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作 者:唐骅[1] 李灿明[1] 赖渭妍[1] 饶嘉玲[1] 叶增纯[1] 娄探奇[1]
机构地区:[1]中山大学附属第三医院肾内科,广州510630
出 处:《新医学》2015年第3期144-148,共5页Journal of New Medicine
基 金:广东省医学科研基金(A2011186)
摘 要:目的探讨晚期糖基化终产物(AGEs)对大鼠肾小球内皮细胞(r GEn C)通透性的影响及线粒体功能障碍在其中的作用。方法采用原代培养的r GEn C,予AGEs 80 mg/L作用24 h,采用跨内皮细胞电阻抗和异硫氰酸荧光素标记的牛血清白蛋白滤过率观察通透性的变化,Mito SOX试剂盒检测细胞内活性氧,JC-1荧光标记法检测线粒体膜电位(△Ψm),荧光素酶检测系统测定三磷酸腺苷(ATP)的生成,蛋白免疫印迹法检测NF-E2相关因子2(Nrf2)的表达。结果 AGEs可引起r GEn C通透性升高,活性氧产生增加,采用叔丁基对苯二酚(t BHQ)(20μmol/L)、N-乙酰半胱氨酸(NAC)(10 mmol/L)和抗AGE受体抗体(100 mg/L)预处理后,上述AGEs作用被抑制。AGEs可引起r GEn C△Ψm下降,ATP和Nrf2减少,而anti-RAGE抗体可抑制△Ψm下降和ATP减少,t BHQ则能抑制△Ψm下降和Nrf2减少。结论 AGEs可导致r GEn C线粒体功能障碍,引起活性氧产生增加,从而导致r GEn C间通透性增加。Objective To investigate the effect of advanced glycation end products( AGEs) on the permeability of glomerular endothelial cells( r GEn C) in rats and the role of mitochondrial dysfunction in this pathological process. Methods Primary cultured r GEn C were incubated with AGEs( 80 mg / L) for 24 h. The changes in endothelial permeability were investigated by transendothelial electrical resistance and the flux of fluorescein isothiocyanate-conjugated bovine serum albumin. Mito SOX kit was used to detect intracellular reactive oxygen species( ROS). Mitochondrial membrane potential( △Ψm) was measured by JC1 fluorescein staining.The generation of adenosine triphosphate( ATP) was evaluated by luciferase assay system. The expression of NF-E2-related factor 2( Nrf2) was detected by western blotting. Results The monolayer permeability and the generation of ROS of r GEn C were increased by AGEs. Pretreatment with tert-Butyl-hydroquinone( t BHQ)( 20μmol /L),N-acetylcysteine( NAC)( 10 mmol /L) and anti-RAGE antibody( 100 mg /L) could suppress the detrimental effect of AGEs. The decline of △Ψm,ATP and Nrf2 were induced by AGEs,whereas the decreases of △Ψm and ATP could be blocked by pretreatment with anti-RAGE antibody and the decline of △Ψm and Nrf2 inhibited by t BHQ pretreatment. Conclusions AGEs can cause mitochondrial dysfunction,leading to elevated levels of ROS,which contributes to increase of permeability in r GEn C.
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