Maresin 1对脓毒症相关急性肾损伤的保护作用及机制实验研究

Experimental study on the protective effect of Maresin 1 on sepsis-associated acute kidney injury and its mechanism

目的:研究Maresin 1对脓毒症相关急性肾损伤(S-AKI)的保护作用及相关机制。方法:SPF级雄性SD大鼠,采用盲肠结扎穿孔法(CLP)制备脓毒症大鼠模型。模型大鼠按随机数字表法分成CLP组、CLP+Maresin 1组、CLP+Maresin 1+Nigericin(NLRP3激活剂)组。另取正常大鼠作为Sham组。CLP+Maresin 1组、CLP+Maresin 1+Nigericin组大鼠于术后1 h单次腹腔注射Maresin 1(4 ng/g,溶于200μl 0.9%氯化钠溶液)。CLP+Maresin 1+Nigericin组在给予Maresin 1基础上联合单次Nigericin(1 mg/kg)给药。Sham组和CLP组给予200μl 0.9%氯化钠溶液。12 h后,采血用于血肌酐(Scr)和血尿素氮(BUN)生化指标和血清中性粒细胞明胶酶相关载脂蛋白(NGAL)、肾损伤分子1(KIM1)及炎症因子ELISA检测。HE染色检测肾脏病理改变。免疫组化和Western blot检测NOD样受体热蛋白结构域3(NLRP3)、半胱氨酸蛋白酶-1(Caspase-1)、白介素-1β(IL-1β)、白介素-18(IL-18)表达变化。另取大鼠按照上述方法分组及治疗,统计大鼠7 d存活率。结果:与Sham组相比,CLP组Scr、BUN、尿中性粒细胞明胶酶相关脂质运载蛋白(NGAL)、肾损伤分子-1(KIM-1)及炎症因子水平明显增加,NLRP3、Caspase-1、IL-1β、IL-18表达明显上调,肾脏出现炎性损伤改变,大鼠7 d存活率明显降低(均P<0.05)。与CLP组相比,CLP+Maresin 1组Scr、BUN、NGAL、KIM-1及炎症因子水平明显降低,NLRP3、Caspase-1、IL-1β、IL-18表达明显下调,肾脏炎性损伤减轻,大鼠7 d存活率明显增加(均P<0.05)。然而,CLP+Maresin 1组给予Nigericin后,Maresin 1的保护作用被部分逆转。结论:Maresin 1对S-AKI具有保护作用,该机制与抑制NLRP3/Caspase-1轴有关。

Objective:To study the protective effect of Maresin 1 on sepsis-associated acute renal injury(S-AKI)and its mechanism.Methods:A total of 103 SPF grade male SD rats were used to establish sepsis rat model by cecal ligation and puncture(CLP).The model rats were randomly divided into CLP group(20 rats),CLP+Maresin 1 group(15 rats)and CLP+Maresin 1+Nigericin(NLRP3 activator)group(18 rats).Another 10 normal rats were taken as Sham group.A single intraperitoneal injection of Maresin 1(4 ng/g,dissolved in 200μl 0.9%sodium chloride solution)was given to rats in group CLP+Maresin 1 and group CLP+Maresin 1+Nigericin at 1 hour after operation.In the CLP+Maresin 1+Nigericin group,Maresin 1 was given in addition to a single dose of Nigericin(1 mg/kg).Sham group and CLP group were given 200μl 0.9%sodium chloride solution.After 12 hours,blood samples were collected for Scr and BUN biochemical indexes and ELISA for serum NGAL,KIM1 and inflammatory factors.HE staining was used to detect the pathological changes of kidney.The expression of NLRP3,Caspase-1(IL-1βand IL-18 were detected by immunohistochemistry and Western blot.Another rats were grouped and treated according to the above methods,and the 7-day survival rate of rats was calculated.Results:Compared with the Sham group,in the CLP group,the levels of Scr,BUN,NGAL,KIM-1 and inflammatory factors were significantly increased,and the expressions of NLRP3,Caspase-1,IL-1βand IL-18 were significantly up-regulated(all P<0.05);the kidney showed inflammatory changes;the 7-day survival rate of the rats was significantly decreased(P<0.05).Compared with the CLP group,in the CLP+Maresin 1 group,the levels of Scr,BUN,NGAL,KIM-1 and inflammatory factors were significantly decreased,the expressions of NLRP3,Caspase-1,IL-1βand IL-18 were significantly down-regulated(all P<0.05);the inflammatory injury was alleviated;the 7-day survival rate of rats was significantly increased(P<0.05).However,the protective effect of Maresin 1 was partially reversed after Nigericin administration in the CLP+Maresin 1 group.Conclusion:Maresin 1 has a protective effect on S-AKI,which is related to the inhibition of NLRP3/Caspase-1 axis.