ACOT1-specific expression modulates metabolic reprogramming in diabetic cardiomyopathy: The role of SREBP1c lactylation in CD36-mediated lipotoxicity

Figures

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  Fig. 1. From lipid uptake to oxidation: Comprehensive transcriptomic analysis reveals lipid metabolism as the nexus of metabolic reprogramming in diabetic cardiomyopathy. A. Transcriptomic analysis of amino acid metabolism-related genes in db versus Tr hearts; B. Transcriptomic profiling of glucose metabolism-related genes in db versus Tr myocardium; C-E. Transcriptomic analysis of oxidative phosphorylation-, lactate metabolism-, and lipid metabolism-related genes in diabetic cardiomyopathy; F—H. Transcriptomic profiling of 18 critical genes across six core metabolic pathways in DCM and detailed analysis of lactate metabolism- and TCA cycle-related genes among these pathways; I—K.Analysis of regulatory patterns of the six core metabolic pathways, identification of core driver genes, and schematic illustration of the mechanistic cascade of metabolic reprogramming in DCM.

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  Fig. 2. ACOT1 as a critical regulator of lipid metabolism in diabetic cardiomyopathy. A. KEGG pathway enrichment analysis revealing lipid metabolism and energy-sensing signaling pathways as the core transcriptional signature of DCM, with AMPK as a central integrator; B—C. KEGG pathway enrichment analysis of DCM heart transcriptomic data, highlighting biosynthesis of unsaturated fatty acids as the most significantly enriched metabolic pathway and identifying ACOT1 as a critical node; D-G. Prognostic screening of lipid metabolism candidates via KM Plotter, showing elevated ACOT1 expression is associated with improved survival and adequate statistical power throughout the observation period.

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  Fig. 3. ACOT1 upregulation in early-stage DCM confers cardioprotection. A-C. ACOT1 protein and mRNA expression are progressively induced in DCM hearts from weeks 16–18 (P < 0.05–0.01). D-E.HFD/STZ-induced DCM mice exhibit obesity and hyperglycemia. F-G. Cardiac-specific ACOT1 overexpression reduces ANP (P < 0.001) and LDH (P < 0.05) in DCM but not control mice. H—I.ACOT1 improves FS and EF in DCM. J-K. ACOT1 attenuates fibrotic (Col1a1, Col3a1, Tgfβ, Ctgf) and hypertrophic (Nppa, Nppb, Myh7, Acta1) gene expression. Data represent mean ± SEM; P values determined by Student's t-test or one-way ANOVA.

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  Fig. 4. ACOT1 downregulation in late-stage DCM marks transition to decompensated heart failure. A-B. ACOT1 protein and mRNA expression progressively decline in DCM hearts from weeks 19–22 (P < 0.05–0.01). C. Sustained ACOT1 overexpression impairs FS and EF in late-stage DCM. D-E. ACOT1 elevation increases ANP and LDH levels (P < 0.05) in DCM but not control mice. F. Schematic illustrating biphasic ACOT1 function. G-H. ACOT1 upregulates fibrotic (Col1a1, Col3a1, Tgfβ, Ctgf) and hypertrophic (Nppa, Nppb, Myh7, Acta1) gene expression. I.ACOT1 selectively elevates circulating FFA (P < 0.05) without altering TC, TG, or HDL—C. Data represent mean ± SEM; P values determined by one-way ANOVA or Student's t-test.

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  Fig. 5. ACOT1 modulates lactate-induced SREBP-1c activation and CD36 upregulation. A. Lactate dose-dependently induces Pre-SREBP1c in H9C2 cells. B. Co-immunoprecipitation showing physical interaction between endogenous ACOT1 and SREBP-1c in AC16 cells under high-lactate conditions. C. ACOT1 overexpression synergizes with lactate to enhance Pre-SREBP1c protein levels. D. ACOT1 knockdown attenuates lactate-induced Pre-SREBP1c expression. E-G. Lactate elevates Srebp1c mRNA, with ACOT1 overexpression producing synergistic activation (~2.5-fold). H. SREBP-1c knockdown reverses ACOT1-mediated induction of Srebp1c, Cd36, and nSrebp1c. I. Combined ACOT1 overexpression and lactate maximally induces Pre-SREBP1c, nSREBP1c, and CD36 proteins. I. Immunofluorescence confirms ACOT1 is required for lactate-driven CD36 upregulation. Data represent mean ± SEM; P values determined by one-way ANOVA.

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  Fig. 6. SREBP-1c undergoes lactylation at conserved lysine 716 residue. A-C. Immunoprecipitation and Pan-Kla detection experiments confirming Pre-SREBP1c is post-translationally modified by lactylation, with lactylation intensity increasing concentration-dependently with exogenous lactate and signal specificity verified by IgG controls; E-H. Computational prediction of candidate lactylation sites on Pre-SREBP1c, mass spectrometry analysis identifying K716 as the predominant lactylation site, evolutionary conservation analysis of K716 across species, and site-directed mutagenesis confirming K716 as the primary lactylation residue.

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  Fig. 7. ACOT1 Modulates lipotoxicity via SREBP-1c K716 lactylation-dependent CD36 upregulation. A. ACOT1 enhances Pre-SREBP1c lactylation and CD36 expression; knockdown abrogates this effect. B.K716R mutation or HDAC treatment blocks ACOT1-SREBP-1c-CD36 axis. C.ACOT1 synergizes with lactate to induce CD36 mRNA; knockdown eliminates lactate responsiveness.D.ACOT1-lactate cooperation requires intact SREBP-1c and K716. E-F.ACOT1 upregulates cardiac CD36 protein and mRNA specifically in DCM. G.ACOT1 coordinately induces CD36 (uptake) and SCD1 (synthesis). H. Myocardial FFA peaks with sustained ACOT1 in late DCM. I. Immunofluorescence confirms K716 lactylation is essential for CD36 membrane localization. Data represent mean ± SEM; P values determined by one-way ANOVA.