鸡肺泡上皮Ⅱ型细胞的培养与鉴定  

Culture and identification of alveolar epithelial type Ⅱ cells from chicken

作　　者：侯海燕[1] 赵立红[1] 乔健[1] 金继昌[1] 赵慧颖[1] 胡永峰[1] 

机构地区：[1]中国农业大学动物医学院,北京100094

出　　处：《中国农业大学学报》2008年第1期6-10,共5页Journal of China Agricultural University

摘　　要：为研究禽流感等敏感病毒及其他致病因子对肺泡上皮Ⅱ型细胞（AEC-Ⅱ）的致病性,建立了鸡AEC-Ⅱ细胞体外培养方法。采用胰蛋白酶和Ⅳ型胶原酶联合消化法对18日龄鸡胚肺组织进行消化,经差速离心和免疫黏附,获得了鸡AEC-Ⅱ细胞,再经纯化、体外培养和碱性磷酸酶染色法鉴定。结果表明,鸡AEC-Ⅱ细胞在接种后18 h开始贴壁进入适应期,细胞为圆形,呈岛屿状生长;接种后24-72 h,进入对数生长期,细胞为多边形,并融合成单层,胞浆内有明显的颗粒;接种后第4天细胞数量达到高峰（1.38×10^5个/mL）,并进入稳定生长期,胞浆内颗粒开始减少;接种后6-7 d,细胞进入衰退期,胞浆内颗粒显著减少,细胞逐渐死亡、脱落。另外,4代（F4）之前的AEC-Ⅱ传代细胞贴壁时间早,形态均一,纯度高。说明接种后24-72 h内获得的AEC-Ⅱ原代细胞和F4之前的传代细胞可以供体外试验研究。The aim of the study was to develop a method for the isolation, purification, identification and primary culture of alveolar epithelial type Ⅱ cells （AEC-Ⅱ ）as a new model for studying the pathogenicity of some etiopathogenisis （such as avian influenza virus） on AEC-Ⅱ , The lung tissue of 18 day embryonic chickens was digested with trypsin and collagenase Ⅳ. AEC- Ⅱ was isolated, purified by differential centrifugation and specific immunoabsorption, then cultured, The AEC-Ⅱ was identified by alkaline phosphatase （AKP） staining based on cell biochemical properties, The results showed that excellent yields and highly purified, culturable AEC-Ⅱ could be obtained. AEC-Ⅱ adhered 18 hour post culture （h, p. c） and reached the adaptation phase, when cells were rotund-shaped and grew in islands shape, At 24 to 72 h, p, c, cells were polygon-shaped, formed confluent monolayers and enter the logarithmic phase, with fine particles within the cytoplasm. At 4 days post culture, we obtained 1.38×10^5 cells per milliliter, with cells entering the stationary phase, when the particles began to decrease. At 6 to 7 days post culture, particles decreased significantly and cells were at the declining phase. Before onset of the 4 phases, cells were better than primary culture cells in purity and morphology. Thus, we considered that a reliable method for primary culture of AEC- Ⅱ from chickens was developed, and the primary culture cell during 24 to 72 h, p, c as well as cells before the 4 phases are suitable for in vitro studies .

关 键 词：鸡胚 肺泡上皮Ⅱ型细胞 原代培养 传代培养 细胞生长曲线 细胞鉴定 

分 类 号：S852.2[农业科学—基础兽医学]