Apelin/APJ系统在肢体缺血再灌注所致肺损伤中的保护作用

Protective effect of Apelin/APJ system on lung injury induced by limb ischemia-reperfusion in rats

目的探讨Apelin/APJ系统在肢体缺血再灌注(LIR)所致肺损伤中的保护作用及其可能机制。方法将24只成年SD大鼠按照随机数字表法分为假手术组(CON组)、肢体缺血再灌注组(IR组)、Apelin处理组(APL组)、Apelin+APJ拮抗剂组(APLF组),每组6只。除CON组外,其余各组大鼠采用腹主动脉夹闭3 h再灌注3 h制备LIR模型。CON组分离但不夹闭腹主动脉;IR组缺血及再灌注前分别给予0.9%氯化钠注射液0.75 ml/kg;APL组缺血前给予0.75 ml/kg 0.9%氯化钠注射液,再灌注前给予150μg/kg[Pyr1]-Apelin-13;APLF组缺血前给予150μg/kg APJ拮抗剂F13A,再灌注前给予150μg/kg[Pyr1]-Apelin-13。分别于平衡末及再灌注期末进行血气分析,实验终点取动脉血标本和肺组织,检测血浆炎症指标IL-6、TNF-α及氧化应激指标丙二醛(MDA)、超氧化物歧化酶(SOD);检测肺组织炎症指标髓过氧化物酶(MPO)及氧化应激指标(MDA、SOD);计算各组肺组织湿干重比(W/D);采用HE染色法观察肺组织病理结果;采用Western blot法检测肺组织解偶联蛋白2(UCP2)蛋白表达及RT-PCR法检测UCP2 mRNA表达。结果和IR组和APLF组比较,APL组pH值及PaO2值显著升高(均P<0.05),PaCO2值显著降低(P<0.01),肺组织W/D显著降低(均P<0.05),肺泡结构显著改善,血浆IL-6、TNF-α、MDA均显著降低,SOD显著升高(均P<0.01),肺组织MDA、MPO显著降低,SOD显著升高(均P<0.05),同时肺组织UCP2蛋白及mRNA表达显著升高(P<0.05)。结论Apelin/APJ系统可减轻大鼠LIR引起的肺损伤,其机制可能与减轻炎症反应和氧化应激相关,通过上调线粒体UCP2蛋白发挥作用。

Objective To explore the protective effect of Apelin/APJ system on lung injury induced by limb ischemia-reperfusion(LIR)and its mechanism.Methods Twenty-four adult SD rats were randomly divided into four groups(n=6 in each):CON group(sham operation group),IR group(limb ischemia-reperfusion group),APL group(Apelin treatment group),APLF group(Apelin+APJ inhibitor group).LIR model was established by clamping abdominal aorta for 3 h and reperfusion for 3 h.The abdominal aorta was isolated but not clamped in the CON group;the 0.75 ml/kg normal saline was administrated before ischemia and reperfusion in the IR group;the 0.75 ml/kg normal saline was administrated before ischemia and 150μg/kg[Pyr1]-Apelin-13 before reperfusion in the APL group;the 150μg/kg APJ inhibitor F13A was administrated before ischemia and 150μg/kg[Pyr1]-Apelin-13 before reperfusion in the APLF group.Blood gas analysis was carried out at the end of balance and the end of reperfusion.At the end of the experiment,the arterial blood samples and lung tissues were obtained.The plasma inflammatory indexes Interleukin-6(IL-6),tumor necrosis factor-α(TNF-α)and oxidative stress indexes malondialdehyde(MDA),superoxide dismutase(SOD)were detected.The inflammatory indexes myeloperoxidase(MPO)and oxidative stress indexes(MDA,SOD)of lung tissues were detected.The ratio of wet to dry weight(W/D)of lung tissues in each group was calculated.The pathological results of lung tissues were observed through HE staining.The expression of UCP2 protein and mRNA were detected by Western blot and RT-PCR.Results Compared with IR group and APLF group,pH and PaO2 value in APL group increased significantly(all P<0.05),PaCO2 value decreased significantly,W/D value decreased significantly(all P<0.05),alveolar structure improved significantly,plasma IL-6,TNF-α,MDA decreased significantly(P<0.01),SOD increased significantly(P<0.01),MDA and MPO in lung tissue decreased significantly,SOD increased significantly(all P<0.05).At the same time,the expression of UCP2 protein and mRNA in lung tissue increased significantly(P<0.05).Conclusion The Apelin/APJ system can reduce lung damage caused by LIR in rats,which may be associated with the reduced inflammatory response and oxidative stress,through up-regulaiton of mitochondrial UCP2 proteins.