Clin. exp. Immunol. (1975) 22, 22-29.

SMOOTH-MUSCLE ANTIBODIES AND OTHER TISSUE ANTIBODIES IN CYTOMEGALOVIRUS INFECTION P. ANDERSEN AND H. K. ANDERSEN Institute of Medical Microbiology, University of Aarhus, Denmark (Received 20 December 1974) SUMMARY

Smooth-muscle antibodies (SMA) were present in 1600 of sixty-three patients with cytomegalovirus (CMV) antibodies in serum and absent in forty CMV antibodynegative blood donors (P = 0 005). The SMA were of the IgG and IgM class, while no IgA antibodies were found. In patients with CMV infection, SMA, mainly of the IgM class, were present in the early stages of the disease, and the titre decreased faster than the complement-fixing CMV antibody titre. Anti-nuclear antibodies (ANA) were also found more often in CMV antibody-positive sera than in CMV antibodynegative sera, and ANA were usually present in sera which also contained SMA. Parietal cell antibodies, mitochondrial antibodies and other cytoplasmic antibodies did not occur more frequently in CMV antibody-positive than in CMV antibodynegative sera. INTRODUCTION Cytomegalovirus (CMV) infection and infectious mononucleosis are caused by two related viruses and have many clinical and laboratory features in common. Clinical or biochemical signs of liver involvement may be present in both diseases, and serological abnormalities, such as the occurrence of antinuclear antibodies, rheumatoid factors, cold agglutinins and cryoglobulins, have been found in both CMV infection and in infectious mononucleosis (Carter, 1966; Kaplan, 1968; Kaplan & Tan, 1968; Wager et al., 1968; Kantor et al., 1970). Smooth-muscle antibodies (SMA) occur most often in patients with liver disease (Johnson, Holborow & Glynn, 1965; Farrow et al., 1970; Ajdukiewicz et al., 1972). They have also been demonstrated in patients with infectious mononucleosis and in healthy subjects, and it has been suggested that other viral infections may give rise to SMA (Ajdukiewicz et al., 1972; Holborow, Hemsted & Mead, 1973; Sutton et al., 1974). We have studied the incidence of SMA and certain other tissue antibodies in patients with CMV infection, especially in relation to the onset of a recovery from the disease. Correspondence: Dr P. Andersen, Institute of Medical Microbiology, University of Aarhus, DK-8000 Aarhus C, Denmark,

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Smooth-muscle antibodies in CMV infection

23

MATERIALS AND METHODS Patients. A total of 152 sera from 120 subjects were investigated for CMV antibodies and tissue antibodies. Based on clinical diagnoses and the findings of CMV antibodies, the subjects were divided into the following four groups: Group 1. Forty blood donors whose serum did not contain complement-fixing (CF) or neutralizing (NT) CMV antibodies. The blood donors had never been jaundiced; they were in good health and their haemoglobin concentrations were within normal limits. The group consisted of thirty-two female and eight male donors aged from 21 to 54 years. One serum sample from each of these individuals was investigated. Group 2. Sixty-three patients, half of them renal allograft recipients, whose serum had been sent to the laboratory for investigation for CMV antibodies. The diagnoses of the patients were unknown to us. One serum sample was obtained from each of these individuals, and all sera contained CF-CMV antibodies in titres between 16 and > 256. The group consisted of forty-two women and twenty-one men aged from 2 to 78 years. Group 3. Nine adult patients with CMV infection. The group consisted of two men aged 25 and 67 years, an 1 1-year-old boy and six women whose ages ranged from 26 to 65 years. The age and sex of the individual patients are shown in Table 2. CMV was also isolated from urine and/or throat swabs in patients numbers 1-3. No attempts to isolated virus were done in the remaining six patients. Heterophil antibodies were not revealed in any of the patients. Serum transaminase values were elevated in four (numbers 1-3 and 8), normal in two (numbers 5 and 6) and not determined in three patients (numbers 4, 7 and 9). Relative lymphocytosis ( > 40%) was found in four (numbers 1-3 and 8); the lymphocyte count was normal in three (numbers 5-7), and was not performed in two patients (numbers 4 and 9). No clinical or biochemical signs of renal involvement were observed in any of the patients. Group 4. Eight infants with CMV infection. CMV was isolated from the urine in all cases. The patients' ages at the time of investigation and the results of serlogical tests are shown in Table 3. Tissue antibody tests. Unfixed tissue sections of rat stomach and kidney were used as antigens for the tissue antibody tests. SMA, glomerular antibodies (GA), parietal cell antibodies (PA), cytoplasmic antibodies (CA) reacting with rat renal tubular cells, but not with rat gastric parietal cells, and mitochondrial antibodies (MTA) were demonstrated by the indirect immunofluorescent method (11F) as previously described (Andersen & Beutner, 1972; Andersen, 1974). The sera were investigated for antibodies of the IgG, IgA and IgM classes by means of monospecific fluorescein (FITC) conjugated anti-human immunoglobulins obtained from Wellcome Research Laboratories. The molar F: P ratios of the conjugates were between 2-9 and 4- 1, and the antibody content was between 2 and 8 u/ml. The conjugates were used in a dilution corresponding to an antibody content between 1/4 and 1/8 u/ml. All sera from one patient were investigated in the same experiment, and the slides were read blindly. The sera were tested in doubling dilutions, starting with 1:10. CMV antibody tests. The sera were investigated for CMV antibodies by the complement-fixation reaction and virus-neutralization test using strain Ad 169 as described earlier (Andersen, 1971). Isolation and identification of CMV from urine and throat swabs were performed as previously described (Andersen & Spencer, 1969).

RESULTS SMA and GA in sera with and without CMV antibodies SMA were found in ten out of sixty-three (16%) unselected patients with CF-CMV antibodies in serum (group 2), but could not be demonstrated in forty CMV antibody-negative blood donors (group 1). This difference is statistically significant (Fisher's exact test, P = 0 005). The SMA titres were low, i.e. 10 and 20, and the antibodies were of IgG class (five cases) and IgM class (five cases), while IgA antibodies were not found. GA in titres of 10 were found in only two sera, which also contained SMA in titres of 10 and 20. It is seen from Table 1 that SMA were found more often in sera with CF-CMV antibodies in high titres than in sera with low titres, but this correlation is not significant (P = 0 06).

P. Andersen and H. K. Andersen

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TABLE 1. The relationship between SMA and CF-CMV antibodies in 103 subjects CMV antibody-positive subjects (group 2) CF-CMV antibody titre

CMV antibody-negative subjects (group 1) No.

%

SMA-positive

0

0

SMA-negative

40

100

32

64

128

No. %

No. %

No. %

No. %

0

2 10

2 15

4 36

9 100

18 90

11 85

7 64

16

0

>256

Y

Total No.

%

2 20

10

16

8 80

53

84

No.

SMA and GA in patients with CMV infection SMA were found in five of the nine adult patients with clinical CMV infection (group 3), but not in the remaining four patients (Table 2). SMA of the IgM class were found in titres between 10 and 40 in four patients (numbers 1-4) in the first serum sample, which was obtained 3-5 weeks after the onset of symptoms. The titres decreased during the following weeks; in three of the patients (numbers 1-3) the antibodies disappeared within 6-22 weeks, while the titre had decreased from 20 in week 4 to 10 in week 9 in patient number 4. In two patients (numbers 3 and 5), IgG-SMA in titres of 10 and 40 were found in the 3rd week of illness, and in another two (numbers 2 and 4) IgG antibodies appeared later in the course of illness as seen from Table 2. The IgG-SMA had disappeared from serum after 13 and 26 weeks in two patients (numbers 2 and 3), while they were still present in patient number 5 after 12 weeks. Thus IgG-SMA appeared later and could be demonstrated at a later stage of disease than SMA of the IgM class. Four patients with CMV infection (numbers 1, 3-5) had GA with the highest titre in the first serum sample obtained, and the GA disappeared faster from serum than SMA. The GA with maximum titres of 20 were of the IgM class in patients numbers 1, 3 and 4, and of the IgG class in patient number 5. CF-CMV antibodies were present in the highest titres in the early phase of the disease in five patients (numbers 1-4 and 9), and in four of them (numbers 1-4) SMA were present in the first serum sample obtained. The CF-CMV antibody titre decreased during the course of illness in four patients (numbers 1-3 and 9), while it remained almost constant in patient number 4. who was studied only during the first 9 weeks of illness. In four cases (numbers 5-8), the CF-CMV antibody titre increased and in only one of them (number 5) could SMA be demonstrated. It appears from Table 2 that in adults with CMV infection in whom SMA developed the IgM-SMA seemed to appear as early or almost as early as CF-CMV antibodies, and that SMA disappeared faster from serum than CF-CMV antibodies. NT-CMV antibodies developed more slowly than CF-CMV antibodies and SMA, and the highest NT-CMV antibody titre was not reached until after 7-26 weeks of illness when SMA had disappeared from serum. Four (numbers 1-4) of the five SMA-positive patients were under 34 years of age; three of them (numbers 1-3) had elevated serum transaminase values and relative lymphocytosis, while these parameters were not determined in patient number 4. The last SMA-positive

Smooth-muscle antibodies in CMV infection

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TABLE 2. Tissue antibody titres and CMV antibody titres during the course of CMV infection in six female (F) and three male (M) patients

Patient number

Age (yr)

Sex

1

25

M

2

27

F

3

33

F

4

11

M

SMA titre ANA Weeks after onset of GA titre IgG IgM titre IgM disease 40 20 10

3 6 8 12 14 18 34 5 8 10 22 26 3 4 6 13 -5* -3*

20 10 10 10 10 10

20t

10 20 10 10

10 10 10

20 20

20t 20

20 10

10t 10 10

_1*

5

61

F

6

59

F

7

65

F

8

67

M

9

26

F

4 6 9 3 5 8 12 4 8 3 7 3 7 7 11 27 41

-

10

40 40 40 40

10

20t

-

-

10

10 10

10

-

-

Titre < 10. Investigated prior to the onset of illness. t Antibody of IgM class. I Antibody of IgG class. -

*

=

CMV antibody titre CF

NT

64 32 64 32 16 8 256 16 64 4 32 16 64 > 256 128 -16 32

Smooth-muscle antibodies and other tissue antibodies in cytomegalovirus infection.

Clin. exp. Immunol. (1975) 22, 22-29. SMOOTH-MUSCLE ANTIBODIES AND OTHER TISSUE ANTIBODIES IN CYTOMEGALOVIRUS INFECTION P. ANDERSEN AND H. K. ANDERSE...
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