Further Experimental Study Of Human Umbilical Cord Blood Mesenchymal Stem Cells Differentiating Into Schwann-like Cells In Vitro

Background:The clinical treatment of patients with peripheral nerve injury isunresolved challenges of the world in the field of surgery. Autogenous nervetransplantation is the standard method for the treatment of peripheral nerve defect it,but can cause nerve damage. The repair of peripheral nerve defect by tissueengineered artificial neural is the clinical study on the treatment of hotspots. Seedcells of artificial nerve are Schwann cells (SCs), but source issue has not beenresolved. Human umbilical cord blood mesenchymal stem cells (HUCBMSCs)as analternative source for human bone marrow derived mesenchymal stem cells (MSCs)can differentiate into Schwann-like cells under the induced culture conditions in vitro,but also require further study.Objective: This study aims to establish isolation, culture, purification andidentification of HUCBMSCs system to explore in vitro inducing differentiation ofHUCBMSCs as Schwann-like cells, and induction programmes; further compoundinduced successful Schwann-like cells with acellular nerve basal membrane in vitroco-cultured and traced.Methods: Human umbilical cord blood specimens,6%Hetastarch(HES) settlementred blood cells were taken as a sample, then using human lymphocytes separationliquid Ficoll (density for1.077g/mL) separation cord blood mononuclear cells bycultured in the Mesencult full medium, The expression of phenotype onHUCBMSCs was detected by flow cytometry (FCM). Potential of multi-directionaldifferentiation was identified by osteogenic and adipogenic differentiation; taking the3rd generation cell for directed induced differentiation, Identification of glial cellmarkers S100b, GFAP, P75used Immunocytochemistry, and RT-PCR, and WesternBlotting method for inducing cells; Repeated freeze-thaw oscillation washing preparation of acellular basal lamina tubes of sciatic nerve, after inducedSchwann-like cells were injected into acellular basal lamina tubes bymicro-injection,then placed in six wells plate in vitro and removed the specimens byHE staining after0,1,2w.Results: Isolation training out of cells low express or do not express CD34, highexpress CD44, and CD73. Immunocytochemistry find that almost all cells areexpressed glial cell markers S100b (98±1.63%), and GFAP (95.33±2.05%) and P75(90.67±1.7%), which more cells are presented bipolar, spindle-shaped classicmorphology of Schwann cells after induced4d, while differentiated cells express glialcell markers S100b, GFAP and P75by RT-PCR and Western Blotting in proof ofgene and protein, and undifferentiated cells do not express; HE staining detectionresults show that cells can survive and migrate trends in acellular nerve basal laminatubes.Conclusions:1.The successful isolation of MSCs are from human umbilical cord blood invitro,while HUCBMSCs not only have strong proliferation and self-renewalcapacity,but also have multilineage differentiation potential.2. HUCBMSCs can be differentiated into Schwann-like cells in vitro.3. Schwann-like cells are expected as the new source of seed cells and provided abasis for artificial nerve graft next step since they were survival and migration inthe acellular nerve basal lamina tubes.