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目的:对人类睾丸组织中表达血红素加氧酶(HO)的细胞进行定位;通过测定原发性无精子症及梗阻性无精子症患者睾丸组织中血红素加氧酶1(HO-1)的表达量与正常睾丸组织中HO-1表达量的差异性,来探讨其与无精子症发病的相关性。方法:应用免疫组化方法对人类睾丸组织中表达HO的细胞进行定位;采用逆转录-荧光定量PCR(FQ-PCR)方法定量检测无精子症患者与正常人睾丸组织HO-1及HO-2基因水平的表达量;应用W est-ern印迹检测各组之间HO蛋白水平表达量。结果:在正常睾丸组织,HO-1主要表达在支持细胞上;而HO-2在支持细胞和各级生精细胞中均有表达;FQ-PCR结果显示非梗阻性无精子症患者睾丸组织HO-1、HO-2的表达量均显著低于正常组及梗阻性无精子症组(P<0.05),差异具有统计学意义。而梗阻性无精子症患者睾丸组织表达HO-1、HO-2的量与正常组相比无显著性差异。W estern印迹结果显示HO-1蛋白水平的表达量差异与基因水平一致。而HO-2的蛋白水平在各组之间表达没有显著性差异。结论:非梗阻性无精子症患者睾丸组织中HO的表达量显著性降低,且HO-1无论是蛋白水平还是基因水平的差异一致。HO-1可以通过抗炎、抗氧化、抗凋亡的机制保护睾丸组织免受各种应激的损伤,从而维护正常的生精功能。可见,HO-1的减少可能与生精功能低下相关,这可能是非梗阻性无精子症的发病机制之一。
OBJECTIVE: To locate the heme oxygenase (HO) -positive cells in human testis tissue. By measuring the expression of heme oxygenase 1 (HO-1) in testis of patients with primary azoospermia and obstructive azoospermia, And the difference of HO-1 expression in normal testes to explore its correlation with the development of azoospermia. Methods: Immunohistochemistry was used to locate the cells that expressed HO in human testis. HO-1 and HO-2 were detected by reverse transcription-fluorescence quantitative polymerase chain reaction (FQ-PCR) in testicular tissue of azoospermia patients The expression level of HO protein was detected by W est-ern blot. Results: In normal testicular tissue, HO-1 mainly expressed on the supporting cells; while HO-2 was expressed in the supporting cells and spermatogenic cells at all levels; FQ-PCR results showed that the non-obstructive azoospermia testis HO -1, HO-2 were significantly lower than the normal group and obstructive azoospermia group (P <0.05), the difference was statistically significant. The obstructive azoospermia testicular tissue expression of HO-1, HO-2 compared with the normal group no significant difference. The result of Western blot showed that the difference of HO-1 protein level was consistent with the gene level. There was no significant difference in HO-2 protein expression between the groups. CONCLUSION: The expression of HO in testis tissue of non-obstructive azoospermia patients is significantly decreased, and the difference of HO-1 protein level or gene level is consistent. HO-1 protects testicular tissue against various stress through anti-inflammatory, anti-oxidant and anti-apoptotic mechanisms, thus maintaining normal spermatogenesis. It can be seen that the decrease of HO-1 may be related to the poor function of spermatogenesis, which may be one of the pathogenesis of non-obstructive azoospermia.