Uwe Kühl

The Role of Endomyocardial Biopsy in the Management of Cardiovascular Disease A Scientific Statement From the American Heart Association, the American College of Cardiology, and the European Society of Cardiology

The role of endomyocardial biopsy (EMB) in the diagnosis and treatment of adult and pediatric cardiovascular disease remains controversial, and the practice varies widely even among cardiovascular centers of excellence. A need for EMB exists because specific myocardial disorders that have unique prognoses and treatment are seldom diagnosed by noninvasive testing.1 Informed clinical decision making that weighs the risks of EMB against the incremental diagnostic, prognostic, and therapeutic value of the procedure is especially challenging for nonspecialists because the relevant published literature is usually cited according to specific cardiac diseases, which are only diagnosed after EMB. To define the current role of EMB in the management of cardiovascular disease, a multidisciplinary group of experts in cardiomyopathies and cardiovascular pathology was convened by the American Heart Association (AHA), the American College of Cardiology (ACC), and the European Society of Cardiology (ESC). The present Writing Group was charged with reviewing the published literature on the role of EMB in cardiovascular diseases, summarizing this information, and making useful recommendations for clinical practice with classifications of recommendations and levels of evidence. The Writing Group identified 14 clinical scenarios in which the incremental diagnostic, prognostic, and therapeutic value of EMB could be estimated and compared with the procedural risks. The recommendations contained in the present joint Scientific Statement are derived from a comprehensive review of the published literature on specific cardiomyopathies, arrhythmias, and cardiac tumors and are categorized according to presenting clinical syndrome rather than pathologically confirmed disease. The ultimate intent of this document is to provide an understanding of the range of acceptable approaches for the use of EMB while recognizing that individual patient care decisions depend on factors not well reflected in the published literature, such as local availability of specialized facilities, cardiovascular pathology expertise, and operator experience. The use of EMB …

Viral Persistence in the Myocardium Is Associated With Progressive Cardiac Dysfunction

Background—Cardiotropic viral infections have been suspected as one possible cause of myocarditis and dilated cardiomyopathy. Although adverse outcomes in dilated cardiomyopathy patients have been documented, the natural course of heart diseases caused by cardiotropic viruses is unknown. Methods and Results—Consecutive patients (n=172) with biopsy-proven viral infection in endomyocardial biopsies (EMBs) were followed up by reanalysis of EMBs and hemodynamic measurements after a median period of 6.8 months (range, 5.4 to 11.9). Nested polymerase chain reaction (PCR) and reverse transcription–PCR were performed to analyze the genomic sequences. Myocardial inflammation was assessed by histology and immunohistology. At baseline, 32.6% of EMBs in the study group contained enteroviral (EV) RNA, 8.1% adenovirus (ADV) DNA, 36.6% parvovirus B19 (PVB19) DNA, and 10.5% human herpesvirus type 6 (HHV6) DNA. In 12.2% of the samples, dual infection with PVB19 and HHV6 was present. Follow-up analysis of EMBs by PCR documented spontaneous clearance of viral genomes in 36.2% (55/151) of all patients with single infections. Virus-specific clearance rates were 50% for EV, 35.7% for ADV, 22.2% for PVB19, and 44.4% for HHV6. In patients with dual infection with PVB19+ and HHV6+-, HHV6 was cleared in 42.8% (9/21), whereas PVB19 persisted in all 21 patients. Clearance of viral genomes was associated with a significant improvement in left ventricular ejection fraction (LVEF), improving from 50.2±19.1% to 58.1±15.9% (P<0.001). In contrast, LV function decreased in patients with persisting viral genomes (LVEF, 54.3±16.1% versus 51.4±16.1%, P<0.01). Conclusions—In this first biopsy-based analysis of the course of viral heart disease, we show that EV, ADV, PVB19, and HHV6 persistence detected in the myocardium of patients with LV dysfunction was associated with a progressive impairment of LVEF, whereas spontaneous viral elimination was associated with a significant improvement in LV function.

Interferon-β Treatment Eliminates Cardiotropic Viruses and Improves Left Ventricular Function in Patients With Myocardial Persistence of Viral Genomes and Left Ventricular Dysfunction

Background—Viral infections are important causes of myocarditis and may induce cardiac dysfunction and finally lead to dilated cardiomyopathy. We investigated whether interferon (IFN)-&bgr; therapy is safe and may achieve virus clearance and prevent deterioration of left ventricular (LV) function in patients with myocardial virus persistence. Methods and Results—In this phase II study, 22 consecutive patients with persistence of LV dysfunction (history of symptoms, 44±27 months) and polymerase chain reaction–proven enteroviral or adenoviral genomes were treated with 18×106 IU/week IFN-&bgr; (Beneferon) subcutaneously for 24 weeks. Histological and immunohistological analysis of endomyocardial biopsies was used to characterize myocardial inflammation. LV diameters and ejection fraction were assessed by echocardiography and angiography, respectively. During the treatment period, IFN-&bgr; was well tolerated by all patients. No patient deteriorated. Clearance of viral genomes was observed in 22 of 22 of patients after antiviral therapy. Virus clearance was paralleled by a significant decrease of LV end diastolic and end systolic diameters, decreasing from 59.7±11.1 to 56.5±10.0 mm (P <0.001) and 43.2±13.6 to 39.4±12.1 mm (P <0.001), respectively. LV ejection fraction increased from 44.6±15.5% to 53.1±16.8% (P <0.001). Conclusions—A 6 months, IFN-&bgr; treatment was safe in patients with myocardial enteroviral or adenoviral persistence and LV dysfunction and resulted in elimination of viral genomes (22 of 22 patients) and improved LV function (15 of 22 patients).

Cardiac Troponin T in Patients With Clinically Suspected Myocarditis

OBJECTIVES The present study investigated whether myocyte injury can be assessed sensitively by measurement of serum levels of cardiac troponin T (cTnT) in patients with clinically suspected myocarditis and whether cTnT levels may predict the results of histologic and immunohistologic analysis of endomyocardial biopsy specimens. BACKGROUND Conventionally used laboratory variables often fail to show myocyte injury in patients with clinically suspected myocarditis, possibly because of a low extent of myocardial injury in these patients. Sensitive variables for myocyte injury have not yet been investigated. METHODS Eighty patients with clinically suspected myocarditis were screened for creatine kinase (CK) activity, MB isoform of CK (CK-MB) activity and cTnT. Endomyocardial biopsy specimens were examined histologically and immunohistologically. RESULTS cTnT was elevated in 28 of 80 patients with clinically suspected myocarditis, CK in 4 and CK-MB in 1. Histologic analysis alone of the endomyocardial biopsy specimen revealed evidence of myocarditis in only five patients, all with elevated cTnT levels. Twenty-three of 28 patients with elevated cTnT levels had histologically negative findings for myocarditis. Additional immunohistologic analysis revealed evidence of myocarditis in 26 (93%) of 28 patients with elevated cTnT levels and in 23 (44%) of 52 patients with normal cTnT levels. Mean cTnT levels were higher in patients with myocarditis proved histologically or immunohistologically, or both, than in patients without myocarditis (0.59 +/- 1.68 vs. 0.04 +/- 0.05, p < 0.001). CONCLUSIONS Measurement of serum levels of cTnT provides evidence of myocyte injury in patients with clinically suspected myocarditis more sensitively than does conventional determination of cardiac enzyme levels. Myocardial cell damage may be present even in the absence of histologic signs of myocarditis. Additional immunohistologic analysis often shows lymphocytic infiltrates in these patients. Elevated levels of cTnT are highly predictive for myocarditis in this group.

Dilated Cardiomyopathy Is Associated With Significant Changes in Collagen Type I/III ratio

BACKGROUND It is controversial whether myocardial fibrosis in end-stage dilated cardiomyopathy (DCM) is associated with altered collagen type I/type III (Col I/Col III) ratio. METHODS AND RESULTS Patients with DCM (ejection fraction [EF] <50%, n=12) and with mild global left ventricular dysfunction (EF >50%, n=18) were examined. Col I, Col III, and transforming growth factors-beta1 (TGF-beta1) and -beta2 (TGF-beta2) gene expression in endomyocardial biopsies was evaluated by quantitative competitive reverse transcriptase-polymerase chain reaction (qRT-PCR). Collagen content was quantified after picrosirius red and immunohistological staining and by hydroxyproline assay. In patients with EF <50%, there was a pronounced 2- to 6-fold increase of myocardial Col I mRNA abundance (P<0.01), with a corresponding 1.6-fold increase at the protein level versus that found in patients with EF >50%. The Col III mRNA abundance showed a 2.0-fold increase (P<0.04). There was a relevant shift in the Col I/Col III mRNA ratio for DCM patients (Col I/Col III, 8.2) compared with patients with an EF >50% (Col I/Col III, 6. 4). In addition, total collagen content was increased in patients with EF <50% (n=3) (4.3+/-0.1%) compared with patients with EF >50% (n=8) (2.7+/-0.9%) (P<0.004). The biochemically determined ratio of hydroxyproline/total protein (n=12) was correlated to the Col I mRNA abundance (P<0.05, r=0.77). TGF-beta1 and TGF-beta2 showed elevated myocardial mRNA abundances (1- to 7-fold and 4- to 5-fold, respectively) in DCM patients. CONCLUSIONS Differential increase of Col I and Col III leads to an increased Col I/Col III ratio in DCM myocardium. Because Col I provides substantial tensile strength and stiffness, this may contribute to systolic and in particular diastolic dysfunction in DCM.

Parvovirus B19 Infection Mimicking Acute Myocardial Infarction

Background—Enteroviruses (EVs) and adenoviruses (ADVs) have been considered common causes of myocarditis and dilated cardiomyopathy. In the present study, we report on the association of parvovirus B19 (PVB19) genomes in the clinical setting of acute myocarditis. Methods and Results—This study included 24 consecutive patients admitted to our hospital within 24 hours after onset of chest pain. Acute myocardial infarction had been excluded in all patients by coronary angiography. Endomyocardial biopsies were analyzed by nested polymerase chain reaction/reverse transcriptase-polymerase chain reaction for EV, ADV, PVB19, human cytomegalovirus, Epstein-Barr virus, Chlamydia pneumoniae, influenza virus A and B, and Borrelia burgdorferi genomes, respectively, followed by direct sequencing of the amplification products. All patients presented with acute onset of angina pectoris and ST-segment elevations or T-wave inversion mimicking acute myocardial infarction. Mean baseline peak creatinine kinase and creatine kinase-isoenzyme fraction were 342±241 U/L and 32±20 U/L, respectively. Mean troponin T was increased to 7.5±15.0 ng/mL and C-reactive protein to 91±98 mg/mL. Eighteen patients had global or regional wall motion abnormalities (ejection fraction 62.5±15.5%). Histological analysis excluded the presence of active or borderline myocarditis in all but one patient. PVB19, EV, and ADV genomes were detected in the myocardium of 12, 3, and 2 patients, respectively (71%). Follow-up biopsies of virus-positive patients (11 of 17) demonstrated persistence of PVB19 genomes in 6 of 6 patients, EV genomes in 2 of 3 patients, and ADV genomes in 1 of 2 patients, respectively. Conclusions—Virus genomes can be demonstrated in 71% of patients with normal coronary anatomy, clinically mimicking acute myocardial infarction. In addition to EVs and ADVs, PVB19 was the most frequent pathogen.

Suspected Chronic Myocarditis at Cardiac MR: Diagnostic Accuracy and Association with Immunohistologically Detected Inflammation and Viral Persistence

PURPOSE To retrospectively compare the diagnostic accuracy of three cardiac magnetic resonance (MR) imaging approaches for the detection of histologic and immunohistologic criteria (reference standard) proved myocardial inflammation in patients clinically suspected of having chronic myocarditis (CMC). MATERIALS AND METHODS Cardiac MR imaging was performed in 83 consecutive patients (55 male, 28 female; mean age, 44.8 years +/- 17.7 [standard deviation]) clinically suspected of having CMC, after written informed consent was obtained according to guidelines of the local ethics committee, which approved the study. T2-weighted triple-inversion-recovery imaging to calculate the edema ratio (ER), T1-weighted imaging before and after contrast agent administration to calculate the myocardial global relative enhancement (gRE), and inversion-recovery gradient-echo imaging to evaluate areas of late gadolinium enhancement (LE) were performed. The MR results were correlated with the endomyocardial biopsy (EMB) findings to detect intramyocardial inflammation and cardiotropic viral genomes analyzed at polymerase chain reaction assay. For statistical analyses, receiver operating characteristic analysis and the Wilcoxon test for unpaired data were used because the Kolomogorov-Smirnov test revealed a distribution of data that was different from normality. RESULTS Intramyocardial inflammation and cardiotropic viral persistence were confirmed at immunohistologic analysis in 48 and 49 of the 83 patients, respectively. The sensitivity, specificity, and diagnostic accuracy of the MR parameters, as compared with the immunohistologic detection of inflammation, were, respectively, 62%, 86%, and 72% for gRE; 67%, 69%, and 68% for ER; and 27%, 80%, and 49% for LE. Cardiac MR-derived gRE, ER, and LE were not associated with polymerase chain reaction proof of viral genomes. CONCLUSION In patients clinically suspected of having CMC, increased gRE and ER indicating inflammation were common findings that could be confirmed at immunohistologic analysis, whereas LE had low sensitivity and accuracy. Cardiac MR imaging may be helpful in detecting intramyocardial inflammation noninvasively, but it fails to depict viral persistence.

Immunohistological evidence for a chronic intramyocardial inflammatory process in dilated cardiomyopathy.

OBJECTIVE: To determine whether immunohistochemical analysis of cardiac biopsies from patients presenting clinically as dilated cardiomyopathy (DCM) show a chronic inflammatory process. DESIGN: Comparative case control study. SETTING: Tertiary referral centre. PATIENTS: Biopsies from 170 patients with DCM and 85 control patients with other cardiac diseases. RESULTS: Nine patients had sufficient interstitial inflammatory cells to be called borderline myocarditis on conventional histology, leaving 161 patients with DCM. In 78 patients with DCM (48%) there were T lymphocytes in the myocardium. In 48 (62%) of these 78 T lymphocyte densities were in the range 2-14 per high power field (HPF), equivalent to 7-50 per mm2 of tissue. In 43 (89%) interstitial and endothelial immune activation was demonstrated by MHC expression. In 30 patients with T cell counts in the range 1.5-2.0 per HPF, 80% also showed endothelial activation. Lymphocyte density correlated with increased expression of MHC class I and II antigens and the adhesion molecules ICAM, VCAM, ELAM, LFA-3, and GMP140. In all control biopsies the T lymphocyte density was less than 1.0 per HPF (less than 2-5 per mm2 of tissue). CONCLUSIONS: Nearly half the patients with DCM had increased T lymphocyte density and immune activation of endothelial and interstitial cells in their cardiac biopsies. A chronic autoimmune process is still active within the myocardium in a significant percentage of patients with DCM. Immunohistochemical analysis of cardiac biopsies will enhance the sensitivity of cardiac biopsy and is essential for the diagnosis of myocarditis.

High Prevalence of Cardiac Parvovirus B19 Infection in Patients With Isolated Left Ventricular Diastolic Dysfunction

Background—The etiology of left ventricular (LV) isolated diastolic dysfunction often remains unclear. In the present study, we report a strong association between parvovirus B19 (PVB19) genomes and isolated LV diastolic dysfunction. Methods and Results—In 70 patients (mean±SD age, 43±11 years) admitted with exertional dyspnea and/or reduced exercise tolerance despite preserved LV systolic contractility (ejection fraction=68%), isolated diastolic dysfunction was clinically suspected. Patients with classic risk factors for diastolic dysfunction such as hypertension, coronary heart disease, diabetes mellitus, or pulmonary disease had been excluded. Diastolic function was assessed by echocardiography and LV and RV catheterization. Endomyocardial biopsies (EMBs) were analyzed for the presence of storage or infiltrative diseases or myocarditis, including molecular screening for cardiotropic virus genomes. In a substudy of 24 patients who reported atypical angina, coronary endothelial function was additionally investigated with a coronary Doppler flow-wire technique. In 37 of 70 patients (53%), isolated diastolic dysfunction was confirmed as the cause of their clinical symptoms. No evidence for cardiac storage or infiltrative diseases was found in these cases, but in 35 of 37 of these patients (95%), cardiotropic virus genomes were detected in EMBs (P<0.001). PVB19 was the most frequent pathogen in 31 of 37 patients (84%). In a subgroup of 10 patients with diastolic dysfunction and coexisting endothelial dysfunction, all 10 (100%) were PVB19 positive. Conclusions—PVB19 genomes were predominant in patients with unexplained, isolated diastolic dysfunction. A strong association with the incidence of endothelial dysfunction was obvious, consistent with the hypothesis that PVB19-induced endothelial dysfunction may be a possible pathomechanism underlying diastolic dysfunction.

Expression of Cell Adhesion Molecules in Dilated Cardiomyopathy Evidence for Endothelial Activation in Inflammatory Cardiomyopathy

BACKGROUND Dilated cardiomyopathy (DCM) is pathogenically linked to inflammatory cardiomyopathy (InfCM), which is characterized by intramyocardial infiltration. The transendothelial migration of immunocompetent cells is mediated by cell adhesion molecules (CAMs). METHODS AND RESULTS We investigated the expression pattern of CAMs (immunoglobulin superfamily, 32 selectins, and beta1- and beta2-integrins) in endomyocardial biopsies from DCM patients (n=152; left ventricular ejection fraction <40%) using immunohistochemistry. Whereas few specimens obtained at autopsy (controls; n=14) presented enhanced expression regarding single endothelial CAMs (human leukocyte antigen [HLA] class I, 7%; HLA-DR, 14%; CD29, 14%), none demonstrated concurrent abundance of >3 CAMs (inflammatory endothelial activation), nor did any control tissue prove positive for InfCM (>7.0 CD3+ lymphocytes per 1 mm2). In comparison, 64% (n=97) of the DCM biopsies were evaluated positive for InfCM and 67% (n=101) for inflammatory endothelial activation, respectively. Whereas expression of HLA class I, HLA-DR, intercellular cell adhesion molecule-1, and CD29 was distributed homogeneously within a patients serial sections, immunoreactivity of vascular cell adhesion molecule-1, lymphocyte function antigen-3, and the selectins was accentuated on single vascular endothelia. Sixty-six percent of the DCM biopsies presented CD29 abundance also within the extracellular matrix and the sarcolemma. CD62P and CD62E were present in 16% and 40% of the DCM patients, respectively. Endothelial CAM representatives correlated with one another (P<0.05), except for CD62P with HLA. Endothelial CAM expression correlated with intramyocardial infiltrates phenotyped by the corresponding counterreceptors. CONCLUSIONS Inflammatory endothelial activation is present in 67% of DCM patients. Because CAM expression correlates with the immunohistological diagnosis of InfCM and counterreceptor-bearing intramyocardial infiltrates, evaluation of endothelial CAMs might be of diagnostic significance in InfCM.