Inhibiting HIV-1 infection in human T cells by lentiviral-mediated delivery of small interfering RNA against CCR5
Inhibiting HIV-1 infection in human T cells by lentiviral-mediated delivery of small interfering RNA against CCR5
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DOI：
10.1073/pnas.232688199
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发表时间：
2003-01-07
影响因子：
11.1
通讯作者：
Baltimore, D
中科院分区：
文献类型：
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作者：
Qin, XF;An, DS;Baltimore, D
长度约为21个核苷酸的双链RNA[小干扰RNA（siRNA）]被认为是降低特定基因表达的有效试剂。要将它们用作保护细胞免受病毒感染的试剂，就需要将siRNA导入原代细胞的有效方法。在此，我们描述了成功构建一种基于慢病毒的载体，用于将针对HIV - 1共受体CCR5的siRNA导入人外周血T淋巴细胞。利用高滴度的载体储备液，可转导超过40%的外周血T淋巴细胞，并且在不进行筛选的情况下，一种有效的CCR5 - siRNA的表达在2周内可使细胞表面CCR5的表达抑制高达10倍。相比之下，另一种主要的HIV - 1共受体CXCR4的表达未受影响。重要的是，通过siRNA阻断CCR5的表达为淋巴细胞群免受嗜CCR5的HIV - 1病毒感染提供了显著的保护，使感染细胞减少3到7倍；而对嗜CXCR4病毒的感染仅观察到极小的影响。因此，我们的研究证明了慢病毒载体介导的siRNA递送作为一种细胞内免疫的通用手段用于治疗HIV - 1和其他病毒性疾病的可行性和潜力。
Double-stranded RNAs approximate to21 nucleotides long [small interfering RNA (siRNA)] are recognized as powerful reagents to reduce the expression of specific genes. To use them as reagents to protect cells against viral infection, effective methods for introducing siRNAs into primary cells are required. Here, we describe success in constructing a lentivirus-based vector to introduce siRNAs against the HIV-1 coreceptor, CCR5, into human peripheral blood T lymphocytes. With high-titer vector stocks, >40% of the peripheral blood T lymphocytes could be transduced, and the expression of a potent CCR5-siRNA resulted in up to 10-fold inhibition of CCR5 expression on the cell surface over a period of 2 weeks in the absence of selection. In contrast, the expression of another major HIV-1 coreceptor, CXCR4, was not affected. Importantly, blocking CCR5 expression by siRNAs provided a substantial protection for the lymphocyte populations from CCR5-tropic HIV-1 virus infection, dropping infected cells by 3- to 7-fold; only a minimal effect on infection by a CXCR4-tropic virus was observed. Thus, our studies demonstrate the feasibility and potential of lentiviral vector-mediated delivery of siRNAs as a general means of intracellular immunization for the treatment of HIV-1 and other viral diseases.