p62 expression is enhanced in AMD mice model.

APOE4 mice at the age of 18–20 months were fed a high fat, cholesterol enriched diet (HFC) for 2 months and control age-matched APOE4 mice were kept on a normal diet (ND), (A) Paraffin embedded mouse retina sections from each group (n = 6, disease and control groups) were incubated with monoclonal p62 antibody followed by secondary antibodies conjugated with AlexaFluor® 594 dye. Sections were covered with Vectashield mounting medium/DAPI. Photographs were taken by Zeiss AX10.1 Observer fluorescent microscope. (B) Protein lysates from RPE/Choroid layers (pooled left and right eye cup for each mouse) were analyzed by Western blot against p62 and LC3. 10 mice were used per group. β-actin was used as an internal control. Lane numbers denote the animal number in each group. Densitometric quantification of p62 band intensity was analyzed. Statistical significance between the normal diet (n = 10) and high-fat diet-fed groups (n = 10) was determined by ANOVA, p<0.05. (C) Protein lysates from the neural retina (pooled left and right retina for each mouse) were analyzed by Western blot against p62 and LC3. 10 mice were used per group. β-actin was used as an internal control. Densitometric quantification of p62, LC3 (II/I) ratio and total LC3 were plotted. Statistical significance between the normal diet (n = 10) and high-fat diet-fed groups (n = 10) was determined by ANOVA, p<0.05. (D) Real Time PCR analyses for Lc3 and p62 in mouse RPE/Choroid was performed with Gapdh as internal control.