[Mechanism of TNF-alpha in bone defect of chronic apical periodontitis].

PURPOSE: To investigate the effect of lipopolysaccharides(LPS) extracted from Porphyromonas endodontalis(P.e) on the expression of tumor necrosis factor-alpha(TNF-alpha) mRNA in MC3T3-E1 cells and the role of NF-kappaB signaling on the expression of macrophage colony stimulating factor (M-CSF) induced by TNF-alpha in MC3T3-El cells． METHODS: MC3T3-E1 cells were treated with different concentrations of P.e-LPS(0-50 mg/L) and 10 mg/L P.e-LPS for different time (0-24 h). The expression of TNF-alpha mRNA was detected by reverse transcription polymerase chain reaction(RT-PCR). MC3T3-E1 cells were treated with different concentrations of TNF-alpha(0-10 ng/L) for 6 h. The expression of M-CSF mRNA and protein was detected by RT-PCR and enzyme-linked immunoadsordent assay(ELISA).The expression of M-CSF protein was also detected in 10 ng/L TNF-alpha treated MC3T3-E1 cells after pretreated with BAY 11-7082 for 1 h, a special NF-kappaB inhibitor . Statistical analysis was performed using Multi-way ANOVA and Dunnett t test with SPSS 13.0 software package. RESULTS: The level of TNF-alpha mRNA increased significantly after treatment with different concentrations of P.e-LPS(0-50 mg/L)，which indicated that P.e-LPS induced osteoblasts to express TNF-alpha mRNA in dose dependent manners. Maximal induction of TNF-alpha mRNA expression was seen in the MC3T3-E1 cells treated with 10 mg/L P.e-LPS for 6 h. After 6 h, the expression of TNF-alpha mRNA decreased gradually .The expression of M-CSF mRNA and protein was increased in a does- dependent manner by different concentrations of TNF-alpha treatment（0-10 ng/L）. The expression of M-CSF protein increased from (37+/-2) ng/L(control group) to (301+/-8) ng/L(10 ng/L group).The protein of M-CSF decreased significantly after pretreatment with 10 mumol/L BAY 11-7082 for 1 h, and the expression of M-CSF proteins was reduced from (253+/-14) ng/L to (154+/-2) ng/L .BAY group had no significant difference from the control group. CONCLUSIONS: The expression of TNF-alpha mRNA was increased by P. endodontalis LPS treatment in osteoblast. TNF-alpha may induce the expression of M-CSF in MC3T3-E1 cells through the signaling of NF-kappaB. It suggests that TNF-alpha affect osteoblasts through autocrine way for bone destruction in chronic apical periodontitis induced by P.e-LPS.