This study attempted to establish miRNA expression profiles in acute myocardial infarct (AMI) sheep model with left ventricular assist device (LVAD) unloading. AMI was established in sheep model and FW-II type axial flow pump was implanted to maintain continuous unloading for 3 days. The cardiomyocyte survival, inflammatory cell infiltration, and myocardial fibrosis were detected by tissue staining, and cardiomyocyte apoptosis was detected by TUNEL assay. High throughput sequencing technique was used to detect miRNA expression in cardiomyocytes and to establish miRNA expression profile. The Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses were established. miRNA sequencing results identified 152 known mature miRNAs and 1582 new mature miRNAs. The unloading and control groups differentially expressed genes, of which RT-PCR verified oar-miR-19b and oar-miR-26a. The GO and KEGG pathway annotation and enrichment established that the regulating functions and signaling pathways of these miRNAs were closely related to cardiovascular diseases (CVD). In this study, LVAD effectively reduced the cell death degree of cardiomyocyte in MI. The established miRNA expression profiles of AMI and LVAD intervention in this study suggest that the expression profile could be used to explore the unknown miRNA and the regulatory mechanisms involved in AMI.