DRSAb对缺血再灌注损伤大鼠肾脏的保护作用

doi:10.6040/j.issn.1671-7554.0.2017.493

摘要/Abstract

摘要： 目的研究钠钾ATP酶DR区特异性抗体(DRSAb)对肾脏缺血再灌注损伤的保护作用及其机制。方法合成DR区多肽,免疫SD雄性大鼠制备特异性抗血清。采用Western blotting和流式细胞技术测定DRSAb的生物学活性、细胞信号传导机制;采用MTT法测定肾小管上皮细胞HK-2活性;采用缺血再灌注损伤模型检测DRSAb对缺血再灌注损伤大鼠肾脏的保护作用。结果 90.02%的HK2细胞可以和所制备的DRSAb结合;DRSAb免疫血清可增强HK-2细胞对缺氧的耐受能力,与对照血清比较差异有统计学意义(OD: 0.50±0.03 vs 0.10±0.02,P<0.001);DRSAb可以激活PI3K/AKT激酶和PKCε激酶(P<0.001),LY294002和PEAVSLKPT可以抑制这种激活作用(P<0.001);动物实验结果显示,DRSAb组大鼠术后3~6 d血清肌酐、尿素氮水平显著低于Control组(P<0.001)。肾组织切片显示,Control组大鼠肾脏可见明显的血管充血、上皮细胞水肿和小管坏死;DRSAb组的大鼠肾脏可见轻微血管充血、上皮细胞轻度水肿,但无小管坏死。结论 DRSAb对缺血再灌注损伤大鼠肾脏具有一定的保护作用,这种保护作用与PI3K/AKT、PKCε信号通路活化密切相关。

关键词: 缺血再灌注损伤, 钠钾ATP酶, DR区特异性抗体, PI3K/AKT, PKCε

Abstract: Objective To explore the protective effects of Na⁺/K⁺-ATPase DR region specific antibody(DRSAb)on ischemia/reperfusion induced renal injury and its mechanism. Methods Spragur-Dawley(SD)male rats were immunized with synthesised DR region peptides to prepare DRSAb. The biological activity of DRSAb was detected with Western blotting, and the mechanism of cell signal transduction was determined with flow cytometry. The activity of HK-2 cells was assessed with MTT. The protective effect of DRSAb on ischemia-reperfusion injury in rat kidney was evaluated with animal models of ischemia-reperfusion injury. Results The results showed that 90.02% of HK-2 cells could bind with DRSAb. The DRSAb immune sera could significantly enchance the tolerance of HK-2 cells to hypoxia, compared with control seara(OD: 0.50±0.03 vs 0.10±0.02, P<0.001). DRSAb could activate PI3K/AKT and PKCε kinase(P<0.001), which could be inhibited by LY294002 and PEAVSLKPT(P<0.001). Animal experiments indicated that DRSAb group demonstrated a significant improvement in renal function with a lower serum creatinine(sCr)and blood urea nitrogen(BUN)levels 3-6 days postoperative compared with the Control group(P<0.001). Histological evaluation showed that the Control group had visible vascular congestion, epithelial cell swelling and extensive tubular necrosis. 山东大学学报 (医学版)56卷7期 -孙晶晶,等. DRSAb对缺血再灌注损伤大鼠肾脏的保护作用 \=-However, DRSAb group only had slight vascular congestion, mild epithelial cell edema, and no tubular necrosis. Conclusion DRSAb has protective effects against renal ischemia/reperfusion injury in rats, which is associated with the activiation of PI3K/AK and PKCε signalling pathways.

Key words: Ischemia/reperfusion, Na⁺/K⁺-ATPase, DR region specific antibody, PI3K/AKT, PKCε

中图分类号:

R392.7

孙晶晶,张江伟,匡培丹,张颖,薛武军,郑瑾. DRSAb对缺血再灌注损伤大鼠肾脏的保护作用[J]. 山东大学学报 (医学版), 2018, 56(7): 21-27.

SUN Jingjing, ZHANG Jiangwei, KUANG Peidan, ZHANG Ying, XUE Wujun, ZHENG Jin. Protective effects of DRSAb on ischemia reperfusion induced renal injury in rats[J]. Journal of Shandong University (Health Sciences), 2018, 56(7): 21-27.

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