基于NF-κB信号途径的褪黑素对SD大鼠肾脏热缺血及再灌注损伤的疗效及其机制研究

Role of melatonin in NF-κB signal pathway-mediated inflammation after kidney warm ischemia-reperfusion in rats

目的探讨褪黑素抑制炎性反应和减轻肾脏缺血再灌注损伤的可能机制。方法64只成年雄性SD大鼠分为溶媒治疗的假手术组(Sham组)、褪黑素治疗的假手术组(M+Sham组)、溶媒治疗的缺血再灌注损伤组(I/R组)、褪黑素治疗的缺血再灌注损伤组(M+I/R组),褪黑素或溶媒于术前15 min经腹腔注射。检测各组大鼠血清中尿素氮(BUN)和血肌酐(Scr)水平,苏木素-伊红(HE)染色观察肾组织病理变化,原位末端标记法(TUNEL)检测肾小管上皮细胞(TEC)凋亡情况,免疫组织化学检测肾组织中白细胞介素-6(IL-6)和肿瘤坏死因子-α(TNF-α)的表达水平,Western blot评估核因子-κB(NF-κB)信号通路活性和下游炎症因子IL-6的表达水平,观察术后24 h生存率。结果与I/R组比较,M+I/R组大鼠术后生存率有所提高,血清BUN和Scr水平明显降低,肾组织病理损伤明显改善,TEC凋亡数量明显减少,IL-6和TNF-α的表达水平明显降低,差异均有统计学意义(P<0.05)。M+I/R组大鼠p-NF-κBp65/t-NF-κBp65、p-IκB-α/t-IκB-α的比值和IL-6的表达水平较I/R组明显降低,差异有统计学意义(P<0.05)。结论褪黑素可下调NF-κB信号通路活性,抑制炎性反应,降低细胞凋亡水平,减轻肾脏缺血再灌注损伤。

Objective To explore the possible mechanisms of melatonin in inhibiting inflammatory response and reducing renal ischemia-reperfusion injury.Methods Sixty-four adult of male SD rats were randomly divided into Sham group(Sham),ischemia-reperfusion injury group(I/R),melatonin treatment Sham group(M+Sham),and melatonin treatment I/R group(M+I/R).Melatonin or solvent was injected intraperitoneally 15 min before surgery.Levels of serum urea nitrogen(BUN)and serum creatinine(Scr)were detected in each group,HE staining was used to observe the tissue pathological changes in kidney,TUNEL assay to detect the apoptosis quantity of renal tubular epithelial cells,and immunohistochemical staining to detect interleukin-6(IL-6)and tumor necrosis factor-α(TNF-α)expression levels,Western blot to assess activity of nuclear factor-κB(NF-κB)signaling pathway and IL-6 expression level.Survival rate was observed at 24 h after surgery.Results Compared with the I/R group,the survival rate in the M+I/R group significantly increased,levels of BUN and Scr notably decreased,pathological damage of renal tissue were significantly mitigated,the apoptosis of renal tubular epithelial cells was significantly reduced,expression levels of IL-6 and TNF-αwere significantly decreased(P<0.05).In comparison with the I/R group,the ratios of p-NF-κBp65/t-NF-κBp65,p-IκB-α/t-IκB-αand IL-6 expression levels were significantly down-regulated in the M+I/R group(P<0.05).Conclusion Melatonin can down-regulate the activity of NF-κB signaling pathway,alleviate inflammation,decrease apoptosis and reduce the renal ischemia-reperfusion injury.