PRECLINICAL VALIDATION OF HIPSC-DERIVED INHIBITORY INTERNEURONS AS A REGENERATIVE THERAPY FOR CORTICAL STROKE

Stroke remains the leading cause of adult disability, with no therapies currently available that effectively promote recovery. Conventional transplantation approaches using neural progenitors have been limited by poor survival, migration, and integration in the poststroke environment. To address these challenges, we developed an innovative cell-based therapy using human induced pluripotent stem cell (hiPSC)–derived inhibitory interneurons. These cells were generated via small molecule inhibition of Wnt, TGF-β, and BMP pathways (termed hiPSC-3is), resulting in a differentiation trajectory resembling median ganglionic eminence (MGE)-derived interneurons. In vitro characterization confirmed expression of interneuron markers, including LHX6, somatostatin (SST), and GABA, along with functional GABA secretion, migratory responses, and inhibitory synaptic activity validated by microelectrode array (MEA) recordings. Single-cell RNA sequencing provided a comprehensive transcriptomic profile supporting interneuron identity. For in vivo studies, hiPSC-3i cells were transplanted into the stroke cavity of mice, encapsulated within a hyaluronan (HA) hydrogel containing clustered VEGF immobilized on heparin nanocapsules to enhance survival and integration. Transplants administered during both subacute (7 days) and chronic (1 month) stages post-stroke led to significant behavioral recovery, as demonstrated by the gridwalking task. Confocal imaging revealed structural integration, and electrophysiological recordings confirmed functional incorporation of transplanted interneurons into host circuitry. These findings demonstrate the feasibility of generating functionally mature interneurons from hiPSCs and provide preclinical evidence supporting their potential as a regenerative therapy for cortical stroke.