Prevalence and Demographic Predictors

Despite several recent advances, accurate epidemiologic data are lacking because of a deficiency in population-based studies and reliable noninvasive screening tools. The prevalence of NAFLD in children is affected by many factors, and it is determined by a complex interaction of genetic and environment influences, and is therefore difficult to define. In general, however, the risk of liver disease increases with the weight of the patient. The increasing prevalence of childhood obesity in the United States is alarming, affecting 15% of children between 6 and 19 years of age, with an additional 30% considered overweight. A large prospective study has shown that up to 50% of severely obese children have associated metabolic syndrome characterized by hypertension, high serum triglyceride levels, low high-density lipoprotein (HDL) levels, and impaired glucose tolerance. A number of studies in children have used surrogate markers of NAFLD (elevated serum liver enzymes and/or liver ultrasound, or both) to evaluate the prevalence in the pediatric population. Both, American and Asian surveys report that approximately 3% of adolescents have elevated serum alanine aminotransferase (ALT) levels. With use of liver ultrasound, in a large study of children 4 to 12 years of age in Japan that included more than 800 children, the prevalence of NAFLD was 2.6%, with a strong correlation with other measurements of obesity. Several studies in various countries have selectively examined the prevalence of NAFLD in groups of obese children and found that it may range from 23% to up to 77%. Noninvasive techniques were used for diagnosis in these studies; thus, none was able to determine the proportion of isolated hepatic steatosis versus more advanced forms of the disease including NASH or cirrhosis. A large autopsy study found that 9.6% of American population aged 2 to 19 years have NAFLD, and that figure increased to 38% among those who were obese.

Affected adults generally present between the fourth and sixth decades of life and are more frequently women (50% to 80%). Children most commonly present in the pubertal age group and have a male predominance, with a higher incidence in children of Hispanic origin. Several studies have hypothesized that hormonal changes during puberty are associated with increased serum insulin levels and insulin resistance, especially in boys, and thus a propensity for accumulation of fat in the liver. Estrogens seem to be protective through their effect on reduction of cell death, lipid peroxidation, inflammation, and fibrosis, major mechanisms responsible for the progression of the disease. Several differences in ethnic predisposition to NAFLD and NASH have been reported. The highest rates of NAFLD and signs of liver damage on histology (higher grades of ballooning and Mallory bodies) are found in Mexican Americans as well as Asian Indians and Native Americans, probably because of higher rates of insulin resistance and increased visceral adiposity at an equivalent body mass index (BMI). African American patients have lower rates of NAFLD, NASH, and less severe fibrosis, suggesting a protective genetic or metabolic effect in this group. These differences may also be influenced by environmental factors including the type of diet, exercise choice, socioeconomic status, and living location.

Natural History

Several long-term longitudinal studies in both the United States and Europe have examined the outcome and prognosis of adult patients with NAFLD. Currently, there is only one available study with these characteristics addressing the course and natural history of the disease in children. At the time of initial biopsy, a substantial number of adult patients with NASH may have advanced hepatic fibrosis, and one in 10 may show well-established cirrhosis. In addition, it is now recognized that a large proportion of adult patients with cryptogenic cirrhosis have burned-out NASH. Available data suggest that the natural history of NAFLD seems to be determined by the severity of the histologic damage. A large study of 106 adult patients with pure steatosis without inflammation showed a benign clinical course. Only one patient developed cirrhosis over a median follow-up period of 9 years. On the other hand, patients with NASH are believed to be at increased risk of advanced disease. Progression of fibrosis was seen in one-third to one-half of patients over a 3- to 5-year follow-up period and cirrhosis, and its complications have been shown to occur in more than 9% to 20% of patients over the same period. It is important to note that fibrosis may also regress in a number of patients. In addition, overall and liver-related mortality was also significantly higher in NAFLD patients than in the general population.

In the pediatric population, cross-sectional studies have described cirrhotic stage disease in children at diagnosis, and there are other reported cases of children with NAFLD who developed cirrhosis in early adulthood. A recent study examined the long-term prognosis of children with NAFLD and compared their survival with the expected survival of the general population. Sixty-six children with NAFLD with a mean age 14 years were followed for up to 20 years, a total of 409.6 person-years of follow-up. The metabolic syndrome was present in 19 children (29%) at the time of NAFLD diagnosis, with 55 (83%) presenting with at least one feature of the metabolic syndrome including obesity, hypertension, dyslipidemia, and/or hyperglycemia. In that series, four children with baseline normal fasting glucose levels developed type 2 diabetes 4 to11 years after NAFLD diagnosis. A total of 13 liver biopsies were obtained from five patients over a mean of about 5 years showing progression of fibrosis stage in four children. During follow-up, two children died and two underwent liver transplantation for decompensated cirrhosis. The observed survival that was free of liver transplantation was significantly shorter in the NAFLD cohort as compared to the expected survival in the general U.S. population of the same age and sex, with a standardized mortality ratio of 13.6 (95% confidence interval [CI] 3.8–34.8). NAFLD recurred in the allograft in the two cases transplanted, with one case progressing to cirrhosis and requiring re-transplantation. This study demonstrated for the first time that children with NAFLD might develop end-stage liver disease with the consequent need for liver transplantation during adolescence or early adulthood.

Not only has NASH-related cirrhosis been a growing indication for liver transplantation, but the disease has been shown to recur after transplantation in some reports. Finally, growing epidemiologic data suggest that as in adults, NAFLD in children may be associated not only with liver-related morbidity, but also with an increased risk of obesity-related metabolic complications as well as being an independent risk factor for cardiovascular disease.

Clinical Presentation

Most patients with NAFLD are asymptomatic, and the liver disease is often discovered incidentally, when laboratory examination shows elevated liver enzyme levels. In adults, it is the most common cause of unexplained persistent elevation of transaminases after viral hepatitis and other chronic liver diseases have been excluded, but this has not been carefully examined in children. The most common symptoms at presentation in children are fatigue and right upper quadrant or diffuse abdominal discomfort. Hepatomegaly may be found on clinical examination in up to 50%, and acanthosis nigricans, a cutaneous marker of insulin resistance, has been described in about 30%. Children with NAFLD are usually obese and have associated features of metabolic syndrome: insulin resistance in most patients, impaired glucose tolerance (10%), type 2 diabetes (2%), and variable incidence of hyperlipidemia and hypertension at diagnosis. When cirrhosis appears, stigmata of chronic liver disease, such as spider angiomata, ascites, splenomegaly, hard liver border, palmar erythema, or asterixis, can be present.

Diagnosis

The diagnosis of pediatric NAFLD is commonly made as a result of additional evaluation for elevated serum aminotransferases found during a routine check-up. NAFLD can also be diagnosed in children by ultrasound or less frequently by liver biopsy. Many centers have adopted a screening program for NAFLD in high-risk subjects, in particular in those presenting features of the metabolic syndrome. Liver biopsy, the current gold standard for the diagnosis of NAFLD, is the only way to distinguish between NASH and hepatic steatosis, determine the severity of liver damage and the presence and extent of fibrosis, and to rule out other diagnoses such as auto­immune hepatitis. However, routine noninvasive evaluation (biochemical parameters, imaging tests, and serum biomarkers) should be used as the first step to confirm the diagnosis of fatty liver disease, especially in the typical patient with characteristic features of metabolic syndrome.

Liver Biopsy

Liver biopsy remains the “the gold standard” for establishing the diagnosis of NAFLD and grading and staging the severity; in other words, distinguishing steatosis from steatohepatitis, and assessing the degree of fibrosis. Moreover, it is helpful in ruling out alternate causes resulting in hepatic steatosis, in particular chronic hepatitis C infection, Wilson disease, autoimmune hepatitis, and other metabolic liver disorders. In addition, histology permits the monitoring of disease progression and the response to therapy, because liver enzymes may decrease during the course of the disease regardless of whether fibrosis progresses or improves. A central limitation for the use of liver biopsy is its invasiveness and the potential association with significant complications such as bleeding and pain. The histologic diagnosis of NASH in pediatric cases may also be a challenging diagnosis, as the features found on liver biopsy in children often differ from those commonly seen in adults. The typical adult pattern (termed NASH type 1) is characterized by the presence of steatosis (mainly macrovesicular) with ballooning degeneration and/or perisinusoidal fibrosis (zone 3 lobular involvement), with the portal tracts being relatively spared. Pediatric type NASH (NASH type 2) is described as the presence of steatosis along with portal inflammation and/or fibrosis in the absence of ballooning degeneration and perisinusoidal fibrosis. However, a large proportion of patients may have overlapping features of both type 1 and type 2 NASH ( Table 74-1 ). Finally, it remains to be studied whether patients with the pediatric pattern differ in natural history, etiopathogenesis, prognosis, or response to treatments compared to patients with adult type or those with overlapping features.

Routine Laboratory Tests and Novel Biomarkers

Circulating biomarkers include indirect markers that are based on the algorithmic evaluation of commonly observed multiparametric liver functional alterations (i.e., aminotransferase levels), and direct markers reflecting the extracellular matrix remodeling and/or the acquisition of a fibrogenic phenotype in liver cells (i.e., production of hyaluronic acid [HA], collagens, tissue inhibitor of metalloproteinase 1 [TIMP-1], and so on). In a patient with suspected NAFLD or NASH, useful baseline testing should include levels of aminotransferases both aspartate aminotransferase (AST) and ALT, total and direct bilirubin, γ-glutamyltransferase (GGT), fasting serum glucose and insulin, as well as a lipid panel. Transaminases may range from normal to 4 to 6 times the upper limit of normal, but mild elevations are usually seen ranging between 1.5 to 2 times the upper limit of normal. Generally, the ratio of AST to ALT is less than 1, but this ratio may increase as fibrosis advances. Circulating levels of aminotransferases may fluctuate over time and may be normal in a large proportion of children with NAFLD and NASH. Furthermore, normal aminotransferase levels do not exclude the presence of fibrosis or even cirrhosis. Serum alkaline phosphatase and GGT levels may also be mildly abnormal. Given that most patients with NAFLD have some components of the metabolic syndrome, lipid profiles as well as fasting glucose and insulin levels should be evaluated. Insulin resistance can be determined by fasting insulin levels or by further studies if necessary (glucose challenge or glucose tolerance test). Albumin, bilirubin, and platelet levels are usually normal, unless the disease has evolved to cirrhosis. As in adults, some children with NAFLD may have positive autoantibody testing (antinuclear and anti-smooth muscle antibody); in the absence of autoimmune hepatitis, the significance of these findings in this setting is still unclear.

Establishing diagnosis and disease severity as well as monitoring children over time remains a major challenge for pediatricians caring for the growing number of children with NAFLD. A liver biopsy is still considered the gold standard; however, this invasive procedure is not suitable for screening and risk stratification of children with this condition. There is a great need to develop noninvasive, simple, and reliable tests that can replace the liver biopsy for these purposes. The currently available noninvasive tests as reviewed earlier have two central limitations, the lack of sensitivity and specificity to distinguish NASH from the more serious form of NAFLD from hepatic steatosis and to stage the presence and extent of liver fibrosis. Thus, identifying and validating potential novel noninvasive biomarkers is a central area of research. Diagnostics development in the area of NAFLD research has been divided into two major groups: those directed to detecting and quantifying the presence of fibrosis and those directed to establish the diagnosis of NASH. Regarding the former, the pediatric NAFLD fibrosis index (PNFI), which is obtained from three simple measures (age, waist circumference [WC], and triglycerides [TG]) was recently developed to predict liver fibrosis in children with NAFLD. This index is easy to calculate with no additional cost to the patient and it has a good positive predictive value to rule in fibrosis; however, its negative predictive value to rule out fibrosis is suboptimal. These limitations can be overcome when used in a sequential algorithm with the enhanced liver fibrosis (ELF) test, which uses a combination of three extracellular matrix components, namely HA, amino-terminal propeptide of type III collagen (PIIINP) and TIMP-1, resulting in an accurate assessment of the presence of liver fibrosis in children. Future studies are still needed to externally cross-validate these findings before the combination of PNFI and ELF can be recommended in children with NAFLD. Moreover, longitudinal studies measuring these panels serially against clinical outcomes will determine if they can be used to measure disease progression and regression.

Hepatocyte apoptosis has been found to be a prominent feature in patients with NASH, making it an interesting focus for biomarker development and for therapeutic intervention. A large body of evidence has demonstrated the utility of a measurement of plasma levels of a specific by-product of apoptosis in liver cells, caspase-generated cytokeratin-18 (CK-18) fragments in the diagnosis of NASH in adult patients. Recently, Fitzpatrick et al. demonstrated that children with biopsy-proven NAFLD also showed considerably elevated levels of the CK-18 fragments as compared to healthy controls. In addition, those with established NASH showed significantly higher numbers versus those with hepatic steatosis or borderline disease. These results suggest that measuring CK-18 fragments may be useful in the workup of children suspected of having NASH, but before we can know how these markers will perform in pediatric clinics, larger validation studies are needed.

Numerous other biomarkers of inflammation, oxidative stress, apoptosis, and fibrosis are under investigation. However, more studies are needed to validate the existing markers and techniques and develop other accurate noninvasive predictors of disease severity.