Long-Term Engraftment of Cryopreserved Human Neurons for In Vivo Disease Modeling in Neurodegenerative Disease.

Abstract

The transplantation of human neurons into the central nervous system (CNS) offers transformative opportunities for modeling neurodegenerative diseases in vivo. This study evaluated the survival, integration, and functional properties of cryopreserved forebrain GABAergic neurons (iGABAs) derived from human induced pluripotent stem cells (iPSCs) across three species used in translational research. iGABAs were stereotactically injected into the striatum of Sprague-Dawley rats, immunodeficient RNU rats, R6/2 Huntington's disease (HD) mice, wild-type controls, and Cynomolgus monkeys. Post-transplantation, long-term assessments revealed robust neuronal survival, extensive neurite outgrowth, and integration with host CNS environments. In immunodeficient rats, iGABAs innervated the neuraxis, extending from the prefrontal cortex to the midbrain, while maintaining mature neuronal phenotypes without uncontrolled proliferation. Similarly, grafts in nonhuman primates showed localized survival and stable phenotype at one month. In the neurodegenerative milieu of HD mice, iGABAs survived up to six months, projecting into the host striatum and white matter, with evidence of mutant huntingtin aggregates localized within the graft, indicating pathological protein transfer. These findings underscore the utility of cryopreserved iGABAs as a reproducible, scalable model for disease-specific CNS investigations and mechanistic studies of proteinopathic propagation. This work establishes a critical platform for studying neurodegenerative diseases and developing therapeutic interventions.