--- - |2- MicroRNA‐224 could alleviate the occurrence and development of acute lower extremity ischemia (ALEXI) in mice through activation of the mammalian target of rapamycin (mTOR) signaling pathway by downregulating C/EBP homologous protein (CHOP). Abstract Acute lower extremity ischemia (ALEXI) is known worldwide as an urgent condition, occurring when there is an abrupt interruption in blood flow into an extremity. This study aims to investigate whether microRNA‐224 (miR‐224) affects the ALEXI mice and the underlying mechanism. The miR‐224 expression and C/EBP homologous protein (CHOP), mammalian target of rapamycin (mTOR), translation initiation factor 4E‐binding protein 1 (4E‐BP1), and phosphoprotein 70 ribosomal protein S6 kinase (p70S6K) messenger RNA (mRNA), as well as protein expressions, were determined. The target gene of miR‐224 was also verified by using a luciferase reporter gene assay. The vascular endothelial cells from the ALEXI mice were transfected with miR‐224 mimics, miR‐224 inhibitors, or small‐interfering RNA against CHOP. Cell proliferation was assessed using a 3‐(4,5‐dimethylthiazol‐2‐yl)‐2,5‐diphenyltetrazolium bromide (MTT) assay. The cell cycle distribution along with the cell apoptosis were both evaluated by using a flow cytometry. The muscle fibers of the lower extremities found in the ALEXI mice were evidently swollen and rounded, presenting with a remarkably narrowed gap. The positive CHOP expression increased in ALEXI mice than normal mice, while the miR‐224 expression and mTOR, 4E‐BP1, and p70S6K mRNA, as well as the protein expression, decreased. Luciferase reporter gene assay validated that the miR‐224 gene directly targeted CHOP. MiR‐224 facilitated cell proliferation but inhibited cell apoptosis; by contrast, CHOP increased cell apoptosis. Moreover, the cells transfected along with miR‐224 mimic exhibited a lower CHOP expression as well as increased mTOR, 4E‐BP1, and p70S6K expression. Our study provided evidence that miR‐224 could alleviate the occurrence and development of ALEXI in mice through activation of the mTOR signaling pathway by downregulating CHOP. - 'Journal of Cellular Physiology, EarlyView. '