基于HMGB1/Beclin-1信号通路探讨温阳利水通络法对肾病综合征模型小鼠TNF-α、IL-4、IL-8的作用机制

Mechanism of Wen Yang Li Shui Draining collaterals therapy on TNF-α,IL-4and IL-8 in nephrotic syndrome model mice based on HMGB1/Beclin-1signaling pathway

目的:基于HMGB1/Beclin-1信号通路探讨温阳利水通络法对肾病综合征模型小鼠TNF-α、IL-4、IL-8的作用机制。方法:实验级小鼠30只,分为对照组、模型组、温阳利水通络法组,每组10只。模型组、温阳利水通络法组注射盐酸阿霉素构建肾病综合征小鼠模型,对照组注射等体积生理盐水。造模结束后第5天,温阳利水通络法组进行给药,对照组和模型组给予同剂量蒸馏水灌胃,1次/d,连续3周。给药停止后24 h,检测小鼠的肾功能指标;HE染色观察小鼠肾组织病理变化;酶联免疫吸附法比较小鼠外周血TNF-α、IL-4、IL-8的表达水平;Western blot检测各组小鼠肾组织中HMGB1/Beclin-1蛋白及相关蛋白的表达变化。结果:模型组小鼠肌酐(CRE)、尿素氮(BUN)、24 h尿蛋白量较对照组增加;温阳利水通络法组CRE、BUN、24 h尿蛋白量较模型组下降。对照组小鼠肾小球形状规则,肾小囊列队排列,界限清晰;模型组小鼠肾小球形态异常肿大,肾小囊皱缩,基底膜加厚伴系膜基质变多,边界模糊;温阳利水通络法组小鼠肾小球形态改善,肿大减小,肾小囊皱缩缓解。模型组小鼠肾组织病理评分较对照组增加;温阳利水通络法组肾组织病理评分较模型组下降。模型组小鼠TNF-α、IL-4、IL-8表达较对照组增加;温阳利水通络法组TNF-α、IL-4、IL-8表达较模型组下降。模型组小鼠HMGB1、LC3、Beclin-1蛋白表达较对照组增加;温阳利水通络法组HMGB1、LC3、Beclin-1蛋白表达较模型组下降。结论:温阳利水通络法能够调节控制HMGB1/Beclin-1信号通路,抑制肾病综合征模型小鼠TNF-α、IL-4、IL-8等炎症因子表达,从而对模型小鼠肾组织起保护作用。

Objective:To investigate the mechanism of Wen Yang Li Shui Draining collaterals therapy on TNF-α,IL-4 and IL-8 in nephrotic syndrome model mice based on HMGB1/Beclin-1 signaling pathway.Methods:Thirty experimental mice were divided into control group,model group and Wen Yang Li Shui Draining collaterals method group,with 10 mice in each group.Model group and Wen Yang Li Shui Draining collaterals method group were injected with doxorubicin hydrochloride to make mouse model of nephrotic syndrome,while the control group was injected with normal saline.On the fifth day after the modeling,the Wen Yang Li Shui Draining collaterals method group was given medicine,and the control group and the model group were given distilled water by gavage,once a day,for three consecutive weeks.He staining was used to observe the pathological changes of kidney tissue.The renal function indexes of mice were detected 24 h after the cessation of drug administration.The expression levels of TNF-α,IL-4,IL-8 in peripheral blood of rats were compared by enzyme-linked immunosorbent assay.The expressions of HMGB1/Beclin-1 protein and related proteins in renal tissues of mice were detected by Western blot.Results:The creatinine(CRE),urea nitrogen(BUN)and 24 h urine protein in model group were higher than those in control group.The CRE,BUN and 24 h urine protein in Wen Yang Li Shui Draining collaterals method group were lower than those in model group.In the control group,the glomeruli were regular in shape,and the renal capsules were arranged in rows with clear boundaries.In model group,the morphology of glomeruli was abnormally enlarged,the renal capsule was shrunk,the basement membrane was thickened with more mesangial matrix,and the boundary was fuzzy.In Wen Yang Li Shui Draining collaterals method group,the morphology of glomeruli was improved,the swelling was reduced,and the shrinkage of renal capsule was alleviated in mice.The pathological score of kidney tissue in model group was higher than that in control group.The pathological score of kidney tissue in the Wen Yang Li Shui Duan collateral group was lower than that in the model group.The expressions of TNF-α,IL-4 and IL-8 in model group were higher than those in control group.The expressions of TNF-α,IL-4 and IL-8 in Wen Yang Li Shui Duan collateral group were lower than those in model group.The protein expressions of HMGB1,LC3 and Beclin-1in model group were higher than those in control group.The protein expressions of HMGB1,LC3 and Beclin-1 in Wen Yang Li Shui Duan collateral group were lower than those in model group.Conclusion:The method of Wen Yang Li Shui Duan collateral can regulate and control the HMGB1/Beclin-1 signaling pathway,inhibit the expressions of TNF-α,IL-4,IL-8 and other inflammatory factors in the model mice with nephrotic syndrome,and thus protect the kidney tissue of the model mice.