Nonalcoholic fatty liver disease (NAFLD) is defined histologically by the accumulation of large droplets of fat, mostly in the form of triglyceride, in hepatocytes (steatosis); the proliferation of inflammatory cells surrounding ballooned injured hepatocytes in the liver parenchyma (steatohepatitis); and, in some patients, the subsequent development of significant bridging hepatic fibrosis and even cirrhosis. It has long been known that certain rare inborn errors of metabolism result in lipid deposition in the liver and that certain drugs that affect mitochondrial fatty acid oxidation and respiratory chain pathways can provoke hepatic steatosis and toxicity.1 Recently, as the epidemic of obesity has spread, an appreciation of the link between obesity and fatty liver has developed, and the importance of NAFLD in the spectrum of chronic liver disease has increased.
Overall, NAFLD has been reported to affect 10% to 24% of the total populations of various countries and up to 74% of obese people.2 NAFLD is now the most common cause of chronic liver disease in many countries and appears to potentiate liver damage induced by other agents, such as alcohol, drugs, industrial toxins, hepatitis C, and other viruses. It is now considered the most common cause of cryptogenic cirrhosis in adults; these patients constitute a large percentage of those undergoing liver transplantation.3
This article describes the increasing prevalence of NAFLD in overweight and obese children, the pathophysiology of steatohepatitis and subsequent fibrosis, and the available and potential therapies to treat this burgeoning longterm threat.
The lack of specific, sensitive, and noninvasive tests for NAFLD limits reliable detection of the disease. Often, the diagnosis is made presumptively when serum aminotransferase (aspartate aminotransferase, AST, and alanine aminotransferase, ALT) and gamma-glutamyl transpeptidase (GGT) levels are elevated in obese children with no other identifiable cause for liver disease, or when imaging studies, including ultrasonography or computed tomography, suggest the intrahepatic accumulation of fat. Conditions that often accompany NAFTLD but are not always present include acanthosis nigricans and hypertriglyceridemia. Fasting hyperinsulinemia and insulin resistance often are present in children with NAFLD and are strong predictors of the severity of steatosis and the development of fibrosis.4
Although recent studies using special magnetic resonance imaging techniques (proton nuclear magnetic resonance spectroscopy) show promise for noninvasive quantification of intrahepatic steatosis, they fail to indicate the degree of inflammation (steatohepatitis) and fibrosis.5 The gold standard for diagnosing NAFLD remains the liver biopsy.
Recent studies have estimated that 21.5 % of black children, 21.8% of Hispanic children, and 12.3% of non-Hispanic white children in the United States are either overweight or obese.6 In series of pediatric patients with NAFLD, most patients were obese, with a range of 1 14% to 192% of ideal weight for height and a body mass index (BMI) greater than the 95th percentile.7 However, gender and ethnicity appear to influence the occurrence of NAFLD in obese children. Males predominate, with a ratio of about 2: 1 in most series, and obese Hispanic children appear most prone to developing NAFLD, while obese black children are least susceptible.8 Between 10% and 25% of obese children have elevated aminotransferases; these findings are more prevalent in boys than girls and increase with the severity of obesity as measured by BMI.9
However, these biochemical abnormalities appear to correlate only with severe steatosis and underestimate the prevalence of NAFLD. Greater percentages of obese patients appear to have fatty livers by sonography, even some with normal liver enzymes.10
Although silent symptomatically, NAFLD may be associated with significant liver injury even in children. A recent study of 332 obese children ages 6 to 16 in Texas demonstrated that 15.7% had elevated aminotransferases greater than 1.5 times normal.11 Approximately 80% of those with elevated enzymes underwent liver biopsy, and 35 of those 40 children demonstrated some degree of fibrosis. Approximately 65% showed at least grade II fibrosis, and 5% showed signs of early cirrhosis.
Most studies of the prevalence of NAFLD in children suffer from selection bias. However, two recent studies looked at unselected populations. A study of 810 schoolchildren from northern Japan showed an overall prevalence of fatty liver by sonography of 2.6%. 12 Schwimmer and colleagues13 recently reported the population prevalence of fatty liver in 954 children ages 2 to 19 in San Diego County, California, who died of external causes (eg, accidents, homicide, suicide) within 48 hours of injury and were autopsied between 1993-2003. Fatty liver was defined as greater than or equal to 5% of hepatocytes with lipid accumulation. Fatty liver was present in 14% of all subjects and increased with increasing age. Boys were 67% more likely than girls to have a fatty liver, and the incidence in Hispanic children (19%) was higher than non-Hispanic white (10%) or black children (5%). Fatty liver was found in 35% of obese children. The overall prevalence of fatty liver in San Diego County adjusted for age and gender was calculated as 8.1%.13
The importance of the predilection to fatty liver in obese Hispanic adolescents is reinforced by data showing that Hispanic adults have an increased rate of liver-related morbidity and mortality. Also, cryptogenic cirrhosis is three times more prevalent in those of Hispanic descent than those of European descent.14
Although obesity is by far the most common association and risk factor for NAFLD, other conditions also predispose children to fatty liver. Patients with hypothalamic damage and hypopituitarism develop central obesity, insulin resistance, and, at times, hyperphagia.15 In children, this phenotype is seen most commonly in patients treated for craniopharyngioma. The acute development of NAFLD has been described after suprasellar tumor resection in previously nonobese children with subsequent development of cirrhosis and portal hypertension.16 Prolonged follow-up of such patients has revealed a high incidence of steatohepatitis and cirrhosis associated with excessive weight gain, impaired glucose tolerance, and dyslipidemia. Sidebar 1 (see page 292) summarizes risk factors for NAFLD in children.
Children with other syndromes characterized by hyperinsulinism and insulin resistance as a component also should be screened for NAFLD. These include patients with polycystic ovarian syndrome, Bardet-Biedl syndrome, Prader-Willi syndrome, and lipodystrophy.17 Genetic polymorphisms unrelated to carbohydrate and lipid metabolism also may play a role, especially in the progression from simple steatosis to steatohepatitis and fibrosis in obese patients. Heterozygosity for hereditary hemochromatosis and alpha- 1 -antitrypsin deficiency appear to predispose patients with NAFLD to inflammation and fibrosis.18 Genes controlling intrahepatic concentrations of cytochrome P-450 enzymes capable of generating oxygen-free radicals, the consumption of alcohol, and the adequacy of anti-oxidant defenses such as hepatic glutathione and vitamin E, beta-carotene, and vitamin C levels also may be involved.19 Finally, genes regulating body weight and controlling hormonal cycles of leptin and adiponectin likely play a role in the clinical diversity of NAFLD.20
A net retention of lipids within the hepatocytes is a prerequisite for the development of NAFLD. Fat accumulates in the liver as a result of an imbalance between the enzymatic systems that promote the uptake and synthesis of fatty acids within the liver and those that promote their oxidation and export from the liver. During the fed state, fatty acids are delivered to the liver and synthesized as a result of protein or carbohydrate excess. Two major pathways of fatty acid disposal balance fat deposition: mitochondrial B-oxidation, generating ATP and ketone bodies, and secretion of triglycerides into the blood as very low-density lipoproteins (VLDL). Disturbances in these processes can be inherited or acquired, resulting in the accumulation of triglycerides within the liver.
Insulin resistance appears to be an important component in the pathogenesis of NAFLD. Consistent with this hypothesis is the fact that mild insulin resistance is very common at the earliest stages of NAFLD, and more severe insulin resistance (Type 2 diabetes) correlates with more advanced stages of NAFLD. In a mouse model of insulin resistance where mice have been genetically manipulated to overexpress lipoprotein lipase in the liver which in turn inhibits hepatic insulin signaling, NAFLD inevitably develops.21 As a result of insulin resistance, the body's metabolic profile mimics the fasting state. Lipolysis is not suppressed, and the delivery of free fatty acids (FFA) to the liver is increased. Augmented uptake of FFA by the hepatocytes overwhelms the intramitochondrial B -oxidation pathway and leads to increased production of intrahepatic triglycerides, resulting in steatosis. Insulin resistance also results in hyperinsulinemia that increases the synthesis of fatty acids in hepatocytes by increasing glycolysis and that favors the accumulation of triglycerides by decreasing hepatic production of apolipoprotein B-IOO, a critical part of VLDL production.
Recent attention has focused on defects in lipoprotein secretion as a paradigm to explain intrahepatic steatosis. Microsomal triglyceride transfer protein (MTP) is present in small bowel and liver, where it plays an important role in lipoprotein assembly and hepatic lipid efflux.22 Mutations in the MTP gene in humans result in abetalipoproteinemia, a rare autosomal recessive disorder characterized by the absence of serum apoprotein B -containing lipoproteins and fatty liver.23 Liver-specific MTP knockout mice develop hepatic steatosis.24 Moreover, it has been shown that several steatogenic drugs inhibit not only intrahepatic mitochondrial B -oxidation but also MTP activity.25 The role of MTP in NAFLD is under active investigation.
The development of NAFLD in patients after resection of craniopharyngiomas suggests that the hypothalamus, which is intimately involved in appetite and weight regulation via the actions of leptin and insulin, may be involved in its pathogenesis. Leptin is secreted by adipose tissue and binds to hypothalamic receptors to reduce appetite and increase energy expenditure. Damage to the hypothalamus raises leptin and insulin levels, induces hyperphagia, and lowers metabolic rate. Leptin levels are increased markedly in most obese patients (leptin resistance), and elevated leptin recently has been found to be an independent predictor of the severity of hepatic steatosis in patients with NAFLD.26
If fat deposition is the "first hit," then most investigators believe that a "second hit" is necessary for hepatic steatosis to progress to inflammation and fibrosis.27 Excess fat deposition in the liver is associated with lipid peroxidation, producing products (4-hydroxynonal and malondialdehyde) that bind covalently to hepatic proteins and act as potent agents for neutrophil Chemotaxis, proinflammatory cytokines, and stimulation of collagen production by hepatic stellate cells. Increased delivery of FFA to the hepatocyte also stresses the intramitochondrial fatty acid oxidation system, as evidenced by the higher levels of ketone bodies found in fasting patients with NAFLD. Increased fatty acid oxidation in the mitochondria could lead to the generation of oxygen free radicals, which in turn damage hepatocytes and induce fibrogenesis via stimulation of cytokine production.28
Patients with steatohepatitis have increased expression of TNF-alpha mRNA in both the liver and adipose tissue compared with obese controls without steatohepatitis; this over-expression is correlated with histologic severity.29 Moreover, damage to the mitochondria themselves may result in damage to the B -oxidation pathway, leading to decreased metabolism of long-chain fatty acids to ketones, increased reconstitution of triglycerides deposited in the hepatocyte, and the generation of potentially toxic metabolites in the B -oxidation pathway that are known inhibitors of ATP production via the intramitochondrial respiratory chain.30
Recently, a novel theory based on data from a mouse model proposed a mechanical factor as a link between pure steatosis and steatohepatitis.31 In this model, fat-loaded hepatocytes cause mechanical compression of sinusoids with resulting pericentral ischemia. This, in turn, is a potent stimulus to attract inflammatory cells that elaborate cytokines leading to liver injury.
A liver filled with fat also appears to be more susceptible to toxins that ordinarily might not cause much inflammation. The genetically obese Zucker rat, when exposed to endotoxin, will show a marked increase in inflammation and hepatotoxicity compared with normal litter mate controls.32 Obesity also lowers the threshold of alcohol-related liver disease, resulting in a doubling of NAFLD and an increase in alcohol-related cirrhosis in obese versus lean alcohol abusers.33 Recent evidence suggests that steatosis and steatohepatitis are major independent risk factors for cirrhosis in patients infected with hepatitis C.34 NAFLD also appears to predispose patients with hereditary hemochromatosis and excess intrahepatic iron deposition to more severe liver disease. Affected patients have a significantly increased risk of cirrhosis compared with NAFLD patients who have a normal genotype.
Sidebar 2 (see page 293) lists criteria used to make a diagnosis of NAFLD in children. The presence of NAFLD, particularly in infants or young children, should prompt consideration of underlying inherited syndromes that make up a very small minority of cases. It is also important to remember that intrahepatic steatosis and steatohepatitis can be an accompanying feature of other primarily liver-based diseases, including Wilson's disease, alpha- 1 -antitrypsin deficiency, and hepatitis C.
A careful physical examination can provide some clues. There is no BMI above which NAFLD always occurs and under which it never appears. However, the higher the BMI, the more likely NAFLD and steatohepatitis with fibrosis will be present. Conversely, it cannot be stressed enough that NAFLD is not only a disease of obese children but may also be found in those with profound malnutrition, rapid weight loss (eg, anorexia nervosa, bariatric surgery, type 1 diabetes), and those taking certain medications. Acanthosis nigricans is observed in only approximately one-third of patients with NAFLD, but it is a strong predictor of the presence of insulin resistance.7 Hepatomegaly is found in 50% of cases but may be underestimated because of the difficulty in assessing liver size in obese patients. Splenomegaly is rare unless cirrhosis and portal hypertension have already supervened. Other stigmata of chronic liver disease, such as scleral icterus, jaundice, clubbing, spider angiomata, ascites, and caput medusae, are almost never seen.
Most patients come to medical attention because of elevated aminotransferases discovered serendipitously Abnormalities are usually in the range of two to five times greater than normal; often, ALT alone is elevated, and it almost always exceeds AST. The only exception is found in those patients who have already developed steatohepatitis and significant fibrosis, in whom AST can exceed ALT. A recent prospective study attempting to develop criteria for ALT and AST elevation predictive of steatohepatitis in children found that an AST or ALT greater than 200 IU/L or any elevation of AST or ALT for more than 6 months in a child with a BMI greater than the 95th percentile was always predictive of the presence of steatohepatitis. However, only 3% of the subjects in obese childhood population studies satisfied these criteria, suggesting that their sensitivity is low.35
GGT also frequently is increased mildly, whereas alkaline phosphatase, bilirubin, albumin, and prothrombin time are normal. Other laboratory abnormalities reflect the underlying metabolic state of insulin resistance, including hypertriglyceridemia, occasional hypercholesterolemia, and elevated fasting insulin levels accompanying a normal serum glucose. However, epidemiological studies suggest that NAFLD in some obese children may precede development of the metabolic syndrome associated with insulin resistance. In adults, up to one-third have diabetes at the time of the diagnosis of NAFLD, but this is uncommon in children.36
Approximately one-third of patients with NAFLD are noted to be positive for autoantibodies (anti-nuclear antibodies and/or anti-smooth muscle antibodies), but liver histology is sufficiently different to allow adequate distinction between NAFLD and autoimmune chronic active hepatitis.37 In some studies, measures of inflammation (C-reactive protein) and oxidative stress (eg, ferritin, erythrocyte glutathione peroxidase) were elevated in about half of those with NAFLD, and serum levels of antioxidants (eg, vitamin E, beta-carotene, vitamin C) were decreased.38*39
Abdominal sonography is positive in 50% of obese patients and supportive of the diagnosis of NAFLD but is neither sensitive nor specific.40 A finding of a homogenous brightly echogenic liver suggests diffuse fat deposition but also can reflect fibrosis. Ultrasound is particularly useful in a patient who has suggestive risk factors and supportive physical findings but normal AST and ALT and it is clearly more sensitive than aminotransferases in predicting hepatic steatosis. However, relying on ultrasound detection of fatty liver will give an incorrect diagnosis in about one-third of patients; studies of ultrasound compared with histologic findings showed a sensitivity of only 67% and a specificity of 77%.41
Other imaging modalities include computed tomography and magnetic resonance imaging. Hepatic magnetic resonance imaging modified to allow a very rapid sequence appears to be very sensitive at detecting intrahepatic fat in obese children.42 Quantitative intrahepatic fat evaluation can be done using this method, and preliminary studies suggest that a fat fraction greater than 18% is associated with elevated ALT in obese patients with steatosis. However, all of these imaging techniques lack the ability to detect steatohepatitis and fibrosis.
Liver biopsy remains the most sensitive and specific method to diagnose NAFLD. Histopathology is characterized by macrovesicular steatosis defined by large droplet fat inside the hepatocyte displacing the nucleus eccentrically. Other features include ballooning degeneration of hepatocytes, perisinusoidal fibrosis, portal inflammation, and even portal fibrosis. In the most comprehensive pathologic study of NAFLD in children, advanced fibrosis was present in 8% of the 100 biopsies reviewed.43 Some investigators have begun to make a distinction between steatohepatitis characterized by steatosis and perisinusoidal fibrosis (Type 1), and those patients with steatosis and portal inflammation and fibrosis (Type 2). Type 2 more often is found in males and among Hispanics, Native Americans, and Asians with NAFLD.
To date, the only modality shown to decrease liver enzymes consistently in patients with NAFLD is weight loss via dieting and exercise. In animals, diets high in sucrose or fat content are more likely to cause insulin resistance and hepatic steatosis than equicaloric diets enriched with protein.44 Long-term studies testing this intervention on large numbers of obese children with NAFLD have not yet been published.
Insulin-sensitizing medications such as metformin and thiazolidinediones have shown promise. Treatment with metformin improved liver enzymes and hepatic steatosis in small groups of obese adult and pediatric patients with biopsyproven steatohepatitis.45 A recently published phase II open-label clinical trial of metformin (500 mg twice daily for 24 weeks) in 10 obese children with nondiabetic NAFLD showed that aminotransferases normalized in 50%. 46 There also were significant improvements in measures of insulin resistance and liver fat quantitated via magnetic resonance imaging.46 These changes occurred in the absence of significant weight loss.
Troglitazone, another insulin-sensitizing agent, significantly improved hepatic histology and liver enzymes in a small series of adult patients with NAFLD, but more widespread use of this drug has been hampered by rare reports of fatal hepatotoxicity.47 Currently, a multicenter randomized controlled trial of the efficacy and safety of metformin, sponsored by the National Institutes of Health, is being conducted in children with NAFLD.
Because the second-hit hypothesis implicates oxidant stress and the generation of oxygen-free radicals via lipid peroxidation, therapeutic trials of antioxidants have been conducted. Large doses of vitamin E improved liver enzyme abnormalities in certain animal models of NAFLD.48 An open-labeled trial of 400 to 1,200 IU/day of vitamin E in 1 1 obese children with elevated aminotransferases for greater than 3 months and an echogenic liver showed significant enzyme reductions but no change in liver echogenicity after 4 to 10 months of treatment.49 Enzyme reductions could not be ascribed to weight loss or to vitamin E deficiency prior to treatment. The antioxidant betaine also appeared efficacious in limited trials.50
Small pilot studies initially suggested that ursodeoxycholic acid improved liver enzymes in patients with NAFLD. However, a recently completed large, multicenter, double-blind, placebo-controlled trial of ursodeoxycholic acid in obese adults with NAFLD failed to show efficacy over and above the effect of diet and weight loss.51 Very preliminary reports of promising effects of 3hydroxy-3 -methyl glutaryl coenzyme A reductase inhibitors (statins) have been published in abstract form, but currently, the routine use of statins to treat NAFLD is not recommended.52
NAFLD likely is the most common liver disease in children and is responsible for significant progression to cirrhosis, portal hypertension, and the need for liver transplantation in adults and even in some adolescents. Early diagnosis and lifestyle interventions appear to be our best hope for controlling progression of disease. The pediatrician is responsible for screening all obese children with measurement of aminotransferases. Those with elevated enzymes (particularly ALT) for longer than 3 months, in the absence of markers of hepatitis B or C, autoimmune chronic active hepatitis, Wilson's disease, hemochromatosis, or alpha- 1 -antitrypsin deficiency, should follow up with an abdominal ultrasound. In patients with a BMI in the morbidly obese range, an ultrasound to search for a diffusely echogenic liver should be performed even if the liver enzymes are normal. Findings suggestive of NAFLD should prompt the institution of appropriate dietary and exercise regimens. If these are unsuccessful after a 3-month trial, the patient should be referred to a pediatric gastroenterologist and hepatologist for further work-up and treatment, preferably in the context of a controlled therapeutic trial. Only by aggressively engaging this current epidemic will we be able to decrease the mounting human cost of NAFLD.
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