Reddy Raghava, Buckley Sue, Doerken Melissa, Barsky Lora, Weinberg Kenneth, Anderson Kathryn D, Warburton David, Driscoll Barbara
Childrens Hospital Los Angeles Research Institute, Smith Research Towe, MS 35, 4650 Sunset Blvd., Los Angeles, CA 90027, USA.
Am J Physiol Lung Cell Mol Physiol. 2004 Apr;286(4):L658-67. doi: 10.1152/ajplung.00159.2003. Epub 2003 Aug 15.
Alveolar epithelial type 2 cells (AEC2) isolated from hyperoxia-treated animals exhibit increases in both proliferation and DNA damage in response to culture. AEC2 express the zonula adherens proteins E-cadherin, -, - and -catenin, desmoglein, and pp120, as demonstrated by Western blotting. Immunohistochemical analysis of cultured AEC2 showed expression of E-cadherin on cytoplasmic membranes varying from strongly to weakly staining. When cultured AEC2 placed in suspension were labeled with fluorescent-tagged antibodies to E-cadherin, cells could be sorted into at least two subpopulations, either dim or brightly staining for this marker. With the use of antibody to E-cadherin bound to magnetic beads, cells were physically separated into E-cadherin-positive and -negative subpopulations, which were then analyzed for differences in proliferation and DNA damage. The E-cadherin-positive subpopulation contained the majority of damaged cells, was quiescent, and expressed low levels of telomerase activity, whereas the E-cadherin-negative subpopulation was undamaged, proliferative, and expressed high levels of telomerase activity.
从高氧处理的动物中分离出的肺泡Ⅱ型上皮细胞(AEC2)在培养时增殖和DNA损伤均增加。蛋白质免疫印迹法显示,AEC2表达紧密连接蛋白E-钙黏蛋白、α-连环蛋白、β-连环蛋白、γ-连环蛋白、桥粒芯糖蛋白和pp120。对培养的AEC2进行免疫组织化学分析显示,E-钙黏蛋白在细胞质膜上表达,染色强度从强到弱不等。当用荧光标记的E-钙黏蛋白抗体标记悬浮培养的AEC2时,细胞可被分为至少两个亚群,该标记物染色呈弱阳性或强阳性。使用与磁珠结合的E-钙黏蛋白抗体,将细胞物理分离为E-钙黏蛋白阳性和阴性亚群,然后分析它们在增殖和DNA损伤方面的差异。E-钙黏蛋白阳性亚群包含大多数受损细胞,处于静止状态,端粒酶活性水平低,而E-钙黏蛋白阴性亚群未受损,具有增殖能力,端粒酶活性水平高。