摘要
目的:探讨糖尿病脑病中,Wnt/β-catenin信号通路在星形胶质细胞表型转化中的作用。方法:通过高脂喂养12周建立小鼠糖尿病模型。将12只C57BL/6小鼠随机分为对照组和糖尿病组,在第0周和12周分别检测小鼠的血糖水平和体质量。免疫荧光检测小鼠海马区A1型星形胶质细胞标记物C3和A2型星形胶质细胞标记物S100A10的表达。选用C8-D1A星形胶质细胞系,分为对照组、棕榈酸(PA)组、PA+Wnt信号通路激动剂(SKL2001)组。CCK-8检测不同浓度PA(0、50、100、200、300μmol/L)对细胞活力的影响;油红O染色检测不同浓度PA对星形胶质细胞中脂质含量的影响;免疫印迹检测星形胶质细胞标记物GFAP、C3、S100A10以及Wnt/β-catenin信号通路中Wnt3a、β-catenin、p-GSK3β、GSK3β蛋白表达水平;免疫荧光检测星形胶质细胞C3/GFAP、S100A10/GFAP、Wnt3a和β-catenin的表达。结果:与对照组小鼠比较,糖尿病组小鼠的血糖水平与体质量均明显升高。糖尿病组小鼠海马区C3的荧光强度明显高于对照组,S100A10的荧光强度在2组中差异无统计学意义。与对照组比较,不同浓度PA对C8-D1A细胞存活率均有抑制作用;油红O染色显示不同浓度的PA组中C8-D1A细胞的细胞质中可见橘红色颗粒着色。免疫印迹结果显示,与对照组比较,PA组的GFAP、C3蛋白表达升高,Wnt3a、β-catenin、p-GSK3β/GSK3β的蛋白表达降低;PA+SKL2001组中SKL2001可部分逆转PA组中蛋白表达变化;S100A10蛋白表达在3组间差异无统计学意义。免疫荧光结果显示,与对照组比较,PA组C3/GFAP的荧光强度升高;与PA组比较,PA+SKL2001组C3/GFAP的荧光强度降低;S100A10的荧光强度在3组中差异无统计学意义。与对照组比较,PA组Wnt3a的荧光强度降低,β-catenin向核内转移减少;与PA组比较,PA+SKL2001组Wnt3a的荧光强度升高,β-catenin向核内转移增加。结论:在糖尿病脑病中,星形胶质细胞表型发生转化可能是通过抑制Wnt/β-catenin信号通路相关蛋白的表达,导致星形胶质细胞向A1型转化,进而加重糖尿病脑病的发展。
Objective:To investigates the role of Wnt/β-catenin signaling pathway in the phenotypic transformation of astrocytes in diabetic encephalopathy.Methods:A diabetic mouse model was established by feeding the mice with high-fat diet for 12 weeks.Twelve C57BL/6 mice were randomly divided into control group and diabetic groups.The blood glucose level and body weight of the mice were detected at 0 and 12 weeks.Immunofluorescence was used to detect the expression of A1 astrocyte marker C3 and A2 astrocyte marker S100A10 in the hippocampus.C8-D1A astrocyte cell line was used to establish three groups:control,palmitic acid(PA)-treated,and PA+Wnt pathway activator(SKL2001).cell viability at various PA concentrations(0,50,100,200,300μmol/L)was assessed through the CCK-8 assay.Lipid accumulation in C8-D1A astrocytes was evaluated by using Oil Red O staining.Western blotting analysis was performed to measure the protein expression levels of GFAP,C3,S100A10,Wnt3a,β-catenin,p-GSK3βand GSK3β.Immunofluorescence was used to visualize the expression of C3/GFAP,S100A10/GFAP,Wnt3a andβ-catenin in astrocytes.Results:Compared with the control group,the blood glucose level and body weight of the diabetic group were significantly increased.The fluorescence intensity of C3 in the hippocampus of diabetic mice was significantly higher than that of the control group,but the fluorescence intensity of S100A10 was not significantly different between the two groups.Compared with the control group,PA treatment significantly inhibited the cell viability in a concentration-dependent manner.Oil Red O staining showed orange-red lipid droplets in the cytoplasm of C8-D1A cells in PA-treated groups.Western blotting results showed that the protein expressions of GFAP and C3 in the PA group were significantly higher than those in the control group,while the protein expressions of Wnt3a,β-catenin,and p-GSK3β/GSK3βin the PA group were significantly lower than those in the control group.In the PA+SKL2001 group,SKL2001 partially reversed the PA-induced changes in protein expression.No significant differences were found in S100A10 protein expression among the three groups.Immunofluorescence results confirmed that compared with the control group,the fluorescence intensity of C3/GFAP in the PA group was increased.Compared with the PA group,the fluorescence intensity of C3/GFAP was decreased in the PA+SKL2001 group,while the fluorescence intensity of S100A10 did not differ significantly between groups.Compared with the control group,the fluorescence intensity of Wnt3a and the translocation ofβ-catenin into the nucleus in the PA group decreased.Compared with the PA group,both indicators above were increased in the PA+SKL2001 group.Conclusion:In diabetic encephalopathy,the transformation of astrocyte phenotype may be caused by inhibiting the expression of Wnt/β-catenin signaling pathway-related proteins,leading to the transformation of astrocytes into type A1,thereby exacerbating disease progression.
作者
丁然
王欣笛
吴磊
高林荫
史宝宝
王海涛
Ding Ran;Wang Xindi;Wu lei;Gao Linyin;Shibaobao;Wang Haitao(Hebei Key Laboratory for Chronic Diseases,Tangshan Key Laboratory for Preclinical and Basic Research on Chronic Diseases,School of Basic Medical Sciences,North China University of Science and Technology,Tangshan 063200,China;School of Psychology and Mental Health,Hebei Key Laboratory of Mental Health and Brain Science,North China University of Science and Technology,Tangshan 063200,China)
出处
《解剖学杂志》
2025年第3期199-203,267,F0002,共7页
Chinese Journal of Anatomy
