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Transl. Med."],"published-print":{"date-parts":[[2026,4,29]]},"abstract":"<jats:p>Optic neuropathies cause irreversible vision loss. Transplantation of pluripotent stem cell (PSC)\u2013derived retinal ganglion cells (RGCs) offers one potential therapeutic avenue to restore vision in patients suffering from optic neuropathies if the donor neurons survive long term in the recipient eye and develop synaptic connections in the retinal inner plexiform layer (IPL) and subcortical visual centers, which has been difficult to achieve. Previous work in mouse retinal explant cultures has shown that enzymatic digestion of the retinal internal limiting membrane (ILM) promotes migration of transplanted RGCs into the recipient retina. Here, we examined donor RGC survival and engraftment in immunosuppressed mice, rats, and rhesus macaques and in postmortem human retinal explant cultures. Using three separate human PSC lines and three independent methods of ILM disruption (including enzymatic, developmental, and mechanical approaches), we demonstrated that the ILM is a barrier to retinal engraftment of intravitreally delivered human PSC-derived RGCs. Across models, ILM disruption was associated with greater donor RGC survival over 2 to 8 weeks and enabled migration of donor neuronal somata into the endogenous RGC layer, where they elaborated dendrites into the IPL and extended axons that followed the course of the endogenous retinal nerve fiber layer into the optic nerve head, findings that were negligible with intact ILM. Further, ILM disruption enabled donor RGCs to synaptically integrate into IPL circuits, conferring light responsivity in rodents. 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