Murakami Yusuke, Ikeda Yasuhiro, Yonemitsu Yoshikazu, Tanaka Sakura, Kondo Haruhiko, Okano Shinji, Kohno Ri-Ichiro, Miyazaki Masanori, Inoue Makoto, Hasegawa Mamoru, Ishibashi Tatsuro, Sueishi Katsuo
Department of Pathology, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan.
J Gene Med. 2008 Feb;10(2):165-76. doi: 10.1002/jgm.1142.
Recombinant Sendai virus vectors (rSeV) constitute a new class of cytoplasmic RNA vectors that have shown efficient gene transfer in various organs, including retinal tissue; however, the related immune responses remain to be overcome in view of clinical applications. We recently developed a novel rSeV from which all envelope-related genes were deleted (rSeV/dFdMdHN) and, in the present study, assess host immune responses following retinal gene transfer.
rSeV/dFdMdHN or conventional F-gene deleted rSeV (rSeV/dF) was injected into subretinal space of adult Wistar rats or C57BL/6 mice. The transgene expression and histopathological findings were assessed at various time points. Immunological assessments, including the expression of proinflammatory cytokines, natural killer (NK)-cell activity, as well as SeV-specific cytotoxic T lymphocytes (CTLs) and antibodies, were performed following vector injection.
rSeV/dFdMdHN showed high gene transfer efficiency into the retinal pigment epithelium at an equivalent level to that seen with rSeV/dF. In the early phase, the upregulation of proinflammatory cytokines, local inflammatory cell infiltration and tissue damage that were all prominently seen in rSeV/dF injection were dramatically diminished using rSeV/dFdMdHN. NK cell activity was also decreased, indicating a reduction of the innate immune response. In the later phase, on the other hand, CTL activity and anti-SeV antibodies were similarly induced, even using rSeV/dFdMdHN, and resulted in transient transgene expression in both vector types.
Deletion of envelope-related genes of rSeV dramatically reduces the vector-induced retinal damage and may extend the utility for ocular gene transfer; however, further studies regulating the acquired immune response are required to achieve long-term transgene expression of rSeV.
重组仙台病毒载体（rSeV）构成了一类新型的细胞质RNA载体，已在包括视网膜组织在内的各种器官中显示出高效的基因转移；然而，鉴于临床应用，相关的免疫反应仍有待克服。我们最近开发了一种新型的rSeV，其中所有包膜相关基因均被删除（rSeV/dFdMdHN），在本研究中，我们评估了视网膜基因转移后的宿主免疫反应。
将rSeV/dFdMdHN或传统的F基因缺失的rSeV（rSeV/dF）注射到成年Wistar大鼠或C57BL/6小鼠的视网膜下间隙。在不同时间点评估转基因表达和组织病理学结果。在载体注射后进行免疫评估，包括促炎细胞因子的表达、自然杀伤（NK）细胞活性以及SeV特异性细胞毒性T淋巴细胞（CTL）和抗体。
rSeV/dFdMdHN在视网膜色素上皮中显示出与rSeV/dF相当的高效基因转移效率。在早期阶段，rSeV/dF注射中显著出现的促炎细胞因子上调、局部炎性细胞浸润和组织损伤，使用rSeV/dFdMdHN时显著减少。NK细胞活性也降低，表明先天免疫反应减弱。另一方面，在后期，即使使用rSeV/dFdMdHN，CTL活性和抗SeV抗体也同样被诱导，并导致两种载体类型中的转基因表达短暂。
rSeV包膜相关基因的缺失显著降低了载体诱导的视网膜损伤，并可能扩展眼内基因转移的效用；然而，需要进一步研究调节获得性免疫反应以实现rSeV的长期转基因表达。
