反义锁核酸与拉米夫定在HepG2.2.15细胞中抗HBV作用的比较

Inhibitory effect of locked nucleic acid antisense oligonucleotides versus lamivudine on HBV replication in HepG2.2.15 cells

目的:应用反义锁核酸与拉米夫定作用HepG2.2.15细胞,对他们抗乙型肝炎病毒(hepatitis B virus,HBV)效果进行比较.方法:设计针对HBV翻译起始区S基因mRNA的反义寡核苷酸,并进行锁核酸修饰,以阳离子脂质体介导反义锁核酸转染HepG2.2.15细胞;拉米夫定组直接作用HepG2.2.15细胞;分别于用药后第2、4、6、8、10天收集细胞培养上清液.用ELISA法和FQ-PCR法检测收集上清液HBsAg、HBeAg和HBVDNA的含量.MTT法分别检测反义锁核酸与拉米夫定对细胞存活率的影响.结果:拉米夫定对HBVDNA具有明显抑制作用,最高可达46.52%,但对HBsAg、HBeAg影响较小;反义锁核酸对HBsAg、HBeAg及HBVDNA均有较强抑制作用,对HBsAg、HBeAg和HBVDNA的最高抑制率分别达67.69%、59.71%和62.96%(P<0.05),且抑制随时间呈增高趋势.反义锁核酸与拉米夫定对细胞代谢均无明显影响.结论:反义锁核酸抗HBV作用机制与拉米夫定不同,反义锁核酸抗HBV作用明显优于拉米夫定.

AIM：To compare the inhibitory effect of locked nucleic acid antisense oligonucleotides（antisense-LNA） and lamivudine on HBV replication in HepG2.2.15 cells.METHODS：Antisense-LNA was introduced into HepG2.2.15 cells by cationic liposome-mediated transfection.Supernatants were collected 2,4,6,8,10 days after medication.The concentrations of HBsAg and HBeAg in cell supernatants were tested by ELISA.HBV DNA levels in cellsupernatants were determined by FQ-PCR.Cell toxicity of antisense-LNA and lamivudine was detected by MTT assay.RESULTS：Lamivudine only inhibited viral DNA synthesis.Antisense-LNA effectively inhibited the expression of HBsAg and HBeAg and the replication of HBV DNA（67.69%,59.71%,62.96%,P 0.05） in a time-dependent manner.Both antisense-LNA and lamivudine showed no obvious cell toxicity.CONCLUSION：The anti-HBV effect of antisense-LNA is more effective than that of lamivudine in HepG2.2.15 cells.