松果菊苷对严重烧伤大鼠急性肾损伤的保护作用及其机制

Protective effects and mechanism of echinoside against acute kidney injury in severely burned rats

目的研究松果菊苷(echinacoside,ECH)对严重烧伤大鼠急性肾损伤的保护作用及其分子机制。方法将60只8周龄雄性SD大鼠随机均分为4组:假烧伤组(sham)、烧伤组(burn)、烧伤+松果菊苷处理组(burn+ECH)和烧伤+松果菊苷处理+抑制剂组(burn+ECH+3-TYP)。burn组、burn+ECH组和burn+ECH+3-TYP组大鼠通过背部烫伤法构建Ⅲ度烧伤大鼠模型。burn+ECH组和burn+ECH+3-TYP组大鼠给予腹腔注射ECH(100 mg/kg)药物处理,且burn+ECH+3-TYP组大鼠在给药处理的同时给予3-TYP(50 mg/kg)腹腔注射。各组大鼠处理完成后48 h,采集大鼠血清以酶联免疫吸附方法检测其肾脏功能(BUN和Cr水平),HE染色观察肾组织形态结构改变,利用特定试剂盒检测肾组织氧化应激水平(SOD、GSH-Px、MDA和gp91 phox表达),实时定量PCR检测肾组织炎症反应水平(IL-1β、IL-6、TNF-α和ICAM-1表达),TUNEL染色检测肾组织细胞凋亡情况,Western blot检测SIRT3通路蛋白表达情况。结果与sham组相比,burn组大鼠肾组织大量炎性细胞浸润,肾小管肿胀伴上皮变性坏死,血清BUN和Cr水平明显升高(P<0.05),肾组织中炎症因子IL-1β、IL-6、TNF-α和ICAM-1的表达明显上升(P<0.05),抗氧化物SOD和GSH-Px的含量显著减低,膜脂过氧化产物MDA含量及氧化应激相关蛋白gp91 phox表达显著增加(P<0.05),同时大鼠肾组织细胞凋亡率明显升高(P<0.05),并伴有SIRT3蛋白表达及去乙酰化活性减低(P<0.05)。与burn组相比,burn+ECH组大鼠肾组织病理改变明显减轻,血清BUN和Cr水平明显减少(P<0.05),炎症因子IL-1β、IL-6、TNF-α和ICAM-1的表达明显下降(P<0.05),SOD和GSH-Px的含量显著升高,MDA含量及gp91 phox表达显著减低(P<0.05),肾组织细胞凋亡明显减少(P<0.05),并伴有SIRT3蛋白表达及去乙酰化活性上升(P<0.05)。而与burn+ECH组相比,burn+ECH+3-TYP组大鼠肾功能明显减退,肾组织氧化应激损伤和炎症反应显著增强,肾组织细胞凋亡率显著上升,同时SIRT3信号明显被抑制(均P<0.05)。结论松果菊苷通过抑制氧化应激损伤、炎症反应和凋亡减轻严重烧伤大鼠急性肾损伤,其分子机制与激活SIRT3信号通路相关。

Objective To investigate the protective effects of echinacoside(ECH)against acute kidney injury(AKI)in severely burned rats and explore its molecular mechanism.Methods Sixty 8-week-old SD rats were randomly divided into four groups:sham group,burn group,burn+ECH group and burn+ECH+3-TYP group.The severe burn rats in burn group,burn+ECH group and burn+ECH+3-TYP group were constructed by subjecting rats to third degree burn comprising 30%of the total burned surface area(TBSA)on the back.Rats in burn+ECH group and burn+ECH+3-TYP group were intraperitoneally injected with ECH(100 mg/kg),and rats in burn+ECH+3-TYP group were intraperitoneally injected with 3-TYP(50 mg/kg)at the same time.All tests were exa-mined 48 h after the treatment.Serum was collected to detect renal function(serum BUN and Cr levels)by enzyme linked immunosorbent assay.Histopathological changes of renal tissue were observed by HE staining.Oxidative stress(SOD,GSH Px,MDA and gp91 phox expression)in renal tissue was detected by corresponding kits.Inflammatory reaction(IL-1β,IL-6,TNF-αand ICAM-1 levels)in renal tissue was detected by real-time quantitative PCR.The expression of SIRT3 pathway protein was detected by Western blot.Results Compared with sham group,serum BUN and Cr levels were significantly increased in rats in burn group(P<0.05),with significantly tubule swelling and inflammatory cell infiltration in renal tissue.Compared with sham group,IL-1β,IL-6,TNF-αICAM-1 and MDA contents were significantly elevated in burn group,SOD and GSH-Px levels in renal tissue were decreased in burn group(P<0.05),the cell apoptosis was significantly increased in burn group,and the expression and the deacetylation activity of SIRT3 were inhibited(P<0.05).Compared with burn group,serum BUN and Cr levels were significantly decreased in rats in burn+ECH group(P<0.05),with significantly relieved pathological changes of renal tissue,IL-1β,IL-6,TNF-αICAM-1 and MDA contents were reduced(P<0.05),SOD and GSH-Px levels in renal tissue in burn+ECH group were increased(P<0.05),the cell apoptosis was decreased(P<0.05),the activated expression and the deacetylation activity of SIRT3 were increased(P<0.05).Compared with burn+ECH group,the renal function was alleviated,the oxidative stress and the inflammatory response in renal tissue were increased in burn+ECH+3-TYP group,the apoptosis rate was increased,and the SIRT3 signaling was inhibited(all P<0.05).Conclusion ECH alleviates AKI in severely burned rats by inhibiting oxidative stress,inflammation and apoptosis,which is mediated partly through the activation of SIRT3 signaling.