RESUMO
This study determined the effects of pcDNA3-beta-nerve growth factor (NGF) gene-modified bone marrow stromal cells (BMSC) on the rat model of Parkinson's disease (PD). The recombinant plasmid pcDNA3-beta-NGF was transfected into BMSC, and NGF expression and its biological activity in vitro were detected. BMSC modified by the NGF gene were then grafted into the corpus striatum of PD rats, and the rotation behavior was evaluated at 1, 2, 4, and 6 weeks post-transplantation. A significant improvement in rotation behavior was observed in PD rats subjected to cell transplantation, especially in PD rats receiving NGF-modified BMSC. The genetically modified BMSC survived and expressed beta-NGF but did not differentiate into tyrosine hydroxylase-positive cells in vivo. The present findings suggested that genetically modified BMSC could be effective for PD treatment, and the mechanisms might involve the neuroprotective effects of beta-NGF.
Assuntos
Transplante de Medula Óssea/métodos , Terapia Genética/métodos , Fator de Crescimento Neural/genética , Transtornos Parkinsonianos/genética , Transtornos Parkinsonianos/terapia , Células Estromais/transplante , Animais , Sequência de Bases , Comportamento Animal , Diferenciação Celular , Corpo Estriado/citologia , Corpo Estriado/fisiologia , Corpo Estriado/cirurgia , Modelos Animais de Doenças , Sobrevivência de Enxerto , Proteínas de Fluorescência Verde/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Dados de Sequência Molecular , Atividade Motora , Plasmídeos , Ratos , Ratos Sprague-Dawley , Proteínas Recombinantes/genética , TransfecçãoRESUMO
OBJECTIVE: To evaluate the effect of different fetal bovine serum (FBS) concentration and first-time exchange of total volume medium on the proliferation of bone marrow stromal cells (BMSCs) from green fluorescence protein (GFP) transgenic mouse in vitro. METHODS: Bone marrow cells isolated from GFP transgenic mice were cultured in DMEM/F12 containing 10%, 20%, 30% FBS respectively; the first exchange of the total volume medium was made at different times (4 h, 6 h, 8 h, 10 h, 12 h, 24 h, 48 h and 72 h) after 3 d primary culture; then the total volume medicum was exchanged every three days. The amplification of BMSCs was determined. The passage 5 BMSCs cultured in DMEM/F12 containing 10% and 20% FBS were examined with the antibodies CD44, CD45 and CD54 at the time of first exchange of the total volume medium. RESULTS: The cultured cells proliferated well in DMEM/F12 containing 10% FBS and 20% FBS. With the extension of the time for first exchange of total volume medium, the density of the adhered cells increased, but the purity of BMSCs decreased. CONCLUSION: The method of making cells adhere to culture plastic in different time for cultivating and purifying BMSCs from GFP transgenic mice is effective, the appropriate concentration of FBS is 10%-20% and the best time for the the first exchange of total volume medium is 8 hour.