大鼠肝细胞分离、原代培养及生物学特性研究

Study on the isolation, primary culture and biologicial functions of rat hepatocytes

目的 :探讨体外原代培养大鼠肝细胞的分离方法、原代培养肝细胞的生物学特性。 方法 :以SD大鼠作肝细胞供者 ,采用胶原酶消化法分离获取肝细胞 ,并进行原代培养 ;以锥虫蓝染色法测细胞活力 ,在倒置显微镜下观察肝细胞形态变化 ,采用Beckman全自动生化分析仪检测各培养体系不同时间培养上清中Albumin、LDH、Urea的含量。 结果 :胶原酶消化法所获取的肝细胞形态完整、贴壁良好、活性高、功能强。LDH漏出量、白蛋白分泌及尿素合成功能在 1周内出现波动性变化 ,第 3天时LDH漏出量最低 ,白蛋白分泌及尿素合成功能较好。 结论 :胶原酶消化法为一较好的肝细胞分离方法 ,分离的肝细胞在培养第

Objectives:To investigate the methods of isolation of rat hepatocytes and observe the biologicial characteristic of primary culture rat hepatocytes. Methods:Hepatocytes were isolated from SD rat by collagenase digestion and cultured in a conditioned medium. The viability were assessed by trypan blue exclusion. The morphologic change of cultured hepatocytes was observed, the concentrations of albumin, urea and lactate dehydrogenase(LDH) in the supernatant in different cultural period were examined. Results: Hepatocytes obtained by collagenase digestion were intact and have a good viability and function. The LDH leakage, albumin synthesis and urea level fluctuate in one week,there is a lesser LDH leakage and higher albumin and urea level on the third day. Conclusions: The collagenase digestion method is feasible for isolation of hepatocytes. The primary culture rat hepatocytes have a good function in the third day.