Protective effect of Toll-like receptor 4 inhibitor TAK-242 on β-amyloid peptide25-35 inducing PC12 cytotoxicity / 中华神经医学杂志

ABSTRACT

Objective To study the role ofTAK-242,a Toll-like receptor 4 (TLR4) specific inhibitor,in β-amyloid peptide (Aβ)25-35 inducing PC12 cytotoxicity and its potential mechanism.Methods PC12 cells were cultured with different concentrations of Aβ25-35 (0,10,20 and 30 μmol/L) for 24 h,and then,the cell survival rate was detected by CCK-8 kit to choose the specific concentration of Aβ25-35 to establish cell AD models.The survival rate of Aβ25-35 inducing PC12 cells was further detected one h after TAK-242 intervention.The PC12 cells were divided into four groupscontrol group,Aβ treatment group,Aβ+TAK-242 pretreatment group and TAK-242 group.The apoptosis of cells was observed with Hoechst 33258 kit.The secretions of interleukin (IL)-1β and tumor necrosis factor-α (TNF-α) were detected with ELISA.The protein expression levels of TLR4,myeloid differentiation factor 88 (MyD88),IκB kinase complexus α/β (IKKα/β) and nuclear factor (NF)-κB were detected by Western blotting.Results The cell survival rate decreased gradually with the increase of Aβ25-35 concentrations after PC12 cells cultured with Aβ25-35 for 1 h.Twentyμmol/L Aβ25-35 was used to establish the AD models,with which the cell survival rate was closely half of the control group.As compared with Aβ treatment group,Aβ+TAK-242 pretreatment group had significantly increased cell survival rate and significantly decreased apoptosis (P＜0.05).The secretions of IL-1β and TNF-α in Aβ treatment group were significantly increased than those in the control group (P＜0.05),and Aβ+TAK-242 pretreatment group had significantly decreased secretions of IL-1β and TNF-α (P＜0.05).As compared with those in the control group,the protein expressions of TLR4,MyD88,IKKα/β and NF-κB in the Aβ treatment group were significantly increased (P＜0.05);as compared with Aβ treatment group,the protein expressions of TLR4,MyD88,IKKα/β and NF-κB in the Aβ+TAK-242 pretreatment group were degraded obviously,with significant differences (P＜0.05).Conclusions Aβ25-35 could reduce the cell survival rate and apoptosis in PC12 cells by up-regulating the expressions of TLR4/MyD88 signal pathway related proteins and increasing the secretions of IL-1β and TNF-α,and the phenomenon is concentration-dependent.TAK-242 could resist Aβ25-35-induced PC12 cytotoxicity through down-regulating the TLR4/MyD88 signal pathway related proteins levels and decreasing the secretions of TNF-α and IL-1β.