黄芩苷对低氧诱导的新生大鼠肺损伤的保护作用

Protective effect of baicalin on hypoxia-induced lung injury of newborn rats

目的探究黄芩苷(baicalin)对低氧诱导的新生大鼠肺损伤的保护作用及其机制。方法SPF级3~5 d新生SD大鼠在10%O2和0.5%CO2低氧舱诱导肺损伤模型,每天低氧8 h,常氧16 h。低氧期每天用自动血气分析仪测动脉血氧合指数(PaO2/FiO2),PaO2/Fi O2<300为发生肺损伤。低氧诱导起,同时ig给予黄芩苷25,50和100 mg·kg-1,连续14 d。测量肺湿/干重比值,HE染色检测肺损伤并进行评分,TUNEL染色检测肺组织细胞凋亡率,ELISA检测大鼠血清中炎症因子肿瘤坏死因子α(TNF-α),白细胞介素6(IL-6),IL-4和IL-10水平,Western印迹检测肺组织中活化胱天蛋白酶3、活化胱天蛋白酶9、Bcl-2、Bax、TNF-α和IL-10蛋白表达及蛋白激酶B(Akt)和NF-κB P65蛋白磷酸化水平。结果与正常对照组相比,低氧组大鼠肺湿/干重比值和肺损伤评分显著升高(P<0.01);HE染色显示肺组织结构紊乱,肺间质及肺泡腔内有炎症细胞浸润;肺细胞凋亡率显著升高(P<0.01);大鼠血清TNF-α,IL-6,IL-4和IL-10水平显著升高(P<0.01),肺组织TNF-α和IL-10蛋白表达显著上调(P<0.01);活化胱天蛋白酶3、活化胱天蛋白酶9和Bax蛋白表达显著上调(P<0.01),Bcl-2表达显著下调(P<0.05);Akt和NF-κB P65蛋白磷酸化水平显著升高(P<0.01)。与低氧组相比,低氧+黄芩苷50和100 mg·kg-1组肺湿/干重比值和肺损伤评分显著降低(P<0.01),肺组织损伤减轻,肺细胞凋亡率显著降低(P<0.01);大鼠血清TNF-α和IL-6表达显著下调(P<0.01),IL-4和IL-10表达显著上调(P<0.01),肺组织中TNF-α蛋白表达显著下调(P<0.01),IL-10蛋白表达显著上调(P<0.01),肺组织中活化胱天蛋白酶3、活化胱天蛋白酶9和Bax蛋白表达显著下调(P<0.01),Bcl-2蛋白表达显著上调(P<0.01),Akt和NF-κB P65蛋白磷酸化水平显著降低(P<0.01)。结论黄芩苷对低氧诱导的新生大鼠肺损伤具有保护作用,并通过调节Akt和NF-κB P65蛋白磷酸化抑制炎症反应和细胞凋亡。

OBJECTIVE To explore the protective effect and mechanism of baicalin on lung injury induced by hypoxia in newborn rats.METHODS A lung injury model was induced in 3-5-d-old SD rats of SPF grade in a 10%O2,0.5%CO2 hypoxic chamber where hypoxia lasted for 8 h and normoxia for16 h.An oxygenation index of arterial blood(Pao2/Fio2)<300 was determined as the symptom of lung injury.Baicalin 25,50 and 100 mg·kg-1 was ig given for 14 d.Lung wet/dry(W/D)mass ratio was measured,while lung injury was detected and scored by HE staining.TUNEL staining was employed to detect the apoptosis rate of lung tissue,ELISA was performed to detect the expressions of tumor necrosis factorα(TNF-α),interleukin-6(IL-6),IL-4 and IL-10 levels in serum,and Western blotting was used to detect the protein expressions of cleaved-caspase 3,cleaved-caspase 9,Bax,Bcl-2,TNF-αand IL-10 and phosphorylation levels of protein kinase B(Akt)and NF-κB P65 protein in lung tissue.RESULTS Compared with the normal control group,the lung W/D mass ratio and lung injury score of the hypoxia group were significantly increased(P<0.01).HE staining showed the rat lung tissue structure was disordered,and there was inflammatory cell infiltration in the lung interstitium and alveolar cavity.The lung cell apoptosis rate was significantly increased(P<0.01),the expressions of TNF-α,IL-6,IL-4 and IL-10 in rat serum were significantly increased(P<0.01),the expressions of TNF-αand IL-10 in lung tissue were significantly increased(P<0.01),the expressions of cleaved-caspase 3,cleaved-caspase 9 and Bax proteins were significantly up-regulated(P<0.01),the expression of Bcl-2 protein was significantly down-regulated(P<0.01),and the phosphorylation levels of Akt and NF-κB P65 were significantly increased(P<0.01).Compared with the hypoxia group,the lung W/D mass ratio and lung injury score were reduced significantly in hypoxia+baicalin 50 and 100 mg·kg-1 group(P<0.01);lung tissue was significantly less damaged,lung cell apoptosis rate was significantly reduced(P<0.05),the expressions of TNF-αand IL-6 were significantly down-regulated(P<0.01)and the expressions of IL-4 and IL-10 were significantly up-regulated in serum(P<0.01),the expression of TNF-αwas significantly down-regulated and the expression of IL-10 was significantly up-regulated(P<0.01),the expressions of cleaved-caspase 3,cleaved-caspase 9 and Bax proteins were significantly down-regulated(P<0.01),the expression of Bcl-2 protein was significantly up-regulated(P<0.01)and the phosphorylation levels of Akt and NF-κB P65 proteins were significantly reduced(P<0.01).CONCLUSION Baicalin has protective effects on hypoxia-induced lung injury in newborn rats,and can inhibit inflammation and apoptosis by regulating Akt and NF-κB P65 proteins phosphorylation.