〔Abstract〕 Objective To investigate the improvement of endoplasmic reticulum stress mediated by microRNA(miR)-34a-5p/transmembrane fusion protein 1(Notch1) signaling axis on hypoxia/reoxygenation(H/R) human cardiomyocytes. Methods Human cardiomyocytes were randomly divided into Control group, H/R group, mimic NC group, mimic group, inhibitor NC group and inhibitor group. Except the Control group, H/R injury model was established in other groups. The expression levels of miR-34a-5p and Notch1 were detected by quantitative real-time polymerase chain reaction(qRT-PCR), cell survival rate was detected by thiazolyl blue(MTT), cell apoptosis rate was detected by flow cytometry, and the targeting relationship between miR-34a-5p and Notch1 was detected by dual luciferase gene reporting method. The expressions of transcriptional activator 6(ATF6), inositol demand enzyme 1(IRE 1), protein kinase-like endoplasmic reticulum kinase(PERK) and glucose regulatory protein 78(GRP78) were detected by Western blot.Results miR-34a-5p targeted Notch1(P<0. 05); compared with Control group, the expression level of miR-34a-5p, apoptosis rate and protein expressions of ATF6, IRE1, PERK and GRP78 in H/R group increased, while the cell survival rate and Notch1 mRNA and protein expressions decreased(P<0. 05). Compared with H/R group and mimic NC group, miR-34a-5p expression, apoptosis rate, and proteinexpressions of ATF6, IRE1, PERK and GRP78 in mimic group increased, while cell survival rate and Notch1 mRNA and protein expressions decreased(P<0. 05). Compared with H/R group and inhibitor NC group, the expression of miR-34a-5p, apoptosis rate and protein expressions of ATF6, IRE1, PERK and GRP78 decreased in inhibitor group, while cell survival rate and Notch1 mRNA and protein expressions increased(P<0. 05).Conclusion miR-34a-5p can inhibit the apoptosis of H/R human cardiomyocytes, possibly through the targeted inhibition of Notch1-mediated endoplasmic reticulum stress.