利用RNAi抑制PARs防治脑缺血性损伤的实验研究

目的：利用RNA干扰技术，通过重组腺病毒导入，抑制脑组织中PARs基因的表达，观察神经功能缺陷评分、缺血脑组织梗死体积的变化，为缺血性脑血管疾病的基因治疗提供实验依据。方法：雄性Wistar大鼠随机分组。以重组腺病毒介导的无序PARs基因shRNA片段、生理盐水进行干预，干预3天后以线栓法建立Wistar大鼠大脑中动脉永久闭塞模型，各组分别于线栓后24h、72h进行神经功能缺陷评分并断头取脑，应用4％TTC染色测脑梗死体积。结果：1．缺血后24h、72h同一时间点：①实验组分别与对照组、生理盐水组对比，神经功能缺陷评分明显降低（P〈0．05），脑梗死体积明显减小（P〈0．05）；②对照组与生理盐水组对比，神经功能缺陷评分、脑梗死体积无明显差异（P〉0．05）。2．各组组内缺血后24h、72h对比，神经功能缺陷评分、脑梗死体积各组均有明显差异（P〈0．05）。结论：重组腺病毒介导的RNAi能有效抑制脑组织中PARs基因的表达，缩小脑梗死体积，改善神经功能缺陷评分。

Research on Inhibition of PARs Gene Expression by RNAi Attenuates Ischemic Brain Injury

Objective： To investigate the neuron-protection effects of brain ischemia on gene expression inhibition of recombinant adeno-associated virus （rAAV）-mediated gene transfer to ischemic brain, and to provide basic evidence for gene therapy on cerebral ischemia. Meflaod： 84 adult male Wistar rats between 16 and 20 weeks of age and weighing 220 to 280g were randomly divided into three groups： the experimental group （EG）, the control group （CG）, the physiological sodium group （PSG）. The rats were given rAAV-RPARs-shRNA in proper sequence, rAAV-RPARs-shRNA in disturbed sequence, sodium chloride 3 days before performing mid- dle cerebral artery occlusion （MCAO）. 24 or 72 hours after ischemia all rats were performed neurological evaluation and then decapitated. We determined brain infarct volume by 2,3,5-triphenyltetrazolium chloride （TTC） staining. Result： （L）At the same time point, 24 or 72 hours aider ischemia, the grade of neuro-functional deficit in the experiment group （EG） decreased compared with that in the control group （CG）（P〈0.05）, and the volume of cerebral infarction was strikingly diminuted （P〈0.05）; the grade of neuro-functional deficit in the experiment group （EG） decreased compared with that in the physiological saline group （PSG） （P〈0.05）, and the volume of cerebral in- farction was strikingly diminuted （P〈0.05）; But when we compared the CG with the PSG, we found that there were no obvious differ- ences of indexes above between these two groups （P〉0.05）.（2）At different time point the grade of neuro-functional deficit decreased com- pared with each other inner groups （P〈0.05）, and the volume of cerebral infarction was strikingly diminuted （P〈0.05）; Conclusions： rAAV-mediated RNA interference could actively suppress the expression of PARs in brain tissues, thereby diminute cerebral infarction volume and improve the grade of neuro-functional deficit.