摘要
目的 :探讨腺病毒介导人肝细胞生长因子 (Ad -HGF)对体外无血清培养所致大鼠皮层神经元损伤的保护作用。方法 :用流式细胞仪测定不同感染强度 (MOI)条件下携带绿色荧光蛋白的重组腺病毒 (Ad GFP) (2 5 ,5 0 ,10 0 ,2 0 0 pfu/cell)对皮层神经元的转染效率 ,确定最佳MOI ;ELISA测定Ad HGF在最佳感染强度下皮层神经元中的表达规律 ;通过中性红染色和PI Hoechst33342双染色法比较转染Ad HGF组与对照组 (转染Ad GFP组和空白对照组 )间无血清培养后 6h、12h、2 4h和 4 8h细胞的存活情况。结果 :在感染强度为 5 0 pfu/cell时 ,重组腺病毒对皮层神经元的转染率达 99.3% ,Ad HGF能在皮层神经元中有效而持久地表达 ,接种后 2h转染Ad HGF组的细胞死亡率和凋亡率在无血清培养后 12h明显低于两对照组 (P <0 .0 5 )。结论 :Ad HGF对接种后 2h转染组无血清培养所致的损伤有显著的保护作用 ,对接种后第 5d转染的皮层神经元无血清所致的损伤虽亦具有一定的保护作用 ,但不明显。
Aim: To investigate the protective effects of adenovirus-mediated hepatocyte growth factor(Ad-HGF) on injury of rat cortex neurons induced by in vitro serum-free culture. Methods: Flow cytometry was used to assay the transfection rate of rat cortex neurons infected by adenovirus-mediated green fluorescent protein(Ad-GFP) at different multiplicity of infection(MOI) to find out the best MOI in experiment. ELISA was used to elucidate the expression patterns of cortex neuron. Neutral red stain and PI-Hoechst 33342 double stain were used to compare the viability of cortex nerons, which were cultured in serum-free medium for 6 h, 12 h, 24 h and 48 h respectively, among the Ad-HGF transfected group, the Ad-GFP transfected group and the control group. Results: It was found that when MOI was 50 PFU per cell, a transfection rate as high as 99.3% was maintained and Ad-HGF was able to express in cortex neurons effectively and persistently. In addition, the death rate and apoptotic rate of cortex neurons(infected 2 hours after seeding)cultured in serum-free medium for 12 h in Ad-HGF transfected group was significantly lower than that in both the Ad-GFP group and the control group (P<0.05). Conclusion: Ad-HGF plays a protective role against in vitro serum-free culture induced injury on rat cortex neurons infected 2 hours after seeding. Though its effects on rat cortex neurons infected 5 days after seeding are not so remarkable, Ad-HGF also has the potential to protect cortex neurons from serum-free culture induced injury.
出处
《中国应用生理学杂志》
CAS
CSCD
北大核心
2004年第2期156-160,共5页
Chinese Journal of Applied Physiology
