Quantification of fusion events between MSC and epithelial cells by flow cytometry analysis.

For direct coculture, eGFP or DsRED MSC (PM7) cells were cultured with epithelial (DsRED AGS, eGFP HFE-145 and DsRED HT-29) cells for 8 days, harvested, stained with anti-ESA antibodies, detected by Alexa-647 labelled secondary antibodies. For indirect coculture, PM7 and epithelial cells (AGS) were separated by a 0.4 µm cell culture insert and processed as for direct coculture. Cells were then analyzed by flow cytometry. Results represent the mean ± SD of one experiment with three replicates representative of at least three different experiments.

于直接共培养体系中，将表达增强型绿色荧光蛋白（enhanced Green Fluorescent Protein, eGFP）或红色荧光蛋白（Discosoma Red Fluorescent Protein, DsRED）的PM7株间充质干细胞（mesenchymal stem cells, MSC），与上皮细胞（包括DsRED标记的AGS细胞、eGFP标记的HFE-145细胞及DsRED标记的HT-29细胞）共同培养8天；收集细胞后，采用抗上皮特异性抗原（Epithelial Specific Antigen, ESA）抗体进行染色，再以Alexa-647标记的二抗完成检测。于间接共培养体系中，将PM7细胞与上皮细胞（AGS）通过0.4 μm孔径的细胞培养插入物进行分隔，其余处理步骤与直接共培养体系一致；随后通过流式细胞术（flow cytometry）对细胞进行分析。实验结果以单次实验三次生物学重复的平均值±标准差表示，该实验可代表至少三次独立重复实验的结果。