血红素氧合酶-1介导NAD表达在大鼠内毒素急性肺损伤中的作用

目的探讨血红素氧合酶-1(HO-1)介导烟酰胺腺嘌呤二核苷酸(NAD)表达在大鼠内毒素急性肺损伤中的作用。方法清洁级健康雄性SD大鼠40只,体重200~220 g,8周龄,采用随机数字表法分为四组:对照组(C组)、内毒素急性肺损伤组(L组)、HO-1激动剂Hemin+内毒素急性肺损伤组(H组)和HO-1阻断剂ZnPP-IX+内毒素急性肺损伤组(Z组),每组10只。L组、H组和Z组采用尾静脉缓慢注射脂多糖(LPS)5mg/kg溶于生理盐水1 ml,制备大鼠内毒素急性肺损伤模型。H组于模型前1 h腹腔注射Hemin 50mg/kg,用0.1 mol/L NaOH溶液稀释至1 ml。Z组于模型前1 h腹腔注射ZnPP-IX 10μmol/kg,50 mmol/L NaHCO3溶液稀释至1 ml。给予LPS后6 h处死大鼠,采集动脉血行血气分析,留取肺组织,观察病理学结果并行肺损伤评分,计算肺组织湿/干重比(W/D),采用NAD/NADH定量检测试剂盒测定NAD含量,Western blot法检测HO-1蛋白含量。结果与C组比较,L组、H组、Z组pH和PaO2明显降低,PaCO2、肺损伤评分、肺组织NAD含量、HO-1蛋白含量明显升高(P<0.05),肺W/D比值明显增大(P<0.05),肺组织病理损伤明显。与L组比较,H组pH和PaO2明显升高,PaCO2、肺损伤评分明显降低(P<0.05),肺W/D比值明显减小(P<0.05),肺组织NAD含量、HO-1蛋白含量明显升高(P<0.05),肺组织病理损伤减轻;Z组pH和PaO2明显降低,PaCO2、肺损伤评分明显升高(P<0.05),肺W/D比值明显增大(P<0.05),肺组织NAD含量、HO-1蛋白含量明显降低(P<0.05),肺组织病理损伤加重。与H组比较,Z组pH、PaO2明显降低,PaCO2明显升高(P<0.05),Z组肺损伤评分明显升高(P<0.05),肺W/D比值明显增大(P<0.05),肺组织NAD含量明显降低(P<0.05),肺组织病理损伤明显。结论内毒素肺损大鼠通过上调HO-1表达水平而增加肺组织NAD含量,降低了肺组织炎症反应,从而减轻大鼠内毒素急性肺损伤。

Effect of heme oxygenase-1-mediated NAD levels in endotoxin-induced acute lung injury in rats

Objective To evaluate the effect of heme oxygenase-1(HO-1)-mediated nicotinamide adenine dinucleotide(NAD) levels in endotoxin-induced acute lung injury in rats. Methods Forty male Sprague-Dawley rats, weighting 200-220 g, aged 8 weeks, were randomly divided into 4 groups(n = 10): control group(group C), lipopolysaccharide(LPS)-induced acute lung injury group(group L), HO-1 agonist hemin plus LPS-induced acute lung injury group(group H), and HO-1 antagonist ZnPP-IX plus LPS-induced acute lung injury group(group Z). LPS 5 mg/kg(dissolved in 1 ml 0.9% normal saline) was injected via caudal vein in group L, group H and group Z to establish the model of endotoxin-induced acute lung injury. The rats were intraperitoneally pretreated with 50 mg/kg hemin(dissolved in 0.1 mol/L NaOH solution)in group H, and 10 μmol/kg ZnPP-IX(dissolved in 50 mmol/L NaHCO3 solution) in group Z 1 h before LPS injection. The rats were sacrificed 6 h after LPS administration. The arterial blood was collected for blood gas analysis. The lung tissue was collected for microscopic examination, evaluation of pathological injury scores, calculation the ratio of wet weight to dry weight(W/D), measurement of NAD content by NAD/NADH quantitative detection kit and determination of HO-1 protein expression by western blot. Results Compared with group C, the pH and PaO2 of group L, group H and group Z were significantly decreased;the PaCO2, lung injury scores, W/D ratio, NAD content, and the HO-1 protein expression in group L, group H and group Z were significantly increased(P < 0.05), the pathological changes of lung tissue in group L, group H and group Z were obvious damage. Compared with group L, the pH and PaO2 in group H were significantly increased;the PaCO2, lung injury score, and lung W/D in group H were significantly reduced(P < 0.05);the NAD content and HO-1 protein expression in the lung tissue of group H were significantly increased(P < 0.05), the pathological changes of lung tissue in group H was alleviated. Compared with group L, the pH and PaO2 in the group Z were significantly reduced;the PaCO2, lung injury score, and lung W/D in group Z were significantly increased(P < 0.05), and the NAD content and HO-1 protein expression in the lung tissue of group Z were significantly reduced(P < 0.05),the pathological damage of lung tissue in group Z was more serious. Compared with group H, pH and PaO2 of group Z were significantly decreased(P < 0.05), PaCO2 was significantly increased(P < 0.05), the ratio of W/D in lung increased significantly(P < 0.05), and the content of NAD in lung decreased significantly, the pathological changes of lung tissue was obvious damage. Conclusion By up regulating the expression of HO-1 in endotoxin induced lung injury rats, the content of NAD in lung tissue was increased, and the inflammatory response in lung tissue was reduced, so as to reduce the acute lung injury induced by endotoxin.