The Role Of MicroRNA-9 In Pancreatic Cancer And Its Correlation Analysis With Prognosis

ObjectiveThe aim of this study,on the one hand,is to detect the expressions of miRNA-9 in different tumors of the abdomen including pancreatic cancer and adjacent normal pancreatic tissues,analyze the relationship between miR-9 and clinicopathological parameters and further reveal the correlation between miR-9 and the prognosis of patients in pancreatic cancer,on the other hand,to demonstrate the functions of miRNA-9 including proliferation,invasion and migration on human pancreatic cancer cell line(Panc-1).MethodsPart I: Analysis of the correlation between miR-9 expression and clinicopathological parameters in patients with pancreatic cancer 1 The expression level of miR-9 in different tumors of the abdomen was detected by RT-PCR;2 The expression level of miR-9 in pancreatic cancer tissues and adjacent normal pancreatic tissues was detected by RT-PCR;3 The relationship between miR-9 expression and clinicopathological parameters by using the four-table ? test and the correlation analysis between miR-9 and the prognosis of patients in pancreatic cancer by using Kaplan-Meier;Part II: Effect of miR-9 on biological behaviors of pancreatic cancer cells 1 Cell scratch assay detects cell migration ability.First,human pancreatic cancer cell line(Panc-1)were seeded on six-well plates and transfected with miR-9-inhibitor or miR-9 inhibitor control at a cell confluence of 70%-80%.Second,when the bottom of 6-well plates were covered with the cells attainting to the confluence of 100%,we scratched.Third,we observed the healing status of the two groups of "wounds" at 0,12,24 h and conducted the comparison between the two groups;2 Transwell assay detects the abilities of cell migration and invasion: Panc-1 cells transfected with miR-9-inhibitor were seeded on the upper transwell chambers with matrigel-coated and matrigel-free and set the control group seperately.After 48 and 60 hours,the numbers of cells passing through the chambers were observed and compared to evaluate the effect of miR-9 on the ability of cancer cells to migrate and invade;3 CCK-8 assay detects cell proliferation ability.First,human pancreatic cancer cell line(Panc-1)were seeded in 96-well plates and transfected with miR-9-inhibitor or miR-9 inhibitor control at a cell confluence of 30-50%.Then,CCK-8 reagent was added into experimental and control group separately after cells were cultured for 0,24,48 and 72 hours respectively.Third,the absorbance of the two groups of cells was measured by the microplate reader at 2 hours after giving CCK-8 so as to evaluate the effect of miR-9 on the proliferation of cancer cells.Finally,the value of absorbance were compared by using t test.ResultsPart I: The relationship between miR-9 expression and clinicopathological parameters of patients diagnosed as pancreatic cancer and the correlation analysis between miR-9 and prognosis 1 The expressions of miR-9 in tumor samples from different organs of the abdomen were detected by RT-PCR.The results showed that the expression of miR-9 in pancreatic cancer was significantly higher than that of liver,colon tumors(P<0.05);2 The expressions of miR-9 in pancreatic cancer and adjacent normal pancreatic tissues were analyzed by RT-PCR and the results showed that the expression level of miR-9 in pancreatic cancer tissues was significantly higher than that of adjacent normal pancreatic tissues(P<0.05);3 The result of the correlation analysis between miR-9 expression in pancreatic cancer tissue specimens and clinicopathological data indicates that the incidence of lymph node metastasis was significantly higher in the group with the high expression of miR-9 than that of the group with the low expression of miR-9.In addition,the tumor was larger and the tumor differentiation was lower in the group with high expression of miR-9(P<0.05).Part II: The analysis results of the effect of miR-9 on biological behaviors of pancreatic cancer cells 1 The results of cell scratches and transwell experimental show that the “wound healing rate” of pancreatic cancer cell line(Panc-1)transfected with miR-9-inhibitor was slower than that of the miR-9 inhibitor control group(P<0.05).PANC-1 cells transfected with miR-9-inhibitor were seeded in Transwell chambers with matrigel-free and matrigel-coated and set the control group separately.After 48 hours and 60 hours respectively,the number of cells passing through the cells in the experimental group was significantly less than that in the control group(P<0.05);2 CCK-8 experimental results suggest: The proliferation of Panc-1 cells transfected with miR-9-inhibitor was significantly weaker than that of the control group(P<0.05).Conclusion 1 The expression of miR-9 in pancreatic cancer tissue specimens was significantly higher than that of other abdominal organs tumors,suggesting that miR-9 is enriched in pancreatic cancer.The expression of miR-9 in pancreatic cancer tissues was significantly higher than that of adjacent normal pancreatic tissues,suggesting that the expression of miR-9 is abnormal in pancreatic cancer;2 The expression level of miR-9 in pancreatic cancer tissue specimens was correlated with the situation of lymph node metastasis.The tumor differentiation,tumor size and the long-term survival of patients in the group with the high expression of miR-9 was significantly lower than that of patients in the group with low expression of miR-9.It is speculated that miR-9 promotes the malignant progress of pancreatic cancer;Multivariate analysis of prognosis of pancreatic cancer suggests that miR-9 expression is an independent risky factor in the prognosis of pancreatic cancer;3 The results of in vitro function experiments of pancreatic cancer cell line PANC-1 suggest that miR-9 plays a role of oncogene in pancreatic cancer.