Mutation in the ßA3/A1-crystallin gene impairs phagosome degradation in the retinal pigmented epithelium of the rat.

Phagocytosis of the shed outer segment discs of photoreceptors is a major function of the retinal pigmented epithelium (RPE). We demonstrate for the first time that ßA3/A1-crystallin, a major structural protein of the ocular lens, is expressed in RPE cells. Further, by utilizing the Nuc1 rat, in which the ßA3/A1-crystallin gene is mutated, we show that this protein is required by RPE cells for proper degradation of outer segment discs that have been internalized in phagosomes. We also demonstrate that in wild-type RPE, ßA3/A1-crystallin is localized to the lysosomes. However, in the Nuc1 RPE, ßA3/A1-crystallin fails to translocate to the lysosomes, perhaps because misfolding of the mutant protein masks sorting signals required for proper trafficking. The digestion of phagocytized outer segments requires a high level of lysosomal enzyme activity, and cathepsin D, the major enzyme responsible for proteolysis of the outer segments, is decreased in mutant RPE cells. Interestingly, our results also indicate a defect in the autophagy process in the Nuc1 RPE, which is probably also linked to impaired lysosomal function, because phagocytosis and autophagy might share common mechanisms in degradation of their targets. ßA3/A1-crystallin is a novel lysosomal protein in RPE, essential for degradation of phagocytosed material.

Expression patterns of beta1-related alpha integrin subunits in murine lens during embryonic development and wound healing.

PURPOSE: To study the expression patterns of b1-related alpha integrin subunits in murine lens epithelial cells, comparing embryonic fiber differentiation with injury-induced epithelial mesenchymal transition (EMT). METHODS: Adult mice type C57BL/6, pregnant as well as with an eye injured, were sacrificed at different time-course intervals. The embryonic and the injured eyes were obtained and deparaffinized sections of these eyes were processed for immunohistochemistry staining for detection of integrin a subunits. RESULTS: Embryonic lens epithelial cells expressed primarily a3 and a5 subunits, whereas embryonic fiber cells expressed a2, a5, and a6 subunits. Adult lens epithelial cells expressed a3, and a6 subunits,whereas injured lens cells expressed a2, a3, and a6 integrin subunits. CONCLUSIONS: The phenotypic changes of lens epithelial cells during embryonic fiber differentiation and EMT are characterized by different expression of integrin subunits as a result both of the altered extracellular matrix conditions and of the altered cell signaling pathways recruited in each process.

Biological compatibility of polymethyl methacrylate, hydrophilic acrylic and hydrophobic acrylic intraocular lenses.

PURPOSE: Extensive clinical investigations of the biocompatibility of different intraocular lenses (IOLs) have been made in an effort to optimize the outcome of modern cataract surgery. The aim of this study was to add animal eye experimental implantation data regarding cellular reaction on the anterior surface of IOLs. METHODS: Thirteen adult albino rabbits had phacoemulsification/aspiration of the crystalline lens followed by implantation of a posterior chamber IOL in each eye. Three types of IOLs were studied: Hydroview (Bausch and Lomb; n = 7), Acrysof (Alcon, USA; n = 7), and polymethyl methacrylate (PMMA; HOYA, Japan; n = 7). The animals were killed by intravenous pentobarbital 1, 4, or 8 weeks later. The IOLs were explanted and stained with hematoxylin and eosin, and observed under a light microscope. The shape of mouse ascites-induced macrophages on the anterior surface of the three different IOL types (Hydroview, PMMA, and Acrysof) was studied after 24 h of oven culture. RESULTS: Hydrophilic acrylic IOLs showed the highest affinity for lens epithelial cell (LEC) outgrowth, and the lowest and slowest maturation rate reaction of macrophages. PMMA IOLs showed the lowest affinity for LEC outgrowth, and the highest reaction of macrophages. Hydrophobic acrylic IOLs showed intermediate results both regarding LECs and macrophages. CONCLUSIONS: Results suggest that IOL biomaterial properties are the key factor that influences the quantity of monocytes/macrophages as well as the process of their maturation/senescence. LEC outgrowth is influenced both by the biomaterial of IOLs and by the monocyte/macrophage reaction.

Accumulation of thrombospondin-1 in post-operative capsular fibrosis and its down-regulation in lens cells during lens fiber formation.

Thrombospondin-1 (TSP-1) is a glycoprotein involved in activation of latent transforming growth factor beta (TGFbeta) expression. We examined changes in its expression pattern during human capsular opacification (PCO) and anterior subcapsular cataractogenesis (ASC), as well as in a healing injured mouse lens. Its expression pattern was also compared in a mouse embryonic lens with that in an adult lens. Based on immunohistochemistry under light microscopy, TSP-1 expression and other matrix components were evident in the anterior epithelium of an uninjured human lens, whereas fiber-differentiating cells in the equator of human lens lack TSP-1 immunoreactivity. In contrast, in post-operative human lens epithelial or fibroblastic cells, there was TSP-1 immunoreactivity, whereas it decreased in fiber-differentiating cells in PCO. Matrix components accumulated on the healing capsule also labeled with anti-TSP-1 antibody like antibodies against collagen I, IV, V and laminin. In uninjured, injured mouse lens epithelial cells and its matrix, there was TSP-1 expression. Embryonic lens cells in the posterior pole, undergoing differentiation to fiber cells, began to express TSP-1 protein at embryonic day (E) 11.5 whereas anterior epithelial cells started to express it at E13.5 in association with marked expression in central fiber cells. At E16.5, TSP-1 was detected in fibers just beneath the anterior epithelium, but the fiber mass showed minimal expression. At E18.5 and post-natally day 1, lens fiber TSP-1 expression was no longer seen. On the other hand, it was evident in both intact human anterior epithelial and dispersed mouse cells. The results indicate that there is TSP-1 expression in uninjured human and mouse lens epithelial cells and their fibrous tissue. In contrast, in post-operative lens cells differentiating to fiber cells, its expression levels decline. Further study is needed to clarify the roles of TSP-1 in modulating lens cell phenotype expression.