腺病毒介导BTEB2基因转染对动脉损伤后新生内膜增生的影响

目的 :构建反义转录元件结合蛋白 2 (BTEB2 )重组腺病毒载体并研究BTEB2反义RNA对动脉损伤后新生内膜增生的影响。方法 :通过聚合酶链反应 (RT PCR)法从培养的大鼠血管平滑肌细胞中制备BTEB2cDNA ,将其反向克隆至腺病毒载体 ,构建反义BTEB2重组腺病毒 ;用重组腺病毒局部转染球囊损伤的大鼠颈动脉 ,观察反义BTEB2基因转染对BTEB2蛋白表达及损伤动脉新生内膜形成的影响。结果 :构建的BTEB2反义重组腺病毒经鉴定正确 ,其滴度为 5× 10 9/ml;反义BTEB2重组腺病毒转染可明显抑制BTEB2蛋白表达及新生内膜增生。结论 :成功构建了反义BTEB2重组腺病毒载体 ;BTEB2反义RNA可明显抑制大鼠颈动脉球囊损伤后的新生内膜增生。

Effects of BTEB2 antisense RNA mediated by recombinant adenoviral vector on neointima hyperplasia after rat carotid artery injury

Objective:To construct recombinant adenovirus vector of antisense BTEB2 (AdASBTEB2) and study the effects of BTEB2 antisense RNA on neointima hyperplasia after rat carotid artery injury.Method:BTEB2 cDNA was prepared by RT-PCR technique and was subcloned reversedly into adenovirus vector to construct AdASBTEB2. The PCR technique was used to detect target gene fragment and adenovirus genomic characteristic fragment. Antisense BTEB2 gene was transduced into rat carotid arteries with AdASBTEB2 after the establishment of rat carotid balloon injury restenosis model. The BTEB2 expression was detected by immunohistochemistry. The intimal/medial area ratio were measured.Result:The recombinant adenovirus proved correct by PCR. AdASBTEB2 delivered into injured rat carotid arteries reduced BTEB2 protein expression significantly from day 7 to 21 after injury. At day 21 after injury, compared with untreatment and treatment with AdLacZ, treatment with AdASBTEB2 reduced intimal hyperplasia by 49% and 47% respectively.Conclusion:BTEB2 antisense RNA reduced BTEB2 protein expression and intimal hyperplasia induced by arterial endothelium injury.