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目的 探讨oxLDL对人内皮细胞的CXC亚家族趋化因子GROα的调节作用及生理意义。方法 超高速离心得LDL ,氧化后得oxLDL。逆转录PCR分析人内皮细胞系ECV30 4细胞GROαmRNA。GAPDH被用作PCR中正常改变的对照物。酶联免疫检测仪测定ECV30 4细胞表面连接及分泌到溶液中的GROα蛋白。静态细胞粘附试验测定ECV30 4细胞表面连接的GROα蛋白的生理意义。结果 正常ECV30 4细胞不表达GROαmRNA ;OxLDL显著调节其表达 ,诱导效应首先出现在处理后 1h ,最大在 2h ,在 4h时下降。相比较 ,LDL对其mRNA表达没有影响。正常ECV30 4细胞低水平表达细胞表面的GROα蛋白。OxLDL呈浓度依赖性、时间依赖性地上调其表面的GROα蛋白。相比较 ,LDL对其表面表达的GROα蛋白无调节作用。OxLDL对ECV30 4细胞分泌到细胞溶液中GROα蛋白很少有影响。 4 0 μg/mloxLDL处理ECV30 4细胞 2 4h ,造成显著的人单核细胞系U937细胞粘附到ECV30 4细胞数目的增加。用GROα抗体预处理oxLDL刺激的ECV30 4细胞 ,可显著减低U937细胞粘附到ECV30 4细胞的U937细胞数目 (大约 2 / 3)。结论 oxLDL可能功能性上调内皮细胞GROα的表达。
Objective To investigate the regulation and physiological significance of oxLDL on the chemokine GROα of CXC subfamily in human endothelial cells. Methods LDL was obtained by ultracentrifugation and oxLDL was obtained after oxidation. Reverse transcription-PCR analysis of human endothelial cell line ECV304 cells GROα mRNA. GAPDH was used as a normal change control in PCR. The enzyme-linked immunosorbent assay was used to determine the GROα protein attached to the surface of ECV304 cells and secreted into the solution. The static cell adhesion assay was used to determine the physiological significance of the GROα protein attached to the surface of ECV304 cells. RESULTS: Normal ECV304 cells did not express GROalpha mRNA; OxLDL significantly regulated its expression. The induction effect first appeared at 1 h after treatment, with a maximum of 2 h, and decreased at 4 h. In contrast, LDL had no effect on its mRNA expression. Normal ECV304 cells express low levels of cell surface GROα protein. OxLDL upregulates its surface GROα protein in a concentration- and time-dependent manner. In contrast, LDL has no regulatory effect on its surface expressed GROα protein. OxLDL has little effect on the secretion of GROα protein from ECV304 cells into the cell solution. Treatment with 40 μg/ml oxLDL for ECV304 cells for 24 h resulted in a significant increase in the number of ECG304 cells adherent to the human monocyte line U937 cells. Pretreatment of oxLDL-stimulated ECV304 cells with GROalpha antibody significantly reduced the number of U937 cells (about 2/3) that U937 cells adhered to ECV304 cells. Conclusion oxLDL may functionally up-regulate the expression of GROα in endothelial cells.