The Study Of Signal Regulation During Human Neural Differentiation In Vitro And The Destiny Of Transplanted Cells Under Different Environments

Differentiation of neural progenitor cells(NPCs)from human pluripotent stem cells(hPSCs)provides an unlimited cell source for replacement therapy for neural disease and drug screen platform.Dual SMAD inhibition(SB431542/Dosomophrin,S/D)has been proven efficient and stable for hPSCs neural differentiation,however,with limited proliferation ability and unclear differentiation mechanism.In this report,we showed that in addition to dual SMAD inhibition,blocking Glycogen synthase kinase3(GSK3)signaling by CHIR99021(S/D/C)generated hindbrain biased NPCs(S/D/C-NPC).S/D/C-NPC possess metencephalic region identity,with significantly expandable property and differentiation bias to posterior neurons.Through human neural regionally induction system,we found that:(1)OTX2 and GBX2 played the key roles during human neural regional pattern at very early stage of induction.The high expression level of OTX2 in human pluripotent stem cells is one of determining factors for the phenomenon that the default neural differentiation goes to the rostral lineage.(2)GSK3 inhibition activated WNT/beta-catenin pathway which further activated GBX2.And then GBX2 antagonized OTX2,resulting in metencephalic formation.(3)OTX2 promoted PAX6 expression during rostral differentiation,while in hindbrain formation,it was the expression of SOX1 that elevated rather than PAX6.Our study based on the neural induction "default model" suggested that,artificially manipulating relevant signaling pathways could produce different NPCs.These manipulating might not exist in normal brain development,but could bring broad practical usage in clinical therapy.We transplanted human induced neural stem cells into Lateral ventricles of wild type SD rats,fetal and adult.The grafts could be survived and differentiation normally.The cells transplanted into fetal migrated from white matter to olfactory bulb along RMS.While the grafts in adult prefer to integrate into hippocampus.