ABSTRACT
Aim To investigate the effect of acacetin on cell proliferation and the influence of acacetin on estrogen receptor expression in vitro.Methods The proliferation rates and the cell cycle changes of acace-tin-treated T47D cells were measured by sulforhodam-ine B(SRB)assay and flow cytometry,respectively. Moreover,the mRNA expressions of estrogen receptor-alpha(ERα),estrogen receptor-beta(ERβ)and pro-liferating antigen(Ki67)were determined by quantita-tive real time PCR (qPCR).Western blot was em-ployed to detect the ERαand ERβprotein expression. Results Acacetin significantly promoted the prolifera-tion and increased the amount of cells arrested in S and G2 /M phase under the concentration of 0.001 ~1 0μmol·L -1 .Ki67 mRNA level and the ERαprotein level in T47D cells were remarkably upregulated after acacetin treatment.To clarify which estrogen receptors played a role in acacetin induced the proliferation of T47D cells,the combination treatment of acacetin and ERαinhibitor (MPP)/ERβ inhibitor (PHTPP) was employed.We found that MPP could reverse the cell proliferation,the cell arrested in S and G2 /M phase and the increased Ki67 mRNA level induced by acace-tin.PHTPP also alleviated the T47D cell proliferation induced by acacetin,whereas no significant changes were found in cell cycle and Ki67 mRNA level.Con-clusion Acacetin stimulates the cell proliferation of T47D cells in the concentration from 0.001 μmol · L -1 to 1 0 μmol·L -1 ,which is mainly mediated by ERα.
ABSTRACT
Aim To investigate the mechanism of the melanoma B16 F10 cells proliferation induced by Lico-chalcone A in vitro. Methods The proliferation of B16 F10 cells induced by Licochalcone A was deter-mined by SRB method. The morphological changes were observed using Giemsa staining under the phase contrast microscope equipped with a digital camera. The melanin level was assessed by colorimetric meth-od. The apoptotic rate was determined by Annexin V-FITC/PI assay. Cell cycle distribution was determined by flow cytometry. The mRNA expression levels of B cell lymphoma/lewkmia-2 ( Bcl-2 ) , Bcl-2 associated X protein ( Bax) , the cell cycle protein CyclinE2 and cyclin-dependent kinase-2 ( CDK2 ) CDK2 were detec-ted using Q-PCR analysis. Results The proliferation of B16 F10 cells treated with Licochalcone A was effec-tively inhibited in a concentration and time-dependent manner. A clear morphological change was observed with the increasing concentration of Licochalcone A in B16F10 cells, the dendrite-like projections changed to the narrowing ball shape, which was associated with the increasing melanin level. The low concentration of Licochalcone A could induce B16F10 differentiation, and the high concentration of Licochalcone A could in-duce B16F10 apoptosis, which was accompanied with the increasing G1 phase in cell cycle. The mRNA ex-pression levels of Bcl-2 /Bax, CyclinE2 and CDK2 were markedly reduced. Conclusion Licochalcone A can effectively inhibit the proliferation of B16 F10 cells, induced cell cycle arrest at G1 phase, and fur-ther induced differentiation and apoptosis.
ABSTRACT
Aim To evaluate the mechanism of apopto-sis induced by the isoliquiritigenin in A375 human ma-lignant melanoma cells. Methods Sulforhodamine B ( SRB) method was used to determine the A375 cell viability;acridine orange/ethidium bromide ( AO/EB) and Hoechst 33258 staining were used to observe the morphological changes of apoptotic cells; flow cytome-try was used to detect A375 cell apoptotic rate;DCFH-DA was applied to determine the changes of total intra-cellular ROS in A375 cells;JC-1 method was used to measure the changes of mitochondrial membrane poten-tial;the kits methods were used to determine the con-tent of ATP, lactic acid and glucose in A375 cell which was treated with different concentrations of isoliquiritigenin. Results Isoliquiritigenin could in-hibit A375 cell proliferation in a concentration-depend-ent manner; A375 cells showed obvious apoptosis charateristics after treatment by isoliquiritigenin, and the apoptosis rate increased with increasing concentra-tion of isoliquiritigenin. The level of total intracellular ROS in A375 cells increased obviously after dealing with different concentrations of isoliquiritigenin;in ad-dition, the mitochondrial membrane potential, the lev-els of intracellular ATP,lactic acid and the level of glu-cose uptake all declined. Conclusions These find-ings demonstrate that isoliquiritigenin can induce apop-tosis of A375 cells. The mechanism may be related to elevation of ROS level and reduction of aerobic glycoly-sis level.