days after injection in both the control and 7-day groups as well as 14 days after injection in among the control,14-day and treatment groups, the blood and testicles of rats anaesthetized by 20%urethane solution were acquired for the following experiments.(2) The weight of the rat testicles in each group was weighed by electronic balance.(3) The testicle tissues of the rats in each group were extracted for DNA, amplified for the conservative urease gene fragments of UU with the PCR method and detected by agarose electrophoresis.(4) The testicle tissues of the rats in each group were fixed for 16-18 h in Bouin’s liquid. Then, they were observed for the pathological changes through the method of HE staining, determined for the cells apoptosis by the TUNEL method and for the levels of apoptosis proteins Bak, Bax, Fas L through the immunohistochemical method.(5) The testis interstitial fluid and plasma of the rats in each group were acquired to test the level of testosterone by the Chemi Luminescence method.(6) The testicle tissues of the rats in each group were fixed in 2% glutaraldehyde solution over 2h, and investigated for the ultrastructure of testicle spermatogenic cells by transmission electron microscope.Result:(1) After UU infection, the conservative urease gene regarded as the special gene fragment of UU was discovered in testicle tissue in the 7-day, 14-day and treatment groups, but not in the control group.(2) Although the testis weights in the 7-day group were slightly decreased, the testicle weights among each group were no statistical significance.(3) In the testis tissues of the 7-day and 14-day groups, HE staining showed the pathology changes including the decreased layers and amounts of the spermatogenic cells, basement membrane lesion, inflammatory cells infiltration and decreased mature sperms, but the pathology changes in the 14-day group were lighter than those in the 7-day group. The structures of testicular tissue in the treatment group were close to the normal.(4) After UU infection, it was found that the level of the testosterone in testis interstitial fluid in the 7-day group was significantly higher than those in the control,14-day and treatment groups(P<0.01). But there was no statistical significance among the control, 14-day and treatment groups.(5) Compared with the levels of the testosterone in the plasma in the control and treatment groups after UU infection, those in the 7-day and 14-day groups were substantially decreased(P<0.01), but the level of the testosterone in the 14-day group was lower than that in the 7-day group(P<0.05).(6) The results by TUNEL method showed that the percentages of the spermatogenic cells apoptosis in the 7-day, 14-day and treatment groups were significantly higher than that in the control group(P<0.01), and the percentages of the spermatogenic cells apoptosis in the 7-day and 14-day groups were significantly higher than that in the treatment group. But the percentage of the spermatogenic cells apoptosis in the 7-day group was markedly higher than that in the 14-day group(P<0.01).(7) The results from transmission electron microscope showed the concentrated and gathered chromatin in the nucleus of the spermatogenic cells in the 7-day and 14-day groups. Furthermore, the nuclear membrane shrinkage and nuclei break were also found in the spermatogenic cells of the 7-day group. The morphological structures of spermatogenic cells in the treatment group were close to the normal.(8) The levels of apoptosis proteins Bak, Bax and Fas L were determined in the testicular tissue through the immunohistochemical method. It was found that the levels of Bak protein expression in the 7-day, 14-day and treatment groups were significantly higher than that in the control group(P<0.01), but the levels in the 7-day and 14-day groups were significantly higher than that in the treatment group(P<0.01 and P<0.05). Compared with the control and treatment groups, the levels of Bax protein expression in the 7-day and 14-day groups were obviously increased(P<0.01),but the level in the 7-day group was remarkably higher than that in the 14-day group(P<0.05). At the same time, it was found that the levels of Fas L protein expression in the 7-day, 14-day and treatment groups were significantly higher than that in the control group(P<0.01 and P<0.05), but the levels in the 7-day and 14-day groups were significantly higher than that in the treatment group(P<0.01 and P<0.05) and the level in the 7-day group was remarkably higher than that in the 14-day group.(P<0.01).Conclusion:(1) UU infection could cause the pathology changes in the testicular tissue.(2) UU infection could cause the increase of the apoptosis rate of the spermatogenic cells and of the expressed levels of the apoptosis proteins, which suggested that UU infection was able to induce the apoptosis of the spermatogenic cells.(3) UU infection could decrease the levels of the testosterone in the plasma and testis interstitial fluid, which may be associated with male infertility.(4) The antibiotic therapeutic intervention could attenuate the pathological changes,decrease the apoptosis of the spermatogenic cells and increase the levels of the testosterone. |
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