Castleman W L, Northrop P J, McAllister P K
Department of Pathobiological Sciences, School of Veterinary Medicine, University of Wisconsin, Madison 53706.
Am J Vet Res. 1991 Jun;52(6):880-5.
The objectives of our research were to determine whether bovine pulmonary type-II alveolar epithelial cells could be isolated from bovine lung and maintained in tissue culture and to determine whether isolated bovine type-II alveolar epithelial cells would support productive viral replication of bovine parainfluenza type-3 virus and bovine respiratory syncytial virus. Type-II alveolar epithelial cells were isolated from lungs of 4- to 7-day-old male Holstein calves by enzymatic dissociation of pulmonary tissue with trypsin and by separation of cells with the use of filtration and centrifugation on continuous Percoll gradients. Cells were further separated by panning on IgG-coated plastic plates and by lectin binding. Isolated type-II alveolar cells were maintained on basement membrane-coated tissue cultured plates. In culture, type-II cells formed alveolar structures and maintained other cytologic features of type-II cells, including osmiophilic lamellar inclusions. Cell cultures were inoculated with and supported productive replication of bovine parainfluenza type-3 virus and bovine respiratory syncytial virus. This was determined by recovery of infectious viruses from inoculated cell cultures and by identification of viral structures in type-II alveolar epithelial cells by transmission electron microscopy.
我们研究的目的是确定能否从牛肺中分离出牛肺II型肺泡上皮细胞并在组织培养中维持其生长，以及确定分离出的牛II型肺泡上皮细胞是否支持牛副流感3型病毒和牛呼吸道合胞病毒的有效病毒复制。通过用胰蛋白酶对肺组织进行酶解，并利用过滤和在连续Percoll梯度上离心分离细胞，从4至7日龄雄性荷斯坦犊牛的肺中分离出II型肺泡上皮细胞。通过在IgG包被的塑料板上淘选和凝集素结合进一步分离细胞。将分离出的II型细胞培养在基底膜包被的组织培养板上。在培养过程中，II型细胞形成肺泡结构，并保持II型细胞的其他细胞学特征，包括嗜锇性板层小体。用牛副流感3型病毒和牛呼吸道合胞病毒接种细胞培养物，并支持其有效复制。这是通过从接种的细胞培养物中回收感染性病毒以及通过透射电子显微镜在II型肺泡上皮细胞中鉴定病毒结构来确定的。
