Inhibitory effect of small interfering RNA on dengue virus replication in mosquito cells.
Inhibitory effect of small interfering RNA on dengue virus replication in mosquito cells.
复制标题
小干扰RNA对蚊子细胞中登革热病毒复制的抑制作用
DOI：
10.1186/1743-422x-7-270
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发表时间：
2010-10-14
期刊：
影响因子：
4.8
通讯作者：
Yang X
中科院分区：
文献类型：
--
作者：
Wu X;Hong H;Yue J;Wu Y;Li X;Jiang L;Li L;Li Q;Gao G;Yang X
背景登革病毒（DEN）是全球热带和亚热带地区最广泛传播的蚊媒病原体。感染登革病毒可导致严重的流感样疾病和潜在的致命性出血热。虽然RNA干扰引发的长长度的dsRNA被认为是一个有效的抗病毒途径在蚊子，只有有限的研究的价值小干扰RNA（siRNA）已经进行了carried.ResultsA 21 nt siRNA靶向膜糖蛋白前体基因DEN-1的合成和转染到蚊子C6/36细胞，然后与DEN的挑战。通过流式细胞术监测siRNA在细胞中的稳定性。MTT法检测细胞存活率，实时定量RT-PCR检测病毒RNA含量。转染后0.25、1、3、5、7 d的siRNA阳性率分别为66.0%、52.1%、32.0%、13.5%和8.9%。与DEN孵育7天后，细胞病变效应降低，提高细胞存活率（76.9 ± 4.5% vs23.6 ± 14.6%）和病毒RNA拷贝减少在转染的C6/36细胞中检测到（Ct值为19.91 ± 0.63vs14.56 ± 0.39）。1膜糖蛋白前体基因能有效抑制DEN-1病毒RNA的复制，提高C6/36细胞的存活率。siRNA可能为DEN感染的预防和治疗提供一种潜在的新策略。
BackgroundDengue viruses (DENs) are the wildest transmitted mosquito-borne pathogens throughout tropical and sub-tropical regions worldwide. Infection with DENs can cause severe flu-like illness and potentially fatal hemorrhagic fever. Although RNA interference triggered by long-length dsRNA was considered a potent antiviral pathway in the mosquito, only limited studies of the value of small interfering RNA (siRNA) have been conducted.ResultsA 21 nt siRNA targeting the membrane glycoprotein precursor gene of DEN-1 was synthesized and transfected into mosquito C6/36 cells followed by challenge with DEN. The stability of the siRNA in cells was monitored by flow cytometry. The antiviral effect of siRNA was evaluated by measurement of cell survival rate using the MTT method and viral RNA was quantitated with real-time RT-PCR. The presence of cells containing siRNA at 0.25, 1, 3, 5, 7 days after transfection were 66.0%, 52.1%, 32.0%, 13.5% and 8.9%, respectively. After 7 days incubation with DEN, there was reduced cytopathic effect, increased cell survival rate (76.9 ± 4.5%vs23.6 ± 14.6%) and reduced viral RNA copies (Ct value 19.91 ± 0.63vs14.56 ± 0.39) detected in transfected C6/36 cells.ConclusionsOur data showed that synthetic siRNA against the DEN-1 membrane glycoprotein precursor gene effectively inhibited DEN-1 viral RNA replication and increased C6/36 cell survival rate. siRNA may offer a potential new strategy for prevention and treatment of DEN infection.