Adeno-Associated Virus-Mediated Gene Transferinto Taste Cells In Vivo
Adeno-Associated Virus-Mediated Gene Transferinto Taste Cells In Vivo
复制标题
腺相关病毒介导的基因转移到体内味觉细胞
DOI：
10.1093/chemse/bjw101
复制
发表时间：
2017
期刊：
影响因子：
3.5
通讯作者：
Marunaka Y
中科院分区：
文献类型：
--
作者：
Taruno A;Kashio M;Sun H;Kobayashi K;Sano H;Nambu A;Marunaka Y
味觉是通过味觉细胞中表达的许多信号分子的合作来实现的，这些信号分子编码并将有关味觉质量和强度的信息传递到神经系统。病毒载体介导的基因转移技术已被证明是有用的研究和控制功能的基因生产在体内。然而，在活体动物中没有味觉细胞的转导方法。本文建立了一种利用重组腺相关病毒（AAV）载体在小鼠味觉细胞中诱导外源基因表达的方法。首先，使用增强型绿色荧光蛋白（EGFP）作为报告基因，我们筛选了6种AAV血清型沿着重组慢病毒载体，以了解它们对味觉细胞的能力。病毒注射到舌粘膜下层一周后，仅在注射AAV-DJ（一种合成血清型）的动物中观察到真菌状味觉细胞中的EGFP表达。接下来，评估AAV-DJ介导的EGFP在真菌状味觉细胞中表达的时间过程。胚内的EGFP信号出现后延迟，迅速增加，直到7天注射后，并在接下来的几周内逐渐下降，可能是因为细胞周转。最后，表征了对AAV-DJ转导敏感的味细胞类型。在PLCβ2免疫反应性II型和芳香族氨基酸脱羧酶（AADC）免疫反应性III型味觉细胞中以及在PLCβ2和AADC免疫阴性的细胞中观察到EGFP表达，表明AAV-DJ不能区分功能性味觉细胞类型。总之，本研究建立的方法将是一个很有前途的工具，研究味觉的机制。
The sense of taste is achieved by cooperation of many signaling molecules expressed in taste cells, which code and transmit information on quality and intensity of taste to the nervous system. Viral vector-mediated gene transfer techniques have been proven to be useful to study and control function of a gene productin vivo. However, there is no transduction method for taste cells in live animals. Here, we have established a method for inducing foreign gene expression in mouse taste cellsin vivoby recombinant adeno-associated virus (AAV) vector. First, using enhanced green fluorescent protein (EGFP) as a reporter, we screened 6 AAV serotypes along with a recombinant lentivirus vector for their ability to transduce taste cells. One week after viral injection into the submucosa of the tongue, EGFP expression in fungiform taste cells was observed only in animals injected with AAV-DJ, a synthetic serotype. Next, time course of AAV-DJ-mediated EGFP expression in fungiform taste cells was evaluated. Intragemmal EGFP signals appeared after a delay, rapidly increased until 7 days postinjection, and gradually decreased over the next few weeks probably because of the cell turnover. Finally, the taste cell types susceptible to AAV-DJ transduction were characterized. EGFP expression was observed in PLCβ2-immunoreactive type II and aromaticl-amino acid decarboxylase (AADC)-immunoreactive type III taste cells as well as in cells immunonegative for both PLCβ2 and AADC, demonstrating that AAV-DJ does not discriminate functional taste cell types. In conclusion, the method established in this study will be a promising tool to study the mechanism of taste.