Long-lasting survival, migration and functional maturation of transplanted human reprogrammed neurons in vivo

Direct reprogramming of glial progenitor cells (GPCs) into induced neurons (iNs) represents a promising strategy for future brain regenerative therapies. This approach uses forced expression of neuronal fate-determining genes to convert non-neuronal cells into neurons, bypassing a pluripotent intermediate stage and enabling potential in vivo applications. However, evidence for long-term survival and maturation of human reprogrammed neurons in vivo remains limited. Here, we present a longitudinal in vivo analysis of human iNs following transplantation into the immunodeficient mouse cortex. Using 3D cultures of reprogrammed human iNs to enhance graft survival, we demonstrate survival of human iNs for up to 5 months post-transplantation with a proportion of cells remaining up to 10 months. The transplanted cells exhibit widespread migration from the grafted core into anatomically connected brain regions and protracted neuronal maturation, reflected by increased cellular complexity and expression of neuronal markers. Functional analyses reveal gradual electrophysiological maturation and the establishment of afferent synaptic connectivity with host circuits. Importantly, non-reprogrammed GPC grafts retain their glial identity. Together, these findings provide in vivo evidence for long-term survival, neuronal maturation and long-range migration in the intact mouse brain, supporting their potential for future cell repair strategies.