BACKGROUND:Olfactory ensheathing cells can promote oriented differentiation and proliferation of neural stem cells by cell-secreted neural factors. OBJECTIVE:To observe the effect of olfactory ensheathing cells on the differentiation and proliferation of neural stem cells. DESIGN,TIME AND SETTING:Cytology was performed at the Department of Neurology,Tongji Medical College,Huazhong University of Science and Technology,China,from September 2007 to October 2008. MATERIALS:Mouse anti-nestin polyclonal antibody(Chemicon,USA),mouse anti-glial fibrillary acidic protein(GFAP) IgG_1,mouse anti-2’,3’-cyclic nucleotide 3’-phosphodiesterase(CNPase) IgG_1, mouse anti-Tubulin Class-ⅢIgG_1(Neo Markers,USA),Avidin-labeled Cy3(KPL,USA),and goat anti-mouse IgG_1:fluorescein isothiocyanate(FITC)(Serotec,UK) were used in this study. METHODS:Tissues were isolated from the embryonic olfactory bulb and subependymal region of Wistar rats.Serum-free DMEM/F12 culture media was used for co-culture experiments.Neural stem cells were incubated in serum-free or 5%fetal bovine serum-containing DMEM/F12 as controls. MAIN OUTCOME MEASURES:After 7 days of co-culture,neural stem cells and olfactory ensheathing cells underwent immunofluorescent staining for nestin,tubulin,glial fibrillary acidic protein,and CNPase. RESULTS:Olfactory ensheathing cells promoted proliferation and differentiation of neural stem cells into neuron-like cells,astrocytes and oligodendrocytes.The proportion of neuron-like cells was 78.2%,but the proportion of neurons in 5%fetal bovine serum DMEM/F12 was 48.3%.In the serum-free DMEM/F12,neural stem cells contracted,unevenly adhered to the glassware wall,or underwent apoptosis at 7 days. CONCLUSION:Olfactory ensheathing cells promote differentiation of neural stem cells mainly into neuron-like cells,and accelerate proliferation of neural stem cells.The outcome is better compared with serum-free medium or medium containing 5%fetal bovine serum. BACKGROUND: Olfactory ensheathing cells can promote oriented differentiation and proliferation of neural stem cells by cell-secreted neural factors. OBJECTIVE: To observe the effect of olfactory ensheathing cells on the differentiation and proliferation of neural stem cells. DESIGN, TIME AND SETTING: Cytology was performed at the Department of Neurology, Tongji Medical College, Huazhong University of Science and Technology, China, from September 2007 to October 2008. MATERIALS: Mouse anti-nestin polyclonal antibody (Chemicon, USA), mouse anti-glial fibrillary acidic protein ) IgG1, mouse anti-2 ’, 3’-cyclic nucleotide 3’-phosphodiesterase (CNPase) IgG_1, mouse anti-Tubulin Class- IIIIgG_1 (Neo Markers, USA), Avidin-labeled Cy3 (KPL, USA) METHODS: Tissues were isolated from the embryonic olfactory bulb and subependymal region of Wistar rats. Serum-free DMEM / F12 culture media was used for co- culture experiments. Neural stem cells were incubated in serum-free or 5% fetal bovine serum-containing DMEM / F12 as controls. MAIN OUTCOME MEASURES: After 7 days of co-culture, neural stem cells and olfactory ensheathing cells underwent immunofluorescent staining for nestin, tubulin, glial fibrillary acidic protein, and CNPase. RESULTS: Olfactory ensheathing cells promoted proliferation and differentiation of neural stem cells into neuron-like cells, astrocytes and oligodendrocytes. proportion of neuron-like cells was 78.2%, but the proportion of neurons in 5% fetal bovine serum DMEM / F12 was 48.3% .In the serum-free DMEM / F12, neural stem cells contracted, unevenly to the glassware wall, or underwent apoptosis at 7 days. CONCLUSION: Olfactory ensheathing cells promote differentiation of neural stem cells mainly into neuron-like cells, and accelerate proliferation of neural stem cells. The outcome is better than with serum-free medium or medium containing 5% fetal bovine serum.