Transplanted Iron Oxide Nanoparticle-Labeled Mesenchymal Stem Cells Exhibit ex vivo Neuronal Firing Activity in Ischemic Stroke Rats

Mesenchymal stem cell (MSC) therapy shows promise in preclinical ischemic stroke models, yet clinical translation remains inconsistent. To address this gap, we investigated whether labeling MSCs with Ferucarbotran enables magnetic resonance imaging (MRI) tracking and enhances neural differentiation and functional integration, particularly focusing on the novel observation of spontaneous neuronal firing activity in transplanted cells. Rat MSCs (rMSCs) were transduced with red fluorescent protein (RFP) and labeled with Ferucarbotran to generate Fer-RFP⁺ rMSCs. These were transplanted into rats subjected to middle cerebral artery occlusion. MRI tracked cell migration and localization. Behavioral recovery was evaluated via the corner test, modified neurological severity score (mNSS), and infarct volume analysis. Post-transplantation, Fer-RFP⁺ rMSCs were magnetically isolated for ex vivo electrophysiological and immunocytochemical analyses. Ferucarbotran labeling did not impair rMSC viability and enhanced in vitro proliferation. MRI effectively visualized Fer-RFP⁺ rMSC migration to ischemic regions. Rats receiving Fer-RFP⁺ rMSCs showed significantly improved functional recovery and reduced infarct volumes compared to controls. Remarkably, ex vivo isolated Fer-RFP⁺ rMSCs exhibited spontaneous neuronal firing on multi-electrode array recordings and expressed the neuronal marker NeuN. Ferucarbotran-labeled MSCs not only serve as MRI-visible tracers but also exhibit neuronal electrophysiological properties post-transplantation in an ischemic stroke model. The emergence of spontaneous neuronal firing in ex vivo transplanted MSCs suggests functional neuronal differentiation, potentially underpinning the observed therapeutic effects. These findings offer new mechanistic insights into MSC-mediated stroke recovery and may enhance the translational relevance of MSC-based therapies.