David A. Neumann

Circulating heart-reactive antibodies in patients with myocarditis or cardiomyopathy

Heart-reactive antibodies are commonly observed in patients with myocarditis or cardiomyopathy. Such antibodies may be important in the pathogenesis of these disorders, yet the specific antigens recognized have not been studied systematically. This report characterizes circulating heart autoantibodies from patients with myocarditis (n = 17) or idiopathic cardiomyopathy (n = 71) and from healthy volunteers (n = 15). Indirect immunofluorescence demonstrated that high titer (greater than or equal to 1:20) immunoglobulin G (IgG) antibody activity occurred in 59% of the myocarditis samples, 20% of the cardiomyopathy samples and none of the normal samples. All samples were tested by Western immunoblotting for IgG activity against a normal human heart extract. The number of antigens recognized by each sample was enumerated and the molecular weight of each antigen estimated; the prevalence of reactivity against antigens in selected molecular weight classes was determined. There was no difference in the mean number of heart antigens recognized by serum from each group. For most weight classes, prevalence either did not differ significantly among the various groups or subgroups or was greatest among samples from healthy volunteers. Prevalence of reactivity with 190 to 199 kilodalton (kd) antigens was greatest (p less than 0.05) among low titer serum samples from patients with myocarditis. High titer cardiomyopathy serum differed from normal serum by an increased (p less than 0.05) prevalence of antibodies to 40 to 49 and 100 to 109 kd antigens. These results suggest that western immunostaining may ultimately contribute substantively to identifying patients with myocarditis or cardiomyopathy.

Prevalence and incidence of left ventricular dysfunction in patients with human immunodeficiency virus infection

The prevalence and incidence of left ventricular (LV) dysfunction was examined in patients infected with the human immunodeficiency virus (HIV). Sixty-nine randomly selected patients diagnosed with HIV infection who were followed in HIV clinics were prospectively evaluated by 2-dimensional echocardiography. Mean follow-up duration was 11 months. Additionally, 39 consecutive HIV-infected patients referred to the Cardiomyopathy Service and found to have LV dysfunction by 2-dimensional echocardiography were also studied. Of the 39 referred patients, 34 (87%) were referred for recent onset, unexplained, congestive heart failure. During this time, the HIV clinic population comprised 1,819 alive and actively followed patients; the 39 cardiomyopathy referrals therefore constituted a crude rate of 2.1% for this population. Of the 69 prospectively studied patients without clinical heart disease, a 14.5% prevalence of global LV hypokinesia and an incidence of 18%/patient-year were found. During a maximal 18-month follow-up period, 4 prospective patients (5.8%) developed symptoms of congestive heart failure. A greater proportion of prospective and referred patients with LV dysfunction had CD4 counts < 100/mm3 (62 and 79%, respectively) than did that of those without LV dysfunction (35%). In conclusion, the high rate of unexpected LV dysfunction in this HIV-infected population suggests that early cardiac contractile abnormalities may involve a significant number of patients, most of whom have low CD4 counts. A subgroup of these patients appears to progress to symptomatic congestive heart failure.

Induction of major histocompatibility complex antigens within the myocardium of patients with active myocarditis: A nonhistologic marker of myocarditis

The histologic diagnosis of active myocarditis is frequently difficult to establish. A nonhistologic marker of immune activation would be clinically useful in identifying cases of immune-mediated myocarditis. A viral etiology with subsequent autoimmunity to cardiac antigens has been implicated in human myocarditis. Because autoimmunity and viral disease are commonly associated with increased expression of major histocompatibility complex (MHC) antigens on targeted tissue, we examined endomyocardial biopsy samples from patients with active myocarditis for abnormal levels of MHC antigen expression. Thirteen patients with active myocarditis and eight control patients with other well-defined cardiac diagnoses (coronary disease, amyloidosis or neoplasm) were studied. A sensitive radioimmunoassay was developed that utilized monoclonal antibodies to human MHC class I and class II antigens in order to quantitate the expression of both of these antigens within each biopsy. Abnormal MHC class I and class II antigen expression was present in 11 of 13 myocarditis specimens and 1 of 8 control samples (specificity 88%, sensitivity 84.6%). Active myocarditis samples had approximately a 10-fold increase in MHC class I and class II expression. Immunoperoxidase staining localized abnormal MHC expression primarily within microvascular endothelium and along myocyte surfaces (11 of 13). This study is the first to demonstrate a marked increase in major histocompatibility complex antigen expression within the myocardium of patients with active myocarditis. The identification of abnormal histocompatibility antigen expression within an endomyocardial biopsy may prove a useful adjunct to the histologic diagnosis of myocarditis.

Demographic Features and Prevalence of Idiopathic Myocarditis in Patients Undergoing Endomyocardial Biopsy

From January 1985 through December 1990, 534 patients underwent endomyocardial biopsy at Johns Hopkins Hospital for suspected myocarditis. One hundred thirty-eight (26%) biopsy specimens were diagnosed histologically by 2 cardiac pathologists as either active (n = 85, 16%) or borderline (n = 53, 10%) myocarditis. Of the 138 patients, 60 were excluded based on either specific concurrent clinical conditions or noncongestive heart failure presentations. Immunohistochemical staining for common leukocyte antigen infiltrating cells performed on the remaining 78 specimens confirmed the presence of focal or multifocal inflammatory infiltrates in 58, of which 49 had histologic evidence of active myocarditis. All 49 patients presented with congestive heart failure and left ventricular ejection fractions of < 40%. Compared with patients with either idiopathic dilated cardiomyopathy (n = 207) or ischemic cardiomyopathy (n = 44), these patients with myocarditis had a less striking male predominance (58 vs 69 and 83%, respectively) (p = 0.02) and were younger (43 +/- 16 vs 50 +/- 17 and 55 +/- 13 years, respectively, p = 0.005). Racial distributions were similar. A recent history of a discrete flu-like illness was obtained in 52%, two-thirds of which were clustered between the months of December and March. Onset of heart failure peaked between December and April (63% and was low between May through September (22%). A peak in the proportion of patients found to have active myocarditis on biopsy occurred in 1986 (17 vs 7 to 10% in other years).(ABSTRACT TRUNCATED AT 250 WORDS)

LPS promotes CB3-induced myocarditis in resistant B10.A mice.

Coxsackie virus B3 (CB3) infection of A/J or A.SW mice results in autoimmune myocarditis characterized by a diffuse mononuclear cell infiltrate and heart-specific autoantibodies. C57BL/10 congenic mice that are identically treated are resistant to this disease. CB3-infected resistant B10.A mice were treated with LPS to determine if this immunomodulator alters disease susceptibility. In contrast to mice infected only with CB3 or treated only with LPS. CB3-infected/LPS-treated (CB3/LPS) B10.A mice developed autoimmune myocarditis similar to that observed in susceptible A/J or A.SW mice. By Day 14, CB3/LPS-induced disease was characterized by significant mortality, myocardial immunoglobulin deposition, and mononuclear cell infiltration of the heart. Immunohistochemical examination revealed deposits of IgG in the heart tissue and serum IgG autoantibodies reactive with sarcolemmal and fibrillary antigens in normal heart tissue. This serum IgG reacted with normal mouse cardiac antigens of a wide range of molecular weights by Western immunoblotting. Because LPS treatment is capable of increasing cytokine levels as well as MHC Class I and Class II expression in heart tissue, it suggests that these factors may contribute to susceptibility to autoimmune myocarditis in CB3-infected mice.

In situ detection of human cytomegalovirus immediate-early gene transcripts within cardiac myocytes of patients with HIV-associated cardiomyopathy

Objective.Recent clinical and echocardiographic studies have identified dilated cardiomyopathy in 10–20% of HIV-infected adults. The purpose of this study was to determine the role of cardiotropic cytomegalovirus (CMV) infection in the development of HIV-associated cardiomyopathy. Design.We generated sense and antisense digoxigenin-labeled riboprobes derived from the CMV immediate-early (IE) and delayed-early (DE) genes and applied them retrospectively to endomyocardial biopsy samples and control autopsy cardiac samples from HIV-infected patients. Setting.Tertiary care, referral hospital. Patients.Twelve consecutive HIV-infected patients with global left ventricular hypokinesis demonstrated on two-dimensional echocardiography; eight randomly selected control autopsy cardiac samples from HIV-infected patients without cardiac disease during life. Measurements and main results.Of the 12 endomyocardial biopsy specimens, six (50%) were found to have specific myocyte nuclear and perinuclear hybridization for transcripts of the CMV IE gene, consistent with non-permissive or latent infection. Similar patterns were not found in any of the eight autopsy control samples. All six patients presented with unexplained congestive heart failure and had CD4 counts <100 x 106/1; all six biopsy samples had immunohistochemical evidence of increased myocardial major histocompatibility complex (MHO class I expression, a finding typical of non-HIV myocarditis. None of the endomyocardial biopsy samples had characteristic CMV inclusions and no specific hybridization was noted with the DE gene riboprobe, suggesting that no active viral DNA replication was present. Only two of the six patients with myocyte hybridization with the IE riboprobe had clinical evidence of solid organ infection with CMV at the time of cardiovascular presentation. Conclusions.This study is the first to demonstrate the expression of the IE gene of CMV within myocytes from HIV-infected patients with cardiomyopathy, suggesting a non-permissive infection of myocytes without classical intranuclear inclusions. Myocyte infection may be necessary to trigger cellular and humoral-mediated cardiac injury and may be best identified using in situ hybridization techniques.

Autoimmune myocarditis: concepts and questions.

There has been a resurgence of interest in immunologically-mediated heart disease, culminating in a recent meeting. This interest was sparked off by new experimental models of autoimmune myocarditis that have served two useful functions: first, as good models of human myocarditis and second, as paradigms of infection-induced autoimmune disease.

Heart-specific autoantibodies can be eluted from the hearts of coxsackievirus B3-infected mice

This study was undertaken to determine if immunoglobulin G (IgG) antibodies could be eluted from the hearts of mice with Coxsackievirus B3‐induced autoimmune myocarditis and to characterize the immunorcactivity of any elutable autoantibodies. Susceptible (A/J) and resistant (B10.A) mice were administered the virus or the control treatment and killed at various times after treatment. Acid eluates from pooled heart tissue from each treatment group and each time were tested for IgG reactivity with normal heart tissue by immunohistochemistry and with normal heart extracts by Western immunostaining. Eluates from infected A/J mice reacted strongly with syngeneic heart and modestly with syngeneic skeletal muscle tissue. Eluates from infected B10.A or control mice of either strain exhibited little reactivity with either tissue. Tissue reactivity was similar when allogeneic tissue was used as the substrate. Eluates from infected A/J mice recognized the heavy chain of cardiac myosin and several other cardiac antigens by Western immunostaining while eluates from the other treatment groups exhibited little or no reactivity with any normal heart constituents. These results indicate that in vivo IgG deposition occurs in the hearts of mice with post‐infectious autoimmune myocarditis and that the specificity of these antibodies is similar to that reported for serum from animals with this disease. The mechanism(s) leading to myocardial IgG deposition and its possible role in pathogenesis remain to be elucidated.

Myocarditis: A Postinfectious Autoimmune Disease

An association of autoimmune disease with infection, including viral infection, is strongly supported by clinical observation and epidemiologic study. A cause-and-effect relationship, however, between a particular infectious agent and a specific autoimmune response has been difficult to establish. A number of plausible mechanisms have been proposed, including molecular mimicry, changes induced in endogenous antigens, and disturbance in the host’s immune response. Coxsackievirus (CV) infections frequently initiate autoimmune response in humans and provide a tool for understanding postinfection autoimmune disease. The first step is to delineate the specificity of the autoimmune response and identify the corresponding antigens. In the case of postinfectious autoimmunity, this task may be complicated by the fact that only a minor proportion of patients go on to develop significant autoimmune disease. Analysis of the pathogenetic potential of the autoimmune response requires establishing a malleable model.

Induction of Major Histocompatibility Antigens on Myocardial Cells in Patients with Active Myocarditis and Idiopathic Cardiomyopathy

The histologic diagnosis of active myocarditis on endomyocardial biopsy has now been standardized and is defined by the presence of interstitial myocardial inflammation associated with myocyte necrosis and degeneration of adjacent myocytes [1]. The use of arbitrary histologic criteria to define a disease whose pathogenesis is not fully unterstood has raised several important diagnostic issues: 1. Defining myocarditis by histology alone may lump together diseases of different etiologies. 2. The histologic criteria may not be sensitve enough to identify all cases of immune-mediated heart disease. 3. The histologic pattern of myocardial injury in most cases of active myocarditis cannot easily explain the severe cardiac dysfunction experienced by these patients.