Lysosomal Acid Lipase Deficiency Unmasked in Two Children With Nonalcoholic Fatty Liver DiseaseRyan W. Himes, MD, a Sarah E. Barlow, MD, MPH, a Kevin Bove, MD, b Norma M. Quintanilla, MD, c Rachel Sheridan, MD, b Rohit Kohli, MBBS, MSd

aSection of Gastroenterology and Hepatology, Department

of Pediatrics and cDepartment of Pathology and

Immunology, Texas Children’s Hospital, Baylor College

of Medicine, Houston, Texas; and bDepartment of

Pathology, Cincinnati Children’s Hospital and dSection

of Gastroenterology and Hepatology, Department of

Pediatrics, University of Cincinnati, Cincinnati, Ohio

Dr Himes conceptualized the report and drafted the

initial manuscript; Drs Barlow, Bove, Quintanilla,

and Sheridan reviewed and revised drafts; Dr Kohli

conceptualized the report and revised the initial

draft; and all authors approved the fi nal manuscript

as submitted.

DOI: 10.1542/peds.2016-0214

Accepted for publication Jun 7, 2016

Address correspondence to Ryan W. Himes, MD,

6701 Fannin St, Houston, TX 77030. E-mail: himes@

bcm.edu

PEDIATRICS (ISSN Numbers: Print, 0031-4005; Online,

1098-4275).

Copyright © 2016 by the American Academy of

Pediatrics

FINANCIAL DISCLOSURE: Dr Himes has served on

an advisory board, speakers’ bureau, and been

a site investigator for an epidemiology study

on lysosomal acid lipase defi ciency, for Alexion

Pharmaceuticals; Dr Barlow has served as site

investigator for an observational database on

lysosomal acid lipase defi ciency, for Alexion

Pharmaceuticals; Dr Kohli has served on an

advisory board and speakers’ bureau for Alexion

Pharmaceuticals; and Drs Bove, Quintanilla, and

Sheridan have indicated they have no fi nancial

relationships relevant to this article to disclose.

FUNDING: No external funding.

Lysosomal acid lipase deficiency

(LAL-D) is an autosomal recessive,

single gene disorder, caused by

mutations in LIPA (OMIM #278000)

resulting in lysosomal accumulation

of cholesteryl ester and triglyceride

in hepatocytes, endothelium, and

myeloid-derived cells. Depending

on ethnicity and race, prevalence

estimates range from 1:40 000 to

1:300 000. 1, 2 LAL-D may present in

early infancy with malabsorption and

catastrophic liver failure culminating

in death before 6 months of age, a

clinical phenotype formerly called

Wolman disease. On the other hand,

LAL-D may present insidiously

in children and adolescents with

variable and nonspecific findings like

hepatosplenomegaly, hepatic steatosis,

elevations in aminotransferases, or a

lipid profile characterized by elevated

low-density lipoprotein cholesterol

(LDL-c) and depressed high-density

lipoprotein cholesterol levels. In spite of

the later presentation in these patients,

there is evidence of clinically significant

liver disease and liver-related mortality

among children and adolescents with

LAL-D, underscoring the importance

of identifying affected patients. 3

The diagnosis of LAL-D, particularly in

children and adolescents, is hampered

by the lack of specific clinical findings,

the broad spectrum of disease

presentation, and significant overlap

with more common diseases. In

primary care practices, as well as

specialty clinics, features of LAL-

D, like hepatic steatosis, elevated

aminotransferases, and dyslipidemia,

are more often seen coexisting with

conditions like nonalcoholic fatty liver

disease (NAFLD), metabolic syndrome,

abstractLysosomal acid lipase deficiency (LAL-D) is a classic lysosomal storage

disorder characterized by accumulation of cholesteryl ester and

triglyceride. Although it is associated with progressive liver injury, fibrosis,

and end-stage liver disease in children and adolescents, LAL-D frequently

presents with nonspecific signs that overlap substantially with other, more

common, chronic conditions like nonalcoholic fatty liver disease (NAFLD),

metabolic syndrome, and certain inherited dyslipidemias. We present 2

children with NAFLD who achieved clinically significant weight reduction

through healthy eating and exercise, but who failed to have the anticipated

improvements in aminotransferases and γ-glutamyl transferase. Liver

biopsies performed for these “treatment failures” demonstrated significant

microvesicular steatosis, prompting consideration of coexisting metabolic

diseases. In both patients, lysosomal acid lipase activity was low and LIPA

gene testing confirmed LAL-D. We propose that LAL-D should be considered

in the differential diagnosis when liver indices in patients with NAFLD fail

to improve in the face of appropriate body weight reduction.

CASE REPORTPEDIATRICS Volume 138 , number 4 , October 2016 :e 20160214

To cite: Himes RW, Barlow SE, Bove K, et al.

Lysosomal Acid Lipase Defi ciency Unmasked

in Two Children With Nonalcoholic Fatty Liver

Disease. Pediatrics. 2016;138(4):e20160214

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HIMES et al

and certain inherited dyslipidemias.

NAFLD, linked tightly to overweight

and obesity, is estimated to

affect 33% of adults 4 and 9.6% of

children 5 in the general population,

making it the most common

chronic liver disease in the United

States. Unfortunately, there are no

laboratory-based diagnostic tests for

NAFLD, and although routine liver

imaging such as ultrasound may be

suggestive of hepatic steatosis, its

sensitivity is poor until steatosis

exceeds 30%. 6 Thus, only liver biopsy

can discriminate between simple

hepatic steatosis, a nonprogressive

or slowly progressive form of NAFLD,

and the more aggressive nonalcoholic

steatohepatitis (NASH), which is

defined by inflammation and/or

fibrosis. Given the invasiveness,

cost, and potential risks of applying

diagnostic liver biopsy to a condition

whose prevalence is nearly 10% of all

children, NAFLD is often diagnosed

on clinical grounds in overweight

or obese individuals after ruling out

other conditions through standard

investigations.

Herein, we report 2 children

with LAL-D, who were initially

presumed to have NAFLD alone. We

highlight the diagnostic challenge

of identifying a rare, but important

disease, which may coexist with, or

be misdiagnosed as, the much more

common, NAFLD.

PATIENT 1

An 8-year-old Hispanic girl

presented to her pediatrician after

nuchal acanthosis nigricans was

identified during a school-wide

screening program. Her weight was

38.1 kg (89th percentile), height

134.7 cm (53rd percentile), and

BMI 21.1 (94th percentile). Based

on her BMI and the presence of

acanthosis nigricans, screening

laboratory tests were performed that

revealed the following: aspartate

aminotransferase (AST), 207 U/L

(normal, 15–40 U/L); alanine

aminotransferase (ALT), 401 U/L

(normal, 7–35 U/L); total cholesterol,

235 mg/dL (normal, <170 mg/dL);

LDL-c, 167 mg/dL (normal, <110

mg/dL), prompting referral to a

pediatric gastroenterologist and

recommendations about weight

reduction. Her examination at

the gastroenterology clinic was

remarkable only for the presence

of nuchal acanthosis nigricans; she

did not have hepatosplenomegaly.

A diagnostic evaluation included

normal results for α-1 antitrypsin

PI-type, antiactin antibody,

antinuclear antibody, liver–

kidney microsomal antibody,

total immunoglobulin (Ig) G level,

ceruloplasmin, infectious hepatitis

(A/B/C), and tissue transglutaminase

IgA. An abdominal ultrasound

revealed normal hepatic echogenicity

and no organomegaly.

After 6 months, the patient was seen

in follow-up; her BMI was stable

(21.1, 93rd percentile); however,

her aminotransferases remained

elevated (AST, 131 U/L; ALT, 171

U/L), so a diagnostic liver biopsy

was performed. The biopsy revealed

mixed micro and macrovesicular

steatosis with minimal lobular

inflammation and stage 1 lobular

and portal fibrosis ( Fig 1 A and

B), interpreted as consistent with

NAFLD.

Over the next 3 years, the patient

participated in structured programs

for weight management and

ultimately reduced her BMI to the

85th percentile; follow-up laboratory

testing revealed the following:

AST, 61 U/L; ALT, 83 U/L; total

cholesterol, 200 mg/dL; and

LDL-c, 136 mg/dL. Given the

persistent aminotransferase

elevations in spite of significant

BMI improvement, a follow-up liver

biopsy was obtained. Light and

electron microscopy (EM) revealed

widespread microvesicular steatosis

and lipid-filled cytoplasmic vesicles

in hepatocytes and Kupffer cells ( Fig

1C). Minimal lobular inflammation

and stage 1 lobular and portal

fibrosis were unchanged. Based on

the predominantly microvesicular

pattern of steatosis, the differential

diagnosis was broadened to include

metabolic diseases, including LAL-D.

Her lysosomal acid lipase activity

e2

FIGURE 1Patient 1: A, Initial liver biopsy reveals predominantly microvesicular steatosis. Macrovesicular steatosis is distributed in ~10% of hepatocytes (hematoxylin and eosin, original magnifi cation, ×400). B, Masson Trichrome stain highlights mild fi brous expansion of portal tracts (×100). C, Follow-up liver biopsy EM demonstrates numerous membrane-bound lipid vesicles in hepatocytes and Kupffer cells (uranyl acetate and lead citrate).

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PEDIATRICS Volume 138 , number 4 , October 2016

was low, 0.011 nmol/punch/hour,

and confirmatory Sanger sequencing

of LIPA revealed a homozygous

mutation, c.894G>A, which alters a

splice site and leads to skipping exon

8, resulting in ~97% reduction in

enzyme activity. 7

PATIENT 2

A 16-year-old white girl presented

to the emergency department with

fever and symptoms suggestive of

a urinary tract infection. She had

severe obesity, her weight was

137 kg (>99th percentile), height

160.5 cm (35th percentile), with

a resultant BMI of 53.18 (>99th

percentile).

She underwent a renal ultrasound

that confirmed pyelonephritis

but also incidentally revealed

splenomegaly. Her blood counts

suggested hypersplenism with

low platelet (78 000/μL) and

leukocyte (3100/μL) counts. Based

on these initial observations, the

treating inpatient service requested

laboratory tests and consulted

gastroenterology. Relevant initial

test results included the following:

AST, 57 U/L (normal, 15–45 U/L);

ALT, 87 U/L (normal, 7–35 U/L);

γ-glutamyl transferase (GGT),

108 U/L (normal, 7–32 U/L);

LDL-c, 102 mg/dL (normal,

<200 mg/dL); high-density

lipoprotein cholesterol, 43 mg/dL

(normal, >45 mg/dL); and

triglycerides, 83 mg/dL (normal,

<126 mg/dL).

The gastroenterology team

continued the diagnostic evaluation

of elevated aminotransferases

and GGT in the setting of morbid

obesity. This included normal

results for α-1 antitrypsin PI-type,

antismooth muscle antibody,

antinuclear antibody, liver–kidney

microsomal antibody, total IgG

level, ceruloplasmin, infectious

hepatitis (A/B/C/cytomegalovirus/

Epstein-Barr virus), iron, ferritin,

and tissue transglutaminase IgA.

An abdominal ultrasound revealed

normal hepatic echogenicity and no

organomegaly. An abdominal MRI

with liver elastography demonstrated

splenomegaly, but also reported

a nodular liver with an increased

liver stiffness value of 7 kPa; a

liver stiffness value of more than

2.7 is consistent with the presence

of significant liver fibrosis. 8 The

child was discharged with advice to

improve lifestyle and diet such as to

attempt to lose weight.

Over the next year, the child reduced

her BMI from a peak of 56 to 46, but

her liver indices, though improved,

remained elevated despite significant

weight loss: AST, 33 U/L; ALT, 58

U/L; and GGT, 105 U/L. A repeat

MRI revealed an improved, though

still elevated, liver stiffness (3.83

kPa) value. Given these continued

abnormalities, a liver biopsy was

obtained that revealed panlobular

microvesicular steatosis, minimal

lobular inflammation, and stage 3

to 4 fibrosis ( Fig 2 A, B, and C). EM

revealed prevalent membrane-bound

lipid droplets, as well as diffuse

mitochondrial abnormalities typical

of metabolic stress in obese patients

with metabolic syndrome ( Fig 2D).

Based on the microvesicular pattern

of steatosis and the EM findings, the

differential diagnosis was broadened

to include metabolic diseases,

particularly LAL-D. Her lysosomal

acid lipase activity was 0.0 nmol/

punch/hour and sequencing of

LIPA revealed 2 variants: c.920C>A

(p.A307D) and c.1055_1057delACG

(p.D352del). Each has a population

frequency of <0.0001 in the Exome

Aggregation Consortium database,

with the former predicted by in silico

modeling to be pathogenic and the

latter disrupting an aspartate residue

that is conserved in all mammalian

species.

e3

FIGURE 2Patient 2: A and B, Liver biopsy reveals predominantly microvesicular steatosis distributed in ~30% of the hepatocytes. Patches of macrovesicular lipid were uncommon (hematoxylin and eosin, original magnifi cation, ×200 and ×400). C, Masson Trichrome stain demonstrates the severe nonuniform, dense periportal, and bridging fi brosis (×20). D, EM reveals numerous membrane-bound, empty-appearing, lipid vesicles (lysosomes) in hepatocytes, consistent with LAL-D. The mitochondria are universally contracted with marked dilatation of the inner crystal spaces. This diffuse mitochondriopathic change is not typical of LAL-D and is likely a secondary stress change related to concomitant NAFLD (uranyl acetate and lead citrate, ×12 000).

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HIMES et al

DISCUSSION

We report 2 children with LAL-D,

who were initially diagnosed and

treated for NAFLD. Their diagnoses

were established only after liver

biopsies, performed 1 or more

years into follow-up, demonstrated

findings atypical for NAFLD. Both

were consistent with predominantly

microvesicular steatosis, suggesting

the possibility of a competing or

coexisting diagnosis. Indeed, we

believe that these patients had

both NAFLD and LAL-D, the latter

coming to attention only after

additional studies were performed,

when clinically meaningful weight

reduction failed to achieve expected

improvements of laboratory

parameters. In case 2, the severity

of hepatic fibrosis at such an

early age would not be expected

in NASH alone. Moreover, diffuse

mitochondrial stress changes are

not a component of LAL-D, but are

frequently observed in NASH. These

phenotypically similar diseases are

not necessarily mutually exclusive.

In a practical sense, the diagnostic

evaluation of suspected NAFLD

should, at a minimum, address

conditions that are potentially

life-threatening or those that are

treatable; Wilson disease and

autoimmune hepatitis are examples

of conditions routinely tested for

in this context. LAL-D, which can

be ruled out with a blood-based

enzyme analysis, has not, up to this

point, been part of this diagnostic

evaluation in most centers. This is

in spite of clinical guidelines from

Europe 9 and the United States 10 that

endorse consideration of monogenic

causes of steatosis and utilization

of noninvasive testing initially in

children, before they undergo liver

biopsy.

It is now recognized that LAL-D

may be both underdiagnosed 2

and associated with significant

morbidity and mortality, even among

pediatric and adolescent patients.

In a literature review, Bernstein

et al 3 identified 135 patients with

LAL-D presenting after infancy; 4

of 8 liver-related deaths occurred

in patients under 21 years of age

and children aged 5 to 14 years old

accounted for all 9 liver transplants

reported. Burton et al 11 presented

a similar rate of pediatric liver

transplantation, 8.1%, among a group

of 49 patients with LAL-D. On this

background, there appears to be

value in identifying individuals with

LAL-D, whether for closer medical

supervision or for consideration of

treatment. Encouraging results from

clinical trials 12– 14 of recombinant

lysosomal acid lipase, now approved

for use in the United States, may

portend a viable therapy for this

group of patients. We conclude,

given the accessibility of clinical

laboratory-based testing for LAL-

D, availability of potential curative

treatments, and the data from

our report, there is a supportive

rationale for testing for LAL-D among

patients with suspected NAFLD who

do not respond to routine lifestyle

interventions.

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e4

ABBREVIATIONS

ALT:  alanine aminotransferase

AST:  aspartate aminotransferase

EM:  electron microscopy

GGT:  γ-glutamyl transferase

Ig:  immunoglobulin

LAL-D:  lysosomal acid lipase

deficiency

LDL-c:  low-density lipoprotein

cholesterol

NAFLD:  nonalcoholic fatty liver

disease

NASH:  nonalcoholic steatohep-

atitis

POTENTIAL CONFLICT OF INTEREST: Dr Himes has served on an advisory board, speakers’ bureau, and been a site investigator for an epidemiology study on

lysosomal acid lipase defi ciency, for Alexion Pharmaceuticals; Dr Barlow has served as site investigator for an observational database on lysosomal acid lipase

defi ciency, for Alexion Pharmaceuticals; Dr Kohli has served on an advisory board and speakers’ bureau for Alexion Pharmaceuticals; and Drs Bove, Quintanilla,

and Sheridan have indicated they have no potential confl icts of interest to disclose.

by guest on February 19, 2020www.aappublications.org/newsDownloaded from

PEDIATRICS Volume 138 , number 4 , October 2016

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SD, et al. Use of magnetic resonance

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J Pediatr. 2014;164(1):186–188

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Clinical effect and safety profi le of

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14. Valayannopoulos V, Malinova V, Honzík

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DOI: 10.1542/peds.2016-0214 originally published online September 13, 2016; 2016;138;Pediatrics 

Sheridan and Rohit KohliRyan W. Himes, Sarah E. Barlow, Kevin Bove, Norma M. Quintanilla, Rachel

Nonalcoholic Fatty Liver DiseaseLysosomal Acid Lipase Deficiency Unmasked in Two Children With

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