Naim Alkhouri

Adipocyte Apoptosis, a Link between Obesity, Insulin Resistance, and Hepatic Steatosis

Adipocyte death has been reported in both obese humans and rodents. However, its role in metabolic disorders, including insulin resistance, hepatic steatosis, and inflammation associated with obesity has not been studied. We now show using real-time reverse transcription-PCR arrays that adipose tissue of obese mice display a pro-apoptotic phenotype. Moreover, caspase activation and adipocyte apoptosis were markedly increased in adipose tissue from both mice with diet-induced obesity and obese humans. These changes were associated with activation of both the extrinsic, death receptor-mediated, and intrinsic, mitochondrial-mediated pathways of apoptosis. Genetic inactivation of Bid, a key pro-apoptotic molecule that serves as a link between these two cell death pathways, significantly reduced caspase activation, adipocyte apoptosis, prevented adipose tissue macrophage infiltration, and protected against the development of systemic insulin resistance and hepatic steatosis independent of body weight. These data strongly suggest that adipocyte apoptosis is a key initial event that contributes to macrophage infiltration into adipose tissue, insulin resistance, and hepatic steatosis associated with obesity in both mice and humans. Inhibition of adipocyte apoptosis may be a new therapeutic strategy for the treatment of obesity-associated metabolic complications.

Lipotoxicity in Nonalcoholic Fatty Liver Disease: Not All Lipids Are Created Equal

Nonalcoholic fatty liver disease (NAFLD) is currently the most common form of chronic liver disease affecting both adults and children in the USA and many other parts of the world. NAFLD encompasses a wide spectrum of conditions associated with the overaccumulation of lipids in the liver, ranging from steatosis to nonalcoholic steatohepatitis, to cirrhosis and its feared complications of portal hypertension, liver failure and hepatocellular carcinoma. In this article, we will focus on the growing evidence linking changes in hepatic lipid metabolism and accumulation of specific lipid types in the liver with hepatocellular damage, inflammation and apoptosis, resulting in disease progression to the more serious forms of this condition.

Ultrasonographic quantitative estimation of hepatic steatosis in children With NAFLD.

Background and Aims: The diagnostic accuracy of hepatic ultrasonography (US) for detection and grading of hepatic steatosis in children with suspected nonalcoholic fatty liver disease (NAFLD) remains poorly characterized. The aim of this study was to prospectively evaluate the clinical utility of ultrasonographic quantification of hepatic steatosis. Patients and Methods: Our cohort consisted of 208 consecutive pediatric patients with biopsy-proven NAFLD. Hepatic US was performed within 1 month of the liver biopsy procedure. Steatosis identified by US was scored using a 0 to 3 scale based on echogenicity and visualization of vasculature, parenchyma, and diaphragm, and compared to histological features based on Brunts classification. Results: The median age at time of first visit was 10.8 years and 64% were boys. Sixty-nine percent had moderate to severe steatosis on histology. Ultrasonographic steatosis score (USS) had an excellent correlation with histological grade of steatosis (with a Spearmans coefficient of 0.80). The area under the receiver operating characteristic curve for ultrasonographic detection of moderate-to-severe steatosis was 0.87. The USS did not correlate significantly with inflammatory activity or fibrosis stage; however, there was significant correlation with the NAFLD activity score (NAS), albeit this was in large part the result of the strong correlation with the steatosis component of NAS. Serum alanine transaminase and aspartate transaminase were not associated with histological grade of steatosis and showed no correlation with USS. Conclusions: Our results, which represent the largest prospective pediatric study evaluating the role of hepatic US in children with biopsy-proven NAFLD, demonstrate the utility of this technique for noninvasive diagnosis and estimation of hepatic steatosis in children.

Neutrophil to lymphocyte ratio: a new marker for predicting steatohepatitis and fibrosis in patients with nonalcoholic fatty liver disease.

Nonalcoholic steatohepatitis (NASH), the most severe form of nonalcoholic fatty liver disease (NAFLD), is associated with inflammation and increased oxidative stress. The neutrophil/lymphocyte ratio (N/L) integrates information on the inflammatory milieu and physiological stress.

Apoptosis in nonalcoholic fatty liver disease: diagnostic and therapeutic implications

Pathological increases in cell death in the liver as well as in peripheral tissues has emerged as an important mechanism involved in the development and progression of nonalcoholic fatty liver disease (NAFLD). An increase in hepatocyte cell death by apoptosis is typically present in patients with NAFLD and in experimental models of steatohepatitis, while an increase in adipocyte cell death in visceral adipose tissue may be an important mechanism triggering insulin resistance and hepatic steatosis. The two fundamental pathways of apoptosis, the extrinsic (death receptor-mediated) and intrinsic (organelle-initiated) pathways, are both involved. This article summarizes the current knowledge related to the distinct molecular and biochemical pathways of cell death involved in NAFLD pathogenesis. In particular, it will highlight the efforts for the development of both novel diagnostic and therapeutic strategies based on this knowledge.

Serum Cytokeratin-18 Fragment Levels Are Useful Biomarkers for Nonalcoholic Steatohepatitis in Children

OBJECTIVES:Nonalcoholic steatohepatitis (NASH) is the most aggressive form of nonalcoholic fatty liver disease (NAFLD). Noninvasive methods to identify children with NASH are urgently needed. The aim of this study was to evaluate the use of plasma cytokeratin-18 (CK18) fragment levels, a marker of increased hepatocyte apoptosis, as a non-invasive biomarker for pediatric NASH.METHODS:Consecutive children with biopsy-proven NAFLD were included and blood samples were collected at the time of the biopsy. The diagnosis of NASH was based on Brunts criteria. Histological features were scored: steatosis (0–3), lobular inflammation (0–3), ballooning (0–2), and portal inflammation (0–2). NAFLD activity score was calculated (0–8) and fibrosis stage was scored (0–4). We measured plasma CK18 levels using the M30-Apoptosense enzyme-linked immunosorbent assay kit.RESULTS:A total of 201 subjects were included in the study. The mean age was 10.7±2.5 years and 37% were male. NASH was diagnosed in 140 patients with a mean NAFLD activity scoring of 4.4±1.3. CK18 levels were significantly higher in subjects with NASH compared with not NASH (322.1 U/l±104.8 vs. 164.2 U/l±62, respectively; P<0.001). The risk of having NASH on liver biopsy increased with increased CK18 levels (P<0.001). For every 10 U/l increase in CK18 levels, the likelihood of having NASH increased by 70% after adjusting for multiple confounders. The performance of CK18 level for the diagnosis of NASH was excellent with an area under the receiver operating characteristics curve of 0.933.CONCLUSIONS:CK18 is a promising non-invasive biomarker for NASH in children with fatty liver disease.

Nonalcoholic Fatty Liver Disease: A Challenge for Pediatricians

Nonalcoholic fatty liver disease (NAFLD) is the hepatic manifestation of the metabolic syndrome and is considered the most common form of chronic liver disease in children. Several factors contribute to NAFLD development, including race/ethnicity, genetic factors, environmental exposures, and alterations in the gut microbiome. The histologic spectrum of NAFLD ranges from simple steatosis to the more aggressive nonalcoholic steatohepatitis (NASH). Fibrosis and eventually cirrhosis can develop from NAFLD during childhood. Diagnosing advanced disease is challenging and may require a liver biopsy, highlighting the urgent need for reliable, noninvasive markers of disease severity. The mainstay of treatment for NAFLD remains lifestyle modifications and weight loss. Probiotics and ω-3 fatty acids may ameliorate disease progression. Recent data have suggested that vitamin E may be considered as a NASH-specific therapy in children, and there are several ongoing human studies evaluating different therapeutic targets for NAFLD. We provide an up-to-date review of the risk factors, diagnosis, and treatment to manage this common disease in children.

A combination of the pediatric NAFLD fibrosis index and enhanced liver fibrosis test identifies children with fibrosis.

BACKGROUND & AIMS Nonalcoholic fatty liver disease (NAFLD) encompasses diseases from simple steatosis, to steatohepatitis, to fibrosis, and cirrhosis. The pediatric NAFLD fibrosis index (PNFI) and the enhanced liver fibrosis (ELF) test are potential noninvasive markers for fibrosis. We prospectively evaluated the performance of PNFI and ELF in assessing fibrosis in children with biopsy-proven NAFLD. METHODS We analyzed 111 consecutive children with NAFLD. The stage of fibrosis was scored according to the Nonalcoholic Steatohepatitis Clinical Research Network. PNFI was calculated based on age, waist circumference, and levels of triglycerides. The ELF test was used to determine levels of hyaluronic acid, the amino-terminal propeptide of type III collagen, and tissue inhibitor of metalloproteinase-1. RESULTS Some degree of fibrosis was detected in 68.5% of patients (62 had stage 1, 5 had stage 2, and 9 had stage 3). PNFI and ELF test values was higher among patients with fibrosis (P < .001). The area under the receiver operating characteristic (ROC) curve for predicting fibrosis using the PNFI and ELF test was 0.761 and 0.924, respectively. The best performance was obtained by combining PNFI and ELF test with (area under the receiver operating characteristic curve = 0.944). The combined results from the PNFI and ELF test predicted the presence or absence of fibrosis in 86.4% of children with NAFLD. CONCLUSIONS In children with NAFLD, the combined results from the PNFI and ELF test can accurately assess the presence of liver fibrosis and identify patients that should be evaluated by liver biopsy.

Retinol-Binding Protein 4: A Promising Circulating Marker of Liver Damage in Pediatric Nonalcoholic Fatty Liver Disease

BACKGROUND & AIMS Noninvasive methods are needed to identify pediatric nonalcoholic fatty liver disease (NAFLD), the most frequent chronic liver disease in children and adolescents in industrialized countries. Retinol-binding protein 4 (RBP4) is an adipocytokine that has been associated with the pathogenesis of insulin resistance. We tested the serum levels of RBP4 to assess their associations with the metabolic profile and histologic features in a large well-characterized group of children with NAFLD. METHODS The study included 59 children with biopsy-proven NAFLD. Histologic analyses were performed by an experienced hepatopathologist; the NAFLD activity score and fibrosis score were calculated for each patient. RBP4 levels in serum samples were measured by an enzyme-linked immunosorbent assay analysis. Anthropometric, blood pressure, and metabolic profile analyses (including glucose tolerance, fasting glucose, insulin, and lipid panel tests) were performed on samples from all patients. RESULTS Decreasing levels of RBP4 were associated significantly with increasing levels of serum triglyceride. High levels of RBP4 were associated significantly with low necroinflammatory activity, a low NAFLD activity score, and a low fibrosis score. Furthermore, serum RBP4 levels decreased significantly as disease severity increased; there was a stepwise decrease in RBP4 from children with steatosis (3.8 mg/dL) to borderline nonalcoholic steatohepatitis (2.9 mg/dL) to definitive nonalcoholic steatohepatitis (1.9 mg/dL) (P < .0001). This association remained significant after adjusting for other relevant clinical variables. CONCLUSIONS Our study shows an inverse relationship between RBP4 levels and degree of liver damage. RBP4 therefore might be a potential novel noninvasive marker of severity of pediatric NAFLD.

Serum Retinol-binding Protein 4 Levels in Patients With Nonalcoholic Fatty Liver Disease

Background Nonalcoholic fatty liver disease (NAFLD), one of the most common forms of chronic liver disease, is closely associated with obesity and insulin resistance (IR). Recent studies suggest serum retinol-binding protein 4 (RBP4) plays a key role in the pathogenesis of IR. The aims of this study were to determine serum RBP4 levels in patients with biopsy proven NAFLD, and to correlate these levels with the metabolic profile and histologic features in this population. Methods Our cohort consisted of 51 consecutive patients undergoing liver biopsy for clinical suspicion of NAFLD. Patients were subsequently divided into 3 groups: simple steatosis (n=16), borderline nonalcoholic steatohepatitis (NASH) (n=2) and NASH (n=33). The stage of fibrosis was measured using a 4-point scale. RBP4 was measured in triplicates by a specific enzyme-linked immunosorbent assay. The degree of insulin resistance was determined by the homeostatic model assessment. Results Serum RBP4 levels did not correlate with body mass index, homeostatic model assessment, fasting glucose, or insulin levels in patients with simple steatosis and NASH. Moreover, RBP4 levels were lower in patients with NASH compared with those with simple steatosis (21.3 and 26.8 mg/L, respectively) although the difference did not reach statistical significance (P=0.21). A stepwise decrease in RBP4 levels from patients without fibrosis (27.9 mg/L) to patients with cirrhosis (14.1 mg/L) was noted (P=0.03). Conclusions Our study demonstrates that in adult patients with NAFLD, serum RBP4 levels do not correlate with body mass index or insulin resistance and identifies a novel association between serum RBP4 levels and hepatocellular injury in these patients.