Nonalcoholic Fatty Liver Disease in Children with Obesity

C H A P T E R

20 Nonalcoholic Fatty Liver Disease in Children with Obesity Shima A. Dowla, Amy M. Goss, and Ambika P. Ashraf University of Alabama School of Medicine, University of Alabama at Birmingham, Birmingham, AL, United States

20.1 INTRODUCTION In the United States, childhood obesity prevalence has tripled in the past 30 years, with one in every six children now classified as obese [1–3]. In parallel to the rapid growth of childhood obesity, nonalcoholic fatty liver disease (NAFLD) has become the most common form of chronic liver disease in children and adolescents in the United States and industrialized nations [4–6]. Although prevalence estimates vary between studies, research using gold standard liver biopsy suggests that more than one in three children with obesity are affected with this condition [4, 6]. The most commonly accepted definition of NAFLD is macrovesicular lipid accumulation in
5% hepatocytes due to reasons other than alcohol consumption [6]. It can be understood as being on a spectrum, ranging from minimal lipid accrual in the liver, hepatic steatosis, to lipid accumulation causing hepatic inflammation, or nonalcoholic steatohepatitis (NASH). NAFLD is particularly alarming in the pediatric population because long-term exposure and buildup of lipids in the liver promotes hepatocellular injury and fibrosis, and can lead to significant morbidity and mortality [7]. Indeed, epidemiological research in this field has shown strong associations between NAFLD diagnosed in childhood and cirrhosis, hepatocellular carcinoma, and end-stage liver disease [8–12]. Unfortunately, NAFLD remains underdiagnosed in children and adolescents due to its asymptomatic nature, invasive and expensive diagnostic procedures, and lack of standardized screening guidelines for pediatricians [5, 13]. Although NAFLD is a newly emerging disease in the pediatric population, many developments in our understanding of its epidemiology, pathogenesis, diagnosis, and treatment have been made in recent decades. This chapter reviews these advances and highlights areas where more research is needed.

20.2 EPIDEMIOLOGY 20.2.1 Prevalence Estimates Epidemiological studies from around the world have reported prevalence estimates for pediatric NAFLD [4]. These estimates vary, however, due to many reasons such as differences in the demographic makeup of the sample population, prevalence of comorbid conditions such as obesity and insulin resistance, and methodology used to define the disease. The majority of prevalence studies in the pediatric literature use elevated serum alanine aminotransferase (ALT) levels to define NAFLD despite its low sensitivity and specificity for detecting the presence of disease [4, 14]. As such, prevalence reports using serum aminotransferase levels may provide an inaccurate estimate of the true rates of NAFLD in the pediatric population. Section 20.4.1 further details the issues with diagnosing NAFLD using liver transaminases. Few pediatric studies have used diagnostic approaches like ultrasound (US), magnetic resonance imaging (MRI), and gold standard liver biopsy to estimate the rates of pediatric NAFLD [4]. The most robust and rigorous NAFLD prevalence study to date in the pediatric population was performed by Schwimmer et al., which used autopsy

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specimens (n ¼ 742) to histologically evaluate the prevalence of fatty liver, which they defined as
5% of hepatocytes with macrovesicular fat [6]. Prevalence of hepatic steatosis in the sample population was 13%, ranging from <1% in young children (ages 2–4 years) to 17% in adolescents (ages 15–19 years), and 38% among children with obesity. The results of this study should be interpreted with caution, however, as the high percentage of Hispanic children in the sample makes the results less generalizable to populations with greater heterogeneity.

20.2.2 Histological Characteristics Schwimmer et al. first examined the histopathology associated with pediatric NAFLD/NASH and found several histological distinctions from the adult form of the disease [8]. One hundred biopsies were reviewed and classified into two histological subgroups, deemed “type 1” and “type 2”. Among all patients, the prevalence of hepatic steatosis was 16% and advanced fibrosis was 8%. Characteristics associated with the type 1 phenotype of the disease included steatosis, lobular inflammation, ballooning degeneration, and perisinusoidal fibrosis. This phenotype characterized 17% of children with NAFLD and was more common among Caucasians. Features present in the type 2 phenotype included macrovesicular steatosis, portal inflammation, and portal fibrosis. In comparison to type 1 NASH, type 2 NASH had negligible or minimal amounts of ballooning degeneration. However, advanced fibrosis was more prevalent in this subgroup. More than one-half of the subjects (51%) fit into the type 2 phenotype and were more commonly male, younger, overweight, and of Asian, Native American, and Hispanic race/ethnicity. The final one-third of the sample population had liver biopsies with histological characteristics consistent with both types of NASH. Section 20.4.3 contains more information on the histopathology and histological scoring system associated with pediatric NAFLD and NASH.

20.2.3 Sex In both adults and children, the prevalence of NAFLD is higher in males than females, although no sex-related differences have been reported in the development of NASH [5, 6]. Several reasons for this sex disparity have been hypothesized, including differences in hormones, body composition, fat distribution, and metabolic profile [5]. However, an emerging body of literature suggests that sex-differences in endogenous estrogen production and action may be the primary underlying factor creating the sex discrepancy present in NAFLD. Results from studies in humans and animal models support the hypothesis that estrogen confers protection against NAFLD [15–19]. In rodent models with liver injury, exogenous administration of estrogen has been shown to attenuate the effects of lipid peroxidation, oxidative stress, mitochondrial dysfunction, and immune related hepatic injury in later stages of the disease, and even reverse hepatic steatosis [15–17, 20]. Puberty and menopause may be particularly salient time periods to study to establish the causal effects of estrogen on NAFLD development and progression. A prospective study conducted by Suzuki and colleagues demonstrated that children who had undergone puberty had less severe NAFLD compared to prepubescent children, including reduced steatosis, steatohepatitis, portal inflammation, and fibrosis [19]. Although estrogen levels were higher in postpubescent females than males, results indicated that estrogen conferred some protection to all children during and after puberty. Additional prospective studies have also studied the relationship between age (menopause) and NAFLD severity [5, 18]. Nevertheless, the prospective nature of the reported studies limits causal inference; randomized clinical trials are needed in both adult and pediatric populations to further establish the role of estrogen in the development, progression, and treatment of NAFLD.

20.2.4 Age Estrogen may confer a degree of protection against NAFLD. Paradoxically, however, the prevalence of NAFLD has been shown to be higher in adolescence than childhood. This may be attributed to the deleterious effects of other hormones during puberty in males. Some research has demonstrated that insulin resistance, which is strongly implicated in NAFLD pathogenesis, is higher in boys during the pubertal transition and persists onward [21, 22]. This relationship was also significantly associated with dyslipidemia (low high-density lipoprotein cholesterol [HDL-C], increased triglyceride levels) and increased systolic blood pressure, even after controlling for reductions in fat mass [22]. In addition to biological influences, diagnostic and social factors may also impact the prevalence of NAFLD in childhood. Prevalence of the disease may simply be higher in adolescence because it is screened for and diagnosed at a later age. NAFLD is commonly asymptomatic and may only be detected at much later stages of the disease. In addition, the time frame from initial screening to final diagnosis could take months to years, depending on physician or specialist

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availability, as well as patient-specific factors (i.e., insurance access, socioeconomic status). Moreover, screening guidelines established by the Expert Committee for the Prevention, Assessment, and Treatment of Child and Adolescent Overweight and Obesity recommend biannual screening of NAFLD starting at the age of 10 for overweight and obese children, despite several studies indicating that NAFLD is present in children as young as 2 years of age [6, 23].

20.2.5 Race/Ethnicity Little research has examined racial and ethnic differences in pediatric NAFLD. Some studies have demonstrated that pediatric NAFLD is more common among Hispanic and Asian children and less common in African Americans, but causes behind these racial and ethnic disparities have yet to be elucidated [5, 6, 24, 25]. Although African American children have higher rates and more severe clinical presentations of insulin resistance and obesity, they paradoxically have the lowest prevalence of NAFLD [5, 6, 25, 26]. Genetic differences between races may help this discrepancy. In fact, genome-wide association studies have been conducted specifically to understand racial and ethnic differences in pediatric NAFLD and have found several genetic variants that may be implicated in disease prevalence and severity [5]. One such variant in African Americans, which enhances lipoprotein lipase activity and attenuates hepatic lipase, may help explain the low prevalence of dyslipidemia, a strong correlate of NAFLD, in this population [26].

20.2.6 Disease Correlates NAFLD is often regarded as the hepatic manifestation of metabolic syndrome, as it is highly correlated with abdominal obesity, insulin resistance, hypertension, and dyslipidemia [24, 27–29]. The critical roles of obesity and insulin resistance in the pathogenesis of NAFLD are more fully described in Section 20.3. In addition to metabolic syndrome, NAFLD is strongly linked to cardiovascular disease (CVD) and is associated with several CVD risk factors [30, 31]. In fact, CVD is the leading cause of death in individuals with NAFLD [32]. Moreover, dyslipidemia, a known CVD risk factor and component of metabolic syndrome, is prevalent in pediatric NAFLD and has been implicated in NAFLD severity and CVD risk in those with NAFLD [24, 29, 33–36]. Dyslipidemia is characterized by elevated cholesterol, low-density lipoprotein (LDL-C), triglycerides (TG), and low HDL-C. Recent studies have reported that the prevalence of elevated total cholesterol, TG, LDL-C, non-HDL-C, and low HDL is as high as 20%–80% in children with NAFLD [24, 27–35].

20.2.7 Complications Because pediatric NAFLD is a newly emerging disease, little is known about the long-term morbidity and mortality associated with the condition. In adults with NAFLD, the most common causes of death are CVD, cancer, and chronic liver disease [32]. Using 20 years of follow-up data, Feldstein and colleagues investigated the long-term prognosis of 66 children with NAFLD [9]. At baseline, 66% of the sample population had obesity and 83% had one condition related to metabolic syndrome. Among all subjects at follow-up, two developed cirrhosis and two died. Researchers determined that this cohort was 13.6 times more likely than healthy children of the same age and sex to die or need liver transplantation.

20.3 PATHOGENESIS NAFLD exists on a spectrum from simple hepatic steatosis to NASH, a progressive medical condition involving extensive hepatocellular inflammation, injury, and potentially fibrosis [37]. It is not well understood how NAFLD progresses in severity from simple steatosis, a relatively benign condition, to NASH. This section describes what is currently known about its pathogenesis and highlights most recent contributions to the literature.

20.3.1 Introduction The “two-hit” hypothesis, proposed by Day and colleagues almost two decades ago, remains the most recognized and accepted of all hypotheses involving the pathogenesis and progression of NAFLD [38]. The first “hit” in this hypothesis involves triglyceride accumulation in the hepatocyte, whereas the second includes additional insults leading to hepatocellular inflammation, injury, and death. Such mechanisms of injury include mitochondrial dysfunction,

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oxidative stress, and production of reactive oxygen species (ROS). Findings from rodent models have provided great insight into the cellular and molecular changes associated with each phase of the disease and are detailed in the following subsections.

20.3.2 Hepatic Steatosis The first step in the development of NAFLD is fat accumulation in the liver, or hepatic steatosis. Although not fully elucidated, the initiation of steatosis is believed to be associated with a cascade of molecular events involving altered nutrient metabolism [7]. Two processes are most strongly implicated: (1) enhanced hormone sensitive lipase (HSL) activity, resulting in increased concentration of plasma free fatty acids (FFA) and (2) hepatic de novo lipogenesis (DNL), resulting in increased concentrations of hepatic FFA and a blockade of fatty acid oxidation [7]. In the context of insulin resistance, HSL activity is enhanced, which hydrolyzes available fat in adipocytes (lipolysis) and releases FFA into the circulation [39]. These FFA are then imported into the liver for proper storage or breakdown via beta oxidation. Individuals who are obese or disproportionately accumulate fat in their abdomen have increased adipocyte number and mass, allowing for greater concentrations of plasma FFA and uptake into the liver. Several epidemiological studies have consistently shown that these individuals are also at greater risk for developing NAFLD [27, 28]. Once inside the liver, FFA can either be converted into triglycerides or broken down via fatty acid oxidation to form adenosine triphosphate (ATP). Newly converted triglycerides either remain in the hepatocyte for storage, or are released from the liver via very low-density lipoproteins (VLDL) or intermediate-density lipoproteins (IDL) [7]. One of the primary purposes of the liver is to maintain glucose homeostasis in the plasma. In an effort to do so, excess glucose in the circulation, often from dietary ingestion of carbohydrates, is imported into the liver and other tissues such as skeletal muscle. In contrast, in an energy-deficient state, glucose is generated in the liver and released into the bloodstream. Hepatic DNL is the process by which the liver produces FFA from excess glucose present in the circulation. Hyperinsulinemia, a physiological consequence of insulin resistance, promotes hepatic DNL by activating sterol regulatory element-binding protein-1c or SREBP-1c [7, 40, 41]. This lipogenic transcription factor is responsible for inducing expression of all cellular machinery needed for fatty acid synthesis from glucose. Increased concentrations of glucose in the blood, hyperglycemia, further promote hepatic DNL by activating the carbohydrate response element binding protein (ChREBP) [42]. The actions of ChREBP synergize with those of SREBP-1c and lead to the increased production of FFA in the liver. In a healthy liver, available FFA is esterified or oxidized to produce energy for the cell. In NAFLD, however, fatty acid oxidation is bypassed, and FFA substrates are converted into triglycerides. This is attributed to the increased production of a de novo biosynthesis byproduct, malonyl Co-A, a coenzyme that inhibits a key enzyme necessary for beta oxidation (carnitine palmitoyl transferase-1) and effectively shunts all biochemical pathways toward triglyceride accumulate in the liver [7]. Other abnormalities in lipid metabolism may also contribute to hepatic triglyceride accumulation present in NAFLD [43]. Typically, hepatocytes package cholesterol and triglycerides into VLDL, which are subsequently secreted into plasma. The triglycerides in VLDL are later metabolized to release FFA for energy, and the excess triglycerides are stored in locations in the body such as muscle, adipose tissue, and organs such as the heart. In NAFLD, there is increased production and secretion of VLDL in an effort to match TG production [43]. However, this ratio is not always balanced, and lower quantities of VLDL allow for residual triglyceride and precursors of triacyl glyceride to accumulate in the liver [43]. Emerging literature suggests that VLDL production and secretion may also be impaired in very advanced stages of NAFLD and contribute to hepatic dysfunction. In particular, a study by Fujita et al. found that adults with NASH, in comparison those with NAFLD, had similar hepatic lipid profiles but reduced secretion of VLDL, which was attributed to impairments in VLDL synthesis [44]. As lipid profile or concentrations did not significantly vary between the two groups, the researchers proposed that dysregulation of VLDL metabolism contributed to the oxidative damage present in NASH.

20.3.3 Steatosis to Steatohepatitis Hepatic steatosis is necessary but not sufficient for progression of NAFLD to NASH. The etiological mechanisms underlying this progression remain poorly understood and are restricted to inferences made using studies with animal models. Although several mechanisms of hepatic injury have been cited in the literature, the most established, and the focus of this section, will be mechanisms of cellular injury induced by the presence of hepatic fatty acid. Increased concentrations of fatty acid in the liver prompt cellular organelles such as the endoplasmic reticulum (ER) and peroxisomes to degrade fatty acid via oxidation [7]. In this process of removal, ROS, namely hydrogen peroxide

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and other superoxide anions are formed [7]. Although less studied, defects in the mitochondrial respiratory chain in patients with NASH have also been shown to increase the production of ROS [45, 46]. ROS can attack specific lipid particles, polyunsaturated fatty acids, to begin the process of lipid peroxidation [43]. Aldehyde byproducts of this biochemical reaction are more reactive, have longer half-lives, and can diffuse further than ROS to affect neighboring cells [47]. Together, these molecules and ROS activate mechanisms involved in cell injury and death such as glutathione depletion and DNA damage. Moreover, they promote inflammation by generating potent cytokines and attracting immune cells such as neutrophils. Third, they initiate fibrosis by engaging hepatic stellate cells, which produce collagen and scar tissue to contain liver injury [7].

20.3.4 New Insights in Pathogenesis Although obesity and insulin resistance remain integral to the pathogenesis of the NAFLD, additional mechanisms and mediators of injury, particularly in the pediatric population, have been implicated in recent years. This has led to an expansion of the “two-hit” hypothesis, into the “multiple-hit” model, which involves interactions between several genes, cells, tissues, organs, and environmental factors [5]. Hepatic progenitor cells are stem cells of the liver that can differentiate into any cell type and be activated during hepatic stress or injury. They have most recently been shown to be associated with pediatric NAFLD, working to attenuate the effects of oxidative stress on the liver and contain hepatic injury but paradoxically increasing levels of fibrosis and severity of NASH [48]. A growing body of literature has also demonstrated the role of Kupffer cells, macrophages of the liver, in hepatic steatosis, potentially through the suppression of peroxisome proliferator-activated receptor alpha activity [49, 50]. Finally, although only recently recognized, CD8 + T cells may play a critical role in the development of pediatric NASH. In comparison to adults with the condition, children with NASH demonstrated an overproliferation and activation of this cell line in response to cellular injury [51].

20.4 DIAGNOSIS The diagnosis of NAFLD in childhood can be particularly challenging because of its asymptomatic nature, the lack of sensitivity and specificity of most commonly used screening tests, and often, lack of awareness of the condition by health care professionals. Indeed, NAFLD has been shown to be underscreened for by general pediatricians and specialists [13]. The time frame from initial screening to final diagnosis could potentially take months to years, depending on physician or specialist availability as well as patient-specific factors. Traditional methods to screen for NAFLD include an assessment of known risk factors (demographics, family history, obesity, insulin resistance, metabolic syndrome), blood tests evaluating liver function, additional tests to rule out other liver pathologies, and finally, radiological and/or histological analysis if deemed necessary. Accordingly, this section will describe these methods of screening and diagnosis.

20.4.1 Serum Biomarkers Several biomarkers are associated with NAFLD and NASH [52]. These can be organized into the following categories: (1) liver transaminases (ALT, AST, GGT); (2) other markers of hepatocyte inflammation and injury (i.e., CK-18, TNF-a, C-reactive protein); and (3) indicators of liver fibrosis (i.e., hyaluronic acid). Clinically, however, only ALT and AST are commonly measured to screen for and assess severity of NAFLD. Liver transaminases are routinely obtained because they are widely available, inexpensive, and can be elevated in NAFLD. However, they are elevated in a wide range of liver conditions. Consequently, they have low sensitivity (45%) and specificity (85%) for diagnosing NAFLD [14]. In fact, many studies have reported that more than half of children with NAFLD do not present with elevations in liver transaminases [53–55]. Moreover, there is no consensus in the medical or scientific community as to what serum transaminase concentration is considered an elevated value. Past studies investigating NAFLD have used an ALT level that was 1.5 to 2 times the upper limit of the reference range to define the disease [14, 23]. However, reference ranges vary between institutions and even laboratories, making a numerical cutoff difficult to quantify. Finally, children with and without elevations in liver enzymes are at equivalent risk of progression to more severe disease [56]. Given these statistics, the use of a liver function test as the primary and single screening tool for pediatric NAFLD is problematic.

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20.4.2 Radiological Imaging The gold standard method for diagnosing NALFD is liver biopsy. However, it is often not practical, feasible, nor ethical to obtain a highly invasive liver biopsy from a child for the purposes of diagnosis, particularly because there are no approved pharmacological therapies to treat the condition. As such, several imaging modalities have been adapted for clinical and/or scientific use to assess hepatic fat infiltration. Advantages and disadvantages of each modality are presented in Table 20.1 [57]. Of note, the majority of the literature on imaging modalities for NAFLD comes from adult and animal studies; more research is needed in the pediatric population to validate these methods for clinical practice. 20.4.2.1 Ultrasound Ultrasound (US) is the most popular imaging modality used to assess NAFLD as it is relatively inexpensive, widely accessible, and safe for use in children. During an US evaluation, hepatic tissue is compared to neighboring tissue (renal) to determine differences in brightness. Tissue with fatty infiltration appears brighter on US than tissue without fat; this tissue is considered to have increased echogenicity [57]. Severity of hepatic steatosis can be rated on a fourpoint scale from 0 (normal liver) to 3 (severe steatosis). However, this assessment is more qualitative than quantitative by nature, making it difficult to use US as a method for tracking hepatic fat improvement or worsening over time [58]. More sophisticated technologies have been developed to address this limitation, although they are not currently implemented in the clinical setting [57]. Sensitivity of this radiologic modality depends on the degree of hepatic steatosis and existence of other liver conditions. In the absence of other liver pathology, hepatic triglyceride content of
30% can be detected with high TABLE 20.1

Summary of Imaging Modalities for NAFLD

Imaging Modality

Strengths

Limitations

Ultrasound (US)

• • • • •

• Qualitative assessment • Highly operator dependent • Cannot differentiate between various levels of steatosis • Unable to assess fibrosis • Lower sensitivity and specificity in the presence of other liver conditions • Evaluates steatosis using surrogate measures

Computed tomography (CT)

• Accessible • Moderate training needed to perform • High sensitivity for moderate steatosis

• • • •

Magnetic resonance imaging (MRI)

• • • • • •

More direct measure of hepatic fat (than US or CT) Quantitative Assessment High sensitivity and specificity for all levels of hepatic steatosis Can distinguish between steatosis and NASH Easier than MRS to perform scan and interpret data Assessment of multiple regions of the liver

• Expensive • Small space, potential for claustrophobia

Magnetic resonance spectroscopy (MRS)

• Same strengths as MRI (objectivity, reproducibility, sensitivity, specificity, and ability to distinguish disease)

• Expensive • Small space, potential for claustrophobia • Expert is needed to conduct scan and analyze images • Image analysis is time consuming • Single voxel used to interpret hepatic fat

Elastography (US)

• Can differentiate between various levels of fibrosis

• Cannot exclusively evaluate degree of hepatic steatosis or inflammation • Cannot accurately evaluate severely obese patients

Elastography (MR)

• Can differentiate between various levels of fibrosis and steatohepatitis • Accuracy of technique is BMI dependent

• Cannot exclusively evaluate degree of hepatic steatosis or inflammation

Safe Inexpensive Accessible Least training needed to perform High sensitivity for moderate steatosis

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Ionizing radiation Low sensitivity for mild steatosis Unable to assess fibrosis Evaluates steatosis using surrogate measures

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sensitivity (80%–100%) and specificity (98%) [58, 59]. However, it is not as precise at detecting or distinguishing between lower levels of hepatic steatosis or more complicated pathology like fibrosis [60–62]. Moreover, in the presence of other liver conditions, such as hepatic tumors and inflammatory disease, US has been shown to have reduced sensitivity and specificity at detecting moderate steatosis [63–65]. The final and likely greatest limitation associated with US is its variability in interpretation, which differs based on personnel. In fact, a prospective study found that the results of 22% of US readings differed by interpreter, and the number of diagnoses of hepatic steatosis made by radiologists were significantly different [58]. Taken together, this body of work suggests that US may be best utilized as a screening method to detect moderate hepatic steatosis in children and adults at risk for or with clinical suspicion of NAFLD. 20.4.2.2 Magnetic Resonance MRI and MRS are currently the most accurate noninvasive methods of measuring hepatic fat infiltration and have even been shown to be more reliable than liver biopsy at determining the degree of hepatic steatosis. MR technology uses the ratio of protons bound to fat, to all protons (those bound to fat and water) to estimate hepatic fat. This ratio, known as the proton density fat fraction, relies on the chemical-shift phenomenon to differentiate between the types of bound protons [57, 66]. More recently, MR methods have also developed techniques to directly estimate hepatic content. The physics and other technical components of both technologies are further described by Lee et al. [57]. Several studies have compared these imaging modalities to available technologies used to diagnose NAFLD (i.e., CT and US) and have determined that they are superior at differentiating the severity of hepatic steatosis and between steatosis and steatohepatitis [58, 60]. Unlike US or CT, MRI and MRS can detect very mild steatosis (
5% of fat in liver) with high sensitivity and specificity [58, 60]. Moreover, unlike US, MR does not rely on qualitative interpretations to assess degree of steatosis; instead, it uses the quantitatively derived proton density fat fraction. In many ways, MR rivals liver biopsy and has been suggested by many as a better alternative for the diagnosing NAFLD [67–69]. In comparison to liver biopsy, MR techniques have been shown to be more accurate, reproducible, and flexible in terms of grading disease severity [67–69]. These technologies, particularly the MRI, use larger portions of the liver to determine the extent of hepatic fat infiltration, which may explain their precision. Finally, and likely most relevant for the pediatric population, both MRI and MRS are noninvasive techniques used to assess hepatic fat. Although past research comparing MRI and MRS technologies suggest that MRS is superior in accurately detecting and quantifying hepatic steatosis, newer work has demonstrated that both imaging modalities are comparable [70]. MRI is the more commonly practiced technique, as it more widely accessible, involves a shorter and less complicated protocol, and takes less time to analyze. Moreover, MRI quantifies multiple regions of the liver rather than a single voxel [57]. MRS, in contrast, requires extensive expertise and time to both operate and analyze data [71]. Nonetheless, advances in these techniques have been developed in recent years to incorporate both MR imaging and spectroscopy. However, such methods are relatively new, highly specialized, and used primarily for scientific purposes, making them an impractical and unlikely option for use in the clinical setting in the near future. For these reasons, MRI may currently be the most appropriate method for diagnosing NAFLD in the pediatric population. 20.4.2.3 Other Imaging Modalities Similar to the US, computed tomography is a highly accurate imaging modality used to detect moderate steatosis and is equivalently inadequate at detecting lower levels of hepatic fat. Its greatest limitation, however, which has effectively eliminated it as a viable imaging option in children, is its emission of high levels of ionizing radiation. Because it is comparable to the US in terms of detecting steatosis, it is not an appropriate radiological technique to diagnose NAFLD [57]. Despite advancements made in the radiologic evaluation of NAFLD, current imaging modalities cannot yet accurately detect advanced stages of steatohepatitis. However, new technologies, namely US and MR elastography, hold great promise in the management of NASH. Both techniques, which measure shear wave velocity to estimate liver stiffness, have been shown to differentiate between various degrees of fibrosis with high sensitivity and specificity [72–76]. MR elastography may be moderately superior to US elastography, however, due to its ability to differentiate between liver fibrosis with and without hepatic steatosis and inflammation [77]. Moreover, the accuracy of the technique does not dependent on the body habitus of the patient [78]. The main limitation of both of these techniques is their inability to detect less severe stages of NAFLD [5, 57]. As such, they should be used in conjunction with other imaging modalities to fully evaluate a patient with NAFLD at risk for fibrosis.

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20.4.3 Liver Biopsy Liver biopsy remains the gold standard diagnostic technique for the assessment of pediatric NAFLD and NASH. To systematically evaluate the histopathology associated with these liver conditions, many scoring systems have been developed. However, the most comprehensive and well-cited of all scoring systems is the NAFLD scoring system designed by the NASH Clinical Research Network, which has been validated for use in pediatric populations [37]. The NAFLD scoring system is a histopathological scoring system that assesses the severity of NAFLD using 13 characteristics: hepatic steatosis, hepatocellular ballooning, lobular inflammation, fibrosis, microvesicular steatosis, glycogenated nuclei, microgranulomas, megamitochondria, acidophil bodies, lipogranulomas, portal inflammation, pigmented macrophages, and Mallory hyaline. The first four listed histological characteristics are evaluated using a partially quantitative scale, and the final nine are scored on a binary (present/absent) scale. This classification scheme also offers an activity score (“NAFLD Activity Score”) that has been widely used in epidemiological studies to evaluate the prevalence of NAFLD. The NAFLD Activity Score, which ranges from 0 to 8, is calculated by combining the individual scores for steatosis, lobular inflammation, and ballooning degeneration [37, 71]. Scores <3 are considered to be free of NASH and scores
5 are associated with NASH. A score of 3 or 4 is borderline and requires additional assessment to determine the extent of disease progression.

20.5 TREATMENT Because NAFLD is a newly emerging disease, mechanistic insight into the pathophysiology of NAFLD in children and adults remains limited. For this reason, primary management of the condition involves the treatment of comorbidities, particularly weight, insulin resistance, metabolic syndrome, and other associated factors, to improve longterm morbidity and mortality of patients. To date, there are no pharmacological therapies approved to treat pediatric NAFLD.

20.5.1 Lifestyle Intervention The mainstay of therapy for the management of pediatric NAFLD is weight loss through caloric restriction and exercise. Based on a report from the American Association for the Study of Liver Diseases, American College of Gastroenterology, and American Gastroenterological Association, a gradual weight loss of 3%–5% of one’s body weight is recommended to improve hepatic steatosis; however, weight loss of 5%–10% may be necessary to reduce hepatic inflammation [78]. These recommendations use evidence from adult studies however, and may not be generalizable to pediatric and adolescent populations. Lifestyle modification may improve liver fat, liver function, and comorbid conditions in children with NAFLD in the short and long term. In a European study, 73 obese adolescents with US-confirmed NAFLD were placed on a 12-week multidisciplinary treatment program, consisting of diet change, physical activity, and lifestyle coaching [79]. Researchers found significant improvements in liver enzymes, visceral adiposity, insulin resistance, and NAFLD prevalence using ultrasonography. Two studies by Nobili et al. have also demonstrated long-term efficacy of lifestyle intervention on improvements in NAFLD and NASH in children. In one such study, 90 children were placed in either the lifestyle intervention group, or vitamin E and vitamin C supplementation group for 12 months. Although liver-related effects were not significantly different between the two groups at follow-up, subjects who reduced their total body weight by
20% showed significant reductions in hepatic fat as well as ALT levels [80]. The latter study, an extension of the first trial, randomized 53 children to the same two groups for 24 months to assess changes in liver histology [81]. This study did not show any significant differences between the two groups in terms of primary endpoints. Nonetheless, both treatment groups did demonstrate significant reductions in the degree of steatosis, lobular inflammation, hepatocyte ballooning, and NAFLD activity score. Similarly, a study conducted by Reinehr et al. found that participating in lifestyle intervention was associated with significant reductions in liver transaminases (AST, ALT), BMI, and NAFLD prevalence [82]. The differential effect of macronutrient composition on improvements in NAFLD in children remains unknown. However, guidelines from the American Association for the Study of Liver Diseases recommend that diet choice should be dependent on one’s ability to adhere to the chosen diet in the long term. To date, only one study has tested the effects of diet quality in this population. Ramon-Krauel et al. compared the effect of a low glycemic load diet vs. conventional low fat diet in 17 obese children and adolescents with NAFLD [83]. Among the 16 children who completed the intervention, there were no statistically significant differences between the two diet groups in terms of

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reductions in liver fat (quantified using MRS), visceral adiposity, BMI, ALT, or insulin resistance. It is difficult, however, to draw conclusions about the beneficial effect of macronutrient modification using the results of one single pilot study. A two-arm, parallel design randomized controlled trial is currently underway that tests the effect of a carbohydrate (CHO)-restricted diet (< 25: > 50:25% daily calories from CHO: fat: protein) vs. fat-restricted diet (55, 20:25% daily calories from CHO: fat: protein) on improvements in hepatic lipid (also measured MRS), liver transaminases, markers of inflammation, body composition, visceral adipose tissue, and insulin resistance in adolescents with NAFLD [84].

20.5.2 Pharmacological Therapies Several pharmacological therapies have been evaluated in the treatment of pediatric NAFLD. These include, but are not limited to, insulin sensitizers (metformin and thiazolidinediones), antioxidant and cytoprotective agents (vitamin E, ursodiol), antiobesity medications (orlistat), statins, and probiotics. Although short-term results are encouraging, more research, particularly in children and adolescents, is needed to determine the therapeutic potential of these medications. 20.5.2.1 Insulin Sensitizers Insulin resistance plays an integral role in the pathogenesis of NAFLD. Accordingly, pharmacological therapies used to treat insulin resistance (biguanides and thiazolidinediones) have been tested in adults and children with NAFLD [5]. Metformin, a principal therapy in the treatment of type 2 diabetes, is an insulin-sensitizing medication that has most recently been rigorously evaluated as a potential treatment for pediatric NAFLD. Although its mechanisms of action are not fully understood, metformin suppresses hepatic gluconeogenesis, promotes glucose uptake in several peripheral locations, including the muscle, and improves insulin sensitivity [85]. Open-label clinical trials evaluating metformin as a treatment for NAFLD have shown significant improvements in liver enzymes, steatosis, steatohepatitis, and fibrosis, as well as metabolic profile in adults [5, 85]. To date, four studies have tested the effect of metformin on improvements in NAFLD in children and have demonstrated inconsistent findings [86–89]. Of note, two of four trials were pilot studies consisting of 10 total participants. The most robust study conducted to evaluate the role metformin in the treatment of pediatric NAFLD, named the Treatment of Nonalcoholic Liver Disease in Children (TONIC) trial, was a multicenter, double-blind, placebocontrolled randomized clinical trial in which 173 children were randomized to vitamin E (n ¼ 58), metformin (n ¼ 57), or placebo (n ¼ 58) [89]. The primary outcome measure of this clinical trial was improvements in ALT; secondary outcomes included histological improvements and/or resolution of NAFLD and NASH. Results demonstrated no significant differences between the three groups in terms of ALT reduction. In addition, other than significant differences in hepatocellular ballooning, metformin did not improve the histopathology associated with NAFLD in comparison to the placebo [89]. Several studies have also evaluated the efficacy of thiazolidinediones, namely pioglitazone and rosiglitazone, on improvements in NAFLD in adults, and have shown encouraging results [5, 85]. This group of insulin-sensitizing medications act on the peroxisome proliferator-activated receptor gamma nuclear transcription regulator to improve glucose uptake in the periphery and promote fat redistribution into adipose tissue [5]. However, the alarming side effect profile of thiazolidinedione has largely prevented its utilization as viable treatment option for patients with NAFLD, particularly among children. Rosiglitazone, for example, has been shown to significantly increase the risk for heart attack and death from cardiovascular complications [90]. In addition, thiazolidinediones promote weight gain and may therefore inadvertently worsen NAFLD [5]. 20.5.2.2 Antioxidant and Cytoprotective Agents Hepatocyte injury induced by lipid peroxidation, mitochondrial dysfunction, and oxidative stress is necessary for the progression of steatosis to steatohepatitis. By limiting the extent of hepatocyte inflammation and injury, antioxidants and cytoprotectants may prevent the development or reduce the severity of NASH. Vitamin E is the most popular antioxidant therapy investigated and clinically prescribed for pediatric NAFLD [91]. In adults, vitamin E has unequivocally demonstrated efficacy in improving liver transaminases, hepatic steatosis, inflammation, and pathology contributing to the development of NASH [92, 93]. Studies in the pediatric population show mixed results. A double-blind randomized clinical trial, which compared lifestyle modification to lifestyle modification with vitamin C (500 mg/day) and vitamin E (600 IU/day) for 12 months, did not find significant changes in

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ALT, insulin resistance, or weight loss between the two groups [80]. Moreover, in the TONIC multicenter clinical trial, which evaluated the efficacy of metformin and vitamin E in improving NAFLD-related outcomes, neither vitamin E nor metformin demonstrated significant improvements in liver transaminases in comparison to the placebo. Nonetheless, vitamin E did show encouraging findings related to hepatocellular ballooning, NAFLD severity, and NASH resolution [89]. Ursodeoxycholic acid, more commonly known as ursodiol, is a widely prescribed medication that competitively inhibits the absorption of hydrophobic bile acids, thereby protecting hepatocytes from bile acid-induced mitochondrial injury [5]. Ursodiol has not been shown to be efficacious in children with NAFLD, although only one clinical trial has been conducted in children to date [94]. In this study, Vajro et al. [94] assigned 31 children to a low-calorie diet alone, ursodiol alone, low-calorie diet with ursodiol, and untreated control. Ursodiol did not show any therapeutic benefit above the diet alone. However, this study had several notable limitations including small sample size overall and per study group, lack of randomization, and inclusion of children without histological or radiological confirmation of NAFLD. Adult studies evaluating ursodiol have shown improvements in liver-related outcomes associated with NAFLD; therefore additional studies are needed in the pediatric population to assess its therapeutic potential. 20.5.2.3 Other Treatments Orlistat is a weight loss medication that inhibits the enzyme responsible for fat absorption in the gut and is approved for use in adolescents [5]. Harrison et al. tested the effect of calorie restriction (1400 Kcal/day) and vitamin E (800 IU/day) with and without orlistat (120 mg/TID) on 50 adults with biopsy proven NASH for 36 weeks [95]. The two groups did not significantly differ in terms of changes in liver enzymes or histology. Uncomfortable side effects of this medication related to the increased concentration of fat in the stool may also limit their use in the pediatric population. Although there are no well-defined guidelines regarding the use of statins for management of NAFLD, statins are considered to be safe and without any added risk for hepatotoxicity when used to manage dyslipidemia [96]. Use of statins has been shown to improve hepatic fat content, normalize liver function abnormalities, and reduce cardiovascular events in some pilot studies in adults [97–99]. There are no clinical trials to date in children evaluating the efficacy of statins to manage NAFLD. Probiotics are a newly emergent therapy for several gastrointestinal-related disorders. In the treatment of NAFLD, probiotics may be beneficial by helping attenuate immune-related injury to hepatocytes. Vajro et al. tested the effect of 8 weeks on probiotic therapy (lactobacillus rhamnosus strain GG) on 20 children with US-confirmed fatty liver [100]. In comparison to those in the placebo group, those on probiotic therapy had a significant reduction in ALT and antipeptidoglycan-polysaccharide antibodies even after adjusting for changes in body mass index and visceral fat. However, there were no significant changes in liver echogenicity. More research in the pediatric population is necessary to evaluate the effects of this promising therapy on liver-related outcomes in NAFLD.

20.6 CONCLUSION NAFLD has become the most common chronic pediatric liver disease in the developed world in recent years. Emerging epidemiological evidence suggests that it disproportionately affects males, Hispanics, Asians, and those with insulin resistance and metabolic disease. NAFLD is particularly alarming in the pediatric population because long-term exposure and buildup of fat in the liver promotes hepatocellular injury and fibrosis, which can lead to significant morbidity and mortality [7]. Indeed, NAFLD in childhood is strongly associated with cirrhosis, hepatocellular carcinoma, end-stage liver disease, as well as CVD and metabolic syndrome in adulthood. Although the pathophysiological mechanisms leading to hepatocellular injury and subsequent cell death have yet to be fully elucidated, increased concentrations of hepatic lipid accrual have been shown to cause cellular dysfunction through oxidative processes. New imaging modalities, such as MRS and elastography, hold promise for the diagnosis and management of pediatric NAFLD. Nonetheless, little guidance is available on how to best screen, diagnose, and treat children at risk of developing and diagnosed with this condition. Given the increased risk of morbidity and all-cause mortality associated with NAFLD, more research in the pediatric population is urgently needed to address this emerging and evergrowing health threat.

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REFERENCES

265

References [1] Ogden CL, Carroll MD, Curtin LR, McDowell MA, Tabak CJ, Flegal KM. Prevalence of overweight and obesity in the United States, 1999-2004. JAMA 2006;295(13):1549–55. [2] Ogden CL, Carroll MD, Kit BK, Flegal KM. Prevalence of obesity and trends in body mass index among US children and adolescents, 19992010. JAMA 2012;307(5):483–90. [3] Ogden CL, Carroll MD, Kit BK, Flegal KM. Prevalence of childhood and adult obesity in the United States, 2011-2012. JAMA 2014;311 (8):806–14. [4] Widhalm K, Ghods E. Nonalcoholic fatty liver disease: a challenge for pediatricians. Int J Obes (Lond) 2010;34(10):1451–67. [5] Temple JL, Cordero P, Li J, Nguyen V, Oben JA. A guide to non-alcoholic fatty liver disease in childhood and adolescence. Int J Mol Sci 2016;17 (6)E947. [6] Schwimmer JB, Deutsch R, Kahen T, Lavine JE, Stanley C, Behling C. Prevalence of fatty liver in children and adolescents. Pediatrics 2006;118 (4):1388–93. [7] Browning JD, Horton JD. Molecular mediators of hepatic steatosis and liver injury. J Clin Invest 2004;114(2):147–52. [8] Schwimmer JB, Behling C, Newbury R, Deutsch R, Nievergelt C, Schork NJ, Lavine JE. Histopathology of pediatric nonalcoholic fatty liver disease. Hepatology 2005;42(3):641–9. [9] Feldstein AE, Charatcharoenwitthaya P, Treeprasertsuk S, Benson JT, Enders FB, Angulo P. The natural history of non-alcoholic fatty liver disease in children: a follow-up study for up to 20 years. Gut 2009;58(11):1538–44. [10] Molleston JP, White F, Teckman J, Fitzgerald JF. Obese children with steatohepatitis can develop cirrhosis in childhood. Am J Gastroenterol 2002;97:2460–2. [11] Vajro P, Lenta S, Socha P, Dhawan A, McKiernan P, Baumann U, Durmaz O, Lacaille F, McLin V, Nobili V. Diagnosis of nonalcoholic fatty liver disease in children and adolescents: position paper of the ESPGHAN hepatology committee. J Pediatr Gastroenterol Nutr 2012;54(5):700–13. [12] Suzuki D, Hashimoto E, Kaneda K, Tokushige K, Shiratori K. Liver failure caused by non-alcoholic steatohepatitis in an obese young male. J Gastroenterol Hepatol 2005;20:327–9. [13] Riley MR, Bass NM, Rosenthal P, Merriman RB. Underdiagnosis of pediatric obesity and underscreening for fatty liver disease and metabolic syndrome by pediatricians and pediatric subspecialists. J Pediatr 2005;147(6):839–42. [14] Rinella ME. Nonalcoholic fatty liver disease: a systematic review. JAMA 2015;313(22):2263–73. [15] Yasuda M, Shimizu I, Shiba M, Ito S. Suppressive effects of estradiol on dimethylnitrosamine-induced fibrosis of the liver in rats. Hepatology 1999;29(3):719–27. [16] Kozlov AV, Duvigneau JC, Hyatt TC, Raju R, Behling T, Hartl RT, Staniek K, Miller I, Gregor W, Redl H, Chaudry IH. Effect of estrogen on mitochondrial function and intracellular stress markers in rat liver and kidney following trauma-hemorrhagic shock and prolonged hypotension. Mol Med 2010;16(7–8):254–61. [17] Hanada S, Snider NT, Brunt EM, Hollenberg PF, Omary MB. Gender dimorphic formation of mouse Mallory-Denk bodies and the role of xenobiotic metabolism and oxidative stress. Gastroenterology 2010;138(4):1607–17. [18] Goh GB, Pagadala MR, Dasarathy J, Unalp-Arida A, Pai RK, Yerian L, Khiyami A, Sourianarayanane A, Sargent R, Hawkins C, Dasarathy S, McCullough AJ. Age impacts ability of aspartate-alanine aminotransferase ratio to predict advanced fibrosis in nonalcoholic fatty liver disease. Dig Dis Sci 2015;60(6):1825–31. [19] Suzuki A, Abdelmalek MF, Schwimmer JB, Lavine JE, Scheimann AO, Unalp-Arida A, Yates KP, Sanyal AJ, Guy CD, Diehl AM, Network NSCR. Association between puberty and features of nonalcoholic fatty liver disease. Clin Gastroenterol Hepatol 2012;10(7):786–94. [20] Hewitt KN, Pratis K, Jones ME, Simpson ER. Estrogen replacement reverses the hepatic steatosis phenotype in the male aromatase knockout mouse. Endocrinology 2004;145(4):1842–8. [21] Potau N, Ibañez L, Riqu
e S, Carrascosa A. Pubertal changes in insulin secretion and peripheral insulin sensitivity. Horm Res 1997;48(5):219–26. [22] Moran A, Jacobs Jr. DR, Steinberger J, Steffen LM, Pankow JS, Hong CP, Sinaiko AR. Changes in insulin resistance and cardiovascular risk during adolescence: establishment of differential risk in males and females. Circulation 2008;117(18):2361–8. [23] Barlow SE, Committee E. Expert committee recommendations regarding the prevention, assessment, and treatment of child and adolescent overweight and obesity: summary report. Pediatrics 2007;120(Suppl 4):S164–92. [24] Dowla S, Aslibekyan S, Goss A, Fontaine K, Ashraf AP. Dyslipidemia is associated with pediatric nonalcoholic fatty liver disease. J Clin Lipidol 2018;S1933–2874. [25] Pan JJ, Fallon MB. Gender and racial differences in nonalcoholic fatty liver disease. World J Hepatol 2014 May 27;6(5):274–83. https://doi.org/ 10.4254/wjh.v6.i5.274 Review. PubMed PMID: 24868321; PubMed Central PMCID: PMC4033285. [26] Sumner AE. Ethnic differences in triglyceride levels and high-density lipoprotein lead to underdiagnosis of the metabolic syndrome in black children and adults. J Pediatr 2009;155(3). S7.e7-11. [27] Manco M, Marcellini M, Devito R, Comparcola D, Sartorelli MR, Nobili V. Metabolic syndrome and liver histology in paediatric non-alcoholic steatohepatitis. Int J Obes (Lond) 2008;32(2):381–7. [28] Schwimmer JB, Deutsch R, Rauch JB, Behling C, Newbury R, Lavine JE. Obesity, insulin resistance, and other clinicopathological correlates of pediatric nonalcoholic fatty liver disease. J Pediatr 2003;143(4):500–5. [29] Schwimmer JB, Pardee PE, Lavine JE, Blumkin AK, Cook S. Cardiovascular risk factors and the metabolic syndrome in pediatric nonalcoholic fatty liver disease. Circulation 2008;118(3):277–83. [30] Targher G, Arcaro G. Non-alcoholic fatty liver disease and increased risk of cardiovascular disease. Atherosclerosis 2007;191(2):235–40. [31] Stepanova M, Younossi ZM. Independent association between nonalcoholic fatty liver disease and cardiovascular disease in the US population. Clin Gastroenterol Hepatol 2012;10(6):646–50. [32] Lazo M, Hernaez R, Bonekamp S, Kamel IR, Brancati FL, Guallar E, Clark JM. Non-alcoholic fatty liver disease and mortality among US adults: prospective cohort study. BMJ 2011;343:d6891. [33] Alkhouri N, Tamimi TA, Yerian L, Lopez R, Zein NN, Feldstein AE. The inflamed liver and atherosclerosis: a link between histologic severity of nonalcoholic fatty liver disease and increased cardiovascular risk. Dig Dis Sci 2010;55(9):2644–50.

IV. CONSEQUENCES

266

20. NONALCOHOLIC FATTY LIVER DISEASE IN CHILDREN WITH OBESITY

[34] Nobili V, Alkhouri N, Bartuli A, Manco M, Lopez R, Alisi A, Feldstein AE. Severity of liver injury and atherogenic lipid profile in children with nonalcoholic fatty liver disease. Pediatr Res 2010;67(6):665–70. [35] Alkhouri N, Carter-Kent C, Elias M, Feldstein AE. Atherogenic dyslipidemia and cardiovascular risk in children with nonalcoholic fatty liver disease. Clin Lipidol 2011;6(3):305–14. [36] Cali AM, Zern TL, Taksali SE, de Oliveira AM, Dufour S, Otvos JD, Caprio S. Intrahepatic fat accumulation and alterations in lipoprotein composition in obese adolescents: a perfect proatherogenic state. Diabetes Care 2007;30(12):3093–8. [37] Kleiner DE, Brunt EM, Van Natta M, Behling C, Contos MJ, Cummings OW, Ferrell LD, Liu YC, Torbenson MS, Unalp-Arida A, Yeh M, McCullough AJ, Sanyal AJ, Nonalcoholic Steatohepatitis Clinical Research Network. Design and validation of a histological scoring system for nonalcoholic fatty liver disease. Hepatology 2005;41(6):1313–21. [38] Day CP, James OF. Steatohepatitis: a tale of two “hits”? Gastroenterology 1998;114(4):842–5. [39] Lewis GF, Carpentier A, Adeli K, Giacca A. Disordered fat storage and mobilization in the pathogenesis of insulin resistance and type 2 diabetes. Endocr Rev 2002;23(2):201–29. [40] Shimomura I, Bashmakov Y, Ikemoto S, Horton JD, Brown MS, Goldstein JL. Insulin selectively increases SREBP-1c mRNA in the livers of rats with streptozotocin-induced diabetes. Proc Natl Acad Sci U S A 1999;96(24):13656–61. [41] Foretz M, Guichard C, Ferr
e P, Foufelle F. Sterol regulatory element binding protein-1c is a major mediator of insulin action on the hepatic expression of glucokinase and lipogenesis-related genes. Proc Natl Acad Sci U S A 1999;96(22):12737–42. [42] Yamashita H, Takenoshita M, Sakurai M, Bruick RK, Henzel WJ, Shillinglaw W, Arnot D, Uyeda K. A glucose-responsive transcription factor that regulates carbohydrate metabolism in the liver. Proc Natl Acad Sci U S A 2001;98(16):9116–21. [43] Fon Tacer K, Rozman D. Nonalcoholic fatty liver disease: focus on lipoprotein and lipid deregulation. J Lipids 2011;2011:783976. [44] Fujita K, Nozaki Y, Wada K, Yoneda M, Fujimoto Y, Fujitake M, Endo H, Takahashi H, Inamori M, Kobayashi N, Kirikoshi H, Kubota K, Saito S, Nakajima A. Dysfunctional very-low-density lipoprotein synthesis and release is a key factor in nonalcoholic steatohepatitis pathogenesis. Hepatology 2009;50(3):772–80. [45] García-Ruiz C, Colell A, Morales A, Kaplowitz N, Fernández-Checa JC. Role of oxidative stress generated from the mitochondrial electron transport chain and mitochondrial glutathione status in loss of mitochondrial function and activation of transcription factor nuclear factorkappa B: studies with isolated mitochondria and rat hepatocytes. Mol Pharmacol 1995;48(5):825–34. [46] P
erez-Carreras M, Del Hoyo P, Martín MA, Rubio JC, Martín A, Castellano G, Colina F, Arenas J, Solis-Herruzo JA. Defective hepatic mitochondrial respiratory chain in patients with nonalcoholic steatohepatitis. Hepatology 2003;38(4):999–1007. [47] Esterbauer H, Schaur RJ, Zollner H. Chemistry and biochemistry of 4-hydroxynonenal, malonaldehyde and related aldehydes. Free Radic Biol Med 1991;11(1):81–128. [48] Nobili V, Carpino G, Alisi A, Franchitto A, Alpini G, De Vito R, Onori P, Alvaro D, Gaudio E. Hepatic progenitor cells activation, fibrosis, and adipokines production in pediatric nonalcoholic fatty liver disease. Hepatology 2012;56(6):2142–53. [49] Stienstra R, Saudale F, Duval C, Keshtkar S, Groener JE, van Rooijen N, Staels B, Kersten S, M€ uller M. Kupffer cells promote hepatic steatosis via interleukin-1beta-dependent suppression of peroxisome proliferator-activated receptor alpha activity. Hepatology 2010;51(2):511–22. [50] De Vito R, Alisi A, Masotti A, Ceccarelli S, Panera N, Citti A, Salata M, Valenti L, Feldstein AE, Nobili V. Markers of activated inflammatory cells correlate with severity of liver damage in children with nonalcoholic fatty liver disease. Int J Mol Med 2012;30(1):49–56. [51] Ferreyra Solari NE, Inzaugarat ME, Baz P, De Matteo E, Lezama C, Galoppo M, Galoppo C, Cherñavsky AC. The role of innate cells is coupled to a Th1-polarized immune response in pediatric nonalcoholic steatohepatitis. J Clin Immunol 2012;32(3):611–21. [52] Hyysalo J, M€ annist€ o VT, Zhou Y, Arola J, K€arj€a V, Leivonen M, Juuti A, Jaser N, Lallukka S, K€akel€a P, Venesmaa S, Simonen M, Saltevo J, Moilanen L, Korpi-Hy€ ovalti E, Kein€anen-Kiukaanniemi S, Oksa H, Orho-Melander M, Valenti L, Fargion S, Pihlajam€aki J, Peltonen M, YkiJ€ arvinen H. A population-based study on the prevalence of NASH using scores validated against liver histology. J Hepatol 2013;60(4):839–46. [53] Fracanzani AL, Valenti L, Bugianesi E, Andreoletti M, Colli A, Vanni E, Bertelli C, Fatta E, Bignamini D, Marchesini G, Fargion S. Risk of severe liver disease in nonalcoholic fatty liver disease with normal aminotransferase levels: a role for insulin resistance and diabetes. Hepatology 2008;48(3):792–8. [54] Mofrad P, Contos MJ, Haque M, Sargeant C, Fisher RA, Luketic VA, Sterling RK, Shiffman ML, Stravitz RT, Sanyal AJ. Clinical and histologic spectrum of nonalcoholic fatty liver disease associated with normal ALT values. Hepatology 2003;37(6):1286–92. [55] Vernon G, Baranova A, Younossi ZM. Systematic review: the epidemiology and natural history of non-alcoholic fatty liver disease and nonalcoholic steatohepatitis in adults. Aliment Pharmacol Ther 2011;34(3):274–85. [56] Maximos M, Bril F, Portillo Sanchez P, Lomonaco R, Orsak B, Biernacki D, Suman A, Weber M, Cusi K. The role of liver fat and insulin resistance as determinants of plasma aminotransferase elevation in nonalcoholic fatty liver disease. Hepatology 2015;61(1):153–60. [57] Lee SS, Park SH. Radiologic evaluation of nonalcoholic fatty liver disease. World J Gastroenterol 2014;20(23):7392–402. [58] Lee SS, Park SH, Kim HJ, Kim SY, Kim MY, Kim DY, Suh DJ, Kim KM, Bae MH, Lee JY, Lee SG, Yu ES. Non-invasive assessment of hepatic steatosis: prospective comparison of the accuracy of imaging examinations. J Hepatol 2010;52(4):579–85. [59] Saadeh S, Younossi ZM, Remer EM, Gramlich T, Ong JP, Hurley M, Mullen KD, Cooper JN, Sheridan MJ. The utility of radiological imaging in nonalcoholic fatty liver disease. Gastroenterology 2002;123(3):745–50. [60] van Werven JR, Marsman HA, Nederveen AJ, Smits NJ, ten Kate FJ, van Gulik TM, Stoker J. Assessment of hepatic steatosis in patients undergoing liver resection: comparison of US, CT, T1-weighted dual-echo MR imaging, and point-resolved 1H MR spectroscopy. Radiology 2010;256(1):159–68. [61] de Moura Almeida A, Cotrim HP, Barbosa DB, de Athayde LG, Santos AS, Bitencourt AG, de Freitas LA, Rios A, Alves E. Fatty liver disease in severe obese patients: diagnostic value of abdominal ultrasound. World J Gastroenterol 2008;14(9):1415–8. [62] Palmentieri B, de Sio I, La Mura V, Masarone M, Vecchione R, Bruno S, Torella R, Persico M. The role of bright liver echo pattern on ultrasound B-mode examination in the diagnosis of liver steatosis. Dig Liver Dis 2006;38(7):485–9. [63] Hepburn MJ, Vos JA, Fillman EP, Lawitz EJ. The accuracy of the report of hepatic steatosis on ultrasonography in patients infected with hepatitis C in a clinical setting: a retrospective observational study. BMC Gastroenterol 2005;5:14. [64] Wu S, Tu R, Liu G, Huang L, Guan Y, Zheng E. Focal fatty sparing usually does not arise in preexisting nonalcoholic diffuse homogeneous fatty liver. J Ultrasound Med 2014;33(8):1447–52.

IV. CONSEQUENCES

REFERENCES

267

[65] Yilmaz Y, Ergelen R, Akin H, Imeryuz N. Noninvasive detection of hepatic steatosis in patients without ultrasonographic evidence of fatty liver using the controlled attenuation parameter evaluated with transient elastography. Eur J Gastroenterol Hepatol 2013;25(11):1330–4. [66] Deng J, Fishbein MH, Rigsby CK, Zhang G, Schoeneman SE, Donaldson JS. Quantitative MRI for hepatic fat fraction and T2* measurement in pediatric patients with non-alcoholic fatty liver disease. Pediatr Radiol 2014;44(11):1379–87. [67] Urdzik J, Bjerner T, Wanders A, Weis J, Duraj F, Haglund U, Nor
en A. The value of pre-operative magnetic resonance spectroscopy in the assessment of steatohepatitis in patients with colorectal liver metastasis. J Hepatol 2012;56(3):640–6. [68] Roldan-Valadez E, Favila R, Martínez-López M, Uribe M, Ríos C, M
endez-Sánchez N. In vivo 3T spectroscopic quantification of liver fat content in nonalcoholic fatty liver disease: correlation with biochemical method and morphometry. J Hepatol 2010;53(4):732–7. [69] Raptis DA, Fischer MA, Graf R, Nanz D, Weber A, Moritz W, Tian Y, Oberkofler CE, Clavien PA. MRI: the new reference standard in quantifying hepatic steatosis? 61(1):117–27. [70] Hu HH, Kim HW, Nayak KS, Goran MI. Comparison of fat-water MRI and single-voxel MRS in the assessment of hepatic and pancreatic fat fractions in humans. Obesity (Silver Spring) 2010;18(4):841–7. [71] Loomba R, Sirlin CB, Schwimmer JB, Lavine JE. Advances in pediatric nonalcoholic fatty liver disease. Hepatology 2009;50(4):1282–93. [72] Ochi H, Hirooka M, Koizumi Y, Miyake T, Tokumoto Y, Soga Y, Tada F, Abe M, Hiasa Y, Onji M. Real-time tissue elastography for evaluation of hepatic fibrosis and portal hypertension in nonalcoholic fatty liver diseases. Hepatology 2012;56(4):1271–8. [73] Nobili V, Vizzutti F, Arena U, Abraldes JG, Marra F, Pietrobattista A, Fruhwirth R, Marcellini M, Pinzani M. Accuracy and reproducibility of transient elastography for the diagnosis of fibrosis in pediatric nonalcoholic steatohepatitis. Hepatology 2008;48(2):442–8. [74] Orlacchio A, Bolacchi F, Antonicoli M, Coco I, Costanzo E, Tosti D, Francioso S, Angelico M, Simonetti G. Liver elasticity in NASH patients evaluated with real-time elastography (RTE). Ultrasound Med Biol 2012;38(4):537–44. [75] Wong VW, Vergniol J, Wong GL, Foucher J, Chan HL, Le Bail B, Choi PC, Kowo M, Chan AW, Merrouche W, Sung JJ, de L
edinghen V. Diagnosis of fibrosis and cirrhosis using liver stiffness measurement in nonalcoholic fatty liver disease. Hepatology 2010;51(2):454–62. [76] Palmeri ML, Wang MH, Rouze NC, Abdelmalek MF, Guy CD, Moser B, Diehl AM, Nightingale KR. Noninvasive evaluation of hepatic fibrosis using acoustic radiation force-based shear stiffness in patients with nonalcoholic fatty liver disease. J Hepatol 2011;55(3):666–72. [77] Chen J, Talwalkar JA, Yin M, Glaser KJ, Sanderson SO, Ehman RL. Early detection of nonalcoholic steatohepatitis in patients with nonalcoholic fatty liver disease by using MR elastography. Radiology 2011;259(3):749–56. [78] Chalasani N, Younossi Z, Lavine JE, Diehl AM, Brunt EM, Cusi K, Charlton M, Sanyal AJ. The diagnosis and management of non-alcoholic fatty liver disease: practice guideline by the American Association for the Study of Liver Diseases, American College of Gastroenterology, and the American Gastroenterological Association. Hepatology 2012;55(6):2005–23. [79] Tock L, Prado WL, Caranti DA, Cristofalo DM, Lederman H, Fisberg M, Siqueira KO, Stella SG, Antunes HK, Cintra IP, Tufik S, de Mello MT, D^ amaso AR. Nonalcoholic fatty liver disease decrease in obese adolescents after multidisciplinary therapy. Eur J Gastroenterol Hepatol 2006;18 (12):1241–5. [80] Nobili V, Manco M, Devito R, Ciampalini P, Piemonte F, Marcellini M. Effect of vitamin E on aminotransferase levels and insulin resistance in children with non-alcoholic fatty liver disease. Aliment Pharmacol Ther 2006;24(1112):1553–61. [81] Nobili V, Manco M, Devito R, Di Ciommo V, Comparcola D, Sartorelli MR, Piemonte F, Marcellini M, Angulo P. Lifestyle intervention and antioxidant therapy in children with nonalcoholic fatty liver disease: a randomized, controlled trial. Hepatology 2008;48(1):119–28. [82] Reinehr T, Schmidt C, Toschke AM, Andler W. Lifestyle intervention in obese children with non-alcoholic fatty liver disease: 2-year follow-up study. Arch Dis Child 2009;94(6):437–42. [83] Ramon-Krauel M, Salsberg SL, Ebbeling CB, Voss SD, Mulkern RV, Apura MM, Cooke EA, Sarao K, Jonas MM, Ludwig DS. A low-glycemicload versus low-fat diet in the treatment of fatty liver in obese children. Child Obes 2013;9(3):252–60. [84] Dowla S, Pendergrass M, Bolding M, Gower B, Fontaine K, Ashraf A, Soleymani T, Morrison S, Goss A. Effectiveness of a carbohydrate restricted diet to treat non-alcoholic fatty liver disease in adolescents with obesity: trial design and methodology. Contemp Clin Trials 2018;68:95–101. [85] Mazza A, Fruci B, Garinis GA, Giuliano S, Malaguarnera R, Belfiore A. The role of metformin in the management of NAFLD. Exp Diabetes Res 2012;2012:716404. [86] Schwimmer JB, Middleton MS, Deutsch R, Lavine JE. A phase 2 clinical trial of metformin as a treatment for non-diabetic paediatric nonalcoholic steatohepatitis. Aliment Pharmacol Ther 2005;21(7):871–9. [87] Nadeau KJ, Ehlers LB, Zeitler PS, Love-Osborne K. Treatment of non-alcoholic fatty liver disease with metformin versus lifestyle intervention in insulin-resistant adolescents. Pediatr Diabetes 2009;10(1):5–13. [88] Nobili V, Marcellini M, Devito R, Ciampalini P, Piemonte F, Comparcola D, Sartorelli MR, Angulo P. NAFLD in children: A prospective clinical-pathological study and effect of lifestyle advice. Hepatology 2006;44(2):458–65. [89] Lavine JE, Schwimmer JB, van Natta ML, Molleston JP, Murray KF, Rosenthal P, Abrams SH, Scheimann AO, Sanyal AJ, Chalasani N, € Tonascia J, Unalp A, Clark JM, Brunt EM, Kleiner DE, Hoofnagle JH, Robuck PR, Nonalcoholic Steatohepatitis Clinical Research Network. Effect of vitamin E or metformin for treatment of nonalcoholic fatty liver disease in children and adolescents: the TONIC randomized controlled trial. JAMA 2011;305(16):1659–68. [90] Nissen SE, Wolski K. Effect of rosiglitazone on the risk of myocardial infarction and death from cardiovascular causes. N Engl J Med 2007;356 (24):2457–71. [91] Li J, Cordero P, Nguyen V, Oben JA. The role of vitamins in the pathogenesis of non-alcoholic fatty liver disease. Integr Med Insights 2016;11:19–25. [92] Sanyal AJ, Chalasani N, Kowdley KV, McCullough A, Diehl AM, Bass NM, Neuschwander-Tetri BA, Lavine JE, Tonascia J, Unalp A, Van Natta M, Clark J, Brunt EM, Kleiner DE, Hoofnagle JH, Robuck PR, NASH CRN. Pioglitazone, vitamin E, or placebo for nonalcoholic steatohepatitis. N Engl J Med 2010;362(18):1675–85. [93] Hoofnagle JH, van Natta ML, Kleiner DE, Clark JM, Kowdley KV, Loomba R, Neuschwander-Tetri BA, Sanyal AJ, Tonascia J, Non-alcoholic Steatohepatitis Clinical Research Network (NASH CRN). Vitamin E and changes in serum alanine aminotransferase levels in patients with non-alcoholic steatohepatitis. Aliment Pharmacol Ther 2013;38(2):134–43.

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20. NONALCOHOLIC FATTY LIVER DISEASE IN CHILDREN WITH OBESITY

[94] Vajro P, Franzese A, Valerio G, Iannucci MP, Aragione N. Lack of efficacy of ursodeoxycholic acid for the treatment of liver abnormalities in obese children. J Pediatr 2000;136(6):739–43. [95] Harrison SA, Fecht W, Brunt EM, Neuschwander-Tetri BA. Orlistat for overweight subjects with nonalcoholic steatohepatitis: A randomized, prospective trial. Hepatology 2009;49(1):80–6. [96] European Association for the Study of the Liver (EASL), European Association for the Study of Diabetes (EASD), European Association for the Study of Obesity(EASO). EASL-EASD-EASO Clinical Practice Guidelines for the management of non-alcoholic fatty liver disease. J Hepatol 2016;64(6):1388–402. [97] Athyros VG, Tziomalos K, Gossios TD, Griva T, Anagnostis P, Kargiotis K, Pagourelias ED, Theocharidou E, Karagiannis A, Mikhailidis DP. GREACE Study Collaborative Group. Safety and efficacy of long-term statin treatment for cardiovascular events in patients with coronary heart disease and abnormal liver tests in the Greek atorvastatin and coronary heart disease evaluation (GREACE) study: a post-hoc analysis. Lancet 2010;376(9756):1916–22. [98] Kargiotis K, Katsiki N, Athyros VG, Giouleme O, Patsiaoura K, Katsiki E, Mikhailidis DP, Karagiannis A. Effect of rosuvastatin on nonalcoholic steatohepatitis in patients with metabolic syndrome and hypercholesterolaemia: a preliminary report. Curr Vasc Pharmacol 2014;12(3):505–11. [99] Hyogo H, Tazuma S, Arihiro K, Iwamoto K, Nabeshima Y, Inoue M, Ishitobi T, Nonaka M, Chayama K. Efficacy of atorvastatin for the treatment of nonalcoholic steatohepatitis with dyslipidemia. Metabolism 2008;57(12):1711–8. [100] Vajro P, Mandato C, Licenziati MR, Franzese A, Vitale DF, Lenta S, Caropreso M, Vallone G, Meli R. Effects of lactobacillus rhamnosus strain GG in pediatric obesity-related liver disease. J Pediatr Gastroenterol Nutr 2011;52(6):740–3.

IV. CONSEQUENCES