| Objective: To explore the proliferation,migration and invasion of breast cancer MDAMB-453 cells under the intervention of Yanghe Decoction drug-containing serum,as well as the mechanism and influence of Akt/CREB/MMP13 pathway in the treatment of breast cancer,further explain the target and rule of Yanghe Decoction in the treatment of breast cancer,and provide theoretical and experimental basis for the clinical application of Yanghe Decoction in the treatment of breast cancer.Methods: Decoct the Chinese prepared medicinal herb of Yanghe Decoction into a medicinal liquid with a crude drug amount of 1.4 g/m L.Thirty-two female SD rats were randomly divided into a blank control group(10 m L/kg body weight saline),a high dose group of Yanghe Tang(24 g crude drug/kg/d),a medium dose group(12 g crude drug/kg/d),and a low dose group(6 g crude drug/kg/d).Blood was taken through the abdominal aorta,serum was isolated,inactivated,subpackaged,and frozen for later use.Resuscitated MDA-MB-453 cells were carefully cultured using DMEM high glucose medium containing 10% fetal bovine serum.After blowing,digestion,centrifugation,and resuspension,MDA-MB-453 cells were divided into blank control groups and low,medium,and high dose drug groups,and corresponding rat serum was used for intervention treatment.The effects of different concentrations of serum containing drugs on the proliferation of MDA-MB-453 cells were detected through CCK-8 and plate cloning experiments.The effect of different concentrations of drug containing serum on the lateral migration ability of MDA MB-453 cells was detected through scratch healing experiments.The effects of different concentrations of serum containing drugs on the vertical migration and invasion ability of MDA-MB-453 cells were detected by Transwell experiment.The effects of different concentrations of drug-containing serum on the expression of Akt,CREB,and MMP13 mRNA in MDA-MB-453 cells were detected using qRT-PCR.Western blot was used to detect the effects of different concentrations of serum containing drugs on the expression of Akt,p-Akt,CREB,p-CREB,and MMP13 proteins in MDA-MB-453 cells.Results: The results of the CCK-8 experiment showed that compared to the blank control group,the 10% concentration of Yanghe Decoction drug-containing serum can inhibit the proliferation of MDA-MB-453 cells,which is positively correlated with dose and time.The inhibition rates of the high-dose group at 24 hours,48 hours,and 72 hours reached 25.70%,41.65%,and 57.41%,respectively(P<0.01).The results of the plate cloning experiment showed that compared with the blank group,the mid dose and high dose groups showed a decrease in the ability of cells to clone into spheres after intervention with Yanghe Decoction drugcontaining serum(P<0.01).The results of the scratch healing experiment showed that compared with the blank group,the serum containing Yanghe Decoction in each group could inhibit the lateral migration of cells,The low-dose group had an inhibitory effect at 48 hours(P<0.01),the medium dose group had an inhibitory effect at 48 hours and 72 hours(P<0.01),and the highdose group had a significant inhibitory effect at 24 hours,48 hours,and 72 hours(P<0.01).The Transwell experiment showed that compared to the blank control group,the medium and highdose Yanghe Decoction drug-containing serum could inhibit the vertical migration ability of MDA-MB-453 cells(P<0.01)and weaken their invasion ability(P<0.01).The qRT PCR experiment showed that compared to the blank control group,the medium and high-dose Yanghe Decoction drug-containing serum could reduce the expression levels of Akt,CREB,and MMP13 mRNA in MDA-MB-453 cells(P<0.05,P<0.01),and showed a close relationship with the dose.Western blot experiments showed that compared to the blank control group,the expression levels of p-Akt,CREB,p-CREB,and MMP13 proteins in MDA-MB-453 cells after intervention with drug-containing serum decreased(P<0.05,P<0.01),and showed a certain positive correlation with dose.Conclusion: Yanghe Decoction drug-containing serum can inhibit the proliferation and migration ability of MDA-MB-453 cells,weaken the invasiveness of MDA-MB-453 cells,and reduce the amount of associated p-Akt,CREB,p-CREB,and MMP13 protein expression.This suggests that the effects of Yanghe Decoction drug-containing serum on the proliferation,migration and invasion abilities of MDA-MB-453 cells may involve the Akt/CREB/MMP13 signaling pathway. |
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