Corneal keratocytes: Phenotypic and species differences in abundant protein expression and in vitro light-scattering
Corneal keratocytes: Phenotypic and species differences in abundant protein expression and in vitro light-scattering
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DOI：
10.1167/iovs.04-1225
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发表时间：
2005-07-01
影响因子：
4.4
通讯作者：
Huang, JY
中科院分区：
文献类型：
--
作者：
Jester, JV;Budge, A;Huang, JY
目的。先前的研究表明，受伤后的角膜混浊涉及角膜细胞光散射特性的变化，这可能与水溶性蛋白质的丰富表达有关。本研究的目的是确定不同物种和不同培养兔角膜细胞表型的角膜细胞的蛋白表达模式，并评估体外光散射的差异。方法。从多个物种的角膜上皮细胞和角膜细胞中分离出水溶性蛋白质，包括人 (Hu)、小鼠 (Mo)、兔 (Ra)、鸡 (Ch) 和猪 (P) 以及不同培养的兔角膜细胞表型。然后通过 SDS-PAGE、胰蛋白酶肽序列分析和蛋白质印迹分析对蛋白质进行表征。分别用共焦反射和荧光显微镜测定培养细胞的光散射和肌动蛋白组织。结果。蛋白质表达模式在物种和细胞类型之间存在很大差异，其中鉴定出五种新的丰富表达蛋白质，包括 LDH (Ra, Ch)、G3PDH (Hu, Ch)、丙酮酸激酶 (Ch)、膜联蛋白 II (Ch) 和蛋白质二硫键异构酶 (Ch)。不同的兔角膜细胞表型也表现出不同水平的ALDH1A1和TKT表达，其中肌成纤维细胞的表达减少最多。肌成纤维细胞显示显着更大（P
Purpose. Previous studies suggest that corneal haze after injury involves changes in the light-scattering properties of keratocytes that are possibly linked to the abundant expression of water-soluble proteins. The purpose of this study was to determine the protein expression pattern of keratocytes from different species and different cultured rabbit keratocyte phenotypes and to assess differences in light-scattering in vitro.Methods. Water-soluble proteins were isolated from corneal epithelial cells and keratocytes of several species, including human (Hu), mouse (Mo), rabbit (Ra), chicken (Ch), and pig (P) and different cultured rabbit keratocyte phenotypes. Proteins were then characterized by SDS-PAGE, tryptic peptide sequence analysis, and Western blot analysis. Light-scattering and actin organization from cultured cells were determined with confocal reflectance and fluorescence microscopy, respectively.Results. Protein expression patterns varied substantially between species and cell types, with five new abundantly expressed proteins identified including, LDH (Ra, Ch), G3PDH (Hu, Ch), pyruvate kinase (Ch), Annexin II (Ch), and protein disulfide isomerase (Ch). Different rabbit keratocyte phenotypes also showed different levels of expression of ALDH1A1 and TKT, with myofibroblasts showing the greatest reduction. Myofibroblasts showed significantly greater (P