Developmental pattern of urinary bile acid profile in preterm infants

Background:  Bile acid metabolism in preterm infants is yet to be fully characterized. We compared the developmental pattern of urinary bile acid profiles in ten infants born at gestational ages from 25 to 33 weeks with previous data from full-term infants from birth to about 7 months of age.
Methods:  Gas chromatography–mass spectrometry was performed on serial samples.
Results:  Total urinary bile acid concentrations gradually increased until 1 to 2 months of age. After this peak of excretion (30 to 60 µmol/mmol creatinine), total urinary bile acid concentrations gradually decreased to less than 20 µmol/mmol creatinine. The percentage of usual bile acids (mainly cholic acid) relative to total urinary total bile acids gradually deceased from approximately 30% at birth to less than 15% at 7 months of age. On the other hand, 1β-hydroxylated bile acids (mainly 1β,3α,7α,12α-tetrahydroxy-5β-cholan-24-oic acid) relative to total urinary bile acids were increased gradually from 60% at birth to reach 70% to 80% at 1 month of age. The percentage of 1β-hydroxylated bile acids relative to total urinary bile acids then remained stable at a high percentage (70% to 90%) until the age of 7 months.
Conclusion:  Physiological cholestasis in preterm infants persists longer than in full-term infants. Moreover, as large amounts of cholic and 1β,3α,7α,12α-tetrahydroxy-5β-cholan-24-oic acids were detected in urine from preterm infants during this study, the 25-hydroxylation pathway may be particularly important for bile acid synthesis in early preterm infants.     

Urinary bile alcohol profile in infants with intrahepatic cholestasis: identification of 5beta-cholestane-3alpha,7alpha,24,25-tetrol

Urinary bile acids and bile alcohols were examined in six infants aged between 1 and 6 mo who had intrahepatic cholestasis. Following extraction, hydrolysis and solvolysis, cholanoids were analysed by gas-liquid chromatography and gas-liquid chromatography-mass spectrometry. The relative ratio of the urinary excretion of bile alcohols to bile acids was very low (0.07-0.22) in three patients with mild to severe cholestasis, whereas the urinary excretion of bile alcohols was 2-4 times greater than that of the total bile acids in three patients with slight cholestasis. The urinary bile alcohol spectrum in infants appears to be quite different from that in adults. Although the major bile alcohol was 27-nor-5beta-cholestane-3alpha,7alpha,12alpha,24 ,25-pentol, comprising more than 50% of total urinary bile alcohols in healthy adults, it accounted for only 35% of total urinary bile alcohols in our patients. In addition, bile alcohols carrying chenodeoxycholic acid type nucleus were detected in our patients by comparison of the retention times and mass spectra with those of authentic standards. The presence of 5beta-cholestane-3alpha,7alpha,24,25-tetrol confirmed for the first time in this study may represent an alternative pathway for chenodeoxycholic acid biosynthesis via a "25-hydroxylation pathway" in early life.     

Unconjugated, Glycine-conjugated, Taurine-conjugated Bile Acid Nonsulfates and Sulfates in Urine of Young Infants with Cholestasis

ABSTRACT. A direct assay system for conjugated bile acids using an enzymatic procedure and high-performance liquid chromatography was used for the analysis of urinary bile acid profiles in young infants with intrahepatic cholestasis (idiopathic neonatal hepatitis syndrome) or extra-hepatic biliary atresia. The major urinary bile acids were cholate and chenodeoxycholate conjugates, but a small amount of deoxycholate and 3β-hydroxy-5-cholenate conjugates were detected. Although there was no significant difference in total bile acid excretion between patients with intrahepatic cholestasis and extrahepatic biliary atresia, mean ratios of cholate to chenodeoxycholate and sulfated to total urinary bile acids were different between the two groups examined (5.63±2.83 vs. 2.50±1.25, p <0.05, 15.8±9.9 vs. 34.5±9.9%, p < 0.005). The proportion of taurine-conjugated chenodeoxycholate in the sulfate fraction to the total bile acid was lower in intrahepatic cholestasis, compared with that in biliary atresia (7.7±7.5 vs. 22.7±7.8 %, p < 0.005). The greater ratio of cholate to chenodeoxycholate and the reduced excretion of sulfated urinary bile acids in intrahepatic cholestasis was due to decreased taurine-conjugated chenodeoxycholate sulfate excretion.     

Maternal and fetal circulation of unusual bile acids: A pilot study

Background: Large amounts of unusual bile acids are synthesized by the fetal liver in late gestation. These compounds are mostly transferred from fetus to mother, although some are excreted into the amniotic fluid. We investigated the role of placental transfer of bile acids in fetal bile acid metabolism, particularly with respect to the unusual bile acids (1β‐hydroxylated and ketonic bile acids). Methods: We measured concentrations of bile acids in umbilical cord blood and urine of newborn infants, and in perinatal maternal serum and urine, using gas chromatography‐mass spectrometry. Serum and urine specimens from healthy non‐pregnant women were used as controls. Results: In newborn infants at delivery, cord blood and urine contained mostly primary and 1β‐hydroxylated bile acids, respectively. We also detected large amounts of ketonic bile acids in their urine, and the urinary concentration of total bile acids was elevated. Main maternal bile acids at 30 and 35 weeks of gestation and at delivery were 1β‐hydroxylated bile acids. After delivery, main bile acids changed from 1β‐hydroxylated bile acids to primary bile acids (P < 0.03), which also predominated in healthy non‐pregnant women. Conclusion: Fetally synthesized unusual bile acids were transported from fetus to mother. Pregnant women appear to excrete these bile acids into the urine, lowering both fetal and maternal serum bile acid concentrations.     

Bile Acid Abnormalities and the Diagnosis of Cerebro-Hepato-Renal Syndrome (Zellweger Syndrome)

ABSTRACT. The Zellweger or cerebro-hepato-renal syndrome (CHRS) is a congenital disorder characterized by cerebral dysfunction, craniofacial dysmorphic features, transient cholestasis and renal cysts. Patients fail to thrive, and usually die in their first year of life. In some cases, a definite diagnosis on purely clinical signs might not be possible. Several biochemical abnormalities have been observed in these patients and some of them have been tested as diagnostic markers. The aim of this study is to evaluate bile acid metabolites as biochemical markers of the CHRS. From a study of 20 CHRS patients, we conclude that screening for the presence of coprostanic acids and the C-29 dicarboxylic bile acid in serum or urine is a reliable method for detection of CHRS and confirmation of the diagnosis.     

Perinatal bile acid metabolism: analysis of urinary unsaturated ketonic bile acids in preterm and full-term infants

AIM: To compare urinary concentrations of unsaturated ketonic bile acids in preterm and full-term infants. METHODS: Urinary unsaturated ketonic bile acids were determined using gas chromatography-mass spectrometry. RESULTS: Urinary concentrations of total bile acids in early preterm infants (of less than 29wk gestational age) exceeded concentrations in late preterm (between 30 and 37 wk) and full-term infants (between 38 and 41 wk; p < 0.01). The percentage of ketonic bile acids (7alpha, 12alpha-dihydroxy-3-oxo-4-cholenoic acid and 7alpha-hydroxy-3-oxo-4-cholenoic acid) among total urinary bile acids in full-term infants (20.2 +/- 14.1%) was higher than that in early preterm infants (8.94 +/- 8.1%; p < 0.05). The percentage of unsaturated bile acids (3beta-hydroxy-delta5-bile acids) among total bile acids in urine did not differ greatly between groups. CONCLUSION: The percentage of 3-oxo-delta4 bile acids among total bile acids in urine gradually increased from early to late preterm infants, while healthy full-term infants excreted large amounts of 3-oxo-delta4 bile acids in urine at delivery.     

Tyrosinemia type Mike disease: A possible manifestation of 3-oxo-Δ4-steroid 5β-reductase deficiency

Abstract Background: It has been suggested that quantitative analysis of urinary bile acids may help to distinguish primary 3-oxo-δ⁴-steroid 5β-reductase deficiency from the excretion of 3-oxo-δ⁴ bile acids that occurs as a result of liver damage. Methods: Urinary bile acids were quantitatively analyzed by gas chromatography-mass spectrometry in four Japanese patients with severe neonatal cholestasis associated with hypertyrosinemia without urinary succinyl-acetone (i.e. tyrosinemia type I-like disease). These four patients represented sporadic cases. Results: Large amounts of 3-oxo-δ⁴ bile acids were detected, which comprised greater than 80% of the total urinary bile acids. Small amounts of allo-bile acids and primary bile acids were also detected, comprising less than 1% and 15% of the total urinary bile acids, respectively. Conclusions: It was suspected that these four patients had a primary 3-oxo-δ⁴-steroid 5β-reductase deficiency. However, it is possible that some patients in this study may have had a secondary 3-oxo-δ⁴-steroid 5β-reductase deficiency, caused by idiopathic neonatal cholestatic liver failure.     

Bile acids in serum and bile of infants with cholestatic syndromes

The concentration of individual bile acids in serum was measured in 18 neonates and infants with various cholestatic conditions (extrahepatic biliary atresia, neonatal hepatitis syndrome, chronic intrahepatic cholestasis and posthemolytic cholestasis). The cholate/chenodeoxycholate ratio in serum was smaller than one in all patients with neonatal hepatitis syndrome or extrahepatic biliary atresia, cholestatic conditions which were accompanied by signs of liver cell injury. It was greater than one in the patients with chronic intrahepatic cholestasis. Administration of cholestyramine to patients with patent extrahepatic bile ducts decreased the total concentration bile acids in serum and elevated the cholate/chenodeoxycholate ratio. Thus, cholestyramine administration may be of diagnostic value for evaluation of bile duct patency in cholestasis of infancy. Differences between the bile acid pattern in serum and bile were observed. Thus, the cholate/chenodeoxycholate ratio was always higher in bile than in serum. 3beta-hydroxy-5-cholenoic acid found in serum was not detectable in bile. This finding suggests that impairment of biliary excretion rather than increased hepatic synthesis is responsible for elevation of this monohydroxy bile acid in serum.     

Investigation of serum bile acids; seven patients with Alagille syndrome

To clarify whether an abnormal bile acid pattern has a role in the pathogenesis of Alagille syndrome, we compared serum bile acid patterns in seven with Alagille syndrome with those of patients with congenital biliary atresia (CBA), neonatal hepatitis (NH) and normal infants.Of the seven patients with Alagille syndrome, four patients were younger and three were older than 1 year. The mean total serum bile acid level in the infants was higher than in older subjects. There was a dissociation between the levels of serum total bile acid and bilirubin in three of the seven cases. The mean total bile acid levels in serum were in the following decreasing order: CBA, Alagille syndrome, NH and controls.The ratio of cholate to chenodeoxycholate in the younger patients with Alagille syndrome was significantly higher than CBA (P<0.001). However, no specific bile acid pattern was found in Alagille syndrome by high-performance liquid chromatography (HPLC).Abbreviations TBA total bile acids - FBA free bile acids - conj-BA conjugated bile acids - C/CDC ratio of cholate to chenodeoxycholate - G/T ratio of glycine conjugates to taurine conjugates - GPT glutamic pyruvic transaminase - CBA congenital biliary atresia - NH neonatal hepatitis - HPLC high performance liquid chromatography - GCA glycocholate - TCA taurocholate - GCDCA glycochenodeoxycholate - TCDCA taurochenodeoxycholate     

Evidence for defective primary bile acid secretion in children with progressive familial intrahepatic cholestasis (Byler disease)

To clarify the relationship of progressive familial intrahepatic cholestasis (Byler disease) to bile acid metabolism, we analysed, by high performance liquid chromatography, the bile acid composition of serum and bile in seven children with Byler disease and in eight control children with other cholestatic diseases. In serum, total bile acid concentration was increased in patients with Byler disease (0.30±0.05 mmol/l) and in control patients (0.21±0.08 mmol/l). Cholate (C) and chenodeoxycholate (CDC) cornprised the major proportion of total bile acids in patients with Byler disease as in control patients. Hyocholate (HC) was only detected in patients with Byler disease and lithocholate was only present in control children. In bile, total bile acid concentration was very low in patients with Byler disease (1.1±1.4 mmol/l) compared to control patients (88.9±83.2 mmol/l). C and CDC were the major bile acids in control patients, whereas C and HC comprised the major proportion of bile acids in patients with Byler disease. These results suggest the existence of a defect of primary bile acid secretion in Byler disease characterized by the presence of high concentration of bile acids in serum and absence or very low concentration of bile acids in bile.     

BILE ACID METABOLISM IN LOW BIRTHWEIGHT INFANTS

Strandvik, B. (Department of Paediatrics and Research Center, Karolinska Institute, Huddinge University Hospital, Stockholm, Sweden). Bile acid metabolism in low birth weight infants. Acta Paediatr Scand, Suppl. 296: 71, 1982.—The early synthesis of bile acids is described. From 28 weeks of gestation cholic acid is the predominating bile acid, but the bile acid pool is very small in preterm infants, leading to a low intraduodenal concentration of bile acids, especially during digestion of meals. Preliminary results indicate that preterm infants excrete less cholic and more 3β0H5-cholenoic acid in the urine than fullterm infants indicating a functional minor pathway in the synthesis of chenodeoxycholic acid. The tetrahydroxylated bile acids, which tend to dominate in the urine of newborn, have been shown to be products of the most common bile acid in cholestatic adults, and may thus be the result of a "physiological cholestasis" in the newborn.     

Bile and acid reflux in the pathogenesis of reflux oesophagitis in children

Aim:   The aim of this study was to investigate the role of bile and acid reflux in the pathogenesis of reflux oesophagitis (RE) in children.
Methods:   A total of 44 patients aged 5–17 years with gastro-oesophageal reflux symptoms were enrolled. Simultaneous 24-h oesophageal Bilitec 2000 (Medtronic Instruments, Minneapolis, MN, USA) bilirubin monitoring and pH monitoring, in biopsy of oesophageal mucosa by gastro-endoscopy, were performed in all patients.
Results:   According to the diagnostic criteria of pathological acid reflux and pathological bile reflux, 10 of 44 cases (22.7%) had acid reflux, 10 (22.7%) had isolated bile reflux, 16 (36.4%) had mixed acid and bile reflux, and the other eight (18.2%) had no reflux. Significant difference was observed in the ratio of different patterns of reflux between the RE group (26 cases) and the non-erosive reflux disease (NERD) group (18 cases) (χ² = 9.096, P  < 0.01). All the parameters of acid reflux in the RE group were higher significantly than that in the NERD group ( P  < 0.05 or P  < 0.01). A total of 20 out of 26 cases (76.9%) with RE had oesophageal acid reflux as against six out of 18 cases (33.3%) in patients with NERD ( P  < 0.01). The difference of each parameter of bile reflux had not reached significance between the two groups.
Conclusions:   Mixed reflux is the predominant form of reflux in the causation of oesophageal mucosal injury in children. Isolated bile reflux also plays a role in the development of RE, although only in patients without acid reflux.     

Cholestasis in neonatal intensive care unit: incidence, aetiology and management

Aim:  Prevalence, aetiology, management and outcome of cholestasis were evaluated in infants admitted to neonatal intensive care unit (NICU).
Methods:  Medical records of all infants admitted to two Italian level III NICUs from January 2005 to August 2007 were retrospectively reviewed. The role of ursodeoxycholic acid (UDCA) therapy was also investigated.
Results:  Twenty-seven of 1289 enrolled infants developed cholestasis. In 25 infants, cholestasis had a multifactorial basis, while in two, no aetiology was found. UDCA did not significantly affect clinical and biochemical course of cholestasis. During a period of 12 months, eight cholestatic infants died, one underwent liver transplantation and 18 fully recovered.
Conclusion:  Infants admitted in NICU have a rate of cholestasis higher than that reported in the general population of live births; in most cases, cholestasis is associated to multiple risk factors and shows a favourable outcome. UDCA does not seem to affect clinical course of cholestasis in this setting.     

Differential diagnosis of cholestasis following allogeneic hematopoietic stem cell transplantation in children: the contribution of serum bile acid levels in relation to other liver function tests

Hepatic complications associated with cholestasis occur frequently in hematopoietic stem cell transplant recipients. Since bile acid seems to be a sensitive indicator of beginning cholestasis, the authors monitored total serum bile acid levels in addition to the standard liver function tests in 23 recipients of allogeneic transplants between June 1999 and September 2000. The observations suggest that bile acid is an early and sensitive marker of hepatic GvHD but not as specific as bilirubin. For cholestasis in absence of hepatic GvHD bile acid seems to be more sensitive than bilirubin. Routinely monitoring of bile acid after hematopoietic stem cell transplantation is not indicated.     

Unconjugated, glycine-conjugated, taurine-conjugated bile acid nonsulfates and sulfates in urine of young infants with cholestasis

A direct assay system for conjugated bile acids using an enzymatic procedure and high-performance liquid chromatography was used for the analysis of urinary bile acid profiles in young infants with intrahepatic cholestasis (idiopathic neonatal hepatitis syndrome) or extra-hepatic biliary atresia. The major urinary bile acids were cholate and chenodeoxycholate conjugates, but a small amount of deoxycholate and 3 beta-hydroxy-5-cholenate conjugates were detected. Although there was no significant difference in total bile acid excretion between patients with intrahepatic cholestasis and extrahepatic biliary atresia, mean ratios of cholate to chenodeoxycholate and sulfated to total urinary bile acids were different between the two groups examined (5.63 +/- 2.83 vs. 2.50 +/- 1.25, p less than 0.05, 15.8 +/- 9.9 vs. 34.5 +/- 9.9%, p less than 0.005). The proportion of taurine-conjugated chenodeoxycholate in the sulfate fraction to the total bile acid was lower in intrahepatic cholestasis, compared with that in biliary atresia (7.7 +/- 7.5 vs 22.7% +/- 7.8%, p less than 0.005). The greater ratio of cholate to chenodeoxycholate and the reduced excretion of sulfated urinary bile acids in intrahepatic cholestasis was due to decreased taurine-conjugated chenodeoxycholate sulfate excretion.     

Phenobarbital can aggravate a cholestatic bile acid pattern in infants with obstructive cholangiopathy

The effect of phenobarbital on urinary bile acid excretion in intrahepatic cholestasis was studied in four boys 4-43 months of age who received 10 mg/kg of body weight of phenobarbital for a period of 3 weeks-3 years. One child was observed at two different periods: with and without histologically proven cirrhosis. Before the treatment period, the infants excreted 10-fold higher amounts of bile acids in urine than healthy children. The primary bile acids predominated, and there were also increased amounts of polyhydroxylated bile acids, 3 beta-hydroxy-5-cholenoic acid, and ketonic bile acids but small amounts of secondary bile acids. After the phenobarbital treatment, the patients further increased their urinary bile acid excretion, including all kinds of bile acids except the secondary ones. The sulfated fraction did not increase in absolute amounts, and its relative percentage decreased from a mean of 60-33%. Liver function test results generally did not improve, although serum concentration of bilirubin decreased. Most of these changes suggested a worsening of the cholestatic state after phenobarbital treatment. The results indicate that at our present state of knowledge, phenobarbital should not be given routinely to infants or children with intrahepatic cholestasis.     

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??Objective??To detect the level of fecal primary and secondary bile acids in infants with infantile cholestatic hepatopathy??ICH??and analyze its clinical value. Methods??Thirty infants with ICH were enrolled in this study??who were diagnosed with infantile cholestatic hepatopathy. Thirty infants with good health condition were enrolled as the healthy control group. The fecal samples were collected respectively in the preparatory treatment phase and treatment phase from infants with ICH and from the healthy infants. Bile acids were extracted from infants’ feces and were quantitatively analyzed by liquid chromatography-mass spectroscopy. Results??Among the fecal primary bile acids??the level of cholic acid??chenodeoxycholic and glycochenodeoxycholic acid both in the ICH preparatory treatment group and ICH treatment group was significantly lower than that in the healthy control group??P??0.016??.The level of fecal cholic acid and chenodeoxycholic acid of ICH treatment group was higher than in the ICH preparatory treatment group??P??0.016??. Among the fecal secondary bile acids??the level of lithocholic acid both in the ICH preparatory treatment group and ICH treatment group was significantly lower than that in the healthy control group??P??0.016????and the level of ursodeoxycholic acid in the ICH preparatory treatment group was lower than that in the ICH treatment group and healthy control group??P??0.016??. Conclusion??In infants with ICH, the changes of fecal primary bile acids and fecal secondary bile acids have their own characteristics at the early stage of treatment, which may be caused by the short-term treatment, the prognosis of the disease itself and the changes of intestinal function, including intestinal bacteria. Clinical attention should be paid to these changes.     

Profile of urinary bile acids in familial intrahepatic cholestasis with Coombs'negative haemolytic anaemia

We present two male siblings with intrahepatic cholestasis and prolonged indirect hyperbilirubinaemia. Their familial intrahepatic cholestasis syndrome was characterized by Coombs'negative haemolytic anaemia, without giant cell transformation of hepatocytes and high concentrations of serum γ-glutamyl transpeptidase and cholesterol. By gas chromatography-mass spectrometry, we detected large amounts of 1β-hydroxylated bile acids, especially lβ,3α,7α,12α-tetrahydroxy-5β-cholan-24-oic acid (25.5-67.9% of total urine bile acids) in the urine during phenobarbital therapy. However, the amount of urinary 1β-hydroxylated bile acids gradually decreased as the disease progressed. At the end-stage, we detected large amounts of 7α,12α-dihydroxy-3-oxochol-4-en-24-oic acid (19.6% of total urine bile acids). The ratio of 7α,12α-dihydroxy-3-oxochol-4-en-24-oic acid to cholic acid in the urine was 0.8. We conclude that in infants with end-stage liver failure, the microsomal hydroxylation of bile acids is impaired and the excretion of Δ⁴-3-oxo bile acids is increased. Familial intrahepatic cholestasis, Coombs'negative haemolytic anaemia, 1β-hydroxylated bile acids, unsaturated ketonic bile acids     

BILE ACID EXCRETION AND MALABSORPTION IN INTRAHEPATIC CHOLESTASIS OF INFANCY ("NEONATAL HEPATITIS")

Bile acid excretion has been studied in four patients with intrahepatic cholestasis of infancy neonatal hepatitis) after intramuscular administration of cholic acid-24-¹⁴.C Bile acid secretion to the intestines was found to be highly impaired and the main route of excretion was via the urine. Practically all of the administered labeled cholic acid was conjugated prior to excretion. The main conjugates were glycocholic and taurocholic acid. At least three additional conjugates of cholic acid were isolated from the urine. Analysis of bile obtained from three of the patients in connection with operative cholangiography showed a very low concentration of bile acids, phospholipids and cholesterol. The bile was of normal colour owing to the presence of bilirubin. Severe steatorrhea and markedly impaired absorption of vitamin A was demonstrated when the patients were jaundiced. The impairment of bile acid excretion to the gut and the degree of steatorrhea were well correlated. In some of the patients steatorrhea persisted after the disappearance of jaundice. In those instances, the impairment of bile acid excretion to the gut was found to persist.     

3beta-hydroxy-delta5 -C27-steroid dehydrogenase deficiency: diagnosis and treatment

The aim of this study was to evaluate the effects of bile acid treatment and to obtain further information about the pathway of bile acid biosynthesis in a patient with 3beta-hydroxy-delta5-C27-steroid dehydrogenase/isomerase (3beta-HSD) deficiency by gas chromatography-mass spectrometry. Results showed that at 2 months of age, 3beta-hydroxy-5-cholen-24-oic acid (3.0 micromol/mmol Cr, 7.9%) was detected in the urine in essentially the same relative amount as 3beta,7alpha-dihydroxy- and 3beta,7alpha,12alpha-trihydroxy-5-cholen-24-oic acids (3.7 micromol/mmol Cr, 9.8%) during ursodeoxycholic acid treatment combined with prednisolone. As a result, diagnosis was delayed until 18 months of age. One month later with substitution of chenodeoxycholic acid treatment, urinary 3beta,7alpha-dihydroxy- and 3beta,7alpha,12alpha-trihydroxy-5-cholen-24-oic acids decreased significantly, and subsequent improvement of liver dysfunction was accelerated. Chenodeoxycholic acid treatment is useful in 3beta-HSD deficiency. However, in the diagnosis of this disease in early life, it should be noted that the acidic pathway may be the major route for bile acid biosynthesis in the neonatal period. Diagnosis of 3beta-HSD deficiency may have been delayed by administration of ursodeoxycholic acid, resulting in prolonged diagnostic investigation in this child with cholestasis. Further, use of prednisolone may have been contraindicated.