Occurrence of Australia Antigen in Chronic Hepatitis in Italy

Vol. 63, No. 3 Printed in U.S.A.

GA~TftOENTEitO I.OC. y © 1972 hy· Thc Willinms & Wilk ins Co.

C opy ri~h t

OCCURRENCE OF AUSTRALIA ANTIGEN IN ClffiONIC HEPATITIS IN ITALY P. BIANCHI, BLANCO,

c.

U.

c.

SPANO, AND

L. DARDANONI, C. DEL VECCHIO R. NACCARATO, L. PAGLIARO,

BIANCHI PORRO, M . COLTORTI,

FAGIOLO,

G.

R.

FARINI,

l.

MENOZZI,

VERME

Istituto di Patologia Medica I, Milan ; Istituto di Clinica Medica I, Naples ; Istituto di Igiene, Palermo; Istituto di Patologia M edica , Padova; Istituto di Patologia Medica, Palermo; and Divisione di Gastroenterologia, Ospedale S . Giovanni Battista, Torino, Italy

This paper deals with a cooperative study, fostered by Associazione Italiana per lo Studio del Fegato, concerning the distribution of Australia antigen in patients with chronic aggressive hepatitis, chronic persistent hepatitis, and primary biliary cirrhosis in Italy.

Patients and Methods Fifty-nine patients with chronic aggressive hepatitis (CAH), 20 patients with chronic persistent hepatitis (CPH) , and 5 patients with primary biliary cirrhosis were studied. The patients were observed in one of the following clinical institutes : Istituto di Patologia Medica I, Universita di Milano; Istituto di Clinica Medica I, Universita di Napoli; Istituto di Patologia Medica, Universita di Padova; or Istituto di Patologia Medica, Universita di Palermo. The diagnosis fulfilled the generally accepted clinical and humoral criteria 1 • 2 ; in all cases it was ultimately confirmed by histological examination of one or more liver biopsies. The histological examination of liver biopsies was carried out by the pathologist of the institute where the patients had been observed. Furthermore, the liver biopsies of 43 patients with CAH and of 5 patients with CPH were sent for a blind review and detailed histological analysis to an independent pathologist. Other cases with conflicting or uncertain histological diagnoses were excluded. One or more specimens from each patient were assayed for antinuclear factor, mitochondrial antibody, and smooth muscle antibody at the Istituto di Patologia Medica of Padova. The technique used was that proposed by the World Health Organization (July, 1969)." Other specimens of the same sera were asReceived December 14, 1971. Accepted April 4, 1972. Address requests for reprints to: Dr. Remo Naccarato, Istituto de Patologia Medica, Padova, Italy.

sayed for Australia antigen (Au antigen) at the Istituto di Igiene of Palermo, by using double immunodiffusion (DI), quantitative ·. complement fixation (CF) , • and high voltage counterelectrophoresis (CE)!. All the assays were carried out on coded samples, including .those of control sera. Most of the specimens were tested for .autoantibodies and Au antigen in more than one laboratory, with invariably coincidental results. Samples were decoded at the end of the study.

Results Au antigen was detected in one or more specimens of 51 % of the patients observed with CAH, 25% with CPH, and 20% with primary biliary cirrhosis (table 1) . Since few cases with CPH or primary biliary cirrhosis were studied, only the results of patients with CAH are further reported. In these patients, CF titers ranged from 1:4 to 1: 1024. CF titers lower than 1: 16 were found persistently in 8 patients, only 3 of which were positive 'by the DI or the CE test. · On the other hand, DI and CE tests detected 2 Au antigen-positive cases out of 12 with persistently anticomplementary sera as compared with CF assays (table 2) _In some Au antigen-positive patients, Au antigen was detected only in a few of the many specimens assayed. In most patients; Au anti-

482

TABLE

483

LIVER PHYSIOLOGY AND DISEASE

September 1972

1. Australia antigen (Au antigen) in patients with chronic aggressiv e hepatitis (CA H) , chronic persistent hepatitis (CPH), (md primary biliary cirrhosis (PBC) CA H

PHC

CPH

Hosp it al

Milano .. .. Padova .. .. .. Napoli .. .. .. . Palermo ...... Total ...... . TABLE

No. of cases

Au ailt igen +

No. of cases

8 22 8 21 59

4 9 7 10 30 (51 %)

2 5 13 30

2. Th e relationship of the complement fixation

(CF) test. to the doubl e immunodiffusion (Dl) and counterelectrophoresis (CE) tests in patients with

Au antigen +

No. of cases

2 0 3 5 (25%)

Au antigen

1

3

1

2 5

0 1 (20%)

3. Clinical features in 30 Australia antigen (Au antigen)-positive and 29 Au antigen-negative patients with chronic aggressive h epatitis

TABLE

chronic aggressive hepatitis (CA H) positive

Au ant igennegat ive

25 5 34.6 ±2.86 14

15 14 43.2 ±2.70 3

0 .01 "

19 11

5 24

0.01"

9 18 6

14 15 13

NS•· ' NS"· '" NS•· '

Au antigen-

No. of cases Au antigen-

No. of cases Au anti- 28 gen-positive as detected by C F test Peak CF titers < 1:32 8 1 :32-1:128 17 > 1 : 128 3 No. of cases Au anti- 20 gen-negative as detected by .CF test No. of cases with per- 12 sistently anticomplementary _sera

NO.

positive as

or cases Au antigenposit ive as

detected by th e Dltest ·

detected by t he CE test

19

22

13 .3 0

2

3 14 3 0

2

gen was detected . in all serum samples serially obtained, whereas in some cases, particularly of long-standing, severe CAH, only a few of many repeated samples gave low titer positive results; the . others were anticomplementary or, rarely; negative. In 2 Au antigen-positive and in 2 Au antigennegative patients, the Au antigen antibody was detected both by CE and .CF tests in repeated assays. In CAH, Au antigen was found to be significantly more frequent iri younger males, with acute onset of disease and history of exposure to hepatitis virus(es), occurring usually by parenteral route. No other clinical or laboratory-significant differences were found between Au antigen-positive and Au antigen-negative cases. However, the Au antigen-negative patients showed

Sex Mal es .. ....... Females . Age (mean ± so) . Exposure history . Onset of disease . Acute . Insid ious ...... Extrahepatic systemic pathology . .. J aundi ce Ascites, edema .

p

0 .02" o.o5•

• According to Fisher. • According to Student. ' NS, statistically not significant.

some trend to exhibit fluid retention, extrahepatic systemic pathology, mitochondrial antibody, and slightly higher r-globulin values more frequently. These differences, however, were not statistically significant (tables 3 and 4) . Blind histological analysis of 43 liver biopsies showed, in the Au antigen-negative patients with CAH, a higher incidence of cirrhotic transformation, fibrosis, liver cell damage, inflammatory infiltration, and ductular proliferation (table 5). Within the group of the Au antigen-positive cases, cirrhosis and severe liver changes were usually associated with lower CF titers (table 6). Both differences were statistically significant.

484 TARLE 4.

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LIVER PHYSIOLOGY AND DISEASE

Laboratory data in 30 Australia antigen (Au antigen)-positive and 29 Au antigen-negative patients with chronic aggressive hepatitis" Au antige npositi ve

Serum glutamic oxa!oacetic transaminase, miU" (mean ± so) . . . . . . . . . . . . . . . . . . . Serum glutamic pyruvic transaminase, miU (mean ± so) .... -y-globulin, g/100 ml (mean ±so) .. .. ' . Antinuclear factor-positive cases . . . . . . . . . . . Mitochondrial antibody-positive cases . Smooth muscle antibody-positive cases ........

..

....

Au a nti ge nnegative

p

o .o5c

80.0 ± 10.8

47.5

70.1 ± 8.9 2.46 ± 0.10 5 18 27

39 .3 ± 7.7 2 . 74 ± .14 7 24 28

± 9.5

o . o5c NSc.• NS•· • NS•· • NS•· •

No data obtained during steroid or immunosuppressive treatment are included. • miU, milli-international unit. c Student's t-test. • NS, statistically not significant. • x' according to Fisher. a

5. Histological findings in liver biopsies from 24 Australia antigen (Au antigen)-positive and 19 Au antigen-negative patients with chronic aggressive hepatitis"

TABLE

Au antigenpositi ve pati ents

Liver architecture Preserved• Cirrhotic Fibrosis +or ++ +++ or ++++ Parenchymal damage + ++ Mononuclear infiltration (parenchymal areas) + ++ Mononuclear infiltration (fibrotic areas) + or ++ +++ or ++++ Ductular proliferation ± or + ++

6. Relationship between complement fixation (CF) peak titers and histological hepatic changes in Australia antigen-positive patients with chronic aggressive hepatitis"

TABLE

Au antigennegative patients

CF peak titers I: 4- 1: 16 (10 patients)

12 12

4 15

15 9

7 12

13 11

7 12

14 10

9 10

14 10

10 9

19 5

13 6

" According to the Cochran test, the statistical analysis was positive to 1%. • Lobular arrangement was usually disturbed but still recognizable.

Discussion The finding of the Au antigen in as many as 50% of patients with CAH is partially due to the sensitivity of the CF test and

Liver architecture Preserved• Cirrhotic Fibrosis + or+ + +++ or ++++ Parechymal damage + ++ Mononuclear infiltration (parenchymal areas) + ++ Mononuclear infiltration (fibrotic areas) + or ++ +++ or ++++ Ductular proliferation ±or + ++

I ::12- 1:1024 ( 14 patient s)

3 7

9 5

5 5

10 4

5 5

8 6

6 4

8 6

5 5

9 5

6 4

13

a According to the Cochran test, the statistical analysis was positive to 1%. • Lobular arrangement was usually disturbed but still recogni zable.

to the serial repetition of the assays, as previously mentioned. However, lower figures have been reported by others, 6 • 7 who also used the CF test in repeated

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LIVER PHYSIOLOGY AND DISEASE

485

assays. Our results confirm that a high It is our opinion that the Au antigenfrequency of Au antigen associated with negative cases, the low titer Au antigenCAH does exist in Italy, as suggested by positive cases, and the high titer Au antiprevious studies. 4 • 8 gen-positive cases are but a manifestation The clinical and laboratory differences of a continuous spectrum of a single Au between Au antigen-positive and Au an- antigen-associated disease, the different tigen-negative patients with CAH are forms being a reflection of the various less impressive than those found in other phases of the disease or a variation of the studies. Our results do not confirm the · immunological response. mutual exclusion between the presence of REFERENCES Au antigen and smooth muscle antibody or 1. DeGroote J, Desmet VJ, Gedigk P , et al: A the significantly higher prevalence of sysclassification of chronic hepatitis. Lancet 2:626temic pathology that has been reported 628, 1968 2. Mistilis SP, Blackburn CR: Active chronic hepby others in Au antigen-negative paatitis. Am J Med 48:484- 495, 1970 tients. 6 • 9 - 11 These conflicting results may 3. Bull World Health Organization, July 1969, be partially due to the fa'ct that in some PA/66.272 previous studies the DI test, which fails to detect low titers of Au antigen, was 4. Dardanoni L, Pagliaro L, Spano C: Complement fixation test in the study of Australia antigen. used. 9 - 1 1 In fact, our findings are in agreeArch Gesamte Virusforsch, 34:51, 1971 ment with the observations of Vernace 5. Prince AM, Burke K: Serum hepatitis antigen 12 et al., who also used the CF test, and (SH): rapid detection by high voltage immunoalso with those of Grob et al., 1 3 who used electroos mophoresis. Science 169:563-565, 1970 immunoelectrophoresis. 6. Bulkley BH, Goldfinger SE, Heizer WD, et a!: Distinctions in chronic active hepatitis based on As previously reported by others, 14 we circulating hepatitis-associated antigen . Lancet also observed more severe histological le2:1323- 1326, 1970 sions of the liver, and more frequent evi7. Kaplan MM, Grady G: Serum-hepatitis antigen dence of cirrhotic architecture, in Au anin chronic hepatitis and primary biliary cirrhosis. tigen-negative patients. Furthermore, we Lancet 1:159- 161, 1971 found that, within the group of the Au 8. Naccarato R, Fagiolo U, Farini R, et al: antigen-positive cases, severe liver damage L'antigene Australia in alcune epatopatie acute and cirrhosis were usually associated with e cronische. G Clin Med 50:170-179, 1969 lower CF titers of Au antigen. This result 9. Wright R : Australia antigen and smooth-muscle is similar to Dudley et al.'s 15 observations antibody in acute and chronic hepatitis. Lancet 1:521-522, 1970 concerning the finding of higher titers of Au antigen in persistent, rather than in 10. Sherlock S, Fox RA, Niazi SP, et al: Chronic liver disease and primary liver-cell cancer with aggressive, chronic hepatitis. hepatitis-associated (Australia) antigen in serum . Our results indicate that the use of a 1:1243- 1247, 1970 sensitive method combined with serial 11. Lancet Vischer TL: Australia antigen and autoantibodies assays enabled us to detect the Au antigen in chronic hepatitis. Br Med J 2:695-698, 1970 in 50% of the patients affected by CAH. 12. Vernace SJ, Paronetto F, Schaffner F: Immune No important differences, serological or reactions and Australia (HAA) antigen in chronic histological, were discovered between the aggressive hepatitis (abstr). Gastroenterology 60:182, 1971 Au antigen-positive and the Au antigen13. Grob PJ, Jemelka HJ, Muller JW: B,A and SH negative patients. antigen in chronic hepatit is. Gastroenterology The Au antigen antibody was detected 61:91-95, 1971 in 2 Au antigen-negative patients. The use of an even more sensitive 14. Sherlock S, Fox RA, Niazi SP, et al: The relationship of hepatitis-associated antigen to method might increase the number of Au chronic hepatit is. Atti Convegno Farmitalia su antigen-positive assays. However, the Au "Antigene Australia ed epatiti virali." Milan, antigen might never become detectable in Minerva Medical Publishers, 1970, p llO some patients because of the particular im- 15. Dudley FJ, Fox RA, Sherlock S: Relationship munological response or the stage of the of hepatitis-associated antigen (H. A.A.) to acute disease at that particular time. and chronic liver injury. Lancet 2:1-3, 1971