摘要
目的探讨哺乳动物雷帕霉素靶分子复合物(mTORC)在糖尿病肾病(DN)小鼠肾组织中的分布、表达。方法 14只C57BL/6小鼠随机分成对照组和DN组,每组各7只。DN组小鼠予以链脲菌素腹腔注射建立小鼠DN模型,采用生化技术检测小鼠血、尿肌酐以及白蛋白水平,组织学染色检测肾脏病理变化,免疫荧光以及免疫印迹技术检测肾组织中mTOR、mTOR第2448位丝氨酸磷酸化修饰后mTOR(p-mTOR)、mTORC1(Raptor)、mTORC2(Rictor)以及mTOR信号通路下游的效应蛋白磷酸化S6K1(p-S6K1)的分布和表达。结果 DN组小鼠血糖、尿白蛋白/肌酐比值明显增加(P<0.01),肾小球明显增大(P<0.05)。mTOR、Raptor以及Rictor在正常以及DN小鼠肾皮质和髓质中均有表达,主要表达在肾小球系膜区、毛细血管袢、皮质近曲小管以及外髓和内髓集合管上皮细胞中。其中正常小鼠内髓肾组织中未见p-S6K1表达,正常以及DN小鼠肾小球中未见p-mTOR表达。免疫印迹检测表明,DN小鼠肾组织中mTOR、p-mTOR、Raptor、Rictor以及p-S6K1均明显上升(P<0.05)。结论 mTORC广泛分布于小鼠肾组织且参与DN的发生发展,但mTOR第2448位丝氨酸磷酸化并不直接参与高血糖介导的肾小球损伤。
Objective To investigate the different distribution and expression of mammalian target of rapamycin complex （mTORC） in the kidney of diabetic nephropathy （DN） mice.Methods Fourteen eight-week-old male C57BL/6 mice were assigned to 2 groups： the control group （ n=7 ） and the streptozotocin （ STZ ）-induced DN group （ n=7 ） . Blood and urinary variables including glucose , albumin, creatinine and albumin/creatinine ratio were assessed 2 weeks after STZ injection.Hematoxylin-eosin staining was performed for renal pathological analyses .The distributions of mTOR , phosph-ser2448-mTOR（p-mTOR）, mTORC1（Raptor）, mTORC2（Rictor） and phosph-ser240/244-S6K1 （p-S6K1） were determined by immunofluorescence.The expression levels of mTOR, p-mTOR, mTORC1（Raptor）, mTORC2（Rictor）, S6K1 and p-S6K1 were detected by Western blotting .Results Two weeks after STZ injection , the diabetic mice developed albuminuria （P＆lt;0.01） and renal hypertrophy （P＆lt;0.05）.The immunofluorescence positive staining for mTOR , Raptor, and Rictor was distributed in the epithelial cells of proximal tubules , glomerular mesangium and capillary loops as well as the medullary collecting ducts of the control mouse kidney .These positive signals increased in the DN mouse kidney （ P＆lt;0.05）.However, pS6K1 was not detected in the inner medulla of control mouse and p-mTOR was not found in the glomeruli of both control and DN mice .Conclusion mTORC is widely expessed in the mouse kidney and participates in the development of DN , whereas the 2448 serine phosphorylation of mTOR may be not implicated in the hyperglycemia mediated glomerular injury .
出处
《解剖学报》
CAS
CSCD
北大核心
2014年第4期555-560,共6页
Acta Anatomica Sinica
基金
国家自然科学基金资助项目(81370818
30871173)
关键词
哺乳动物雷帕霉素靶分子
糖尿病肾病
免疫荧光
免疫印迹法
小鼠
Rictor
Raptor
Mammaliam target of rapamycin
Diabetic nephropathy
Rictor
Raptor
Immunofluorescence
Western blotting
Mouse
