angiogenin and Liver-Neoplasms

Topics: Animals; Humans; Liver Neoplasms; Myocardial Ischemia; Neovascularization, Physiologic; Proteins; Ribonuclease, Pancreatic

Angiogenin (ANG) is known to alter multiple cell behaviors by directly targeting downstream targets, but its role in hepatocellular carcinoma (HCC) remains to be elucidated. The expression of ANG in HCC cell lines was measured by quantitative real-time polymerase chain reaction (qRT-PCR) and western blot. The effects of ANG expression on cell proliferation, cell migration, and hallmarks of epithelial-mesenchymal transition (EMT) process were also investigated. The relationship between ANG and high mobility group AT-hook 2 (HMGA2) was evaluated. ANG expression was increased in HCC cell lines. Downregulating of ANG inhibits proliferation, migration, and EMT of HCC cells. The direct regulation of ANG on HMGA2 was verified by luciferase activity reporter assay and western blot assay. Furthermore, overexpression of HMGA2 reversed the inhibitory effects of ANG downregulation on HCC cell behaviors. Our results illustrated the mechanism that ANG promote the EMT of HCC through targeting HMGA2.

Topics: Carcinoma, Hepatocellular; Cell Line, Tumor; Cell Movement; Cell Proliferation; Down-Regulation; Epithelial-Mesenchymal Transition; HMGA2 Protein; Humans; Liver Neoplasms; Ribonuclease, Pancreatic; RNA, Small Interfering; Transfection; Up-Regulation

Patients with hepatitis B virus (HBV) infection are at a high risk of developing hepatocellular carcinoma (HCC). In this study, we aim to investigate the roles of HBV on angiogenin (ANG), as well as the effects on cell proliferation in presence of ANG down-regulation.. Serum ANG was determined by ELISA. The expression of ANG mRNA and protein in HCC cell lines with or without HBV/HBx were determined. Western blot and ELISA were conducted to determine the effects of HBV/HBx on IL-6 expression. The role of IL-6 on ANG was evaluated by IL-6 recombinant protein or IL-6 neutralizing antibody. Immunofluorescence staining was used to detect the nuclear translocation of ANG. MTT was performed to evaluate the relative inhibition ratio.. In vivo experiments showed elevation of serum ANG in patients infected with HBV. In vitro experiments showed HBV and HBx contributed to the transcription and translation of ANG. ANG expression showed increase after IL-6 stimulation, and ANG protein decreased in the presence of IL-6 blocking with its antibody. HBV promoted nuclear translocation of ANG. Inhibiting ANG expression or blocking of nuclear transfer of ANG attenuated the 45S rRNA synthesis and cell proliferation.. HBV and HBx protein can increase the level of ANG through IL-6. HBV and HBx contributed to the nuclear translocation of ANG. Cell proliferation was inhibited after inhibiting the expression or nuclear transfer of ANG.

Topics: Adult; Antibodies; Carcinoma, Hepatocellular; Cell Line, Tumor; Cell Nucleus; Cell Proliferation; Female; Hep G2 Cells; Hepatitis B; Hepatitis B virus; Humans; Interleukin-6; Liver Neoplasms; Male; Middle Aged; Ribonuclease, Pancreatic; RNA Interference; RNA, Ribosomal; Trans-Activators; Up-Regulation; Viral Regulatory and Accessory Proteins

Persistent infections with hepatitis B virus (HBV) or hepatitis C virus (HCV) account for the majority of cases of hepatic cirrhosis and hepatocellular carcinoma (HCC) worldwide. Small, non-coding RNAs play important roles in virus-host interactions. We used high throughput sequencing to conduct an unbiased profiling of small (14-40 nts) RNAs in liver from Japanese subjects with advanced hepatitis B or C and hepatocellular carcinoma (HCC). Small RNAs derived from tRNAs, specifically 30-35 nucleotide-long 5' tRNA-halves (5' tRHs), were abundant in non-malignant liver and significantly increased in humans and chimpanzees with chronic viral hepatitis. 5' tRH abundance exceeded microRNA abundance in most infected non-cancerous tissues. In contrast, in matched cancer tissue, 5' tRH abundance was reduced, and relative abundance of individual 5' tRHs was altered. In hepatitis B-associated HCC, 5' tRH abundance correlated with expression of the tRNA-cleaving ribonuclease, angiogenin. These results demonstrate that tRHs are the most abundant small RNAs in chronically infected liver and that their abundance is altered in liver cancer.

Topics: Animals; Base Sequence; Carcinoma, Hepatocellular; Hepatitis B, Chronic; Hepatitis C, Chronic; High-Throughput Nucleotide Sequencing; Humans; Immunoprecipitation; Liver; Liver Neoplasms; MicroRNAs; Molecular Sequence Data; Pan troglodytes; Real-Time Polymerase Chain Reaction; Ribonuclease, Pancreatic; RNA, Transfer; Sequence Analysis, RNA

Resection of a primary colorectal carcinoma (CRC) can be accompanied by rapid outgrowth of liver metastases, suggesting a role for angiogenesis. The aim of this study is to investigate whether the presence of a primary CRC is associated with changes in angiogenic status and proliferation/apoptotic rate in synchronous liver metastases and/or adjacent liver parenchyma.. Gene expression and localization of CD31, HIF-1α, members of the vascular endothelial growth factor (VEGF) and Angiopoietin (Ang) system were studied using qRT-PCR and immunohistochemistry in colorectal liver metastases and nontumorous-adjacent liver parenchyma. Proliferation and apoptotic rate were quantified. Three groups of patients were included: (1) simultaneous resection of synchronous liver metastases and primary tumor (SS-group), (2) resection of synchronous liver metastases 3 to 12 months after resection of the primary tumor [late synchronous (LS-group)], and (3) resection of metachronous metastases >14 months after resection of the primary tumor (M-group).. In all 3 groups a higher expression of the angiogenic factors was encountered in adjacent liver parenchyma as compared to the metastases. VEGFR-2 gene expression was abundant in adjacent liver parenchyma in all 3 groups. VEGF-A and VEGFR-1 were prominent in adjacent parenchyma in the SS-group. The SS-group showed the highest Ang-2/Ang-1 ratio both in the metastases and the adjacent liver. This was accompanied by a high turnover of tumor cells.. In the presence of the primary tumor, the liver parenchyma adjacent to the synchronous liver metastases provides an angiogenic prosperous environment for metastatic tumor growth.

Topics: Adult; Aged; Apoptosis; Cell Proliferation; Colorectal Neoplasms; Disease Progression; Female; Gene Expression Regulation, Neoplastic; Humans; Hypoxia-Inducible Factor 1, alpha Subunit; Liver; Liver Neoplasms; Male; Membrane Proteins; Middle Aged; Neoplasms, Multiple Primary; Neoplasms, Second Primary; Neovascularization, Pathologic; Platelet Endothelial Cell Adhesion Molecule-1; Prognosis; Real-Time Polymerase Chain Reaction; Ribonuclease, Pancreatic; Vascular Endothelial Growth Factor A; Vascular Endothelial Growth Factor Receptor-2

Staphylococcal nuclease domain-containing 1 (SND1) is a multifunctional protein that is overexpressed in multiple cancers, including hepatocellular carcinoma (HCC). Stable overexpression of SND1 in Hep3B cells expressing a low level of SND1 augments, whereas stable knockdown of SND1 in QGY-7703 cells expressing a high level of SND1 inhibits establishment of xenografts in nude mice, indicating that SND1 promotes an aggressive tumorigenic phenotype. In this study we analyzed the role of SND1 in regulating tumor angiogenesis, a hallmark of cancer. Conditioned medium from Hep3B-SND1 cells stably overexpressing SND1 augmented, whereas that from QGY-SND1si cells stably overexpressing SND1 siRNA significantly inhibited angiogenesis, as analyzed by a chicken chorioallantoic membrane assay and a human umbilical vein endothelial cell differentiation assay. We unraveled a linear pathway in which SND1-induced activation of NF-κB resulted in induction of miR-221 and subsequent induction of angiogenic factors Angiogenin and CXCL16. Inhibition of either of these components resulted in significant inhibition of SND1-induced angiogenesis, thus highlighting the importance of this molecular cascade in regulating SND1 function. Because SND1 regulates NF-κB and miR-221, two important determinants of HCC controlling the aggressive phenotype, SND1 inhibition might be an effective strategy to counteract this fatal malady.

Topics: Carcinoma, Hepatocellular; Cell Nucleus; Chemokine CXCL16; Chemokines, CXC; Endonucleases; Enzyme-Linked Immunosorbent Assay; Gene Expression Regulation, Neoplastic; Human Umbilical Vein Endothelial Cells; Humans; Liver Neoplasms; MicroRNAs; Neoplasm Metastasis; Neovascularization, Pathologic; NF-kappa B; Nuclear Proteins; Receptors, Scavenger; Ribonuclease, Pancreatic; RNA, Small Interfering

Hepatocellular carcinoma (HCC) is one of the most common cancers worldwide. Basic fibroblast growth factor (bFGF), which is highly expressed in developing tissues and malignant cells, regulates cell growth, differentiation, and migration. Its expression is essential for the progression and metastasis of HCC. This study aims to investigate the effects of bFGF on the expression of angiogenin, another growth factor, which plays an important role in tumor angiogenesis, and on cell proliferation in H7402 human hepatoma cells. The bFGF sense cDNA or antisense cDNA was stably transfected into H7402 cells. Genomic DNA PCR analysis demonstrated that human bFGF sense cDNA or antisense cDNA was inserted into the genome. Furthermore, the expression of bFGF and angiogenin was examined by RT-PCR and Western blot assays. MTT and colony formation assays were employed to determine cell proliferation. Stable bFGF over-expressing and under-expressing transfectants were successfully established. Expression of angiogenin was decreased in the over-expressing bFGF cells (sense transfectants) and was increased in the under-expressing bFGF cells (antisense transfectants). Cell proliferation increased in the bFGF sense transfectants and decreased in the bFGF antisense transfectants. These results demonstrated that the endogenous bFGF may not only negatively regulate the angiogenin expression but also contribute to the overall cell proliferation in H7402 human hepatoma cells. This study may be helpful in finding a potential therapeutic approach to HCC.

Topics: Carcinoma, Hepatocellular; Cell Line, Tumor; Cell Proliferation; DNA; Fibroblast Growth Factor 2; Gene Expression Regulation, Neoplastic; Genome, Human; Humans; Liver Neoplasms; Ribonuclease, Pancreatic; Transgenes

Physiological angiogenesis occurs during liver regeneration, leading to the formation of new functional sinusoids. Pathological angiogenesis occurs in hepatocellular carcinoma (HCC). We aimed to evaluate the expression of angiogenic factors in hepatitis C virus (HCV)-HCC tissues and the utility of angiogenesis soluble factors as noninvasive markers of HCC and tumor growth.. Thirty-eight HCV-HCC tumors with 10 corresponding nontumor cirrhotic tissues, as well as 42 independent HCV cirrhotic and 6 normal liver tissues were studied using high-density oligonucleotide arrays. Human angiogenesis microarray was used for the protein detection of EGF, TIMP-1, TIMP-2, HGF, angiopn-1, angiopn-2, VEGF-A, IP-10, PDGF, KGF, angiogenin, VEGF-D, ICAM-1, and FGF in plasma samples from 40 patients (30 HCCs and 10 HCV cirrhosis).. From the gene expression analysis of the HCV-HCC tumors compared to normal livers, we found an important number of genes related to angiogenesis differentially expressed (alpha=0.01), including VEGF, PDGF, AGPTL2, ANG, EGFL6, EGFR, angiopn-1, angiopn-2, ICAM2, TIMP-2, among others. Moreover, angiogenic genes were also differentially expressed when HCV-HCC samples were compared to HCV cirrhotic tissues (alpha=0.01; VEGF, EGFL3, EGFR, VEGFB, among others). Ten out of 14 angiogenic proteins analyzed were statistically differentially expressed between HCV cirrhosis and HCV-HCC groups (TIMP-1, TIMP-2, HGF, angiopn-1, angiopn-2, VEGF-A, IP-10, PDGF, KGF, and FGF; P<0.05). In addition, we observed that angiopn-2 was the most significant predictor (area under the curve: 0.83).. Differentially expressed angiogenesis genes were observed between HCV patients with and without HCC. Soluble angiogenic factors might be useful for monitoring high-risk HCV patients.

Topics: Adult; Aged; Carcinoma, Hepatocellular; Female; Fibroblast Growth Factors; Hepatitis C; Humans; Liver Cirrhosis; Liver Neoplasms; Liver Transplantation; Male; Middle Aged; Neovascularization, Physiologic; Oligonucleotide Array Sequence Analysis; Platelet-Derived Growth Factor; Ribonuclease, Pancreatic; Tissue Inhibitor of Metalloproteinase-1; Tissue Inhibitor of Metalloproteinase-2; Vascular Endothelial Growth Factor A; Waiting Lists

Hepatocellular carcinoma (HCC) is a hypervascular malignancy. Vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF), and angiogenin (ANG) are important angiogenic factors of neoangiogenesis. This study investigated the predictive value of serum VEGF, bFGF, and ANG in tumor recurrence, disease-free survival (DFS), and overall survival (OS) in HCC patients.. Preoperative serum VEGF, bFGF, and ANG were measured in 98 patients with resectable HCC and in 15 healthy controls. The median follow-up time was 43 months.. Preoperative serum VEGF was increased in patients with resectable HCC compared with healthy controls (P <.05). Increased serum VEGF was correlated with tumor recurrence (P =.001). Univariate analysis showed that serum VEGF, tumor-node-metastasis stage, tumor size and number, macroscopic portal vein invasion, and microscopic vascular invasion were correlated with OS and DFS. Serum bFGF and ANG were not associated with survival. Multivariate analysis showed that serum VEGF was the most significant predictor of DFS (relative risk, 2.35; 95% confidence interval, 1.26-4.39; P =.007) and OS (relative risk, 3.44; 95% confidence interval, 1.81-6.57; P <.001) in HCC patients after surgical resection.. Preoperative serum VEGF is a significant independent predictor of tumor recurrence, DFS, and OS in patients with resectable HCC.

Topics: Carcinoma, Hepatocellular; Endothelial Growth Factors; Female; Fibroblast Growth Factor 2; Humans; Intercellular Signaling Peptides and Proteins; Liver Neoplasms; Lymphokines; Male; Middle Aged; Multivariate Analysis; Neoplasm Invasiveness; Neoplasm Recurrence, Local; Portal Vein; Prognosis; Ribonuclease, Pancreatic; Survival Analysis; Vascular Endothelial Growth Factor A; Vascular Endothelial Growth Factors

Neovascularization is known to be one of the major characteristics of human hepatocellular carcinoma (HCC). Angiogenin (ANG), originally discovered in a human colon cancer cell line, is a liver-derived polypeptide that shows strong angiogenic activity in vivo. However, the role of ANG on the development of HCC remains unknown. The present study was designed to examine the implication of ANG in the neovascularization of human HCC.. Forty-one HCC patients who had undergone conventional celiac angiography were used in this study. ANG protein expression and microvessel density (MVD) in HCC specimens obtained by liver biopsy or surgical resection were examined by immunohistochemistry, and the levels were quantified by the KS-400 image analyzing system. ANG mRNA expression in liver tissues was evaluated by in situ hybridization. Serum ANG concentrations were measured by an ELISA. Survival rates were calculated using the Kaplan-Meier method.. Immunohistochemistry and in situ hybridization showed greater increments of ANG protein expression and mRNA expression, respectively, in HCC tissues than in the surrounding nontumorous tissues. MVD within tumorous tissues increased according to dedifferentiation of the histological grade of HCC, showing a significant correlation (r = 0.877, P = 0.0009) with ANG expression levels. Mean +/- SD serum ANG levels of healthy subjects and chronic hepatitis (CH) patients were 362.3 +/- 84.1 ng/ml and 331.9 +/- 133.8 ng/ml, respectively, with no significant difference. Serum ANG levels of liver cirrhosis patients (242.4 +/- 126.9 ng/ml) were lower than those of healthy subjects or CH patients and decreased as the fibrosis grade advanced. In HCC patients, despite the cirrhotic background, serum ANG levels increased as the tumor vascularity increased (197.8 +/- 64.9 ng/ml for hypovascular, 326.7 +/- 148.6 ng/ml for hypervascular, and 405.0 +/- 121.3 ng/ml for very hypervascular), in good accordance with histological grading, and significantly decreased (P = 0.015) after successful treatment with transcatheter arterial embolization or percutaneous ethanol injection. HCC patients were conventionally divided into two groups according to the serum level of ANG, those with values higher than the mean level (332.9 +/- 143.8 ng/ml) and those with values lower than the mean,; the 5-year survival rate of the latter group was determined to be significantly higher than that in the former group.. These results suggest that ANG is one of the neovascularization defining factors of HCC. Thus, measuring serum ANG may assist in monitoring the disease, and targeting ANG may provide a new strategy for treating advanced HCC.

Topics: Adult; Aged; Aged, 80 and over; Carcinoma, Hepatocellular; Cell Differentiation; Cell Line, Tumor; Enzyme-Linked Immunosorbent Assay; Female; Humans; Immunohistochemistry; In Situ Hybridization; Liver; Liver Neoplasms; Male; Middle Aged; Neovascularization, Pathologic; Ribonuclease, Pancreatic; RNA, Messenger; Time Factors

It has been suggested that angiogenin binds to an actin-like molecule present on the surface of endothelial cells. Actin inhibits plasmin activity, but the angiogenin-actin complex is not active. In this report, we found that plasmin inhibits the interaction between angiogenin and actin suggesting a possibility that both angiogenin and plasmin may bind to a similar site on actin. Here we report that chANG, an antiangiogenin peptide that binds to the actin-binding site of angiogenin, inhibits the proteolytic activity of plasmin without any apparent effect on the activities of plasminogen activators and matrix metalloproteases. Its antiplasmin activity is comparable with that of actin. chANG inhibits plasmin activity via its binding to plasmin kringle domains while scrambled chANG does not bind to plasmin. chANG also inhibits the invasion of angiogenin-secreting human fibrosarcoma and colorectal carcinoma cells without effecting migration. Furthermore, chANG blocks angiogenesis induced by fibrosarcoma cells and metastasis of colorectal carcinoma cells to the liver. Therefore, the 11-amino acid peptide chANG has both antiangiogenin and antiplasmin activity, and could be useful in the development of anticancer agents.

Topics: 3T3 Cells; Actins; alpha-2-Antiplasmin; Animals; Anticarcinogenic Agents; Binding Sites; Biotinylation; Cell Movement; Dose-Response Relationship, Drug; Enzyme-Linked Immunosorbent Assay; Fibrinolysin; Humans; Liver Neoplasms; Mice; Neoplasm Invasiveness; Neovascularization, Pathologic; Peptides; Protein Binding; Protein Structure, Tertiary; Ribonuclease, Pancreatic; Time Factors; Tumor Cells, Cultured

Using digoxigenin-labelled, synthetic oligonucleotide probe cocktails of angiogenin and bFGF genes, the expression of the two genes was observed by in situ hybridization in ten colonic adenocarcinomas, seven gastric adenocarcinomas, and four hepatocellular carcinomas. The angiogenin gene was expressed in eight of the ten cases of colonic adenocarcinoma and in all of the four cases where dysplastic glands were found. Angiogenin expression was evident in four of the seven cases of gastric adenocarcinoma. bFGF expression was detected in only five of the seven gastric carcinoma cases. The mRNAs for angiogenin and bFGF were mainly cytoplasmic in distribution and were only occasionally seen in the nuclei of the positive cells. Neither the angiogenin gene mRNA nor the bFGF mRNA was expressed in the four cases of hepatocellular carcinoma. It is postulated that the angiogenin gene may play an important role in angiogenesis in colonic adenocarcinomas; in gastric cancers, both angiogenin and bFGF were involved in this process. For hepatocellular carcinomas, neither angiogenin nor bFGF production appeared to be related to angiogenesis.

Topics: Adenocarcinoma; Angiogenesis Inducing Agents; Carcinoma, Hepatocellular; Colonic Neoplasms; Fibroblast Growth Factor 2; Gastrointestinal Neoplasms; Humans; In Situ Hybridization; Liver Neoplasms; Neoplasm Proteins; Neovascularization, Pathologic; Proteins; Ribonuclease, Pancreatic; RNA, Messenger; Stomach Neoplasms