ABSTRACT
The purpose of this study was to investigate the inhibitory effect of the main 9,10-dihydrophenanthrene orchinol isolated from Spiranthes sinensis Radix et Herba on the invasion and migration of human gastric cancer SGC-7901 cells and its preliminary molecular mechanism. SGC-7901 cells were cultured in vitro, after the cells were treated with different final concentrations(5, 10, 20, 40, 80 μmol·L⁻¹) of orchinol for 24, 48 or 72 hours, the effect of orchinol on cell viability was measured by MTT assay. Wound healing and Transwell assays were performed to determine the effects of different final concentrations(5, 10, 20, 40 μmol·L⁻¹) of orchinol for 48 hour on invasion and migration abilities of SGC-7901 cells, respectively. The protein expression levels of β-catenin, Wnt-3α, DvL2, cyclinD1 and GSK-3β were detected by Western blot. The results showed that 5-80 μmol·L⁻¹ orchinol inhibited the viability of SGC-7901 cells in a dose-dependent and time-dependent manner, and the IC₅₈ values of 24, 48 and 72 hours were 77.79, 42.96 and 7.85 μmol·L⁻¹, respectively. Compared with the control group, the ability of invasion and migration of SGC-7901 cells was significantly inhibited after treated with 5, 10 and 20 μmol·L⁻¹ orchinol for 48 hours (<0.05, <0.01), and the dose-effect relationship was observed. The results of Western blot showed that orchinol could significantly down-regulate the protein expression levels of β-catenin, Wnt3a, DvL2 and cyclinD1, and up-regulate the protein expression level of GSK-3β(<0.05, <0.01, <0.001). The above results suggest that orchinol can obviously inhibit the invasion and migration of SGC-7901 cells, which may be related to its inhibition of Wnt3a/β-catenin signaling pathway and the proteins expression of downstream genes.
Subject(s)
Humans , Cell Line, Tumor , Cell Movement , Cell Proliferation , Phenanthrenes , Stomach Neoplasms , Wnt Signaling Pathway , Wnt3A Protein , beta CateninABSTRACT
Objective: To investigate the effects of silymarin on the proliferation, migration and invasion of human gastric cancer cell line SGC 7901 and its role in promoting apoptosis, and to clarify their possible mechanisms. Methods: The human gastric cancer SGC 7901 cells in logarithmic growth phase were selected and divided into control group (without silymarin) and different concentrations (15, 30, and 60 mg · L-1) of silymarin groups. Inverted microscope was used to observe the morphology of cells. MTT method was used to detect the cell cycle and the apoptotic rates of cells in various groups. The inhibitory rates of proliferation of cells in various groups were detected by flow cytametry. Scratch test and Trans well chamber assay were used to detect the cell migration and invasion abilities. Results: After the SGC 7901 cells were treated with silymarin for 24 h, compared with control group, the adherent densities of the cells in 30 and 60 mg · L-1 silymarin groups were smaller, the cell shape was irregular and the volume was smaller, resulting in a large number of cell debris. Compared with control group, the inhibitory rates of cells in different concentrations of silymarin groups were increased significantly (P<0. 05); the percentages of cells in Gi phase and the apoptosis rates of SGC 7901 cells were increased significantly (P<0. 05); the migration distances were decreased (P<0. 05) and the number of transmembrane cells was decreased significantly (P<0. 05). Conclusion: Silymarin can inhibit the proliferation of human gastric cancer SGC 7901 cells by inducing the G1 phase cell arrest and early apoptosis.
ABSTRACT
OBJECTIVE: To investigate the effect and mechanism of juglone on the apoptosis of human gastric cancer SGC-7901 cells. METHODS: The antiproliferative effect of juglone on SGC-7901 cells was tested by the MTT assay. The apoptosis rate and in-tracellular reactive oxygen species(ROS) level were detected by flow cytometry (FCM). The expression of JNK, p-JNK, p38, and p-p38 proteins were examined by Western blot. In order to clarify the role of ROS in the apoptosis induced by juglone on SGC-7901 cells, the combination of the juglone and ROS inhibitor NAC groups were set up in each experiment above. RESULTS Juglone could effectively inhibit the proliferation of SGC-7901 cells (IC50 for 72 h was 24.16 μmol·L-1). When combination juglone with NAC, the IC50 raised to 36.91 μmol·L-1. After 72 h of exposure to 5-20 μmol·L-1 of juglone, the cell apoptosis rate increased gradually with the increase of juglone concentration. After adding NAC, the apoptosis rates declined and the apoptosis rate of 20 μmol·L-1 group decreased from 32.06% to 11.56%. After SGC-7901 cells were treated with juglone for 24 h, the ROS level increased and mitochondrial transmembrane potential decreased which were inhibited by the pretreatment of NAC. After 48 h of exposure to different con-centration of juglone, the expressions of p-p38 and p-JNK proteins were up-regulated which could also be inhibited by the adding of NAC. Meanwhile, there were no significant changes in p38 and JNK protein expression in all groups. CONCLUSION: Juglone can induce apoptosis of human gastric cancer SGC-7901 cells by JNK and P38 pathway mediated by reactive oxygen species.
ABSTRACT
Objective To explore the effects of humulon on kinetic parameters of N-acetyltransferase-1(NAT1) of human gastric cancer SGC-7901.Methods Employing HPLC,using para-aminobenzoic acid(PABA) as substrate,in intact SGC-7901 cells and their cytoplasm,taking the speed of PABA being acetylated to Ac-PABA by NAT1 as the rate of NAT1,using double reciprocal plot,taking the reciprocal of concentration of PABA and reaction rate of NAT1 as coordinates,regression equation was obtainied and the Michaelis constant(Km) and maximum reaction velocity(Vmax) were calculated.Results Study on enzyme kinetics demonstrated,as for intact SGC-7901 cells,Km and Vmax of control group were(3.910?0.087) ?mol/L and(0.306 0?0.006 7) pmol/L(1?106 cells),respectively,Km and Vmax of the humulon group were(3.830?0.123) ?mol/L and(0.275 0?0.005 8) pmol(1?106 cells),respectively.As for the cytoplasm of SGC-7901 cells,Km and Vmax of control group were(760.2?210.2) ?mol/L and(0.191 0?0.043 7) pmol/(mg?min),Km and Vmax of the humulon group were(449.0?72.9) ?mol/L and(0.094 0?0.010 4) pmol/(mg?min).Statistically,as for intact SGC-7901 cells or their cytoplasm,there was no difference of the Km between control group and humulon group,but there was remarkable difference of Vmax between control group and humulon group,P