The Study Of Induction Of Apoptosis And Inhibition Of MMPs Expression In The Gastric Carcinoma Cell Lines By Trametes Extract

Objective:This study focused on the effect of Trametes extract on inhibiting the proliferation, apoptosis, and expression levels of MMP-2and MMP-9in gastric carcinoma cell lines MKN28, SGC7901. It also provided theoretical and practical basis for clinical treatment of gastric cancer.Methods:Human gastric carcinoma cell line, MKN28and SGC7901, were well and moderate differentiation, respectively. The analysis of proliferation inhibition of gastric carcinoma cell lines induced by Trametes extract with the concentration of0.5mg/ml, lmg/ml,3mg/ml,5mg/ml and10mg/ml was detected through microscopy and MTT method. The analysis of apoptosis of gastric carcinoma cell lines incubated with Trametes extract for48h at the concentration of0mg/ml,5mg/ml,10mg/ml was detected using immunofluoresent double staining (AO/EB) and flow cytometry. Expression level of MMP-2and MMP-9mRNA of MKN28cell line was detected by RT-PCR after incubation with the Trametes extract for48h at the concentration of Omg/ml,5mg/ml,1Omg/ml.Results:After MKN28and SGC7901cell lines were incubated with Trametes extract at the concentration of0.5mg/ml,1mg/ml,3mg/ml,5mg/ml and10mg/ml for24h,48h,72h, the cultured cells looked shrinkage, membrane rupture, nuclear condensation and fragmentation observed under microscope in all groups. In MKN28cell line, compared with control group, the inhibition rate of cell line at the concentration of0.5mg/ml of Trametes extract was5.73%,7.50%,20.50%, respectively. At the concentration of lmg/ml. the inhibition rate was11.70%,13.83%,15.59%, respectively. At the concentration of3mg/ml. the inhibition rate was16.06%,23.79%,24.73%, respectively. At the concentration of5mg/ml the inhibition rate was24.91%,32.59%,34.67%, respectively. At the concentration of10mg/ml, the inhibition rate was30.98%,45.01%,46.15%. respectively. In SGC7901cell line, the inhibition rate of cell line at the concentration of0.5mg/ml of Trametes extract was2.43%,2.61%,5.60%. respectively; At the concentration of1mg/ml. the inhibition rate was5.63%,5.92%.13.70%. respectively; At the concentration of3mg/ml. the inhibition rate was of12.48%,19.83%,22.02%, respectively: At the concentration of5mg/ml, the inhibition rate was23.10%,30.37%,33.93%, respectively; At the concentration of10mg/ml, the inhibition rate was29.68%,35.28%,37.03%, respectively.After MKN28and SGC7901cells incubated with the Trametes extract at the concentration of Omg/ml,5mg/ml,10mg/ml, respectively, the degree of apoptosis depended on drug concentration using the immunofluorescence AO/EB double staining. Apoptosis rate of MKN28cell line was41.76%,95.62%,94.13%, respectively using flow cytometry; Apoptosis rate of the SGC7901cell line was33.77%,83.74%,92.77%, respectively; MKN28cell line incubated with the Trametes extract for48h at the concentration of Omg/ml,5mg/ml,10mg/ml, respectively, the analysis of gray degree indicated that ratio of MMP-2/β-actin mRNA expression was0.321,0.260,0.205, respectively, and that of MMP-9/β-actin mRNA expression was0.244,0.192,0.135using RT-PCR. With increase of drug concentration, the expression level of MMP-2and MMP-9mRNA decreased.Conclusion:Growth inhibition and induction of apoptosis in human gastric carcinoma cell lines, MKN28and SGC7901was affected by Trametes extract with dose-dependent manner; With the extention of drug incubation time, the inhibition rate was time-dependent, increasing level of inhibition rate at48h is higher than that of inhibition rate at72h. Under the same condition, the ratio of apoptosis of well differentiation MKN28cell line was higher than that of moderate differentiation SGC7901cell line. Expression level of MMP-2. MMP-9mRNA in MKN28cell line was both inhibited by Trametes extract and was relative to its concentration. The results of anti-apoptosis and inhibition of MMP-2, MMP-9mRNA expression in gastric carcinoma cell lines by Trametes extract provided evidence for application in the clinical treatment.