Approach Considerations
All patients with impaired bilirubin conjugation have an elevated total serum bilirubin level that is due primarily to the unconjugated form; however, the level of elevation varies according to the underlying disease process.
Transient elevations of plasma bilirubin may be seen in healthy neonates. The plasma bilirubin usually returns to normal within 10 days. For infants in whom the plasma bilirubin level remains elevated, search for the inherited disorders of bilirubin metabolism.
In neonates, transcutaneous devices that use multiwavelength spectral reflectance can be used to estimate total serum bilirubin levels and avoid blood sampling. At higher total serum bilirubin levels, the transcutaneous measurements may underestimate the total serum bilirubin concentration; therefore, serum measurements should be obtained. Also, the transcutaneous measurements are not reliable in infants undergoing phototherapy.
In a study, Cakmak et al suggested that by measuring haptoglobin levels from the cord blood, neonatologists and pediatricians could stratify neonates into high- versus low-risk groups for developing jaundice, leading to earlier intervention. Because haptoglobin levels decrease during hemolysis, which in turn plays a significant role in raising the serum bilirubin levels in neonates, the investigators examined the relationship between decreasing haptoglobin levels and the risk of jaundice in 84 term babies. [66]
The average gestational age of the mothers was 39.5 ±1.5 weeks. The authors noted a negative correlation between haptoglobin levels drawn from the umbilical cord blood and bilirubin values on the fifth postpartum day.
Crigler-Najjar Syndrome
Although no simple, widely available clinical test is available to confirm the diagnosis of Crigler-Najjar syndrome, unconjugated hyperbilirubinemia in the presence of normal liver function test findings is characteristic of the disease.
Direct bilirubin is less than 15% of the total serum bilirubin in Crigler-Najjar syndrome. High-performance liquid chromatography analysis of duodenal bile reveals that, in Crigler-Najjar syndrome type 1, negligible bilirubin diglucuronides or monoglucuronides are present; in the type 2 syndrome, these conjugates are present but in low concentrations. DNA analysis can be very helpful in establishing the correct diagnosis.
Persistent unconjugated hyperbilirubinemia levels of more than 20 mg/dL after the first week of life in the absence of liver disease or hemolysis strongly suggests UGT deficiency.
Findings on abdominal imaging studies, such as plain radiography, computed tomography (CT) scanning, and ultrasonography, are normal in Crigler-Najjar syndrome.
Definitive diagnosis of Crigler-Najjar syndrome requires high-performance liquid chromatography of bile or a tissue enzyme assay of a liver biopsy sample.
Crigler-Najjar syndrome type 1
Except for the presence of high serum unconjugated bilirubin levels, the results of liver tests in Crigler-Najjar syndrome type 1 are normal. Serum bilirubin levels range from 20-50 mg/dL. Conjugated bilirubin is absent from serum, and bilirubin is not present in urine.
Bile collected through duodenal aspiration is light yellow because of small amounts of unconjugated bilirubin. Bilirubin conjugates are nearly absent from the bile.
Crigler-Najjar syndrome type 2
Crigler-Najjar syndrome type 2 results in lower bilirubin concentrations than does type I, with levels ranging from 7-20 mg/dL. Higher bilirubin levels may be seen if coexisting hemolysis or an intercurrent illness is present.
This disorder may be distinguished definitively from type 1 by chromatographic analysis of pigments excreted in bile. In type 2, bile contains significant amounts of conjugated bilirubin, although the proportion of bilirubin monoglucuronide in bile is increased.
Transferase activity measurements and the response to phenobarbital treatment can also distinguish Crigler-Najjar syndrome type 1 from type 2. Phenobarbital has no effect in type 1 but causes an approximately 25% reduction in plasma bilirubin level in most patients with type 2.
Liver function testing
Liver enzyme levels are usually within the reference range. Occasionally, however, these levels may be somewhat elevated, as a result of intrahepatic cholestasis.
Percutaneous liver biopsy
Liver biopsy reveals normal histology other than the occasional bile plugs in the bile canaliculi. Bile is sometimes observed in the portal triad, in dilated bile canaliculi, in hepatocytes, and in the Kupffer cells.
Enzymatic assay of liver tissue reveals absent UGT activity in Crigler-Najjar syndrome type 1 and diminished activity in Crigler-Najjar syndrome type 2.
Gilbert Syndrome
As a rule, Gilbert syndrome can be diagnosed by a thorough history and physical examination and confirmed by standard blood tests. Repeated investigations and invasive procedures are not usually justified for establishing a diagnosis.
Hyperbilirubinemia is the only biochemical serum abnormality in Gilbert syndrome. Serum bilirubin concentrations range from 1-5 mg/dL. Two provocative tests, energy deprivation and nicotinic acid administration, have been used to diagnose the condition. However, a significant number of false-positive and false-negative results limit the value of these tests in patients with marginal elevation of serum bilirubin concentration.
A polymerase chain reaction (PCR) assay has also been introduced to identify TA repeats and may be used as a screening test.
A complete blood count (CBC), including a reticulocyte count and a blood smear, is a useful screening test for excluding hemolysis. Rarely, red blood cell abnormalities resembling variegate porphyria, possibly resulting from the increased hepatocellular bilirubin concentration, have been described in persons with Gilbert syndrome.
Serum lactate dehydrogenase (LDH) levels are elevated in persons with hemolysis but are normal in those with Gilbert syndrome.
A familial increase in serum alkaline phosphatase (ALP) levels has been reported in some persons with Gilbert syndrome.
Additional diagnostic tests are rarely required, because a diagnosis of Gilbert syndrome can generally be made on the basis of the following findings:
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Unconjugated hyperbilirubinemia noted on several occasions
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Normal results from the CBC, reticulocyte count, and blood smear
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Normal liver function test (LFT) results
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An absence of other disease processes
Nevertheless, certain specialized tests (including some that are of historical interest, as well as the newer molecular genetic techniques) are occasionally performed to confirm a diagnosis of Gilbert syndrome. These tests are described below to introduce the clinician to the broad diagnostic armamentarium available for diagnosing Gilbert syndrome. Recourse to these specialized tests should be rare and is usually difficult to justify in clinical practice, given that the diagnosis of Gilbert syndrome is generally straightforward.
Fasting test
Within 48 hours of the start of a fast, there is usually a 2- to 3-fold rise in the plasma unconjugated bilirubin level, which returns to normal levels within 24 hours after resumption of a normal diet. Although unconjugated bilirubin levels also rise with fasting in patients with hemolysis or liver disease, the magnitude of the rise is less than that observed with Gilbert syndrome. A similar rise in plasma bilirubin is also observed with normocaloric diets deficient in lipids and reverses promptly with lipid replacement.
Nicotinic acid test
Intravenous (IV) administration of 50 mg of nicotinic acid results in a 2- to 3-fold rise in plasma unconjugated hyperbilirubinemia within 3 hours. The mechanisms are multifactorial and probably related to the following:
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Elevated osmotic fragility of red blood cells
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Increase in splenic production of bilirubin
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Transient inhibition of hepatic bilirubin-UGT activity
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Increased splenic heme oxygenase activity
Because a similar, if less impressive, increase is observed in healthy individuals and those with hemolysis or liver disease, the nicotinic test, like the fasting test, does not clearly distinguish patients with Gilbert syndrome from those who are healthy or who have other disease processes.
Phenobarbital test
Phenobarbital and other enzyme inducers of the bilirubin-UGT system will normalize plasma bilirubin levels in patients with Gilbert syndrome. This effect is predominantly due to the accelerated bilirubin clearance from enzyme induction, but it is also due to reduced bilirubin turnover. Steroids can also reduce plasma bilirubin levels in Gilbert syndrome, by increasing the hepatic uptake and storage of bilirubin.
Radiolabeled chromium test
The radiolabeled chromium test is used to measure red blood cell survival. As many as 60% of patients with Gilbert syndrome have a mild and fully compensated state of hemolysis together with increased hepatic heme production. This means that hyperbilirubinemia may develop due to reduced clearance, as well as increased production, of bilirubin, with higher production resulting from increased erythroid or hepatic heme turnover.
Thin-layer chromatography
Thin-layer chromatography is diagnostic of Gilbert syndrome when it shows a significantly higher proportion of unconjugated bilirubin than is seen in individuals with chronic hemolysis or liver disease or is found in healthy individuals. If confirmation of the diagnosis is truly essential, chromatographic determination is of potential use. In Gilbert syndrome, this shows an increased ratio of bilirubin monoglucuronide to diglucuronide, reflecting reduced bilirubin-UGT activity.
Drug clearance test
In approximately 30% of patients with unconjugated hyperbilirubinemia, there is impaired clearance of bromosulfophthalein, indocyanine green, and free fatty acid, suggesting an abnormality in hepatic uptake, transport, or both. Metabolic clearance of tolbutamide is also reduced in persons with Gilbert syndrome, but because the drug does not undergo glucuronidation, hepatic uptake appears to be defective.
Plasma clearance of most drugs that undergo glucuronidation (eg, benzodiazepines) is unaffected. With regard to acetaminophen, however, patients with Gilbert syndrome are a heterogeneous group, with some demonstrating normal metabolism and others exhibiting marked reduction in glucuronidation and an increase in oxidation. [67, 68] These changes suggest that people in this subgroup may be more susceptible to liver injury after an acetaminophen overdose, though no such adverse events have been reported.
Polymerase chain reaction assay
PCR assay is a novel and rapid method of identifying genetic polymorphisms in the TATA box of the UGT1A1 gene by using fluorescence resonance energy transfer. It is well recognized that genetic testing can confirm the diagnosis of Gilbert Syndrome in settings where there is diagnostic confusion.
Imaging studies
Japanese researchers have reported that patients with schizophrenia associated with Gilbert syndrome have specific changes in the signal intensities on fluid-attenuated inversion-recovery magnetic resonance imaging (FLAIR MRI) scans. This suggests that schizophrenia with associated Gilbert syndrome may produce changes in the frontotemporal cortex, limbic system, and basal ganglia.
Percutaneous liver biopsy
Liver biopsies are not performed routinely in Gilbert syndrome and are rarely necessary. Histologically, the liver is normal in persons with this disorder, except for occasional accumulation of a lipofuscin-like pigment around the terminal hepatic venules.
Neonatal Jaundice and Ineffective Erythropoiesis
Physiologic jaundice
In physiologic jaundice, the peak total serum bilirubin level is 5-6 mg/dL (86-103 µmol/L), occurs at age 48-120 hours, and does not exceed 17-18 mg/dL (291-308 µmol/L).
Breast milk jaundice
In breast milk jaundice, the bilirubin can increase to levels as high as 20 mg/dL, necessitating the need for phototherapy and the discontinuation of breastfeeding.
Increased production of bilirubin
Ineffective erythropoiesis (ELB production) is characterized by a marked increase in fecal urobilinogen excretion and a normal or near-normal red blood cell lifespan.
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Production of bilirubin.
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Enterohepatic circulation of bilirubin.
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Conjugation of bilirubin.
