Macrophages overexpressing interleukin-10 alleviate atherosclerosis by targeting the lactate-CEBPB axis to inhibit pathogenic VSMC phenotypic transitions
M Wang, S Zhou, Y ChenAbstract
Background
Vascular smooth muscle cell (VSMC) plasticity is a critical determinant of atherosclerotic plaque vulnerability. Within the plaque microenvironment, contractile VSMCs switch into pathogenic states, notably macrophage-like and fibroblast activation protein (FAP)-positive phenotypes, accelerating necrotic core expansion and fibrous cap thinning. However, the overarching metabolic-epigenetic mechanisms governing this divergent reprogramming remain insufficiently characterized.
Purpose
This study aimed to elucidate whether the ox-LDL-lactate-histone H3 lysine 18 lactylation (H3K18la)-CCAAT/enhancer-binding protein beta (CEBPB) axis epigenetically drives VSMC phenotypic transitions, and to investigate the therapeutic efficacy of plaque-homing, interleukin-10-overexpressing engineered macrophages (IL-10M) in attenuating this cascade.
Methods
In vivo models utilized smooth muscle lineage-tracing mice (TgMyh11-CreERT2/ROSAtdT/+) with PCSK9-AAV8 and a high-fat diet. Primary murine VSMCs stimulated with ox-LDL were used in vitro. Methodologies included single-cell RNA sequencing (scRNA-seq), Cleavage Under Targets and Tagmentation (CUT&Tag), Seahorse metabolic flux analysis, adeno-associated virus 9 (AAV9)-mediated VSMC-specific CEBPB knockdown, and the administration of IL-10M.
Results
scRNA-seq analysis identified CEBPB as one of the central transcriptional hubs enriched in the intermediate VSMC phenotype. Mechanistically, ox-LDL triggered metabolic reprogramming in VSMCs, characterized by enhanced glycolysis and lactate accumulation. This stress prompted a p300-dependent elevation in global pan-lysine lactylation, specifically enriching nuclear H3K18la, which directly localized to the Cebpb promoter to drive its transcription. Furthermore, CUT&Tag and RT-qPCR analyses demonstrated that ox-LDL enhanced CEBPB binding at promoters proximal to Fap, Reck, and Cxcr2 driving a pro-atherogenic program. Concurrently, increased CEBPB enrichment at Abca1 suggested parallel activation of compensatory cholesterol efflux pathways. In vivo, AAV9-mediated VSMC-specific CEBPB knockdown preserved the contractile phenotype and diminished modulated VSMC proportions. Therapeutically, intravenously delivered IL-10M successfully homed to plaques, remodeling the immune-metabolic microenvironment. Sustained IL-10 release via STAT3 signaling reversed the VSMC lactate burden, suppressing the downstream H3K18la-CEBPB program. Consequently, IL-10M therapy—particularly combined with direct CEBPB inhibition—synergistically reduced macrophage-like and FAP+ VSMC generation, yielding substantial plaque stabilization through decreased necrotic cores and enhanced fibrous caps.
Conclusions
The lactate-H3K18la-CEBPB axis acts as a critical mechanism orchestrating VSMC phenotypic divergence. Targeted delivery of IL-10M effectively modulates this metabolic-epigenetic cascade, offering a promising cellular intervention paradigm for alleviating atherosclerosis.