中文摘要：

目的：探讨肌肉注射Notexin是否影响C57BL/6小鼠骨骼肌引流淋巴结内的外源性CD8＋T细胞（OT-I细胞）的活性和增殖。方法分别采用Notexin注射或机械挤压法制备B6小鼠的胫骨前肌（TA）急性肌损伤模型，并获得性转移（adoptive transfer, i.v.）OVA特异的OT-I细胞（CFSE标记），随后肌注可溶性OVA蛋白。收集损伤侧TA引流淋巴结（腘、腹股沟淋巴结），流式分析OT-I细胞的增殖程度。OVA静脉内注射免疫B6鼠，收集激活的脾脏树突状细胞（cDCs），与CFSE标记的OT-I细胞体外联合培养，并添加不同稀释度的Notexin,流式检测OT-I细胞的活性和增殖。结果 CFSE的荧光递减结果证实，机械挤压损伤鼠TA引流淋巴结内OT-I细胞迅速活化增殖，但Notexin诱发的肌损伤小鼠引流淋巴结内OT-I细胞在4 d时无增殖反应，7 d的增殖率与非注射组无显著差异。与活化的cDCs细胞共培养的OT-I细胞活性良好，增殖显著，但即使添加高度稀释（1：1000）的Notexin也会严重干扰OT-I细胞的活性和增殖。结论肌内注射Notexin注射将干扰CD8＋T细胞的活性，这表明蛇毒血清诱导的骨骼肌损伤模型不适用于肌损伤诱发的T细胞功能改变的相关研究。

英文摘要：

Objective To explore the effects of intramuscular injection of Notexin on transferred OT-I cells in muscle draining lymph nodes of C57BL/6 mice. Methods Tibialis anterior（TA）injury were performed by intramuscular injection of Notexin, or crush skill on B6 mice, separately. CFSE-labeled OT-I cells were adoptively transferred to myoinjury mice, and followed by OVA i.m. injection. TA draining lymph nodes were received, and lymphocytes were collected and analyzed the proliferation of OT-I cells by CFSE fluorescence intensity evaluation using flow cytometry. Activated spleen cDC cells were collected and purified from one B6 mouse received OVA i.v. injection. Purified cDC cells were co-cultured with CFSE-labeled OT-I cells. Notexin with different concentration were added into co-culturing system. After 72h, the survival and proliferation of OT-I cells were detected. Results According to the detection results of CFSE fluorescence intensity, OT-I cells in crush induced-damaged muscle draining lymph nodes were activated and proliferated rapidly under the stimulation of OVA antigen. For Notexin injection-induced muscle damage, no cell proliferation of OT-I cells were detected in TA draining lymph nodes at the day 4, but the proliferation of OT-I cells recovered at the day 7. Co-culturing with activated cDCs cells induced a obvious proliferation of OT-I cells in vitro, however, OT-I cells lost the proliferation ability if the co-culturing system were added with Noteixn, even if the concentration of Notexin was very lower（1：1000）. Conclusions The activity and proliferation of CD8＋T cells will be interfered by intramuscular injection of Notxin. Therefore, we suggest that Notexin injection is unsuitable for investigating T cell response associated with myoinjury.