肢体缺血-再灌注大鼠脑组织iNOS基因表达的半定量检测

目的检测大鼠肢体缺血-再灌注后脑组织iNOS基因表达的变化.方法SD大鼠,随机分为肢体缺血-再灌注(I-R)组、肢体单纯缺血(I)组及正常对照(N)组,通过夹闭腹主动脉末端4?h,或/和开放2～24?h,复制I、I-R组动物模型,半定量RT-PCR方法检测脑组织iNOSmRNA表达的变化,免疫组化染色法观测脑组织内iNOS及过氧亚硝基阴离子(ONOO-)的硝基化产物硝基酪氨酸(NT)的生成与分布.结果N组脑组织iNOSmRNA未检出,I组及I-R组脑组织iNOSmRNA均有表达,再灌注2?h,iNOSmRNA表达量显著升高(P<0.01,vsN组),再灌注6?h达到高峰,随后下降,24?h仍有少量表达;I及I-R组脑组织均有iNOS阳性细胞,I-R组见于大脑皮层各区、海马、尾状核,I组仅见于皮层后肢区及尾状核.I-R组脑组织可见弥散分布的NT阳性神经元,I组偶见NT阳性神经元.结论大鼠肢体缺血-再灌后脑组织iNOS基因表达显著增强,且有时相变化特征.

Semiquantitative Detection of iNOS Gene Expression in Brain from Limb Ischemia-Reperfused Rats

Objective To detect the expression of iNOS gene in brain tissues from limb ischemia\|reperfused SD rats.Method SD rats were randomly divided into three groups:limb ischemia\|reperfusion group(I\|R), simple ischemia group(I) and normal control group(N).The expression of iNOS mRNA in brain was semiquantitatively detected by using of RT\|PCR.The distribution and the expression of iNOS and peroxynitrite protein in brain were measured by immunohistochemistry.Results iNOS mRNA was not detected in brain from control rats,while it was detected in both I and I\|R groups.After I\|R, the expression of iNOS mRNA gradually increased at 2h and peaked at 6h( P <0.01 vs I group),then decreased,but there was also a little expression at 24h.The morphologic results showed that there were more iNOS positive cells and more dispersal NT positive neurons in I\|R group than I group.Conclusion The expression of iNOS mRNA in brain significantly increased after limb ischemia\|reperfusion in a time\|dependent manner.