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目的观察小干扰RNA(siRNA)7816在小鼠体内的分布及吸收情况,探讨siRNA7816对呼吸道合胞病毒(RSV)感染的抑制效果。方法将5’端带有三氢吲哚氰(cy3)荧光基团的siRNA7816,通过滴鼻的方法散布到小鼠的肺组织中,并于滴鼻后的12 h将小鼠处死,无菌条件下取小鼠肺组织、肝脏组织及肠组织,并迅速行组织冷冻切片,荧光显微镜下观察siRNA7816在Balb/c小鼠体内的分布情况。将48只Balb/c小鼠随机分为6组,每组8只,各组均予以1010pfu.L-1的RSVA2标准株病毒行滴鼻处理,并于感染后的第2天分别予治疗组小鼠0.5 nmol、1.0 nmol及2.0 nmol的治疗量siRNA7816,阴性对照组则予以1.0 nmol的阴性siRNA,PBS组予PBS滴鼻处理,空白对照组不予任何处理。于感染后的第3天将小鼠处死,并于无菌条件下取小鼠肺组织,通过光学显微镜观察小鼠肺组织的病理改变和免疫组织化学检测光密度值的方法,了解siRNA7816对RSV病毒的抑制效果。结果siRNA7816通过滴鼻的方法能够顺利到达Balb/c小鼠肺部,并分布于支气管、肺泡等处,且分布水平较高;病理结果显示siRNA7816能显著减轻RSV感染小鼠的炎症,免疫组织化学分析结果显示siRNA7816对RSV病毒蛋白的表达有抑制作用,与PBS组比较差异有统计学意义(P<0.05)。结论采用滴鼻的方法可以将siRNA投送到小鼠肺组织的有效部位并形成一定浓度,siRNA7816对RSV的复制有一定抑制作用。
Objective To observe the distribution and absorption of small interfering RNA (siRNA) 7816 in mice and to investigate the inhibitory effect of siRNA7816 on respiratory syncytial virus (RSV) infection. Methods siRNA7816 with cy3 fluorescent group at the 5 ’end was disseminated to the lung tissue of mice by intranasal method, and the mice were sacrificed 12 h after intranasal instillation. Sterile conditions The lung, liver and intestine tissues of mice were removed and the frozen section was rapidly dissected. The distribution of siRNA7816 in Balb / c mice was observed under a fluorescence microscope. Forty-eight Balb / c mice were randomly divided into 6 groups with 8 mice in each group. Each group received intranasal administration of 1010pfu.L-1 RSV A2 standard virus and were treated on the second day after infection The mice were treated with 0.5 nmol, 1.0 nmol and 2.0 nmol siRNA7816, negative control group was given 1.0 nmol of siRNA negative, PBS group treated with PBS nasal drops, the blank control group without any treatment. The mice were sacrificed on the third day after infection and the lung tissues of the mice were taken under aseptic conditions. The pathological changes in the lung tissues of the mice were observed by optical microscopy and the optical density of the mice was examined by immunohistochemistry. The inhibitory effect of the virus. Results siRNA7816 successfully delivered to the lungs of Balb / c mice via nasal route and distributed in the bronchi, alveoli and other places, and its distribution was high. The results of pathology showed that siRNA7816 could significantly reduce the inflammation of mice infected with RSV, and the immunohistochemistry The results showed that siRNA7816 could inhibit the expression of RSV virus, which was significantly different from PBS group (P <0.05). Conclusion The method of intranasal administration of siRNA can be delivered to the effective part of the lung tissue of mice and to form a certain concentration, siRNA7816 certain inhibition of RSV replication.