The Anti-pancreatic Cancer Effect Of Cimicifugae Rhizoma In Vitro Based On Network Pharmacology

Objective:Pancreatic cancer is one of the most common malignant tumors of the digestive tract,it also with poor prognosis.The 5-year survival rate of patients is now less than 10 %.The purpose of this study is to find natural drugs with high efficiency and low toxicity from traditional Chinese medicine to explore potential role and mechanism of active medicine Cimicifuga Rhizoma in the treatment of pancreatic cancer by comprehensive application of network pharmacology,so as to provide new ideas for the intervention of traditional Chinese medicine in pancreatic cancer.Methods:Determine the target active Chinese medicine : CCK8 method was used to investigate the inhibitory activity of commonly used Chinese medicine(100 μg / ml)on Aspc-1 and Panc-1 cells.The IC50 of active Chinese medicine Cimicifuga Rhizoma inhibiting Aspc-1 and Panc-1 was determined.In vitro evaluation of the anti-pancreatic cancer activity of Cimicifuga : The effect of Cimicifuga on the migration of Aspc-1 and Panc-1 cells was investigated by cell scratch assay.The effect of Cimicifuga on the proliferation of Aspc-1 and Panc-1 cells was investigated by colony formation assay.The expression of apoptosis-related proteins(Bcl-2,BAX,casepase-3,casepase-9)in Aspc-1 and Panc-1 cells was detected by Western Blot to further verify the effect of Cimicifuga on apoptosis of pancreatic cancer cells.Using network pharmacology to explore the potential active ingredients and mechanism of anti-tumor effect of Cimicifuga : Search the existing data in TCMSP,Dis Genet,Gene Cards,OMIM and other databases,and screen according to oral bioavailability,drug-likeness and other indicators to obtain active ingredients and related targets,pancreatic cancer disease-related targets;the results obtained from the above steps were imported by Cytoscape software to construct a ’ Cimicifuga-pancreatic cancer-target’ network.The PPI network was constructed by importing the target information in the ’Cimicifuga-pancreatic cancer-target ’ dataset into the String database,and the ’ proteinprotein interaction ’ contained in this dataset was integrated.The obtained core targets of Cimicifuga-pancreatic cancer were imported into the gene list,and the cell components and KEGG pathway analysis were performed to obtain the biological function annotation of the core targets.Through Western Blot verification of the factors in the regulatory network diagram,the mechanism of anti-pancreatic cancer activity of Cimicifuga was determined.Result:In the screening of a variety of traditional Chinese medicine,Cimicifuga can reduce the survival rate of Aspc-1 and Panc-1 cells at the same time,and the survival rate of both cells is less than 30 % at 100 μg / ml;moreover,the inhibition of Cimicifuga Rhizoma on the survival rate of pancreatic cancer cells was concentration-dependent and timedependent,and the IC50 after 72 h treatment was 10.78μg / ml(Aspc-1)and 48.76μg / ml(Panc-1),respectively.The results of cell scratch migration assay showed that compared with the control group,the cell fusion on both sides of the scratch was inhibited in the experimental group with cohosh,and the inhibition was more obvious with the increase of cohosh concentration.The results of colony formation assay showed that compared with the control group,the number of crystal violet-stained cells in the experimental group was significantly reduced,and the number of stained cells decreased with the increase of drug concentration.Western Blot results showed that the expression of apoptosis-related proteins in pancreatic cancer cells treated with Cimicifuga increased significantly with time,while the expression of anti-apoptosis-related proteins decreased significantly.These results suggest that Cimicifuga can inhibit the proliferation and migration of pancreatic cancer cells and induce apoptosis of pancreatic cancer cells.The network pharmacology analysis showed that the targets related to Cimicifuga and pancreatic cancer were sorted according to oral bioavailability and drug-likeness,and 21 intersection targets including ’ PPARγ ’ and GSK3β ’ were obtained.These 21 targets were correlated with the active components of Cimicifuga and pancreatic cancer.The expression of Cycling D1,Cycling E1,COX2,NF-κB and other related proteins was detected by Western Blot.It was found that these proteins related to PPARγ and GSK3β were affected by Cimicifuga,which verified the anti-tumor activity of Cimicifuga.Conclusion:A variety of traditional Chinese medicines have the activity of inhibiting pancreatic cancer cells in vitro,among which Cimicifuga has strong anti-tumor activity.Cimicifugae Rhizoma inhibited the proliferation and migration of Aspc-1 and Panc-1cells and induced apoptosis of two pancreatic cancer cells.Cimicifuga may exert anti-pancreatic cancer activity in vitro through PPARγ and GSK3β signaling pathways.