关键词: 膀胱癌, 　外泌体, 　增强型绿色荧光蛋白, 　示踪

Abstract: Objective　To obtain EGFP labeling exosomes of bladder cancer cells for tracking exosomes in vitro by constructing a bladder cancer cell line stably expressing CD63-EGFP fusion protein. Methods　The CD63 gene was inserted into pLVX-EGFP-puro plasmid by molecular cloning to construct the expression vector of CD63-EGFP fusion protein. Bladder cancer cell lines stably expressing CD63-EGFP fusion protein were obtained by lentivirus transfection. Exosomes were extracted by ultracentrifugation and identified by transmission electron microscopy, particle size analysis and Western blot. The uptake of bladder cancer cell exosomes by SV-HUC-1 cells and peripheral blood mononuclear cell (PBMC) was detected through co-culture. Results　A Bladder cancer cell line stably expressing the CD63-EGFP fusion protein was constructed, and the secreted exosomes were consistent with the classic characteristics of exosomes. SV-HUC-1 cells and Monocytes in PBMC can uptake bladder cancer cell-derived exosomes in vitro. Conclusion　Bladder cancer cells expressing CD63-EGFP fusion protein can secrete fluorescent exosomes, which can be used for tracing of exosomes in vitro.

Key words: Bladder cancer, 　Exosomes, 　Enhanced green fluorescent protein, 　Tracing