Wagner L M, Takemoto D J
Department of Anatomy and Physiology, Kansas State University, Manhattan, KS 66506, USA.
Mol Vis. 2001 Mar 7;7:57-62.
To determine if Protein Kinase C (PKC) plays a role in the initiation of lens epithelial cell differentiation into a lens fiber cell.
PKCalpha or PKCgamma was overexpressed in N/N 1003A lens epithelial cells for up to 7 days. Phase contrast microscopy was used to observe morphological changes associated with PKCalpha or PKCgamma overexpression. Cell cycle changes in cells overexpressing PKCalpha or PKCgamma were measured using acridine orange staining and flow cytometry. Crystallin levels in cells overexpressing PKCalpha or PKCgamma were measured using Western blots and RT-PCR.
Significant differences in cell cycling were observed between untransfected cells and those overexpressing PKCalpha or PKCgamma. Overexpression of PKCalpha and PKCgamma caused the cells to lose their epithelial-like appearance and elongate. alphaB-crystallin expression was detected in all the samples while alphaA-crystallin was detected only in cells after 7 days of PKCalpha or PKCgamma overexpression.
The observations that alphaA-crystallin is only found in N/N 1003A cells overexpressing PKCalpha or PKCgamma for 7 days along with the finding that a block in the G0/G1 phase of the cell cycle and the consequent morphological changes are observed, indicate that PKCalpha and PKCgamma may have a role in the initiation of differentiation in lens epithelial cells.
确定蛋白激酶C（PKC）是否在晶状体上皮细胞分化为晶状体纤维细胞的起始过程中发挥作用。
在N/N 1003A晶状体上皮细胞中过表达PKCα或PKCγ长达7天。利用相差显微镜观察与PKCα或PKCγ过表达相关的形态学变化。使用吖啶橙染色和流式细胞术测量过表达PKCα或PKCγ的细胞中的细胞周期变化。使用蛋白质免疫印迹法和逆转录-聚合酶链反应测量过表达PKCα或PKCγ的细胞中的晶状体蛋白水平。
未转染细胞与过表达PKCα或PKCγ的细胞之间在细胞周期方面观察到显著差异。PKCα和PKCγ的过表达导致细胞失去其上皮样外观并伸长。在所有样品中均检测到αB-晶状体蛋白表达，而仅在过表达PKCα或PKCγ 7天后的细胞中检测到αA-晶状体蛋白。
αA-晶状体蛋白仅在过表达PKCα或PKCγ 7天的N/N 1003A细胞中被发现，同时还观察到细胞周期的G0/G1期阻滞以及随之而来的形态学变化，这些观察结果表明PKCα和PKCγ可能在晶状体上皮细胞分化的起始过程中发挥作用。
