不同疗程高压氧治疗对大鼠肾脏缺血再灌注损伤的保护作用对比分析

Comparison of protective effects of different courses of hyperbaric oxygen therapy on renal ischemia-reperfusion injury in rats

目的探讨不同疗程高压氧(hyperbaric oxygen,HBO)治疗对大鼠肾脏缺血再灌注损伤(IRI)的保护作用。方法按照随机数字表法将40只健康成年SD大鼠分为假手术组(A组:n=8)、肾缺血再灌注组(B组:n=8)、肾缺血再灌注+高压氧治疗组(C组:n=24);根据高压氧疗程不同,C组又进一步分为1疗程组(C1组:n=8)、2疗程组(C2组:n=8)、3疗程组(C3组:n=8)。B组、C组采用切除右肾、结扎左肾血管再灌注建立肾脏IRI大鼠模型,A组单纯切除右肾。苏木精-伊红(HE)染色观察肾脏病理学改变,生化试剂盒检测血尿素氮(BUN)、肌酐(Scr)水平,酶联免疫吸附试剂盒检测血清白细胞介素6(IL-6)、白细胞介素8(IL-8)、肿瘤坏死因子α(TNF-α)水平,蛋白免疫印迹检测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)、胞内磷脂酰肌醇激酶(PI3K)、磷酸化PI3K(p-PI3K)、蛋白激酶B(AKT)、磷酸化AKT(p-AKT)等蛋白表达水平。结果与A组相比,B组大鼠HE染色结果提示肾显微结构明显损伤,血清BUN、Scr、IL-6、IL-8、TNF-α水平显著升高(P<0.05),肾脏组织NGAL、KIM-1表达水平显著增加(P<0.05),p-PI3K/PI3K、p-AKT/AKT值显著降低(P<0.05);与B组相比,C1、C2、C3组大鼠HE染色结果提示肾显微结构损伤减轻,血清BUN、Scr、IL-6、IL-8、TNF-α水平显著降低(P<0.05);肾脏组织NGAL、KIM-1表达水平显著下降(P<0.05),p-PI3K/PI3K、p-AKT/AKT值显著提升(P<0.05),且随疗程增加,效应增强。结论HBO治疗能够通过调控PI3K/AKT信号通路减轻肾脏IRI大鼠炎性反应,降低肾脏损伤标志物NGAL、KIM-1表达水平,改善肾功能,且随着HBO疗程增加,效应增强。

Objective To compare the protective effects of different courses of hyperbaric oxygen(HBO)therapy on renal ischemia-reperfusion injury(IRI)in rats.Methods According to the random number table method,a total of 40 healthy adult SD rats were divided into sham operation group(group A,n=8),renal IRI group(group B,n=8),and renal IRI+HBO treatment group(group C,n=24).According to different courses of HBO treatment,group C was further divided into one-course-treatment group(group C1,n=8),two-course-treatment group(group C2,n=8),and three-course-treatment group(group C3,n=8).The rat model of renal IRI in group A was established by removing the right kidneys of rats,while such models in groups B and C were established by removing the right kidneys,ligating the left renal blood vessels,and then performing reperfusion.Hematoxylin and Eosin(H&E)staining was used to observe the pathological changes of the kidney.The biochemical kit was used to detect the levels of blood urea nitrogen(BUN)and serum creatinine(Scr).The enzyme-linked immunosorbent assay(ELISA)kit was used to detect the serum levels of interleukin 6(IL-6),interleukin 8(IL-8),and tumor necrosis factor alpha(TNF-α).Western blotting was used to detect the expression levels of neutrophil gelatinase-associated lipocalin(NGAL),kidney injury molecule-1(KIM-1),intracellular phosphatidylinositol kinase(PI3K),and protein kinase B(PKB/Akt).Results Compared with group A,the results of H&E staining in group B showed obvious damage of renal microstructure,the serum BUN,Scr,IL-6,IL-8,and TNF-αlevels were significantly increased(P<0.05),the expression levels of NGAL and KIM-1 in kidney tissues were significantly increased(P<0.05),and the values of p-PI3K/PI3K and p-AKT/AKT in kidney tissues were significantly decreased(P<0.05);while compared with group B,the results of H&E staining of rats in groups C1,C2,and C3 showed reduced damage of renal microstructure,and the levels of serum BUN,Scr,IL-6,IL-8,and TNF-αwere significantly decreased(P<0.05),the expression levels of NGAL and KIM-1 in kidney tissues were significantly decreased(P<0.05),and the values of p-PI3K/PI3K and p-AKT/AKT were significantly increased(P<0.05),and with the increase of HBO treatment courses,better improvement in the above indicators were shown in groups C1,C2,and C3.Conclusion By regulating the PI3K/AKT signaling pathway,HBO therapy can relieve the inflammatory response,reduce the expression levels of kidney injury markers NGAL and KIM-1,and improve renal function in renal IRI rats,and such effects are enhanced with the increase of HBO treatment course.