TRESK基因重组腺病毒载体转染大鼠背根神经节神经元的效果

目的 评价TRESK基因重组腺病毒载体(pAD/CMV/V5- DEST-TRESK)转染大鼠背根神经节(DRG)神经元的效果.方法 原代培养大鼠DRG神经元,调整细胞密度为6× 104个/ml.采用随机数字表法,将细胞随机分为3组,每组6孔,每孔1ml细胞悬液,TRESK基因重组腺病毒载体组(T组)每孔加入3×107 TU的pAD/C MV/V5-DEST- TRESK；腺病毒对照组(A组)每孔加入3× 107TU的阴性对照腺病毒( pAD/CMV/V5- DEST)；对照组(C组)不加入腺病毒.于72 h后采用RT-PCR法检测TRESKmRNA表达,Western blot法检测TRESK表达.结果 与C组和A组比较,T组DRG神经元IRESK及其mRNA表达上调(P＜0.05),C组和A组上述指标差异无统计学意义(P＞0.05).结论 pAD/CMV/V5-DEST-TRESK可有效转染大鼠DRG神经元,上调其TRESK表达.

Transfection of rat dorsal root ganglion neurons with TRESK gene recombinant adenovirus vector

Objective To transfect rat dorsal root ganglion (DRG) neurons with TRESK gene recombinant adenovirus vector.Methods Primary cultured DRG neurons were enzymatically isolated from newborn SD rats of both sexes and cultured in DMEM liquid culture medium.The density of the cells in suspension was 6 × 104/ml.The cells were randomly assigned to one of 3 groups ( n =6 wells each):group TRESK gene recombinant adenovirus vector (group T) ; group adenovirus (group A) and group control (group C).In group T TRESK gene recombinant adenovirus vector (pAD/CMV/V5-DEST-TRESK) 3 × 107 TU was added to each well.In group A simple adenovirus 3 × 107 TU was added to each well and in group C no adenovirus was added.After 72 h incubation the expression of TRESK mRNA and protein was detected by RT-PCR and Western blot respectively.Results The expression of TRESK mRNA and protein was significantly up-regulated in group T as compared with groups C and A.Conclusion The rat DRG neurons can be effectively transfected with TRESK gene recombinant adenovirus vector.