Inducing Human Umbilical Cord Blood Cells Differentiate Into Neural Cells In Vitro

Objectives: Human umbilical cord blood cells (HUCBC) is a rich source of hematopoietic stem/progenitor cells, mesenchymal stem cells and endothelial progenitor cells which are capable of expansion, self-renewal and differentiate into multiple cell lineages. Its capable of colony formation, the velocity of enter cell cycle and autocrine production level are higher than BM and HPC .Meanwhile, umbilical cord blood are widely available, low antigen expression and long time survive. It has attracted widely attention being a source of stem/ progenitor cells.Neural cells in central nervous system have been thought to be terminal differentiate cell. They can't regenerate once been damaged. However some research indicate that embryonic stem cell and neural stem cell can differentiate to neuron and glial cells. Mesenchymal stem cells coming from marrow can differentiate to neuron and glial cells. When they transplant into the animal with brain injury, cells were found surviving and migrating. The animal's behavioral deficits can significantly be reduced. Some research indicate that HUCBC can be induced into neuron and glial cells with β-mercaptoethanol and poly-l-lysine or RA and NGF. However,if there are better method inducing HUCBC differentiating into neural cells ,and how its mechanism process aren't clearly. In order to get more neural cells to implore in clinic,we should do more further research. Our experiment first use brain derived neurotrophic factor(BDNF) and solvable receptor activator of NF-KappaB ligand(sRANKL) induce HUCBC differentiating to neuron and glial cells in vitro. Using immunocytochemical staining identifie the GFAP and NeuN marker expressing cells and compare the different method. The results may offer reference for the applying of HUCBC in the field of neuroscience. Material and Methods: Term pregnancy and normal labored fresh umbilical cord blood was used. First we use 6.0% dextran fixed into umbilical cord blood to sediment the RBCs.The mononuclear cells are separate by standard Ficoll-Hypaque techniques with Ficoll (density 1.077) and cultured in vitro by randomly divided into five groups:(1) the control group, cultured by DMEM medium with the addition of 10% fetal bovine, 1%L-glutamine, penicilline100u/ml, streptomycin100u/ml;(2)the neural differentiation medium group, cultured by neural differentiation medium only; (3)BDNF group, cultured by differentiation medium with BDNF; (4)sRANKL group, cultured by differentiation medium with sRANKL; (5)BDNF+sRANKL group, cultured by differentiation medium with BDNF and sRANKL.Cultured cells were observed by invert phase contrast microscope. When cultured for ten days, cultured cells' expression of glial fibrillary acidic protein (GFAP) and neuron-specific nuclear protein (NeuN) was detected by immunocytochemical stain method to identify the cells type. Count the average cells in the four hole in every group. All data are presented as mean values±standard error of the mean. The t test were employed for statistical analysis and a probability value less than 0.05 was considered significant. Res...