Fig. 4.

The effect of flavonoids on levels of photoreceptor-specific markers in eyes of Abca4^−/−Rdh8^−/− mice exposed to bright light. Total RNA was isolated from the eyes of mice unexposed to light and mice treated with either DMSO, quercetin, or myricetin prior to illumination. Four or five mice were used for each treatment group. Eyes were collected on days 1, 3, and 7 post-treatment. RT-qPCR was performed to determine the mRNA expression levels of photoreceptor’s specific genes, such as rhodopsin (Rho) (A), M cone opsin (B), S cone opsin (C), and Mef2c transcription factor regulating the expression of Rho and cone opsins (D). Relative fold change of these genes’ expression was normalized to the expression of GAPDH. The mean of data from three independent experiments is shown as a fold change of NT control. The expression of these genes was slightly downregulated in DMSO-treated and illuminated mice, whereas it was significantly upregulated upon treatment with flavonoids. The statistically different changes in the expression of the specific gene compared on different days after treatment between DMSO-treated and flavonoid-treated mice are indicated with asterisk (*). The nonstatistically different changes are indicated as NS. Error bars indicate S.D. Statistical analysis was performed for each gene separately, combining data for all days and using the multivariate two-way ANOVA analysis and Turkey’s post hoc tests. Immunoblot analysis examining the changes in the protein expression of rhodopsin, cone opsins, and Mef2c in mouse eyes on day 1 and day 7 post-treatment. Twenty mice were used with at least four mice for each treatment group. Representative immunoblots are shown (E). Quantification of rhodopsin (F), M cone opsin (G), and Mef2c (H) protein expression levels. Protein bands detected in (E) were quantified by densitometry analysis with ImageJ software. Band intensities were normalized to the intensity of GAPDH. The mean of data from three independent experiments is shown. The statistically different changes in the expression of the specific protein compared between DMSO-treated control and flavonoid-treated mice on different days are indicated with asterisk (*). The nonstatistically different changes are indicated as NS. Error bars indicate S.D. Statistical analysis was performed for each gene separately, including data for all days together, using the multivariate two-way ANOVA analysis and Turkey’s post hoc tests. D, treated with DMSO vehicle; M, treated with myricetin; NT, nontreated; Q, treated with quercetin.