黄芪多糖对高糖刺激小鼠肾小球足细胞Nephrin和Desmin表达的影响

The Effect of Astragalus Polysaccharide on the Expressions of Nephrin and Desmin in High Sugar Stimulation Mice Podocytes

目的模拟体外高糖病理微环境,探讨黄芪多糖对实验鼠肾脏肾小球足细胞Nephrin及Desmin表达的影响.方法试验(1):将培养的实验鼠肾脏肾小球足细胞分为5个组,分别是D-葡萄糖30 mmol/L+黄芪多糖干预组(0.0 g/L,0.1 g/L,0.2 g/L,0.4 g/L及0.8 g/L);干预时间为72 h,筛选出黄芪多糖干预的最佳浓度点0.Z g/L.试验(2):将培养的实验鼠肾脏肾小球的足细胞分7个组,分别是D-葡萄糖30 mmol/L+黄芪多糖的干预(干预浓度是0.Z g/L),干预的时间分别是0 h,3 h,6 h,12 h,24 h,48 h及72 h;筛选出干预的最佳时间点Y h.试验(3):根据上述试验(1)、(2)结果,将培养的小鼠肾小球足细胞分为:正常对照组、甘露醇高渗对照组、高糖组和黄芪多糖干预组(黄芪多糖的干预浓度是0.Z g/L,干预的时间是Y h),采用流式细胞术与实时聚合酶链反应(real-Time Polymerase Chain Reaction,real-Time PCR)分别检测七组足细胞Nephrin、Desmin蛋白与m RNA的表达.结果随着黄芪多糖干预梯度浓度的递增,实验鼠肾脏肾小球的足细胞Nephrin蛋白表达逐渐上调,Desmin蛋白表达逐渐下调,呈浓度依赖性(P<0.05);随着黄芪多糖干预时间的延长,实验鼠肾脏肾小球的足细胞Nephrin的表达逐渐上调,但肾小球的足细胞Desmin的表达逐渐下调,呈时间依赖性.相比正常对照组和甘露醇高渗对照组,高糖组小鼠肾小球足细胞Nephrin蛋白和m RNA表达均下降,Desmin蛋白和m RNA表达均增加(P<<0.05);而相较高糖组,黄芪多糖干预组Nephrin蛋白和m RNA表达明显上升,Desmin蛋白和m RNA表达明显降低(P<0.05).结论在高糖刺激下,黄芪多糖干预可上调肾小球足细胞Nephrin表达,降低肾小球足细胞Desmin表达,是黄芪治疗DN蛋白尿的可能作用靶点.

Objective To study the effect of Astragalus polysaccharide on the expressions of Nephrin and Desmin in high sugar stimulation mice podocytes. Me thods Test one： the cultured mouse podocytes were divided into 5 groups： astragalus polysaccharide group [D-glucose 30 mmol/L ＋ astragalus polysaccharide（0.0 g/L, 0.1 g/L,0.2 g/L,0.4 g/L and 0.8 g/L） ]. Test two： In test one, we chose the concentration of astragalus polysaccharide which most affected both expression of Nephrin and Desmin in podocytes and named it as astragalus polysaccharide 0.X g/L. The cultured mouse podocytes were divided into 7 groups： astragalus polysaccharide group（D-glucose 30 mmol/L ＋ astragalus polysaccharide 0.X g/L） had been cultured for 0 h, 3 h, 6 h, 12 h,24 h,48 h and 72 h, respectively. Test three： Basted on test two, we chose the time which most affected both expression of Nephrinand Desmin in podocytes and named it as Yh. The cultured mouse podocytes were divided into 4 groups： the control group,mannitol hypertonic glucose control group, high sugar group,and astragalus polysaccharide group（high sugar group ＋ astragalus polysaccharide 0.X g/L）. Each group had been cultured for Yh. Nephrin and Desmin of each subgroup was detected by flow cytometry podocytes and real-time PCR. Re s ults With the intervention of polysaccharide, Nephrin in podocyte cell surface gradually increased, Desmin gradually reduced in a concentration-dependent manner（ P〈0.05）. With the intervention of astragalus polysaccharide, Nephrin gradually increase and Desmin expression gradually decreased in a concentration-dependent manner（P〈0.05）.Compared to the control group and the high sugar group,Nephrin mR NA increased and Desmin mR NA decreased in0.8 g/L astragalus polysaccharide co-cultured for 72 h（ P〈0.05）. Conclus ion Astragalus polysaccharide intervention can increase Nephrin and reduce Desmin in podocytes, which are possible targets of Astragalus polysaccharide in treatment of DN proteinuria.