Size-adjustable self-assembled nanoparticles through microfluidic platform promotes neuronal differentiation of mouse embryonic stem cells

Neuronal differentiation from stem cells is one of the most potent therapeutic approaches for recovering neurological function in individuals with neurodegenerative disorders. Herein, an on-demand intracellular retinoic acid released nanoparticles with tunable size and accurately controlled physico-biological properties have been prepared for achieving efficient neuronal differentiation. The amphiphilic chitosan oligosaccharide-cholesterol copolymers were synthesized by varying cholesterol content and self-assembled into spherical micelle in a microfluidic chip with different flow rates. Notably, the results indicated that by increasing the lipophilicity of the chitosan chain as well as mixing rate, the size of micelles was decreased. Retinoic acid (RA) was efficiently encapsulated in the core of micelles. The retinoic acid-containing nanoparticles could escape lysosome, accumulate in the cytoplasm, and release payload with a sustained pattern. The cytotoxicity assay of free retinoic acid and retinoic acid-loaded formulations against P19 embryonic stem cells confirmed the desirable safety of micelles. The result obtained from the uptake study showed that internalization of micelles occurs predominantly via lipid-raft endocytosis in the presence of higher cholesterol content. Moreover, the intracellular RA release upregulated the expression levels of neuronal factors. The micelles described here offer a promising nanomedicine strategy for neuronal differentiation of stem cells.