Yu Keming, Ma Ping, Ge Jian, Willey Christopher D, Yang Peizeng, Wang Zhichong, Gao Qianying
Key Laboratory of Ophthalmology, Zhongshan Ophthalmic Center, Sun Yat-sen University, Ministry of Education, Guangzhou 510060, China.
Graefes Arch Clin Exp Ophthalmol. 2007 Jul;245(7):993-9. doi: 10.1007/s00417-006-0467-3. Epub 2006 Nov 24.
Protein kinase C (PKC) is involved in both physiological and pathophysiological processes and plays an important role in signal transduction. The present studies were designed to examine the 12 isoforms (PKCalpha, PKCbetaI, PKCbetaII, PKCgamma, PKCdelta, PKCepsilon, PKCeta, PKCtheta, PKCmu, PKCxi, PKClambda and PKCiota) of PKC expressed in cultured human retinal pigment epithelium (RPE) cells.
Human RPE cells were investigated for 12 PKC isoforms at the mRNA, protein and cellular levels by reverse transcription (RT)-PCR, Western blot analysis and laser scanning confocal microscope (LSCM), respectively.
RT-PCR and Western blot analyses showed similar results for specific PKC isoforms in that both revealed that PKCalpha, PKCbetaI, PKCbetaII, PKCdelta, PKCepsilon, PKCtheta, PKCmu, PKCxi, PKClambda and PKCiota, but not PKCgamma and PKCeta, were constantly expressed in RPE cells, with the exception of PKCbetaI at the protein level. Confocal microscopy showed that ten PKC isoforms - PKCalpha, PKCbetaI, PKCbetaII, PKCdelta, PKCepsilon, PKCtheta, PKCxi, PKCiota, PKClambda and PKCmu - appeared almost exclusively in the cytoplasm of the cells. However, PKCgamma and PKCeta were not detected by staining.
This study characterized the expression pattern of all 12 PKC isoforms and showed that ten of these (PKCalpha, PKCbetaI, PKCbetaII, PKCdelta, PKCepsilon, PKCtheta, PKCmu, PKCxi, PKClambda and PKCiota) are present in cultured human RPE cells. This identification provides the first step towards elucidating their roles in RPE cell proliferation.
蛋白激酶C（PKC）参与生理和病理生理过程，在信号转导中起重要作用。本研究旨在检测培养的人视网膜色素上皮（RPE）细胞中表达的12种PKC亚型（PKCalpha、PKCbetaI、PKCbetaII、PKCgamma、PKCdelta、PKCepsilon、PKCeta、PKCtheta、PKCmu、PKCxi、PKClambda和PKCiota）。
分别通过逆转录（RT）-PCR、蛋白质印迹分析和激光扫描共聚焦显微镜（LSCM）在mRNA、蛋白质和细胞水平上研究人RPE细胞中的12种PKC亚型。
RT-PCR和蛋白质印迹分析对特定PKC亚型显示出相似的结果，即两者均显示PKCalpha、PKCbetaI、PKCbetaII、PKCdelta、PKCepsilon、PKCtheta、PKCmu、PKCxi、PKClambda和PKCiota在RPE细胞中持续表达，但PKCgamma和PKCeta除外，蛋白质水平的PKCbetaI除外。共聚焦显微镜显示，十种PKC亚型——PKCalpha、PKCbetaI、PKCbetaII、PKCdelta、PKCepsilon、PKCtheta、PKCxi、PKCiota、PKClambda和PKCmu——几乎只出现在细胞的细胞质中。然而，通过染色未检测到PKCgamma和PKCeta。
本研究对所有12种PKC亚型的表达模式进行了表征，并表明其中十种（PKCalpha、PKCbetaI、PKCbetaII、PKCdelta、PKCepsilon、PKCtheta、PKCmu、PKCxi、PKClambda和PKCiota）存在于培养的人RPE细胞中。这一鉴定为阐明它们在RPE细胞增殖中的作用迈出了第一步。
