Proliferation Inhibory Effect Of Annexin A7on Hepatoma Cell By Binding With IGFBP2

HCC is a complex disease caused by multiple genes and influenced by multiple factors. The fatality rate of HCC in malignant tumor is ranked top in our country and even the worldwide. The molecular mechanism of hepatoma, including the factors related to hepatocarcinogenesis, the search of the biological marker and treatment target referred to the early diagnosis and prediction, metastasis and recurrence has important clinical significance for diagnosis, treatment and predictions of the disease. Annexins are a large group of calcium-dependent phospholipid binding proteins participating in diverse important physiological regulation, inflammation response, anti-coagulation and play an important role in tumorigenesis. Recently, more and more studies suggest that several annexins are involved in the occurrence and development of a wide variety of tumor including hepatocellular carcinoma.Annexin A7, also known as synexin, is the protein that can aggregate chroman cellar granules, which is the first annexin to be isolated. They are widely expressed in various kinds of cells. Tumor suppressor role of Annexin A7has been shown in hormone-relevant tumors, such as prostate cancer, metastatic breast carcinoma, and glioma etc.But the exact mechanism is still unclear in its tumorigenesis and development. The existing research suggests that the abnormal expression of Annexin A7in HCC may play an important role in the regulating the development of cancer. Whether Annexin A7is abnormally expressed in hepatocellular carcinoma? Whether it is related to the phenotype of hepatocellular carcinoma cells? Those questions should be answered after further studies.In the present study, the ANXA7mRNA was detected in48HCC samples by real-time quantitative PCR. These results showed that ANXA7is down regulated in HCC and hepatoma cell lines. Decreased the expression of ANXA7by RNAi can down-regulate tumor-suppressor gene PTEN. In biological function test A7could inhibit hepatoma cell proliferation, but showed no significantly effect on migration and apoptosis. In order to find the mechanism of ANXA7in hepatocellular carcinoma, the eukaryotic expression vector with ANXA7gene and HA fusion protein was constructed successfully. Co-immunopricipitation (co-IP) was used to detect the specific binding protein of Annexin A7in HCC cells, Mass spectrometry analysis showed the difference of IGFBP2(Insulin-like growth factor binding protein2) among the two groups IGFBP2is a part of the insulin-like growth factor (IGF) system, it is important in the regulatory network, controlling essential cellular processes such as proliferation, differentiation and cell migration. IGFBP2is highly expressed in many human malignancies, including HCC, especially in the serum and liver organizations of hepatocellular carcinoma patients. It may be as a kind of independence growth regulation factor, which can regulate cell proliferation and attack by many ways. Then we further verify this interaction using specific antibodies of IGFBP2by Western blot. Besides, in order to facilitate us to better understand the interaction of them, we have respectively detected the distribution of the Annexin A7and IGFBP2in the cell level by cell immune fluorescence. The Results suggest that A7could specifically interact with IGFBP2.After confirming the relationship of them, we analyzed the correlation expression of A7and IGFBP2by Western Blot. These results showed that the expression of IGFBP2can be increased by siANXA7in SMMC7721cell line. However, up-regulated IGFBP2did not affect the expression of ANXA7. Furthermore increased IGFBP2could raise the phosphorylation of ERK1/2, while increased ANXA7could inhibit the effect. MAPK is an important signal pathway, which participates in cell proliferation, growth and differentiation. In addition, it is downstream pathways of the IGFBP2pathways. Our research suggested that Annexin A7may participate in and mediated the IGFBP2pathways through ERK1/2. If ERK1/2is inhibited, the activity of MAPK pathway can be affected, which lead to the inhibition of cell proliferation.In conclusion, we analyzed the expression and biological function of Annexin A7in HCC. The potential tumor suppressive role of Annexin A7in hepatoma could improve the knowledge of Annexin A7in malignant tumor, and is helpful for the further study of liver cancer treatments.