目的:研究高压氧对大鼠糖尿病睾丸病变的保护作用并探讨其机制.方法:32只大鼠,其中8只Wistar正常大鼠,24只2型糖尿病GK大鼠,分4组,分别为对照组、模型组、二甲双胍对照组及高压氧组.对照组和模型组予纯净水5 mL/(kg·d)灌胃,二甲双胍组予二甲双胍混悬液250 mL/(kg·d)灌胃,高压氧组予纯净水5 mL/(kg·d)灌胃,同时加以稳压纯氧0.15 MPa处理30 min.3周后集中处死,摘取睾丸组织.睾丸组织经甲醛溶液固定后制成切片,分别行HE染色观察组织结构变化;TUNEL法检测细胞凋亡情况,免疫组织化学法检测NOS及SOD蛋白表达水平.结果:对照组睾丸曲细精管HE染色后光镜下见丰富饱满,生精细胞5~6层,腔内见丝状精子;模型组睾丸曲细精管萎缩,生精细胞数量减少,只有2~3层,腔内精子减少;高压氧组睾丸结构破坏较模型组明显减轻,曲细精管内精子数量明显增多,结构较完整.睾丸的凋亡主要发生在生精细胞,模型组凋亡指数最高,与其他3组相比差异具有统计学意义(P<0.05);高压氧干预后凋亡细胞数较模型组减少,但仍高于对照组(P<0.05),与二甲双胍组类似.模型组NOS表达最强,对照组最弱,与其他3组比较差异均有统计学意义(P<0.05);高压氧组NOS表达介于对照组和模型组之间(P<0.05),与二甲双胍组类似.模型组SOD表达最弱,对照组最强,高压氧组介于两者之间,与两组比较差异均有统计学意义(P<0.05).结论:糖尿病时氧自由基增多,并引起凋亡增加是引起睾丸损伤的重要原因,高压氧可以促进SOD的合成,使其清除多余氧自由基,通过此机制可以抑制细胞过度凋亡,从而达到保护睾丸组织的目的.%Objective: To explore the effect and the mechanism of hyperbaric oxygen therapy on testis of GK rats.Methods: Twenty-four GK rats were randomly divided into a model group, a metformin hydrochloride group and a hyperbaric oxygen group, 8 normal Wistar male rats were in the control group. The rats in the control group,the model group and the hyperbaric oxygen group received purified water 5 mL/kg once a day, while the rats in the metformin group received intragastric administration with metformin hydrochloride 250 mg/kg once a day.In addition, the rats in the hyperbaric oxygen group inhaled pure oxygen under a constant pressure (0.15 MPa) for 30 minutes. After 3-week treatment, the testis of all rats were excised for examination. For each sample, the formalin fixed testicular tissue were made into slices. Then the slices were stained by HE to observe the tissue structure. Subsequently, the apoptosis and expression level of NOS and SOD were determined by TUNEL and immunohistochemistry. Results: Under the microscopy, the testis seminiferous tubules (HE staining) of rats in the control group were full with filamentous sperm in spermatogenic cell lumen and 5- to 6-layer sperm cells.In the model group, the rats' seminiferous tubules were atrophied, and quantity of spermatogenic cell decreased (2-3 layers). Compared with the model group, the testis structure of rats in the hyperbaric oxygen group was more integrated, and the quantity of sperm increased. The apoptosis of testis mainly occurs in the androgone.The amounts of apoptotic cells increased significantly in the model group (P<0.05), while the apoptotic cells deceased after intervention of hyperbaric oxygen (more than the control group) whose effect was similar to the metformin group. The NOS level of the rats in the model group was much higher than that in the other group,while the level of the control group was the lowest (P<0.05). The NOS expression of the hyperbaric oxygen group was weaker than the model group, but stronger than the control group (P<0.05). Moreover, the SOD expression in the hyperbaric oxygen group was stronger than the model group, but weaker than the control group. Conclusion:Increased oxygen free radicals and apoptosis is an important cause of testicular injury in diabetes. However,hyperbaric oxygen can induce the generation of SOD, remove excess oxygen free radicals, and inhibit excessive apoptosis cells so as to protect the testis tissue.
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