Richards R J, Davies N, Atkins J, Oreffo V I
Lung. 1987;165(3):143-58. doi: 10.1007/BF02714430.
A method is described for the isolation of rat lung epithelial Type II cells using trypsin digestion of tissue to release cells for subsequent separation by Percoll gradient centrifugation. Both the concentration of trypsin and the age (body weight) of the rat affect the yield from primary digestion and the final number of Type II cells obtained. A lung weighing 1 g from a 200 g rat yields approximately 30 X 10(6) washed Type II cells (approximately 25% of the total estimated lung population). These cells have a plating efficiency of 40-50% after 48 h of culture. The cells have a high alkaline to acid phosphatase ratio (usually greater than 4.0) compared with that of alveolar macrophages (0.1) and accumulate putrescine by an active transport mechanism with an apparent KM between 8 and 14 microM. Together with studies of [3H]thymidine uptake into DNA, which is maximal between 48 and 72 h of culture, these quantitative measurements form a good basis for investigating the interactions between a number of chemical agents and Type II cells in vitro.
本文描述了一种分离大鼠肺上皮II型细胞的方法，即通过胰蛋白酶消化组织来释放细胞，随后利用Percoll梯度离心法进行分离。胰蛋白酶的浓度以及大鼠的年龄（体重）都会影响初次消化的产量和最终获得的II型细胞数量。一只体重200克大鼠的1克肺组织大约可产生30×10⁶个经洗涤的II型细胞（约占估计的肺细胞总数的25%）。这些细胞在培养48小时后的接种效率为40 - 50%。与肺泡巨噬细胞（0.1）相比，这些细胞的碱性磷酸酶与酸性磷酸酶的比值较高（通常大于4.0），并且通过一种主动转运机制积累腐胺，其表观Km值在8至14微摩尔之间。结合对培养48至72小时期间DNA中[³H]胸苷摄取量的研究，这些定量测量为体外研究多种化学试剂与II型细胞之间的相互作用提供了良好的基础。
