EPO在大鼠缺血再灌注急性肾损伤模型中对内质网应激的影响

发布时间：2018-09-09 19:59

【摘要】：背景：急性肾功能衰竭主要表现为急性肾功能的改变,其主要原因之一为急性肾小管损伤,是临床常见的疾病之一,总体死亡率高,目前以支持或替代治疗为主。红细胞生成素(Erythropoietin, EPO)通过结合红系组细胞表面的EPO受体(EPOR),促使红细胞存活、增殖和分化。近年来发现神经细胞、心肌细胞和肾小管上皮细胞表面均具有EPOR。研究发现EPO对脑缺血和心肌梗塞均具有一定的治疗作用。近年来EPO在肾脏缺血再灌注中的研究也越来越多。EPO的保护机制可能是激活JAK2下游抗凋亡途径。内质网应激(Endoplasmic reticulum stress,ERS)是组织细胞在亚细胞层面应对外界刺激的常见反应,主要通过未折叠蛋白反应(Unfolded protein response,UPR)起始PERK、ATF6和IRE1信号途径,UPR可能具有两面性,早期起细胞保护作用,晚期则通过CHOP、JNK及Caspase12等促进细胞凋亡。目的：探讨EPO对缺血再灌注急性肾损伤的保护作用及EPO对内质网应激和全身炎症因子的影响方法：将104只300-350g雄性SD大鼠随机分组,根据干预情况分为假手术+生理盐水组(Sham+NS,n=20)、假手术+给药组Sham+EPO,n=20)、缺血再灌注+生理盐水组(IR+NS,n=32)及缺血再灌注+给药组(IR+EPO,n=32)、同时根据再灌注时间分为再灌注1、3、6和24小时组。其中假手术组在每个再灌注时间点为5只大鼠,缺血再灌注组在每个灌注时间为8只大鼠。在手术前2小时根据分组给予EPO或NS。真手术组,夹闭双侧肾蒂缺血90分钟后行再灌注。各组分别于再灌注1、2、6及24小时后,留取下腔静脉血及肾脏组织标本并处死动物。检验科协助静脉血检测血清肌酐和尿素氮。右肾组织行HE病理检测、免疫组化测定CHOP,以及ELISA测定血清MCP-1、IL-1β、IL-6和TNF-α水平。结果：(1)与Sham+NS组比较,IR+NS组大鼠SCr及BUN水平于再灌注1小时即有显著上升(P0.001),并且随着再灌注时间的延长逐渐升高。(2)在缺血再灌注损伤组的各个时间点(1、3、6和24小时)中,EPO干预后的SCr、BUN、 CHOP以及全身炎症因子MCP-1、IL-1β、 IL-6和TNF-α相较生理盐水对照组均有下降趋势。与IR+NS组比较,IR+EPO组的SCr、CHOP于再灌注3小时、24小时具有统计学意义(P0.05),BUN、MCP-1、IL-1β、IL-6于再灌注24小时具有统计学意义(P0.05)。(3)CHOP在再灌注1小时组中,缺血再灌注组与假手术组无明显的差异；在再灌注3、6和24小时组中,IR+NS组与Sham+NS组间具有显著性差异(P0.05) 结论：(1)缺血再灌注损伤对大鼠肾组织及肾功能造成显著损害。(2)EPO预处理后,损伤组SCr、BUN及全身炎症因子MCP-1、IL-1β、IL-6和TNF-a均出现下降趋势。(3)急性缺血再灌注可以诱导内质网应激,EPO下调了ERS标志蛋白CHOP的水平。EPO的肾脏保护作用可能同抑制ERS有关,此需要进一步研究。
[Abstract]:Background: acute renal failure (ARF) is characterized by acute renal function change, one of the main causes is acute renal tubular injury (ART), which is one of the common clinical diseases with high overall mortality. At present, support or replacement therapy is the main cause. Erythropoietin (Erythropoietin, EPO) promotes erythrocyte survival, proliferation and differentiation by binding EPO receptor (EPOR), on erythroid cell surface. In recent years, EPOR. has been found on the surface of nerve cells, cardiomyocytes and renal tubular epithelial cells. It is found that EPO has some therapeutic effects on cerebral ischemia and myocardial infarction. In recent years, more and more studies on the role of EPO in renal ischemia-reperfusion have been carried out. The protective mechanism of EPO may be the activation of JAK2 downstream anti-apoptosis pathway. Endoplasmic reticulum stress (Endoplasmic reticulum stress,ERS) is a common response of tissue cells to external stimuli at the subcellular level. The initiation of PERK,ATF6 and IRE1 signaling pathway mainly through unfolded protein response (Unfolded protein response,UPR) may have two sides and play an early role in cell protection. In the late stage, apoptosis was promoted by CHOP,JNK and Caspase12. Objective: to investigate the protective effect of EPO on acute renal injury induced by ischemia and reperfusion and the effect of EPO on endoplasmic reticulum stress and systemic inflammatory factors. Methods: 104 male SD rats with 300-350 g were randomly divided into three groups. According to the intervention conditions, they were divided into three groups: sham operation saline group (Sham NS,n=20), sham operation administration group (Sham EPO,n=20), ischemia reperfusion saline group (IR NS,n=32) and ischemia-reperfusion administration group (IR EPO,n=32). There were 5 rats in sham operation group and 8 rats in ischemia reperfusion group at each reperfusion time point. EPO or NS. were given in groups 2 hours before surgery. In the true operation group, bilateral renal pedicles were subjected to reperfusion after 90 minutes of ischemia. The blood and kidney tissues of inferior vena cava were collected and killed after reperfusion for 6 and 24 hours. The laboratory assists in the detection of serum creatinine and urea nitrogen in venous blood. HE was performed in the right kidney, CHOP, was detected by immunohistochemistry, and serum MCP-1,IL-1 尾 -IL-6 and TNF- 伪 were measured by ELISA. Results: (1) compared with Sham NS group, the levels of SCr and BUN in IR NS group increased significantly at 1 hour after reperfusion (P0. 001), and gradually increased with the prolongation of reperfusion time. The levels of SCr,BUN, CHOP, MCP-1,IL-1 尾, IL-6 and TNF- 伪 were decreased compared with those of normal saline control group. Compared with IR NS group, the SCr,CHOP of IR EPO group had statistical significance at 3 hours and 24 hours after reperfusion (P0.05). (P0.05) there was no significant difference between ischemia reperfusion group and sham operation group in CHOP at 24 hours after reperfusion (P0.05). (3). There was significant difference between IR NS group and Sham NS group in 3h and 24h reperfusion group (P0.05) conclusion: (1) Ischemia-reperfusion injury caused significant damage to renal tissue and renal function in rats. (2) after EPO preconditioning, there was significant damage to renal tissue and renal function in rats. (3) acute ischemia-reperfusion could induce endoplasmic reticulum stress to down-regulate the level of ERS marker CHOP. The renal protective effect of EPO may be related to the inhibition of ERS, which needs further study.
【学位授予单位】：北京协和医学院
【学位级别】：博士
【学位授予年份】：2010
【分类号】：R-332;R692.5

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