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| A primary culture method of human renal carcinoma cells and normal cells based on RNA-seq validation |
| ZHOU Xiaoguang WANG Haozhou LI Yansheng SONG Liming |
| Department of Urology, Beijing Chaoyang Hospital, Capital Medical University, Beijing 100020, China |
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Abstract Objective To establish a more rapid and convenient method for primary culture of renal tumor and normal renal tubular epithelial cells. Methods Fresh clear renal cell carcinoma tissue and renal cortical tissue from patients undergoing radical nephrectomy and partial nephrectomy in Beijing Chaoyang Hospital, Capital Medical University from January to March 2019 were selected. The tumor and normal primary cells were purified by mechanical segmentation, collagenase digestion and continuous cell screening and purification. The genetic background of the two primary cells was identified by transcriptomic sequencing (RNA-seq) technique, the expression of cell markers was identified by immunocytochemistry, and the ultrastructure of the primary cells was observed by transmission electron microscopy. Finally, the tumorigenesis characteristics of tumor cells were identified by the model of nude mice bearing tumors. Results A total of three human proximal tubular epithelial primary cells and eight renal clear cell carcinoma primary cells were isolated and purified. RNA-seq showed that the sequencing results were consistent with the genetic background of the two types of cells, and immunocytochemistry could identify the expression of marker proteins in the proximal renal tubular epithelial cells. Two kinds of cell specific ultrastructure could be observed by transmission electron microscope. The tumor bearing model of nude mice confirmed the tumorigenic properties of primary renal cell carcinoma cells. Conclusion A rapid and convenient processes and methods for primary culture of renal carcinoma cells and renal tubular epithelial cells is successfully established.
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