Post on 08-Sep-2020
Supplementary Digital Content (SDC)
S1: Screening evaluation in all patients
Etiological Evaluation
◦ Hepatotrophic viruses- IgM Hepatitis A, IgM Hepatitis E, HbsAg, IgM Anti
Hbc.
Other viruses- IgM Epstein Barr virus, IgM Cytomegalovirus, IgM
Parvo virus (where indicated).
◦ Wilson disease-Serum Ceruloplasmin, 24 hour urinary copper, DCT, Evidence
of Hemolysis, KF Ring (If possible).
◦ Autoimmune- DCT, ANA/ASMA/LKM-1/SLA, IgG.
◦ Hemophagocytosis- Serum triglyceride, cholesterol, ferritin and bone marrow
biopsy (If Fever/Splenomegaly/Bicytopenia)
Subject Evaluation
Detailed historical evaluation including age, gender, clinical
symptomatology, detailed past, development, birth and family history was
elicited from the subjects/parents.
Laboratory evaluation
All the subjects underwent detailed lab evaluation including complete liver
function test (serum bilirubin, aspartate aminotransferase, alanine
aminotransferase, alkaline phosphatase, gamma glutamyl transpeptidase,
albumin), international normalized ratio, serum creatinine, blood urea, serum
electrolytes, serum uric acid, metabolic Screen (random plasma glucose,
arterial ammonia, arterial blood gas analysis, urinary ketones, urine non
glucose reducing substances, serum creatine phosphokinase).
Echocardiography and Ultrasonography of abdomen was performed by trained
and experienced operators. Samples for metabolic testing included plasma
sample for Tandem Mass Spectrometry (TMS) for Carnitine/Acylcarnitine
Profile (and amino-acid analysis) and urine sample for Gas Chromatography
and Mass Spectrometry (GCMS) were collected within 4 hours of admission.
Age wise cut offs were used for analyzing metabolites’ results [1-4]
S2: Specific metabolic abnormalities detected in fatty acid oxidation disorders
Fatty acid oxidation disorder Plasma acylcarnitine Urine organic acidsCarnitine uptake defect Decreased C0 (free
carnitine), Decreased long-chainacylcarnitines
Normal or Elevated dicarboxylic acids
Carnitine palmitoyltransferase-I Elevated C0, Decreased long-chain acylcarnitines
Usually normal
Carnitine palmitoyltransferase-II Elevated C16, C18:1, C14, low C2
Normal
Carnitine acylcarnitine translocase Elevated C16, C18:1, C14, low C2
Normal or Elevated dicarboxylic acids
Very-long-chain acyl-CoAdehydrogenase
Elevated C14, C14:1/C12:1
Elevated Dicarboxylic acids
Long-chain 3-hydroxyacyl-CoAdehydrogenase/mitochondrialtrifunctional protein
Elevated C16 OH, C18:1 OH, C14 OH
Elevated Dicarboxylic acids, Elevated hydroxydicarboxylic acids (3-hydroxyadipic acid, 3-hydroxysebacic acid)
Medium-chain ketoacyl-CoA thiolase
Unknown Elevated Lactic, 3-hydroxybutyric, saturated and unsaturated C6–C16dicarboxylic acids
Multiple acyl-CoA dehydrogenase/glutaric aciduria II
Elevated C4, C5, C5 DC, C6, C8, C10, C12, C14
Elevated Glutaric acid, isobutyrylglycine, ethylmalonic acid,dicarboxylic acids, acylglycines (phenylpropionylglycine),2-hydroxyglutaric acid
Succinyl-CoA: oxoacid transferase Normal NormalMedium-chain acyl-CoAdehydrogenase
Elevated C8, C10, C10:1 Elevated Dicarboxylic acids, acylglycines (suberylglycine,hexanoylglycine), 5-hydroxyhexanoic acid, octanedioic acid, decanedioic acid
Medium/short-chain acyl-CoAdehydrogenase
Elevated 3-OH-C4 Elevated Dicarboxylic acids, 2-hydroxyglutaric acid
Short-chain acyl-CoA dehydrogenase
Elevated C4 (butyrylcarnitine and butyrylglycine)
Elevated Ethylmalonate, methylsuccinate
Ethylmalonic encephalopathy Elevated C4, C5 Elevated Lactic acid, ethylmalonate, methylsuccinate
2,4-Dienoyl reductase Hypocarnitinemia, 2-trans, 4-cisdecadienoylcarnitinein both urine and blood
Normal
3-Hydroxy-3-methylglutaryl-CoAsynthase 2
Normal Normal
3-Hydroxy-3-methylglutaryl-CoAlyase
Elevated C5OH Elevated 3-Methylglutaric acid, 3-methylglutaconic acid, 3-hydroxy-3-methylglutaric acid, 3-methylcrotonylglycine
β-Ketothiolase Normal or Decreased C0 Tiglylglycine, acetoacetic acid, 2-methylacetoacetic acid
S3: Consort diagram depicting patient selection
Abbreviation: ALF- Acute liver failure
Group 1Children (0-18 years age)
with ALF(n= 38)
Group 2Adults (> 18 Years age) with ALF
(n=26)
Step 1History , Baseline and Etiological Evaluation
All ALF Cases
(n= 55)Pediatric Group(n= 33)
Adult Group(n = 22)
Step 2 Metabolic
Testing
Excluded:On Valproate Therapy (n=2)Delayed sampling beyond 4 hours (n = 3)
Excluded:Delayed sampling beyond 4 hours (n = 4)
S4- Comparison of proportions for predicting outcome (Death or Liver transplantation/LT vs Survival) in the overall group (n = 55)
Parameter(Normal Value)
Death/LT(n= 27)
Survived (n= 28)
OR, 95 % CI , p-Value
High Methionine 24 17 5.18, 1.25 to 21.41, 0.016
High Phenyalanine 8 1 11.37, 1.31 to 98.59, 0.012
High Tyrosine 15 6 4.58, 1.41 to 14.91, 0.009
High Arginine 4 2 2.26, 0.38 to 13.51, 0.422
Hyperaminoacidemia
(elevation of > 1 aminoacid)
24 19 3.789, 0.89 to 15.97, 0.059
Low Free Carnitine (C0) 12 11 1.236, 0.42 to 3.61, 0.698
Abnormal Carnitine profile
(any abnormality)
14 17 0.69, 0.24 to 2.03, 0.508
Abbreviations- OR- Odd’s Ratio, CI- Confidence interval.
S5: Liver biopsy figure showing Diffuse Hepatocyte steatosis with minimal fibrosis and inflammation in Case 1
S6: Liver biopsy figure showing Steatohepatitis with Pericellular fibrosis in Case 2
a b c
a b
References
1. Schmidt-Sommerfeld E, Werner E, Penn D: Carnitine plasma concentrations in 353
metabolically healthy children. Eur J Pediatr 1988;147:356-360.
2. http://geneticslab.emory.edu/support//AR_reference_ranges.pdf. Accessed January 2,
2015.
3. http://geneticslab.emory.edu/support//AA_reference_ranges.pdf. Accessed January 2,
2015.
4. http://geneticslab.emory.edu/support//OA_reference_ranges.pdf. Accessed January 2,
2015.