The Role And Mechanism Of Evodiamine On MiR-489 Expression In Pancreatic Cancer Cells

Objective: Although it has been reported that evodiamine(Evo)can inhibit the proliferation of pancreatic cancer cells,induce autophagy,decrease the activity of Akt and ERK signaling pathways,and enhance the sensitivity of cancer cells to the traditional drug kitasabine,the research on its mechanism is not comprehensive,such as whether it is involved in the regulation of pancreatic cancer glycolysis pathway,whether it induces the expression of functional miRNAs,and whether it can inhibit the proliferation of pancreatic cancer cells,In view of the above aspects,this project intends to further study the mechanism of evodiamine in pancreatic cancer cells.Methods: Firstly,protein samples of pancreatic cancer cells treated with Evo and control group were prepared.Western blot was used to detect the expression levels of glycolysis and invasion related proteins in pancreatic cancer cells.Secondly,by reading the literature and combining with the prediction software targetscan7.2,we screened out the miRNAs that had corresponding target sites with different proteins,and further detected the correlation between candidate miRNAs and target genes by q PCR,Western blot and dual luciferase experiments.QPCR was used to detect the effect of evodiamine on the expression of candidate miRNAs,and to study the function of candidate miRNAs in pancreatic cancer,such as the role of glycolysis,proliferation,migration and invasion of pancreatic cancer cells,and the mechanism of related signaling pathways.Results: 1.After the pancreatic cancer cells were treated with evodiamine,the expression levels of several glycolysis and invasion related proteins were detected,and the differences were compared.It was found that evodiamine could significantly inhibit the protein expression of ldh A and mmp16 in pancreatic cancer cells.2.Further study found that both ldh A and mmp16 had mir-489-3p potential binding sites.Ldh A was confirmed to be the target gene of mir-489 by dual luciferase assay.Furthermore,q PCR and Western blot showed that mir-489 could only inhibit ldh A.QPCR results also confirmed that the expression level of mir-489-3p in pancreatic cancer cells treated with Evo was significantly increased.3.Overexpression of mir-489 can reduce the proliferation,migration and invasion of pancreatic cancer cells.Overexpression of mir-489 decreased glucose uptake and ATP production,decreased Akt signaling pathway activity and inhibited glycolysis..Conclusion: Evodiamine up regulates the expression of mir-489 in pancreatic cancer cells,thereby inhibiting the expression of its target gene ldh A.After transfection of mir-489 mimics alone,the glycolysis,proliferation,migration and invasion of pancreatic cancer cells were inhibited,and the activity of Akt signaling pathway was decreased.In conclusion,this study is the first to prove that evodiamine can induce the expression of mir-489,thereby regulating the progress of glycolysis pathway and related biological characteristics of pancreatic cancer cells.In the future,related in vivo experiments will be carried out to verify the conclusions.The current research results enrich the anti pancreatic cancer mechanism of Evo and have certain research significance.