超声及超声微泡介导基因转染的安全性研究

Security of gene delivery mediated by ultrasound and microbubbles

目的:探讨超声及超声微泡介导基因转染的安全性。方法:将人脐静脉内皮细胞分别按超声辐照时长及所加入的微泡剂量分组,超声辐照后分别培养24h及48h,应用台盼蓝拒染法进行细胞计数,与对照组进行比较。超声辐照含有不同微泡剂量的细胞后,行扫描电镜观察细胞膜形态。结果:不同辐照时长组经超声辐照后培养24h,10min、15min、30min组细胞计数比对照组降低,差异有统计学差异(P<0.05);继续培养48h,各组与对照组比较无统计学差异。不同微泡剂量组经超声辐照5min后培养24h,200μl、500μl组细胞计数比对照组降低,差异有统计学差异(P<0.05);继续培养48h,500μl组细胞计数比对照组降低,差异有统计学差异(P<0.05)。扫描电镜可见超声辐照后细胞膜完整性变差,继续培养24h后有一定程度改善。结论:超声辐照+微泡对细胞增殖有一定的抑制作用,对细胞膜有一定程度的损害,但这些作用在超声辐照时长不超过5min及微泡剂量不超过100μl时是轻微、可逆的。超声及超声微泡介导基因转染是相对安全的。

Objective：To investigate the security of ultrasound-and microbubble-mediated gene delivery.Methods：24hor 48 hafter HUVEC,which was divided into different groups according to exposure time or microbubble does,was exposed to ultrasound,the number of living cells was counted by trypan blue exclusion assay,and comparison was made.Then,Observations were done by scanning electron microscopy.Results：After 24h-culture,compared with that in control group,cell count was decreased significantly in 10 min,15min and 30 min group（P〈0.05）,and after 48h-culture,there were no differences among the different exposure-time groups.24h-culture after the cells in different microbubble-does groups were exposed to ultrasound for 5min,compared with that in control group,cell count was decreased significantly in 200μl and 500μl group（P〈0.05）,and after 48h-culture,cell count was decreased significantly in 500μl group（P〈0.05）.Observed by scanning electron microscopy after exposure,we found that the integrity of the cell membrane became worse,and after 24h-culture,it could improve to some extent.Conclusion：Exposure by ultrasound with microbubble can inhibit the proliferation of cell and impaired the cell membrane to some extent.However,these functions are limited and reversible when the exposure time is no more than5 min and the microbubble does is no more than 100μl.Ultrasound-and microbubble-mediated gene delivery is relatively safe.