特异性反义寡核苷酸抗小鼠呼吸道合胞病毒感染的研究

目的 比较研究特异性抑制呼吸道合胞病毒(respiratory syncytial virus,RSV)的反义核苷酸(antisense oligodeoxynucleotide,AS-ODN)和利巴韦林对RSV感染小鼠的治疗作用,寻求安全有效的抗RSV药物.方法 特异性ASODN和利巴韦林滴鼻治疗RSV感染BALB/c鼠,空斑形成实验检测肺组织病毒滴度,RT-PCR和免疫组织化学分析RSVmRNA和蛋白表达水平,支气管肺泡灌洗液(BALF)白细胞计数、肺组织病理学检查了解气道炎症和治疗副作用.结果 0.2、0.4 mg AS-ODN和0.8 mg利巴韦林能降低RSV感染小鼠肺组织病毒滴度,空斑抑制率分别为34.48%、46.75%和23.02%,与感染对照组比较差异显著(P＜0.05).0.4 mg AS-ODN抑制RSV mRNA和蛋白表达,抑制率分别为30.54%和29.41%,0.4 mg利巴韦林治疗降低17.65%的RSV蛋白表达.AS-ODN和利巴韦林都降低RSV感染小鼠BALF中白细胞总数,对肺组织病理性改变没有明显改善作用,与感染对照组比较差异不显著(P＞0.05).单独AS-ODN和利巴韦林滴鼻不会引发明显肺炎症病理损害.结论 AS-ODN在小鼠体内有比利巴韦林更强的抗RSV作用,经滴鼻治疗没有明显毒副作用,是安全而有效的抗RSV制剂.

Study on specific antisense oligonucleotide against respiratory syncytial virus in mice

Objective To compare the effects of antisense oligodeoxynucleotide (AS-ODN) and Ribavirin against respiratory syncytial virus (RSV) in mice. Methods BALB/c mice infected of RSV were nasally dropped with AS-ODN (0.2, 0.4 mg) or Ribavirin (0.4, 0.8 mg). Seventy-two hours after infection, pulmonary viral titer was detected by plaque forming experiment, pulmonary RSV N gene mRNA expression was assayed by RT-PCR, and viral protein was detected by immunohistochemistry. To represent respiratory tract inflammation and side-effect caused by AS-ODN or Ribavirin, leukocytes and the subgroup cells in bronchoalveolar lavage fluid (BALF) were counted and pulmonary histopathologic changes were examined. Results Pulmonary titer of mice treated with 0.2, 0.4 mg AS-ODN or 0.8 mg Ribavirin decreased as compared with that of infected control mice, and the plaque forming inhibition ratio was 34.48%, 46.75% and 23.02% respectively. AS-ODN (0.4 mg) inhibited the expressions of pulmonary RSV N gene mRNA and protein by 30.54% and 29.41% respectively. Ribavirin (0.4 mg) decreased RSV protein expression by 17.65%. Furthermore, AS-ODN and Ribavirin both at dose of 0.4 mg dramatically decreased total number of leukocytes in BALF, but did not significantly improve pulmonary histopathologic changes. The mice merely dropped with AS-ODN or Ribavirin both at dose of 0.4 mg appeared no visible pulmonary histopathologic changes. Conclusion AS-ODN shows more potent anti-RSV activity than Ribavirin in mice, and their application through nasal drop has no obvious noxious and side effects. So AS-ODN is an efficient and safe anti-RSV agent.