An Experiment On Role Mechanism Of MiRNA-423-3p Targeting For ISG15 To Regulate PD-L1 In Pancreatic Cancer

ObjectivePancreatic cancer(PC)is one of the top ten malignant tumors in China.PC is hard to determine,early diagnosis and treatment is the key to improve survival rate of patients with PC.We detected the expression of miRNA-423-3p(miR-423-3p)in the serum of PC patients and healthy people,analyzed its relationship with the clinicopathology and prognosis of patients with PC,explored the effects of miR-423-3p on the proliferation,migration,invasion,adhesion of PC cells and tumor growth in vivo by regulating ISG15 and PD-L1,To clarify the mechanism of the occurrence and development of PC,provide strategies for early diagnosis and treatment of PC.MethodsqRT-PCR was measured to detect the expression level of miR-423-3p in the serum of PC patients and healthy people,analyzed its relationship with the clinicopathological characteristics and prognosis of PC patients,and analyzed the correlation between serum miR-423-3p and ISG15 in PC tissues.Western blot,qRT-PCR,dual-luciferase reporter assay were carried out to study the regulation of miR-423-3p on the expression of ISG15 and PD-L1.The CCK-8 assay,wound-healing assay,Transwell chamber assay and adhesion assay were used to detect the effect of miR-423-3p on the proliferation,migration,invasion and adhesion ability of PC cell.A nude mouse PC xenograft model was constructed,and miR-423-3p agomir and antagomir were injected into the tail vein to study the effect of miR-423-3p on PC in vivo.Results1.The expression of miR-423-3p in the serum of PC patients is lower than that of healthy people.The expression of miR-423-3p in serum is related to clinicopathological characteristics such as TNM stage and resectability in PC patients,and low expression of miR-423-3p in serum is related to poor prognosis of PC patients.2.The immunohistochemistry showed that the expression of ISG15 and PD-L1 in the cancer tissue of PC patients was significantly higher than that in the adjacent tissues.The expression of miR-423-3p in the serum of PC patients was negatively correlated with the expression of ISG15 in the PC tissue.3.The expression of miR-423-3p in PC cell lines is lower than that in human normal pancreatic ductal epithelial cell lines.4.Transfection of synthetic oligonucleotides can stably and effectively regulate the expression of miR-423-3p in cells.5.miR-423-3p can target the 3’ UTR of ISG15 to downregulate ISG15.Down-regulation of the ISG15 pathway reduces the expression of PD-L1 in PC.6.miR-423-3p inhibits the proliferation,migration,invasion and adhesion ability of PC cells.7.miR-423-3p inhibits the growth of PC subcutaneously transplanted tumors in nude mice Conclusions1.The low expression of miR-423-3p in PC patients is negatively correlated with TNM stage,and its low expression indicates poor prognosis.2.ISG15 is highly expressed in human PC tissues and promotes the occurrence and development of PC.The combined detection of miR-423-3p and ISG15 may be a tumor marker for early diagnosis and prognosis of PC.3.miR-423-3p targeted down-regulation of ISG15 in PC,thereby reducing the expression of PD-L1,inhibiting PC cell proliferation and invasive metastasis,and playing a tumor suppressor role in PC.4.Down-regulation of the ISG15 pathway reduces the expression of PD-L1 in PC,which may improve the efficacy of immune checkpoint inhibitors and increase the overall survival of PC patients who were treated with immunotherapy.