p27Kip1 and RPE Cell Cycle

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DESCRIPTION (provided by applicant): Abnormal growth or turnover of the retinal pigment epithelium (RPE) is associated with an alarming number of ocular disorders (i.e., proliferative vitreo-retinopathy (PVR), age-related macular degeneration, developmental hamartomas, congenital hypertrophy of the RPE), accounting for significant health costs. An understanding of the basic mechanisms for establishing and maintaining appropriate epithelial cell number and size is crucial to finding ways to alleviate disease consequences and to facilitate their repair. The aim of the proposed research is to define the mechanisms that regulate growth, through cell multiplication and enlargement of cell size, during RPE development in situ. While many exogenous influences can regulate cell proliferation, they do so by ultimately impinging on machinery of the cell division cycle. Consequently, the proposed studies focus on mechanisms operating in the RPE cells as they withdraw from the cell cycle during differentiation of the epithelial layer in vivo. Experiments will make use of a mutant mouse with targeted disruption of the gene coding for the cell cycle regulatory protein p27Kip1, in which there is an abnormal expansion of the RPE rnonolayer. The overall hypothesis is that loss of p27Kip1 leads to an increase in the number or density of epithelial cells as a result of an enhancement in the rate of cell proliferation, or of a delay in the timing of cell cycle exit prior to differentiation. This hypothesis will be tested using immunohistochemical techniques to determine the magnitude of cell division, as well as cell death, at various developmental ages. The alternative hypothesis that the thickening of the RPE layer is due to an increase in cell size will be examined by morphometric analysis of images obtained by confocal microscopy.