Myocardial ischemia-reperfusion injury (MIRI) is a common pathophysiological process in reperfusion therapy following myocardial infarction. It exacerbates myocardial damage and negatively impacts patient prognosis. This study aims to explore potential regulatory targets and related mechanisms in MIRI. Using the GSE123342 dataset, differentially expressed genes (DEGs) related to myocardial infarction-associated conditions were screened and a protein-protein interaction network was constructed. Core genes were identified through weighted gene co-expression network analysis (WGCNA), followed by GO and KEGG enrichment analyses. The GSE6381 dataset was used to examine core gene expression, and ROC curve analysis was performed as an exploratory assessment of group separation. A transcription factor regulatory network for CFTR was constructed using the KnockTF database. An oxygen-glucose deprivation/reoxygenation (OGD/OGR) model was established in AC16 cells, and plasmid overexpression of YTHDF3 and CFTR was used to examine their effects on OGD/OGR-induced cell injury. The interaction between YTHDF3 and CFTR mRNA was examined by RIP-qPCR experiments, and CFTR mRNA stability was assessed using an actinomycin D assay. Additionally, the effects of YTHDF3 and CFTR overexpression were examined in a rat I/R model. We found in the GSE123342 dataset, 5444 differentially expressed mRNAs were identified, with CFTR significantly upregulated. Correlation analysis revealed 6651 genes related to CFTR. WGCNA identified 120 hub genes, with the MEyellow module significantly associated with MIRI. These genes were mainly enriched in ABC transporters and the AMPK pathway. In the GSE6381 dataset, CFTR and YTHDF3 showed differential expression and ROC analysis showed preliminary group separation. CFTR was closely related to multiple transcription factors, including TP53, STAT3 and TFAP4. In AC16 cells, OGD/OGR decreased YTHDF3 expression and increased CFTR expression. YTHDF3 overexpression reduced CFTR expression and aggravated OGD/OGR-induced injury, as shown by reduced cell viability and proliferation and increased apoptosis. Additional CFTR overexpression partly attenuated these changes. In the rat I/R model, YTHDF3 overexpression increased myocardial infarct size and impaired cardiac function, whereas CFTR overexpression attenuated the injury associated with YTHDF3 overexpression. CFTR knockdown aggravated OGD/OGR-induced cellular injury and was associated with changes in AMPK signalling. Together, these findings suggest that YTHDF3 may be associated with MIRI and CFTR-related changes. Changes in AMPK signalling were also observed, but the underlying mechanism still needs further validation. These results provide preliminary evidence for a possible relationship between YTHDF3 and CFTR in MIRI.