This study investigated the role of microRNA-21 (miR-21) and microRNA-24 (miR-24) in the pathological processes that follow cerebral ischemic injury and examined the potential use of miR-21 and -24 in stroke diagnostics as sensitive plasma biomarkers. An oxygen-glucose deprivation (OGD) model was constructed using mouse N2A neuroblastoma cells (N2A). Western blot analysis and quantitative polymerase chain reaction (qPCR) were employed to detect protein and miRNA expression levels. miR-21 and miR-24 were analyzed in the plasma from 68 patients with acute cerebral infarction (ACI) and 21 healthy individuals. In the present study, it was identified that plasma miR-21 and miR-24 were lower in ACI patients than in the controls (P<0.05). A positive correlation was demonstrated between plasma miR-21 and miR-24, and a negative correlation was revealed between miR-21, miR-24 and the National Institutes of Health Scales Score (NIHSS) within the first day following stroke. In addition, the expression of miR-21 and miR-24 was upregulated by 3.3- and 4.9-fold, respectively, when the reoxygenation time persisted up to 24 h following 3 h of OGD. The expression of Bcl-2 was upregulated following gain of miR-21 function, while X-linked inhibitor of apoptosis protein (XIAP) was downregulated after gain of miR-24 function in N2A cells. The data suggested that miR-21 may have an antiapoptotic effect in N2A neuroblastoma cells following OGD and reoxygenation, while miR-24 may have a pro-apoptotic effect. Therefore, these microRNAs may be potential therapeutic targets for the treatment of post-ischemic injury and may act as diagnostic markers during the early stage of ACI.