Intracerebral delivery of small interfering RNAs (siRNAs) using adenoviral vector protects mice against lethal peripheral rabies challenge.
Intracerebral delivery of small interfering RNAs (siRNAs) using adenoviral vector protects mice against lethal peripheral rabies challenge.
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DOI：
10.1016/j.virusres.2011.08.004
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发表时间：
2012-01
期刊：
影响因子：
5
通讯作者：
Saini M
中科院分区：
文献类型：
--
作者：
Gupta PK;Sonwane AA;Singh NK;Meshram CD;Dahiya SS;Pawar SS;Gupta SP;Chaturvedi VK;Saini M
►我们研究了针对狂犬病病毒核蛋白(N)和聚合酶(L)基因的两个小干扰RNA(SiRNA)作为抗病毒药物的作用。►腺病毒携带的siRNAs在BHK-21细胞中的抗病毒作用表现出明显的抑制RV增殖、RV滴度下调RV基因转录的作用。表达RV-N和RV-L基因的腺病毒对致死性狂犬病病毒攻击的保护率分别为66.6%和33.3%。为探讨RNA干扰作为抗狂犬病药物的可能性，设计并评价了针对狂犬病病毒核蛋白(N)和聚合酶(L)基因的两种小干扰RNA。在基于质粒的瞬时表达模型中，两个siRNAs都击倒或沉默了目标RABV基因。为了有效地传递siRNAs，构建了表达siRNAs的腺病毒，并研究了所传递的siRNAs在BHK-21细胞中的抗病毒作用。以RABV-N和L基因为靶点的siRNA腺病毒感染细胞后，RABV感染细胞的RABV增殖分别下降了88.35±22.4%和41.52±99.3%。实时定量RT-PCR检测结果显示，RABV-N和L基因的转录均被下调，但只有腺病毒表达的针对RABV-N的siRNA显著降低。脑内注射表达siRNAs腺病毒的小鼠经咬肌肌内注射致死性RABV后，对RABV-N和L基因的保护率分别为66.6%和33.3%。这些结果表明，表达针对RABV-N的siRNA的腺病毒在体内外都有效地抑制了RABV的增殖，并对致死性RABV攻击具有显著的保护作用。这支持了基于针对RABV-N基因的siRNA的RNAi可以预防RABV感染的假设，并保持了RNAi作为预防狂犬病感染的一种途径的潜力。
► We investigated RNA interference (RNAi) as antiviral agent against rabies using two small interfering RNAs (siRNAs) targeting rabies virus (RV) nucleoprotein (N) and polymerase (L) genes. ► The antiviral potential of siRNAs delivered using adenoviruses in BHK-21 cells showed marked inhibition in RV multiplication, RV titer knockdown of RV gene transcripts. ► Mice treated with adenoviruses expressing siRNAs showed 66.6% and 33.3% protection with adenoviruses expressing siRNAs against RV-N and RV-L genes, respectively against lethal rabies virus challenge. To investigate the potential of RNA interference (RNAi) as antiviral agent against rabies, two small interfering RNAs (siRNAs) targeting rabies virus (RABV) nucleoprotein (N) and polymerase (L) genes were designed and evaluated. Both siRNAs knockdown or silenced the target RABV genes as evaluated in a plasmid based transient expression model. For efficient delivery, adenoviruses expressing the siRNAs were constructed and antiviral potential of the delivered siRNAs was investigated in BHK-21 cells. When cells treated with adenoviruses expressing siRNAs were challenged with RABV, there was 88.35 ± 2.4% and 41.52 ± 9.3% reduction in RABV multiplication in infected cells with siRNAs targeting RABV-N and L genes, respectively. Relative quantification of RABV transcripts using real-time PCR revealed knockdown of both RABV-N and L gene transcripts, however, significant reduction was observed only with adenovirus expressing siRNA against RABV-N. When mice treated intracerebrally with adenoviruses expressing siRNAs were challenged peripherally with lethal RABV by the intramuscular route in masseter muscle, there was 66.6% and 33.3% protection with adenoviruses expressing siRNAs against RABV-N and L genes, respectively. These results demonstrated that adenovirus expressing siRNA against RABV-N efficiently inhibited the RABV multiplication both, in vitro and in vivo and conferred significant protection against lethal RABV challenge. This supported the hypothesis that RNAi, based on siRNA targeting RABV-N gene can prevent RABV infection and holds the potential of RNAi as an approach to prevent rabies infection.
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