Nonalcoholic fatty liver disease risk and histologic severity are associated with genetic polymorphisms in children.

BACKGROUND AND AIMS: NAFLD is the most common chronic liver disease in children. Large pediatric studies identifying single nucleotide polymorphisms (SNPs) associated with risk and histologic severity of NAFLD are limited. Study aims included investigating SNPs associated with risk for NAFLD using family trios and association of candidate alleles with histologic severity. APPROACH AND RESULTS: Children with biopsy-confirmed NAFLD were enrolled from the NASH Clinical Research Network. The Expert Pathology Committee reviewed liver histology. Genotyping was conducted with allele-specific primers for 60 candidate SNPs. Parents were enrolled for trio analysis. To assess risk for NAFLD, the transmission disequilibrium test was conducted in trios. Among cases, regression analysis assessed associations with histologic severity. A total of 822 children with NAFLD had mean age 13.2 years (SD 2.7) and mean ALT 101 U/L (SD 90). PNPLA3 (rs738409) demonstrated the strongest risk ( p = 2.24 × 10 -14 ) for NAFLD. Among children with NAFLD, stratifying by PNPLA3 s738409 genotype, the variant genotype associated with steatosis ( p = 0.005), lobular ( p = 0.03) and portal inflammation ( p = 0.002). Steatosis grade associated with TM6SF2 ( p = 0.0009), GCKR ( p = 0.0032), PNPLA3 rs738409 ( p = 0.0053), and MTTP ( p = 0.0051). Fibrosis stage associated with PARVB rs6006473 ( p = 0.0001), NR1I2 ( p = 0.0021), ADIPOR2 ( p = 0.0038), and OXTR ( p = 0.0065). PNPLA3 rs738409 ( p = 0.0002) associated with borderline zone 1 NASH. CONCLUSIONS: This study demonstrated disease-associated SNPs in children with NAFLD. In particular, rs6006473 was highly associated with severity of fibrosis. These hypothesis-generating results support future mechanistic studies of development of adverse outcomes such as fibrosis and generation of therapeutic targets for NAFLD in children.

Prevalence of Elevated ALT in Adolescents in the US 2011-2018.

OBJECTIVE: The objective of this study is to characterize suspected nonalcoholic fatty liver disease (NAFLD) using elevated alanine aminotransferase (ALT) in a diverse and nationally representative cohort of adolescents and to characterize higher ALT elevation in adolescents with obesity. METHODS: Data from the National Health and Nutrition Examination Survey 2011-2018 were analyzed for adolescents 12-19 years. Participants with causes for elevated ALT other than NAFLD were excluded. Race and ethnicity, sex, body mass index (BMI), and ALT were examined. Elevated ALT was defined as >22 U/L (females) and >26 U/L (males) using the biologic upper normal limit (ULN). Elevated ALT thresholds up to 2X-ULN were examined among adolescents with obesity. Multivariable logistic regression was used to determine the association of race/ethnicity and elevated ALT, adjusting for age, sex, and BMI. RESULTS: Prevalence of elevated ALT in adolescents was 16.5% overall and 39.5% among those with obesity. For White, Hispanic, and Asian adolescents, prevalence was 15.8%, 21.8%, and 16.5% overall, 12.8%, 17.7%, and 27.0% in those with overweight, and 43.0%, 43.5%, and 43.1% in those with obesity, respectively. Prevalence was much lower in Black adolescents (10.7% overall, 8.4% for overweight, 20.7% for obesity). Prevalence of ALT at 2X-ULN was 6.6% in adolescents with obesity. Hispanic ethnicity, age, male sex, and higher BMI were independent predictors of elevated ALT. CONCLUSIONS: Prevalence of elevated ALT in U.S. adolescents is high, affecting 1 in 6 adolescents during 2011-2018. The risk is highest in Hispanic adolescents. Asian adolescents with elevated BMI may comprise an emerging risk group for elevated ALT.

An Open Label, Randomized, Multicenter Study of Elafibranor in Children With Nonalcoholic Steatohepatitis.

OBJECTIVES: Nonalcoholic fatty liver disease is the most common chronic liver disease in children. Elafibranor, a dual peroxisome proliferator-activated receptor α/δ agonist, has been proposed as a treatment for nonalcoholic steatohepatitis (NASH). The aims were to (1) describe pharmacokinetics (PK), safety, and tolerability of oral elafibranor at 2 doses (80 and 120 mg) in children 8-17 years and (2) assess changes in aminotransferases. METHODS: Children with NASH were randomized to open-label elafibranor 80 mg or 120 mg daily for 12 weeks. The intent-to-treat analysis included all participants who received at least 1 dose. Standard descriptive statistics and PK analyses were performed. RESULTS: Ten males [mean 15.1 years, standard deviation (SD) 2.2] with NASH were randomized to 80 mg (n = 5) or 120 mg (n = 5). Baseline mean alanine aminotransferase (ALT) was 82 U/L (SD 13) and 87 U/L (SD 20) for 80 mg and 120 mg groups, respectively. Elafibranor was rapidly absorbed and well tolerated. Elafibranor plasma exposure increased between the 80 mg and 120 mg dose with a 1.9- and 1.3-fold increase in median Cmax and AUC 0-24 , respectively. End of treatment mean ALT was 52 U/L (SD 20) for the 120 mg group, with a relative mean ALT change from baseline of -37.4% (SD 23.8%) at 12 weeks. CONCLUSIONS: Once daily dosing of elafibranor was well tolerated in children with NASH. There was a 37.4% relative reduction from mean baseline ALT in the 120 mg group. Decreasing ALT may be associated with improvement in liver histology, thus could be considered a surrogate for histology in early phase trials. These results may support further exploration of elafibranor in children with NASH.

Incidence of Depression and Anxiety in a Cohort of Adolescents With Nonalcoholic Fatty Liver Disease.

OBJECTIVES: To determine the incidence of clinically diagnosed depression and anxiety in adolescents with nonalcoholic fatty liver disease (NAFLD). METHODS: This was a prospective, longitudinal cohort study between January 1, 2012 and July 1, 2018 conducted in a Children's Hospital Pediatric Gastroenterology Clinic. Participants included adolescents 12 to 17 years old at baseline with biopsy-confirmed NAFLD. The primary outcomes were having depression and/or anxiety based upon a clinical diagnosis established by a physician or psychologist. The rates of depression and anxiety were measured at baseline and longitudinally throughout follow-up. RESULTS: A total of 160 adolescents with NAFLD were followed for a mean of 3.8 years. At baseline, 8.1% had a diagnosis of depression. During follow-up, an additional 9.5% (95% confidence interval, 4.7-14.3) developed depression. The incidence density of depression was 27 new cases per 1000 person-years at risk. In adolescents with NAFLD, 6.3% had anxiety at baseline and 6.7% (95% confidence interval, 2.6-10.7) developed anxiety during follow-up. The incidence density of anxiety was 18 new cases per 1000 person-years at risk. The change in alanine aminotransferase was significantly worse for adolescents with NAFLD who developed depression compared to those who did not develop depression (P < 0.01). CONCLUSIONS: Adolescents with NAFLD had a high incidence of clinically diagnosed depression and anxiety. The rates were higher than expected relative to the available data in the general population. Addressing this mental health burden will require efforts at both the patient level and the systems level.

Dairy Fat Intake, Plasma Pentadecanoic Acid, and Plasma Iso-heptadecanoic Acid Are Inversely Associated With Liver Fat in Children.

OBJECTIVES: We sought to evaluate the relevance of pediatric dairy fat recommendations for children at risk for nonalcoholic fatty liver disease (NAFLD) by studying the association between dairy fat intake and the amount of liver fat. The effects of dairy fat may be mediated by odd chain fatty acids (OCFA), such as pentadecanoic acid (C15:0), and monomethyl branched chain fatty acids (BCFA), such as iso-heptadecanoic acid (iso-C17:0). Therefore, we also evaluated the association between plasma levels of OCFA and BCFA with the amount of liver fat. METHODS: Observational, cross-sectional, community-based sample of 237 children ages 8 to 17. Dairy fat intake was assessed by 3 24-hour dietary recalls. Plasma fatty acids were measured by gas chromatography-mass spectrometry. Main outcome was hepatic steatosis measured by whole liver magnetic resonance imaging proton density fat fraction (MRI-PDFF). RESULTS: Median dairy fat intake was 10.6 grams/day (range 0.0--44.5 g/day). Median liver MRI-PDFF was 4.5% (range 0.9%-45.1%). Dairy fat intake was inversely correlated with liver MRI-PDFF (r = -0.162; P = .012). In multivariable log linear regression, plasma C15:0 and iso-C17:0 were inverse predictors of liver MRI-PDFF (B = -0.247, P = 0.048; and B = -0.234, P = 0.009). CONCLUSIONS: Dairy fat intake, plasma C15:0, and plasma iso-C17:0 were inversely correlated with hepatic steatosis in children. These hypothesis-generating findings should be tested through clinical trials to better inform dietary guidelines.

Microbiome Signatures Associated With Steatohepatitis and Moderate to Severe Fibrosis in Children With Nonalcoholic Fatty Liver Disease.

BACKGROUND & AIMS: The intestinal microbiome might affect the development and severity of nonalcoholic fatty liver disease (NAFLD). We analyzed microbiomes of children with and without NAFLD. METHODS: We performed a prospective, observational, cross-sectional study of 87 children (age range, 8-17 years) with biopsy-proven NAFLD and 37 children with obesity without NAFLD (controls). Fecal samples were collected and microbiome composition and functions were assessed using 16S ribosomal RNA amplicon sequencing and metagenomic shotgun sequencing. Microbial taxa were identified using zero-inflated negative binomial modeling. Genes contributing to bacterial pathways were identified using gene set enrichment analysis. RESULTS: Fecal microbiomes of children with NAFLD had lower α-diversity than those of control children (3.32 vs 3.52, P = .016). Fecal microbiomes from children with nonalcoholic steatohepatitis (NASH) had the lowest α-diversity (control, 3.52; NAFLD, 3.36; borderline NASH, 3.37; NASH, 2.97; P = .001). High abundance of Prevotella copri was associated with more severe fibrosis (P = .036). Genes for lipopolysaccharide biosynthesis were enriched in microbiomes from children with NASH (P < .001). Classification and regression tree model with level of alanine aminotransferase and relative abundance of the lipopolysaccharide pathway gene encoding 3-deoxy-d-manno-octulosonate 8-phosphate-phosphatase identified patients with NASH with an area under the receiver operating characteristic curve value of 0.92. Genes involved in flagellar assembly were enriched in the fecal microbiomes of patients with moderate to severe fibrosis (P < .001). Classification and regression tree models based on level of alanine aminotransferase and abundance of genes encoding flagellar biosynthesis protein had good accuracy for identifying case children with moderate to severe fibrosis (area under the receiver operating characteristic curve, 0.87). CONCLUSIONS: In an analysis of fecal microbiomes of children with NAFLD, we associated NAFLD and NASH with intestinal dysbiosis. NAFLD and its severity were associated with greater abundance of genes encoding inflammatory bacterial products. Alterations to the intestinal microbiome might contribute to the pathogenesis of NAFLD and be used as markers of disease or severity.

Normal range for MR elastography measured liver stiffness in children without liver disease.

BACKGROUND: Magnetic resonance elastography (MRE) can determine the presence and stage of liver fibrosis. Data on normative MRE values, while reported in adults, are limited in children. PURPOSE: To determine the distribution of MRE-measured liver stiffness in children without liver disease. STUDY TYPE: Prospective, observational. POPULATION: Eighty-one healthy children (mean 12.6 ± 2.6 years, range 8-17 years). FIELD STRENGTH/SEQUENCE: 3.0T Signa HDxt, General Electric MR Scanner; 2D GRE MRE sequence. ASSESSMENT: History, examination, laboratory evaluation, and (MR) exams (proton density fat fraction, PDFF, and MRE) were performed. MR elastograms were analyzed manually at two reading centers and compared with each other for agreement and with published values in healthy adults and thresholds for fibrosis in adult and pediatric patients. STATISTICAL TESTS: Descriptive statistics, Bland-Altman analysis, t-test to compare hepatic stiffness values with reference standards. RESULTS: Stiffness values obtained at both reading centers were similar, without significant bias (P = 0.362) and with excellent correlation (intraclass correlation coefficient [ICC] = 0.782). Mean hepatic stiffness value for the study population was 2.45 ± 0.35 kPa (95th percentile 3.19 kPa), which was significantly higher than reported values for healthy adult subjects (2.10 ± 0.23 kPa, P < 0.001). In all, 74-85% of subjects had stiffness measurements suggestive of no fibrosis. DATA CONCLUSION: Mean liver stiffness measured with MRE in this cohort was significantly higher than that reported in healthy adults. Despite rigorous screening, some healthy children had stiffness measurements suggestive of liver fibrosis using current published thresholds. Although MRE has the potential to provide noninvasive assessment in patients with suspected hepatic disease, further refinement of this technology will help advance its use as a diagnostic tool for evidence of fibrosis in pediatric populations. LEVEL OF EVIDENCE: 1 Technical Efficacy: 5 J. Magn. Reson. Imaging 2020;51:919-927.

Evaluation of Quantitative Imaging Biomarkers for Early-phase Clinical Trials of Steatohepatitis in Adolescents.

OBJECTIVES: Early-phase pediatric nonalcoholic fatty liver disease (NAFLD) clinical trials are designed with noninvasive parameters to assess potential efficacy. Increasingly, these parameters include magnetic resonance imaging (MRI)-derived proton density fat fraction (PDFF) and MR elastography (MRE)-derived shear stiffness as biomarkers of hepatic steatosis and fibrosis, respectively. Understanding fluctuations in these measures is essential for calculating trial sample sizes, interpreting results, and planning clinical drug trials in children with NAFLD. Lack of such data in children constitutes a critical knowledge gap. Therefore, the primary aim of this study was to assess whole-liver MRI-PDFF change in adolescents with nonalcoholic steatohepatitis (NASH) over 12 weeks. METHODS: Adolescents 12 to 19 years with biopsy-proven NASH undergoing standard-of-care treatment were enrolled. Baseline and week-12 assessments of anthropometrics, transaminases, MRI-PDFF, and MRE stiffness were obtained. RESULTS: Fifteen adolescents were included (mean age 15.7 [SD 2.9] years). Hepatic MRI-PDFF was stable over 12 weeks (mean absolute change -0.8%, P = 0.24). Correlation between baseline and week-12 values of MRI-PDFF was high (ICC = 0.97, 95% CI 0.90-0.99). MRE stiffness was stable (mean percentage change 2.7%, P = 0.44); correlation between baseline and week-12 values was moderate (ICC = 0.47; 95% CI 0-0.79). Changes in weight, BMI, and aminotransferases were not statistically significant. CONCLUSION: In adolescents with NASH, fluctuations in hepatic MRI-PDFF and MRE stiffness over 12 weeks of standard-of-care were small. These data on the natural fluctuations in quantitative imaging biomarkers can serve as a reference for interventional trials in pediatric NASH and inform the interpretation and planning of clinical trials.

Prevalence of Nonalcoholic Fatty Liver Disease in Children with Obesity.

OBJECTIVES: To determine the prevalence of nonalcoholic fatty liver disease (NAFLD) in children with obesity because current estimates range from 1.7% to 85%. A second objective was to evaluate the diagnostic accuracy of alanine aminotransferase (ALT) for NAFLD in children with obesity. STUDY DESIGN: We evaluated children aged 9-17 years with obesity for the presence of NAFLD. Diseases other than NAFLD were excluded by history and laboratories. Hepatic steatosis was measured by liver magnetic resonance imaging proton density fat fraction. The diagnostic accuracy of ALT for detecting NAFLD was evaluated. RESULTS: The study included 408 children with obesity that had a mean age of 13.2 years and mean body mass index percentile of 98.0. The study population had a mean ALT of 32 U/L and median hepatic magnetic resonance imaging proton density fat fraction of 3.7%. The estimated prevalence of NAFLD was 26.0% (95% CI 24.2%-27.7%), 29.4% in male patients (CI 26.1%-32.7%) and 22.6% in female patients (CI 16.0%-29.1%). Optimal ALT cut-point was 42 U/L (47.8% sensitivity, 93.2% specificity) for male and 30 U/L (52.1% sensitivity, 88.8% specificity) for female patients. The classification and regression tree model with sex, ALT, and insulin had 80% diagnostic accuracy for NAFLD. CONCLUSIONS: NAFLD is common in children with obesity, but NAFLD and obesity are not concomitant. In children with obesity, NAFLD is present in nearly one-third of boys and one-fourth of girls.

Clinical Practice Approach to Nonalcoholic Fatty Liver Disease by Pediatric Gastroenterologists in the United States.

OBJECTIVES: Nonalcoholic fatty liver disease (NAFLD) is common; however, no information is available on how pediatric gastroenterologists in the United States manage NAFLD. Therefore, study objectives were to understand how pediatric gastroenterologists in the US approach the management of NAFLD, and to identify barriers to care for children with NAFLD. METHODS: We performed structured one-on-one interviews to ascertain each individual pediatric gastroenterologist's approach to the management of NAFLD in children. Responses were recorded from open-ended questions regarding screening for comorbidities, recommendations regarding nutrition, physical activity, medications, and perceived barriers to care. RESULTS: Response rate was 72.0% (486/675). Mean number of patients examined per week was 3 (standard deviation [SD] 3.5). Dietary intervention was recommended by 98.4% of pediatric gastroenterologists. Notably, 18 different dietary recommendations were reported. A majority of physicians provided targets for exercise frequency (72.6%, mean 5.6 days/wk, SD 1.6) and duration (69.9%, mean 40.2 minutes/session, SD 16.4). Medications were prescribed by 50.6%. Almost one-half of physicians (47.5%) screened for type 2 diabetes, dyslipidemia, and hypertension. Providers who spent more than 25 minutes at the initial visit were more likely to screen for comorbidities (P = 0.003). Barriers to care were reported by 92.8% with 29.0% reporting ≥3 barriers. CONCLUSIONS: The majority of US pediatric gastroenterologists regularly encounter children with NAFLD. Varied recommendations regarding diet and exercise highlight the need for prospective clinical trials. NAFLD requires a multidimensional approach with adequate resources in the home, community, and clinical setting.