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Diagnosis of primary BILE ACID SYNTESIS DISORDERS

Early identification and diagnosis of bile acid synthesis disorders (BASDs), is vital to prevent liver failure or progressive chronic liver disease  1
  • Deficiency of 3β-HSD is the most common BASD, presenting as both neonatal cholestasis and as chronic liver disease in older patients 1
  • Deficiency of Δ4-3-oxoR can present as either neonatal cholestasis or as liver failure resembling haemochromatosis; in cases of delayed diagnosis, mortality may reach up to 50%1
Suspicion of primary BASD 2,5

Essential clues for suspicion of primary BASD:

  • Cholestatic infant with normal serum GGT activity
  • Chronic cholestasis or liver disease of unknown aetiology
  • Neonatal/infant liver failure (for Δ4-3-oxoR deficiency liver disease is rapidly progressing to liver failure)
  • Malabsorption not related to pancreatic or digestive disease: steatorrhoea, fat-soluble vitamin deficiencies
  • Family history of cholestasis or liver failure, consanguinity

Confirmation of primary BASD 2

Mass spectrometry analysis of urinary bile acids

AND

Genetic sequencing of the HSD3B7 and AKR1D1 genes

In practice, examinations must be carried out at the same time to exclude other possible causes with a comparable clinical picture. 4

BASDs may be suspected based on a combination of clinical and laboratory signs, and histological signs of the liver. When all these signs are observed together, a specific urinary bile acids analysis test should be performed. Genetic testing provides final confirmation of the diagnosis. Furthermore, as these diseases are inherited, it is important to investigate the patient’s family history, particularly in cases of unexplained liver problems or deaths in young children. 2-4

  • Cholestasis and/or hepatocellular insufficiency during the first months of life or in childhood
  • Progressive and prolonged jaundice
  • Hepatomegaly, splenomegaly
  • Signs of liver failure
  • And/or malabsorption syndrome
  • Steatorrhoea
  • Clinical signs associated with fat-soluble vitamin deficiency: A, D, E, K (eye disorders, rickets, neurological disorders, haemorrhage)
  • Or cirrhosis
  • No pruritus
  • Elevation in serum transaminase levels (ALT, AST)
  • Conjugated hyperbilirubinaemia
  • Normal serum GGT activity
  • Normal or low serum total bile acids

Other indications: 4

  • Normal or low cholesterolaemia
  • Low serum concentrations of 25-hydroxyvitamin D, vitamin E and vitamin A
  • Prolonged prothrombin time / decreased prothrombin levels (vitamin K deficiency)
  • Unexplained liver problems
  • Deaths in young children
  • Consanguineous parents
  • Canalicular cholestasis
  • Without bile duct proliferation
  • Possibly with signs of giant cell hepatitis
  • Portal and lobular fibrosis with features of septal fibrosis or cirrhosis, depending on the stage
Specific diagnosis is based on mass spectrometry (MS) analysis of urinary bile acids showing typical bile acid profiles and on the identification of disease-causing mutations in the HSD3B7 or AKR1D1 genes.2

Bile acid analysis by mass spectrometry

Bile acid analysis in serum, urine, and bile are is usually performed with 4

  • Fast atom bombardment mass spectrometry (FAB-MS)
  • Gas chromatography mass spectrometry (GC-MS)
  • Electrospray ionisation tandem mass spectrometry (ESI-MS/MS)
  • Liquid chromatography tandem mass spectrometry (LC-MS/MS)
  • Or other similar techniques

In 3β-HSD deficiency, urinary analysis shows high excretion of bile acids, that is consistent with cholestasis, but qualitatively abnormal: 4

  • Absence or low concentrations (traces) of primary bile acids
  • Presence of high levels of atypical intermediate metabolites (3β-hydroxy-Δ5).

These assays are usually carried out by highly specialised laboratories. They provide a specific urinary excretion profile for each BASD, enabling diagnosis. 4

In Δ4-3-oxoR deficiency, urine analysis by mass spectrometry shows the absence or low concentrations (traces) of primary bile acids and the presence of atypical metabolites, with a predominance of conjugated hydroxy-oxo-bile acid and allo bile acid derivatives. However, the metabolic profile is not pathognomonic of a primary Δ4-3-oxoR deficiency as the enzyme is sensitive to any advanced liver damage, irrespective of the origin – it may result from secondary deficiency in its activity associated with a closely related urinary excretion profile. A primary genetic deficiency of Δ4-3-oxoR is very likely if the proportion of urinary Δ4-3-oxo bile acid reaches or exceeds 70%.4

Genetic analysis

Genetic sequence analysis of the HSD3B7 and AKR1D1 genes is required to confirm diagnosis.1,4 Currently, two methods are available: Sanger sequencing or targeted gene sequencing panel analysis.4 The gene encoding the enzyme HSD3B7 has been cloned, and the deleterious mutations associated with dysfunction of the enzyme have been identified. About fifteen homozygous or compound heterozygous mutations have been found to date, though no correlation has been identified between the types of mutation and the phenotypic expression of the disease.4

Following identification of the structure of the gene coding for the enzyme (AKR1D1), the primitive character of the deficiency is demonstrated by providing evidence of the mutations in affected patients.4

Suspicion of primary BASD 2,5

Essential clues for suspicion of primary BASD:

  • Cholestatic infant with normal serum GGT activity
  • Chronic cholestasis or liver disease of unknown aetiology
  • Neonatal/infant liver failure (for Δ4-3-oxoR deficiency liver disease is rapidly progressing to liver failure)
  • Malabsorption not related to pancreatic or digestive disease: steatorrhoea, fat-soluble vitamin deficiencies
  • Family history of cholestasis or liver failure, consanguinity

BASDs may be suspected based on a combination of clinical and laboratory signs, and histological signs of the liver. When all these signs are observed together, a specific urinary bile acids analysis test should be performed. Genetic testing provides final confirmation of the diagnosis. Furthermore, as these diseases are inherited, it is important to investigate the patient’s family history, particularly in cases of unexplained liver problems or deaths in young children. 2-4

Clinical signs 2-4

  • Cholestasis and/or hepatocellular insufficiency during the first months of life or in childhood*
  • Progressive and prolonged jaundice
  • Hepatomegaly, splenomegaly
  • Signs of liver failure
  • And/or malabsorption syndrome
  • Steatorrhoea
  • Clinical signs associated with fat-soluble vitamin deficiency: A, D, E, K (eye disorders, rickets, neurological disorders, haemorrhage)
  • Or cirrhosis
  • No pruritus

Laboratory signs 2-4

  • Elevation in serum transaminase levels (ALT, AST)
  • Conjugated hyperbilirubinaemia
  • Normal serum GGT activity**
  • Normal or low serum total bile acids

Other indications: 4

  • Normal or low cholesterolaemia
  • Low serum concentrations of 25-hydroxyvitamin D, vitamin E and vitamin A
  • Prolonged prothrombin time / decreased prothrombin levels (vitamin K deficiency)

family history 2-4

  • Unexplained liver problems
  • Deaths in young children
  • Consanguineous parents

histological signs 2-4

  • Canalicular cholestasis
  • Without bile duct proliferation
  • Possibly with signs of giant cell hepatitis
  • Portal and lobular fibrosis with features of septal fibrosis or cirrhosis, depending on the stage

Confirmation of primary BASD 2

Mass spectrometry analysis of urinary bile acids

AND

Genetic sequencing of the HSD3B7 and AKR1D1 genes

In practice, examinations must be carried out at the same time to exclude other possible causes with a comparable clinical picture. 4

Specific diagnosis is based on mass spectrometry (MS) analysis of urinary bile acids showing typical bile acid profiles and on the identification of disease-causing mutations in the HSD3B7 or AKR1D1 genes.2

Bile acid analysis by mass spectrometry

Bile acid analysis in serum, urine, and bile are is usually performed with 4

  • Fast atom bombardment mass spectrometry (FAB-MS)
  • Gas chromatography mass spectrometry (GC-MS)
  • Electrospray ionisation tandem mass spectrometry (ESI-MS/MS)
  • Liquid chromatography tandem mass spectrometry (LC-MS/MS)
  • Or other similar techniques

In 3β-HSD deficiency, urinary analysis shows high excretion of bile acids, that is consistent with cholestasis, but qualitatively abnormal: 4

  • Absence or low concentrations (traces) of primary bile acids
  • Presence of high levels of atypical intermediate metabolites (3β-hydroxy-Δ5).

These assays are usually carried out by highly specialised laboratories. They provide a specific urinary excretion profile for each BASD, enabling diagnosis. 4

In Δ4-3-oxoR deficiency, urine analysis by mass spectrometry shows the absence or low concentrations (traces) of primary bile acids and the presence of atypical metabolites, with a predominance of conjugated hydroxy-oxo-bile acid and allo bile acid derivatives. However, the metabolic profile is not pathognomonic of a primary Δ4-3-oxoR deficiency as the enzyme is sensitive to any advanced liver damage, irrespective of the origin – it may result from secondary deficiency in its activity associated with a closely related urinary excretion profile. A primary genetic deficiency of Δ4-3-oxoR is very likely if the proportion of urinary Δ4-3-oxo bile acid reaches or exceeds 70%.4

Genetic analysis

Genetic sequence analysis of the HSD3B7 and AKR1D1 genes is required to confirm diagnosis.1,4 Currently, two methods are available: Sanger sequencing or targeted gene sequencing panel analysis.4 The gene encoding the enzyme HSD3B7 has been cloned, and the deleterious mutations associated with dysfunction of the enzyme have been identified. About fifteen homozygous or compound heterozygous mutations have been found to date, though no correlation has been identified between the types of mutation and the phenotypic expression of the disease.4

Following identification of the structure of the gene coding for the enzyme (AKR1D1), the primitive character of the deficiency is demonstrated by providing evidence of the mutations in affected patients.4

1. Sundaram SS, Bove KE, Lovell MA, Sokol RJ. Mechanisms of disease: Inborn errors of bile acid synthesis. Nat Clin Pract Gastroenterol Hepatol 2008;5:456-68.

2. Jahnel J, Zöhrer E, Fischler B, et al. Attempt to determine the prevalence of two inborn errors of primary bile acid synthesis: results of a European survey. J Pediatr Gastroenterol Nutr 2017;64:864-8.

3. Bove KE, Heubi JE, Balistreri WF, Setchell KD. Bile acid synthetic defects and liver disease: a comprehensive review. Pediatr Dev Pathol 2004;7:315-34.

4. Protocole national de diagnostic et de soins : Déficits de synthèse des acides biliaires primaires. Centre de Référence Coordonnateur de l’Atrésie des Voies Biliaires et des Cholestases Génétiques; 2019.

5. Monte MJ, Marin JJG, Antelo A, Vasquez-Tato J. Bile acids: chemistry, physiology, and pathophysioly. World J Gastroenterol 2009;15:804-16.

​TH-BAS12EN/01/02/2024

Content reviewed in conjunction with experts in paediatric gastroenterology and hepatology

03/2024

General condition

The gene encoding CYP7A1 is highly regulated by a negative feedback involving the farnesoid X receptor (FXR)-dependent induction of fibroblast growth factor 15/19 (FGF15/19) by bile acids in enterocytes.

FGF15/19 binds to the fibroblast growth factor receptor 4 (FGFR4)/b-klotho complex in hepatocytes, activating signaling pathways that transcriptionally repress CYP7A1 expression.

Bile acids are transported to the liver in the portal blood and enter in the hepatocytes via sodium taurocholate cotransporting polypeptide (NTCP).

Bile acids are recovered from the intestines by the apical sodium-dependent bile acid transporter (ASBT).

 Cholic acid and chenodeoxycholic acid are conjugated to glycine or taurine, which are substrates for the bile acid transport pump (BSEP). Canalicular transport of bile acid is the rate-limiting step of bile secretion.

Biosynthesis of the primary bile acids, cholic acid (CA) and chenodeoxycholic acid (CDCA), involves numerous enzymes including 3β-HSD and Δ4-3-oxoR. The neutral pathway, the major pathway in adults, is initiated by CYP7A1, which is the rate-limiting step in the conversion of cholesterol to CA and CDCA. During the first months of life, however, the acidic pathway is essential for the synthesis of primary bile acids. The acidic pathway is initiated by hydroxylation of cholesterol by the sterol 27-hydroxylase (CYP27). Its regulation is not fully understood. .

Suppression of the negative feedback loop further represses biosynthesis, increasing the production of bile acid intermediates, which are excreted in the urine.

Bile acid intermediates that are hepatotoxic and cholestatic accumulate.

Absence of primary bile acids in the intestine results in fat and fat-soluble vitamin malabsorption.

 

Biliary secretion is strongly decreased by a reduction in the bile acid-dependent fraction of bile secretion. This results in cholestasis with normal serum γ-glutamyl-transferase (GGT) activity.

Defects in the enzymes 3β-HSD and ∆4-3-oxoR, catalysing key reactions in the formation of the primary bile acids (cholic acid and chenodeoxycholic acid), lead to inadequate synthesis of primary bile acids. Consequently, primary bile acids are not synthesized but instead atypical and hepatotoxic bile acid intermediates are formed.

Tight junction protein 2 (TJP2) deficiency or progressive familial intrahepatic cholestasis type 4 (PFIC4)

TJP2 deficiency or PFIC4 is an autosomal recessive disorder caused by a mutation in the TJP2 gene, encoding the tight junction protein-2, which is involved in the organisation of epithelial and endothelial cell junctions.59 PFIC4 is characterised by early-onset cholestasis with severe progression.4

More information about tight junction protein 2 (TJP2) deficiency or progressive familial intrahepatic cholestasis type 4 (PFIC4) and references
Benign recurrent intrahepatic cholestasis type 2 (BRIC2)

BRIC2 is a hereditary liver disorder characterised by intermittent attacks of intrahepatic cholestasis, generally without progression to chronic liver damage.31,32

More information about benign recurrent intrahepatic cholestasis type 2 (BRIC2) and references

Benign recurrent intrahepatic cholestasis type 1 (BRIC1)

BRIC1 is a hereditary liver disorder characterised by intermittent attacks of intrahepatic cholestasis, generally without progression to chronic liver damage. 31,32

More information about benign recurrent intrahepatic cholestasis type 1 (BRIC1) and references

Progressive familial intrahepatic cholestasis type 1 (PFIC1)

PFIC1 is an autosomal recessive childhood bile formation disorder of hepatocellular origin associated with extrahepatic features.31

More information about progressive familial intrahepatic cholestasis type 1 (PFIC1) and references

Progressive familial intrahepatic cholestasis type 2 (PFIC2)

PFIC2 is an autosomal recessive childhood disorder of bile formation in hepatocytes that is not associated with extrahepatic features.32 It is caused by impaired bile salt secretion due to defects in ABCB11 encoding the bile salt export pump protein (BSEP).58

More information about progressive familial intrahepatic cholestasis type 2 (PFIC2) and references

Choledochal cysts

Choledochal cysts are rare congenital bile duct abnormalities. 33,34

More information about choledochal cysts and references

Biliary lithiasis

Biliary lithiasis or gallstone disease is characterised by the presence of stones in the gallbladder, the biliary ducts, or both.24,25

More information about the biliary lithiasis and references
PRIMARY BILE ACID SYNTHESIS DEFECTS (3β-HYDROXY-Δ5-C27-STEROID DEHYDROGENASE (3β-HSD) DEFICIENCY, OR Δ4-3-OXOSTEROID 5β-REDUCTASE (Δ4-3-OXO-R) DEFICIENCY)

These inborn errors of primary bile acid synthesis are rare autosomal recessive cholestatic disorders, leading to inadequate synthesis (or complete absence) of the primary bile acids and resulting in the build-up of atypical and hepatotoxic bile acid intermediates.52

More information about primary bile acid synthesis defects 3β-HSD and Δ4-3-oxoR deficiencies and references

Cystic fibrosis

Cystic fibrosis is an autosomal recessive disorder caused by mutations in the CF transmembrane conductance regulator (CFTR) gene. CFTR is expressed in the apical surface of cholangiocytes.42

More information about cystic fibrosis and references

Alpha-1-antitrypsin deficiency (AATD)

AATD is a rare autosomal recessive condition caused by a mutation in the SERPINA1 gene leading to decreased production of the neutrophil-elastase protective protein alpha-1 antitrypsin, thereby increasing the risk of serious lung and liver disease.4,5

More information about alpha-1-antitrypsin deficiency (AATD) and references

Drug toxicity

Drug-induced liver injury (DILI) is a major cause of paediatric liver disease including cholestasis, accounting for almost 20% of cases of acute liver failure in children, and is a major reason for liver transplantation.45

More information about drug toxicity and references 

Cortisol deficiency

Cortisol deficiency may result from a primary adrenal process, owing to maldevelopment, malfunction, or destruction of the gland or be secondary to dysfunction of the hypothalamic-pituitary-adrenal axis.39

More information about cortisol deficiency and references 

Other extrahepatic causes

Such as spontaneous perforation of the common bile duct or congenital stenosis, etc1

  1. Adapted from, Protocole national de diagnostic et de soins : Déficits de synthèse des acides biliaires primaires. In: Génétiques CdRCdlAdVBedC, ed., 2019.

Choledochal cysts

Choledochal cysts  are rare congenital bile duct abnormalities. 21,22

More information about choledochal cysts and references

Cholestasis and / or cholestatic jaundice

In newborns:

First, confirm the aetiology:1

  Those requiring urgent treatment:

       Biliary atresia

        Urinary infection or sepsis

        Cortisol deficiency

  Those with an easily detectable aetiology:

     Alagille syndrome

       Choledochal cyst

       Cystic fibrosis

       Alpha-1-antitrypsine deficiency 

 

Neonatal sclerosing cholangitis

Neonatal sclerosing cholangitis is a rare biliary tract disease characterised by severe neonatal-onset cholangiopathy with patent bile ducts.4

More information about neonatal sclerosing cholangitis and references 

Biliary Lithiasis

Biliary lithiasis or gallstone disease is characterised by the presence of concretions in the gallbladder, the biliary ducts, or both.15,16

More information about biliary lithiasis and references

Alpha-1-antitrypsin deficiency (AATD)

AATD is a rare autosomal recessive condition caused by a mutation in the SERPINA1 gene leading to decreased production of the neutrophil-elastase protective protein alpha-1 antitrypsin, thereby increasing the risk of serious lung and liver disease.1,5

More information about alpha-1-antitrypsin deficiency and references

Cystic fibrosis

Cystic fibrosis is an autosomal recessive disorder caused by mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene. CFTR is expressed in the apical surface of cholangiocytes.30

More information about cystic fibrosis and references 

Cortisol deficiency

Cortisol deficiency may result from a primary adrenal process, owing to maldevelopment, malfunction, or destruction of the gland, or be secondary to dysfunction of the hypothalamic-pituitary-adrenal axis.27

More information about cortisol deficiency and references 

Biliary atresia

Biliary atresia is a progressive obliterative cholangiopathy of the intrahepatic and extrahepatic bile ducts. It occurs in the perinatal period causing severe, persistent jaundice and acholic stool and, if untreated, is associated with a poor prognosis.4

More information about biliary atresia and references 

Arthrogryposis-renal dysfunction-cholestasis syndrome (ARC syndrome)

ARC syndrome is an autosomal recessive disease characterised by immobility of the limbs and fixation of joints with muscle wasting (neurogenic arthrogryposis multiplex congenita), renal tubular dysfunction and neonatal cholestasis.8

More information about arthrogryposis-renal dysfunction-cholestasis syndrome (ARC syndrome) and references 

Hepatitis A

Hepatitis A virus (HAV) is an RNA picornavirus which is transmitted through faecal-oral contamination.47

More information about hepatitis A and references

Progressive familial intrahepatic cholestasis type 3 (PFIC3)

PFIC3 is an autosomal recessive childhood disorder of bile formation in hepatocytes caused by mutations in the ABCB4 gene coding for the phospholipid transporter MDR3, which is expressed in the canalicular membrane of hepatocytes.32,58

More information about progressive familial intrahepatic cholestasis type 3 (PFIC3) and references

Autoimmune cholangitis

Autoimmune cholangitis (AIC), also known as autoimmune cholangiopathy, is a chronic inflammation of the liver and a variant syndrome of autoimmune hepatitis.9

More information about autoimmune cholangitis and references

Biliary atresia

Biliary atresia is a progressive obliterative cholangiopathy of the intrahepatic and extrahepatic bile ducts. It occurs in the perinatal period causing severe, persistent jaundice and acholic stool and, if untreated, is associated with a poor prognosis.3

More information about biliary atresia and references 

sclerosing cholangitis

Primary sclerosing cholangitis (PSC) is a rare, idiopathic, heterogeneous, chronic, cholestatic liver disease characterised by inflammation and fibrosis of both the intrahepatic and extrahepatic bile ducts, leading to the formation of multifocal bile duct strictures.54-56

More information about primary sclerosing cholangitis and references 

Alagille syndrome (AGS)

Alagille syndrome is a multisystemic, autosomal dominant disorder caused by a defect in the Notch signalling pathway leading to intrahepatic bile duct paucity and resulting in significant cholestasis.2

More information about Alagille syndrome and references

Progressive familial intrahepatic cholestasis type 5 (PFIC5)

PFIC5 is a severe autosomal recessive liver disorder caused by a mutation in the NR1H4 gene.50

More information about progressive familial intrahepatic cholestasis type 5 (PFIC5) and references 

MYOSIN 5B DEFICIENCY

Myosin 5B deficiency causes microvillus inclusion disease. It is a severe autosomal recessive disease caused by a myosin 5B (MYO5B) mutation and characterised by malabsorption and diarrhoea.48

More information about myosin 5B deficiency and references

UNC45A DEFICIENCY (OSTEO-OTO-HEPATO-ENTERIC SYNDROME (O2HE SYNDROME))

UNC45A deficiency causes osteo-oto-hepato-enteric syndrome. It is an autosomal recessive syndrome secondary to loss of function mutations in the UNC45A gene.60

More information about UNC45A deficiency (osteo-oto-hepato-enteric syndrome (O2HE syndrome)) and references

Alagille syndrome

Alagille syndrome is a multisystemic, autosomal dominant disorder caused by a defect in the notch signalling pathway leading to intrahepatic bile duct paucity and resulting in significant cholestasis.1

More information about Alagille syndrome and references

Bilirubin metabolism5-7

Unconjugated
bilirubin

Bilirubin is mainly
produced from the
break down of red
blood cells.

Red cell breakdown
produces unconjugated
(“indirect”) bilirubin,
which circulates mostly
bound to albumin,
although some is
“free” and hence
able to enter the brain.

The terms “direct”
and “indirect” refer
to the way laboratories
measure the different
forms.

Unconjugated bilirubin
is metabolised in the
liver to produce
conjugated (“direct”)
bilirubin by uridine
diphosphate
glucuronosyltransferase.

The activity of this
enzyme only rises after
birth to reach adult like
values around the age
of three months.

Conjugation of bilirubin
increases its solubility
and facilitates its
secretion into bile.

Conjugated (“direct”)
bilirubin then passes
into the gut and is
largely excreted.

In the gastrointestinal
tract, bilirubin is modified
by digestive bacteria and
transformed into
urobilinogens . A large
portion remains in the
intestine and is
converted into stercobilin
(responsible for the
brown colour of faeces).
Some is reabsorbed into
the bloodstream and the
remainder is excreted in
the urine (urobilin is
responsible for the
yellow colour of urine).

Reticuloendothelial
system

Red blood cells

albumin

Haeme

Blood

“Indirect bilirubin”

“Direct bilirubin”

Liver

Intestine

Conjugated
bilirubin

Urobilinogens

Uridine
diphosphate
glucuronosyl-
transferase

Excreted in bile

Gut bacteria

Enterohepatic
circulation

Excreted in urine
(urobilin)

Excreted in faeces
(stercobilin)

Unconjugated
bilirubin-albumin
complex

​TH-BAS07EN/01/02/2024

Normal values:

Total bilirubin: 0-1.1 mg/dL

Vitamin A: 20-43 µg/dL

Vitamin E: 2.9 – 16.6 mg/L

Vitamin K1: 80-160 pg/mL

Normal values:

Serum calcium: 8.7 – 9.8 mg/dL

Serum phosphorous: 3.9 – 6.5 mg/dL

PTH: 15 – 65 pg/mL

Normal values:

Vitamin A: 30–120 μg/dL

Vitamin D: 20–100 ng/mL

Vitamin E: 5–20 μg/mL

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Enzymatic pathways of bile acid synthesis

classic pathway

adapted from Sundaram 20081 and Monte 20092

Cholesterol

Cholesterol 7α-hydroxylase
CYP7A1

3β-hydroxy-Δ5 -C27-steroid dehydrogenase
HSD3B7

Δ4-3-oxosteroid-5β-reductase
AKR1D1

Δ4-3-oxosteroid-5β-reductase
AKR1D1

3α-hydroxysteroid dehydrogenase
AKR1C4

Sterol 27-hydroxylase
CYP27A1

Bile acid CoA synthetase (BACS)
or very long chain acyl CoA
synthetase (VLCS)

Side-chain modification by
4 peroxisomal enzymes
(AMACR, BCOX, BDP, SCPx)

3α-hydroxysteroid dehydrogenase
AKR1C4

Sterol 27-hydroxylase
CYP27A1

Bile acid CoA synthetase (BACS)
or very long chain acyl CoA
synthetase (VLCS)

Side-chain modification by
4 peroxisomal enzymes
(AMACR, BCOX, BDP, SCPx)

Amino acid N-acyltransferase
(BAAT)

Amino acid N-acyltransferase
(BAAT)

7α-hydroxycholesterol

7α-hydroxy
4 cholesten-3-one

7α-12α-dihydroxy
-4 cholesten-3-one

7α-dihydroxy-5β-
cholestan-3-one

5β-cholestan-
3α,7α -diol

3α,7α-dihydroxy-5β-
cholestanoic acid (DHCA)

DHCA-CoA

Chenodeoxycholic
acid

Glyco or tauro-
chenodeoxycholic acid

Glyco or tauro-
cholic acid

Cholic acid

3α,7α,12α-trihydroxy-5β-
cholestanoic acid (THCA)

5β-cholestan-
3α,7α,12α-triol

7α-12α-dihydroxy-
5β-cholestan-3-one

THCA-CoA

Microsomes

Cytosol

Mitochondria

peroxisomes

hepatocyte

Endoplasmic
reticulum

Sterol 12α-hydroxylase
CYP8B1

AMARC: alpha methylacyl-CoA racemase

BCOX: Branched-chain acyl CoA oxydase

BDP: D-bifunctional protein hydratase 

SCPx: Sterol carrier protein

Alternative pathway

Cholesterol

Oxysterol 7α-hydroxylase
CYP7B1

3β-hydroxy-Δ5-C27-steroid dehydrogenase
HSD3B7

Side-chain modifications

Sterol 27-hydroxylase
CYP27A1

Amino acid N-acyltransferase
(BAAT)

3β-hydroxy-5-cholestanoic acid

3-oxo-7α-hydroxy-4-cholestanoic acid

3β,7α-dihydroxy-5-cholestanoic acid

Chenodeoxycholic acid

Glyco or tauro-
chenodeoxycholic acid

adapted from Sundaram 20081

1. Sundaram SS, Bove KE, Lovell MA, Sokol RJ. Mechanisms of disease: Inborn errors of bile acid synthesis. Nat Clin Pract Gastroenterol Hepatol 2008;5:456-68.

2. Monte MJ, Marin JJG, Antelo A, Vazquez-Tato J. Bile acids: chemistry, physiology, and pathophysiology. World J Gastroenterol 2009;15:804-16

​TH-BAS10EN/01/02/2024

 

REMINDER ABOUT BILE ACIDS

The bile acid family is a group of acid steroids synthesised from cholesterol in the liver. Although their best-known role is to aid with the emulsion, digestion and absorption of fats and liposoluble vitamins, other important physiological roles have been identified.1

On secretion of bile acids into bile canaliculi, osmotic pressure is created that accounts for the bile-acid-dependent fraction of bile flow. Bile acids stimulate biliary lipid secretion and form mixed micelles with biliary phospholipids, allowing the solubilisation of cholesterol and other lipophilic compounds in the bile. The mixed micelles also emulsify dietary fats in the intestines, facilitating their absorption.1

Bile acid synthesis

The primary bile acids, cholic acid and chenodeoxycholic acid are synthesised from cholesterol by an enzymatic cascade involving nearly 20 enzymes and two complementary chemical pathways, the classic “neutral” pathway and the alternative “acidic” pathway.2,3 Although the neutral pathway is believed to be the major pathway for bile acid synthesis in adults, in the first months of life the acidic pathway is thought to be more important.2,5 In humans and under normal conditions, the acidic pathway contributes little (approximately 10%) to the restitution of daily loss of bile acid.1 It may become the major bile acid biosynthetic pathway in patients with liver diseases.1

The primary bile acids produced include cholic acid (CA), which accounts for approximately 70% of the circulating pool of bile acids, and chenodeoxycholic acid (CDCA), which accounts for approximately 30% of the pool.4

Enzymatic pathways of bile acid synthesis

classic pathway

adapted from Sundaram 20083 and Monte 20091

Cholesterol

Cholesterol 7α-hydroxylase
CYP7A1

3β-hydroxy-Δ5 -C27-steroid dehydrogenase
HSD3B7

Δ4-3-oxosteroid-5β-reductase
AKR1D1

Δ4-3-oxosteroid-5β-reductase
AKR1D1

3α-hydroxysteroid dehydrogenase
AKR1C4

Sterol 27-hydroxylase
CYP27A1

Bile acid CoA synthetase (BACS)
or very long chain acyl CoA
synthetase (VLCS)

Side-chain modification by
4 peroxisomal enzymes
(AMACR, BCOX, BDP, SCPx)

3α-hydroxysteroid dehydrogenase
AKR1C4

Sterol 27-hydroxylase
CYP27A1

Bile acid CoA synthetase (BACS)
or very long chain acyl CoA
synthetase (VLCS)

Side-chain modification by
4 peroxisomal enzymes
(AMACR, BCOX, BDP, SCPx)

Amino acid N-acyltransferase
(BAAT)

Amino acid N-acyltransferase
(BAAT)

7α-hydroxycholesterol

7α-hydroxy
4 cholesten-3-one

7α-12α-dihydroxy
-4 cholesten-3-one

7α-dihydroxy-5β-
cholestan-3-one

5β-cholestan-
3α,7α -diol

3α,7α-dihydroxy-5β-
cholestanoic acid (DHCA)

DHCA-CoA

Chenodeoxycholic
acid

Glyco or tauro-
chenodeoxycholic acid

Glyco or tauro-
cholic acid

Cholic acid

3α,7α,12α-trihydroxy-5β-
cholestanoic acid (THCA)

5β-cholestan-
3α,7α,12α-triol

7α-12α-dihydroxy-
5β-cholestan-3-one

THCA-CoA

Microsomes

Cytosol

Mitochondria

peroxisomes

hepatocyte

Endoplasmic
reticulum

Sterol 12α-hydroxylase
CYP8B1

AMACR: alpha methylacyl-CoA racemase

BCOX: Branched-chain acyl CoA oxydase

BDP: D-bifunctional protein hydratase 

SCPx: Sterol carrier protein

The classic pathway, also known as the “neutral” pathway because its intermediate metabolites are neutral sterols, is the main pathway for bile acid synthesis.1-3 It is present only in the liver and synthesises cholic acid and chenodeoxycholic acid.1 This pathway consists of a cascade of reactions catalysed by enzymes located in microsomes, the cytosol, mitochondria and peroxisomes.1 The final step in the synthesis of bile acid is the conjugation of cholic acid and chenodeoxycholic acid to taurine or glycine.3

Alternative pathway

Cholesterol

Oxysterol 7α-hydroxylase
CYP7B1

3β-hydroxy-Δ5-C27-steroid dehydrogenase
HSD3B7

Side-chain modifications

Sterol 27-hydroxylase
CYP27A1

Amino acid N-acyltransferase
(BAAT)

3β-hydroxy-5-cholestanoic acid

3-oxo-7α-hydroxy-4-cholestanoic acid

3β,7α-dihydroxy-5-cholestanoic acid

Chenodeoxycholic acid

Glyco or tauro-
chenodeoxycholic acid

adapted from Sundaram 20083  and Monte 20091

The alternative pathway involves C27-hydroxylation of cholesterol by sterol 27-hydroxylase as the initial step: side-chain oxidation of cholesterol precedes steroid ring modification.1,3 Thus, acidic intermediate metabolites are formed: this is why this pathway is also known as the “acidic” pathway.1 It primarily produces chenodeoxycholic acid.3

Regulation of bile acid synthesis

Bile acid synthesis is tightly regulated to ensure homeostatic levels of cholesterol are produced and to provide adequate emulsification in the intestine. An excess of bile acids has a negative feedback effect, repressing further synthesis; conversely, when bile acids levels are low, synthesis is increased.2,6

In the ‘neutral’ pathway, the rate-limiting step is the modification of the steroid nucleus, which takes place in hepatic microsomes and is catalysed by cholesterol 7α-hydroxylase (CYP7A1). The neutral pathway facilitates the transformation of cholesterol to cholic acid and chenodeoxycholic acid, which are further conjugated to glycine or taurine and used as substrates for the bile acid transport pump. The canalicular transport of bile acid is the rate-limiting step of bile secretion. Bile acids are recovered from the intestines by the apical sodium-dependent bile acid transporter and return to the liver via the portal blood. The gene encoding CYP7A1 is highly regulated by negative feedback involving farnesoid X receptor (FXR)-dependent induction of fibroblast growth factor 15/19 (FGF15/19) expression by bile acids in the enterocytes. FGF15/19 binds to the fibroblast growth factor receptor 4-β-klotho complex in hepatocytes, activating signalling pathways that transcriptionally repress CYP7A1 expression.2

Enterohepatic circulation of bile acids

Bile acids are mostly restricted to enterohepatic circulation, circulating between the liver, the biliary tree, the intestine, and the portal blood which returns them to the liver. Almost all (95%) the bile acids are recovered from the intestine, mostly in the ileum.1 The liver converts around 500 mg of cholesterol into bile acids per day. This accounts for 90% of the cholesterol that is actively metabolized by the body. The remaining 10% of cholesterol that is synthesised is biosynthesised from steroid hormones.6

Newly synthesised bile acids are secreted into the bile where they are transported to the lumen of the small intestine, where they act as lipid emulsifiers, solubilising nutrients. These nutrients are incorporated into lipoproteins, and are delivered via the portal vein to the liver and metabolised. About 95% of the bile acids are recycled and secreted back into the bile. The remaining 5% are excreted into the faeces.6-7

Enterohepatic circulation of bile acids

adapted from van Mil 20057

ENTEROHEPATIC CYCLE

  • Fichier 101 Approximately 500 mg of cholesterol is converted into bile acids each day in the adult human liver. Newly synthesized bile acids are stored in the gallbladder.
  • Fichier 102 Upon digestion of a meal, bile salts are delivered to the lumen of the small intestine where they act as emulsifiers of dietary lipids, cholesterol, and fat-soluble vitamins.
  • Fichier 104 Bile salts are converted by intestinal bacteria into secondary bile acids.
  • Fichier 105 Bile salts are transported from the intestine to the liver via the portal circulation, and subsequently resecreted into bile. Approximately 95% of bile salts are recovered in the gut during each cycle of this enterohepatic circulation.
  • Fichier 103 5% of bile acids pool that are lost via the faeces are replaced by de novo synthesis in the liver.

1. Monte MJ, Marin JJG, Antelo A, Vazquez-Tato J. Bile acids: chemistry, physiology, and pathophysiology. World J Gastroenterol 2009;15:804-16.

2. Jahnel J, Zöhrer E, Fischler B, et al. Attempt to determine the prevalence of two inborn errors of primary bile acid synthesis: results of a European survey. J Pediatr Gastroenterol Nutr 2017;64:864-8.

3. Sundaram SS, Bove KE, Lovell MA, Sokol RJ. Mechanisms of disease: Inborn errors of bile acid synthesis. Nat Clin Pract Gastroenterol Hepatol 2008;5:456-68.

4. Ashby K, Navarro Almario EE, Tong W, Borlak J, Mehta R, Chen M. Review article: therapeutic bile acids and the risks for hepatotoxicity. Aliment Pharmacol Ther 2018;47:1623-38.

5. Clayton PT. Disorders of bile acid synthesis. J Inherit Metab Dis 2011;34:593-604.

6. Russell DW. The enzymes, regulation, and genetics of bile acid synthesis. Annu Rev Biochem 2003;72:137-74.

7. van Mil SW, Houwen RH, Klomp LW. Genetics of familial intrahepatic cholestasis syndromes. J Med Genet 2005;42:449-63.

​TH-BAS10EN/01/02/2024

Normal values:

Vitamin A: 1,09-3,07 μmol/L

Vitamin E: 25-42 μmol/L

AST: <39 U/L

ALT: <34 U/L

GGT: <38 U/L

Total bilirubin: <17 μmol/L

Conjugated bilirubin level: <5 μmol/L

Normal values:

AST: < 45 mU/mL

International normalized ratio: < 1.2

Prothrombin ratio: 70-120 %

Factor VII: 70-120 %

Factor X: 70-120 %

Vitamin E: 300-1200 μg/dL

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Causes of jaundice in newborn or infant5

Jaundice of the newborns or infants

Unconjugated hyperbilirubinaemia

Mechanical haemolysis

Other causes of haemolysis

Intramedullary haemolysis

Infection-related haemolysis

Immune haemolysis

Constitutional haemolysis

Haemolysis

Default in the conjugation of
indirect bilirubin (UGT1A gene)

Rare genetic deficiency of canalicular
or sinusoidal specific transporter of
conjugated bilirubin

Moderate or severe hereditary
deficiency of enzyme activity

Enzyme immaturity or
enzymatic inhibition

Hepatocellular insufficiency

Cholestasis (decreased bile flow)

Other (hypothyroidism, trisomy 21)

Conjugated hyperbilirubinaemia

​TH-BAS07EN/01/02/2024

Bilirubin metabolism5-7

Important things to check: the degree and duration of stool discolouration. The colour of the stool should ideally be recorded after elimination of any component likely to change the colour and with the help of a colour chart:2, 14

This chart is used with the kind permission of the AMFE, Association Maladies Foie Enfants, a French association dedicated to liver diseases in children.3

You can also refer to the stool chart provided by the Children’s Liver Disease Foundation (CLDF), a UK charity committed to fighting all childhood liver diseases :

CLDF-Yellow-Alert-Stool-Chart

Persistently pale coloured stools may indicated liver disease.14  Complete and prolonged discolouration of stools for 7 days is suggestive of biliary atresia until proven otherwise. However, the sign is not specific to biliary atresia and can also occur, among others, in conditions such as cystic fibrosis, Alagille syndrome, alpha-1 antitrypsin deficiency and neonatal sclerosing cholangitis.2

​TH-BAS08EN/01/02/2024

What causes (rare) cholestasis
in paediatric patients

?

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Content reviewed by experts in paediatric gastroenterology and hepatology.

This website was created by Theravia. Theravia is a leading international pharmaceutical laboratory specializing in rare or neglected diseases. Formed through the merger of Addmedica and CTRS, we are dedicated to address the unmet medical needs of patients with these challenging conditions

03/2024

What causes (rare) cholestasis
in paediatric patients ?

Menu

To access the platform and the
diagnostic algorithms,
please confirm:

YES, I'm a healthcare professional

If you are not a healthcare professional, please do not access the website as the content is not suitable.
We invite you to continue your research through another website

This website was created by Theravia. Theravia is a leading international pharmaceutical laboratory specializing in rare or neglected diseases. Formed through the merger of Addmedica and CTRS, we are dedicated to address the unmet medical needs of patients with these challenging conditions

Created by

Content reviewed by experts in paediatric gastroenterology and hepatology.

03/2024