Correlation of MicroRNA-16, MicroRNA-21 and MicroRNA-101 Expression with Cyclooxygenase-2 Expression and Angiogenic Factors in Cirrhotic and Noncirrhotic Human Hepatocellular Carcinoma

BACKGROUND: Hepatocellular carcinoma (HCC) is a classical example of inflammation-linked cancer and is characterized by hypervascularity suggesting rich angiogenesis. Cycloxygenase-2 (COX-2) is a potent mediator of inflammation and is considered to upregulate angiogenesis. The aims of the study are (1) to analyze expression of Cox-2 mRNA, Cox-2 protein, miR-16, miR-21 and miR-101 in HCC and adjacent liver parenchyma in cirrhotic and noncirrhotic liver, (2) to investigate the relation between COX-2 expression, miR-21 expression and angiogenic factors in these tissues and (3) to investigate the association between miR-16 and miR-101 and COX-2 expression. METHODS: Tissue samples of HCC and adjacent liver parenchyma of 21 noncirrhotic livers and 20 cirrhotic livers were analyzed for COX-2 expression at the mRNA level (qRT-PCR) and at the protein level by Western blot and immunohistochemistry. Gene expression of VEGFA, VEGFR1, VEGFR2, Ang-1, Ang-2 and Tie-2 were correlated with COX-2 levels. miR-16, miR-21 and miR-101 gene expression levels were quantified in HCC tumor tissue. RESULTS: COX-2 mRNA and protein levels were lower in HCC as compared to adjacent liver parenchyma both in cirrhotic and noncirrhotic liver. COX-2 protein localized mainly in vascular and sinusoidal endothelial cells and in Kupffer cells. At the mRNA level but not at the protein level, COX-2 correlated with mRNA levels of angiogenic factors VEGFR1, Ang-1, and Tie2. miR-21 expression was higher in cirrhotic tissues versus noncirrhotic tissues. MiR-101 expression was lower in cirrhotic versus noncirrhotic adjacent liver parenchyma. None of the miRNAs correlelated with COX-2 expression. miR-21 correlated negatively with Tie-2 receptor in adjacent liver parenchyma. CONCLUSIONS: In human HCC, COX-2 mRNA but not COX-2 protein levels are associated with expression levels of angiogenic factors. MiR-21 levels are not associated with angiogenic molecules. MiR-16 and miR-101 levels do not correlate with COX-2 mRNA and protein levels

Molecular Characterization of the Vascular Features of Focal Nodular Hyperplasia and Hepatocellular Adenoma: A Role for Angiopoietin-1

Focal nodular hyperplasia (FNH) and hepatocellular adenoma (HCA) are two hepatic nodular lesions of different etiologies. FNH, a polyclonal lesion, is assumed to be a regenerative reaction following a vascular injury, whereas HCA is a monoclonal, benign neoplastic lesion. In addition to features that are predominantly found in either FNH or HCA (e.g., dystrophic vessels in FNH and single arteries in HCA), FNH and HCA share morphological vascular abnormalities such as dilated sinusoids. We hypothesized that these anomalous vascular features are associated with altered expression of growth factors involved in vascular remodeling. This was based on reports of morphologically abnormal hepatic vasculature and nodular lesions in transgenic models of hepatocytic overexpression of angiopoietin-1 (Ang-1), a member of the angiopoietin family, which is crucially involved in vascular morphogenesis and homeostasis. We investigated gene and protein expression of members of the angiopoietin system and vascular endothelial growth factor A (VEGF-A) and its receptors in 9 FNH samples, 13 HCA samples, and 9 histologically normal livers. In comparison with normal samples, a significant increase in Ang-1 was found in FNH (

Hepatocellular Carcinomas in Cirrhotic and Noncirrhotic Human Livers Share Angiogenic Characteristics

The antiangiogenic drug sorafenib has been shown to be an effective treatment for hepatocellular carcinoma (HCC) in patients with liver cirrhosis. It might also be effective in noncirrhotic HCC provided that the angiogenic properties of both tumor types are comparable. The aim of this study is to compare endothelial cell dynamics, microvessel density (MVD), and vessel maturation as indirect markers of angiogenesis in human HCC in cirrhotic and noncirrhotic livers. In a tertiary care setting, 70 consecutive HCC tumors were analyzed for endothelial cell dynamics. CD34 was applied to identify tumor microvessels, double immunolabeling Ki67/CD34 and activated caspase-3/CD34 to assess endothelial cell proliferation and apoptosis, and alpha-smooth muscle actin/CD34 for pericyte coverage. These characteristics were compared in cirrhotic (n = 33) and noncirrhotic HCCs (n = 37). Microvessel density was correlated with radiological signs of hypervascularity as obtained with dynamic four-phase CT scans during the arterial and portal phase of contrast enhancement. Microvessels in cirrhotic and noncirrhotic HCC were mainly mature. In both groups endothelial cell turnover was low and MVD was not different. There was no correlation between MVD and venous invasion, tumor size, and turnover of tumor cells or endothelial cells. MVD was negatively correlated with contrast washout in the portal venous phase of CT scanning. In transplanted patients, MVD was not correlated with survival, whereas in patients after liver resection a high MVD was associated with a better prognosis. Angiogenic characteristics of HCC in cirrhotic and noncirrhotic livers have a remarkable similarity