Raf kinase inhibitor protein inhibits cholangiocarcinoma cell metastasis by downregulating matrix metalloproteinase 9 and upregulating tissue inhibitor of metalloproteinase 4 expression.

Junji Ma,Wenbin Wang,Xiaoyu Jiang,Lijuan Cheng,Junli Shi,Fangfang Li,Dongqiang Zhao,Huiqing Jiang

doi:10.3892/ol.2014.2637

Abstract

Cholangiocarcinoma cells originate in the biliary epithelium. The cells easily metastasize and cause relapse. The effect of Raf kinase inhibitor protein (RKIP) on the biological behavior of cholangiocarcinoma cells is not yet clear. In the present study, RKIP and cytokeratin 19 expression was detected in the extrahepatic tissues of cholangiocarcinoma patients by immunohistochemistry. RKIP small interfering (si)RNA or an RKIP-overexpressing adenoviral vector were used to infect the human cholangiocarcinoma RBE cell line. RKIP protein or gene expression was analyzed by western blotting or reverse transcription-quantitative polymerase chain reaction (RT-qPCR), respectively. The cells were assayed for proliferation, apoptosis, invasion and migration. Matrix metalloproteinase 9 (MMP-9) and tissue inhibitor of metalloproteinase 4 (TIMP-4) mRNA was assayed by RT-qPCR. RKIP expression was reduced in the extrahepatic cholangiocarcinoma tumor compared with the adjacent uninvolved peritumoral tissues. The current study revealed that RKIP expression was positively correlated with cell differentiation, but negatively correlated with lymph node or distant metastasis (P<0.05). RKIP siRNA treatment promoted RBE cell invasion, but RKIP overexpression prevented cell invasion. In the pDC316-siRNA recombinant vector group, the cells migrated more quickly compared with the siRNA-negative control group, and in the RKIP-expressing adenoviral vector group, the cells migrated more slowly compared with the adenoviral negative control group. RKIP inhibited the invasive and metastatic ability of the cholangiocarcinoma cell line, RBE, by downregulating MMP-9 and upregulating TIMP-4 mRNA expression. RKIP is negatively associated with cholangiocarcinoma distant metastasis and prevents cholangiocarcinoma cell metastasis through downregulating MMP-9 expression and upregulating TIMP-4 expression.

Highlights

Cholangiocarcinoma is a malignant tumor that originates in the intra‐ and extrahepatic biliary epithelium and is commonly found in the elderly, being more frequent in males than females
CK19 was constitutively expressed in the normal cholangiocytes and cholangiocarcinoma cells and there was no significant difference between the two groups (χ2 test, P>0.05; Fig. 1A and B)
The frequency of Raf kinase inhibitor protein (RKIP)‐positive expression was 73 vs. 33% in the normal cholangiocytes and cholangiocarcinoma, respectively (Fig. 1C and D). These results indicated that the RKIP expression was significantly lower in the cholangiocarcinoma cells compared with the normal cholangiocytes (χ2 test; P

Summary

Introduction

Cholangiocarcinoma is a malignant tumor that originates in the intra‐ and extrahepatic biliary epithelium and is commonly found in the elderly, being more frequent in males than females. Certain common symptoms, including jaundice, usually do not present until late in the course of the disease, at which time they are relatively resistant to chemotherapeutic agents. As such, these symptoms are difficult to treat and exhibit a poor prognosis [1,2,3,4]. The formation of cholangiocarcinoma often proceeds through several steps, including interaction among the cancer‐promoting environmental factors, oncogene activation and tumor‐suppressor gene inactivation. During the tumorigenesis of cholangiocarcinoma, certain growth factors, receptors and signaling pathway molecules may be involved [1]

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Conclusion