Effects of LPS and TNF-α on proliferation and osteogenic differentiation of human dental pulp stem cells
Objective To investigate the effects of lipopolysaccharide(LPS)and tumor necrosis factor-α(TNF-α)on the proliferation and differentiation of human dental pulp stem cells(hDPSCs).Methods HDPSCs were isolated and cultured in vitro.Cells were incubated with different concentrations of LPS(0,0.1,1,10 μg/mL)and TNF-α(0,1,10,100 ng/mL),respectively.Cell counting kit-8(CCK-8)was applied to detect the proliferation of hDPSCs at 24,48,72 h,respectively.Alizarin red(AR)staining kit and 5-bromo-4-chloro-3-indolyl phosphate p-toluidine salt(BCIP)/nitrotetrazolium blue chloride(NBT)alkaline phosphatase(ALP)chro-mogenic kit were applied to detect the changes of AR staining by the unaided eye,quantification of calcium nodules,ALP staining,and ALP activity at 7,14,21 d after cultured with different concentrations of LPS or TNF-α,respectively.Results ①CCK-8 results showed that 1,10,and 100 ng/mL TNF-α decreased the proliferation of hDPSCs at 24,48,and 72 h(P＜0.05).AR osteogenic induc-tion staining showed no significant difference in AR staining by the naked eye after treated with different concentrations of TNF-α at 7,14 d.At day 21 of incubation,the mineralization was decreased in 10 ng/mL and 100 ng/mL TNF-α groups compared to 0 ng/mL group(P＜0.05).ALP staining and ALP activity kit analysis showed lighter ALP staining and lower ALP activity at 7,14,21 din1 ng/mL,10 ng/mL,and 100 ng/mL TNF-α groups than in 0 ng/mL group(P＜0.05).②CCK-8 results showed that there was no difference in the proliferative activity of hDPSCs at 24 h and 48 h after treated with different concentrations of LPS,while the OD450 values at 72 h in 1,10 μg/mL LPS groups were higher than that in 0 μg/mL group(P＜0.05).ALP osteogenic induction staining showed no significant changes in ALP staining and ALP activity at 7,14,21 d after treatment with different concentrations of LPS.AR osteogenic induction staining showed no significant mineralization at 7 d after treatment with different concentrations of LPS,and the mineralization was lower at 14,21 d in 10 μg/mL LPS group than in 0 μg/mL LPS group(P＜0.05).Conclusion High concentration of LPS can promote the proliferation of hDPSCs,and the effect on osteogenic differentiation of hDPSCs depends on the concentration of LPS and the duration of action.TNF-α inhibit the proliferation and differentiation of hDPSCs.
万方数据
维普
