The Role And Mechanism Of MiR-592 In Colorectal Cancer

Objective: Colorectal cancer(CRC)is the second leading cause of cancer-related mortality worldwide.Currently available diagnostic biomarkers are neither sensitive nor specific.And dysregulated micro RNAs(mi RNAs)are significantly associated with the malignancy progress of CRC.Our study aimed to identify novel circulating micro RNA(mi RNA)as biomarkers for the diagnosis of CRC,and investigate the expression and regulatory role of the mi RNA in CRC.Methods:1.Candidate mi R-592 was identified by using the online tool GEO2 R.q RT-PCR was implemented to test the expression of mi R-592 in the serums in CRC patients and HCs.The diagnostic value of serum mi R-592 was identified by using receiver operating characteristic curve(ROC)analysis.2.The expression of mi R-592 in colon cancer tissues and cells were confirmed by q RT-PCR analysis.The colon cancer cells were transfected with mi R-592 inhibitor and NC,respectively.The transfection inhibition effect was confirmed by q RT-PCR analysis.The expression of SPARC was detectd by Western blot after transfection.Respectively,CCK-8,Ed U and transwell test were used to analyze colon cancer cell proliferation,invasion and metastasis.3.The dual luciferase assay was used to verify whether SPARC is the direct target of mi R-592 in colorectal cancer.The expression of SPARC in colon cancer tissues and cells were confirmed by q RT-PCR analysis.The colon cancer cells were transfected with si SPARC and NC,respectively.The transfection inhibition effect was confirmed by Western blot.The colon cancer cells were co-transfected with mi R-592 inhibitor and si SPARC,and the expression of SPARC was detected by Western blot after co-transfection.Respectively,CCK-8,Ed U and transwell test were used to analyze colon cancer cell proliferation,invasion and metastasis after transfection.4.The colon cancer cells were transfected with control group,si SPARC group and mi R-592 inhibitor si SPARC co-transfection group,respectively.The expression of E-cadherin and N-cadherin in colon cancer cells were detected by Western blot analysis after transfection.Results:1.We found that the expression of the serum mi R-592 was remarkably upregulated in CRC patients.ROC analysis showed that the serum mi R-592 could discriminate CRC patients from HCs with AUC value of 0.844(95% CI: 0.78-0.91,p<0.01),with 82.8% sensitivity and 78.0% specificity.To explore the value of the serum mi R-592 as an diagnostic biomarker of CRC,we compared its expression levels in serums of HCs and CRC patients with TNM I-II stages.We found that the serum mi R-592 was increased in peripheral blood of CRC patients with TNM I-II stages and could differentiate early stage CRC patients from HCs with an AUC value of 0.801(95% CI: 0.73-0.88,p<0.01),with 78.6% sensitivity and 80.0% specificity.2.The expression of mi R-592 was significantly upregulated in both CRC tissues and cell lines when compared to ANTs and FHC cells,respectively.The expression of SPARC in colon cancer cells was increased after transfection with mi R-592 inhibitor.Inhibiting the expression of mi R-592 could suppress proliferation and metastasis of CRC cells.3.We identifyed SPARC as a direct target of mi R-592 in CRC by the dual-luciferase assay.The expression of SPARC was significantly decreased in CRC tissues and cell lines compared to ANTs and FHC cell,respectively.The expression of SPARC in colon cancer cells was increased after transfection with mi R-592 inhibitor.Inhibiting the expression of SPARC could promote proliferation and metastasis of CRC cells.The expression of SPARC in colon cancer cells which were co-transfected with mi R-592 inhibitor and si SPARC was higher than that in the si SPARC group.Silencing the expression of mi R-592 significantly reversed the promotion effect of si SPARC on CRC cells.4.We found that the expression of E-cadherin was decreased in the si SPARC group compared with the control group,while the expression of N-cadherin was increased.The expression of E-cadherin was increased in the mi R-592 inhibitor si SPARC co-transfection group compared with the si SPARC group,while the expression of N-cadherin was decreased.Conclusion:Serum mi R-592 may be a novel potential biomarker for diagnosis of CRC.Mi R-592 acts as an oncogene which directly inhibits the expression of SPARC to induce EMT in colon cancer cells,thereby promoting cell proliferation,invasion and metastasis.