Characterizations Of HTERT-driven Human Fetal Neural Stem Cell Lines And The Role Of BFGF In Human Fetal Neural Stem Cells

Background: For neural stem cells have the capability of selfrenewing and can generate all types of neural cells, they will be used in cell therapy for nervous system dsease. But now the use of neural stem cells for research and clinical appliantion has been restricted by their limited proliferative potential. Establishment of hTERT-introduced immortal neural stem cell line is a good way to resolve the above problem, but it brings the other problem: whether the hTERT-introduced neural stem cells are cancer cells? bFGF plays the important role in the proliferation and differentiation of neural stem cells, but further research on how this growth factor has double functions is worth doing.Objective: we characterize the hTERT-introduced immortal neural stem cell line HNG and HNG-E, and research on the role of bFGF in neural stem cells.Methodology: HNG and HNG-E cells are characterized by tumorigenic analysis, flow cytometre analysis, fluo3 Ca2+ imaging ,gene chip, cytochemistry immunofluorescence, RT-PCR and Western blot. Recombinant bFGF expression vectors, including pRevTRE/IgK-bFGF and pcDNA3.1(-)/bFGF, are introduced into neural stem cells and HNGE cells. These bFGF-introduced cells are characterized by cytochemistry mmunofluorescenceand Western blot.Results: HNG cells expressed ectogenic telermerase and did not possess tumorigenic growth. They were CD90 positive cells and co-expressed Nestin and GFAP. After induction by RA, these cells expressed MAP2 and O1(oligophrenin-l). HNG-E cells were from early HNG cells, characterizied as tumorigenic cells and lose the phenotype of HNG cells. But these cells can be induced to express dopamine receptor 2 (D2) by RA. We introduced bFGF expression vectors into neural stem cells.. After antibiotic screening, the survival cells can't form any clone and differentiated into neurons. We introduce bFGF expression vectors into HNG-E cells and established a cell line called HNGE\bFGF. HNG-E\bFGF cells expressed GFAP and excogenic bFGF.Conclusion: HNG cells were characterized as immortal neural stem cells which had long life-span and the normal phenotype of neural stem eels. HNG-E cells were characterized as transformed neural stem cells, and can be differentiated into dopaminergic neurons by RA induction. bFGF can induce neural stem cells to differentiate into neurons And bFGF had double function in HNG-E cells. It can promote the proliferation of HNG-E cells by addition of bFGF. On the other hand, after we introduced the ectogenic bFGF into HNG-E cells, these cells changed its phenotype and express the markers of neural progenitor cells.