Effects of tacrolimus on the expression of nuclear factor-κB and glucocorticoid receptor by HaCaT cells in vitro

Objective To investigate the effect of tacrolimus on the expression of nuclear factor-κB (NF-κB) in HaCaT cells stimulated by tumor necrosis factor-α(TNF-α),and on the expression of glucocorticoid receptor (GR)α and β in untreated HaCaT cells in vitro.Methods Cultured Ha CaT cells were treated with TNF-α(10μg/L) only,combination of TNF-α(10μg/L) and various concentrations (10-8mol/L, 10-7mol/L,10-6moL/L) of tacrolimus or tacrolimus of different concentrations only.After additional 12-,24-, 36- or 48-hour cnlture, Westem blot and immunofluorescenee-confocal laser scanning microscopy were used to detect the expressions of NF-κB,GRα and GRβ in HaCaT cells.Those untreated HaCaT cells served as the control.Results The relative protein expression level of NF-κB was increased in HaCaT cells after treatment with TNF-α for 24 and 48 hours zompared with untreated ceils (0.73±0.0316 and 0.8925±0.0171 vs 0.4988±0.03506,both P<0.05);however,the increase in NF-κB expression was inhibited by the combination treatment with tacrolimus,and the relative expression level of NF-κB protein was 0.6825±0.0263.0.6200±0.0163 and 0.5575±0.0299 in HaCaT cells treated with TNF-α plus tacrolimus of 10-8mol/L 10-7mol/L and 10-6mol/L,respectively;the difference was significant etween TNF-α-treated cells and those dealt with the combination of NF-α and tacrolimus of 10-7 or 10-6 mol/L (both P<0.05).No significant difference was observed in the expression of NF-κB by HaCaT cells between different time oints treated with tacrolimus of 10-8,10-7 or 10-6 mol/L.Also,there was no zignificant difference in the expression of GRα or GRβ between untreated HaCaT cells and those treated with tacrolimus of 10-8, 10-7 or 10-6 mol/L at any time point.Conclusions Tacrolimus ould inhibit the expression of NF-κB by TNF-α-stimulated HaCaT cells,but does not affect the expression of GRα or GRβ,in untreated HaCaT cells.