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沙发
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发表于 2015-8-26 13:54:29
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rojjer:
谢谢侯兄分享。siRNA之治疗应用。
siRNA抗病毒治疗，怎么排除是因为siRNA在哺乳动物体内激活天然免疫系统，诱导干扰素等炎症因子的分泌而产生的非特异抗病毒作用，还是因为siRNA诱导靶基因沉默后导致病毒复制周期无法完成从而达到抗病毒的目的？
对于流感病毒来说，这一点更是尤为重要，因为其本身的非结构蛋白就有起对抗干扰素作用（可以说流感病毒天生对干扰素就比较“敏感”），siRNA也必将影响NS1功能的发挥，其诱导的干扰素也会严重影响病毒复制。而这种抗病毒都是非特异的，这里siRNA是起到了干扰素诱生剂的作用还是诱导靶基因沉默的作用，我想研究者的出发点还是后者。
那么以上研究选择的靶基因是什么，如何证明是引起了靶基因的沉默？
呵呵，请比较关注的朋友谈谈看法。
一只蜗牛:
以后应用，“DNA环”怎么进入体内？或者如何靶向？电转？效率也是一个问题。
而且DNA环进入机体内，机体天然的siRNA机制就不把你这个外源进来的DNA也来个沉默？还能等你去沉默H5N1？
有点挑刺，交流，呵呵。
对于Rojier提到的问题，我想作者可能会设置一个不相干的DNA环作为对照，然后看是哪个环节出了问题。
而靶基因，我想应该是核内的保守蛋白。类似于人流感里的M2啊，NP啊之类的吧。
Rojjer:
发现一篇文章，CDC的！
Inhibitory effect of small interfering RNA specific for a novel candidate target in PB1 gene of influenza A virus.Cheng C, Yao L, Chen A, Jia R, Huan L, Guo J, Bo H, Shu Y, Zhang Z.
State Key Laboratory for Infectious Disease Prevention and Control, National Institute for Viral Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing, P.R. China.
Influenza, mainly caused by influenza virus, is becoming one of the major concerns in the world. Limitation in vaccines necessitates the urgent development of new therapeutic options against this virus. In the present study, we designed small interfering RNA (siRNA) targeting overlapping gene of PB1 and PB1-F2 gene of the influenza A virus and investigated its effect against influenza A virus infection. A reduction in virus-associated cell apoptosis was observed in A549 cells treated with this siRNA. Furthermore, its antiviral effect was confirmed by different methods. Also, a marked decrease of virus titer in chicken embryos treated with the siRNA was observed. The findings of this work highlight the potential of this shared region to be an additional therapeutic target for the treatment of influenza virus infection.
Rojjer:
For more information:
1,J Biotechnol. 2008 Jun 1;135(2):140-4. Epub 2008 Mar 26.
RNA interference of avian influenza virus H5N1 by inhibiting viral mRNA with siRNA expression
2,Avian Dis. 2005 Dec;49(4):562-73.
Construction of influenza virus siRNA expression vectors and their inhibitory effects on multiplication of influenza virus.Li YC, Kong LH, Cheng BZ, Li KS.
3,Virus Res. 2004 Jun 1;102(1):37-42.
Use of siRNAs to prevent and treat influenza virus infection.Ge Q, Eisen HN, Chen J.
kousyouki:
我也不大了解miRNA的操作方法，仅知道原理而已。
对于中科院动物所这篇文章我也没有找到，所以也不好说，我也来个抛砖引玉呵。呵呵，刚从图书馆借了两本书来看看。
只是觉得自己需要了解一下siRNA在对抗流感病毒的应用。
如蜗牛兄所说，机体内的siRNA机制会沉默外源“DNA环”，是否也存在瞬时全沉默呢？如果不是的话，那应该也是一种有效的沉默influenza virus 的方法吧。
呵呵，我是siRNA外行，问问~~
zhangtao1012450:
你说的应该是这篇文章，我记得我的电脑里有，但是没有找到
J Biotechnol. 2008 Jun 1;135(2):140-4. Epub 2008 Mar 26.
RNA interference of avian influenza virus H5N1 by inhibiting viral mRNA with siRNA expression plasmids.
Zhou K, He H, Wu Y, Duan M.
National Research Center For Wildlife Born Diseases, Key Laboratory of Animal Ecology and Conservation Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing 100101, PR China.
还是没有找到，把摘要给你看看吧
Abstract
Avian influenza virus H5N1 causes widespread infection in the birds and human respiratory tract, but existing vaccines and drug therapy are of limited value. Here we show that small interfering RNAs (siRNAs) specific for conserved regions of the viral genome can potently inhibit influenza virus production in cell lines, embryonated chicken eggs and BALB/c mice. siRNA expression plasmid pBabe-Super was chosen in the study, which directed the synthesis of small interfering RNAs in cells. The inhibition depended on the presence of a functional antisense strand in the small interfering RNA duplex, suggesting that viral mRNA is the target of RNA interference (RNAi). Among the three small interfering RNA expression plasmids we designed, we found that small interfering RNA for nucleocapsid protein (NP) had a specific effect in inhibiting the accumulation of RNAs in infected cells because of a critical requirement for newly synthesized nucleocapsid proteins in avian influenza viral RNA transcription and replication. The findings reveal that newly synthesized nucleocapsid, polymerase A (PA) and polymerase B1 (PB1) proteins are required for avian influenza virus transcription and replication and provide a basis for the development of small interfering RNAs as prophylaxis and therapy for avian influenza infection in birds and humans.
Keywords: H5N1; Avian influenza virus (AIV); RNA interference
ara189:
本来就有这个方法的呀 利用基因保守区域 直接沉默病毒基因（PS，病毒的突变率比较高，这个区域比较难找的说）
天天游泳:
回复 1# kousyouki 的帖子
这种文章没什么意义吧。
文章没有看过，不过估计就是设计了序列针对流感病毒某段基因序列，转染进细胞之后选择性沉默病毒的某些个基因。
这个方法临床的可能性基本上约等于0吧。
而且这种简直是为了文章而文章，浪费纳税人的钱。
wanghai8282:
大家讨论吧，希望能有收获！
这是大家讨论的文章，希望对大家有帮助！
RNAi 我也不大清楚，希望各位高手指导！
tatamige:
真的吗？我最近一直在考虑这个问题……结果发现还是有人最先发现了
其实我觉得吧 与其用质粒DNA环搭载siRNA片段 不如直接用病毒载体
虽然原理是一样.
再补充一点啊 我所谓的病毒载体不是腺病毒载体 应该是机体缺乏先天免疫的载体 如人体内应用新城疫载体 |
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发表于 2015-8-26 13:48:14
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