The Small Chemical Compound Repsox Potentiates Oct4-Driven Astrocyte-to-Neural Stem Cell Reprogramming via Notch1/Hes1/Smurf2 Pathway.

Abstract

Central nervous system (CNS) injury is a prevalent and devastating neurological disorder characterized by progressive and irreversible neuronal loss, leading to persistent neurological deficits. Cell-replacement therapy using pluripotent neural stem cells (NSCs) offers considerable promise for treating CNS injury treatment. Our recent studies have demonstrated that mature astrocytes can be directly reprogrammed to a pluripotent state through specific stimuli. However, acquiring sufficient quantities of functional induced NSCs (iNSCs) derived from astrocytes for clinical applications remains challenging due to the low efficiency and instability in previous methodologies. Consequently, it is of critical importance to improve the reprogramming efficiency of astrocytes into iNSCs. In this study, we showed that the reprogramming of astrocytes into iNSCs via a single transcription factor Oct4 is significantly enhanced by continuous treatment with Repsox, a small molecule inhibitor of transforming growth factor-β (TGF-β) signaling. This enhancement was substantiated by increased efficiency in both reprogramming and conversion processes towards genuine NSCs, as demonstrated by the acquisition of distinctive hallmark NSC properties, including distinct morphological features, self-renewal capacity, expression of NSC-specific markers, and multipotency. Furthermore, the resulting iNSCs successfully differentiated into astrocytes, neurons, and oligodendrocytes. Notably, compared to iNSCs generated by Oct4 alone, Oct4/Repsox-induced NSCs exhibited a greater propensity to give rise to more neurons with neuronal functional properties, and relative fewer glial cells. Mechanistically, the activation of Notch1/Hes1/Smurf2 signaling cascades was involved in this enhanced intricate cell reprogramming events. The efficient reprogramming of astrocyte into iNSCs will provide a promising autologous cell-based therapeutic strategy for patients with CNS injury.