摘要目的 研究羧肽酶E如何促进淋巴细胞及其亚群跨血管内皮细胞进行迁移.方法利用CRISPR/Cas9技术对MS1细胞上的基因cpe进行敲除;淋巴细胞与血管内皮细胞黏附试验比较淋巴细胞对MS1细胞和敲除细胞Cpe-/-MS1的黏附能力,RT-PCR及流式方法比较MS1细胞和敲除细胞Cpe-/-MS1表面黏附分子的表达;Transwell试验比较淋巴细胞及其亚群穿过MS1细胞和敲除细胞Cpe-/-MS1的能力.结果我们成功获得了敲除细胞系Cpe-/-MS1;在TNF-α刺激下,淋巴细胞对MS1细胞的黏附能力显著高于敲除细胞Cpe-/-MS1;在TNF-α刺激下,MS1细胞表面黏附分子的表达量显著高于敲除细胞Cpe-/-MS1;在TNF-α刺激下,淋巴细胞及其亚群穿过MS1细胞的能力显著高于敲除细胞Cpe-/-MS1.结论羧肽酶E影响淋巴细胞与血管内皮细胞的黏附及跨血管内皮细胞迁移,本研究对于深入理解淋巴细胞跨血管内皮细胞向外周淋巴结迁移具有重要指导意义.

abstractsObjective To study the mechanism of carboxypeptidase E ( CPE ) in promoting the migration of lymphocytes and their subsets through vascular endothelial cells. Methods CRISPR/Cas9 technology was used to prepare cpe gene-knockout MS1 (Cpe-/-MS1) cells. Adhesion ability of lymphocytes to MS1 and Cpe-/-MS1 cells was analyzed with adhesion assay. Expression of adhesion molecules on these cells were detected by RT-PCR and flow cytometry. Transwell model was used to compare the difference in the transmigration of lymphocytes and their subsets through MS1 and Cpe-/-MS1 cells. Results Cpe-/-MS1 cells were successfully obtained. Under the stimulation of TNF-α, the adhesion ability of lymphocytes to MS1 cells was much better than that of Cpe-/-MS1 cells. Moreover, adhesion molecules expressed on MS1 cells were significantly more than those on Cpe-/-MS1 cells. The percentages of lymphocytes and their sub-sets that transmigrated through MS1 cells were significantly higher than those through Cpe-/-MS1 cells. Con-clusion CPE involved in the adhesion of lymphocytes to vascular endothelial cells and the transmigration of them through vascular endothelial cells, which was of great significance for understanding the migration of lymphocytes across vascular endothelial cells to peripheral lymph nodes.