Hepatitis Type A and Comparison with

Transcription

1 GASTROENTEROLOGY 1982;82: Light Microscopic Findings of Liver Biopsy Specimens from Patients with Hepatitis Type A and Comparison with Type B HIROHIKO ABE, PAUL R. BENINGER, NAOKI IKEJIRI, HIROSHI SETOY AMA, MICHIO SAT A, and KYUICHI T ANIKA W A Second Department of Internal Medicine, Kurume University School of Medicine, Kurl.!me, Japan We campared the light microscapic features af liver biapsy specimens taken within 15 days af anset af symptams from 17 patients with sero1agically verified hepatitis type A and 10 patients. with sero1agi-. cally verified hepatitis type B. On admissian, patients with hepatitis type A experienced higher serum transaminase cancentratians and 1awer tata1 serum bilirubin cancentratians than patients with type B. On examinatian af H &- E preparatians, specimens fram bath type A and type B displayed simultaneaus hepatacellu1ar degeneratian, parenchymal and parta1 tract inflammatary cell infi1tratian, and sinusaida1lining cell activatian. Hepatitis type A was characterized by conspicuaus, mananuclear inflammatary cell infi1tratian af the parta1 tract with frequent disruptian af the limiting plate, and periparta1 hepp.tacyte faca1 necrosis with virtual sparing of parenchyma abaut the central vein tributary. Specimens from 10 patients also displayed mild chalestasis. In cantrast, hepatitis type B was characterized by madest portal tract infi1tratian, and extensive parenchymal changes and infiltration, particularly abaut the central vein tributary. Viral hepatitis has became the facus af renewed interest with the advent af seralagic markers far hepatitis type B (1) and mare recently far hepatitis type A (2). This has enabled warkers to. more clearly identify marphalagic lesions with their respective viral etialagies, and has encauraged wark with primates to. determine pathaphysialagic mechanisms. The marmaset became the first successful animal madel far hepatitis type A. Deinhardt et al. described facal hepatacyte necrasis and severe partal tract mananuclear cell inflammatian (3). Recently, Krawczynski et al. further described parenchymal necrotic and degenerative changes primarily lacalized to. peripartal tissue (4). The chimpanzee has also. proven to. be a susceptible species far hepatitis A virus (HA V) infectian. Dienstag et al. nated a facal hepatacyte necrasis that develaped mainly in peripartal areas and a predaminant, mananuclear inflammatary cell infiltrate af the portal tract (5). The precise nature af the marphalagic lesian af HA V in the human is nat fully established. Experiments dane with Jaliet prisan inmates in 1970 revealed a mainly partal distributian af hepatitis (6). Recently, investigatian af a natural autbrellk in Japan reve&led partal tract mananuclear cell inflammatian and peripartal hepatacyte facal necrasis in liver biapsy specimens af children (8,9). This report describes distinctly different patterp.s af marphalagic lesians in human liver biapsy tissue from adult patients with seralagically canfirmed hepatitis type A and type B. Canspicuaus partal inflammatian and peripartal hepatacyte alteratians characterize type A, while marked, centrilabular parenchymal changes characterize type B. Received September 4, Accepted January 8, AddrE)ss requests for reprints to: Hirohiko Abe, M.D., Second Department of Internal Medicine, Kurume University School of Medicine, Asahi-machi, 64, Kurume City, 830 Japan. Dr. Beninger is presently at the Department of Internal.Medicine, University of California, Davis, Medical Center, Sacramento, California by the American Gastroenterological Association /82/ $02.50 Materials and Methods Patients Twenty-seven patients with prodromal symptoms su~estive of hepatitis, such as fever, abdominal pain, and diarrhea, were admitted to Kurume University hospital for evaluation and supportive care. Evaluation included total

2 May 1982 HISTOLOGY OF HEPATITIS TYPE A 93S bilirubin, serum glutamic oxaloacetic transaminase (SGOT), serum glutamic pyruvic transaminase (SGPT), lactic dehydrogenase (LDH), alkaline phosphatase (AP), thymol turbidity test (TTT), and serology. We employed radioimmunoassay (RIA) to detect immunoglobulin M hepatitis A virus antibody (IgM anti-ha V) (HA V AB-M kit, Abbott Laboratories, North Chicago, Ill.) for the diagnosis of hepatitis type A and reverse passive hemagglutination (RPHA) to detect hepatitis B surface antigen (HBsAg) (Auscell kit, Abbott Laboratories), and RIA to detect hepatitis B core antibody (anti-hbe) (CORAB kit, Abbott Laboratories) for the diagnosis of hepatitis type B. Patients with positive serology for hepatitis type B showed a return to undetectable levels of HBsAg during convalescence. Liver biopsy specimens were obtained from all 27 patients within 15 days of onset of symptoms. Biopsy specimens were fixed in 10% neutral buffered formalin, embedded in paraffin, and stained with H & E. Pathology Evaluation Procedure Liver biopsy findings of each patient were rated independently by three of the authors (H.A., H.S., and N.L). The following parameters were evaluated in a semiquantitative manner: inflammatory cell infiltration at the portal tract, necrosis (each at the periportal, intermediate, and central zone), and activation of sinusoidal lining cells (SLC). Most categories were rated from 0 to 3 +: (0) absent, (1 +) minimally present, (2+) present in moderate quantity, and (3 +) present in large quantity. Results Patient data base is summarized in Table 1 and presented in detail in Tables 2 and 3. The mean age of patients with hepatitis type A was 30 yr, with a range of yr. The average age of patients with hepatitis type B was 25 yr, with a range of yr. The male/female ratio was 10:7 for type A and 9:1 for type B. The mean length of time from onset of symptoms to liver biopsy was 12 days, with a range of 5-15 days for type A and 12 days, with a range of 6-15 days for type B. No patient had a prior history of liver disease. Only 1 patient, from the group of patients with hepatitis type A, was noted to be an HBsAg carrier. Serum glutamic oxaloacetic transaminase, SGPT, LDH, and TTT levels were generally higher in patients with hepatitis type A than in patients with type B. Total bilirubin and alkaline phosphatase were generally lower in patients with type A than with type B. Immunoglobulin M anti-hay was positive in all patients with type A and negative in all patients with type B. Hepatitis B surface antigen was positive in all patients with type B and in 1 patient with type A. This patient also demonstrated an initial, high-titer anti-hb e, consistent with a diagnosis of HBsAg carrier state. Hepatitis B surface antigen returned to Table 1. Summary of Patient Data Base Viral Viral hepatitis hepatitis Diagnosis type A type B no Age (yr) Mean ± SD 29.8 ± ± 5.7 Range MalelFemale /1 Day of liver biopsyb Mean ± SD 11.7 ± ± 2.7 Range Liver function tests Total bilirubin (mg/dl) 5.4 ± ± 7.5 SGOT (Karmen units, n < 40) 1262 ± ± 339 SGPT (Karmen units, n < 35) 1422 ± ± 535 Alkaline phosphatase 16.5 ± ± 2.3 (King-Armstrong units, n < 10) TIT (Kunkel units, n < 5) 6.2 ± ± 3.3 o n = Number of cases. b After onset of symptoms. undetectable levels in all patients with type B during convalescence but remained positive in the 1 patient with type A. Histologic features are summarized in Tables 4 and 5. The hallmark of viral hepatitis is simultaneous hepatocellular degeneration, parenchymal and portal tract inflammatory cell infiltration, and sinusoidal lining cell activation. Hepatitis type A is characterized by conspicuous portal tract inflammation, involvement of surrounding parenchyma (corresponding to zone 1 of the acinus of Rappaport), and minimal involvement of the central area (Figures la and 2A). Portal inflammation is usually severe and often associated with proliferation of bile ductules amidst an inflammatory exudate (Figure IB). Portal tracts are swollen with an almost exclusively mononuclear cell infiltrate, including plasma cells. The infiltrate frequently disrupts the limiting plate and sometimes reaches out to neighboring portal tracts. The periportal zone shows conspicuous hepatocellular alterations in hepatitis type A (Figures IB and 2B). There is marked anisocytosis, variation in cytoplasmic staining quality and nuclear changes. Ballooning degeneration is marked by sparse, granular cytoplasm, an often twofold or greater increase in cellular diameter, and frequently an increase in nuclear diameter. Other hepatocytes undergo acidophilic degeneration, manifested by cellular shrinkage with deepening of cytoplasmic staining and nuclear pyknosis. Cellular changes are usually found in a focal distribution, though certain areas of biopsy

3 940 ABE ET AL. GASTROENTEROLOGY Vol. 82, No.5, Part 1 Table 2. Laboratory Data on Admission of Patients with Hepatitis Type A Case No. Age (yr) Sex Clin day TB (mg/dl) SGOT (KU) SGPT (KU) TTT (KuU) 1 23 M M F M M M M F F F M F M F M M M NOTE. Clin day: day of biopsy after onset of clinical symptoms; TB: total bilirubin; SGOT: serum glutamic oxaloacetic transaminase; SGPT: serum glutamic pyruvic transaminase; KU: Karmen units; TTT: thymol turbidity test; KuU: Kunkel units. specimens show more severe alterations than other areas in the same specimens. Lymphocytes are sometimes seen in close proximity to altered hepatocytes (Figure IB). Regenerative hepatocellular changes are also seen, notably mitoses and binucleation. Sinusoidallining cell activation is in evidence; SLCs are increased in number and appear somewhat increased in size. The extent of change is demonstrably less in the intermediate zone and minimal in the central zone about the hepatic vein or venule. Ballooning degeneration and acidophilic degeneration are uncommon in the central zone, though slight variation in cytoplasmic staining and small variability in cellular and nuclear size are encountered. There is evidence of SLC activation, though less than in the periportal area. There is little or no inflammatory cell infiltration. There are no differences seen for case 10 (Table 2), a patient who experienced hepatitis type A and was also shown to be an HBsAg carrier. The geographic distribution of periportal necrosis and minimal change about the central vein tributary is superimposed upon a background of "ground-glass" hepatocytes (Figure 3). Mild cholestasis is present in 10 of 17 cases with hepatitis type A. Its presence does not correlate with the degree of hyperbilirubinemia or clinical jaundice. In case 11 (Table 2). however, cholestasis is pronounced and is associated with periportal hepatic cell necrosis (Figure 4). Approximately 4 wk after onset of symptoms, this patient underwent a second liver biopsy which demonstrated cholestasis without periportal hepatocyte necrosis. Hepatitis type B is characterized by modest portal tract inflammation and extensive parenchymal changes, particularly about the central venule (Figure 5A). Portal tract mononuclear cell infiltration is less severe than in type A. Inflammatory cells are also seen sporadically throughout the parenchyma. Table 3. Laboratory Data on Admission of Patients with Hepatitis Type B Case No. Age (yr) Sex Clin day TB (mg/dl) SGOT (KU) SGPT (KU) TTT (KuU) 1 27 F M M M M M M M M M NOTE: Clin day: day of biopsy after onset of clinical symptoms; TB: total bilirubin; SGOT: serum glutamic oxaloacetic transaminase; SGPT: serum glutamic pyruvic transaminase; l(u: Karmen units; TTT: thymol turbidity test; KuU: Kunkel units.

4 May 1982 HISTOLOGY OF HEPATITIS TYPE A 941 Table 4. Liver Biopsy Findings in 17 Patients with Hepatitis Type A Hepatic cell necrosis Case No. Periportal Intermediate Central Inflammatory cell infiltration at the portal tract Activation of sinusoidal cells Hepatocellular alterations are comparable to those seen for type A: anisocytosis, variation in cytoplasmic staining, and nuclear changes. These parenchymal changes are more diffusely distributed throughout the lobule than is the case for type A; however, the more severe lesions of focal necrosis are found about hepatic vein tributaries (Figure 5B). Regenerative changes are also seen. Sinusoidal lining cell activation is more conspicuous in type B than in type A. None of the 17 patients with hepatitis type A and the 10 patients with hepatitis type B exhibited histologic evidence of fibrosis. Discussion Hepatitis type A and hepatitis type B demonstrate significant morphologic differences in liver biopsy specimens taken within 15 days of onset of symptoms. In hepatitis type A, hepatocyte alter- ations, particularly hepatocellular necrosis, are primarily restricted to the periportal area, parenchymal changes are less severe than portal inflammation, and SLC activity is modestly increased. In hepatitis type B, however, hepatocyte alterations are seen predominantly in the centrilobular area, and parenchymal changes are more remarkable than in hepatitis type A. The timing of the biopsy proved to be of importance in detailing the morphologic alterations. This is best exemplified by case 11 (Table 2), for which a biopsy specimen taken within 15 days of onset of symptoms reveals cholestasis and periportal hepatocellular alterations with minimal centrilobular changes, but for which a repeat biopsy specimen taken at 30 days demonstrates only findings of cholestasis. Case 10 (Table 2) afforded an uncommon opportunity to investigate simultaneous HA V and hepatitis B virus (HBV) infections. Many hepatocytes show Table 5. Liver Biopsy Findings in 10 Patients with Hepatitis Type B Hepatic cell necrosis Case No. Periportal Intermediate Central Inflammatory cell infiltration at the portal tract Activation of sinusoidal cells

5 942 ABE ET AL. GASTROENTEROLOGY Vol. 82, No.5, Part 1 Figure 1. H & E preparation of a liver biopsy specimen taken on the 11th day after clinical onset from patient No. 12 with hepatitis type A. A. Note the marked portal tract mononuclear cell infiltration with disruption of the limiting plate, the bile ductule proliferation, the periportal hepatocyte ballooning and acidophilic degeneration, and the sinusoidal lining cell activation (x 50). B. Note the detail of periportal hepatocyte ballooning and acidophilic degeneration (x 200).

6 May 1982 HISTOLOGY OF HEPATITIS TYPE A '.-" ~:~ -:=:. -...".... ~... :... -:..:.... o :"".... "' ':. o... -, : ! '\~... ~.. : '. -',..,.., ~ t... ':. ' "......' Figure 2. H & E preparation of a liver biopsy specimen taken on the 13th day after clinical onset from patient No. 9 with hepatitis type A. A. Note the moderate portal tract infiltration with disruption of the limiting plate and the spotty periportal hepatocyte necrosis with virtual sparing of parenchyma about the central venule (x 50). B. Note the detail of dense portal tract infiltrate with disruption of the limiting plate, and the spotty hepatocyte necrosis (x 100).

7 944 GASTROENTEROLOGY Vol. 82, No.5, Part 1 ABE ET AL... ' 'f... ; :t. '- «.. 11' Figure 3. H & E preparation of a liver biopsy specimen taken on the fifth day after clinical onset from patient No. 10 with hepatitis type A and HB.Ag carrier state. Note the numerous "ground-glass" hepatocytes (arrow) (x 50). Figure 4. H & E preparation of a liver biopsy specimen taken on the 14th day after clinical onset from patient No. 11 with hepatitis type A. Note the conspicuous bile plugging at the centrilobular area (x 50).

8 May 1982 HISTOLOGY OF HEPATITIS TYPE A 945 Figure 5. H & E preparation of a liver biopsy specimen taken on the 15th day after clinical onset from patient No.3 with hepatitis type B. A. Note the modest portal tract mononuclear cell infiltrate, the extensive parenchym'al changes, particularly about the central venule, and the marked infiltration about the central venule (x 50). B. Note the hepatocyte ballooning and acidophilic degeneration, the marked infiltration about the central venule, and the sinusoidal lining cell activation (x 100).

9 946 ABE ET AL. GASTROENTEROLOGY Vol. 82, No.5, Part 1 the characteristic ground-glass appearance of the HBsAg carrier state; there is also evidence of periportal focal necrosis with minimal centrilobular changes. Since periportal necrosis is not known to occur in patients who are only HBsAg carriers, these features cannot be attributed to HBV or its antigen in this case, but can be better explained by the concurrent HA V infection. Morphologic descriptions of liver tissue from marmosets inoculated orally with HA V have been recently provided by Krawczynski et a1. (4). They have shown a predominantly periportal localization of injury. In addition, they have demonstrat~d the presence of hepatitis A antigen (HAAg) in the cytoplasm of hepatocytes adjacent to portal tracts. In chimpanzees inoculated either orally or percutaneously with HAV, Dienstag et a1. and Popper et a1. have shown that extensive destruction of hepatocytes characteristically takes place in the periportal zone, almost sparing the hepatic vein tributaries and surrounding tissue (5,10). Our findings are consistent with these subhuman primate studies. In their hepatitis type B studies, Popper et a1. have described hepatocyte focal necrosis about hepatic vein tributaries with activation of SLCs, as well as dense inflammatory infiltration of portal tracts (10). Our findings parallel this description. Further support for the proposition that there are two distinct patterns of hepatic lesions comes from Drucker et a1. in their report on simultaneous HA V and HBV infections in a chimpanzee (11). They believe the apparent absence of viral interference and the occurrence of two separate rises in transaminase levels speak for the presence of two separately affected populations of hepatocytes. In the classic 1954 paper on the biopsy diagnosis of acute hepatitis in humans, Smetana described focal hepatocyte necrosis and a dense mononuclear cell infiltration of the portal tract (12). Conrad et ai., in 1964, similarly reported a portal mononuclear cell infiltrate with foci of hepatocyte necrosis in their study of infectious hepatitis (13). Ishak, in his 1976 description of viral hepatitis, emphasized the similarities in morphologic changes, whether caused by type A or type B (14). He noted that lobular disarray ami activitation of SLCs characterized acute viral hepatitis. Boggs et a1. (6) provided the first description of a mainly portal distribution of hepatitis, resulting from or'!.l inoculation with MS-l serum (7) which eventually proved to contain HAV (2). Age may influence the clinical, laboratory, and histologic course of hepatitis type A. In comparing laboratory data from our patients with those of children involved in an outbreak of serologically verified hepatitis type A in an elementary school in Japan (8,9), serum bilirubin and transaminase levels are notably higher in adults. Comparing histologic descriptions, almost all of the biopsy specimens in our series demonstrate less severe portal inflammation. Except for a few adult cases that demonstrate conspicuous bile stasis, morphology is comparable, particularly the periportal hepatocyte alterations. It remains a matter of conjecture whether these morphologic differences reflect differences in pathophysiologic mechanisms. Hepatitis A antigen has consistently been detected in marmoset (4,15,16) and chimpanzee (17,18) hepatic tissue, implicating the liver as the sole site of HA V replication. Further, Krawczynski et a1. found no deposits of IgG, IgM, IgA, Clq, or C3 in any of their marmoset liver specimens, interpreting this to mean that hepatic parenchymal lesions are a direct result of viral replication (4). While the geographic location of parenchyillal lesions with regard to portal blood flow suggests enteral passage of the virus and primary contact with periportal hepatocytes, as proposed by Krawczynski et ai., two lines of evidence go against a direct, cytopathic role for HAV. First, Provost and Hilleman reported no cytopathic changes in their marmoset liver cell tissue used to culture HAV (19), suggesting that the virus itself is incapable of the necrotic changes seen in situ. Second, Popper et a1. have demonstrated lymphocytes in close proximity to hepatocytes, particularly altered hepatocytes (10). In addition, we and others (9,10) have shown that SLCs, presumably Kupffer cells, enlarge and increase in number. Furthermore, HAAg virus like particles have been clearly demonstrated in Kupffer cells by Shimizu et al. (20) and Huang et a1. (21), despite the absence of similar findings by Krawczynski and colleagues. Together, these points suggest a role for cell-mediated immunity; A cell-mediated immune response has also been proposed for hepatitis type B (22). However, the morphologic lesion is postulated to result from cellular attack on surface antigens of HBV expressed on the hepatocellular membrane surface. The relationship to the morphologic distribution of the lesions is not clearly understood. In conclusion, hepatitis type A is associated with specific morphologic features, namely conspicuous portal inflammation and periportal hepatocyte alterations. In contrast, hepatitis type B is characterized by centrilobular changes and less conspicuous portal inflammation. References 1. Blumberg as, Alter HJ, Visnich S. A "new" antigen in leukemia sera. JAMA 1965;191: Feinstone SM, Kapikian AZ, Purcell RH. Hepatitis A: detec-

10 May 1982 HISTOLOGY OF HEPATITIS TYPE A 947 tion by immune electron microscopy of a virus-like antigen associated with acute illness. Science 1973;182: Deinhardi F, Holmes AW, Capps RB, Popper H. Studies on the transmission of human viral hepatitis to marmoset monkeys. I. Transmission of disease, serial passages, and description of liver lesions. J Exp Med 1967;125: Krawczynski KK, Bradley DW, Murphy BL, et a1. Pathogenetic aspects of hepatitis A virus infection in enterally inoculated marrd.osets. Am J Clin Pathol 1981;76: Dienstag JL, Feinstone SM, Purcell RH, et a1. Experimental infection of chimpanzees with hepatitis A virus. J Infect Dis 1975;132: Boggs JD, MelnickJL, Conrad MD, Felsher BF. Viral hepatitis: clinical and tissue culture studies. JAMA 1970;214: Krugman S, Giles JP, Hammond J. Infectious hepatitis: evidence for two distinctive clinical, epidemiological and immunological types of infection. JAMA 1967;200: Tanikawa K. Acute viral hepatitis. Type A hepatitis. Its epidemioiogy, clinical picture and pathologic changes of the liver. Gastroenterol Jpn 1979;14: Abe H, Ikejiri N, Sata M, et a1. Histological findings of the liver in viral hepatitis type A. A comparison with viral hepatitis type B. Acta Hepatol Jpn 1981;22:22-3i. 10. Popper H, Dienstag JL, Feinstone SM, et a!. Pathology of viral hepatitis in chimpanzees. Virchows Arch A Pathol Anat HistoI1980;387: Drucker J, Tabor E, Gerety RJ, et a1. Simultaneous infections with hepatitis A and B viruses in a chimpanzee. J Infect Dis 1979;139: Smetana HF. The histologic diagnosis of viral hepatitis by needle biopsy. Gastroenterology 1954;26: Conrad ME, Schwartz FD, Young AA. Infectious hepatitis-a generalized disease. A study of renal, gastrointestinal and hematologic abnormalities. Am J Med 1964;37: Ishak KG. Light microscopic morphology of viral hepatitis. Am J Clin PathoI1976;65: Mathiesen LR, Drucker J, Lorenz D, et a1. Localization of hepatitis A antigen in marmoset organs during acute infection with hepatitis A virus. J Infect Dis 1978;138: Mathiesen LR, Meller AM, Purcell RH, et a1. Hepatitis A virus in the liver and intestine of marmosets after oral inoculation. Infect Immun 1980;28: Schulman AN, Dienstag JL, Jackson DR, et a1. Hepatitis A antigen particles in liver, bile, and stool of chimpanzees. J Infect Dis 1976;18: Mathiesen LR, Feinstone SM, Purcell RH, Wagner JA. Detection of hepatitis A antigen by immunofluorescence. Infect Immun 1977;18: Provost PJ, Hilleman MR. Propagation of human hepatitis A virus in cell culture in vitro. Proc Soc Exp Bioi Med 1979;160: Shimizu YK, Mathiesen LR, Lorenz D, et a1. Localization of hepatitis A antigen in liver tissue by peroxidase-conjugated antibody method: light and electron microscopic studies. J ImmunoI1978;121: Huang S, Lorenz D, Gerety RJ. Electron and immunoelectron microscopic study on liver tissues of marmosets infected with hepatitis A virus. Lab Invest 1979;41: Edgington TS, Chisari FV. Immunological aspects of hepatitis B virus infection. Am J Med Sci 1975;270: