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InternalMed_Harrison. TATA Box stat administration for 4–6 weeks to 16 patients with Crigler-Najjar syndrome FIGURE 359-2 Structural organization of the human UGT1 gene complex. This large complex was associated with a 10–20% decrease on chromosome 2 contains at least 13 substrate-specific first exons (A1, A2, etc.). Since four of these in serum bilirubin in 7 patients, the cost are pseudogenes, nine UGT1 isoforms with differing substrate specificities are expressed. Each exon and side effects (i.e., diarrhea) may obvi 1 has its own promoter and encodes the amino-terminal substrate-specific ∼286 amino acids of the ate the small benefit achievable with this various UGT1-encoded isoforms, and common exons 2–5 that encode the 245 carboxyl-terminal treatment.


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InternalMed_Harrison. amino acids common to all of the isoforms. mRNAs for specific isoforms are assembled by splicing a particular first exon such as the bilirubin-specific exon A1 to exons 2 to 5. The resulting message Renal Excretion of Bilirubin Conjugates encodes a complete enzyme, in this particular case bilirubin-UDP-glucuronosyltransferase (UGT1A1). Unconjugated bilirubin is not excreted Mutations in a first exon affect only a single isoform. Those in exons 2–5 affect all enzymes encoded in urine, as it is too tightly bound to by the UGT1 complex. albumin for effective glomerular filtra tion and there is no tubular mechanism for its renal secretion. In contrast, the bilirubin conjugates are readily filtered other substrates have been designated the UGT1 family. These at the glomerulus and can appear in urine in disorders characterizedare expressed from a single gene complex by alternative promoter by increased bilirubin conjugates in the circulation.usage. This gene complex contains multiple


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InternalMed_Harrison. in urine in disorders characterizedare expressed from a single gene complex by alternative promoter by increased bilirubin conjugates in the circulation.usage. This gene complex contains multiple substrate-specific first exons, designated A1, A2, etc. (Fig. 359-2), each with its own DISORDERS OF BILIRUBIN METABOLISM LEADING promoter and each encoding the amino-terminal half of a specific TO UNCONJUGATED HYPERBILIRUBINEMIA isoform. In addition, there are four common exons (exons 2–5) that encode the shared carboxyl-terminal half of all of the UGT1 isoforms. The various first exons encode the specific aglycone Hemolysis Increased destruction of erythrocytes leads to increased substrate binding sites for each isoform, while the shared exons bilirubin turnover and unconjugated hyperbilirubinemia; the hyper-encode the binding site for the sugar donor, UDP-glucuronic bilirubinemia is usually modest in the presence of normal liver funcacid, and the transmembrane domain. Exon A1 and the four


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InternalMed_Harrison. the hyper-encode the binding site for the sugar donor, UDP-glucuronic bilirubinemia is usually modest in the presence of normal liver funcacid, and the transmembrane domain. Exon A1 and the four tion. In particular, the bone marrow is only capable of a sustained common exons, collectively designated the UGT1A1 gene (Fig. eightfold increase in erythrocyte production in response to a hemo359-2), encode the physiologically critical enzyme bilirubin-UDP-lytic stress. Therefore, hemolysis alone cannot result in a sustained glucuronosyltransferase (UGT1A1). A functional corollary of the hyperbilirubinemia of more than ∼68 μmol/L (4 mg/dL). Higher values organization of the UGT1 gene is that a mutation in one of the imply concomitant hepatic dysfunction. When hemolysis is the only first exons will affect only a single enzyme isoform. By contrast, a abnormality in an otherwise healthy individual, the result is a purely mutation in exons 2–5 will alter all isoforms encoded by the UGT1


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InternalMed_Harrison. exons will affect only a single enzyme isoform. By contrast, a abnormality in an otherwise healthy individual, the result is a purely mutation in exons 2–5 will alter all isoforms encoded by the UGT1 unconjugated hyperbilirubinemia, with the direct-reacting fraction as gene complex. measured in a typical clinical laboratory being ≤15% of the total serum 4. Biliary excretion: It has been thought until recently that biliru-bilirubin. In the presence of systemic disease, which may include a bin monoand diglucuronides are excreted directly across the degree of hepatic dysfunction, hemolysis may produce a component canalicular plasma membrane into the bile canaliculus by an ATP-of conjugated hyperbilirubinemia in addition to an elevated uncondependent transport process mediated by a canalicular membrane jugated bilirubin concentration. Prolonged hemolysis may lead to the protein called multidrug resistance–associated protein 2 (MRP2). precipitation of bilirubin salts within the gallbladder


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InternalMed_Harrison. membrane jugated bilirubin concentration. Prolonged hemolysis may lead to the protein called multidrug resistance–associated protein 2 (MRP2). precipitation of bilirubin salts within the gallbladder or biliary tree, Mutations of MRP2 result in the Dubin-Johnson syndrome (see resulting in the formation of gallstones in which bilirubin, rather than below). However, studies in patients with Rotor syndrome (see cholesterol, is the major component. Such pigment stones may lead to below) indicate that after formation, a portion of the glucuro-acute or chronic cholecystitis, biliary obstruction, or any other biliary nides are transported into the portal circulation by a sinusoidal tract consequence of calculous disease. membrane protein called multidrug resistance–associated protein


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InternalMed_Harrison. Ineffective Erythropoiesis During erythroid maturation, small amounts 3 (MRP3) and subjected to reuptake into the hepatocyte by the of hemoglobin may be lost at the time of nuclear extrusion, and a fracsinusoidal membrane uptake transporters organic anion transport tion of developing erythroid cells is destroyed within the marrow. protein 1B1 (OATP1B1) and OATP1B3.


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InternalMed_Harrison. protein 1B1 (OATP1B1) and OATP1B3. These processes normally account for a small proportion of bilirubin that is produced. In various disorders, including thalassemia major, megaloblastic anemias due to folate or vitamin B12 deficiency, congeni- tal erythropoietic porphyria, lead poisoning, and various congenitalBilirubin in the Gut Following secretion into bile, conjugated bilirubin and acquired dyserythropoietic anemias, the fraction of total bilirubinreaches the duodenum and passes down the gastrointestinal tract with-production derived from ineffective erythropoiesis is increased, reach-out reabsorption by the intestinal mucosa. An appreciable fraction is ing as much as 70% of the total. This may be sufficient to produceconverted by bacterial metabolism in the gut to the water-soluble col-modest degrees of unconjugated hyperbilirubinemia.


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InternalMed_Harrison. orless compound urobilinogen. Urobilinogen undergoes enterohepatic cycling. Urobilinogen not taken up by the liver reaches the systemic Miscellaneous Degradation of the hemoglobin of extravascular colleccirculation, from which some is cleared by the kidneys. Unconjugated tions of erythrocytes, such as those seen in massive tissue infarctions bilirubin ordinarily does not reach the gut except in neonates or, by ill-or large hematomas, may lead transiently to unconjugated hyperbilidefined alternative pathways, in the presence of severe unconjugated rubinemia.


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InternalMed_Harrison. DECREASED HEPATIC BILIRUBIN CLEARANCE Decreased Hepatic Uptake Decreased hepatic bilirubin uptake is believed to contribute to the unconjugated hyperbilirubinemia of Gilbert syndrome (GS), although the molecular basis for this finding remains unclear (see below). Several drugs, including flavaspidic acid, novobiocin, and rifampin, as well as various cholecystographic contrast agents, have been reported to inhibit bilirubin uptake. The resulting unconjugated hyperbilirubinemia resolves with cessation of the medication.


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InternalMed_Harrison. Impaired Conjugation • pHysioloGic neonatal JaunDice Bilirubin produced by the fetus is cleared by the placenta and eliminated by the maternal liver. Immediately after birth, the neonatal liver must assume responsibility for bilirubin clearance and excretion. However, many hepatic physiologic processes are incompletely developed at birth. Levels of UGT1A1 are low, and alternative excretory pathways allow passage of unconjugated bilirubin into the gut. Since the intestinal flora that convert bilirubin to urobilinogen are also undeveloped, an enterohepatic circulation of unconjugated bilirubin ensues. As a consequence, most neonates develop mild unconjugated hyperbilirubinemia between days 2 and 5 after birth. Peak levels are typically <85–170 μmol/L (5–10 mg/dL) and decline to normal adult concentrations within 2 weeks, as mechanisms required for bilirubin disposition mature. Prematurity, often associated with more profound immaturity of hepatic function and hemolysis, can result in


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InternalMed_Harrison. concentrations within 2 weeks, as mechanisms required for bilirubin disposition mature. Prematurity, often associated with more profound immaturity of hepatic function and hemolysis, can result in higher levels of unconjugated hyperbilirubinemia. A rapidly rising unconjugated bilirubin concentration, or absolute levels >340 μmol/L (20 mg/dL), puts the infant at risk for bilirubin encephalopathy, or kernicterus. Under these circumstances, bilirubin crosses an immature blood-brain barrier and precipitates in the basal ganglia and other areas of the brain. The consequences range from appreciable neurologic deficits to death. Treatment options include phototherapy, which converts bilirubin into water-soluble photoisomers that are excreted directly into bile, and exchange transfusion. The canalicular mechanisms responsible for bilirubin excretion are also immature at birth, and their maturation may lag behind that of UGT1A1; this can lead to transient conjugated neonatal


InternalMed_Harrison

InternalMed_Harrison. The canalicular mechanisms responsible for bilirubin excretion are also immature at birth, and their maturation may lag behind that of UGT1A1; this can lead to transient conjugated neonatal hyperbilirubinemia, especially in infants with hemolysis.


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InternalMed_Harrison. acquireD conJuGation Defects A modest reduction in bilirubin conjugating capacity may be observed in advanced hepatitis or cirrhosis. However, in this setting, conjugation is better preserved than other aspects of bilirubin disposition, such as canalicular excretion. Various drugs, including pregnanediol, novobiocin, chloramphenicol, and gentamicin, may produce unconjugated hyperbilirubinemia by inhibiting UGT1A1 activity. Bilirubin conjugation may be inhibited by certain fatty acids that are present in breast milk but not serum of mothers whose infants have excessive neonatal hyperbilirubinemia (breast milk jaundice). Alternatively, there may be increased entero-2001 hepatic circulation of bilirubin in these infants. A recent study has correlated epidermal growth factor (EGF) content of breast milk with elevated bilirubin levels in these infants; however, a cause-and-effect relationship remains to be established. The pathogenesis of breast milk jaundice appears to differ from that of


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InternalMed_Harrison. milk with elevated bilirubin levels in these infants; however, a cause-and-effect relationship remains to be established. The pathogenesis of breast milk jaundice appears to differ from that of transient familial neonatal hyperbilirubinemia (Lucey-Driscoll syndrome), in which there is a UGT1A1 inhibitor in maternal serum.


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InternalMed_Harrison. Three familial disorders characterized by differing degrees of unconjugated hyperbilirubinemia have long been recognized. The defining clinical features of each are described below (Table 359-1). While these disorders have been recognized for decades to reflect differing degrees of deficiency in the ability to conjugate bilirubin, recent advances in the molecular biology of the UGT1 gene complex have elucidated their interrelationships and clarified previously puzzling features.


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InternalMed_Harrison. Crigler-Najjar Syndrome, Type I CN-I is characterized by striking uncon jugated hyperbilirubinemia of about 340–765 μmol/L (20–45 mg/dL) that appears in the neonatal period and persists for life. Other conventional hepatic biochemical tests such as serum aminotransferases and alkaline phosphatase are normal, and there is no evidence of hemolysis. Hepatic histology is also essentially normal except for the occasional presence of bile plugs within canaliculi. Bilirubin glucuronides are virtually absent from the bile, and there is no detectable constitutive expression of UGT1A1 activity in hepatic tissue. Neither UGT1A1 activity nor the serum bilirubin concentration responds to administration of phenobarbital or other enzyme inducers. In the absence of conjugation, unconjugated bilirubin accumulates in plasma, from which it is eliminated very slowly by alternative pathways that include direct passage into the bile and small intestine. These account for the small amounts of urobilinogen


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InternalMed_Harrison. in plasma, from which it is eliminated very slowly by alternative pathways that include direct passage into the bile and small intestine. These account for the small amounts of urobilinogen found in feces. No bilirubin is found in the urine. First described in 1952, the disorder is rare (estimated prevalence, 0.6–1.0 per million). Many patients are from geographically or socially isolated communities in which consanguinity is common, and pedigree analyses show an autosomal recessive pattern of inheritance. The majority of patients (type IA) exhibit defects in the glucuronide conjugation of a spectrum of substrates in addition to bilirubin, including various drugs and other xenobiotics. These individuals have mutations in one of the common exons (2–5) of the UGT1 gene (Fig. 359-2). In a smaller subset (type IB), the defect is limited largely to bilirubin conjugation, and the causative mutation is in the bilirubin-specific exon A1. Estrogen glucuronidation is mediated by UGT1A1 and is


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InternalMed_Harrison. subset (type IB), the defect is limited largely to bilirubin conjugation, and the causative mutation is in the bilirubin-specific exon A1. Estrogen glucuronidation is mediated by UGT1A1 and is defective in all CN-I patients. More than 30 different genetic lesions of UGT1A1 responsible for CN-I have been identified,


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InternalMed_Harrison. The Hyperbilirubinemias 2002 including deletions, insertions, alterations in intron splice donor and acceptor sites, exon skipping, and point mutations that introduce premature stop codons or alter critical amino acids. Their common feature is that they all encode proteins with absent or, at most, traces of bilirubin-UDP-glucuronosyltransferase enzymatic activity. Prior to the availability of phototherapy, most patients with CN-I died of bilirubin encephalopathy (kernicterus) in infancy or early childhood. A few lived as long as early adult life without overt neurologic damage, although more subtle testing usually indicated mild but progressive brain damage. In the absence of liver transplantation, death eventually supervened from late-onset bilirubin encephalopathy, which often followed a nonspecific febrile illness. Although isolated hepatocyte transplantation has been used in a small number of cases of CN-I, early liver transplantation (Chap. 368) remains the best hope to prevent


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InternalMed_Harrison. a nonspecific febrile illness. Although isolated hepatocyte transplantation has been used in a small number of cases of CN-I, early liver transplantation (Chap. 368) remains the best hope to prevent brain injury and death.


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InternalMed_Harrison. Crigler-Najjar Syndrome, Type II (CN-II) This condition was recognized as a distinct entity in 1962 and is characterized by marked unconjugated hyperbilirubinemia in the absence of abnormalities of other conventional hepatic biochemical tests, hepatic histology, or hemolysis. It differs from CN-I in several specific ways (Table 359-1): (1) Although there is considerable overlap, average bilirubin concentrations are lower in CN-II; (2) accordingly, CN-II is only infrequently associated with kernicterus; (3) bile is deeply colored, and bilirubin glucuronides are present, with a striking, characteristic increase in the proportion of monoglucuronides; (4) UGT1A1 in liver is usually present at reduced levels (typically ≤10% of normal) but may be undetectable by older, less sensitive assays; and (5) while typically detected in infancy, hyperbilirubinemia was not recognized in some cases until later in life and, in one instance, at age 34. As with CN-I, most CN-II cases exhibit abnormalities


InternalMed_Harrison

InternalMed_Harrison. (5) while typically detected in infancy, hyperbilirubinemia was not recognized in some cases until later in life and, in one instance, at age 34. As with CN-I, most CN-II cases exhibit abnormalities in the conjugation of other compounds, such as salicylamide and menthol, but in some instances, the defect appears limited to bilirubin. Reduction of serum bilirubin concentrations by >25% in response to enzyme inducers such as phenobarbital distinguishes CN-II from CN-I, although this response may not be elicited in early infancy and often is not accompanied by measurable UGT1A1 induction. Bilirubin concentrations during phenobarbital administration do not return to normal but are typically in the range of 51–86 μmol/L (3–5 mg/dL). Although the incidence of kernicterus in CN-II is low, instances have occurred, not only in infants but also in adolescents and adults, often in the setting of an intercurrent illness, fasting, or another factor that temporarily raises the serum bilirubin


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InternalMed_Harrison. have occurred, not only in infants but also in adolescents and adults, often in the setting of an intercurrent illness, fasting, or another factor that temporarily raises the serum bilirubin concentration above baseline and reduces serum albumin levels. For this reason, phenobarbital therapy is widely recommended, a single bedtime dose often sufficing to maintain clinically safe serum bilirubin concentrations.


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InternalMed_Harrison. Over 77 different mutations in the UGT1 gene have been identified as causing CN-I or CN-II. It was found that missense mutations are more common in CN-II patients, as would be expected in this less severe phenotype. Their common feature is that they encode for a bilirubin-UDP-glucuronosyltransferase with markedly reduced, but detectable, enzymatic activity. The spectrum of residual enzyme activity explains the spectrum of phenotypic severity of the resulting hyperbilirubinemia. Molecular analysis has established that a large majority of CN-II patients are either homozygotes or compound heterozygotes for CN-II mutations and that individuals carrying one mutated and one entirely normal allele have normal bilirubin concentrations.


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InternalMed_Harrison. Gilbert Syndrome (GS) This syndrome is characterized by mild unconjugated hyperbilirubinemia, normal values for standard hepatic biochemical tests, and normal hepatic histology other than a modest increase of lipofuscin pigment in some patients. Serum bilirubin concentrations are most often <51 μmol/L (<3 mg/dL), although both higher and lower values are frequent. The clinical spectrum of hyperbilirubinemia fades into that of CN-II at serum bilirubin concentrations of 86–136 μmol/L (5–8 mg/dL). At the other end of the scale, the distinction between mild cases of GS and a normal state is often blurred. Bilirubin concentrations may fluctuate substantially in any given individual, and at least 25% of patients will exhibit temporarily normal values during prolonged follow-up. More elevated values are associated with stress, fatigue, alcohol use, reduced caloric intake, and intercurrent illness, while increased caloric intake or administration of enzyme-inducing agents produces lower


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InternalMed_Harrison. values are associated with stress, fatigue, alcohol use, reduced caloric intake, and intercurrent illness, while increased caloric intake or administration of enzyme-inducing agents produces lower bilirubin levels. GS is most often diagnosed at or shortly after puberty or in adult life during routine examinations that include multichannel biochemical analyses. UGT1A1 activity is typically reduced to 10–35% of normal, and bile pigments exhibit a characteristic increase in bilirubin monoglucuronides. Studies of radiobilirubin kinetics indicate that hepatic bilirubin clearance is reduced to an average of one-third of normal. Administration of phenobarbital normalizes both the serum bilirubin concentration and hepatic bilirubin clearance; however, failure of UGT1A1 activity to improve in many such instances suggests the possible coexistence of an additional defect. Compartmental analysis of bilirubin kinetic data suggests that GS patients have a defect in bilirubin uptake as well as in


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InternalMed_Harrison. such instances suggests the possible coexistence of an additional defect. Compartmental analysis of bilirubin kinetic data suggests that GS patients have a defect in bilirubin uptake as well as in conjugation. Defect(s) in the hepatic uptake of other organic anions that at least partially share an uptake mechanism with bilirubin, such as sulfobromophthalein and indocyanine green (ICG), are observed in a minority of patients. The metabolism and transport of bile acids that do not utilize the bilirubin uptake mechanism are normal. The magnitude of changes in the serum bilirubin concentration induced by provocation tests such as 48 hours of fasting or the IV administration of nicotinic acid have been reported to be of help in separating GS patients from normal individuals. Other studies dispute this assertion. Moreover, on theoretical grounds, the results of such studies should provide no more information than simple measurements of the baseline serum bilirubin concentration. Family


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InternalMed_Harrison. dispute this assertion. Moreover, on theoretical grounds, the results of such studies should provide no more information than simple measurements of the baseline serum bilirubin concentration. Family studies indicate that GS and hereditary hemolytic anemias such as hereditary spherocytosis, glucose-6-phosphate dehydrogenase deficiency, and β-thalassemia trait sort independently. Reports of hemolysis in up to 50% of GS patients are believed to reflect better case finding, since patients with both GS and hemolysis have higher bilirubin concentrations, and are more likely to be jaundiced, than patients with either defect alone.


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InternalMed_Harrison. GS is common, with many series placing its prevalence at ≥8%. Males predominate over females by reported ratios ranging from 1.5:1 to >7:1. However, these ratios may have a large artifactual component since normal males have higher mean bilirubin levels than normal females, but the diagnosis of GS is often based on comparison to normal ranges established in men. The high prevalence of GS in the general population may explain the reported frequency of mild unconjugated hyperbilirubinemia in liver transplant recipients. The disposition of most xenobiotics metabolized by glucuronidation appears to be normal in GS, as is oxidative drug metabolism in the majority of reported studies. The principal exception is the metabolism of the anti-tumor agent irinotecan (CPT-11), whose active metabolite (SN-38) is glucuronidated specifically by bilirubin-UDP-glucuronosyltransferase. Administration of CPT-11 to patients with GS has resulted in several toxicities, including intractable diarrhea and


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InternalMed_Harrison. (SN-38) is glucuronidated specifically by bilirubin-UDP-glucuronosyltransferase. Administration of CPT-11 to patients with GS has resulted in several toxicities, including intractable diarrhea and myelosuppression. Some reports also suggest abnormal disposition of menthol, estradiol benzoate, acetaminophen, tolbutamide, and rifamycin SV. Although some of these studies have been disputed, and there have been no reports of clinical complications from use of these agents in GS, prudence should be exercised in prescribing them, or any agents metabolized primarily by glucuronidation, in this condition. It should also be noted that the HIV protease inhibitors indinavir and atazanavir (Chap. 226) can inhibit UGT1A1, resulting in hyperbilirubinemia that is most pronounced in patients with preexisting GS.


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InternalMed_Harrison. Most older pedigree studies of GS were consistent with autosomal dominant inheritance with variable expressivity. However, studies of the UGT1 gene in GS have indicated a variety of molecular genetic bases for the phenotypic picture and several different patterns of inheritance. Studies in Europe and the United States found that nearly all patients had normal coding regions for UGT1A1 but were homozygous for the insertion of an extra TA (i.e., A[TA]7TAA rather than A[TA]6TAA) in the promoter region of the first exon. This appeared to be necessary, but not sufficient, for clinically expressed GS, since 15% of normal controls were also homozygous for this variant. While normal by standard criteria, these individuals had somewhat higher bilirubin concentrations than the rest of the controls studied. Heterozygotes for this abnormality had bilirubin concentrations identical to those homozygous for the normal A[TA]6TAA allele. The prevalence of the A[TA]7TAA allele in a general Western


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InternalMed_Harrison. studied. Heterozygotes for this abnormality had bilirubin concentrations identical to those homozygous for the normal A[TA]6TAA allele. The prevalence of the A[TA]7TAA allele in a general Western population is 30%, in which case 9% would be homozygotes. This is slightly higher than the prevalence of GS based on purely phenotypic parameters. It was suggested that additional variables, such as mild hemolysis or a defect in bilirubin uptake, might be among the factors enhancing phenotypic expression of the defect.


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InternalMed_Harrison. Phenotypic expression of GS due solely to the A[TA]7TAA promoter abnormality is inherited as an autosomal recessive trait. A number of CN-II kindreds have been identified in whom there is also an allele containing a normal coding region but the A[TA]7TAA promoter abnormality. CN-II heterozygotes who have the A[TA]6TAA promoter are phenotypically normal, whereas those with the A[TA]7TAA promoter express the phenotypic picture of GS. GS in such kindreds may also result from homozygosity for the A[TA]7TAA promoter abnormality. Seven different missense mutations in the UGT1 gene that reportedly cause GS with dominant inheritance have been found in Japanese individuals. Another Japanese patient with mild unconjugated hyperbilirubinemia was homozygous for a missense mutation in exon 5. GS in her family appeared to be recessive. Missense mutations causing GS have not been reported outside of certain Asian populations.


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InternalMed_Harrison. In hyperbilirubinemia due to acquired liver disease (e.g., acute hepatitis, common bile duct stone), there are usually elevations in the serum concentrations of both conjugated and unconjugated bilirubin. Although biliary tract obstruction or hepatocellular cholestatic injury may present on occasion with a predominantly conjugated hyperbilirubinemia, it is generally not possible to differentiate intrahepatic from extrahepatic causes of jaundice based on the serum levels or relative proportions of unconjugated and conjugated bilirubin. The major reason for determining the amounts of conjugated and unconjugated bilirubin in the serum is for the initial differentiation of hepatic parenchymal and obstructive disorders (mixed conjugated and unconjugated hyperbilirubinemia) from the inheritable and hemolytic disorders discussed above that are associated with unconjugated hyperbilirubinemia.


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InternalMed_Harrison. FAMILIAL DEFECTS IN HEPATIC EXCRETORY FUNCTION Dubin-Johnson Syndrome (DJS) This benign, relatively rare disorder is characterized by low-grade, predominantly conjugated hyperbilirubinemia (Table 359-2). Total bilirubin concentrations are typically between 34 and 85 μmol/L (2 and 5 mg/dL) but on occasion can be in the normal range or as high as 340–430 μmol/L (20–25 mg/dL) and can fluctuate widely in any given patient. The degree of hyperbiliru-2003 binemia may be increased by intercurrent illness, oral contraceptive use, and pregnancy. Because the hyperbilirubinemia is due to a predominant rise in conjugated bilirubin, bilirubinuria is characteristically present. Aside from elevated serum bilirubin levels, other routine laboratory tests are normal. Physical examination is usually normal except for jaundice, although an occasional patient may have hepatosplenomegaly.


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InternalMed_Harrison. Patients with DJS are usually asymptomatic, although some may have vague constitutional symptoms. These latter patients have usually undergone extensive and often unnecessary diagnostic examinations for unexplained jaundice and have high levels of anxiety. In women, the condition may be subclinical until the patient becomes pregnant or receives oral contraceptives, at which time chemical hyperbilirubinemia becomes frank jaundice. Even in these situations, other routine liver function tests, including serum alkaline phosphatase and transaminase activities, are normal. A cardinal feature of DJS is the accumulation in the lysosomes of centrilobular hepatocytes of dark, coarsely granular pigment. As a result, the liver may be grossly black in appearance. This pigment is thought to be derived from epinephrine metabolites that are not excreted normally. The pigment may disappear during bouts of viral hepatitis, only to reaccumulate slowly after recovery.


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InternalMed_Harrison. Biliary excretion of a number of anionic compounds is compromised in DJS. These include various cholecystographic agents, as well as sulfobromophthalein (Bromsulphalein, BSP), a synthetic dye formerly used in a test of liver function. In this test, the rate of disappearance of BSP from plasma was determined following bolus IV administration. BSP is conjugated with glutathione in the hepatocyte; the resulting conjugate is normally excreted rapidly into the bile canaliculus. Patients with DJS exhibit characteristic rises in plasma concentrations at 90 minutes after injection, due to reflux of conjugated BSP into the circulation from the hepatocyte. Dyes such as ICG that are taken up by hepatocytes but are not further metabolized prior to biliary excretion do not show this reflux phenomenon. Continuous BSP infusion studies suggest a reduction in the time to maximum plasma concentration (t max) for biliary excretion. Bile acid disposition, including hepatocellular uptake and biliary


InternalMed_Harrison

InternalMed_Harrison. Continuous BSP infusion studies suggest a reduction in the time to maximum plasma concentration (t max) for biliary excretion. Bile acid disposition, including hepatocellular uptake and biliary excretion, is normal in DJS. These patients have normal serum and biliary bile acid concentrations and do not have pruritus.


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InternalMed_Harrison. By analogy with findings in several mutant rat strains, the selective defect in biliary excretion of bilirubin conjugates and certain other classes of organic compounds, but not of bile acids, that characterizes DJS in humans was found to reflect defective expression of MRP2, an ATP-dependent canalicular membrane transporter. Several different mutations in the MRP2 gene produce the Dubin-Johnson phenotype, which has an autosomal recessive pattern of inheritance. Although The Hyperbilirubinemias Abbreviations: BRIC, benign recurrent intrahepatic cholestasis; BSEP, bile salt excretory protein; DJS, Dubin-Johnson syndrome; γ-GT, γ-glutamyltransferase; MRP2, multidrug resistance– associated protein 2; OATP1A/1B, organic anion transport proteins 1B1 and 1B3; PFIC, progressive familial intrahepatic cholestasis; ↑↑, increased.


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InternalMed_Harrison. 2004 MRP2 is undoubtedly important in the biliary excretion of conjugated bilirubin, the fact that this pigment is still excreted in the absence of MRP2 suggests that other, as yet uncharacterized, transport proteins may serve in a secondary role in this process. Patients with DJS also have a diagnostic abnormality in urinary coproporphyrin excretion. There are two naturally occurring coproporphyrin isomers, I and III. Normally, ∼75% of the coproporphyrin in urine is isomer III. In urine from DJS patients, total coproporphyrin content is normal, but >80% is isomer I. Heterozygotes for the syndrome show an intermediate pattern. The molecular basis for this phenomenon remains unclear.


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InternalMed_Harrison. Rotor Syndrome This benign, autosomal recessive disorder is clinically similar to DJS (Table 359-2), although it is seen even less frequently. A major phenotypic difference is that the liver in patients with Rotor syndrome has no increased pigmentation and appears totally normal. The only abnormality in routine laboratory tests is an elevation of total serum bilirubin, due to a predominant rise in conjugated bilirubin. This is accompanied by bilirubinuria. Several additional features differentiate Rotor syndrome from DJS. In Rotor syndrome, the gallbladder is usually visualized on oral cholecystography, in contrast to the nonvisualization that is typical of DJS. The pattern of urinary coproporphyrin excretion also differs. The pattern in Rotor syndrome resembles that of many acquired disorders of hepatobiliary function, in which coproporphyrin I, the major coproporphyrin isomer in bile, refluxes from the hepatocyte back into the circulation and is excreted in urine. Thus, total


InternalMed_Harrison

InternalMed_Harrison. disorders of hepatobiliary function, in which coproporphyrin I, the major coproporphyrin isomer in bile, refluxes from the hepatocyte back into the circulation and is excreted in urine. Thus, total urinary coproporphyrin excretion is substantially increased in Rotor syndrome, in contrast to the normal levels seen in DJS. Although the fraction of coproporphyrin I in urine is elevated, it is usually <70% of the total, compared with ≥80% in DJS. The disorders also can be distinguished by their patterns of BSP excretion. Although clearance of BSP from plasma is delayed in Rotor syndrome, there is no reflux of conjugated BSP back into the circulation as seen in DJS. Kinetic analysis of plasma BSP infusion studies suggests the presence of a defect in intrahepatocellular storage of this compound. This has never been demonstrated directly. Recent studies indicate that the molecular basis of Rotor syndrome results from simultaneous deficiency of the plasma membrane transporters OATP1B1 and


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InternalMed_Harrison. This has never been demonstrated directly. Recent studies indicate that the molecular basis of Rotor syndrome results from simultaneous deficiency of the plasma membrane transporters OATP1B1 and OATP1B3. This results in reduced reuptake of conjugated bilirubin that has been pumped out of the cell into the portal circulation by MRP3 (Fig. 359-1).


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InternalMed_Harrison. Benign Recurrent Intrahepatic Cholestasis (BRIC) This rare disorder is characterized by recurrent attacks of pruritus and jaundice. The typical episode begins with mild malaise and elevations in serum aminotransferase levels, followed rapidly by rises in alkaline phosphatase and conjugated bilirubin and onset of jaundice and itching. The first one or two episodes may be misdiagnosed as acute viral hepatitis. The cholestatic episodes, which may begin in childhood or adulthood, can vary in duration from several weeks to months, followed by a complete clinical and biochemical resolution. Intervals between attacks may vary from several months to years. Between episodes, physical examination is normal, as are serum levels of bile acids, bilirubin, transaminases, and alkaline phosphatase. The disorder is familial and has an autosomal recessive pattern of inheritance. BRIC is considered a benign disorder in that it does not lead to cirrhosis or end-stage liver disease. However, the episodes


InternalMed_Harrison

InternalMed_Harrison. is familial and has an autosomal recessive pattern of inheritance. BRIC is considered a benign disorder in that it does not lead to cirrhosis or end-stage liver disease. However, the episodes of jaundice and pruritus can be prolonged and debilitating, and some patients have undergone liver transplantation to relieve the intractable and disabling symptoms. Treatment during the cholestatic episodes is symptomatic; there is no specific treatment to prevent or shorten the occurrence of episodes.


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InternalMed_Harrison. A gene termed FIC1 was recently identified and found to be mutated in patients with BRIC. Curiously, this gene is expressed strongly in the small intestine but only weakly in the liver. The protein encoded by FIC1 shows little similarity to those that have been shown to play a role in bile canalicular excretion of various compounds. Rather, it appears to be a member of a P-type ATPase family that transports aminophospholipids from the outer to the inner leaflet of a variety of cell membranes. Its relationship to the pathobiology of this disorder remains unclear. A second phenotypically identical form of BRIC, termed BRIC type 2, has been described resulting from mutations in the bile salt excretory protein (BSEP), the protein that is defective in progressive familial intrahepatic cholestasis type 2 (Table 359-2). How some mutations in this protein result in the episodic BRIC phenotype is unknown.


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InternalMed_Harrison. Progressive Familial Intrahepatic Cholestasis (FIC) This name is applied to three phenotypically related syndromes (Table 359-2). Progressive FIC type 1 (Byler disease) presents in early infancy as cholestasis that may be initially episodic. However, in contrast to BRIC, Byler disease progresses to malnutrition, growth retardation, and end-stage liver disease during childhood. This disorder is also a consequence of a FIC1 mutation. The functional relationship of the FIC1 protein to the pathogenesis of cholestasis in these disorders is unknown. Two other types of progressive FIC (types 2 and 3) have been described. Progressive FIC type 2 is associated with a mutation in the protein originally named sister of p-glycoprotein, now known as bile salt excretory protein, which is the major bile canalicular exporter of bile acids. As noted above, some mutations of this protein are associated with BRIC type 2, rather than the progressive FIC type 2 phenotype. Progressive FIC type 3 has been


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InternalMed_Harrison. canalicular exporter of bile acids. As noted above, some mutations of this protein are associated with BRIC type 2, rather than the progressive FIC type 2 phenotype. Progressive FIC type 3 has been associated with a mutation of MDR3, a protein that is essential for normal hepatocellular excretion of phospholipids across the bile canaliculus. Although all three types of progressive FIC have similar clinical phenotypes, only type 3 is associated with high serum levels of γ-glutamyltransferase activity. In contrast, activity of this enzyme is normal or only mildly elevated in symptomatic BRIC and progressive FIC types 1 and 2.


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InternalMed_Harrison. Jules L. Dienstag Acute viral hepatitis is a systemic infection affecting the liver predominantly. Almost all cases of acute viral hepatitis are caused by one of five viral agents: hepatitis A virus (HAV), hepatitis B virus (HBV), hepatitis C virus (HCV), the HBV-associated delta agent or hepatitis D virus (HDV), and hepatitis E virus (HEV). All these human hepatitis viruses are RNA viruses, except for hepatitis B, which is a DNA virus but replicates like a retrovirus. Although these agents can be distinguished by their molecular and antigenic properties, all types of viral hepatitis produce clinically similar illnesses. These range from asymptomatic and inapparent to fulminant and fatal acute infections common to all types, on the one hand, and from subclinical persistent infections to rapidly progressive chronic liver disease with cirrhosis and even hepatocellular carcinoma, common to the bloodborne types (HBV, HCV, and HDV), on the other.


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InternalMed_Harrison. VIROLOGY AND ETIOLOGY Hepatitis A HAV is a nonenveloped 27-nm, heat-, acid-, and ether-resistant RNA virus in the Hepatovirus genus of the picornavirus family (Fig. 360-1). Its virion contains four capsid polypeptides, designated VP1 to VP4, which are cleaved posttranslationally from the polyprotein product of a 7500-nucleotide genome. Inactivation of viral activity can be achieved by boiling for 1 min, by contact with formaldehyde and chlorine, or by ultraviolet irradiation. Despite nucleotide sequence variation of up to 20% among isolates of HAV, and despite the recognition of four genotypes affecting humans, all strains of this virus are immunologically indistinguishable and belong to one serotype. Hepatitis A has an incubation period of ~4 weeks. Its replication is limited to the liver, but the virus is present in the liver, bile, stools, and blood during the late incubation period and acute preicteric/ presymptomatic phase of illness. Despite slightly longer persistence of virus


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InternalMed_Harrison. but the virus is present in the liver, bile, stools, and blood during the late incubation period and acute preicteric/ presymptomatic phase of illness. Despite slightly longer persistence of virus in the liver, fecal shedding, viremia, and infectivity diminish rapidly once jaundice becomes apparent. HAV can be cultivated reproducibly in vitro.


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InternalMed_Harrison. template, hepadnaviruses rely on reverse 2005 transcription (effected by the DNA polymerase) of minus-strand DNA from a “pregenomic” RNA intermediate. Then plus-strand DNA is transcribed from the minus-strand DNA template by the DNA-dependent DNA polymerase and converted in the hepatocyte nucleus to a covalently closed circular DNA, which serves as a template for messenger RNA and pregenomic RNA. Viral proteins are translated by the messenger RNA, and the proteins and genome are packaged into FIGURE 360-1 Electron micrographs of hepatitis A virus particles and serum from a patient virions and secreted from the hepatocyte. with hepatitis B. Left: 27-nm hepatitis A virus particles purified from stool of a patient with acute


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InternalMed_Harrison. with hepatitis B. Left: 27-nm hepatitis A virus particles purified from stool of a patient with acute Although HBV is difficult to cultivate hepatitis A and aggregated by antibody to hepatitis A virus. Right: Concentrated serum from a in vitro in the conventional sense from patient with hepatitis B, demonstrating the 42-nm virions, tubular forms, and spherical 22-nm clinical material, several cell lines have particles of hepatitis B surface antigen. 132,000×. (Hepatitis D resembles 42-nm virions of hepatitis been transfected with HBV DNA. Such B but is smaller, 35–37 nm; hepatitis E resembles hepatitis A virus but is slightly larger, 32–34 nm; hepatitis C has been visualized as a 55-nm particle.)


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InternalMed_Harrison. B but is smaller, 35–37 nm; hepatitis E resembles hepatitis A virus but is slightly larger, 32–34 nm; hepatitis C has been visualized as a 55-nm particle.) Antibodies to HAV (anti-HAV) can be detected during acute illness when serum aminotransferase activity is elevated and fecal HAV shedding is still occurring. This early antibody response is predominantly of the IgM class and persists for several (~3) months, rarely for 6–12 months. During convalescence, however, anti-HAV of the IgG class becomes the predominant antibody (Fig. 360-2). Therefore, the diagnosis of hepatitis A is made during acute illness by demonstrating anti-HAV of the IgM class. After acute illness, anti-HAV of the IgG class remains detectable indefinitely, and patients with serum anti-HAV are immune to reinfection. Neutralizing antibody activity parallels the appearance of anti-HAV, and the IgG anti-HAV present in immune globulin accounts for the protection it affords against HAV infection.


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InternalMed_Harrison. Hepatitis B HBV is a DNA virus with a remarkably compact genomic structure; despite its small, circular, 3200-bp size, HBV DNA codes for four sets of viral products with a complex, multiparticle structure. HBV achieves its genomic economy by relying on an efficient strategy of encoding proteins from four overlapping genes: S, C, P, and X (Fig. 360-3), as detailed below. Once thought to be unique among viruses, HBV is now recognized as one of a family of animal viruses, hepadnaviruses (hepatotropic DNA viruses), and is classified as hepadnavirus type 1. Similar viruses infect certain species of woodchucks, ground and tree squirrels, and Pekin ducks, to mention the most carefully characterized. Like HBV, all have the same distinctive three morphologic forms, have counterparts to the envelope and nucleocapsid virus antigens of HBV, replicate in the liver but exist in extrahepatic sites, contain their own endogenous DNA polymerase, have partially double-strand and partially single-strand


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InternalMed_Harrison. and nucleocapsid virus antigens of HBV, replicate in the liver but exist in extrahepatic sites, contain their own endogenous DNA polymerase, have partially double-strand and partially single-strand genomes, are associated with acute and chronic hepatitis and hepatocellular carcinoma, and rely on a replicative strategy unique among DNA viruses but typical of retroviruses. Instead of DNA replication directly from a DNA of hepatitis A virus (HAV). ALT, alanine aminotransferase.


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InternalMed_Harrison. tion of the intact virus and its component proteins. viral proteins anD particles Of the three particulate forms of HBV (Table 360-1), the most numerous are the 22-nm particles, which appear as spherical or long filamentous forms; these are antigenically indistinguishable from the outer surface or envelope protein of HBV and are thought to represent excess viral envelope protein. Outnumbered in serum by a factor of 100 or 1000 to 1 compared with the spheres and tubules are large, 42-nm, double-shelled spherical particles, which represent the intact hepatitis B virion (Fig. 360-1). The envelope protein expressed on the outer surface of the virion and on the smaller spherical and tubular structures is referred to as hepatitis B surface antigen (HBsAg). The concentration of HBsAg and virus particles in the blood may reach 500 μg/mL and 10 trillion particles per milliliter, respectively. The envelope protein, HBsAg, is the product of the S gene of HBV.


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InternalMed_Harrison. Envelope HBsAg subdeterminants include a common group-reactive antigen, a, shared by all HBsAg isolates and one of several subtype-specific antigens—d or y, w or r —as well as other specificities. Hepatitis B isolates fall into one of at least eight subtypes and ten


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InternalMed_Harrison. FIGURE 360-3 Compact genomic structure of hepatitis B virus (HBV). This structure, with overlapping genes, permits HBV to code for multiple proteins. The S gene codes for the “major” envelope protein, HBsAg. Pre-S1 and pre-S2, upstream of S, combine with S to code for two larger proteins, “middle” protein, the product of pre-S2 + S, and “large” protein, the product of pre-S1 + pre-S2 + S. The largest gene, P, codes for DNA polymerase. The C gene codes for two nucleocapsid proteins, HBeAg, a soluble, secreted protein (initiation from the pre-C region of the gene), and HBcAg, the intracellular core protein (initiation after pre-C). The X gene codes for HBxAg, which can transactivate the transcription of cellular and viral genes; its clinical relevance is not known, but it may contribute to carcinogenesis by binding to p53. Hepatitis Virus Type Particle, nm Morphology Genomea Classification Antigen(s) Antibodies Remarks


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InternalMed_Harrison. Hepatitis Virus Type Particle, nm Morphology Genomea Classification Antigen(s) Antibodies Remarks HAV 27 Icosahedral 7.5-kb RNA, Hepatovirus HAV Anti-HAV Early fecal shedding Diagnosis: IgM anti-HAV nonenveloped linear, ss, + Previous infection: IgG anti-HAV HCV Approx. Enveloped 50–80 HDV 35–37 Enveloped hybrid particle with HBsAg coat and HDV core HEV 32–34 Nonenveloped icosahedral 3.2-kb DNA, Hepadnavirus HBsAg Anti-HBs circular, ss/ds 9.4-kb RNA, Hepacivirus HCV Anti-HCV linear, ss, + 1.7-kb RNA, Resembles HBsAg Anti-HBs circular, ss, – viroids and HDAg Anti-HDV plant satellite viruses (genus Deltavirus) 7.6-kb RNA, Hepevirus HEV antigen Anti-HEV linear, ss, + Bloodborne virus; carrier state Acute diagnosis: HBsAg, IgM anti-HBc Chronic diagnosis: IgG anti-HBc, HBsAg Markers of replication: HBeAg, HBV DNA Liver, lymphocytes, other organs


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InternalMed_Harrison. Acute diagnosis: HBsAg, IgM anti-HBc Chronic diagnosis: IgG anti-HBc, HBsAg Markers of replication: HBeAg, HBV DNA Liver, lymphocytes, other organs Nucleocapsid contains DNA and DNA polymerase; present in hepatocyte nucleus; HBcAg does not circulate; HBeAg (soluble, nonparticulate) and HBV DNA circulate—correlate with infectivity and complete virions HBsAg detectable in >95% of patients with acute hepatitis B; found in serum, body fluids, hepatocyte cytoplasm; anti-HBs appears following infection—protective antibody Bloodborne agent, formerly labeled non-A, non-B hepatitis Acute diagnosis: anti-HCV (C33c, C22-3, NS5), HCV RNA Chronic diagnosis: anti-HCV (C100-3, C33c, C223, NS5) and HCV RNA; cytoplasmic location in hepatocytes Defective RNA virus, requires helper function of HBV (hepadnaviruses); HDV antigen (HDAg) present in hepatocyte nucleus Diagnosis: anti-HDV, HDV RNA; HBV/HDV co-infection—IgM anti-HBc and anti-HDV; HDV superinfection—IgG anti-HBc and anti-HDV


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InternalMed_Harrison. Diagnosis: anti-HDV, HDV RNA; HBV/HDV co-infection—IgM anti-HBc and anti-HDV; HDV superinfection—IgG anti-HBc and anti-HDV Agent of enterically transmitted hepatitis; rare in United States; occurs in Asia, Mediterranean countries, Central America Diagnosis: IgM/IgG anti-HEV (assays not routinely available); virus in stool, bile, hepatocyte cytoplasm ass, single-strand; ss/ds, partially single-strand, partially double-strand; −, minus-strand; +, plus-strand. Note: See text for abbreviations.


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InternalMed_Harrison. Note: See text for abbreviations. genotypes (A–J). Geographic distribution of genotypes and subtypes varies; genotypes A (corresponding to subtype adw) and D (ayw) predominate in the United States and Europe, whereas genotypes B (adw) and C (adr) predominate in Asia. Clinical course and outcome are independent of subtype, but genotype B appears to be associated with less rapidly progressive liver disease and cirrhosis and a lower likelihood, or delayed appearance, of hepatocellular carcinoma than genotype C or D. Patients with genotype A are more likely to clear circulating viremia and to achieve HBeAg and HBsAg seroconversion, both spontaneously and in response to antiviral therapy. In addition, “precore” mutations are favored by certain genotypes (see below).


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InternalMed_Harrison. Upstream of the S gene are the pre-S genes (Fig. 360-3), which code for pre-S gene products, including receptors on the HBV surface for polymerized human serum albumin and for hepatocyte membrane proteins. The pre-S region actually consists of both pre-S1 and pre-S2. Depending on where translation is initiated, three potential HBsAg gene products are synthesized. The protein product of the S gene is HBsAg (major protein), the product of the S region plus the adjacent pre-S2 region is the middle protein, and the product of the pre-S1 plus pre-S2 plus S regions is the large protein. Compared with the smaller spherical and tubular particles of HBV, complete 42-nm virions are enriched in the large protein. Both pre-S proteins and their respective antibodies can be detected during HBV infection, and the period of pre-S antigenemia appears to coincide with other markers of virus replication, as detailed below; however, pre-S proteins have little clinical relevance and are not included in


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InternalMed_Harrison. and the period of pre-S antigenemia appears to coincide with other markers of virus replication, as detailed below; however, pre-S proteins have little clinical relevance and are not included in routine serologic testing repertoires.


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InternalMed_Harrison. The intact 42-nm virion contains a 27-nm nucleocapsid core particle. Nucleocapsid proteins are coded for by the C gene. The antigen expressed on the surface of the nucleocapsid core is hepatitis B core antigen (HBcAg), and its corresponding antibody is anti-HBc. A third HBV antigen is hepatitis B e antigen (HBeAg), a soluble, nonparticulate, nucleocapsid protein that is immunologically distinct from intact HBcAg but is a product of the same C gene. The C gene has two initiation codons, a precore and a core region (Fig. 360-3). If translation is initiated at the precore region, the protein product is HBeAg, which has a signal peptide that binds it to the smooth endoplasmic reticulum, the secretory apparatus of the cell, leading to its secretion into the circulation. If translation begins at the core region, HBcAg is the protein product; it has no signal peptide, it is not secreted, but it assembles into nucleocapsid particles, which bind to and incorporate RNA, and which, ultimately,


InternalMed_Harrison

InternalMed_Harrison. at the core region, HBcAg is the protein product; it has no signal peptide, it is not secreted, but it assembles into nucleocapsid particles, which bind to and incorporate RNA, and which, ultimately, contain HBV DNA. Also packaged within the nucleocapsid core is a DNA polymerase, which directs replication and repair of HBV DNA. When packaging within viral proteins is complete, synthesis of the incomplete plus strand stops; this accounts for the single-strand gap and for differences in the size of the gap. HBcAg particles remain in the hepatocyte, where they are readily detectable by immunohistochemical staining and are exported after encapsidation by an envelope of HBsAg. Therefore, naked core particles do not circulate in the serum. The secreted nucleocapsid protein, HBeAg, provides a convenient, readily detectable, qualitative marker of HBV replication and relative infectivity.


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InternalMed_Harrison. HBsAg-positive serum containing HBeAg is more likely to be highly infectious and to be associated with the presence of hepatitis B virions (and detectable HBV DNA, see below) than HBeAg-negative or anti-HBe-positive serum. For example, HBsAg-positive mothers who are HBeAg-positive almost invariably (>90%) transmit hepatitis B infection to their offspring, whereas HBsAg-positive mothers with anti-HBe rarely (10–15%) infect their offspring. Early during the course of acute hepatitis B, HBeAg appears transiently; its disappearance may be a harbinger of clinical improvement and resolution of infection. Persistence of HBeAg in serum beyond the first 3 months of acute infection may be predictive of the development of chronic infection, and the presence of HBeAg during chronic hepatitis B tends to be associated with ongoing viral replication, infectivity, and inflammatory liver injury (except during the early decades after perinatally acquired HBV infection; see below).


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InternalMed_Harrison. The third and largest of the HBV genes, the P gene (Fig. 360-3), codes for HBV DNA polymerase; as noted above, this enzyme has both DNA-dependent DNA polymerase and RNA-dependent reverse transcriptase activities. The fourth gene, X, codes for a small, non-particulate protein, hepatitis B x antigen (HBxAg), that is capable of transactivating the transcription of both viral and cellular genes (Fig. 360-3). In the cytoplasm, HBxAg effects calcium release (possibly from mitochondria), which activates signal-transduction pathways that lead to stimulation of HBV reverse transcription and HBV DNA replication. Such transactivation may enhance the replication of HBV, leading to the clinical association observed between the expression of HBxAg and antibodies to it in patients with severe chronic hepatitis and hepatocellular carcinoma. The transactivating activity can enhance the transcription and replication of other viruses besides HBV, such as HIV. Cellular processes transactivated by X


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InternalMed_Harrison. hepatitis and hepatocellular carcinoma. The transactivating activity can enhance the transcription and replication of other viruses besides HBV, such as HIV. Cellular processes transactivated by X include the human interferon γ gene and class I major histocompatibility genes; potentially, these effects could contribute to enhanced susceptibility of HBV-infected hepatocytes to cytolytic T cells. The expression of X can also induce programmed cell death (apoptosis). The clinical relevance of HBxAg is limited, however, and testing for it is not part of routine clinical practice.


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InternalMed_Harrison. seroloGic anD viroloGic markers After a person is infected with HBV, the first virologic marker detectable in serum within 1–12 weeks, usually between 8 and 12 weeks, is HBsAg (Fig. 360-4). Circulating HBsAg precedes elevations of serum aminotransferase activity and clinical symptoms by 2–6 weeks and remains detectable during the entire icteric or symptomatic phase of acute hepatitis B and beyond. In typical cases, HBsAg becomes undetectable 1–2 months after the onset of jaundice and rarely persists beyond 6 months. After HBsAg disappears, antibody to HBsAg (anti-HBs) becomes detectable in serum and remains detectable indefinitely thereafter. Because HBcAg is intracellular and, when in the serum, sequestered within an HBsAg coat, naked core particles do not circulate in serum, and therefore, HBcAg is not detectable routinely in the serum of patients with HBV infection. By contrast, anti-HBc is readily demonstrable in serum, beginning within the first 1–2 weeks after the appearance of


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InternalMed_Harrison. HBcAg is not detectable routinely in the serum of patients with HBV infection. By contrast, anti-HBc is readily demonstrable in serum, beginning within the first 1–2 weeks after the appearance of HBsAg and preceding detectable levels of anti-HBs by weeks to months.


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InternalMed_Harrison. of acute hepatitis B. ALT, alanine aminotransferase.


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InternalMed_Harrison. Because variability exists in the time of appearance of anti-HBs after 2007 HBV infection, occasionally a gap of several weeks or longer may separate the disappearance of HBsAg and the appearance of anti-HBs. During this “gap” or “window” period, anti-HBc may represent the only serologic evidence of current or recent HBV infection, and blood containing anti-HBc in the absence of HBsAg and anti-HBs has been implicated in transfusion-associated hepatitis B. In part because the sensitivity of immunoassays for HBsAg and anti-HBs has increased, however, this window period is rarely encountered. In some persons, years after HBV infection, anti-HBc may persist in the circulation longer than anti-HBs. Therefore, isolated anti-HBc does not necessarily indicate active virus replication; most instances of isolated anti-HBc represent hepatitis B infection in the remote past. Rarely, however, isolated anti-HBc represents low-level hepatitis B viremia, with HBsAg below the detection threshold, and,


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InternalMed_Harrison. of isolated anti-HBc represent hepatitis B infection in the remote past. Rarely, however, isolated anti-HBc represents low-level hepatitis B viremia, with HBsAg below the detection threshold, and, occasionally, isolated anti-HBc represents a cross-reacting or false-positive immunologic specificity. Recent and remote HBV infections can be distinguished by determination of the immunoglobulin class of anti-HBc. Anti-HBc of the IgM class (IgM anti-HBc) predominates during the first 6 months after acute infection, whereas IgG anti-HBc is the predominant class of anti-HBc beyond 6 months. Therefore, patients with current or recent acute hepatitis B, including those in the anti-HBc window, have IgM anti-HBc in their serum. In patients who have recovered from hepatitis B in the remote past as well as those with chronic HBV infection, anti-HBc is predominantly of the IgG class. Infrequently, in ≤1–5% of patients with acute HBV infection, levels of HBsAg are too low to be detected; in such


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InternalMed_Harrison. well as those with chronic HBV infection, anti-HBc is predominantly of the IgG class. Infrequently, in ≤1–5% of patients with acute HBV infection, levels of HBsAg are too low to be detected; in such cases, the presence of IgM anti-HBc establishes the diagnosis of acute hepatitis B. When isolated anti-HBc occurs in the rare patient with chronic hepatitis B whose HBsAg level is below the sensitivity threshold of contemporary immunoassays (a low-level carrier), anti-HBc is of the IgG class. Generally, in persons who have recovered from hepatitis B, anti-HBs and anti-HBc persist indefinitely.


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InternalMed_Harrison. The temporal association between the appearance of anti-HBs and resolution of HBV infection as well as the observation that persons with anti-HBs in serum are protected against reinfection with HBV suggests that anti-HBs is the protective antibody. Therefore, strategies for prevention of HBV infection are based on providing susceptible persons with circulating anti-HBs (see below). Occasionally, in ~10% of patients with chronic hepatitis B, low-level, low-affinity anti-HBs can be detected. This antibody is directed against a subtype determinant different from that represented by the patient’s HBsAg; its presence is thought to reflect the stimulation of a related clone of antibody-forming cells, but it has no clinical relevance and does not signal imminent clearance of hepatitis B. These patients with HBsAg and such nonneutralizing anti-HBs should be categorized as having chronic HBV infection.


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InternalMed_Harrison. The other readily detectable serologic marker of HBV infection, HBeAg, appears concurrently with or shortly after HBsAg. Its appearance coincides temporally with high levels of virus replication and reflects the presence of circulating intact virions and detectable HBV DNA (with the notable exception of patients with precore mutations who cannot synthesize HBeAg—see “Molecular Variants”). Pre-S1 and pre-S2 proteins are also expressed during periods of peak replication, but assays for these gene products are not routinely available. In self-limited HBV infections, HBeAg becomes undetectable shortly after peak elevations in aminotransferase activity, before the disappearance of HBsAg, and anti-HBe then becomes detectable, coinciding with a period of relatively lower infectivity (Fig. 360-4). Because markers of HBV replication appear transiently during acute infection, testing for such markers is of little clinical utility in typical cases of acute HBV infection. In contrast, markers of


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InternalMed_Harrison. Because markers of HBV replication appear transiently during acute infection, testing for such markers is of little clinical utility in typical cases of acute HBV infection. In contrast, markers of HBV replication provide valuable information in patients with protracted infections.


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InternalMed_Harrison. Departing from the pattern typical of acute HBV infections, in chronic HBV infection, HBsAg remains detectable beyond 6 months, anti-HBc is primarily of the IgG class, and anti-HBs is either undetectable or detectable at low levels (see “Laboratory Features”) (Fig. 360-5). During early chronic HBV infection, HBV DNA can be detected both in serum and in hepatocyte nuclei, where it is present in free or episomal form. This relatively highly replicative stage of HBV infection is type chronic hepatitis B. HBeAg and hepatitis B virus (HBV) DNA can be detected in serum during the relatively replicative phase of chronic infection, which is associated with infectivity and liver injury. Seroconversion from the replicative phase to the relatively nonreplicative phase occurs at a rate of ~10% per year and is heralded by an acute hepatitis–like elevation of alanine aminotransferase (ALT) activity; during the nonreplicative phase, infectivity and liver injury are limited. In HBeAg-negative chronic


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InternalMed_Harrison. and is heralded by an acute hepatitis–like elevation of alanine aminotransferase (ALT) activity; during the nonreplicative phase, infectivity and liver injury are limited. In HBeAg-negative chronic hepatitis B associated with mutations in the precore region of the HBV genome, replicative chronic hepatitis B occurs in the absence of HBeAg.


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InternalMed_Harrison. the time of maximal infectivity and liver injury; HBeAg is a qualitative marker and HBV DNA a quantitative marker of this replicative phase, during which all three forms of HBV circulate, including intact virions. Over time, the relatively replicative phase of chronic HBV infection gives way to a relatively nonreplicative phase. This occurs at a rate of ~10% per year and is accompanied by seroconversion from HBeAg to anti-HBe. In many cases, this seroconversion coincides with a transient, usually mild, acute hepatitis-like elevation in aminotransferase activity, believed to reflect cell-mediated immune clearance of virus-infected hepatocytes. In the nonreplicative phase of chronic infection, when HBV DNA is demonstrable in hepatocyte nuclei, it tends to be integrated into the host genome. In this phase, only spherical and tubular forms of HBV, not intact virions, circulate, and liver injury tends to subside. Most such patients would be characterized as inactive HBV carriers. In


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InternalMed_Harrison. In this phase, only spherical and tubular forms of HBV, not intact virions, circulate, and liver injury tends to subside. Most such patients would be characterized as inactive HBV carriers. In reality, the designations replicative and nonreplicative are only relative; even in the so-called nonreplicative phase, HBV replication can be detected at levels of approximately ≤103 virions with highly sensitive amplification probes such as the polymerase chain reaction (PCR); below this replication threshold, liver injury and infectivity of HBV are limited to negligible. Still, the distinctions are pathophysiologically and clinically meaningful. Occasionally, nonreplicative HBV infection converts back to replicative infection. Such spontaneous reactivations are accompanied by reexpression of HBeAg and HBV DNA, and sometimes of IgM anti-HBc, as well as by exacerbations of liver injury. Because high-titer IgM anti-HBc can reappear during acute exacerbations of chronic hepatitis B, relying on


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InternalMed_Harrison. and HBV DNA, and sometimes of IgM anti-HBc, as well as by exacerbations of liver injury. Because high-titer IgM anti-HBc can reappear during acute exacerbations of chronic hepatitis B, relying on IgM anti-HBc versus IgG anti-HBc to distinguish between acute and chronic hepatitis B infection, respectively, may not always be reliable; in such cases, patient history is invaluable in helping to distinguish de novo acute hepatitis B infection from acute exacerbation of chronic hepatitis B infection.


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InternalMed_Harrison. molecular variants Variation occurs throughout the HBV genome, and clinical isolates of HBV that do not express typical viral proteins have been attributed to mutations in individual or even multiple gene locations. For example, variants have been described that lack nucleocapsid proteins (commonly), envelope proteins (very rarely), or both. Two categories of naturally occurring HBV variants have attracted the most attention. One of these was identified initially in Mediterranean countries among patients with severe chronic HBV infection and detectable HBV DNA but with anti-HBe instead of HBeAg. These patients were found to be infected with an HBV mutant that contained an alteration in the precore region rendering the virus incapable of encoding HBeAg. Although several potential mutation sites exist in the pre-C region, the region of the C gene necessary for the expression of HBeAg (see “Virology and Etiology”), the most commonly encountered in such patients is a single base


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InternalMed_Harrison. mutation sites exist in the pre-C region, the region of the C gene necessary for the expression of HBeAg (see “Virology and Etiology”), the most commonly encountered in such patients is a single base substitution, from G to A in the second to last codon of the pre-C gene at nucleotide 1896. This substitution results in the replacement of the TGG tryptophan codon by a stop codon (TAG), which prevents the translation of HBeAg. Another mutation, in the core-promoter region, prevents transcription of the coding region for HBeAg and yields an HBeAgnegative phenotype. Patients with such mutations in the precore region and who are unable to secrete HBeAg may have severe liver disease that progresses more rapidly to cirrhosis, or alternatively, they are identified clinically later in the course of the natural history of chronic hepatitis B, when the disease is more advanced. Both “wild-type” HBV and precore-mutant HBV can coexist in the same patient, or mutant HBV may arise late during


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InternalMed_Harrison. of the natural history of chronic hepatitis B, when the disease is more advanced. Both “wild-type” HBV and precore-mutant HBV can coexist in the same patient, or mutant HBV may arise late during wild-type HBV infection. In addition, clusters of fulminant hepatitis B in Israel and Japan were attributed to common-source infection with a precore mutant. Fulminant hepatitis B in North America and western Europe, however, occurs in patients infected with wild-type HBV, in the absence of precore mutants, and both precore mutants and other mutations throughout the HBV genome occur commonly, even in patients with typical, self-limited, milder forms of HBV infection. HBeAg-negative chronic hepatitis with mutations in the precore region is now the most frequently encountered form of hepatitis B in Mediterranean countries and in Europe. In the United States, where HBV genotype A (less prone to G1896A mutation) is prevalent, precore-mutant HBV is much less common; however, as a result of


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InternalMed_Harrison. B in Mediterranean countries and in Europe. In the United States, where HBV genotype A (less prone to G1896A mutation) is prevalent, precore-mutant HBV is much less common; however, as a result of immigration from Asia and Europe, the proportion of HBeAg-negative hepatitis B–infected individuals has increased in the United States, and they now represent approximately 30–40% of patients with chronic hepatitis B. Characteristic of such HBeAg-negative chronic hepatitis B are lower levels of HBV DNA (usually ≤105 IU/mL) and one of several patterns of aminotransferase activity—persistent elevations, periodic fluctuations above the normal range, and periodic fluctuations between the normal and elevated range.


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InternalMed_Harrison. The second important category of HBV mutants consists of escape mutants, in which a single amino acid substitution, from glycine to arginine, occurs at position 145 of the immunodominant a determinant common to all HBsAg subtypes. This HBsAg alteration leads to a critical conformational change that results in a loss of neutralizing activity by anti-HBs. This specific HBV/a mutant has been observed in two situations, active and passive immunization, in which humoral immunologic pressure may favor evolutionary change (“escape”) in the virus— in a small number of hepatitis B vaccine recipients who acquired HBV infection despite the prior appearance of neutralizing anti-HBs and in HBV-infected liver transplant recipients treated with a high-potency human monoclonal anti-HBs preparation. Although such mutants have not been recognized frequently, their existence raises a concern that may complicate vaccination strategies and serologic diagnosis.


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InternalMed_Harrison. Different types of mutations emerge during antiviral therapy of chronic hepatitis B with nucleoside analogues; such “YMDD” and similar mutations in the polymerase motif of HBV are described in Chap. 362. extraHepatic sites Hepatitis B antigens and HBV DNA have been identified in extrahepatic sites, including lymph nodes, bone marrow, circulating lymphocytes, spleen, and pancreas. Although the virus does not appear to be associated with tissue injury in any of these extrahepatic sites, its presence in these “remote” reservoirs has been invoked (but is not necessary) to explain the recurrence of HBV infection after orthotopic liver transplantation. The clinical relevance of such extra-hepatic HBV is limited.


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InternalMed_Harrison. Hepatitis D The delta hepatitis agent, or HDV, the only member of the genus Deltavirus, is a defective RNA virus that co-infects with and requires the helper function of HBV (or other hepadnaviruses) for its replication and expression. Slightly smaller than HBV, HDV is a formalin-sensitive, 35to 37-nm virus with a hybrid structure. Its nucleocapsid expresses HDV antigen (HDAg), which bears no antigenic homology with any of the HBV antigens, and contains the virus genome. The HDV core is “encapsidated” by an outer envelope of HBsAg, indistinguishable from that of HBV except in its relative compositions of major, middle, and large HBsAg component proteins. The genome is a small, 1700-nucleotide, circular, single-strand RNA of negative polarity that is nonhomologous with HBV DNA (except for a small area of the polymerase gene) but that has features and the rolling circle model of replication common to genomes of plant satellite viruses or viroids. HDV RNA contains many areas of internal


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InternalMed_Harrison. a small area of the polymerase gene) but that has features and the rolling circle model of replication common to genomes of plant satellite viruses or viroids. HDV RNA contains many areas of internal complementarity; therefore, it can fold on itself by internal base pairing to form an unusual, very stable, rodlike structure that contains a very stable, self-cleaving and self-ligating ribozyme. HDV RNA requires host RNA polymerase II for its replication in the hepatocyte nucleus via RNA-directed RNA synthesis by transcription of genomic RNA to a complementary antigenomic (plus strand) RNA; the antigenomic RNA, in turn, serves as a template for subsequent genomic RNA synthesis effected by host RNA polymerase I. HDV RNA has only one open reading frame, and HDAg, a product of the antigenomic strand, is the only known HDV protein; HDAg exists in two forms: a small, 195-amino-acid species, which plays a role in facilitating HDV RNA replication, and a large, 214-amino-acid species, which


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InternalMed_Harrison. strand, is the only known HDV protein; HDAg exists in two forms: a small, 195-amino-acid species, which plays a role in facilitating HDV RNA replication, and a large, 214-amino-acid species, which appears to suppress replication but is required for assembly of the antigen into virions. HDV antigens have been shown to bind directly to RNA polymerase II, resulting in stimulation of transcription. Although complete hepatitis D virions and liver injury require the cooperative helper function of HBV, intracellular replication of HDV RNA can occur without HBV. Genomic heterogeneity among HDV isolates has been described; however, pathophysiologic and clinical consequences of this genetic diversity have not been recognized. The clinical spectrum of hepatitis D is common to all eight genotypes identified, the predominant of which is genotype 1.


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InternalMed_Harrison. HDV can either infect a person simultaneously with HBV (coinfection) or superinfect a person already infected with HBV (superinfection); when HDV infection is transmitted from a donor with one HBsAg subtype to an HBsAg-positive recipient with a different subtype, HDV assumes the HBsAg subtype of the recipient, rather than the donor. Because HDV relies absolutely on HBV, the duration of HDV infection is determined by the duration of (and cannot outlast) HBV infection. HDV replication tends to suppress HBV replication; therefore, patients with hepatitis D tend to have lower levels of HBV replication. HDV antigen is expressed primarily in hepatocyte nuclei and is occasionally detectable in serum. During acute HDV infection, anti-HDV of the IgM class predominates, and 30–40 days may elapse after symptoms appear before anti-HDV can be detected. In self-limited infec-500 virus polyprotein of ~3000 amino acids, which is cleaved after transla-2009 tion to yield 10 viral proteins. The 5′ end


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InternalMed_Harrison. symptoms appear before anti-HDV can be detected. In self-limited infec-500 virus polyprotein of ~3000 amino acids, which is cleaved after transla-2009 tion to yield 10 viral proteins. The 5′ end of the genome consists of an untranslated region (containing an internal ribosomal entry site, IRES) adjacent to the genes for three structural proteins, the nucleocapsid core protein, C, and two structural envelope glycoproteins, E1 and E2. The 5′ untranslated region and core gene are highly conserved among genotypes, but the envelope proteins are coded for by the hypervariable region, which varies from isolate to isolate and may allow the virus to evade host immunologic containment directed at accessible virus-envelope proteins. The 3′ end of the genome also includes an untranslated region and contains the genes for seven nonstructural (NS) proteins, p7, NS2, NS3, NS4A, NS4B, NS5A, and NS5B. p7 is a membrane ion channel protein necessary for efficient assembly and release of HCV. The NS2


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InternalMed_Harrison. contains the genes for seven nonstructural (NS) proteins, p7, NS2, NS3, NS4A, NS4B, NS5A, and NS5B. p7 is a membrane ion channel protein necessary for efficient assembly and release of HCV. The NS2 cysteine protease cleaves NS3 from NS2, and the NS3-4A serine protease cleaves all the downstream proteins from the polyprotein. Important NS proteins involved in virus replication include the NS3 helicase; NS3-4A serine protease; the multifunctional membrane-associated phosphoprotein NS5A, an essential component of the viral replication membranous web (along with NS4B); and the


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InternalMed_Harrison. NS5B RNA-dependent RNA polymerase (Fig. 360-6). Because HCV does not replicate via a DNA intermediate, it does not integrate into the host genome. Because HCV tends to circulate in relatively low titer, 103−107 virions/mL, visualization of the 50to 80-nm virus particles remains difficult. Still, the replication rate of HCV is very high, 1012 virions per day; its half-life is 2.7 h. The chimpanzee is a helpful but cumbersome animal model. Although a robust, reproducible, small animal model is lacking, HCV replication has been documented in an immunodeficient mouse model containing explants of human liver and in transgenic mouse and rat models. Although in vitro replication is difficult, replicons in hepatocellular carcinoma–derived cell lines support replication of genetically manipulated, truncated, or full-length HCV RNA (but not intact virions); infectious pseudotyped retroviral HCV particles have been shown to yield functioning envelope proteins. In 2005, complete replication of


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InternalMed_Harrison. truncated, or full-length HCV RNA (but not intact virions); infectious pseudotyped retroviral HCV particles have been shown to yield functioning envelope proteins. In 2005, complete replication of HCV and intact 55-nm virions were described in cell culture systems. HCV entry into the hepatocyte occurs via the nonliver-specific CD81 receptor and the liver-specific tight junction protein claudin-1. A growing list of additional host receptors to which HCV binds on cell entry includes occludin, low-density lipoprotein receptors, glycosaminoglycans, scavenger receptor B1, and epidermal growth factor receptor, among others. Relying on the same assembly and secretion pathway as low-density and very-low-density lipoproteins, HCV is a lipoviroparticle and masquerades as a lipoprotein, which may limit its visibility to the adaptive immune system and which may explain its ability to evade immune containment and clearance. After viral entry and uncoating, translation is initiated by the IRES on


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InternalMed_Harrison. its visibility to the adaptive immune system and which may explain its ability to evade immune containment and clearance. After viral entry and uncoating, translation is initiated by the IRES on the endoplasmic reticulum tion, anti-HDV is low-titer and transient, AA rarely remaining detectable beyond the Helicase


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InternalMed_Harrison. Envelope Serine RNA-dependent clearance of HBsAg and HDV antigen. Core glycoproteins protease RNA polymerase antigen in the liver and HDV RNA in serum and liver can be detected during HDV replication. Conserved Hypervariable


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InternalMed_Harrison. Hepatitis C Hepatitis C virus, which, FIGURE 360-6 Organization of the hepatitis C virus genome and its associated, 3000-aminobefore its identification was labeled acid (AA) proteins. The three structural genes at the 5’ end are the core region, C, which codes “non-A, non-B hepatitis,” is a linear, sin-for the nucleocapsid, and the envelope regions, E1 and E2, which code for envelope glycoproteins. gle-strand, positive-sense, 9600-nucleo-The 5’ untranslated region and the C region are highly conserved among isolates, whereas the tide RNA virus, the genome of which envelope domain E2 contains the hypervariable region. At the 3’ end are seven nonstructural (NS) is similar in organization to that of fla-regions—p7, a membrane protein adjacent to the structural proteins that appears to function as an viviruses and pestiviruses; HCV is the ion channel; NS2, which codes for a cysteine protease; NS3, which codes for a serine protease and only member of the genus Hepacivirus an RNA helicase;


InternalMed_Harrison

InternalMed_Harrison. as an viviruses and pestiviruses; HCV is the ion channel; NS2, which codes for a cysteine protease; NS3, which codes for a serine protease and only member of the genus Hepacivirus an RNA helicase; NS4 and NS4B; NS5A, a multifunctional membrane-associated phosphoprotein, in the family Flaviviridae. The HCV an essential component of the viral replication membranous web; and NS5B, which codes for an genome contains a single, large open RNA-dependent RNA polymerase. After translation of the entire polyprotein, individual proteins are reading frame (gene) that codes for a cleaved by both host and viral proteases.


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InternalMed_Harrison. In chronic HDV infection, anti-HDV circulates in high titer, and both IgM and IgG anti-HDV can be detected. HDV 5' 3'C E1 E2 NS2 NS3 NS4B NS5A NS5B 2010 membrane, and the HCV polyprotein is cleaved during translation and posttranslationally by host cellular proteases as well as HCV NS2-3 and NS3-4A proteases. Host cofactors involved in HCV replication include cyclophilin A, which binds to NS5A and yields conformational changes required for viral replication, and liver-specific host microRNA miR-122. At least six distinct major genotypes (and a minor genotype 7), as well as >50 subtypes within genotypes, of HCV have been identified by nucleotide sequencing. Genotypes differ from one another in sequence homology by ≥30%, and subtypes differ by approximately 20%. Because divergence of HCV isolates within a genotype or subtype and within the same host may vary insufficiently to define a distinct genotype, these intragenotypic differences are referred to as quasispecies and differ in


InternalMed_Harrison

InternalMed_Harrison. isolates within a genotype or subtype and within the same host may vary insufficiently to define a distinct genotype, these intragenotypic differences are referred to as quasispecies and differ in sequence homology by only a few percent. The genotypic and quasispecies diversity of HCV, resulting from its high mutation rate, interferes with effective humoral immunity. Neutralizing antibodies to HCV have been demonstrated, but they tend to be short lived, and HCV infection does not induce lasting immunity against reinfection with different virus isolates or even the same virus isolate. Thus, neither heterologous nor homologous immunity appears to develop commonly after acute HCV infection. Some HCV genotypes are distributed worldwide, whereas others are more geographically confined (see “Epidemiology and Global Features”). In addition, differences exist among genotypes in responsiveness to antiviral therapy but not in pathogenicity or clinical progression (except for genotype 3, in


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InternalMed_Harrison. “Epidemiology and Global Features”). In addition, differences exist among genotypes in responsiveness to antiviral therapy but not in pathogenicity or clinical progression (except for genotype 3, in which hepatic steatosis and clinical progression are more likely). Currently available, third-generation immunoassays, which incorporate proteins from the core, NS3, and NS5 regions, detect anti-HCV antibodies during acute infection. The most sensitive indicator of HCV infection is the presence of HCV RNA, which requires molecular amplification by PCR or transcription-mediated amplification (TMA) (Fig. 360-7). To allow standardization of the quantification of HCV RNA among laboratories and commercial assays, HCV RNA is reported as international units (IUs) per milliliter; quantitative assays with a broad dynamic range are available that allow detection of HCV RNA with a sensitivity as low as 5 IU/mL. HCV RNA can be detected within a few days of exposure to HCV—well before the appearance of


InternalMed_Harrison

InternalMed_Harrison. a broad dynamic range are available that allow detection of HCV RNA with a sensitivity as low as 5 IU/mL. HCV RNA can be detected within a few days of exposure to HCV—well before the appearance of anti-HCV—and tends to persist for the duration of HCV infection. Application of sensitive molecular probes for HCV RNA has revealed the presence of replicative HCV in peripheral blood lymphocytes of infected persons; however, as is the case for HBV in lymphocytes, the clinical relevance of HCV lymphocyte infection is not known.


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InternalMed_Harrison. Hepatitis E Previously labeled epidemic or enterically transmitted non-A, non-B hepatitis, HEV is an enterically transmitted virus that causes clinically apparent hepatitis primarily in India, Asia, Africa, and Central America; in those geographic areas, HEV is the most common cause of acute hepatitis; one-third of the global population appears to have been infected. This agent, with epidemiologic features resembling those of hepatitis A, is a 27to 34-nm, nonenveloped, HAV-like virus with a 7200-nucleotide, single-strand, positive-sense RNA genome. HEV has three open reading frames (ORF) (genes), the largest of FIGURE 360-7 Scheme of typical laboratory features during acute hepatitis C progressing to chronicity. Hepatitis C virus (HCV) RNA is the first detectable event, preceding alanine aminotransferase (ALT) elevation and the appearance of anti-HCV.


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InternalMed_Harrison. which, ORF1, encodes nonstructural proteins involved in virus replication. A middle-sized gene, ORF2, encodes the nucleocapsid protein, the major nonstructural protein, and the smallest, ORF3, encodes a structural protein whose function remains undetermined. All HEV isolates appear to belong to a single serotype, despite genomic heterogeneity of up to 25% and the existence of five genotypes, only four of which have been detected in humans; genotypes 1 and 2 appear to be more virulent, whereas genotypes 3 and 4 are more attenuated and account for subclinical infections. Contributing to the perpetuation of this virus are animal reservoirs, most notably in swine. No genomic or antigenic homology, however, exists between HEV and HAV or other picornaviruses; and HEV, although resembling caliciviruses, is sufficiently distinct from any known agent to merit its own classification as a unique genus, Hepevirus, within the family Hepeviridae. The virus has been detected in stool, bile, and


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InternalMed_Harrison. is sufficiently distinct from any known agent to merit its own classification as a unique genus, Hepevirus, within the family Hepeviridae. The virus has been detected in stool, bile, and liver and is excreted in the stool during the late incubation period. Both IgM anti-HEV during early acute infection and IgG anti-HEV predominating after the first 3 months can be detected. Currently, availability and reliability of serologic/virologic testing for HEV infection is limited but can be done in specialized laboratories (e.g., the Centers for Disease Control and Prevention).


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InternalMed_Harrison. Under ordinary circumstances, none of the hepatitis viruses is known to be directly cytopathic to hepatocytes. Evidence suggests that the clinical manifestations and outcomes after acute liver injury associated with viral hepatitis are determined by the immunologic responses of the host. Among the viral hepatitides, the immunopathogenesis of hepatitis B and C has been studied most extensively.


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InternalMed_Harrison. Hepatitis B For HBV, the existence of inactive hepatitis B carriers with normal liver histology and function suggests that the virus is not directly cytopathic. The fact that patients with defects in cellular immune competence are more likely to remain chronically infected rather than to clear HBV supports the role of cellular immune responses in the pathogenesis of hepatitis B–related liver injury. The model that has the most experimental support involves cytolytic T cells sensitized specifically to recognize host and hepatitis B viral antigens on the liver cell surface. Nucleocapsid proteins (HBcAg and possibly HBeAg), present on the cell membrane in minute quantities, are the viral target antigens that, with host antigens, invite cytolytic T cells to destroy HBV-infected hepatocytes. Differences in the robustness and broad polyclonality of CD8+ cytolytic T cell responsiveness; in the level of HBV-specific helper CD4+ T cells; in attenuation, depletion, and exhaustion of


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InternalMed_Harrison. Differences in the robustness and broad polyclonality of CD8+ cytolytic T cell responsiveness; in the level of HBV-specific helper CD4+ T cells; in attenuation, depletion, and exhaustion of virus-specific T cells; in viral T cell epitope escape mutations that allow the virus to evade T cell containment; and in the elaboration of antiviral cytokines by T cells have been invoked to explain differences in outcomes between those who recover after acute hepatitis and those who progress to chronic hepatitis, or between those with mild and those with severe (fulminant) acute HBV infection.


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InternalMed_Harrison. Although a robust cytolytic T cell response occurs and eliminates virus-infected liver cells during acute hepatitis B, >90% of HBV DNA has been found in experimentally infected chimpanzees to disappear from the liver and blood before maximal T cell infiltration of the liver and before most of the biochemical and histologic evidence of liver injury. This observation suggests that components of the innate immune system and inflammatory cytokines, independent of cytopathic antiviral mechanisms, participate in the early immune response to HBV infection; this effect has been shown to represent elimination of HBV replicative intermediates from the cytoplasm and covalently closed circular viral DNA from the nucleus of infected hepatocytes. In turn, the innate immune response to HBV infection is mediated largely by natural killer (NK) cell cytotoxicity, activated by immunosuppressive cytokines (e.g., interleukin [IL] 10 and transforming growth factor [TGF] β), reduced signals from inhibitory


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InternalMed_Harrison. mediated largely by natural killer (NK) cell cytotoxicity, activated by immunosuppressive cytokines (e.g., interleukin [IL] 10 and transforming growth factor [TGF] β), reduced signals from inhibitory receptor expression (e.g., major histocompatibility complex), or increased signals from activating receptor expression on infected hepatocytes. In addition, NK cells reduce helper CD4+ cells, which results in reduced CD8+ cells and exhaustion of the virus-specific T cell response to HBV infection. Ultimately, HBV-HLA-specific cytolytic T cell responses of the adaptive immune system are felt to be responsible for recovery from HBV infection.


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InternalMed_Harrison. Debate continues over the relative importance of viral and host factors in the pathogenesis of HBV-associated liver injury and its outcome. As noted above, precore genetic mutants of HBV have been associated with the more severe outcomes of HBV infection (severe chronic and fulminant hepatitis), suggesting that, under certain circumstances, relative pathogenicity is a property of the virus, not the host. The fact that concomitant HDV and HBV infections are associated with more severe liver injury than HBV infection alone and the fact that cells transfected in vitro with the gene for HDV antigen express HDV antigen and then become necrotic in the absence of any immunologic influences are also consistent with a viral effect on pathogenicity. Similarly, in patients who undergo liver transplantation for end-stage chronic hepatitis B, occasionally, rapidly progressive liver injury appears in the new liver. This clinical pattern is associated with an unusual histologic pattern in the new


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InternalMed_Harrison. for end-stage chronic hepatitis B, occasionally, rapidly progressive liver injury appears in the new liver. This clinical pattern is associated with an unusual histologic pattern in the new liver, fibrosing cholestatic hepatitis, which, ultrastructurally, appears to represent a choking of the cell with overwhelming quantities of HBsAg. This observation suggests that, under the influence of the potent immunosuppressive agents required to prevent allograft rejection, HBV may have a direct cytopathic effect on liver cells, independent of the immune system.


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InternalMed_Harrison. Although the precise mechanism of liver injury in HBV infection remains elusive, studies of nucleocapsid proteins have shed light on the profound immunologic tolerance to HBV of babies born to mothers with highly replicative (HBeAg-positive), chronic HBV infection. In HBeAg-expressing transgenic mice, in utero exposure to HBeAg, which is sufficiently small to traverse the placenta, induces T cell tolerance to both nucleocapsid proteins. This, in turn, may explain why, when infection occurs so early in life, immunologic clearance does not occur, and protracted, lifelong infection ensues.


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InternalMed_Harrison. An important distinction should be drawn between HBV infection acquired at birth, common in endemic areas, such as East Asia, and infection acquired in adulthood, common in the West. Infection in the neonatal period is associated with the acquisition of high-level immunologic tolerance to HBV and absence of an acute hepatitis illness, but the almost invariable establishment of chronic, often lifelong infection. Neonatally acquired HBV infection can culminate decades later in cirrhosis and hepatocellular carcinoma (see “Complications and Sequelae”). In contrast, when HBV infection is acquired during adolescence or early adulthood, the host immune response to HBV-infected hepatocytes tends to be robust, an acute hepatitis-like illness is the rule, and failure to recover is the exception. After adulthood-acquired infection, chronicity is uncommon, and the risk of hepatocellular carcinoma is very low. Based on these observations, some authorities categorize HBV infection into an


InternalMed_Harrison

InternalMed_Harrison. After adulthood-acquired infection, chronicity is uncommon, and the risk of hepatocellular carcinoma is very low. Based on these observations, some authorities categorize HBV infection into an “immunotolerant” phase, an “immunoreactive” phase, and an “inactive” phase. This somewhat simplistic formulation does not apply at all to the typical adult in the West with self-limited acute hepatitis B, in whom no period of immunologic tolerance occurs. Even among those with neonatally acquired HBV infection, in whom immunologic tolerance is established definitively, intermittent bursts of hepatic necroinflammatory activity punctuate the early decades of life during which liver injury appears to be quiescent (labeled by some as the “immunotolerant” phase). In addition, even when clinically apparent liver injury and progressive fibrosis emerge during later decades (the so-called immunoreactive, or immunointolerant, phase), the level of immunologic tolerance to HBV remains substantial. More