Therapeutic approach of ASCs and differentiated lineage cells for the regeneration of nervous system

摘要

Cell-based therapy has been widely applied to promote tissue regeneration and repair especially for degeneration ischemic and inflammatory tissues Adipose-derived stem cells (ASCs) hold the potential for differentiation into multiple lineages under appropriate cytokine and growth factor stimulation and can serve as cell source for cell therapy Our study aimed to investigate the microenvironment for ASCs induction cell-cell interaction and cell-host interaction for promoting regeneration especially nervous system First we evaluated the beneficial effect of mechanical shear stress in the differentiation of endothelial lineage cells (ELCs) from ASCs and the possible intracellular signals to protect hypoxic-ischemic (HI) injury using cell-based therapy in the neonatal rats The successful induction of ASCs into ELCs by combined chemical and mechanical stimulation was demonstrated by increasing endothelial marker and endothelial function The transplanted ELCs can migrate and engraft into the brain tissue especially in vessels where they promoted the angiogenesis The activation of Akt by neuropilin 1 (NRP1) and vascular endothelial growth factor receptor 2 (VEGFR2) were important for ELC migration and following in vivo therapeutic outcomes Second sphere formation of ASCs on chitosan-coated microenvironment promoted differentiation of ASCs into a mixed population of neural lineage-like cells (NLCs) The effect of FGF9 a key regulator of neural cell fate during embryo development was tested by adding FGF9 peptide and discovered the switch of NLCs (FGF9-NLCs) toward Schwann cells (SCs) with expressing of S100β and GFAP Specific silencing FGFR2 diminished the FGF9-induced Akt phosphorylation and inhibited the SCs differentiation The transplanted FGF9-NLCs in nerve conduit participated in myelin sheath formation enhanced the axon regrowth and promoted the functional regeneration after sciatic nerve transection injury This study reveals the importance of FGF9 in SCs fate determination via the FGF9-FGFR2-Akt pathway and demonstrates the therapeutic benefit of FGF9-NLCs Third we further revealed the interaction between ELC and NLC as well as transplanted cell and host by cotransplanting ELCs and NLCs (E+N) into HI injured brain The E+N combination produced significant reduction of brain damage and cell apoptosis and the most comprehensive restoration in neurovascular unit Improvements in cognitive and motor functions were also achieved in the injured rats with E+N therapy Synergistic interactions to facilitate transmigration under in vitro hypoxic microenvironment were discovered with involvement of the NRP1 signal in ELCs and the C-X-C chemokine receptor 4 (CXCR4) and fibroblast growth factor receptor 1 (FGFR1) signals in NLCs Finally we revealed the secretory factors particularly the extracellular vesicle (EV) among different stem/progenitor cells and investigated their therapeutic mechanism The EV isolated from coculture of ELCs and NLCs (E+N) showed best effects to inhibit inflammation and cell apoptosis which is further boosted by in vitro hypoxia The miR-126 was enriched in EV by the synergistic effect of E+N increased the Akt phosphorylation and inhibited the VCAM expression in target cells The EV injection showed comparable protection with E+N cell transplantation to prevent neuronal loss in HI injury model Therefore we showed the characterization of EV profiles in different stem/progenitor cells and illustrated the ELC-NLC interactions and hypoxia to enrich miR-126 for better therapeutic EV which may benefit the future clinical EV therapy Taken together current study demonstrated the roles of microenvironmental cue and EV in ASC differentiation ELC-NLC interaction cell-host interaction and finally resulted in better regeneration and functional outcome

獎項日期	2019
原文	English
監督員	Chia-Ching Wu (Supervisor)