Long Hu, Liao Lina, Zhou Yang, Shan Di, Gao Meiya, Huang Renhuan, Yang Xin, Lai Wenli
Department of Orthodontics, State Key Laboratory of Oral Diseases, West China Hospital of Stomatology, Sichuan University, Chengdu, China.
Department of Stomatology, Shanghai Tenth People's Hospital, Tongji University School of Medicine, Shanghai, China.
Eur J Oral Sci. 2017 Feb;125(1):1-7. doi: 10.1111/eos.12326. Epub 2017 Jan 9.
The objective of this study was to develop a viable and reliable technique of delivering viral vectors to rat trigeminal ganglia. Adult Sprague-Dawley rats (200-300 g) were used, and lentiviral vectors containing enhanced green fluorescence protein and calcitonin gene-related peptide short hairpin RNA (shRNA) were generated. Following general anesthesia, viral vectors were delivered to rat trigeminal ganglia using the technique described in this study. Both X-ray and micro-computed tomography (micro-CT) were employed to verify the position of the needles when injecting the vectors. In vivo fluorescence imaging and immunostaining against enhanced green fluorescence protein were performed to determine the success of viral transduction.The levels of calcitonin gene-related peptide in trigeminal ganglia were determined using real-time PCR, and pain levels following injections were evaluated using the Rat Grimace Scale. Our results show that injection needles can be advanced precisely at the trigeminal fossa and that viral vectors can successfully transduce trigeminal ganglia. Moreover, the levels of calcitonin gene-related peptide at trigeminal ganglia were down-regulated on day 7 after viral transduction. Pain levels returned to baseline by day 7 following injection. Therefore, we suggest that our trigeminal ganglion-targeting technique could be used for delivering genes or drugs to rat trigeminal ganglia.
本研究的目的是开发一种将病毒载体递送至大鼠三叉神经节的可行且可靠的技术。使用成年Sprague-Dawley大鼠（200 - 300克），构建了包含增强型绿色荧光蛋白和降钙素基因相关肽短发夹RNA（shRNA）的慢病毒载体。全身麻醉后，采用本研究中描述的技术将病毒载体递送至大鼠三叉神经节。在注射载体时，使用X射线和微型计算机断层扫描（micro-CT）来验证针头的位置。进行体内荧光成像和针对增强型绿色荧光蛋白的免疫染色以确定病毒转导是否成功。使用实时PCR测定三叉神经节中降钙素基因相关肽的水平，并使用大鼠 grimace量表评估注射后的疼痛程度。我们的结果表明，注射针可以精确地推进到三叉神经窝，并且病毒载体可以成功转导三叉神经节。此外，病毒转导后第7天，三叉神经节中降钙素基因相关肽的水平下调。注射后第7天疼痛程度恢复到基线水平。因此，我们认为我们的三叉神经节靶向技术可用于将基因或药物递送至大鼠三叉神经节。
