FIG. 1.

MN induced rosette formation in immortalized corneal endothelial cells. (A) Schematic representation of MN degradation into a stable quinone in the presence of NQO1. HCEnC-21T cells were treated with various doses of MN and cellular viability was analyzed at 0, 1, 2, and 4-h time points. (B) Cellular viability of MN-treated HCEnC-21T cells in a time and dose–response analysis using trypan blue dye exclusion assay. Phase-contrast images were captured of HCEnC-21T cells treated with (C) STS, 4 h (D) 50 μM MN, showing rosette formation at 4 h. (E) cellular morphology of a postkeratoplasty human FECD specimen (47 male, * guttae). (F) Immunofluorescence image of clusterin staining around guttae (*) in an FECD specimen (74 female) and (J) around rosette-like formation in HCEnCs. Captured images of rosette formation at different time intervals, starting at (G) 1 h, (H) 2 h, and (I) 4 h after 50 μM MN treatment. Data are expressed as the mean ± SE (n = 6). Two-way ANOVA applied for statistical significance comparing treated samples (2, 4 h) versus 1 h (*p < 0.05, ***p < 0.001) and 25, 50, 100 μM treated versus control, 0 (≠p < 0.0001). FECD, Fuchs endothelial corneal dystrophy; HCEnC, human corneal endothelial cell; MN, menadione; NQO1; NAD(P)H quinone dehydrogenase 1; STS, staurosporine. To see this illustration in color, the reader is referred to the web version of this article at www.liebertpub.com/ars