Acid-sensing ion Channel 1 (Asic1) is critical in acidotoxicity and significantly contributes to neuronal death in cerebral stroke. Pharmacological inhibition of Asic1 has been shown to reduce neuronal death. However, the potential of utilising exosomes derived from pluripotent stem cells to achieve inhibition of Asic1 remains to be explored. Developing qualified exosome products with precise and potent active ingredients suitable for clinical application is also ongoing. Here, we adopt small RNA sequencing to interrogate the miRNA contents in pluripotent stem cell-derived induced Mesenchymal Stem Cell (iMSC) derived exosomes. RNAseq was used to compare the oxygen-glucose deprivation (OGD)-damaged neurons before and after the delivery of exosomes. We used western blot to quantify the Asic1 protein abundance in neurons before and after exosome treatment. An in vivo test on rats validated the neuroprotective effect of iMSC-derived exosome and its active potent miRNA hsa-mir-125b-5p. We demonstrate that pluripotent stem cell-derived iMSCs produce exosomes with consistent miRNA contents and sustained expression. These exosomes efficiently rescue injured neurons, alleviate the pathological burden and restore neuron function in rats under oxygen-glucose deprivation (OGD) stress. Furthermore, we identify hsa-mir-125b-5p as the active component responsible for inhibiting the Asic1a protein and protecting neurons. We validated a novel therapeutic strategy to enhance acidosis resilience in cerebral stroke by utilising exosomes derived from pluripotent stem cells with specific miRNA content. This holds promise for cerebral stroke treatment with the potential to reduce neuronal damage and improve clinical patient outcomes.Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.