青风藤活性成分对糖尿病db/db小鼠氧化应激、炎症反应及免疫功能的影响

Effects of Active Ingredients of Sinomenine on Oxidative Stress,Inflammatory Response,and Immune Function in db/db Mice with Diabetes Mellitus

目的:探讨青风藤活性成分对糖尿病(diabetes mellitus,DM)db/db小鼠肾脏的保护作用及其机制研究。方法:30只6周龄雄性db/db小鼠随机分为模型组(等体积生理盐水),阳性组(195 mg/kg/d羟苯磺酸钙),盐酸青藤碱(Sinomenine Hydrochloride,SH)低、中、高剂量组(31.2、62.4、124.8 mg/kg/d SH)5组,同时选取6只同周龄db/m小鼠设为对照组(等体积生理盐水),每日1次,定时灌胃给药,连续6 w。于12 w末观察各组小鼠一般情况,检测尿微量白蛋白(mAlb)、血肌酐(SCr)和血尿素氮(BUN)水平;试剂盒检测肾组织匀浆中丙二醛(MDA)、还原型谷胱甘肽(GSH)和超氧化物歧化酶(SOD)的水平;Luminex多因子检测技术测定血清肿瘤坏死因子-α(TNF-α)、白细胞介素-1β(IL-1β)、白细胞介素-6(IL-6)和白细胞介素-10(IL-10)的含量;ELISA法检测血清免疫球蛋白G(IgG)、免疫球蛋白M(IgM)、免疫球蛋白A(IgA)及补体蛋白3(C3)的水平;苏木精-伊红(HE)和过碘酸-雪夫(PAS)染色观察肾脏组织病理形态变化;透射电镜观察肾脏超微结构的改变。结果:与模型组比较,阳性组和SH低、中、高剂量组小鼠mAlb、SCr和BUN水平均降低(P<0.05);肾组织匀浆中MDA水平降低(P<0.05),GSH水平和SOD活性均升高(P<0.05);血清中TNF-α、IL-1β、IL-6、IgG、IgM、IgA和补体C3水平均降低(P<0.05),IL-10水平升高(P<0.05);镜下肾小球损伤及肾小管上皮水肿情况均有所改善,肾组织损伤减轻。结论:青风藤活性成分对DM db/db小鼠的肾脏具有保护作用,其作用机制可能与改善氧化应激反应,缓解炎症反应及提高免疫功能有关。

Objective:To investigate the protective effects of active ingredients of Sinomenine on the kidneys of db/db mice with Diabetes Mellitus(DM)and its underlying mechanisms.Methods:Thirty 6-week-old male db/db mice were randomly divided into five groups:a model group(treated with equal volume of saline),a positive group(treated with 195 mg/kg/d calcium dobesilate),and low,medium,and high-dose groups of Sinomenine Hydrochloride(SH)(treated with 31.2,62.4,and 124.8 mg/kg/d SH,respectively).Six age-matched db/m mice were selected as the control group and received an equal volume of saline.All mice were administered by gavage once daily for 6 consecutive weeks.At the end of 12 weeks,the general conditions of mice in each group were observed,and levels of urinary microalbumin(mAlb),serum creatinine(SCr),and blood urea nitrogen(BUN)were measured.Kits were used to detect the levels of malondialdehyde(MDA),reduced glutathione(GSH),and superoxide dismutase(SOD)in renal tissue homogenates;Luminex multi-factor detection technology to determine serum levels of tumor necrosis factor-α(TNF-α),interleukin-1β(IL-1β),interleukin-6(IL-6),and interleukin-10(IL-10);ELISA to measure serum levels of immunoglobulin G(IgG),immunoglobulin M(IgM),immunoglobulin A(IgA),and complement protein 3(C3);Hematoxylin-eosin(HE)and periodic acid-Schiff(PAS)staining to observe the renal histopathological changes;and transmission electron microscopy to observe the renal ultrastructural changes.Results:Compared with the model group,the levels of mAlb,SCr,and BUN in the positive group and the low,medium,and high-dose SH groups were all decreased(P<0.05);The level of MDA in renal tissue homogenates was decreased(P<0.05),while the levels of GSH and SOD activity were increased(P<0.05);The serum levels of TNF-α,IL-1β,IL-6,IgG,IgM,IgA,and complement C3 were all decreased(P<0.05),while the level of IL-10 was increased(P<0.05);The glomerular injury and tubular epithelial edema were improved,and the renal tissue damage observed under the microscope was reduced.Conclusion:The active ingredients of Sinomenine have a protective effect on the kidneys of diabetic db/db mice,and its mechanism may be related to improving oxidative stress response,alleviating inflammatory response,and enhancing immune function.