多种类型趋化性细胞因子在小鼠胸腺基质细胞系中的表达

目的分析趋化性细胞因子 SDF- 1α、 IP- 10、 KC、 MCP- 1和 RANTES在小鼠胸腺基质细胞系 MTEC1、 MTDC、 D2SC、 MTECB、 TEC 1C8、 TNC及原代培养胸腺基质细胞的半定量表达情况,并检测重组的趋化性细胞因子纯品 SDF- 1α、 IP- 10、 MCP- 1和 RANTES对小鼠胸腺细胞的趋化活性。方法以β- actin为内对照,用 RT- PCR方法,对上述 5种趋化性细胞因子进行 30个循环 PCR扩增,用凝胶成像系统观察扩增结果 ,并对各电泳带进行积分光密度扫描分析。用 Boyden小室法检测重组趋化性细胞因子 SDF- 1α、 IP- 10、 MCP- 1和 RANTES对全胸腺细胞的趋化活性,并计算其趋化指数。结果 SDF- 1α、 IP- 10、 KC、 MCP- 1和 RANTES在被检测胸腺基质细胞系的表达谱不同,表达强度亦有差异。 SDF- 1α和 MCP- 1在 D2SC和 TEC 1C8中未出现可见扩增条带;在 TEC 1C8中也未见 KC的扩增条带。重组趋化性细胞因子 SDF- 1α、 IP- 10、 MCP- 1和 RANTES对胸腺细胞的趋化指数分别为 3.7、 4.5、 6.2和 2.6。结论不同的胸腺基质细胞系,其趋化性细胞因子的表达谱和表达强度不同, 4种重组趋化性细胞因子对胸腺细胞表现出不同的趋化活性。

Multiple types of chemokines expressed in mouse thymic stromal cell lines

OBJECTIVE: To investigate semi-quantitative mRNA expression of SDF-1 alpha, IP-10, KC, MCP-1, and RANTES in thymic stromal cell lines of MTEC1, MTDC, D2SC, MTECB, TEC 1C8, TNC as well as in the primary thymic stromal cell cultures. The chemotactic activities of recombinant SDF-1 alpha, IP-10, MCP-1, and RANTES to mouse thymocytes were detected. METHODS: Using beta-actin as internal control, the mRNA of the chemokines listed above were amplified for 30 cycles with RT-PCR. The amplified products were observed by agarose electrophoresis, and each band was analyzed with integrated optical density. With the method of Boyden chamber assay, the chemotactic activities of recombinant SDF-1 alpha, IP-10, MCP-1, and RANTES were detected to thymocytes, and the chemotactic indices were calculated. RESULTS: The expression intensity of SDF-1 alpha, IP-10, KC, MCP-1, and RANTES varied from each other in the stromal cell lines detected. The PCR products of SDF-1 alpha and MCP-1 were not seen in D2SC or TEC 1C8, nor was the band of KC observed in TEC 1C8 either. The chemotactic indices of recombinant SDF-1 alpha, IP-10, MCP-1, and RANTES to thymocytes were 3.7, 4.5, 6.2, and 2.6, respectively. CONCLUSIONS: Different thymic stromal cell lines could express different types of chemokines with different expression intensities. To thymocytes, recombinant SDF-1 alpha, MCP-1, and IP-10 showed strong chemotactic activities, while the chemotactic activity of RANTES was very weak.