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. 2014 Apr 14;15(4):6286–6297. doi: 10.3390/ijms15046286

Figure 1.

Figure 1.

Citicoline treatment does not induce cytotoxic effect in primary retinal cultures. Primary retinal cultures, treated or not with 100 μM citicoline for 96 h, were immunolabeled with antibodies against rhodopsin (A,D rhod), GABA (B,E) and CRALBP (C,F), in order to evidence photoreceptors, GABAergic neurons and Müller glia, respectively. Cell nuclei were counterstained with Hoechst 33258. Antibody immunoreactivity was unaffected in citicoline-treated samples, showing that the agent does not influence cell composition and differentiation in primary retinal cultures (D–F); Apoptosis was evaluated by TUNEL assay. Apoptotic nuclei were quantified as percentage of TUNEL-positive nuclei over total nuclei. Bars represent the mean ± SEM of at least five independent experiments (G); Absence of proapoptotic effects after citicoline treatment is also shown by unchanged levels of activated cleaved caspase 3 (H). Whole cell lysates were prepared from primary retinal cultures. Equal amounts of total protein from each lysate were resolved on 15% SDS-PAGE and transferred to nitrocellulose membranes. Membranes were probed with anti-cleaved caspase 3. β-actin levels were used as control of protein loading. Blots are representative of three independent experiments. Bars = 20 μm.