ABSTRACT
Objective To estimate the effect of glycyrrhetinic acid on epidermal growth factor (EGF)-induced proliferation of HaCaT cells,and to investigate its possible mechanism.Methods Methyl thiazolyl tetrazolium (MTT) assay was used to evaluate the proliferation of HaCaT cells treated with different concentrations of EGF (0,1,5,10,25,50,100 μg/L) and glycyrrhetinic acid (0,0.1,1.0,10,25,50,100μmol/L) alone,or the combination of 25 μg/L EGF with 25 μ mol/L glycyrrhetinic acid or 10 μ mol/L U0126 (an inhibitor of MEK1/2).Western blot was carried out to measure the protein expression of proliferating cell nuclear antigen (PCNA),Notch-1,ERK 1/2 and phosphorylated ERK 1/2 in HaCaT cells treated with 25 μg/L EGF,10 μmol/L U0126,25μmol/L glycyrrhetinic acid alone or in combination.Data were statistically analyzed by using t test,analysis of variance and correlation analysis with SPSS 17.0 software.Results EGF of 0-100 μg/L promoted the proliferation of HaCaT cells in a dose-dependent manner (r =0.798,P ＜ 0.05),and there was a linear correlation between the effect and concentration within the concentration range 0-50 μg/L (r =0.859,P ＜ 0.05).However,glycyrrhetinic acid of 10-100 μmol/L inhibited the proliferation of HaCaT cells in a dose-dependent manner (r =-0.945,P ＜0.01),and 10 μmol/L glycyrrhetinic acid could suppress the EGF (25 μg/L)-induced proliferation and phosphorylation of ERK1/2 in HaCaT cells.Also,both 25 μmol/L glycyrrhetinic acid and 10 μmol/L U0126 could attenuate the increase in PCNA and Notch-1 expression in HaCaT cells induced by 25 μg/L EGF.Conclusion Glycyrrhetinic acid can inhibit the EGF-induced proliferation of HaCaT cells,likely by suppressing the activation of ERK1/2 signaling pathway.
ABSTRACT
Objective To evaluate the effect of curcumin on the proliferation of and apoptosis in HaCaT cells induced by tumor necrosis factor α (TNF-α).Methods HaCaT cells were cultured with the presence of different concentrations (0,1,5,10,25,50,100 ng/ml) of recombinant TNF-α,curcumin of 20 μmol/L,or the combination of recombinant TNF-α (25 ng/ml) and curcumin (20 μmol/L),for 24 hours followed by the determination of cell proliferation with methyl thiazolyl tetrazolium (MTT) assay.Western blot was conducted to measure the protein expression of proliferating cell nuclear antigen (PCNA) and Notch-1 in HaCaT cells treated with recombinant TNF-α (25 ng/ml) and curcumin (20 μ mol/L) alone or in combination for 24 hours.Flow cytometry using annexin-V/propidium iodine (PI) was performed to assess the early apoptosis in HaCaT cells incubated with recombinant TNF-α of 25 ng/ml and curcumin of 20 μmol/L alone or in combination for 12 hours.Statistical analysis was carried out with one-way analysis of variance.Results Recombinant TNF-α promoted the proliferation of HaCaT cells in a dose-dependent manner,with the maximum proliferation activity observed in HaCaT cells treated with TNF-α of 25 ng/ml,while curcumin of 20 μmol/L effectively inhibited the proliferation of HaCaT cells induced by TNF-α of 25 ng/ml (P ＜ 0.01).TNF-α of 25 ng/ml had no obvious effect on cell apoptosis,while curcumin of 20 μ mol/L markedly induced the apoptosis in HaCaT cells,and there was a synergy between TNF-α of 25 ng/ml and curcumin of 20 μmol/L in the induction of apoptosis in HaCaT cells,with the apoptosis rate being 2.3％,3.4％,11.6％ and 16.8％ respectively in untreated cells,cells treated with TNF-α,curcumin,and the combination of TNF-α and curcumin,respectively.Conclusions Curcumin could enhance the inductive effect of TNF-α on the apoptosis in,but suppress the promotive effect of TNF-α on the proliferation of,HaCaT cells.
ABSTRACT
ObjectiveTo estimate the clinical efficacy and immunomodulatory effect of polysaccharidenucleic acid fraction of bacillus calmette guerin(BCG-PSN) combined with a traditional Chinese medicine in patients with vitiligo.MethodsThis study recruited 99 patients with vitiligo aged from 13 to 65(35,6 ± 5.8) years.The patients were classified into 3 groups to be treated with BCG-PSN and a traditional Chinese medicine (Baidianfeng granules) alone or in combination.BCG-PSN was intramuscularly injected at a dose of 2 ml every other day and baidianfeng granules were given orally thrice a day,for 3 months.Peripheral blood samples were obtained from the patients at the baseline and after the end of treatment and from 30 healthy controls.Flow cytometry and enzyme linked immunosorbent assay(ELISA) were performed to detect T cell subsets and expression level of Fas and Fas ligand(FasL),respectively.Data were analyzed by t test and chi-square test.Results The response rate was significantly higher in patients treated with BCG-PSN combined with Baidianfeng granules than in those with BCG-PSN alone(82.86％ vs.40.63％,P ＜ 0.01 ).Before the treatment,patients showed a lower percentage of CD3+ cells,CD3+CD4+ cells and CD3+CD8+ cells in peripheral blood(all P ＜0.01 ),weaker expression of Fas (P ＜ 0.01 ),but a higher CD4/CD8 ratio (P ＜ 0.01 ) compared with the controls.The treatment with BCG-PSN and Baidianfeng granules alone or in combinatiou all induced an increase in the percentage of CD3+ cells,CD3+CD4+ cells and CD3+CD8+ cells in peripheral blood(P ＜ 0.05) and in the expression of Fas(P ＜ 0.01),but a decrease in CD4/CD8 ratio(P ＜ 0.05).ConclusionsBCG-PSN may induce the normal apoptosis in lymphocytes via reversing the abnormality in the expression of Fas/FasL by peripheral blood lymphocytes,and the effect of BCG-PSN may be enhanced by a traditional Chinese medicine,Baidianfeng granules.