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(Circulation. 1995;91:631-634.)
© 1995 American Heart Association, Inc.
Articles |
Soluble Interleukin-2 Receptor Levels in Patients With Dilated Cardiomyopathy
Correlation With Disease Severity and Cardiac Autoantibodies
From the Cardiovascular Division, Departments of Medicine, Laboratory Medicine, and Pathology, University of Minnesota School of Medicine, the Minneapolis Heart Institute, and the Department of Veterans Affairs Medical Center, Minneapolis, Minn.
Correspondence to Constantinos J. Limas, MD, FACC, Department of Medicine, University of Minnesota School of Medicine, Box 19, UMHC, 420 Delware St SE, Minneapolis, MN 55455.
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Abstract |
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 Top Abstract IntroductionMethods Results Discussion References |
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Background There is evidence that autoimmunity plays an important role in the initiation and progression of myocardial injury in dilated cardiomyopathy. Abnormalities of both cellular and humoral immunity have been described in this disease. Soluble interleukin-2 receptor (sIL-2R) levels in the serum reflect activation of T lymphocytes in the periphery or in tissues. The present study explored the possibility that activation of cellular immunity is frequent in patients with idiopathic dilated cardiomyopathy and may have functional consequences.
Methods and Results Serum sIL-2R levels were determined with an enzyme-linked immunosorbent assay in 50 dilated cardiomyopathy patients, 30 patients with ischemic heart disease, and 22 normal control subjects. In addition, the presence of anti–ß-receptor and antimyosin antibodies was sought in the serum of cardiomyopathy patients. High sIL-2R levels (>1400 pg/mL) were found in 38% of the dilated cardiomyopathy patients but only 6% of the ischemic heart disease patients. The group of sIL-2R–positive patients was characterized by higher average age, a higher percentage of women, and more severe disease (lower ejection fraction, higher left ventricular filling pressures, and lower cardiac output). Although the prevalence of cardiac autoantibodies did not correlate with the presence of high sIL-2R levels, higher titers of autoantibodies were found predominantly in the sIL-2R–positive group.
Conclusions T-lymphocyte activation, as reflected in elevated sIL-2R levels, is frequent in patients with dilated cardiomyopathy and is associated with more severe disease. Cellular and humoral immune activation may correlate with progression of the disease process.
Key Words: cardiomyopathy • antibodies • receptors • lymphocytes
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Introduction |
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TopAbstractIntroductionMethods Results Discussion References |
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Autoimmunity has been proposed as a major participant in the pathogenesis of human dilated cardiomyopathy on the basis of observed abnormalities involving both cellular and humoral immunity.1 2 3 4 5 6 The former are exemplified by dysfunctional lymphocytes and altered relative proportions of lymphocyte subsets.1 2 3 4 A role for humoral immunity is suggested by reports of enhanced immunoglobulin deposition in the cardiomyopathic myocardium5 and the demonstration of a variety of autoantibodies agonist cardiac cell constituents.6 7 8 The relative contribution of cellular and humoral immune disturbances to the pathogenesis of cardiac injury in cardiomyopathy has not been determined. In experimental autoimmune myocarditis, myocardial damage is thought to be mediated by activated autoreactive T lymphocytes.9 The role of such T-lymphocyte activation in dilated cardiomyopathy is unclear, even though infiltration of the myocardium by lymphocytes is seen in a substantial proportion of patients with this disease.10 11
In the present study, we used the levels of soluble interleukin-2 receptors (sIL-2R) as an index of T-lymphocyte activation in patients with dilated cardiomyopathy and correlated them with hemodynamic severity of disease and the presence of cardiac autoantibodies.
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Methods |
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TopAbstractIntroductionMethods Results Discussion References |
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The study was carried out with 50 patients with idiopathic dilated cardiomyopathy diagnosed according to the WHO criteria12 ; none of the patients had significant coronary artery disease by coronary arteriography, skeletal myopathy, systemic diseases of presumed autoimmune etiology, or history of ethanol abuse. There were 11 women and 39 men, and their mean age was 46.8±5.0 years. All had hemodynamically significant disease as evidenced by an ejection fraction of 23.0±3.0%, pulmonary capillary wedge pressure of 24.0±2.0 mm Hg, and cardiac output of 4.94±0.4 L/min. Two control groups were used: (1) 22 age- and sex-matched normal subjects and (2) 30 patients with ischemic heart disease (27 men and 3 women; age, 48±5 years). The hemodynamic severity of the second group was comparable to that of the cardiomyopathy patients (ejection fraction, 27.0±4.0%; pulmonary capillary wedge pressure, 20.0±2.0 mm Hg; cardiac output, 4.8±0.5 L/min).
Soluble interleukin-2 receptors were assayed with an enzyme-linked immunosorbent assay (ELISA) (Gen Zyme) in the serum of patients and control subjects. Microtiter plates were precoated with a monoclonal antibody to the interleukin-2 receptor followed by a buffer solution containing bovine serum albumin to inhibit nonspecific binding. After removal of the buffer, known concentrations of interleukin-2 standards and serum samples were added and incubated at 37°C for 60 minutes. The wells then were washed five times, and the streptavidin-horseradish peroxidase reagent was added. After incubation at 37°C for 15 minutes, the plates were washed and the reaction was developed for 10 minutes at room temperature. The absorption at 450 nm was read 30 minutes after the reaction was stopped.
Antimyosin antibodies were assayed with an ELISA test against cardiac
myosin prepared by the method of Dalla Libera et al.13 The
conditions of the assay have been described previously.8
Briefly, Nunc microtiter plates from GIBCO were pretreated with 0.2%
(vol/vol) glutaraldehyde in 100 mmol/L sodium phosphate buffer (pH 5.0)
for 4 hours at room temperature and were coated with 20 µg/mL myosin
in 100 mmol/L phosphate buffer, pH 8.0, at 37°C for 3 hours. After
incubation with 1.54 mmol/L L-lysine in phosphate buffer,
pH 8.0, plates were washed three times with phosphate-buffered saline
(PBS) containing 0.05% Tween-20. Serum dilutions (1:50 to 1:800) in
the same buffer containing 0.25% bovine serum albumin were allowed to
absorb at 37°C for 1.5 hours and then were washed three times with
PBS–0.05% Tween-20. Anti-human immunoglobulin G–peroxidase
conjugate
(Sigma Chemical Co) diluted with the above buffer was added in 1:1000
dilution and incubated at 37°C for 1.5 hours. This was followed with
O-phenylenediamine HCl-H2O2
substrate, and incubation was carried out at room temperature for 25
minutes. The reaction was stopped with 4N
H2SO4, and optical density was read at
490 nm using an ELISA Bio-Tek EL 340 plate recorder. Positive results
were considered as the titers with absorbance at 490 nm 
0.1 U above
background.
Anti–ß-receptor antibodies were assayed in the patients' serum with an ELISA test against a ß1-adrenoceptor peptide, as described previously.14
Statistics
Results are presented as mean±SEM. The
Student's
t test was used to compare the two groups of patients, and
P values were considered significant at or below .05.
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Results |
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TopAbstractIntroductionMethods Results Discussion References |
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All normal subjects had interleukin-2 receptor levels <1400 pg/mL (mean, 739±30 pg/mL), while 38% (19 of 50) of the dilated cardiomyopathy patients had values exceeding this level (mean, 2370±30 pg/mL) (Fig 1
). In contrast, only two patients in the
ischemic heart disease group (6%) had elevated sIL-2R levels (one
woman and one man; sIL-2R, 1745 pg/mL and 2822 pg/mL, respectively,
P<.001, compared with the dilated cardiomyopathy patients).
The clinical characteristics of the subgroup of cardiomyopathy patients
with high sIL-2R levels differed from those with normal levels, as
shown in the Table. sIL-2R–positive patients were
older, had a higher proportion of women, and had more severe disease.
For example, only 6.0% of the sIL-2R–positive individuals had an
ejection fraction of 30% compared with 34.6% of the
sIL-2R–negative patients (P<.001). A similar difference
(6.0% versus 34%) was noted when pulmonary capillary wedge pressures
<20 mm Hg were compared. Fig 2 shows a good
correlation between sIL-2R levels and the patients' ejection fraction
and cardiac output.
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Autoantibodies were frequent in the dilated cardiomyopathy
patients, 46% (23 of 50) of whom had anti–ß-receptor antibodies and
68% (34 of 50) of whom had antimyosin antibodies. The prevalence of
these autoantibodies did not differ significantly in the
sIL-2R–positive individuals (42% anti–ß-receptor, 63%
antimyosin)
and sIL-2R–negative (48% anti–ß-receptor, 71% antimyosin)
individuals. However, there was a significant difference in the titers
of these antibodies between sIL-2R–positive and sIL-2R–negative
patients (Figs 3 and 4). For
ß-adrenoceptor autoantibodies, titers of 1600 were found in 20% of
the sIL-2R–positive individuals but only 3% of the sIL-2R–negative
individuals (P<.01). Similarly, titers >200 for antimyosin
antibodies were present in 42% of the sIL-2R–positive patients
versus 15% of the sIL-2R–negative patients (P<.01).
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Discussion |
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TopAbstractIntroductionMethods Results Discussion References |
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The interleukin-2 receptor is a glycoprotein that functions as one of three components (
, ß, 
) of the high-affinity receptor
for the cytokine interleukin-2.15 16 The chain,
also
known as p55, Tac, or CD 25, is a 55-kb transmembrane cell surface
glycoprotein. Upon T-cell activation, this glycoprotein is
proteolytically cleaved to yield a soluble 45-kb glycoprotein (sIL-2R),
which has been used as an index of immune activation.17
Altered levels of sIL-2R have been described in a number of presumed
autoimmune diseases and, in some, have been shown to correlate with
disease
activity.18 19 20 21 22
They are thought to reflect the
severity of inflammation. The finding in the present study that 38% of patients with dilated cardiomyopathy have elevated sIL-2R levels suggests that activation of T cells occurs in a substantial proportion of such patients. Abnormalities in T-cell function have been described in dilated cardiomyopathy1 2 3 4 and include altered proportion of T-cell subpopulations, abnormal blastogenic response to mitogens, and altered interleukin-2 receptor expression. Other indices of T-cell activation, such as neopterin and ß2-microglobulin, also have been reported to be elevated in dilated cardiomyopathy.23 24
Our results also suggest that activation of T-cell lymphocytes, as reflected in elevated sIL-2R levels, correlates with disease severity. Although all the patients included in this study had hemodynamically significant disease, IL-2R–positive individuals had clinical and hemodynamic characteristics of more active and severe disease. Inclusion of patients with milder degrees of myocardial dysfunction might have provided even sharper separation of the sIL-2R–positive and sIL-2R–negative groups. Since patients with ischemic heart disease of comparable hemodynamic severity did not have elevated sIL-2R levels, the findings in cardiomyopathy are not simply the consequence of heart failure. Our results suggest that persistent T-cell activation may contribute to the progression of the disease process, probably through the elaboration of cytokines or direct myocardial damage by inflammatory cells.
Cardiac autoantibodies have been described in dilated cardiomyopathy and may have functional consequences because of their interaction with target antigens. Their presence serves as a marker of disordered humoral immunity in dilated cardiomyopathy, but their relation to cell-mediated immune injury is unclear. The suggestion has been made that in some autoimmune diseases, there is an inverse relation between cellular and humoral immunity.25 In the present series, we found a relatively high prevalence of anti–ß-receptor and antimyosin autoantibodies in both sIL-2R–positive and sIL-2R–negative individuals. It was of interest, however, that high-titer autoantibodies were found predominantly in the sIL-2R–positive group. It might be argued that this is merely the consequence of more extensive or severe tissue injury in this group, with resultant release of intracellular autoantigens. However, we have observed recently (unpublished observation) in a family with cardiomyopathy that antimyosin and anti–ß-receptor antibodies were present in clinically unaffected members, suggesting that elaboration of autoantibodies is not merely an epiphenomenon of tissue damage. A similar observation has been reported in a preliminary report by Caforio et al.26 An alternative explanation would be a concomitant activation of both cellular and humoral immunity in patients with severely compromised myocardium. This immune activation may contribute to the progression of the disease process. In accordance with observations in other autoimmune diseases,18 20 22 elevated sIL-2R and high autoantibody titers may be markers of active disease and may provide prognostic information. This issue can best be addressed in prospective, longitudinal studies.
Received June 15, 1994; accepted September 23, 1994.
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