Journal: Investigative Ophthalmology & Visual Science
Article Title: Essential Role of CD63 in Maintaining Corneal Epithelial Identity in the Human Limbus
doi: 10.1167/iovs.67.3.25
Figure Lengend Snippet: CD63 is expressed in the human limbal epithelium and regulates corneal epithelial gene expression. ( A ) Representative KRT12 (corneal epithelial cell marker, green ), KRT13 (conjunctival epithelial marker, yellow ), and CD63 ( red ) immunostaining in the human conjunctiva, limbus, and central cornea. Hoechst 33342 ( blue ) was used for the nuclei staining ( n = 3 donors). Scale bar = 50 µm. ( B ) The bar graph represents CD63 RNA expression in CD63 siRNA-treated cultured limbal epithelial cells ( n = 4 donors, mean ± SD; *** P < 0.001, **** P < 0.0001). ( C , left panel ) Representative flow cytometry analyses of CD63 protein expression in CD63 siRNA-treated cultured limbal epithelial cells detected by flow cytometry. ( C , right panel ) The bar graph represents quantitative analyses of CD63 protein expression determined by flow cytometry ( n = 3 donors, mean ± SD; * P < 0.05). ( D , left panel ) Representative macroscopic images of colonies formed by CD63 KD cells compared to the control siRNA-transfected cells. ( D , right panel ) The bar graph represents a comparative analysis of the colony-forming efficiency ( n = 6 donors, mean ± SD; * P < 0.05, ** P < 0.01). ( E ) Comparative analyses of the percentage of EdU-positive proliferating cells by CD63 KD in cultured human limbal epithelial cells ( n = 5 donors, mean ± SD; ** P < 0.01). ( F ) A Venn diagram illustrates the overlap of differentially expressed genes identified in both CD63 KD#1 and CD63 KD#2 conditions. ( G ) Bar graphs represent normalized counts of CD63 , KRT12 , CLU , ALDH1A1 , ALDH3A1 , TGFBI , and MYEOV RNA expression in CD63 siRNA-treated cultured limbal epithelial cells ( n = 3 donors, mean ± SD, *adjusted P < 0.05, **adjusted P < 0.01, ***adjusted P < 0.001, ****adjusted P < 0.0001). ( H , left panel ) Western blot analyses of KRT12, CLU, ALDH1A1, ALDH3A1, and TGFBI expression in control and CD63 siRNA-treated limbal epithelial cells. ( H , right panel ) Bar graphs represent quantitative analyses of protein expression ( n = 4 donors, mean ± SD; * P < 0.05, ** P < 0.01, *** P < 0.01, **** P < 0.0001).
Article Snippet: GAPDH (Hs99999905_m1), CD63 (Hs01041237_g1), PAX6 (Hs01088114_m1), KRT12 (Hs00165015_m1), CLU (Hs00971656_m1), ALDH1A1 (Hs00946916_m1), ALDH3A1 (Hs00964880_m1), and TGFBI (Hs00932747_m1) TaqMan probes were used.
Techniques: Gene Expression, Marker, Immunostaining, Staining, RNA Expression, Cell Culture, Flow Cytometry, Expressing, Control, Transfection, Western Blot
R&D Systems is a verified supplier 
R&D Systems manufactures this product 
