Figure 6. Blue light damage induced cell death in the outer nuclear layer of retinal explants. Photoreceptor cell death was quantified
using the terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate (dUTP) nick end labeling (TUNEL) assay.
All vibratome sections are counterstained with 4',6-diamidino-2-phenylindole dihydrochloride (DAPI). Red represents cells
with DNA strand breaks labeled with tetramethylrhodamine (TMR)-dUTP, and blue represents DAPI-stained nuclei. The figure shows
retinal sections with their layers (outer segments [OS], outer nuclear layer [ONL], outer plexiform layer [OPL], inner nuclear
layer [INL], inner plexiform layer [IPL], ganglion cells [GC]). A: The untreated retina (0 h) was incubated with complete TUNEL reaction mixture (negative control). B: Retinas were treated with DNase I before labeling (positive control). Retinas were not treated (C) or were irradiated (D) with visible blue light for 6 h. Cell death initiation was detected in the ONL after 6 h. Cell death initiation was detected,
especially in the outer nuclear layer (ONL). Retinas were not treated (E) or were irradiated (F) with visible blue light for 18 h. The number of TUNEL-positive cells significantly increased in blue light-damaged retinas
after 18 h, although some TUNEL-positive cells were detected in control explants due to the culture conditions. The scale
bar represents 50 µm. In part G, quantitative analysis of cell death in the ONL of retinal explants after blue light exposure for 6 h and 18 h is presented.
The graph displays the number of TUNEL-positive cells in the ONL of 100 µm sections from middle and peripheral parts of the
retina. The mean values of at least eight different counted sections per time point and treatment are shown. TUNEL-positive
cells appear initially at 6 h and increase during prolonged incubation. At 18 h, the TUNEL-positive cells in the ONL of blue
light-treated eyes are significantly more numerous than the cells observed in the time-matched controls (*p<0.05). Error bars
represent SD.