肢体缺血再灌注后肺组织中NOS的表达

Expression of NOS in lung tissue after the hind limb ischemia reperfusion of rats

目的 观察肢体缺血再灌注后肺组织中NOS同工酶活性、分布和mRNA表达的变化。②方法 复制肢体缺血再灌注模型 ,实验动物随机分为 4组 :正常对照组 (Control组 )、损伤组 (IR组 )、L 精氨酸组 (L Arg组 )和氨基胍组 (AG组 ) ,用免疫组织化学方法检测肺组织中NOS蛋白的表达 ;用RT PCR方法检测肺组织中NOSmRNA的表达。③结果 免疫组织化学结果显示 ,Control组iNOS染色少有阳性 ;IR组iNOS染色阳性 ,染色阳性细胞主要分布在支气管上皮细胞、支气管平滑肌细胞 ,支气管周围浸润的中性粒细胞也呈阳性表达 ;AG组iNOS染色较IR组减弱 ;L Arg组则无明显差异。eNOS染色在Control组呈阳性 ,主要分布于毛细血管内皮细胞 ,其余 3组染色均减弱。RT PCR结果显示 ,iNOSmRNA的表达在Control组较少 ,IR组呈高表达 ,AG组、L Arg组表达与IR组相比无明显差异 ;eNOS的表达在Control组呈高表达 ,其余各组表达均降低。④结论 大鼠肢体缺血再灌注后 4小时 ,肺组织中iN OSmRNA高表达 ,eNOSmRNA表达降低 ,肺组织中NO的水平增高 ;给予外源性的L Arg增加NO的生成 ,应用AG不影响NOSmR NA的表达 。

Objective It was observed that the expression of nitric oxide synthase isoenzymes and their mRNA in lung tissue after the hind limb ischemia reperfusion of the rats.Methods The model rats with limb ischemia and reperfusion(IR) were developed and the NO donor L-arginine(L-Arg) and the NOS inhibitor aminoguanidine (AG) were used in the study.The animals were randomly divided into control group,Ischemia reperfusion group (IR group),L-arginine group(L-Arg group) and aminoguanidine group(AG group).At the end of reperfusion, blood samples and lung samples were taken for measurement of NO 2 -, and the nitric oxide synthase(NOS) activity were observed.The RT-PCR was used to detect the level of the NOS mRNA expression and the results were analyzed quantitatively by Bio-Rad 100 gel-imagine density analytical system.The immuno-cytochemical method was used to detect the level of the NOS expression.Results In IR group, the content of NO 2 - in the plasma and in the lung tissue increased.the NOS activity increased too. RT-PCR showed the low level of eNOS mRNA and the high level of iNOS mRNA. It was demonstrated that the upregulation of iNOS expression and the downregulation of eNOS expression contrasting with Control group by immunohistochemistry method. Strong positive stain of iNOS located in bronchium endothelial, smooth muscle cells and infiltrating PMN in pulmonary interstitial tissue. L-Arg had no effects on expression of iNOS and eNOS and their mRNA. The RT-PCR of eNOS and iNOS revealed similar levels in AG group and IR group, and immuno-histochemically the expression of iNOS in AG group was lower than that in I/R group.Conclusions These results indicated that the upregulation of iNOS expression and the downregulation of eNOS expression after limb ischemia reperfusion of rats. The NO production increased by administering L-Arg and the AG had significant effect on the expression of iNOS protein but not its mRNA.