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标题：Increase in the titer of lentiviral vectors expressing potassium channels by current blockade during viral vector production
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作者：Masayoshi Okada ; Naaz Andharia ; Hiroko Matsuda 等
期刊名称：BMC Neuroscience
印刷版ISSN：1471-2202
电子版ISSN：1471-2202
出版年度：2015
卷号：16
期号：1
页码：1
DOI：10.1186/s12868-015-0159-1
语种：English
出版社：BioMed Central
摘要：Background High titers of lentiviral vectors are required for the efficient transduction of a gene of interest. During preparation of lentiviral the vectors, the protein of interest is inevitably expressed in the viral vector-producing cells. This expression may affect the production of the lentiviral vector. Methods We prepared lentiviral vectors expressing inwardly rectifying potassium channel (Lv-Kir2.1), its dominant-negative form (Lv-Kir-DN), and other K⁺ channels, using the ubiquitously active β-actin and neuron-specific synapsin I promoters. Results The titer of Lv-Kir-DN was higher than that of Lv-Kir2.1, suggesting a negative effect of induced K⁺ currents on viral titer. We then blocked Kir2.1 currents with the selective blocker Ba²⁺ during Lv-Kir2.1 production, and obtained about a 5-fold increase in the titer. Higher extracellular K⁺ concentrations increased the titer of Lv-Kir2.1 about 9-fold. With a synapsin I promoter Ba²⁺ increased the titer because of the moderate expression of Kir2.1 channel. Channel blockade also increased the titers of the lentivirus expressing Kv1.4 and TREK channels, but not HERG. The increase in titer correlated with the K⁺ currents generated by the channels expressed. Conclusion In the production of lentivirus expressing K⁺ channels, titers are increased by blocking K⁺ currents in the virus-producing cells. This identifies a crucial issue in the production of viruses expressing membrane channels, and should facilitate basic and gene therapeutic research on channelopathies.
关键词：Lentiviral vector ; Titer ; K + channel ; Method improvement ; Kir2. 1 ; TREK-1 ; Kv1. 4 ; HERG