Inherited Metabolic and Developmental Disorders of the Pediatric and Adult Liver

Architecture

The liver undergoes substantial growth after birth. It normally doubles in weight within the first month of life, doubles again during the first year of life, and does not reach its mature size until late adolescence. The portal tract system grows in parallel with the liver. Therefore the most peripheral aspects of the liver may exhibit developmental residua of fetal histology. For instance, residual bile duct plates may rim the portal tracts, and the latter contain a more cellular mesenchyme and a centrally placed portal vein ( Fig. 55.1 ). The dimensions of hepatic lobules remain constant with growth. However, hepatocyte cords may remain two cells thick well into the fourth postnatal year. This should not be misinterpreted as regenerative hyperplasia in response to tissue injury.

Cell Populations

Hematopoietic elements are commonly present in liver biopsy specimens obtained during the postnatal months. Granulopoiesis predominates in portal tracts, whereas erythropoiesis is common in the parenchyma.

Hepatocyte Content

Until a postnatal age of approximately 3 months, hepatocytes normally contain copper-binding protein and copper (demonstrable by orcein and rhodanine techniques, respectively) and granules of hemosiderin, particularly in periportal hepatocytes. These deposits are considered to be physiological and disperse with time. Conversely, hepatocyte alterations characteristic of various storage disorders may be inconspicuous in early infancy because of the time required to accumulate abnormal substances, such as α ₁ -antitrypsin (A1AT). One dramatic exception to the concept of physiological iron deposition occurs in newborns who exhibit liver failure at birth, which is usually attributable to severe liver injury in utero. In this scenario, marked iron deposits may be present in hepatocytes at birth, giving rise to the term neonatal iron storage disease or perinatal hemochromatosis . A severe degree of necrosis and fibrosis is also present in patients with this condition. The extrahepatic reticuloendothelial system does not exhibit iron accumulation, highlighting the primacy of the liver injury. The finding of severe perinatal hepatic siderosis is nonspecific and indicates the development of liver injury during gestation. A lesser degree of hemosiderosis, with reticuloendothelial system deposits, may be seen in cases of maternal–fetal blood group incompatibility with significant hemolysis.

Response to Injury

Giant multinucleated hepatocytes, with or without bile pigment, are often present in infants with liver disease, regardless of the etiology. This change is considered nonspecific and reactive. Multinucleated hepatocytes are formed by syncytial breakdown of cell-to-cell borders, but with partial preservation of the canalicular aspects of the cell membrane. Canalicular remnants with retained bile may be observed within the cytoplasm. Giant cells exhibit multiple nuclei, either scattered throughout the cytoplasm or clustered toward one pole of the cell. This reaction may persist well into childhood if the inciting disorder is not resolved. Multinucleated giant cell change is unusual in older children and adults, but it may occur in some disorders such as autoimmune hepatitis and paramyxovirus hepatitis. ^,

The histological spectrum of neonatal hepatitis includes giant cell change in hepatocytes, intralobular cholestasis, necrosis of hepatocytes, and intrahepatic hematopoiesis. All of these features are nonspecific events in infancy and can be observed in biliary atresia, A1AT storage disorder, and many other conditions ( Table 55.7 ). With advances in biochemistry and molecular genetics, many conditions formerly grouped within the category of neonatal giant cell hepatitis can now be specifically diagnosed, including progressive familial cholestasis types 2 and 3 and various bile salt synthetic defects.

Clinical Features

Neonatal hemochromatosis (NH), also termed neonatal iron storage disease, is a rare syndrome that is characterized by the presence of congenital cirrhosis and fulminant liver failure. This condition exhibits abundant iron deposition in the liver and in other organs, but not in the reticuloendothelial system (spleen or bone marrow). Clinically, patients with NH, either before birth or shortly thereafter, exhibit liver failure, including hypoglycemia, coagulopathy, hypoalbuminemia, ascites, and hyperbilirubinemia. Although some infants recover with exchange transfusion and some survive to transplantation, most die of their disease. Most cases of NH belong to the group of gestational alloimmune liver disease (GALD).

Pathogenesis

NH has been described in association with various conditions, including metabolic disorders (tyrosinemia, Δ4-3-oxosteroid 5β-reductase deficiency, mtDNA mutations, and Zellweger syndrome); infections (echovirus 9, cytomegalovirus [CMV], herpes simplex virus, rubella, and parvovirus B19); and karyotypic disorders (Down syndrome). ^, As a result, some authors regard NH as a final common phenotype of any gestational insult that culminates in abnormal iron metabolism. ^, At least three patterns of disease transmission have been described: transmission of maternal alloantibodies, autosomal recessive inheritance, and matrilineal inheritance. In the last instance, several reports have documented women with more than one affected child, but the children were fathered by different men. Although gonadal mosaicism in these mothers has not formally been excluded, the possibility of mitochondrial inheritance also has not been excluded. NH is not associated with mutations in the HFE gene, which is involved in most cases of HH (see later discussion).

In 2004, Whitington and Hibbard first reported giving high-dose intravenous γ-globulin to pregnant women with a history of a previously affected infant, and this treatment prevented recurrence in a small series. Their findings have since been confirmed in many centers, and it is now imperative to determine the etiology of all perinatal forms of acute liver injury. Mimicry of hemochromatosis has been seen in cases of mtDNA depletion and other circumstances, which presumably do not benefit from intravenous immunoglobulin therapy.

The antibody responsible for NH has yet to be determined. Complement fixation induced by immunoglobulin G of maternal origin is detected by immunofluorescence for the membrane attack complex on hepatocytes. One patient with cirrhosis at birth did not have hemosiderosis and survived without specific therapy. However, the usual scenario is fatal necrosis with extensive parenchymal siderosis without reticuloendothelial iron, suggesting dysfunction of macrophages. This has been observed even prenatally: Three of eight stillborn or very premature infants had no extrahepatic siderosis. ^, Recently, Whitington and colleagues have provided insight into the severe degree of parenchymal siderosis that accompanies the hepatocellular injury. The injured livers have significantly reduced hepcidin, hemojuvelin, and transferrin gene expression compared with normal livers.

Antibodies to mitochondrial proteins are responsible for primary biliary cirrhosis, a disease that affects women disproportionately. Two patients transmitted this disease transplacentally to their infants, producing transient liver injury. One infant was born with ascites and conjugated hyperbilirubinemia. In the other infant, the presentation was more insidious. His biopsy, at 5 weeks, showed portal inflammation involving bile ducts and ductules, mild portal fibrosis, and multinucleated giant hepatocytes typical of neonatal hepatitis. Immunofluorescence was able to detect immunoglobulin G deposits surrounding hepatocytes. Within 3 months, both infants showed no evidence of liver dysfunction, and the antibodies were undetectable.

NH resembles neonatal hepatitis in infants with systemic lupus erythematosus, in which the mother has high titers of antinuclear anti Ro (SSA) and anti La antibodies.

Pathological Features

The liver in patients with NH (usually seen at postmortem examination) reveals cirrhosis and cholestasis ( Fig. 55.2 ). Confluent areas of hepatocellular loss and a variable degree of hepatocyte regeneration are typical. Residual hepatocytes may demonstrate giant cell or pseudoacinar transformation. Iron deposition is typically coarsely granular and located predominantly within hepatocytes, sparing Kupffer cells. Extrahepatic sites of siderosis include the parenchymal cells of the heart, thyroid, pancreas, adrenal glands, kidneys, and the submucosal glands of the gastrointestinal and upper respiratory tracts. It has now been shown that examples of GALD exist without evidence of hepatic siderosis. While the diagnosis of this entity has been a challenge, the recognition of the deposition of the C5b-9 complex by immunohistochemistry on necrotic hepatocytes in 2010 has led to an increase in recognition of this entity. Dubruc et al. showed 100% expression in GALD though only 26% of their cases showed staining in more than 75% of hepatocytes. This has improved the outcome of future pregnancies because of the ability to administer IV-IG in the prenatal period of subsequent pregnancies. Having said this, there are issues with this staining technique because recent literature suggests overlap of staining in necrotic livers as a result of viral or other etiology, as well as suspected cases of GALD with negative C5b-9 staining. It is therefore important to exclude other causes, especially viral causes, of neonatal acute liver failure.

Natural History

NH carries a high risk of death, and a high index of suspicion is required to diagnose this disorder at first presentation, as this has impact on management of subsequent pregnancies for the mother. NH remains in the differential diagnosis of severely ill neonates who survive beyond the first days of life. Demonstration of siderosis in biopsy samples from oral submucosal glands is a rapid diagnostic method. T2-weighted magnetic resonance imaging (MRI) can be used to assess the presence of iron in various tissues. Siderosis of the liver and extrahepatic organs may also accompany other conditions, such as erythropoietic disorders, sickle cell disease, thalassemia, and erythroblastosis from blood group incompatibility. However, hemosiderosis affects the reticuloendothelial system primarily. These other diseases need to be excluded by other clinical and laboratory studies.

Clinical Features

Biliary atresia accounts for more than 30% of all cases of cholestasis in neonates. This disorder has an incidence of 1 in 5000 to 19,000 live births, occurring more frequently in Asian countries such as Taiwan and Japan when compared with North America and Europe. Most cases are sporadic and do not reveal a positive family history of neonatal cholestasis. A series of 30 sets of twins revealed only 2 sets with both infants affected by EHBA, and both pairs were dizygotic. There are reports of a 20-year-old mother who underwent a portoenterostomy for EHBA at 64 days of age and subsequently gave birth to a daughter with EHBA and of a family cluster of 5 children in which 2 dizygotic twin sisters and a third sibling all had EHBA. Variation in epigenetic modifications of genomic DNA has been suggested for these occasional familial presentations. Genes related to bile duct dysmorphogenesis (including ciliopathies) overlapping with features of biliary atresia in both humans and nonhuman model systems have been proposed, sparking continued interest in identifying potential causative and modifying genes relevant to patients with biliary atresia.

Infants presenting with biliary atresia are usually of normal gestational age and birth weight. Cholestatic jaundice is the main clinical presentation. It typically develops in the first few weeks of life and does not remit, unlike the mild physiological jaundice of early infancy. Furthermore, in physiological jaundice, the mildly elevated serum bilirubin is primarily unconjugated, and the serum levels of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) are normal. The progressive biliary obstruction that characterizes biliary atresia leads to a progressive increase in serum bilirubin levels in which conjugated bilirubin represents 50% to 80% of the total. A recent study of conjugated or direct-reacting bilirubin in the first 48 hours of life revealed significant elevations in all infants with biliary atresia compared with aged-matched controls, even though the total bilirubin level was not increased. Serum levels of GGT are increased several times above normal. Liver biosynthetic function, as indicated by serum albumin levels and prothrombin time, is usually normal at initial presentation.

In keeping with the aforementioned discussion, ultrasound should be performed in suspected cases of biliary atresia to exclude the presence of an extrahepatic biliary tract anomaly, such as a choledochal cyst. Hepatobiliary scintigraphy is also useful to assess the status of biliary tract function, but in many hepatocellular diseases of infants, there also is an absence of excretion of the labeled molecule into the intestine. Ultrasound shear wave elastography has also been shown to be discriminatory for biliary atresia.

Percutaneous liver biopsy is used to determine whether there is histological evidence of large bile duct obstruction. Biopsy features of atresia may also occur with other extrahepatic forms of biliary obstruction ( Box 55.8 ). The overall accuracy rate of percutaneous liver biopsy for diagnosis of biliary atresia has recently been shown to be around 90.1%, with a sensitivity of 88.4% and a specificity of 92.7%. The decision to proceed with a biliary tract exploratory surgical procedure does not rest solely on the pathological findings in a liver biopsy specimen. The presence of acholic stools, an undetectable or irregular contour of the gallbladder on sonographic studies, and failure to excrete into the intestine the radioactive tracer iminodiacetic acid by hepatobiliary scintography (HIDA scan) are findings that lend support to a diagnosis of biliary atresia, but may not be sufficiently discriminating. Magnetic resonance cholangiopancreatography (MRCP) is not optimal for visualization of the extrahepatic biliary tract in children younger than 3 months of age. Intraoperative cholangiography, whether laparoscopic or during open laparotomy, is a confirmatory procedure before actual performance of KPE portoenterostomy.

Pathogenesis

Biliary atresia is not a single disease with a defined etiology. An all-encompassing hypothesis is that a genetically susceptible individual undergoes inflammatory destruction of the extrahepatic biliary system in response to as yet undetermined environmental factors. Potential pathogenetic mechanisms include a genetic defect in morphogenesis with or without defective prenatal hepatic circulation, environmental triggers such as intrauterine or perinatal viral infection or toxin exposure, and immunological dysregulation in the perinatal period ( Table 55.15 ). These hypotheses have been put forth on the basis of epidemiological studies and molecular analyses of tissue specimens from human patients. Mouse models of biliary atresia have provided some additional clues.

Pathological Features

A diagnosis of biliary atresia is favored if a liver biopsy specimen exhibits bile ductular proliferation (neocholangioles), portal edema, and fibrosis but lacks sinusoidal fibrosis. Neutrophilic cholangitis and pericholangitis may or may not be present. The presence of bile plugs within bile duct lumina (which are distinct from the lumina of neocholangioles at the margins of portal tracts), portal edema, and increased numbers of bile duct profiles are helpful diagnostic findings with the largest odds ratio for predicting biliary atresia versus nonbiliary atresia ( Fig. 55.3 ). In fact, the extent of ductular reaction in wedge liver biopsies of children with biliary atresia, obtained at the time of portoenterostomy, is positively associated with improved 1-year survival of the native liver following KPE. Fifteen percent of biliary atresia cases show giant cell transformation of hepatocytes ( Fig. 55.4 ). The progressive fibrosis observed in biliary atresia may be abetted in part by epithelial-mesenchymal transition. ^,

The classic “obstructive” findings of bile duct proliferation and inspissated bile plugs are not always the result of atresia (see Box 55.8 ). Conversely, some cases of biliary atresia may show nonclassic features such as prominent portal tract arteries with medial hypertrophy, associated with disappearance of interlobular bile ducts and an absence of bile duct proliferation ( Fig. 55.5A ,B). ^, In keeping with the developmental theory of biliary atresia, a subset of affected patients exhibit ductal plate malformation (see Fig. 55.5C ). Recapitulating immature fetal portal tracts, this latter entity consists of portal tracts without an interlobular bile duct but with a centrally located hypertrophic hepatic artery and a peripheral rim of bile ductular structures. However, this phenomenon is not limited to the 20% to 25% of patients with a laterality defect (biliary atresia splenic malformation). As a result, biliary atresia can be difficult to diagnose at the time of percutaneous liver biopsy, even for the most experienced pediatric hepatopathologist (see Fig. 55.5D ). The age of the child at biopsy does not help resolve this difficulty. Nevertheless, the pathologist must advise the clinical team whether the histology of the liver biopsy justifies the patient being subjected to surgical intervention, with or without the performance of confirmatory intraoperative cholecystographic imaging before performance of the KPE. The alternative is that the pathologist advises the clinical team that the differential diagnosis of nonsurgical cholestatic disorders is of sufficient concern as to justify further noninvasive studies, rather than moving quickly to surgical intervention.

On performance of a KPE, the bile duct remnant is submitted for pathological examination. Typical findings in the remnant include fibrosis and obstruction of the lumen, a variable degree of periductal inflammation, and apoptotic degeneration of residual bile duct epithelium ( Fig. 55.6 ). These findings do not differ in syndromic cases of biliary atresia, cases discovered by prenatal ultrasound, and nonsyndromic postnatal biliary atresia, supporting the concept of a shared pathogenetic pathway. ^,

Natural History and Treatment

Before 1959, when the KPE procedure was introduced, biliary atresia was considered to be uniformly fatal by 2 years of age, with a median age at death of 10 months. The best survival rate in KPE patients with a native liver is 53% at 10 years, although reported native liver survival rates at 10 years are more commonly in the 35% range. ^, In the first report of the U.S. Biliary Atresia Research Consortium in 2006, a decline in total bilirubin at 3 months after KPE to less than 2 mg/dL was seen in 36.5% of children, and a favorable response (to <6 mg/dL) was seen in another 29%. Most patients in the excellent bilirubin response group had more favorable outcomes (as measured by “native liver survival”), a finding confirmed in more recent reports. ^, The 25% of children with a laterality defect fared more poorly, even if treated earlier. ^, Atresia of the common hepatic duct or ducts at the liver hilum (Ohi types 2 and 3) had a poorer prognosis than an atretic common bile duct (Ohi type 1), similar to the prognosis in biliary atresia patients with other malformations, ascites at surgery, or delay in surgery beyond day 75. Children operated on within the first 30 days had a 2-year 74% native liver survival rate. At King’s College Hospital in London, 56% of infants with EHBA cleared jaundice, and the 2-year survival rate in patients with their native liver was 65%. The 51 patients with biliary atresia splenic malformation or biliary cysts tended to be operated on earlier had poorer outcomes if the procedure was delayed. Nevertheless, the same group reported that even when surgery was performed on day 100 or even later, 45% of patients were alive with their native liver at 5 years, and 40% at 10 years, suggesting that patients may postpone transplantation by the KPE. In a report of native liver histology in 23 patients after a clinically successful KPE with an average follow-up of 4.2 years, resolution in histological cholestasis occurred in a majority of patients (83%), but fibrosis and bile duct proliferation persisted. In these “native liver survivors,” fibrosis had progressed to cirrhosis (stage 4 METAVIR) in 52% of patients, associated with the presence of portal hypertension.

Infants who do not respond to the KPE suffer from recurrent episodes of ascending cholangitis or sepsis caused by the immediate proximity of a bowel segment to the porta hepatis of the residual biliary tree. Regardless of the presence or absence of ascending cholangitis, the liver has a higher likelihood of progression cirrhosis with portal hypertension. Large bile lakes may form at the hilum, indicating a persistent effort at bile secretion by hepatocytes ( Fig. 55.7 ). The pathogenesis of progressive fibrosis in chronic cholestatic diseases, including biliary atresia, PFIC, and Alagille syndrome, has been linked to persistence of Hedgehog (Hh) signaling, which normally promotes embryonal duct differentiation and then shuts off as the liver matures. Immunohistochemical colocalization of the Hh transcription factor Gli2 and the mesenchymal markers vimentin and FSP1 in some of the reactive ductular epithelial cells in these diseases is evidence of reversal of mesenchymal-epithelial transition. Similar colocalization of FSP1 and cytokeratin 7 was shown in ductular epithelium, and this ductal cell derangement may lead to fibroplasia. Growth failure, malnutrition, deficiencies of lipid-soluble vitamins, and altered protein–energy and nutrient utilization are expected complications.

A failed KPE in a patient with biliary atresia is the most frequent indication for liver transplantation in infants and young children, accounting for 32.3% of all pediatric liver transplants in 2016. In such instances, prior performance of a KPE does not adversely affect outcomes of liver transplantation in children with biliary atresia, although patients subjected to laparoscopic KPE are less likely to have adhesions at the time of liver transplantation, when compared with those whose KPE was by open laparotomy. Patient survival after orthotopic transplantation for biliary atresia in the U.S. United Network for Organ Sharing from 1988 to 2003 was 85.8% at 10 years, with graft survival of 72.7%. The decision to transplant a patient with biliary atresia who has progressed to end-stage liver disease is critical; entry of a patient into the “intent-to-transplant” pathway is associated with excellent overall outcomes, with 97% survival at 5 years in one published series. Pathological examination of explanted livers for occult malignancy is crucial for all forms of chronic liver disease in children.

Clinical Features

Persistent unconjugated hyperbilirubinemia raises the possibility of an inherited defect in bilirubin conjugation or hemolysis. Diagnostic assessment and therapeutic intervention is then critically important to avoid kernicterus and permanent neurological damage. In contrast, predominantly conjugated hyperbilirubinemia in newborns should prompt a rigorous diagnostic investigation for biliary atresia. An approach to the workup of neonatal hepatitis was presented earlier (see Cholestatic Pattern and Table 55.8 ).

Pathological Features

The hallmark histological finding of many neonatal liver disorders is the formation of large multinucleated hepatocytes (giant cells), which are part of the broader spectrum of lobular disarray in neonatal hepatitis ( Fig. 55.9 ). Nuclear inclusions in hepatocytes may suggest DNA viruses ( Figs. 55.10 and 55.11 ); those in red cell precursors may suggest parvovirus B19. In some cases, most of the liver parenchyma is transformed into giant cells, and this is referred to as syncytial giant cell hepatitis. When this histological picture predominates, electron microscopy can be extremely helpful and even pathognomonic if NPD type C is the cause. Of 40 infants evaluated for neonatal giant cell hepatitis in a Denver Children’s Series, 27% had NPD type C. Splenomegaly was a useful clue.

In neonatal hepatitis, the relative proportion of inflammatory cells is not considered informative because ongoing apoptosis may lead to extensive hepatocyte destruction and accumulation of residual lymphocytes and macrophages. In some cases, parenchymal inflammation is minimal or absent. In addition, parenchymal neutrophils are an uncommon finding. Occasional islands of erythropoiesis are a normal finding. In all instances, the biopsy should be examined carefully for evidence of viral cytopathic change.

Portal tracts may be inconspicuous, whether normal or abnormal. In general, at least five to seven portal tracts are required to consider the tissue sample adequate for histological evaluation. Ten or more portal tracts are preferred. Bile duct hypoplasia was more common among infants with hypopituitarism, compared with other conditions, in the Johns Hopkins/University of Chicago series. No other biopsy finding was helpful in distinguishing possible etiologies in that series. Exclusion of biliary atresia or A1AT storage disorder requires verification that most, if not all, portal tracts contain a terminal bile duct, a companion hepatic artery, and a portal vein and that bile ductular proliferation, edema, neutrophilic inflammation, and fibrosis are absent. Although the portal tracts in patients with neonatal hepatitis may be expanded by a mixed inflammatory infiltrate, this should not be confused with normal granulocytic hematopoiesis occasionally found within portal tracts.

Occasionally, massive hepatic necrosis, with marked accumulation of parenchymal iron, may be observed in patients with Down syndrome. One infant in the Texas Children’s Hospital series had an exceptional degree of hepatocellular necrosis, and the cause was pyruvate kinase deficiency. The observation of massive necrosis should also prompt consideration of NH (discussed earlier; see Table 55.7 ).

Differential Diagnosis

Once biliary atresia and A1AT mutations have been excluded on the basis of both clinical findings and percutaneous liver biopsy, then the differential diagnosis of neonatal hepatitis syndrome must be pursued. Infections of the newborn have been mentioned, and Chapter 47 addresses some of these disorders. Extensive lobular disease, with or without giant cells, may be seen in patients with biliary atresia (see Fig. 55.4 ). Therefore lobular changes should not divert attention from the features in the portal tracts. Second, nonclassic features of biliary obstruction may be observed in biopsy specimens from patients with biliary atresia, in which ductular reaction has not yet occurred and hepatic arteries exhibit hypertrophy of the media smooth muscle (see Fig. 55.5A ,B). Sometimes, alert detection of acholic stools prompts an early biopsy, but the time of onset and rate of progression can vary greatly. (For a detailed discussion of the differential diagnosis, see Cholestatic Pattern and Table 55.7 .)

Natural History and Treatment

The natural history of neonatal hepatitis depends heavily on the specific etiology ( Table 55.16 ). For instance, pharmacological intervention is imperative for many of the infectious disorders. Many of the other disorders, including hypopituitarism, tyrosinemia, and bile salt synthetic defects, benefit dramatically from prompt intervention. In about a quarter of all infants, no cause is found. The prognosis for “idiopathic” neonatal hepatitis syndrome is considered favorable; the mortality rate is 13% to 25%. Predictors of a poor clinical outcome include severe or prolonged (>6 months) jaundice, acholic stools, familial occurrence, and persistent hepatomegaly. Peak bilirubin level is not predictive of outcome. Sepsis is a devastating complication and is associated with a poor outcome. For infants whose liver disease resolves, the prognosis is quite favorable; most have no residual liver dysfunction. Many children benefit from a cholestatic gene panel evaluation that may pinpoint the exact defect (see Box 55.1 ).

The management of idiopathic neonatal hepatitis syndrome is supportive and includes adequate nutrition. Dietary measures, such as use of lactose-free, low-protein formulas, may mitigate further liver damage until the results of tests for galactosemia, hereditary tyrosinemia type I, and hypopituitarism, for example, become known. Elemental formulas containing medium-chain triglycerides help maintain caloric intake in severely ill patients. Infants with chronic cholestasis require fat-soluble vitamin supplementation. Pruritus associated with chronic cholestasis is difficult to treat in many patients.

Clinical Features

Some infants and young children with conjugated hyperbilirubinemia have associated facial dysmorphism and cardiovascular, vertebral, and ocular malformations. The combination of extrahepatic abnormalities and deficiency of interlobular bile ducts is termed Alagille syndrome . Alagille syndrome is an autosomal dominant disorder with an estimated frequency of 1 in 70,000 live births, increased to an incidence of 1 in 30,000 live births in the molecular era. Sporadic cases account for 45% to 50%. The mortality rate is 15% to 20%. Deaths are mostly caused by hepatic or cardiovascular complications.

Before the advent of genetic testing, a diagnosis of Alagille syndrome required demonstration of paucity of interlobular bile ducts and clinical evidence of at least three major of the following abnormalities: characteristic facies, posterior embryotoxon, butterfly vertebrae, renal disease, and cardiac anomalies. However, bile duct paucity is evident in only 60% of infants who undergo biopsy before 6 months of age. Furthermore, there is wide phenotypic variability in patients with this disorder, ranging from mild subclinical findings to complete liver failure and complex congenital heart disease. ^, Most patients with hepatic involvement are seen clinically within the first 6 months with jaundice, hepatomegaly, pruritus, and failure to thrive. Laboratory studies reveal conjugated hyperbilirubinemia and increased serum levels of GGT, alkaline phosphatase, bile acids, and cholesterol. Serum ALT and AST levels may be normal or mildly elevated. Currently, genetic testing is almost always conclusive and is included as part of the cholestatic gene panel.

Pathogenesis

Mutations in Jagged1 (JAG1), a ligand in the Notch signaling pathway, have been identified in 94% of patients with Alagille syndrome. Mutations in the NOTCH2 gene, which encodes a receptor for Jagged1, have been identified in some JAG1 mutation–negative patients. ^, It is hypothesized that defects in JAG1 result in an arrest of branching and elongation of bile ducts during postnatal liver growth. In support of this theory, Libbrecht and associates reported a case in which an explanted liver from a 16-year-old patient with Alagille syndrome demonstrated paucity of bile ducts in peripheral liver parenchyma but normally developed bile ducts in the perihilar areas. Deletions in a single JAG1 allele are sufficient to cause ALGS, suggesting haploinsufficiency as the disease-causing mechanism. Recent mouse models with mutant JAG1 that have been developed by Andersson et al. and Adams et al. will help better define the mechanism of ALGS as it showed binding of NOTCH2 rather than NOTCH1 by JAG 1 ^Ndr in these mice. The interaction of the NOTCH and WNT pathways in biliary epithelial development is of great interest for aberrant organogenesis and response to injury.

Pathological Features

To diagnose paucity of bile ducts, the liver biopsy specimen should contain at least 7 portal tracts, and preferably at least 10. Normally, on routinely stained tissue sections (H&E or Masson trichrome) of mature liver tissue, approximately 90% of portal tracts contain an interlobular bile duct, either as a single duct or as multiple duct profiles, paired in close apposition to branches of the hepatic artery of similar outside diameter. The presence of multiple portal tracts containing arteries and veins but not bile ducts should prompt consideration of bile duct paucity, which is defined by an absence of, or marked reduction in, the number of interlobular bile ducts within portal tracts. In pediatric patients, a bile duct-to-portal tract ratio of less than 0.8 is often used as a cutoff point between normal and abnormal, although the number of bile ducts may be normally (physiologically) low in preterm infants. However, in any age group, a ratio of less than 0.4 is strongly suggestive of bile duct paucity. When making this assessment, large portal tracts should not be considered in the equation. Furthermore, incomplete portal tracts that are partially transected by the biopsy should be excluded from the analysis because absence of a bile duct may simply represent a sampling artifact. Specimens from patients with Alagille syndrome are conspicuous for their lack of portal tracts and parenchymal inflammation, but they may have abundant giant cells, as in neonatal hepatitis ( Fig. 55.14 ).

Immunohistochemical staining for cytokeratin 7 or 19 is useful to highlight bile duct epithelium. Because hepatocytes in cholestatic livers often acquire cytokeratin 7 immunoreactivity (see Fig. 55.14D ), cytokeratin 19 is preferred for quantitating bile ducts. Immunoreactive ductular profiles at the margins of portal tracts should not be confused with true interlobular bile ducts (see Fig. 55.1 ). A persistent lack of canalicular CD10 immunopositivity, although physiological in infants younger than 2 years of age, is a characteristic finding in Alagille syndrome. The Masson trichrome stain is also helpful in identifying small portal tracts because of its ability to detect delicate amounts of connective tissue. These small portal tracts should be included in the portal tract count when one is assessing bile ducts.

In the evaluation of a liver biopsy specimen for potential paucity of bile ducts, the pathology report should indicate how many portal tracts and bile ducts were identified. The staining method should also be reported. Paucity of interlobular bile ducts is not a specific disorder; instead, it represents a pathological feature that may have a variety of causes.

Differential Diagnosis

When Alagille syndrome cannot be distinguished from biliary obstruction on liver biopsy, and the clinical and radiological features fail to detect the extrahepatic manifestations in an infant with Alagille syndrome, patients have at times still been treated by KPE. Even at a major center with extensive experience with these diseases, 4.4% of patients were mistakenly operated on, and afterward their liver disease progressed more rapidly to end-stage cirrhosis. Alagille histology may show an overlap with EHBA with marked ductular proliferation and fibrosis but shows absence of interlobular bile ducts. Because Alagille disease progression is highly variable, every effort must be made to avoid surgery.

Caution is also required in the interpretation of hepatobiliary scintigraphy and cholangiographic studies. Typically, both extrahepatic and intrahepatic bile ducts in patients with Alagille syndrome are markedly hypoplastic; therefore on hepatobiliary scanning, one may not see excretion of radioisotope into the intestines (mimicking biliary atresia). Similarly, intraoperative cholangiography may not necessarily demonstrate opacification of the proximal extrahepatic ducts. These diagnostic pitfalls underscore the importance of a careful and full physical examination and radiological evaluation in patients with suspected Alagille syndrome.

No genotype-phenotype correlation with severity for JAGGED1 mutations was found among 33 patients with Alagille syndrome who were older than 10 years of age. Those children with elevated total bilirubin (>6.5 mg/dL), conjugated bilirubin (>4.5 mg/dL), and cholesterol (>520 mg/dL) before 5 years of age had significantly worse liver disease later. Sequencing of the entire coding region of JAG1 is now available and should be used in patients in whom there is a strong clinical suspicion and in family members of probands with mutations.

Natural History

Alagille syndrome is a benign illness in many children. However, young children with protracted severe jaundice usually have a poorer prognosis. Between 10% and 50% of patients with Alagille syndrome eventually progress to cirrhosis and liver failure. Cases of HCC in Alagille syndrome have been described, as in every chronic cholestatic disease of infants. The overall mortality rate from Alagille syndrome is estimated to be 20% to 25%; death occurs mainly from cardiac disease, intercurrent infection, or progressive liver disease. Liver transplantation is warranted for patients with severe hepatic disease. Because of the presence of multiple organ defects in Alagille syndrome, the outcomes of liver transplantation are poorer than for patients with EHBA, especially with regard to death during the first 30 postoperative days. No pretransplant factors were identified among the 87% of patients who survived to 1 year, compared with those who did not.

Classification and Pathogenesis

The hepatic conjugating enzyme UGT1A1 is a product of the UGT1A1 gene, which is located on chromosome 2q37. It is a member of a family of UDP-glucuronosyltransferases (UGTs) that catalyze the glucuronidation of an array of substrates such as steroid hormones, carcinogens, and drugs. The various types of UGTs are distributed within a wide range of tissues, including liver, kidney, intestines, skin, lung, olfactory epithelium, and testis. UGT1A1 is located primarily within the smooth and rough endoplasmic reticulum of hepatocytes as a single isoform that catalyzes the glucuronidation of bilirubin to form its monoglucuronidated and diglucuronidated forms. In humans, two members of the UGT1 family possess the capability to glucuronidate bilirubin in vitro, but only one isoform is physiologically relevant in vivo. The bilirubin glucuronidating isoform is termed UGT1A1 because it is generated from the exon 1A of the UGT1 gene locus. Multiple mutations in UGT1A1 cause hereditary unconjugated hyperbilirubinemia: Crigler-Najjar syndrome types I and II and Gilbert syndrome.

Clinical and Pathological Features

In Crigler-Najjar syndrome type I, the liver UGT1A1 enzyme is completely absent. Colorless bile contains only trace amounts of unconjugated bilirubin. Serum unconjugated bilirubin reaches very high levels, leading to severe jaundice and icterus. Without liver transplantation, this condition is invariably fatal (because of kernicterus), usually within 18 months of birth. In a recent review of 22 patients undergoing liver transplant for Crigler-Najjar syndrome type I, the pathological features showed canalicular cholestasis and significant pericentral sinusoidal, periportal, and mixed patterns of fibrosis in almost 41% of the patients. The fibrosis appeared to be progressive and more in older children at the time of transplant. This suggests the need for early recognition of this disease and intervention to arrest progression of fibrosis and need for earlier liver transplantation. ^, This has led to attempts to use AAV8 gene therapy to ameliorate the changes with preliminary success in mouse models.

Crigler-Najjar syndrome type II is a less severe, nonfatal disorder in which the hepatic level of UGT1A1 enzyme activity is greatly reduced but not absent, and the enzyme is capable of forming only monoglucuronidated bilirubin. In contrast with the type I syndrome, the only major clinical consequence is the presence of extraordinarily yellow skin caused by moderate to high levels of circulating unconjugated bilirubin. Phenobarbital treatment may improve bilirubin glucuronidation by inducing hypertrophy of the hepatocellular endoplasmic reticulum.

Gilbert syndrome is a relatively common benign, inherited condition. Affected patients have mild, fluctuating hyperbilirubinemia in the absence of hemolysis or liver disease. Typically, hepatic bilirubin glucuronidating activity is approximately 30% of normal levels. In most patients, the genetic defect consists of two extra bases (TA) in the TATAA element (TATA box) in the promoter region of the UGT1A1 gene. This creates an A(TA) ₇ TAA element, rather than the normal A(TA) ₆ TAA of the UGT1A1 gene (“an extra TA in the TATA box”), and results in reduced expression of UGT1A1. In some cases, patients are heterozygous for missense mutations in the UGT1A1 gene. Gilbert syndrome affects 3% to 10% of the population. Mild hyperbilirubinemia may go unrecognized for many years and is not associated with functional derangements of the liver. When detected in adolescents or adults, it is typically in association with an unrelated stress, such as intercurrent illness, strenuous exercise, or fasting, that reduces hepatic levels of the obligate cofactor for UGT1A1, UDP-glucuronic acid. Gilbert syndrome has no clinical consequence except for anxiety related to persistent jaundice. A 2007 study suggested that diabetic patients with coexistent Gilbert syndrome have a lower incidence of vascular complications and a reduction in serum markers of oxidase stress and inflammation, possibly because of the antioxidant effect of bilirubin.

In all disorders of bilirubin conjugation other than Crigler-Najjar type I, the liver is morphologically normal by both light and electron microscopy.