Fig. 5: BCAAs elicited podocyte metabolic reprogramming by inducing PKM2 depolymerization and inactivation. | Nature Communications

Fig. 5: BCAAs elicited podocyte metabolic reprogramming by inducing PKM2 depolymerization and inactivation.

From: Branched-chain amino acids contribute to diabetic kidney disease progression via PKM2-mediated podocyte metabolic reprogramming and apoptosis

Fig. 5

a, b The protein expression of GCK, PFK, PKM1, and PKM2 in primary podocytes (a) and isolated glomeruli (b) from mice. c Representative Western blots of cross-linked PKM2 in primary podocytes, n = 3. Tetra, tetramer; Di, dimer; Mono, monomer. d Representative Western blots of PKM2 phosphorylation in primary podocytes. e Representative Western blots of cross-linked PKM2 in primary podocytes, 20 mM mannitol (Man) plus 5 mM glucose were used as osmotic control, n = 3. f, g Relative pyruvate kinase activities in primary podocytes. n = 6. h Upper, ECAR in primary podocytes were determined; bottom: measurements of the post-glucose, post-oligomycin A, and reserved capacity of ECAR. n = 5. i Upper, OCR of primary podocytes were determined; bottom, statistical analyses of baseline respiratory capacity, ATP-coupled respiratory capacity, maximum respiratory capacity, and reserve respiratory capacity in the OCR. 2-DG, 2-deoxyglucose. FCCP, cyanide-4-(trifluoromethoxy) phenylhydrazone. RE, rotenone. Resp, respiration. n = 5. j Enrichment pathway analysis via RNA sequencing of MPC-5 podocytes (HG/BCAA + TEPP46 vs HG/BCAA) revealed the top 7 pathways with upregulated component expression (red) and the top 7 pathways with downregulated component expression (green) using Gene Ontology (GO) annotations and modified Fisher’s exact test. The numbers in parentheses indicate the number of genes in each GO term biological process. k The mRNA expression of skeleton-related genes in primary podocytes. n = 6 mice. l Representative immunofluorescence images of F-actin expression in differentiated MPC-5 podocytes. m The expression of 27 out of 33 genes whose expression in the HG/BCAA group was reversed by administration of 10 μM TEPP46. The numbers on the right of the heatmap indicate the fold changes. The data were analyzed by the Mann–Whitney U test. n Schematic metabolic map of glucose metabolism in podocytes treated with HG, HG/BCAA, or HG/BCAA + TEPP46. o The mRNA expression of the indicated genes in primary podocytes. n = 6. p Protein expression in isolated glomeruli from mice. The data were presented as mean ± SD, and analyzed by unpaired two-tailed Student’s t test (j, m) or one-way ANOVA (f–i, k, o).

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