The Roles And Mechanisms Of MiR-183 In Pancreatic Cancer Progression And Chemoresistance

Background The incidence of pancreatic cancer has increased in the past decades all over the world.However,pancreatic cancer decreased slightly in survival,in contrast to other cancer with continuous increases,and it is commonly diagnosed at an advanced stage,leading to a 5-year survival rate of only 6%.At initial diagnosis,50% of patients present with metastatic disease,but 30% of patients have locally advanced tumors,only 20% of cases can be resected.For most patients,curative treatment options do not exist at the time of diagnosis.Surgery is considered the best treatment option,but is restricted to localized pancreatic cancer without evidence of distant metastasis.Recently,the report showed that 5-year survival rates of 15% to30% in patients with margin negative resection and adjuvant chemotherapy with either gemcitabine alone or other methods.Despite of adjuvant chemotherapy,local recurrence of pancreatic cancer occurs in up to 90% of postoperative cases.The gemcitabine resistance has already become the main reason for poor performance of in the current treatment.Therefore,screening of gemcitabine resistance related with biological markers and then improvement of gemcitabine sensitivity are the main huge challenge of pancreatic cancer research.The most common mechanism by which gemcitabine sensitivity arises include abnormal gene expression,loss of signaling pathways,epithelial-mesenchymal transition(EMT),istromal proliferation,induction of drug-detoxifying,all of which serve a crucial role in the development of gemcitabine resistance of cancer cells.Therefore,it is crucial to develop comprehensive understanding of the mechanism by which gemcitabine sensitivity arises in pancreatic cancer cells to identify a new therapeutic strategy to overcome gemcitabine sensitivity and improve the clinical therapeutic response of patients with pancreatic cancer.MicroRNAs post-transcriptionally regulate a variety of target genes by binding to the 3′-untranslated region(3′-UTR)of their target genes.The aberrant expression ofmiRNAs serves either oncogenic or tumor-suppressive roles in the tumorigenesis,progression,metastasis and chemoresistance of various types of cancer include pancreatic cancer.Purpose our study is to investigate the underlying mechanism of its role of miR-183 in pancreatic cancer in tumor progression。Methods The mRNA expression of miR-183 in pancreatic cancer tissues and pancreatic cell lines(PANC-1,ASPC-1,BxPC3)was detected by qRT-PCR,and then according to F test and Chi-square test,the research analyzed the correlation between miR-183 mRNA expression and clinicopathological characteristics of pancreatic cancer.We constructed over-expression of miR-183 and its inhibitors and obsevered the proliferation,apoptosis cell cycle and gemcitabine sensitivity in pancreatic cancer celllines.Biological information prediction and analysis were uesd to identify the putative targets of miR-183,and it will be further affirmed by luciferase reporter assay,qRT-PCR and Westem blotting.The proliferation was assayed by MTT and FCM.the PANC-1 cells transfected by miR-183 were transplanted into subcutaneous of nude miceResults1.The expression level of miR-183 was significantly higher in pancreatic cancer than that in adjacent non-cancerous tissues,and the amount of miR-183 increased progressively with loss of histological differentiation and with the grades of pancreactic cancer.Compare with the human pancreatic ductal epithelial cell line,the expression level of miR-183 is upregulated in pancreatic cancer cell lines(PANC-1,ASPC-1,BxPC3).2.We constructed over-expression of miR-183 and its inhibitors and then transfected into two stable pancreatic cancer cell lines,PANC-1 and BxPC-3.The results of MTT and FCM indicated that downregulation of miR-183 inhibits pancreatic cancer cell proliferation in vitro.Compared with the control,miR-183 significantly increased the population in G1 phase in both PANC-1 cells and BxPC-3cells.In vivo research,PANC-1 cell that were transfected with a control vector ormiR-183 or miR-183 inhibitor were subcutaneously into nude mice.The tumors in the nude mice implanted with miR-183 over-expression cells grew faster and bigger than that in the nude mice with miR-183 inhibitor and control cells.3.To study the effects of miR-183 on the gemcitabine sensitivity of pancreatic cancer cells,various gemcitabine concentrations(0.1、1、10、100 μg/mL)were prepared.Pancreatic cancer cells were transfected with miR-183 mimics or inhibitor 24 h before gemcitabine treatment,and the impact of pancreatic cancer cells of gemcitabine were measured 48 h after treatment by using the MTT.Based on TargetScan miRNA target analysis,BNIP3 L gene is a potential target mRNA of miR-183.The dual-luciferase reporter system result indicated that the coexpression of miR-183 significantly inhibited the firefly luciferase reporter activity of the wild-type BNIP3 L 3’-UTR but not inhibited the activity of the mutant 3’-UTR,and Western blot analysis also revealed that BNIP3 L is a direct target of miR-183.To further confirm the basic mechanism of the effect of BNIP3 L gene in the in tumor progress of miR-183,Western blot analysis showed that miR-183 overexpression increased PI3K/AKT signaling,whereas the ectopic expression of BNIP3 L blocked the effect of miR-183 inclued the effects on cell proliferation,apoptosis and chemoresistance.Conclusion1.miR-183 is upregulated in pancreatic cancer tissues and three pancreatic cancer cell lines and correlates with tumor progression2.miR-183 promotes pancreatic cancer cell proliferation in vitro and in vivo,inhibited cell apoptosis and cell cycle G1 phase and also related with gemcitabine sensitivity3.miR-183 induces pancreatic cancer cell growth,apoptosis and gemcitabine sensitivity inhibition mediated by the BNIP3 L gene through PI3K/AKT signaling pathway.4.miR-183 may be a novel and crucial biomarkers based on digonoiss and therapeutics in pancreatic cancer.