灵芝孢子保护糖尿病模型大鼠附睾及对自噬的影响

Protective effect of Ganoderma lucidum spores on epididymis and authophage in diabetic rats

背景:糖尿病引起的高血糖与男性生殖不育密切相关,灵芝孢子长期以来被认为具有抗衰老、改善血糖水平的功能。然而灵芝孢子对糖尿病附睾的保护作用机制未见详细报道。目的:探讨灵芝孢子对糖尿病附睾的保护作用机制。方法:雄性SD大鼠随机分为正常组(n=10)、高脂高糖组(n=10)和模型组(n=20)。正常组正常喂养,高脂高糖组和模型组大鼠饲以高脂高糖饮食。模型组大鼠在高脂高糖饮食基础上腹腔注射链脲佐菌素30 mg/kg诱导糖尿病大鼠模型,将造模成功的大鼠随机分为糖尿病组和灵芝孢子组(n=10);其中灵芝孢子组大鼠灌胃灵芝孢子300 mg/(kg·d);高脂高糖组与糖尿病组大鼠灌胃等量生理盐水,持续饲以高脂高糖饮食,连续干预12周。称体质量,检测空腹血糖、超氧化物歧化酶活性及丙二醛水平,并检测精子畸形率;苏木精-伊红染色观察大鼠附睾结构变化,Western blot检测大鼠附睾组织裂解半胱氨酸蛋白酶3和beclin1、p62、LC3蛋白表达,免疫组织化学染色法检测大鼠附睾组织裂解半胱氨酸蛋白酶3和p62的蛋白表达。结果与结论:①灵芝孢子可显著恢复附睾功能,明显改善附睾组织病理结构,提高细胞自噬和抗氧化能力;②免疫组织化学染色结果显示,灵芝孢子组大鼠附睾组织p62和裂解半胱氨酸蛋白酶3免疫强度显著下降;③Western blot分析结果显示,与糖尿病组大鼠相比,灵芝孢子可显著上调大鼠附睾组织beclin1和LC3蛋白表达,降低p62和裂解半胱氨酸蛋白酶3的蛋白表达;④提示灵芝孢子可能通过诱导自噬,提高抗氧化能力和抑制细胞凋亡,对糖尿病大鼠附睾产生保护作用。

BACKGROUND:Hyperglycemia caused by diabetes are closely related to male infertility,and Ganoderma lucidum spores are considered to be anti-aging and improve blood glucose levels.However,the protective effect of Ganoderma lucidum spores on diabetic epididymis remains unclear.OBJECTIVE:To investigate the protective effects of Ganoderma lucidum spores on diabetic epididymis.METHODS:Male Sprague-Dawley rats were randomly divided into normal group(n=10),high fat/high glucose group(n=10),and model group(n=20).Rats in the normal group were fed with normal diet,while those in the high fat/high glucose group and model group were fed with high fat and high glucose diet.The rats in the model group were then intraperitoneally injected with streptozotocin(30 mg/kg)to induce diabetes mellitus.Model rats were randomly divided into diabetes mellitus group(n=10)and Ganoderma lucidum spores group(GLS,n=10).The rats in the GLS group were treated with Ganoderma lucidum spores(300 mg/kg per day)by gavage,and the rats in the high fat/high glucose group and diabetes mellitus group were gavaged with the equal volume of normal saline and fed with high fat and high glucose diet for 12 weeks.Body mass,fasting blood glucose,superoxide dismutase activity,malondialdehyde level and sperm deformity rate were measured in each group.Structural changes in the epididymis in diabetic rats were observed by hematoxylin-eosin staining.The protein expression levels of cleaved caspase-3,beclin1,p62,and LC3 in the rat epididymal tissue were detected by western blot and immunohistochemical staining.RESULTS AND CONCLUSION:Ganoderma lucidum spores could significantly recovery epididymal function,improve the epididymal histopathology,and promote cell autophagy and antioxidant capacity.Immunohistochemical staining results showed that the immune intensity of p62 and cleaved caspase-3 decreased significantly.Western blot analysis showed that Ganoderma lucidum spores obviously upregulated beclin1 and LC3 protein expression levels and decreased p62 and Cleaved caspase-3 protein expression levels when compared with the diabetes mellitus group.To conclude,Ganoderma lucidum spores may induce autophagy,increase antioxidant capacity and inhibit apoptosis to protect against epididymal injury in diabetic rats.