Molecular Mechanism Of Astragalin Induced Apoptosis,cell Cycle Arrest And Migration Inhibition In Human Gastric Cancer Cells

Gastric cancer is a malignant tumor that occurs in epithelial gland cells of the gastric antrum.The epidemiological characteristics of gastric cancer in the world are mainly manifested in high morbidity and mortality.It is difficult to diagnose early cases and patients have poor prognosis.China is also a central area with a high incidence of gastric cancer morbidity and death.The number of new patients with gastric cancer each year accounts for 47.13%of the world.At present,the clinical treatment of gastric cancer is still mainly based on traditional surgical resection of cancerous parts of the stomach,combined with radiotherapy and targeted drug therapy targeting tyrosine or growth factor.These treatment methods generally have large side effects,poor targeted,high price,and easy to cause secondary trauma to patients.In addition,the clinical application of first-line gastric cancer treatment drugs has strong toxic effects,which greatly limits the clinical use of related drugs.Therefore,the development of new anti-gastric cancer drugs with high efficacy and low toxicity effects is still one of the problems that need to be solved urgently.The main idea for the development of therapeutic drugs for gastric cancer is to inhibit the proliferation,promote the apoptosis and inhibit the metastasis of cancer cells.Therefore,this project aims to explore the effect of astragalin on the proliferation,apoptosis and migration of gastric cancer AGS cells,and research on the intrinsic association of ROS and astragalin.The cell survival rate of 10 kinds of gastric cancer cells and 4 kinds of normal cells at different concentrations and times was measured by CCK-8 colorimetry.Flow cytometry and Western blot were used to detect the cell cycle and related protein expression.Hoechst/PI dual-fluorescence staining was used to observe the morphology of apoptosis.Flow cytometry and Western blot were used to detect the protein changes induced by mitochondrial pathway.The cell scratch experiment was used to observe the migration of gastric cancer cells and calculate the cell migration rate.The protein expression of apoptosis-related signaling pathways(p38,JNK,ERK,NF-κB),cell cycle signaling pathways(Akt,p53,STAT3)and cell migration signaling pathways(TGF-β/Smad)was detected by Western blot,And further determine the nuclear and cytoplasmic contents of nuclear transcription factors NF-κB and STAT3.Stain with ROS-specific fluorescent probes to determine its intracellular content,then use flow cytometry to detect the intrinsic correlation of ROS and astragalin,and explain the relevance between ROS,apoptosis and cell cycle in the gene and protein levels.Astragalin has a good killing effect on 10 kinds of gastric cancer cells,and it has a low toxic effect on normal lung IMR-90,kidney 293T,liver L-02 and gastric GES-1 cells,Astragalin have stronger proliferation inhibition ability and lower toxic effects on gastric cancer cells.Astragalin upregulates the expression of CKI(p21 and p27),which significantly inhibits the expression of Cyclin B1 and CDK1/2,resulting in G2/M phase arrest in gastric cancer AGS cells.Astragalin increases the mitochondrial membrane protein Bad/Bcl-2 ratio,which increases the permeability of the mitochondrial membrane,the content leaks out,and the homeostasis of the mitochondrial membrane is destroyed,thereby making the downstream Caspase-3 and its action bottom PARP is sheared,which leads to apoptosis of gastric cancer AGS cells through the mitochondrial pathway.Astragalin can promote the expression of p-p53 protein and inhibit the expression of p-Akt and p-STAT3 protein.In addition,the inhibited Akt signaling pathway can further inhibit the transcriptional activity of STAT3,causing cell cycle arrest.Astragalin can inhibit the ERK signaling pathway through.phosphorylation,and activate the p38 and JNK signaling pathways,causing the expression of IκB-α(NF-κB binding protein)in the cytoplasm to increase,leading to cytoplasmic and nuclear NF-κB expression decrease.The biological activity is inhibited,causing apoptosis of gastric cancer cells.Astragalin inhibits the migration of gastric cancer cells by promoting the expression of TGF-β1,Smad2/3,N-cadherin and Vinmentin,and promoting the expression of E-cadherin.Astragalin through its redox activity,causes the accumulation of reactive oxygen species in the cell,which regulates MAPK,Akt,p53,STAT3 and NF-κB signaling pathways,thereby participating in the regulation of apoptosis and cell cycle.In conclusion,astragalin induces AGS cells to undergo mitochondrial-dependent apoptosis by up-regulating ROS levels,regulates MAPK and NF-κB signaling pathways,and causes G2/M phase arrest through Akt,p53 and STAT3 signaling pathway.In addition,astragalin inhibits cell epithelial-mesenchymal transition by down-regulating the TGF-β/Smad signaling pathway,thereby inhibiting the migration of gastric cancer cells.