1. Deborah Cohen, DCN, RD Assistant Professor UNM Nutrition Program
Pediatric Non-Alcoholic Fatty Liver Disease: Etiologies, Features, Treatment and Implications for the RD
Deborah Cohen, DCN, RD
Assistant Professor
UNM Nutrition Program

2. Objectives
Discuss the prevalence of pediatric non-alcoholic fatty liver disease (NAFLD) in the United States.
Identify the major risk factors that contribute to the development of NAFLD.
Review the biochemical and anthropometric features of pediatric NAFLD.
Discuss the recommendations for preventing and treating pediatric NAFLD, based on the current research.

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4. Non-alcoholic fatty liver disease (NAFLD)
Describes a spectrum of steatotic liver disease generally associated with obesity and metabolic syndrome
HTN, dyslipidemia, increased waist circumference, insulin resistance

5. Prevalence
Most common pediatric chronic liver disease in US and globally
2.6%-9.6%
Prevalence depends on population studied
Studies from Europe, Asia, America: Overweight and obese children 24%-77% (severe obesity: up to 90%)
More common in boys vs. girls

6. Study of Child and Adolescent Liver Epidemiology (SCALE)
Retrospective autopsy study (n=742, ages 2-19, , San Diego County)
Fatty liver (≥5% of hepatocytes containing macrovesicular fat) was present in 13% of subjects
Prevalence of fatty liver (adjusted for age, gender, race, and ethnicity) estimated to be 9.6%
Prevalence differed significantly by race and ethnicity
Asian: 10.2%; Black: 1.5%; Hispanic: 11.8%; White: 8.6%
The highest rate of fatty liver seen in obese children (38%).
Schwimmer et al. Prevalence of fatty liver in children and adolescents. Pediatrics. 2006;118(4):

7. Risk Factors Associated with NAFLD
Conditions with established association
Conditions with emerging association
Obesity
Type 2 diabetes mellitus
Dyslipidemia
Metabolic syndrome
Polycystic ovary syndrome
Hypothyroidism
Obstructive sleep apnea
Hypopituitarism
Hypogonadism
Pancreato-duodenal resection
Chalasani et al. 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):

8. Progression of fatty liver disease
Nonalcoholic fatty liver (NAFL)
Progression of hepatic steatosis with no evidence of hepatocellular injury.
Nonalcoholic steatohepatitis (NASH)
Presence of hepatic steatosis with hepatocellular injury (ballooning) with or without fibrosis.
NASH Cirrhosis

10. Pathogenesis
De novo lipogenesis (DNL) Role of Insulin Resistance “Two Hit” Theory Cytokines & Oxidative Stress

11. Increased de novo lipogenesis (DNL)
There are 3 major sources for the increased TG deposition in the liver:
Rate of FFA uptake and synthesis > need for FFA for essential functions
Impaired VLDL export
Increased de novo lipogenesis (DNL)
synthesis of FA from CHO in the liver

13. Role of Insulin Resistance
The liver is the main site of insulin action, in addition to skeletal muscle and adipose tissue.
Fatty liver: ability of insulin to inhibit hepatic glucose production is impaired (hyperglycemia).
Insulin resistance hyperglycemia, hyperinsulinemia…both stimulate DNL by activation of the transcription factors carbohydrate regulatory element-binding protein (ChREBP) & sterol regulatory element-binding proteins (SREBP-1c).
Insulin resistance stimulates the liver to overproduce TG rich VLDL in the fasting state.

14. The metabolic consequences of insulin resistance include: Persistent hyperglycemia Hyperinsulinemia Elevated serum FFA’s Hypertriglyceridemia

15. It is unclear if insulin resistance and hyperglycemia cause NAFLD or are the consequences of the disease. NAFLD can be considered the hepatic manifestation of metabolic syndrome.

16. “Two Hit” Theory
The first “hit” involves the accumulation of fat in the hepatocytes.
Subsequent “hits” involve:
chronic oxidative stress with the production of reactive oxygen species (ROS)
secretion of pro-inflammatory cytokines
mitochondrial dysfunction
liver injury, hepatic apoptosis (liver cell death) and fibrosis

17. Pro-inflammatory cytokines (TNF-α) are produced directly by hepatocytes in response to an increased supply of FFA and/or by adipose tissue macrophages that increase during obesity. Fibrosis is thought to arise as part of the normal healing response to inflammation and injury.

18. Cytokines and Oxidative Stress
Adiponectin:
Inversely associated with obesity, BMI, metabolic syndrome, visceral adiposity, NAFLD
IL-6:
Implicated in insulin resistance, NASH
TNF-α:
Elevated levels with insulin resistance, metabolic syndrome
CRP:
May be a marker for hepatic steatosis --- but not of severity of NAFLD
Pearce S et al. Noninvasive biomarkers for the diagnosis of steatohepatitis and advanced fibrosis in NAFLD. Biomarker Research.2013;1:7.

20. Role of Genetics Global patterns of racial and ethnic distributions.
United States:
Prevalence in Hispanic Americans, Asians and Native Americans higher than in Caucasians & African-Americans.
Schwimmer et al. Prevalence of fatty liver in children and adolescents. Pediatrics 2006;118(4):

21. insulin signaling and the regulation of fat metabolism
Several gene polymorphisms are associated with NAFLD & genes that influence:
insulin signaling and the regulation of fat metabolism
oxidative stress
responses to endotoxins
release of cytokines
severity of fibrosis
Genetic factors may also predispose certain individuals to environmental influences that promote the development of NAFLD.
Alisi et al. Nonalcoholic fatty liver disease and metabolic syndrome in adolescents: Pathogenetic role of genetic background and intrauterine environment. Ann Med. 2012;44(1)29-40.

22. Polymorphisms (single nucleotide polymorphisms or SNPs) in genes encoding proteins involved in the synthesis, storage, and export of TG may play a role in NAFLD susceptibility.

23. HFE gene (human hemochromatosis protein)
PNPLA3 (patatin-like phospholipase domain-containing protein 3, Adiponutrin)
Function not entirely known but is reported to have lipase-like activity and to promote TG hydrolysis in the liver.
SNP (methionine substituted for isoleucine at codon 148 in the gene PNPLA3 has been determined to regulate a variety of the mechanisms involved with the development of NAFLD
HFE gene (human hemochromatosis protein)
Gene involved with insulin sensitivity, oxidative stress.

24. Several investigators have found a genetic association between PNPLA3 polymorphisms and elevated plasma liver enzymes in Hispanic populations.
Romeo et al. Genetic variation in PNPLA3 confers susceptibility to nonalcoholic fatty liver disease. Nature Genetics (2):
Quan et al. PNPLA3 polymorphisms and liver aminotransferase levels in a Mexican American population. Clin Invest Med August 4; 35(4): E237.

25. Romeo et al. The 148M allele of the PNPLA3 gene is associated with indices of liver damage early in life. J Hepatol. 2010;53:
N = 475 overweight/obese children & adolescents (Italy)
age 10 ± 3; genotyped; anthropometric, biochemical and US data obtained
Carriers of two 148M alleles had a 52% increase in circulating ALT levels compared to carriers of two 148I alleles.
Liver steatosis was more prevalent in carriers of two 148M alleles.
Glucose tolerance and insulin sensitivity were similar across all three genotypes.
Carriers of the PNPLA3 148M allele: more likely to exhibit hepatic damage at an early age.

26. Genetics may play an important role particularly at early onset disease.
Use of genetic analysis and genotyping has the potential to become an important noninvasive tool for the screening and diagnosis of NAFLD.

27. Features Histological Classification of steatosis Biochemical
Insulin resistance, Hypertriglyceridemia, elevated ALT
Anthropometric Findings
Overweight, obesity, abdominal obesity
Clinical Features
Acanthosis nigricans
Diagnostic Imaging
MRI, CT, ultrasound

28. Histological
NAFL
presence of hepatic steatosis with no evidence of hepatocellular injury in the form of ballooning of the hepatocytes.
NASH
hepatic steatosis and inflammation with hepatocyte injury (ballooning) with or without fibrosis.

29. Spectrum of disease in NAFLD
A: Nonalcoholic fatty liver (NAFL)
B: Nonalcoholic steatohepatitis (NASH)
C: Cirrhosis

30. Histological Features
Brunt EM et al. Nonalcoholic steatohepatitis: a proposal for grading and staging the histological lesions. Am J Gastroenterol. 1999;94(9):
Kleiner DE et al. Nonalcoholic Steatohepatitis Clinical Research network. Design and validation of a histological scoring system for nonalcoholic fatty liver disease. Hepatology 2005;41(6):

31. Biochemical Features Elevated serum TG Elevated ALT > 150 mg/dL
No universal standards for children
30-45 U/L most commonly used cutoff for abnormal ALT
ALT may be normal
Current standards controversial—range may be too high

32. Insulin resistance
Homeostatic Model Assessment Insulin Resistance (HOMA-IR)
fasting glucose (mg/dl) × fasting insulin (μU/ml)
405
>3

33. Anthropometric Features
Overweight
BMI >95th percentile
Obesity
Abdominal obesity WC > 90th percentile

34. International Diabetes Federation (IDF) Criteria for the Classification of Metabolic Syndrome in Children
Age
years
10 – 16 years
> 16 years
Definition of adiposity
WC > 90th percentile
Blood glucose
No set value
FBG >100 mg/dL
FBG > 100 mg/dL
Dyslipidemia
TG > 150 mg/dL
HDL < 40 mg/dL
Blood Pressure
Systolic >130 mm Hg
Diastolic >85 mm Hg

35. Clinical Features
Acanthosis nigricans
Hepatomegaly on palpation

36. Diagnosis
Liver biopsy invasive, risks, expensive

37. Diagnostic Imaging Computerized Tomography (CT)
exposure to ionizing radiation
Ultrasound
accessible, no ionizing radiation exposure
low sensitivity: mild-moderate steatosis
limited beam penetration in obese individuals
Magnetic Resonance Imaging (MRI) no exposure to ionizing radiation expensive!

38. Ultrasound image of the Liver
MRI of the Liver
CT image of the Liver

39. Dietary Characteristics
overconsumption of fructose & soft drinks
lower consumption of fiber
overconsumption of meat/saturated fat/cholesterol
lower consumption of fish, omega-3 fatty acids, and lower consumption of some vitamins (vitamin E)
Zelber-Sagi et al. Nutrition and physical activity in NAFLD: An overview of the epidemiological evidence. World J Gastroenterol. 2011; 17(29): 3377–3389.

40. Nutritional Factors Role of CHO Sucrose Increases hepatic TG synthesis
Fructose
Increases DNL & insulin resistance in animal models
Fructose overfeeding increases fasting and postprandial plasma TG hepatic DNL, VLDL-TG secretion & decreased VLDL-TG clearance
Tappy L. Does fructose consumption contribute to non-alcoholic fatty liver disease? Clinics Res Gastroent Hepatol. 2012;36(6):

41. A possible explanation:
Insulin resistance and hyperglycemia develops primarily in presence of sustained fructose exposures associated with changes in body composition.
Tappy L. Does fructose consumption contribute to non-alcoholic fatty liver disease? Clinics Res Gastroent Hepatol. 2012;36(6):

42. Objective Materials & Methods
Bravo et al. Consumption of sucrose and high fructose corn syrup does not increase liver fat or ectopic fat deposition in muscles. Appl Physiol Nutr Metab. Doi: /apnm
Objective
To evaluate the effect of the addition of commonly consumed fructose/glucose containing sugars in the usual diet on liver fat and intramuscular adipose tissue (IMAT)
Materials & Methods
64 adults, consumed low fat milk sweetened with either HFCS or sucrose at different levels for 10 weeks
CT: liver fat
MRI: IMAT

43. Results
No increase in liver fat or IMAT
Authors Conclusions
When fructose consumed as part of a typical diet as sucrose or HFCS, liver fat storage is not promoted.
Limitations?

44. Sugar sweetened beverages
Increased soda consumption in US children and adults
~175 calories/day.
Duffey et al. Shifts in patterns and consumption of beverages between 1965 and Obesity. 2007;15:2739–2747.
Caramel coloring (contains advanced glycation end products) which can increase insulin resistance and inflammation.
Gaby AR. Adverse effects of dietary fructose. Altern Med Rev. 2005;10:294–306.

45. Zelber-Sagi S et al. Long term nutritional intake and the risk for non-alcoholic fatty liver disease (NAFLD): a population based study. J Hepatol. 2007;47:711–717.
cross-sectional study of a sub-sample (n = 375) from the Israeli National Health and Nutrition Survey ( ).
semi-quantitative FFQ, abdominal US, biochemical, anthropometrics
The NAFLD group consumed:
almost twice the amount of soft drinks (P = 0.03)
27% more meat (P < 0.001)
Subjects with NAFLD had a higher intake of soft drinks
Higher intake of soft drinks was associated with an increased risk of NAFLD, independent of age, gender, BMI, and total calorie intake.

46. Polyunsaturated n-3 and n-6 fatty acids
Animal models: reduction of steatosis
Studies in adults with NAFLD: improved lipid profiles, reduced inflammation
2 gm fish oil (6 mos, n=40 adults) reduced serum TG, liver enzymes, and TNF-α; regression of steatosis (US)
Spadaro L et al. Omega-3 polyunsaturated fatty acids: a pilot trial in non-alcoholic fatty liver disease. J Hepatol. 2006;44:S264

47. Treatment Lifestyle Interventions Physical activity Weight loss
Pharmacologic agents
Metformin
Vitamin E, omega 3 fats
Bariatric surgery

48. Physical activity
Exercise alone increases LBM, reduces adipose, improves insulin resistance.

49. Weight loss
As little as a 7-10% reduction in total body weight (regardless of diet composition) may reduce hepatic fat accumulation in obese adults and adolescents.
Promrat K et al. Randomized controlled trial testing the effects of weight loss on nonalcoholic steatohepatitis (NASH). Hepatology. 2010;51(1):

50. Reduction in intrahepatic fat has been reported to occur in obese adults with type 2 diabetes in as little as the two weeks of dietary restriction + exercise.
Tamura Y et al. Effects of diet ad exercise on muscle and liver intracellular lipid contents and insulin sensitivity in type 2 diabetic patients. J Endocrinol Metab. 2005;90(6):
Rapid weight loss >1.6 kg per week is not recommended due to the risk of liver damage.

51. Pharmacologic agents Metformin Metabolic effects Safety profile
Few studies in children with fatty liver disease
Not recommended at this time for treatment of NAFLD

52. Vitamin E Potent antioxidant
Children with NASH, NAFLD have lower Vitamin E intakes
TONIC Trial

53. TONIC Trial Objective Design, Setting, and Patients
To determine whether children with NAFLD would improve from therapeutic intervention with vitamin E or metformin.
Design, Setting, and Patients 
Randomized, double-blind, double-dummy, placebo-controlled clinical trial conducted at 10 university clinical research centers
N=173 patients (age 8-17 years)
biopsy-confirmed NAFLD
September March 2010.

54. Interventions 
Daily dosing of 800 IU of vitamin E (58 patients), 1000 mg of metformin (57 patients), or placebo (58 patients) for 96 weeks.

55. Main Outcome Measures 
Sustained reduction in ALT defined as 50% or less of the baseline level or 40 U/L or less at visits every 12 weeks from 48 to 96 weeks of treatment.
Improvements in histological features of NAFLD and resolution of NASH were secondary outcome measures.

56. Conclusion
Neither vitamin E nor metformin was superior to placebo in attaining the primary outcome of sustained reduction in ALT level in patients with pediatric NAFLD.
Lavine JE et al. 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):

57. Bariatric surgery Lap bands not FDA approved for <18 years
American Academy of Pediatrics (2004) Criteria:
failed >6 months of organized attempts at weight management.
attained or nearly attained physiologic or skeletal maturity. Girls age > 13, boys age > 15
BMI > 40 (with serious obesity related conditions) or have a BMI of greater than 50 with less severe obesity-related problems.
Other organizations, including the American Society for Metabolic and Bariatric Surgery (ASMBS) , have less stringent weight criteria for teens.
Proposed as a potential treatment for NASH in adolescents
Inge TH et al. Bariatric surgery for severely overweight adolescents: concerns and recommendations. Pediatrics.2004;114(1):

58. Implications for the RD
Summary of diet and lifestyle recommendations
No consensus as to what diet or lifestyle approach due to lack of scientific evidence
Omega 3 fatty acids, high MUFA, fruits, vegetables, low GI, high fiber, reduced intake of saturated fats, simple CHO and sweetened beverages are universally recommended.

59. Alisi A, Nobili V. Non-alcoholic fatty liver disease in children now: Lifestyle changes
and pharmacologic treatments. Nutrition. 2012;28(7-8):

60. Nutrition Assessment Anthropometric BMI Waist circumference
Skinfolds (?)

61. Biochemical ALT Lipid panel HDL, LDL, triglycerides FBG
Fasting Serum insulin (μIU/mL)
Tanner Stage I (prepubertal)
Tanner Stage II
Tanner Stage III
HOMA-IR
(>3)
fasting glu (mg/dl) × fasting insulin (μU/ml)
405

62. Clinical
Acanthosis nigricans
PMH
PCOS, type 2 diabetes, insulin resistance

63. Diet FFQ 20-60 questions SSB, fruits, vegetables
3, 5 day food records (if possible)
<12 years: caregiver

64. Assess intake of sugar sweetened beverages:
sodas, sweetened waters, teas, flavored milk, fruit juices, fruit drinks, energy drinks, sports drinks.
The type of fat (saturated vs. PUFA)
Fiber intake

65. Research at UNM
Characterization of pediatric outpatients diagnosed with NAFLD at UNMH (anthropometric, biochemical)
Retrospective x 5 years, N=38
PI: Cohen, Gonzales-Pacheco
Genetics, diet and pediatric NAFLD
ages 11-17, recruitment from Health Fit Children's Clinic
Dietary intervention (SSB reduction), measurement of steatosis (US), genetic analysis, biochemical features
Baseline, 4 months
PI: Cohen, Gonzales-Pacheco, Orlando, Garver, Negrete, Kong

66. Concluding Statements
Prevention of overweight and obesity are KEY