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. Author manuscript; available in PMC: 2023 Mar 1.
Published in final edited form as: Free Radic Biol Med. 2022 Feb 5;181:143–153. doi: 10.1016/j.freeradbiomed.2022.01.032

Figure 1. Altered phosphorylation indicating reduced activity of glucose metabolism regulatory enzymes in the retinas of rd10 mice.

Figure 1.

Immunostaining demonstrates expression of the PDHE1-A isoform of pyruvate dehydrogenase (red) in the inner plexiform layer (IPL), outer plexiform layer (OPL) and photoreceptor inner segments (IS) of pig retina (A). There is staining for PDHE1-A within cone matrix sheaths labeled with FITC-labeled peanut agglutinin (green) demonstrating PDHE1-A expression in cones (B, C). Postnatal day (P) 35, wild type C57BL/6 mice and rd10 mice in a C57BL/6 background were euthanized and retinal homogenates were prepared. (D) Immunoblot of retinal homogenates from 7 C57BL/6 mice and 7 rd10 mice probed with anti-pPDHE1-A (S293), anti-PDHE1-A, and anti-Actin antibodies. Densitometry of the blot in (D) and a blot from a repeat experiment demonstrated a significant increase in pPDHE1-A/PDHE1-A ratio (p=0.0339) (E) and a significant decrease in PDHE1-A/Actin ratio (F) in rd10 vs wild type retinas (n=14). (G) Immunoblot of retinal homogenates from 7 C57BL/6 mice and 7 rd10 mice probed with anti-pPKM2 (Y105), anti-PKM2, and anti-Actin antibodies. Densitometry of the blot in (G) and a blot from a repeat experiment demonstrated a significant increase in pPKM2/PKM2 ratio (H) and a significant decrease in PKM2/Actin ratio (p=0.0236) (I) in rd10 vs wild type retinas (n=14). (J) Immunoblot of retinal homogenates from 7 C57BL/6 mice and 7 rd10 mice probed with anti-LDHA (Y10), anti-LDHA, and anti-Actin antibody. Densitometry of the blot in (J) demonstrated a significant reduction in pLDHA/LDHA ratio (K) and a significant decrease in LDHA/Actin ratio (p=0.0274) (L) in rd10 vs wild type retinas (n=7). Real-time RT-PCR showed increased expression of mRNA for pyruvate dehydrogenase kinase 2 (PDK 2, M) and branched chain ketoacid dehydrogenase kinase (p=0.0134, BCKDK, N) in the retinas of rd10 mice compared to wild type retinas.

* p ≤ 0.05, ** p ≤ 0.01, *** p ≤ 0.001, **** p ≤ 0.0001 by Mann-Whitney test

PDHE1-A, Pyruvate dehydrogenase E1-A subunit; PKM2, isoform of pyruvate kinase; LDHA, isoform of lactate dehydrogenase; ONL, outer nuclear layer; INL, inner nuclear layer, IS, inner segment of photoreceptor; RPE, retinal pigmented epithelium.