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目的探讨重组腺相关病毒协同表达双基因:抗菌肽(PR39)和肾上腺髓质素(adrenomedullin,ADM),即r AAVPR39-ADM对大鼠脑缺血/再灌注(ischemia/reperfusion,I/R)损伤的治疗作用。方法离体实验:人脐静脉内皮细胞培养,行血管成型实验。在体实验:建立大鼠脑I/R模型(脑缺血2 h后再灌注)。SD大鼠随机分为:Sham组、生理盐水组(I/R+NS)、空病毒组(I/R+AAV)及治疗组(I/R+r AAV-PR39-ADM组)。I/R组,再灌注24 h后股静脉分别给予等量生理盐水、空病毒及r AAV-PR39-ADM。术后1、2、3、4周,行头颅MRI、神经功能缺损评分、TTC染色及HE染色等方法评价r AAV-PR39-ADM对脑I/R损伤的疗效。结果离体实验中,r AAV-PR39-ADM同对照及空病毒组相比,有明显的促血管成型作用。在体实验中,头颅MRI检查证实模型制备成功。与Sham组相比,各时间点I/R组的神经功能缺损评分及脑组织梗塞的体积均明显增高(P<0.01),I/R组术后即刻神经功能缺损评分差异无统计学意义(P>0.05)。与生理盐水组及空病毒组相比,治疗组各时间点神经功能缺损评分及梗塞体积均明显减小(P<0.01),空病毒组与生理盐水组比较,梗塞面积及神经功能缺损评分差异无统计学意义(P>0.05)。HE染色显示,治疗组的神经元细胞数目与生理盐水组和空病毒组相比明显丰富,缺血区新生血管明显增多。结论 r AAV-PR39-ADM可能通过促进血管新生,改善脑I/R后局部供血及侧枝循环,保护神经元,促进血管再生,减轻I/R脑损伤。
OBJECTIVE: To investigate the effect of recombinant adeno-associated virus (AAV) co-expression on the expression of PR39 and adrenomedullin (ADM) in rat brain after cerebral ischemia / reperfusion (I / R) Therapeutic effect of injury. Methods In vitro experiments: human umbilical vein endothelial cells cultured, angioplasty experiments. In vivo experiment: The rat brain I / R model was established (reperfusion after 2 h of cerebral ischemia). SD rats were randomly divided into sham group, saline group (I / R + NS), empty virus group (I / R + AAV) and treatment group (I / R + r AAV-PR39-ADM group). I / R group, equal volume of saline, empty virus and rAAV-PR39-ADM were given to the femoral vein 24 hours after reperfusion. At 1, 2, 3 and 4 weeks after operation, the effect of rAAV-PR39-ADM on brain I / R injury was evaluated by cranial MRI, neurological deficit score, TTC staining and HE staining. Results In vitro, rAAV-PR39-ADM showed obvious angiogenesis compared with control and empty virus group. In vivo experiments, head MRI examination confirmed the successful preparation of the model. Compared with Sham group, the neurological deficit score and brain infarct volume of I / R group were significantly increased at each time point (P <0.01), there was no significant difference in I / R neurological deficit scores immediately after operation P> 0.05). Compared with the saline group and the empty virus group, the neurological deficit score and the infarct volume of the treated group were significantly decreased at each time point (P <0.01). Compared with the saline group, the difference of infarct size and neurological deficit scores No statistical significance (P> 0.05). HE staining showed that the number of neurons in the treated group was significantly higher than that in the saline group and the empty virus group, and the number of neovascular cells in the ischemic area was significantly increased. Conclusions r AAV-PR39-ADM may protect neurons, promote angiogenesis and alleviate I / R brain injury by promoting angiogenesis, improving local blood supply and collateral circulation after cerebral I / R.