Function And Mechanism Of Small Ras Homologous Protein ARHI In Epithelial Mesenchymal Transition Of Pancreatic Carcinoma Cells

Background and Aim: Pancreatic cancer is a highly malignant tumor of digestive tract, which is difficult for diagnosis and treatment, especially strong invasion and metastasis are important factors for pancreatic cancer mortality. EMT is an important phenomenon in the process of cancer development, not only contribute to the early spread of cancer cells, but alsothe migration and invasion capabilityand cell stemness maintenanceof malignant tumor cells. Small Ras homologuesprotein ARHI haveimportant functional roles in cell cycle arrest, inhibition of tumor cell proliferation andpromotionof autophagy and apoptosisin pancreatic carcinoma cells, but its function and mechanism in EMT of pancreatic carcinoma cells remain poorly understood.In thisstudy, we will investigatethe relation of the expression of ARHI with EMT and cellstemness markersassociated protein, andexplore the regulation mechanismthat ARHI mediated EMT and cell stemess.Methods:First, ARHI expression in different progression of pancreatic cancer,the metastasis and non-metastasis of pancreatic cancer was examined by IHC;Secondly,the correlation of ARHI and EMT associated proteins expression were detected by Western blot in different degree of differentiation of pancreatic cancer cells, andchanges in the expression of EMT marker proteins were studied by changing the expression of endogenous ARHI in the protein and m RNA levels; Thirdly, the related biological functions were studiedafter Transwell assay, scanning electron microscopeand Tumorsphere formation assay.Meanwhile,the liver metastasis model was constructed by spleen injection in nude mice, and the effect of ARHI on the migration and invasion of pancreatic cancer cells was further verified in vivo.Finally,after treatment with inhibitor U0126 of ERK signal pathway, we investigatedthe level of ERK phosphorylation,EMT and cell stemnessmarkers associated proteinexpressionby Western blot.Results:The expression of ARHI was positively correlated with grade malignancy of pancreatic cancer cells, also with EMT and cell stemness level of pancreatic carcinoma cells.In vitro and in vivo biological function study found that ARHI promotes EMT-related phenotypes of pancreatic cancer cells,for example podosome formation,tumorsphere formation and invasion; signal pathway screening experiment showed ERK phosphorylation was significantly increased when ARHI was knocked down; after the application of inhibitors, most EMT markers protein expression were antagonistic,however, theexpressionlevel of Notch-1 protein stilldecreased.Conclusions:Our study suggested thatthe promotion effect ofARHI on EMT and cell stemness maintenanceof pancreatic carcinoma cells is not completely dependent on ERK signal pathway, also closely related to the expression of key protein Notch-1.