The Effect About S100β Gene Modified Olfactory Ensheathing Cells To Inflammatory Environment

Objective To investigate the effects and its related mechanism of S100β protein on olfactory ensheathing cells(OECs)by an established gene modified olfactory ensheathing cells inflammatory model.Methods The olfactory bulb of SD neonatal rats were taken out,and olfactory ensheathing cells(OECs)were cultured and purified.Then OECs were transfected with S100β-LV5(S100β-OECs group)or empty vector(empty-vector-OECs group).After the transfection efficiency was evaluated by flow cytometer,the OECsinflammatory model was set up by treated with recombinant IFN-γ.The proliferation of OECs in S100β-OECs group,empty-vector-OECs group and blank control group were detected by CCK-8 method.After inflammation induced 0h,3h,6h,and 24 h,the apoptosis of OECs in each group was observed by TUNNEL staining.The mRNA expression levels of INOS,IL-1β,TNF-γ,Bax,Bcl-2,and Puma were measured by real time–PCR.Finally,the protein expression levels of S100β,NFκB p65,p38,ERK1/2 and P-JNK in both groups were detected by western blot method.Results 1.The purity rate of primary cultured rat OECs was 82.7%.2.The average transfection efficiency was 74%.3.The growth curve was revealed that there were no significant difference about the proliferation ability in S100β-OECs group,empty-vector-OECs group and OECs group(P > 0.05).4.After treatment with INF-γ 6h,12 h and24h,the m RNA expression level of inflammatory cytokines INOS 、 IL-1β 、 TNF-α and apoptosis related factor Bax were lower in S100β-OECs group than that of in empty-vector-OECs group(P < 0.05),anti apoptotic factor Bcl-2 mRNA expression level was higher in S100β-OECs group than that of in empty-vector-OECs group(P < 0.05).5..After inflammation induced 0h、3h、6h、12h,there was no difference of TUNNEL positive cell number between S100β-OECs group and empty-vector-OEC group(P > 0.05).After 24 h,the apoptotic TUNNEL positive cells were greater in empty-OECs group than that of in S100β-OECs group(P < 0.05).6.The protein expressions level of S100β were increased in both groups after inflammation induced 3h(P < 0.05),but they showed decrease intendency after 6h、12h and 24h(P < 0.05).At the same time point,the protein expression levell of S100β was always higher in S100β-OECs group than that of in empty-vector-OECs group(P<0.05).7.The protein expression level of NFκB p65 became increased in empty-vector-OECs group after inflammation induced 3h,and reached to top at 6h.In S100β-OECs group,expression level of NFκB p65 was decreased at 3h,but it showed increase intendency after 6h.At 3h,6h and 12 h the protein expression levells of NFκB p65 were lower in S100β-OECs group than that of in empty-vector-OECs group(P < 0.05).8.The protein expressions level of p38 were decreased in both groups after inflammation induced 3h.It increased and reached to top at 6h in empty-vector-OECs group.The protein expression levels of p38 were lower in S100β-OECs group than that of in empty-vector-OECs group at 3h、6h、12h(P < 0.05).There were no significance change about the protein expression levels of ERK1/2 and P-JNK in both groups after inflammation induced(P > 0.05).Conclusions 1.S100β could inhibit the OECs release the inflammatory factors.2.S100β could suppress the apoptosis of OECs and increase the survival rate of OECs in inflammatory environment.3.S100β might play roles in NFκB p65/MAPK p38 pathway to modulate the inflammatory response of OECs.