转染CXCR4基因支气管上皮细胞向SDF-1的迁移观察

Observation on chemotaxis towards SDF-1 of bronchi epithelial cell transfected CXCR4 gene

目的观察转染趋化因子受体CXCR4的人支气管上皮细胞HBE向趋化因子SDF-1的定向迁移情况。方法免疫荧光和流式细胞仪检测正常HBE细胞中的CXCR4蛋白。构建CXCR4慢病毒表达载体并转染HBE细胞。实时荧光定量PCR和Western blot检测转染CXCR4的HBE细胞中的CXCR4 mRNA和蛋白,通过Transwell小室实验观察该细胞向SDF-1的定向迁移情况。结果正常HBE细胞的CXCR4蛋白表达量较低。成功构建了CXCR4慢病毒表达载体。转染CXCR4的HBE细胞中CXCR4 mRNA和蛋白都有较高水平的表达,且向SDF-1定向迁移的细胞数[SDF-1终浓度分别为1、3、10、30、100、300 nM时到达膜背面的细胞分别为(42.8±2.8)(、47.6±6.2)(、57.8±3.3)(、62.4±5.0)(、89.8±4.7)(、100.0±4.8)(、62.4±5.0)个]较未加入SDF-1时的细胞数([32.8±2.2)个]显著增加(P均<0.05),并呈浓度依赖性(P<0.05)。结论转染趋化因子受体CXCR4的HBE细胞向趋化因子SDF-1的定向迁移明显增加。

Objective To observe the directional migration towards chemokine SDF-1 of Human Bronchi Epithelial（HBE） cells transfected chemokine receptor-CXCR4.Methods The CXCR4 protein was detected in HBE cells by immunofluorescence and flow cytometry.The CXCR4 lentiviral vector was contructed and transfected to HBE cells.The CXCR4 mRNA and protein in HBE cells transfected CXCR4 was detected by real-time quantitative PCR and Western blot.The HBE cells′ directional migration towards SDF-1 through Transwell chamber experiment was observed.Results The protein expression level of normal HBE cells was low.Succeeded in constructing the lentiviral vector of CXCR4.The CXCR4 mRNA and protein in transfected CXCR4 got a relative high level expression.The amount of cells directionally migrating to SDF-1 increased significantly compared to cells without SDF-1 added（all P0.05.The cells reached to back membrane were 42.8±2.8,47.6±6.2,57.8±3.3,62.4±5.0,89.8±4.7,100.0±4.8 with final concentration of SDF-1 respectively 1,3,10,30,100,300 nM） and this was concentration-dependent（P0.05）.Conclusion The directional migration towards chemokines SDF-1 of HBE cells transfected chemokine receptor-CXCR4 increased significantly.