Fabio Marra

Acute viral hepatitis increases liver stiffness values measured by transient elastography

Liver tissue alterations other than fibrosis may have an impact on liver stiffness measurement. In this study we evaluated 18 patients without a previous clinical history of liver disease, consecutively admitted for acute viral hepatitis. In each patient, aminotransferase determination and liver stiffness measurement were performed on the same study day, at 3 different points: (1) peak increase in aminotransferase; (2) aminotransferase 50% or less of the peak; (3) aminotransferase levels ≤2× the upper limit of normal. In all patients, the degree of liver stiffness at the time of the peak increase in aminotransferases exceeded the cutoff values proposed for the prediction of significant fibrosis or cirrhosis. A progressive significant reduction in liver stiffness values was observed (P < 0.0001) in the follow‐up period in parallel with the reduction of aminotransferase levels (P < 0.0001). Moreover, a statistically significant, positive correlation between aminotransferases and liver stiffness measurement (LSM) at the onset of acute viral hepatitis was found (r = 0.53, P = 0.02 and r = 0.51, P = 0.03 for alanine aminotransferase and aspartate aminotransferase, respectively). In conclusion, the extent of necroinflammatory activity needs to be carefully considered in future studies aimed at further validating transient elastography, particularly in patients with absent or low‐stage liver fibrosis (in other words, F0‐F2 METAVIR). LSM does not represent a reliable instrument to detect the presence of advanced fibrosis and cirrhosis in patients presenting with a clinical picture of acute hepatitis. (HEPATOLOGY 2007.)

Liver stiffness measurement predicts severe portal hypertension in patients with HCV‐related cirrhosis

Measurement of hepatic venous pressure gradient (HVPG) is a standard method for the assessment of portal pressure and correlates with the occurrence of its complications. Liver stiffness measurement (LSM) has been proposed as a noninvasive technique for the prediction of the complications of cirrhosis. In this study, we evaluated the ability of LSM to predict severe portal hypertension compared with that of HVPG in 61 consecutive patients with HCV‐related chronic liver disease. A strong relationship between LSM and HVPG measurements was found in the overall population (r = 0.81, P < 0.0001). However, although the correlation was excellent for HVPG values less than 10 or 12 mm Hg (r = 0.81, P = 0.0003 and r = 0.91, P < 0.0001, respectively), linear regression analysis was not optimal for HVPG values ≥10 mm Hg (r2 = 0.35, P < 0.0001) or ≥12 mm Hg (r2 = 0.17, P = 0.02). The AUROC for the prediction of HVPG ≥10 and ≥12 mm Hg were 0.99 and 0.92, respectively and at LSM cutoff values of 13.6 kPa and 17.6 kPa, sensitivity was 97% and 94%, respectively. In patients with cirrhosis, LSM positively correlated with the presence of esophageal varices (P = 0.002), although no correlation between LSM and esophageal varices size was detected. The area under the ROC for the prediction of EV was 0.76 and at a LSM cutoff value of 17.6 kPa sensitivity was 90%. Conclusion: LSM represents a non‐invasive tool for the identification of chronic liver disease patients with clinically significant or severe portal hypertension and could be employed for screening patients to be subjected to standard investigations including upper GI endoscopy and hemodynamic studies. (HEPATOLOGY 2007;45:1290–1297.)

An Alternatively Spliced Variant of CXCR3 Mediates the Inhibition of Endothelial Cell Growth Induced by IP-10, Mig, and I-TAC, and Acts as Functional Receptor for Platelet Factor 4

The chemokines CXCL9/Mig, CXCL10/IP-10, and CXCL11/I-TAC regulate lymphocyte chemotaxis, mediate vascular pericyte proliferation, and act as angiostatic agents, thus inhibiting tumor growth. These multiple activities are apparently mediated by a unique G protein–coupled receptor, termed CXCR3. The chemokine CXCL4/PF4 shares several activities with CXCL9, CXCL10, and CXCL11, including a powerful angiostatic effect, but its specific receptor is still unknown. Here, we describe a distinct, previously unrecognized receptor named CXCR3-B, derived from an alternative splicing of the CXCR3 gene that mediates the angiostatic activity of CXCR3 ligands and also acts as functional receptor for CXCL4. Human microvascular endothelial cell line-1 (HMEC-1), transfected with either the known CXCR3 (renamed CXCR3-A) or CXCR3-B, bound CXCL9, CXCL10, and CXCL11, whereas CXCL4 showed high affinity only for CXCR3-B. Overexpression of CXCR3-A induced an increase of survival, whereas overexpression of CXCR3-B dramatically reduced DNA synthesis and up-regulated apoptotic HMEC-1 death through activation of distinct signal transduction pathways. Remarkably, primary cultures of human microvascular endothelial cells, whose growth is inhibited by CXCL9, CXCL10, CXCL11, and CXCL4, expressed CXCR3-B, but not CXCR3-A. Finally, monoclonal antibodies raised to selectively recognize CXCR3-B reacted with endothelial cells from neoplastic tissues, providing evidence that CXCR3-B is also expressed in vivo and may account for the angiostatic effects of CXC chemokines.

Adipokines in liver diseases

Adipokines are polypeptides secreted in the adipose tissue in a regulated manner. While some of these molecules are expressed only by adipocytes, resident and infiltrating macrophages and components of the vascular stroma markedly contribute to expression of other adipokines. As a result, adipose tissue inflammation is associated with a modification in the pattern of adipokine secretion. Leptin, adiponectin, and resistin are the best‐studied molecules in this class, but cytokines such as tumor necrosis factor or interleukin‐6 are also secreted at high levels by the adipose tissue. Several other molecules have been recently identified and are actively investigated. Adipokines interfere with hepatic injury associated with fatty infiltration, differentially modulating steatosis, inflammation, and fibrosis. Several studies have investigated plasma levels of adiponectin in patients with nonalcoholic fatty liver disease, to establish correlations with the underlying state of insulin resistance and with the type and severity of hepatic damage. Hepatitis C is another disease where adipokines may represent a link between viral infection, steatosis, and metabolic disturbances. Identification of the mediators secreted by expanded adipose tissue and their pathogenic role is pivotal in consideration of the alarming increase in the prevalence of obesity and of the detrimental role that this condition exerts on the course of liver diseases. (HEPATOLOGY 2009.)

Concomitant Activation of the JAK/STAT, PI3K/AKT, and ERK Signaling Is Involved in Leptin-Mediated Promotion of Invasion and Migration of Hepatocellular Carcinoma Cells

Various epidemiologic studies have shown that obesity is associated with hepatocellular carcinoma. Leptin, the key player in the regulation of energy balance and body weight control, also acts as a growth factor on certain organs in both normal and disease states. It is plausible that leptin acts to promote hepatocellular carcinogenesis directly affecting malignant properties of liver cancer cells. However, a direct role for leptin in hepatocellular carcinoma has not been shown. In this study, we analyzed the role of leptin and the mechanism(s) underlying its action in hepatocellular carcinoma cells, which express both short and long isoforms of leptin receptors. Treatment with leptin resulted in increased proliferation of both HepG2 and Huh7 cells and involves activation of signal transducers and activators of transcription 3 (STAT3), AKT, and extracellular signal-regulated kinase (ERK) signaling pathways. Leptin-induced phosphorylation of ERK and AKT was dependent on Janus-activated kinase (JAK)/STAT activation. Intriguingly, we also found that leptin potently induces invasion of hepatocellular carcinoma cells in Matrigel invasion and electric cell-substrate impedance-sensing assays. Leptin-stimulated invasion was effectively blocked by pharmacologic inhibitors of JAK/STAT and, to a lesser extent, by ERK and phosphatidylinositol 3-kinase (PI3K) inhibition. Importantly, leptin also induced the migration of both HepG2 and Huh7 cells on fibronectin matrix. Inhibition of JAK/STAT, ERK, and PI3K activation using pharmacologic inhibitors effectively blocked leptin-induced migration of HepG2 and Huh7 cells. Taken together, these data indicate that leptin promotes hepatocellular carcinoma growth, invasiveness, and migration and implicate the JAK/STAT pathway as a critical mediator of leptin action. Our findings have potential clinical implications for hepatocellular carcinoma progression in obese patients.

Molecular basis and mechanisms of progression of non-alcoholic steatohepatitis

Non-alcoholic steatohepatitis (NASH), a cause of cirrhosis and hepatocellular carcinoma, is characterized by fatty infiltration of the liver, inflammation, hepatocellular damage and fibrosis. Progress has been made in understanding the molecular and cellular mechanisms implicated in the pathogenesis of this condition, therefore, we here review recent developments regarding the basic mechanisms of NASH development. Accumulation of triglycerides in the hepatocytes is the result of increased inflow of free fatty acids and de novo lipogenesis. Steatosis leads to lipotoxicity, which causes apoptosis, necrosis, generation of oxidative stress and inflammation. The resulting chronic injury activates a fibrogenic response that leads eventually to end-stage liver disease. A better understanding of these mechanisms is crucial for the design of novel diagnostic and therapeutic strategies.

Genome-wide meta-analyses identify three loci associated with primary biliary cirrhosis

A genome-wide association screen for primary biliary cirrhosis risk alleles was performed in an Italian cohort. The results from the Italian cohort replicated IL12A and IL12RB associations, and a combined meta-analysis using a Canadian dataset identified newly associated loci at SPIB (P = 7.9 × 10−11, odds ratio (OR) = 1.46), IRF5-TNPO3 (P = 2.8 × 10−10, OR = 1.63) and 17q12-21 (P = 1.7 × 10−10, OR = 1.38).

HNE interacts directly with JNK isoforms in human hepatic stellate cells.

4-Hydroxy-2,3-nonenal (HNE) is an aldehydic end product of lipid peroxidation which has been detected in vivo in clinical and experimental conditions of chronic liver damage. HNE has been shown to stimulate procollagen type I gene expression and synthesis in human hepatic stellate cells (hHSC) which are known to play a key role in liver fibrosis. In this study we investigated the molecular mechanisms underlying HNE actions in cultured hHSC. HNE, at doses compatible with those detected in vivo, lead to an early generation of nuclear HNE-protein adducts of 46, 54, and 66 kD, respectively, as revealed by using a monoclonal antibody specific for HNE-histidine adducts. This observation is related to the lack of crucial HNE-metabolizing enzymatic activities in hHSC. Kinetics of appearance of these nuclear adducts suggested translocation of cytosolic proteins. The p46 and p54 isoforms of c-Jun amino-terminal kinase (JNKs) were identified as HNE targets and were activated by this aldehyde. A biphasic increase in AP-1 DNA binding activity, associated with increased mRNA levels of c-jun, was also observed in response to HNE. HNE did not affect the Ras/ERK pathway, c-fos expression, DNA synthesis, or NF-kappaB binding. This study identifies a novel mechanism linking oxidative stress to nuclear signaling in hHSC. This mechanism is not based on redox sensors and is stimulated by concentrations of HNE compatible with those detected in vivo, and thus may be relevant during chronic liver diseases.

Increased risk of cardiovascular disease in non-alcoholic fatty liver disease: causal effect or epiphenomenon?

Non-alcoholic fatty liver disease (NAFLD), comprising a spectrum of conditions ranging from pure steatosis to steatohepatitis and cirrhosis, has reached epidemic proportions and represents the most common cause of chronic liver disease in the community. The prevalence of NAFLD has been estimated to be between 20% and 30% in the general population, but this value is much higher (∼70–80%) in type 2 diabetic patients, who are also at higher risk of developing advanced fibrosis and cirrhosis. Increasing recognition of the importance of NAFLD and its strong relationship with the metabolic syndrome has stimulated an interest in the possible role of NAFLD in the development of cardiovascular disease (CVD). Several epidemiological studies indicate that NAFLD, especially in its more severe forms, is linked to an increased risk of CVD, independently of underlying cardiometabolic risk factors. This suggests that NAFLD is not merely a marker of CVD, but may also be actively involved in its pathogenesis. The possible molecular mediators linking NAFLD and CVD include the release of pro-atherogenic factors from the liver (C-reactive protein, fibrinogen, plasminogen activator inhibitor-1 and other inflammatory cytokines) as well as the contribution of NAFLD per se to whole-body insulin resistance and atherogenic dyslipidemia, in turn favouring CVD progression. The clinical impact of NAFLD on CVD risk deserves particular attention in view of the implications for screening and surveillance strategies in the growing number of patients with NAFLD.

Upregulation of proinflammatory and proangiogenic cytokines by leptin in human hepatic stellate cells

Leptin upregulates collagen expression in hepatic stellate cells (HSCs), but the possible modulation of other actions has not been elucidated. The aim of this study was to investigate the expression and function of leptin receptors (ObR) in human HSCs and the biological actions regulated by leptin. Exposure of HSCs to leptin resulted in upregulation of monocyte chemoattractant protein 1 (MCP‐1) expression. Leptin also increased gene expression of the proangiogenic cytokines vascular endothelial growth factor (VEGF) and angiopoietin‐1, and VEGF was also upregulated at the protein level. Activated HSCs express ObRb and possibly other ObR isoforms. Exposure to leptin increased the tyrosine kinase activity of ObR immunoprecipitates and resulted in activation of signal transducer and activator of transcription 3. Several signaling pathways were activated by leptin in HSCs, including extracellular‐signal–regulated kinase, Akt, and nuclear factor κB, the latter being relevant for chemokine expression. Leptin also increased the abundance of hypoxia‐inducible factor 1α, which regulates angiogenic gene expression, in an extracellular‐signal–regulated kinase– and phoshatidylinositol 3‐kinase–dependent fashion. In vivo, leptin administration induced higher MCP‐1 expression and more severe inflammation in mice after acute liver injury. Conversely, in leptin‐deficient mice, the increase in MCP‐1 messenger RNA and mononuclear infiltration was less marked than in wild‐type littermates. Finally, ObR expression colocalized with VEGF and α‐smooth muscle actin after induction of fibrosis in rats. In conclusion, ObR activation in HSCs leads to increased expression of proinflammatory and proangiogenic cytokines, indicating a complex role for leptin in the regulation of the liver wound‐healing response.(HEPATOLOGY 2005;42:1339–1348.)