Identification of caveolae and their signature proteins caveolin 1 and 2 in the lens.
This study shows that caveolae are present in lens epithelia of rabbit and guinea pig under normal conditions. Caveolae are unique lipid membrane microdomains observed in many cell types. They are believed to play crucial roles in a variety of basic physiological functions including signal transduction, lipid and transcellular transport. Using TEM, immunocytochemistry and immunoblotting we show for the first time the existence of caveolae and the co-localization of their signature marker integral proteins, caveolin-1 and caveolin-2, in the intact lens of rabbit and guinea pig. Thin-section TEM shows that among several species studied, lens epithelia of rabbit and guinea pig exhibited a large number of caveolae. The caveolae were pear shaped, approximately 70 nm in diameter, and were found frequently along the lateral membranes of epithelial cells in the intact lens. In the intact cortical fibers, only a small number of caveolae was seen in the superficial cells. In cultured lens epithelial cells, however, caveolae were observed along all membrane surfaces, but were more abundant at the apical membrane of the cells. Immunofluorescence and immunoblot analyses confirmed the presence of caveolin-1 and caveolin-2 in the lens epithelium. In addition, caveolin-1 and caveolin-2 co-exist in the lens epithelium of both rabbit and guinea pig. HRP tracer study demonstrated that caveolae could carry out endocytosis, suggesting their involvement in molecular transport. Cultured rabbit lens epithelial cells (line N/N1003A) were used to examine the response of caveolae to methyl-beta-cyclodextrin (MBCD), a specific cholesterol-depleting drug. The lens epithelial cells were incubated in freshly prepared MEM medium plus 8% rabbit serum containing 10mm MBCD for 0 (control), 15, 30 or 60 min. Controls for MBCD treatment were cultured in MEM plus 8% rabbit serum. MBCD treatment for 30 min revealed that depletion of cholesterol abolished the majority of caveolae in cultured lens epithelial cells. This result strongly suggests that caveolae are cholesterol-rich lipid rafts that are likely to play important roles in the lens.
Lo
Woo-Kuen
WK
Department of Anatomy and Neurobiology, Morehouse School of Medicine, 720 Westview Drive SW, Atlanta, GA 30310, USA. lowk@msm.edu
Zhou
Cheng-Jing
CJ
Reddan
John
J
D000818:Animals; D021941:Caveolae / Q000502:physiology / Q000648:ultrastructure*; D051242:Caveolin 1; D051243:Caveolin 2; D022461:Caveolins / Q000378:metabolism*; D002784:Cholesterol / Q000502:physiology; D003459:Crystallins / Q000378:metabolism*; D046508:Culture Techniques; D004847:Epithelial Cells / Q000378:metabolism / Q000648:ultrastructure; D005260:Female; D006168:Guinea Pigs / Q000033:anatomy & histology / Q000378:metabolism; D006735:Horseradish Peroxidase; D007908:Lens, Crystalline / Q000378:metabolism / Q000648:ultrastructure*; D008297:Male; D008854:Microscopy, Electron / Q000379:methods; D011817:Rabbits / Q000033:anatomy & histology / Q000378:metabolism
D051242:Caveolin 1; D051243:Caveolin 2; D022461:Caveolins; D003459:Crystallins; D002784:Cholesterol; D006735:Horseradish Peroxidase