生理学报Acta Physiologica Sinica, June 25, 2003, 55(3)

研究论文

CD226单克隆抗体诱导人脐静脉内皮细胞胞质钙离子浓度变化的研究

陈丽华, 刘雪松, 刘飞, 金伯泉*

第四军医大学基础部免疫学教研室, 西安 710032

摘要: 为观察CD226单克隆抗体(mAb)对培养人脐静脉内皮细胞(HUVECs)胞质钙离子变化的影响, 我们用Fluo-3作为钙指示剂, 用激光共聚焦显微镜观测不同状态下CD226 mAb作用后HUVECs胞质钙离子[Ca2+]i的变化。结果发现: (1)用Hanks液平衡, CD226 mAb作用后HUVECs [Ca2+]i 水平缓慢升高后回到原位; 加入二抗(羊抗鼠IgG)交联后 [Ca2+]i 水平有较大幅度的升高, 随后回到原位, 与此同时, 细胞外液中[Ca2+]水平有一定程度的下降; (2)用D-Hanks液平衡, CD226 mAb作用后HUVECs [Ca2+]i 水平无显著变化, 加入二抗发生交联作用后, [Ca2+]i 水平有较大幅度的下降; (3)用EGTA预处理后, CD226 mAb及其二抗交联对HUVECs [Ca2+]i变化无显著影响。以上结果提示, CD226 mAb及其二抗交联可诱导不同状态的HUVECs胞质钙离子水平发生不同程度的变化, 从而参与一系列的生理和病理过程。

关键词: 生理学; CD226; 激光共聚焦显微镜; [Ca2+]i ; 人脐静脉内皮细胞

中图分类号: R329; Q582

CD226 monoclonal antibody induces variation of intracytosolic free calcium level in human umbilical vein endothelial cells

CHEN Li-Hua, LIU Xue-Song, LIU Fei, JIN Bo-Quan

Department of Immunology, Fourth Military Medical University, Xi′an, Shanxi 710032

Abstract:In order to study the possible mechanism of CD226 monoclonal antibody (mAb)-mediated intracellular message transduction in human umbilical vein endothelial cells (HUVECs), the influence of CD226 mAb and its cross-linking by secondary antibody (Abs) on the concentration changes in [Ca2+]i in the HUVECs under different conditions were determined by confocal laser scanning microscopy. The main results are as follows. (1) When the culture medium was balanced by Hanks Buffer, [Ca2+]i in HUVECs increased slowly after stimulation by CD226 mAb, whereas [Ca2+]i increase was accompanied by [Ca2+]o decrease after the mAb was cross-linked by goat anti-mouse IgG. Then [Ca2+]i and [Ca2+]o all returned to the normal level. (2) When the culture medium was balanced by D-Hanks buffer, [Ca2+]i in HUVECs showed little variation when the cells were stimulated by CD226 mAb, but [Ca2+]i decreased markedly after cross-linking. (3) When HUVECs were pretreated with EGTA, there was no variation in [Ca2+]i of HUVECs after CD226 mAb stimulation alone or cross-linking of the mAb. Our results suggest that stimulation by CD226 mAb and cross-linking by goat anti-mouse IgG induce the variation of [Ca2+]i in HUVECs under different conditions and the variation of [Ca2+]i in HUVECs may play an important role in many physiological and pathological processes.

Key words: physiology; CD226; confocal laser scanning microscopy; [Ca2+]i; human umbilical vein

endothelial cells (HUVECs)