| Objective: This article aims to discuss the characteristics of proteins and RNA carried by exosomes in the circulating blood volume of patients with colon malignant tumors,and micro RNA-181a-5p in plasma exosomes of colon malignant tumors to the epithelial-mesenchymal transition of colon cancer epithelial cell lines The regulation and influence of the process.Methods: Blood samples from 6 patients with stage III-IV ascending colon cancer who were generally in good condition,had no other underlying diseases,had no tumor metastases,and did not undergo chemotherapy or other treatments were collected from the clinic.Using kit method and ultracentrifugation method to extract exosomes in blood samples,verify the extraction efficiency and purity of exosomes by verifying their morphology,vesicle diameter,concentration and surface molecules,and detect micro RNA-181a-5p in exosomes content.Select 4 indicators closely related to the epithelial-mesenchymal transition of colon cancer cell lines,including E-cadherin,vimentin,SMAD2 protein and SMAD3 protein,using QT-PCR and Western blot The experiment detected the difference in protein expression between the exosome and HCT-116 cell line co-culture system,and verified the induction of micro RNA-181a-5p in plasma exosomes of colon cancer patients by regulating the expression of micro RNA-181a-5p,TGF-β1,SMAD7 The molecular mechanism of epithelial-mesenchymal transition in HCT-116 colon cancer cell line.Results: The expression of micro RNA-181a-5p in plasma-derived exosomes of colon cancer patients was significantly higher than that of normal people.Exosomes can be taken up by the colon cancer HCT-116 cell line.In the paired specimens,the colon cancer cell invasion and migration ability in the co-incubation system of exosomes and HCT-116 colon cancer cell line was significantly higher than that in the control group.However,by transfecting micro RNA-181a-5p agonists and inhibitors in the co-culture system,it can be observed that the expression of micro RNA-181a-5p in the system is negatively correlated with the expression of SMAD7 protein in the system,which is related to the tumor epithelial cell invasion and migration Was positively correlated.Western blot results showed that the content of TGF-β1 in the co-culturesystem was significantly higher than that in the control group,while the expression of SMAD2/3 protein was lower than that in the control group.In an in vitro environment,high expression of TGF-β1 can promote the proliferation and invasion of colon cancer cells to a certain extent,and promote the epithelial-mesenchymal transition process of colon cancer cells by activating the SMAD2/3 signaling pathway,including the expression of EMT-related protein Vimentin Increased and decreased E-cadherin expression,the application of TGF-β1 inhibitors can produce an inhibitory effect on the epithelial-mesenchymal transition of colon cancer HCT-116 cell line.The SMAD7 protein can competitively inhibit the TGF-β1 receptor,promote SMAD2/3phosphorylation and other pathways,antagonize the role of TGF-β1,and thereby inhibit the epithelial-mesenchymal transition of tumor cells.Conclusion: Our results indicate that plasma exosomes of colon cancer patients can be taken up by colon cancer HCT-116 cell line,which in turn affects their biological behavior.It also clarified that micro RNA-181a-5p in exosomes down-regulates the expression of SMAD7 by targeting,thereby promoting TGF-β1 to promote the malignant phenotype of colon cancer cells through the TGF-β / SMAD pathway,thus prompting the plasma of patients with colon malignant tumors Exosomes will provide a new direction for the diagnosis and treatment of colon cancer. |
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