Mechanisms of Diabetic Myocardial Injury Through the Lipid Droplet–Mitochondrial Pathway

Background: Dysregulated lipid metabolism in diabetes contributes to myocardial lipotoxicity, mitochondrial dysfunction, and inflammatory cell death. This study investigated the mechanisms by which abnormal lipid metabolism in diabetes induces mitochondrial oxidative stress, and whether the lipid droplet (LD)–mitochondrial pathway is involved in cardiomyocyte pyroptosis during diabetic cardiomyopathy (DCM). Methods: db/db and db/m mice were evaluated for blood glucose and lipid levels, myocardial histopathologic characteristics, and cardiac function. Differentially expressed genes were identified by transcriptomic analysis, followed by Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses. Immunohistochemistry and TUNEL staining were performed to assess inflammatory responses and cell death, respectively. Myocardial lipid deposition, LD distribution, mitochondrial ultrastructure, and perilipin 5 (PLIN5) expression were examined. In vitro, H9c2 cardiomyocytes were transfected with a PLIN5 overexpression plasmid. Immunofluorescence and Real-Time Quantitative PCR (RT-qPCR) were used to assess pyroptosis-related markers, including NLR Family, Pyrin Domain Containing Protein 3 (NLRP3), caspase 1, and Gasdermin Domain Containing 1 (GSDMD). Lipid peroxidation and reactive oxygen species (ROS) levels were measured using fluorescence-based assays. Oxygen consumption rate (OCR) was assessed using a Seahorse XFe96 analyzer, and LD–mitochondrial colocalization was evaluated by laser confocal microscopy. Results: Compared with db/m mice, db/db mice showed significantly increased body weight and serum levels of glucose, total cholesterol (TC), triglyceride (TG), and low-density lipoprotein cholesterol (LDL-C), together with reduced high-density lipoprotein cholesterol (HDL-C) levels (p < 0.01). The db/db mice also exhibited aggravated pathological changes and reduced ejection fraction (EF) and fractional shortening (FS) (p < 0.01). Transcriptomic analysis showed significant upregulation of fatty acid-binding protein 3 (FABP3), Long-chain acyl-CoA synthetase 1 (ACSL1), Medium-chain acyl-CoA dehydrogenase (ACADM), acyl-CoA thioesterase 2 (ACOT2), and PLIN5 in myocardial tissue. db/db mice exhibited marked myocardial lipid accumulation and increased LD abundance, accompanied by mitochondrial structural abnormalities and altered LD–mitochondrial co-localization. Total PLIN5 protein and Plin5 mRNA expression were significantly increased, whereas mitochondrial-associated PLIN5 protein expression was significantly decreased (p < 0.01). In palmitic acid (PA)-treated H9c2 cardiomyocytes, immunofluorescence showed a significant increase in pyroptosis-positive cells, and protein levels of NLRP3, interleukin-1 beta (IL-1β), interleukin-6 (IL-6), interleukin-18 (IL-18), and tumor necrosis factor-α (TNF-α) (p < 0.05 or p < 0.01) were significantly increased. PLIN5 overexpression significantly reduced lipid peroxidation and ROS accumulation (p < 0.01), decreased maximal OCR, increased LD–mitochondrial co-localization, and reduced pyroptosis (p < 0.01). PLIN5 overexpression also significantly decreased NLRP3, caspase-1, and GSDMD protein and mRNA levels (p < 0.01). Conclusions: PLIN5 overexpression increased LD–mitochondrial co-localization and was associated with reduced lipid peroxidation, ROS accumulation, and activation of the NLRP3/caspase-1/GSDMD pathway in PA-treated cardiomyocytes. The PLIN5-regulated LD–mitochondrial pathway may therefore represent a mechanism linking abnormal lipid metabolism to cardiomyocyte pyroptosis and a potential therapeutic target for further investigation in DCM.