Separation and identification of rat renal tubular epithelial cells and analysis of its proliferation ability

Objective: To establish a method to separate and identify the rat renal tubular epithelial cells (RTECs) and analyze the proliferation ability of the obtained cells, and to provide theoretical basis and technique support for analysis experiment in vitro of kidney diseases such as kidney injury and so on. Methods: The renal tubules were separated with type II collagenase digestion and purified with Percoll density gradient centrifugation and cultured in normal environment, when the RTECs grew out from the tubule, the cytokeratin-18 (CK-18) expression was detected with immunohistochemistry and the expressions of CK-18 in P0, P1 and P2 cells were detected with flow cytometry. The proliferation ability of each generation was evaluated with CCK-8 method. Results: After cultured for 24 h, the RTECs began to grow outside from the tubule and they looked like paving road stones after cultured for 72 h. In the process of passage, the expression efficiencies of CK-18 were 95.9%, 91.5%, and 76.8% in P0, P1 and P2 RTECs, and the RTECs were dead gradually. The proliferation ability of P2 cells was more weaken than that of P1 cells (P<0.05), and more mixed cells existed in the culture system. Conclusion: The method established in this study is cheap, credited as well as convenient. However, the cells just could be passaged for 2 times and couldn't be cultured for long time. Only the cells of P0 and P1 could be applied in analysis experiment in vitro.