| Gastric cancer is one of the most common malignant diseases and the third leading cause of cancer-related deaths worldwide.The 5-year survival rate of patients with gastric cancer is still very low.Recent studies have shown that cancer stem cells(CSCs)play an important role in the development of malignant tumors.Tumor stem cells are progenitor cells of tumor cells.They have self-renewal ability and multi-differentiation potential similar to adult stem cells.They are the starting cells of tumorigenesis,proliferation,recurrence and other processes.The occurrence and maintenance of tumor homeostasis and the invasiveness of tumors and the occurrence of metastasis play a decisive role.In recent years,the use of CSCs theory to explain tumor recurrence and metastasis has been widely recognized by scientists.Therefore,the removal of CSCs is very important in the treatment of tumors.Astrocyte elevated gene-1(AEG-1),also known as Metadherin(MTDH)or LYRIC(Lysine-rich CEACAM1 co-isolated),is the first generation of HIV-infected identified in human astrocytes.In recent years,a large number of studies have supported AEG-1/MTDH,which plays an important role in the development of cancer.It plays a key role in tumor transformation,apoptosis,invasion,angiogenesis and metastasis ? It is associated with the progression of these tumors and poor clinical prognosis.However,the role of AEG-1 in the tumorigenicity of human gastric cancer stem cells remains unclear.Objective:(1)To investigate the expression of AEG-1 in gastric cancer cells.(2)To investigate the effects of AEG-1 on the proliferation,apoptosis and dryness of gastric cancer stem cells,and to clarify the function of AEG-1 gene in gastric cancer stem cells.Methods:(1)Gastric stem cells were sorted by flow cytometry,and the tumor stem cells were sorted by serum-free globule test and tumor-forming experiments in nude mice.(2)The expression levels of mRNA and protein in gastric cancer stem cells(HGC-27,MKN-45)and corresponding non-stem cells were detected by real-time quantitative PCR(qRT-PCR)and Western blot.(3)AEG-1 was treated by transient transfection method,cell proliferation and mechanism of action were detected by MTS,cell cycle,and apoptosis was detected by Annexin V and Caspase 3/7 activity.(4)The tumorsphere formation assay was used to Evaluate the effect of AEG-1silencing on the stemness of gastric stem cells and the tumor formation test in nude mice was used to evaluate the effect of AEG-1 on tumorigenesis of gastric cancer stem cells.Results:(5)The results of serum-free pelleting and tumor formation in nude mice showed that the sorted cells were gastric cancer stem cells.(6)Real-time fluorescence quantification(qRT-PCR)and Western blot showed that the expression level of AEG-1 in gastric cancer stem cells was higher than that in non-stem cell group.(7)The results of MTS test showed that the proliferation of gastric cancer stem cells transfected with AEG-1-siRNA was significantly reduced,while the knockdown of AEG-1 had no significant effect on the growth of non-stem cells of gastric cancer.AEG-1 silencing can lead to the gastric cancer stem cell cycle arrest in the G1 phase.Annexin V and Caspase 3/7 activity detection AEG-1 knockdown can induce gastric stem cell apoptosis.(8)The results of tumorsphere formation assay and tumor formation in nude mice indicated that AEG-1 plays an important role in tumorigenesis of gastric cancer stem cells.Conclusion:The AEG-1 silencing suppressed cell proliferation and induced cell cycle arrest in G1 phase and apoptosis of gastric cancer stem cells,leading to the inhibition of tumorigenesis. |
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