Early transcriptomic responses to revascularization in ischemic human myocardium

Abstract Background Prolonged myocardial ischemia triggers a ’wave front’ of cardiomyocyte death due to oxygen and nutrient deprivation. Thus, timely reperfusion and salvage of ischemic myocardium are the primary objectives of revascularization therapies for patients with flow-limiting coronary artery disease. However, despite its well-established benefits, evidence suggests that reperfusion itself can inflict additional injury to the affected myocardium, which remains a significant concern. Purpose This study aimed to explore the early transcriptomic responses to reperfusion in the myocardium of patients undergoing coronary artery bypass grafting (CABG) surgery. Methods Eleven male patients with NSTE-ACS and significant stenosis (90%) of the left anterior descending (LAD) artery, with an ejection fraction (EF) greater than 40%, were included. During the CABG surgery, myocardial biopsies were collected from the anteroseptal region of the left ventricle at two timepoints: before and five minutes after graft placement (reperfusion). Total RNA was isolated from the cardiac tissue, and RNA libraries were prepared using the NEB Next Ultra kit and sequenced on an Illumina Novaseq platform. Low-quality reads were filtered and mapped to GRCh38 with HISAT2. Transcripts were assembled with StringTie, and read counts were normalized to FPKM. Differential expression analysis (reperfusion vs. ischemia) was performed with DESeq2 (log2FC ≥ ±1, p-adj ≤ 0.05). Protein-coding genes underwent GSEA using GenePattern with MSigDB signatures. Results A paired-sample comparison identified 214 differentially expressed protein-coding genes (DEGs) - 177 significantly downregulated and 37 upregulated during reperfusion (Figure 1A). Of these, 14 DEGs met the criterion ofp-adj 0.05 (Table 1). The most upregulated were the transcripts of early response genes Fos and FosB (FosB Proto-Oncogene, AP-1 Transcription Factor Subunit). The most downregulated DEG was endothelial cell-specific molecule 1 (ESM1; log2FC = -1.94), a pro-inflammatory factor involved in angiogenesis and secreted by inflamed endothelium. Downstream enrichment KEGG and GO analyses revealed reperfusion-induced upregulation of genes related to mitochondrial ATP production, cardiac contraction, and antioxidant activity (GO Molecular Function Antioxidant activity; NES=1.91, p-adj=0.019). Conversely, genes associated with cellular and nuclear division, inflammation, the p53 mediated apoptotic pathway, and cellular response to glucose starvation were downregulated (Figure 1B and C). Conclusions Reperfusion led to beneficial transcriptomic changes, upregulating mitochondrial energy production, contractile and antioxidant mechanisms, while downregulating pro-inflammatory, pro-fibrotic, and pro-apoptotic pathways. These findings highlight the predominantly positive transcriptomic responses that drive myocardial recovery following revascularization.Figure 1. Table 1.