| BackgroundCervical cancer is highly malignant,progresses rapidly and has a poor prognosis.Its occurrence is affected by heredity and life factors.Globally,about 288,000 women die of cervical cancer each year.In China,there are about 135,000 new cases each year,accounting for 1/3 of the world’s total.The current treatment is still mainly radiotherapy,chemotherapy and surgical treatment,but the overall cost is high and the cure rate is low,which brings greater economic pressure and lower quality of life to most patients.Therefore,we urgently need to obtain new drugs for indications or markers for diagnosis and prognosis.At present,traditional Chinese medicine is receiving more and more attention.It can cooperate with radiotherapy and chemotherapy to increase efficacy and reduce toxicity,prevent tumor metastasis and recurrence after surgery,improve clinical symptoms of patients with advanced tumors,and improve the quality of life.There are more successful clinical cases in terms of prolonging survival.Sarcandrae Herba(SH)is a medicine that blindly treats rheumatic SH and has high medicinal value.It has been used for blood-heat rashes,sores,swelling,etc.Modern pharmacological studies have shown that SH has anti-tumor and immune regulation effects.It has been developed into a variety of dosage forms,such as SH injection,which is widely used in clinical adjuvant treatment of cervical cancer,rectal cancer and esophageal cancer.Caoshanhu tablets are used to relieve heat,reduce swelling and pain,and clear the throat.However,the specific anti-cancer active ingredients in SH are unknown,and the targets and signal pathways involved have not been studied in depth.This project is based on these issues to carry out research in order to make new discoveries.ObjectNetwork pharmacology is used to explore the anti-cervical cancer active ingredients of SH,explore the signal pathways and targets involved in the regulation of this ingredient in vitro,and analyze the positive or negative regulatory effects through experimental verification.MethodNetwork pharmacology experiment: Based on the complexity of traditional Chinese medicine components and the characteristics of multi-target treatment,combined with network pharmacology to analyze the in vitro effects of the active ingredients of SH on cervical cancer cells.Search for the chemical components and signal targets related to typhoid in the database to obtain the chemical composition of SH;the molecular structure can be verified,the ID number of the Uni Prot database can be downloaded and the entire document can be converted through Perl code,and the relationship between the drug and the disease can be evaluated Relevance.Search for related terms cervical cancer in the diseaserelated database to identify disease-related genes,and after eliminating duplicate and falsepositive genes,they are matched with potential targets for cervical cancer to find common targets.STRING database was used to predict protein interaction protein interaction network.Put the node 1,node 2 and the comprehensive integration tightness score information in the file into Cytoscape to draw the interactive network diagram,and control the node size according to the style of the statistical tool(Cytoscape→Tool→Network Analyzer→Analysis→Generate Style Based on Statistics)And color settings to reflect the degree of correlation to determine the hub gene.Cell experiment: Through MTT,FACS,Western blotting,Ed U,Transwell and scratch experiment,we can study in vitro the regulation mechanism of the active ingredients of SH on cervical cancer cells and the effect between the involved targets.ResultsThe analysis of network pharmacology results proves that quercetin,as one of the effective components of SH,may play a major anti-tumor effect.The obtained target network action map shows that nodular EGFR is the hub gene(there are higher connectivity in the gene expression network),indicating that EGFR may have important biological significance in the anti-cervical cancer effect induced by quercetin in vitro.Combined with methods such as MTT,FACS,Western blotting,Ed U,Transwell and Scratch experiments,the results show that quercetin can reduce the viability of cervical cancer cells,cause cell cycle arrest and apoptosis in G2/M phase,and inhibit cell growth,migration and invasion.The Tyr-1068 phosphorylation site of EGFR and the corresponding downstream target ERK was selected to participate in the study,and it was found that these two kinases were obviously activated by quercetin.In addition,Afatinib,EGFR inhibitor,and U0126,ERK inhibitor,both further induced cycle arrest and increased apoptosis of cervical cancer cells.Therefore,our results demonstrate that quercetin activates EGFR and ERK to resist the anti-cervical cancer cell activity induced by quercetin for the first time.ConclusionThis study combined with network pharmacological analysis showed that quercetin is one of the active components of SH and may have important research value.It can significantly inhibit the vitality of cervical cancer cells and promote cell cycle arrest and cell cycle arrest in G2/M phase,and inhibit cell migration and invasion.However,both EGFR and ERK inhibitors can enhance the anti-tumor effect induced by quercetin in vitro,indicating that the activation of EGFR and ERK plays a protective role in the anti-cervical cancer activity induced by quercetin,which confirmed the synergistic anti-cervical cancer activity of Afatinib and quercetin indirectly. |
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