Localization of MIP 26 in Nuclear Fiber Cells from Aged Normal and Age-related Nuclear Cataractous Human Lenses

Abstract

The purpose of the current study was to localize the lens membrane protein, MIP 26, in nuclear fiber cells from different regions of aged normal and age-related cataractous human lenses. Adult, juvenile, fetal and embryonic nuclear regions in aged normal and age-related nuclear cataractous lenses were morphologically and biochemically characterized using the technologies of immuno-gold (5 nm) labeling, semi-thin sections (200–500 nm), serial sections, DiI staining following by photobleaching, transmission electron microscopy and spot-blot analysis. Numbers of gold particles per micron length of plasma membrane and numbers of gold particles per square micron of cytosol in the embryonic-fetal and juvenile-adult nuclear regions were quantified. Results showed that the labeling pattern of MIP 26 localized to the cytosol was unique to senescent fiber cells from age-related cataractous lenses. Numbers of gold particles per square micron of cytosol in the embryonic-fetal nucleus of age-related cataractous lenses were significantly elevated (P<0.001) above numbers from fiber cells located within the adult or juvenile nuclei of the same lens or senescent fiber cells from aged normal lenses. Some of the cytosolic labeling in cataracts was localized to lipid vesicles, while the remaining labeling was negative for the lipid specific stain DiI. Spot blot analysis demonstrated that binding of the ant-MIP 26 serum was exclusive to large molecular weight components greater than 10 kDa, and not to small molecular weight fragments of the protein. The results of the current study supply further evidence that damage to membranes occurs in senescent fiber cells during age-related nuclear cataracts, resulting in the internalization of structures containing the membrane protein MIP 26.