CX3CR1对骨髓间充质干细胞修复受损肠上皮细胞的影响

Role of CX3CR1 in repair of injured intestinal epithelial cells by bone marrow mesenchymal stem cells

目的:探讨趋化因子受体CX3CR1对骨髓间充质干细胞(bone marrow-mesenchymal stem cells,BM-MSCs)修复受损肠上皮细胞的影响.方法:体外分离培养、鉴定BM-MSCs.采用肿瘤坏死因子-α(tumor necrosis factor a l p h a,T N F-α)处理结肠癌上皮细胞(c o l o n adenocarcinoma epithelial cell line,Caco-2)建立肠上皮损伤模型.实验分为6组:(1)单纯BM-MSCs组;(2)单纯Caco-2组;(3)Caco-2+TNF-α组;(4)BM-MSCs+Caco-2组;(5)BMMSCs+Caco-2+TNF-α组;(6)anti-CX3CR1-BM-MSCs+Caco-2+TNF-α组,利用Transwell培养板将各组共培养,通过免疫荧光、We s t e r n b l o t及荧光定量RT-P C R方法检测Caco-2上ZO-1(zonula occluden 1)、Occludin和BM-MSCs上CX3CR1蛋白及mR NA表达水平.结果:选择100 ng/mL TNF-α处理Caco-2细胞48 h建立损伤模型后,检测ZO-1和Occludin表达水平明显降低(P<0.05).在BM-MSCs与未受损的Caco-2共培养时ZO-1、Occludin和CX3CR1蛋白及m RNA的表达不受影响,而在BM-MSCs与受损Caco-2共培养时,其表达增加(P<0.05).阻断CX3CR1后,ZO-1、Occludin蛋白及m RNA的表达与未阻断前相比明显减少(P<0.05).结论:CX3CR1参与BM-MSCs对受损肠上皮的修复.

AIM： To investigate the role of chemokine receptor CX3CR1 in the repair of injured intestinal epithelial cells by bone marrow mesenchymal stem cells（BM-MSCs）. METHODS： BM-MSCs were cultured and identified in vitro. Caco-2 cells were exposed to tumor necrosis factor alpha（TNF-α） to establish a cell model of injured intestinal epithelium. Cells were divided into six groups： BM-MSCs, Caco-2 cells, Caco-2 cells treated with TNF-α, co-cultured BM-MSCs and Caco-2 cells, co-cultured BM-MSCs and Caco-2 cells treated with TNF-α, and co-cultured BM-MSCs and Caco-2 cells treated with anti-CX3CR1 and TNF-α. The expression of tight junction proteins and m RNAs in Caco-2 cells, and CX3CR1 protein and m RNA in BM-MSCs was detected by immunofluorescence, Western blot and RT-PCR. RESULTS： We selected 100 ng/mL TNF-α for 48 h to establish the injured model, because the expression of zonula occluden 1（ZO-1） and Occludin was reduced significantly at this time point（P〈0.05）. The protein and mR NA levels of ZO-1, Occludin and CX3CR1 had no significant changes when BM-MSCs were co-cultured with untreated Caco-2 cells, but increased when BMMSCs were co-cultured with injured Caco-2 cells（P〈0.05）. When CX3CR1 was blocked, the protein and mR NA levels of ZO-1 and Occludin decreased significantly.CONCLUSION： CX3CR1 participates in the repair of injured intestinal epithelial cells by BM-MSCs.