摘要
目的研究慢性肾衰竭(CRF)血清对兔主动脉内皮细胞泛素-蛋白酶体途径的影响,初步探讨CRF血清对主动脉内皮细胞作用的可能机制。方法建立CRF兔模型,建模后12周,取CRF模型兔及正常兔下腔静脉血血清备用。原代培养健康兔主动脉内皮细胞,用正常兔血清或CRF兔血清刺激后,分别采用RT-PCR、Western blotting及特异底物反应法检测不同血清对主动脉内皮细胞泛素化蛋白、泛素激活酶(简称E1)mRNA表达,泛素化蛋白、E1、κB抑制因子(IκB)蛋白表达及蛋白酶体活性的影响。结果 CRF血清使主动脉内皮细胞泛素、E1mRNA表达增加,明显高于正常血清(P<0.01)。CRF血清对主动脉内皮细胞泛素化蛋白的表达无明显影响,但使E1蛋白表达增加,使蛋白酶体20S亚基活性增强,明显高于正常血清(P<0.01)。CRF血清使主动脉内皮细胞IκB蛋白表达降低,与正常血清相比,差异有统计学意义(P<0.01)。结论 CRF血清可致兔主动脉内皮细胞泛素-蛋白酶体途径活化,进而影响NF-κB信号通路。
Objective To investigate the effects of chronic renal failure serum on the activation of ubiquitin-proteasome pathway of rabbit arterial endothelial cells （AECs） cultured in vitro. Methods AECs were cultured with growth media supplemented with pooled sera from normal rabbits or rabbits with chronic renal failure. Eighty percent of confluent AECs were incubated for 24h with media supplemented with pools of control or chronic renal failure sera. The expression of mRNA levels for ubquitin （Ub） and ubiquitin activating enzyme （E1） was analyzed by RT-PCR. The levels of protein for ubiquitinated-protein, E1 and IkBa were measured by Western blotting. The three enzyme activities of 20S proteasomes in the total protein was examined using three special fluorogenic peptide suhstrates. Results Compared with normal serum, chronic renal failure serum increased the expressions of E1 mRNA and protein in rabbit arterial endothelial cells （P〈0.01）, and the mRNA expression of Ub was also increased obviously （P〈0.01）, while the expression of IkB decreased obviously （P〈0. 01）. The ubiquitinated-protein was indifference in normal group and chronic renal failure group, but the activity of proteasomes was increased significantly at the same time （P〈0. 01）. Conclusion Chronic renal failure serum may activate the uhiquitinproteasome pathway in arterial endothelial cells.
出处
《解放军医学杂志》
CAS
CSCD
北大核心
2010年第10期1192-1195,共4页
Medical Journal of Chinese People's Liberation Army
基金
国家自然科学基金(30570763)
关键词
肾功能衰竭
慢性
血清
内皮细胞
泛素
kidney failure, chronic
serum
endothelial cells
ubiquitin
