Reinhard Kandolf

Cardiovascular Magnetic Resonance Assessment of Human Myocarditis. A Comparison to Histology and Molecular Pathology

Background—Myocarditis can occasionally lead to sudden death and may progress to dilated cardiomyopathy in up to 10% of patients. Because the initial onset is difficult to recognize clinically and the diagnostic tools available are unsatisfactory, new strategies to diagnose myocarditis are needed. Methods and Results—Cardiovascular MR imaging (CMR) was performed in 32 patients who were diagnosed with myocarditis by clinical criteria. To determine whether CMR visualizes areas of active myocarditis, endomyocardial biopsy was taken from the region of contrast enhancement and submitted to histopathologic analysis. Follow-up was performed 3 month later. Contrast enhancement was present in 28 patients (88%) and was usually seen with one or several foci in the myocardium. Foci were most frequently located in the lateral free wall. In the 21 patients in whom biopsy was obtained from the region of contrast enhancement, histopathologic analysis revealed active myocarditis in 19 patients (parvovirus B19, n=12; human herpes virus type 6 [HHV 6], n=5). Conversely, in the remaining 11 patients, in whom biopsy could not be taken from the region of contrast enhancement, active myocarditis was found in one case only (HHV6). At follow-up, the area of contrast enhancement decreased from 9±11% to 3±4% of left ventricular mass as the left ventricular ejection fraction improved from 47±19% to 60±10%. Conclusions—Contrast enhancement is a frequent finding in the clinical setting of suspected myocarditis and is associated with active inflammation defined by histopathology. Myocarditis occurs predominantly in the lateral free wall. Contrast CMR is a valuable tool for the evaluation and monitoring of inflammatory heart disease.

Presentation, Patterns of Myocardial Damage, and Clinical Course of Viral Myocarditis

Background— Enteroviruses and adenoviruses have been considered the most common causes of viral myocarditis, but parvovirus B19 (PVB19) and human herpesvirus 6 (HHV6) are increasingly found in endomyocardial biopsy samples. Methods and Results— Consequently, our aim was to evaluate the prevalence and clinical presentation of cardiac PVB19 and/or HHV6 infection in a cohort of myocarditis patients and to follow its clinical course. In addition, we sought to demonstrate patterns of myocardial damage and to determine predictors for chronic heart failure. Our study design consisted of a cardiovascular magnetic resonance protocol as well as endomyocardial biopsies in the myocardial region affected as indicated by cardiovascular magnetic resonance. One hundred twenty-eight patients were enrolled by clinical criteria. In the group of myocarditis patients (n=87), PVB19 (n=49), HHV6 (n=16), and combined PVB19/HHV6 infections (n=15) were detected most frequently. The remaining patients were diagnosed with healing myocarditis (n=15) or did not have myocarditis (n=26). Patients with PVB19 presented in a manner similar to that of myocardial infarction; most had typical subepicardial late gadolinium enhancement in the lateral wall and recovered within months. Conversely, patients with HHV6 and especially with HHV6/PVB19 myocarditis presented with new onset of heart failure, had septal late gadolinium enhancement, and frequently progressed toward chronic heart failure. Conclusions— Our data indicate that PVB19 and HHV6 are the most important causes for viral myocarditis in Germany and that the clinical presentation is related to the type of virus. Furthermore, clinical presentation, type of virus, and pattern of myocardial damage are related to the clinical course.

High Prevalence of Viral Genomes and Multiple Viral Infections in the Myocardium of Adults With “Idiopathic” Left Ventricular Dysfunction

Background—For a long time, enteroviruses have been considered to be the most common cause of acute viral myocarditis (MC), with possible transition from MC to dilated cardiomyopathy (DCM). Recent investigations have shown, however, that other viruses are also frequently encountered in MC patients, suggesting that persistence of various virus species may play a pathogenic role in the transition from MC to DCM. The purpose of this study was to screen endomyocardial biopsies (EMBs) from patients with “idiopathic” DCM for the presence of viral genomes by using polymerase chain reaction (PCR) to assess the frequency of cardiac viral infections that may be involved in the pathogenesis of the disease. Methods and Results—EMBs were obtained for PCR analysis from 245 consecutive patients (median left ventricular ejection fraction, 35.0%; range, 9% to 59%). PCR and reverse transcription–PCR were performed to detect the genomic sequences of enterovirus (EV), adenovirus (ADV), human cytomegalovirus (HCMV), herpes simplex virus, Epstein-Barr virus (EBV), human herpesvirus 6 (HHV-6), parvovirus B19 (PVB19), and influenza A and B viruses. Myocardial inflammation was assessed by histological and immunohistological analyses. Viral genomes could be amplified from EMBs of 165 (67.4%) of the 245 DCM patients: EV=23 (9.4%), ADV=4 (1.6%), PVB19=126 (51.4%), HHV-6=53 (21.6%), EBV=5 (2.0%), HCMV=2 (0.8%), including n=45 cases (27.3%) with multiple infections. Active or borderline myocarditis according to the Dallas classification did not exist in any case. Lymphocyte and macrophage infiltrates were not significantly different in virus-positive versus virus-negative patients. Conclusions—Viral genomes were frequently detected in EMBs of patients with systolic left ventricular dysfunction. Our data suggest that myocardial persistence of various viruses, often presenting as multiple infections, may play a role in the pathogenesis of DCM far more frequently than suspected so far.

Predictors of Outcome in Patients With Suspected Myocarditis

Background— The objective of this study was to identify the prognostic indicators in patients with suspected myocarditis who underwent endomyocardial biopsy. Methods and Results— Between 1994 and 2007, 181 consecutive patients (age, 42±15 years) with clinically suspected viral myocarditis were enrolled and followed up for a mean of 59±42 months. Endomyocardial biopsies were studied for inflammation with histological (Dallas) and immunohistological criteria. Virus genome was detected by polymerase chain reaction. The primary end point was time to cardiac death or heart transplantation. In 38% of the patients (n=69), the Dallas criteria were positive. Immunohistological signs of inflammation were shown in 50% (n=91). Genomes of cardiotropic virus species were detected in 79 patients (44%). During follow-up, 22% of the patients (n=40) reached the primary end point. Three independent predictors were identified for the primary end point, namely New York Heart Association class III or IV at entry (hazard ratio, 3.20; 95% confidence interval, 1.36 to 7.57; P=0.008), immunohistological evidence of inflammatory infiltrates in the myocardium (hazard ratio, 3.46; 95% confidence interval, 1.39 to 8.62; P=0.008), and β-blocker therapy (hazard ratio, 0.43; 95% confidence interval, 0.21 to 0.91; P=0.027). Ejection fraction, left ventricular end-diastolic pressure, and left ventricular end-diastolic dimension index were predictive only in univariate, not in multivariate, analysis. Neither the Dallas criteria nor the detection of viral genome was a predictor of outcome. Conclusions— For patients with suspected myocarditis, advanced New York Heart Association functional class, immunohistological signs of inflammation, and lack of β-blocker therapy, but not histology (positive Dallas criteria) or viral genome detection, are related to poor outcome.

Update on myocarditis.

Myocarditis is an inflammatory disease of the heart frequently resulting from viral infections and/or post-viral immune-mediated responses. It is one of the important causes of dilated cardiomyopathy worldwide. The diagnosis is presumed on clinical presentation and noninvasive diagnostic methods such as cardiovascular magnetic resonance imaging. Endomyocardial biopsy remains the gold standard for in vivo diagnosis of myocarditis. The therapeutic and prognostic benefits of endomyocardial biopsy results have recently been demonstrated in several clinical trials. Although remarkable advances in diagnosis, understanding of pathophysiological mechanisms, and treatment of acute myocarditis were gained during the last years, no standard treatment strategies could be defined as yet, apart from standard heart failure therapy and physical rest. In severe cases, mechanical support or heart transplantation may become necessary. There is some evidence that immunosuppressive and immunomodulating therapy are effective for chronic, virus-negative inflammatory cardiomyopathy. Further investigations by controlled, randomized studies are needed to definitively determine their role in the treatment of myocarditis.

Cardiovascular magnetic resonance in clinically suspected cardiac amyloidosis: noninvasive imaging compared to endomyocardial biopsy.

OBJECTIVES We sought to evaluate the diagnostic performance of cardiovascular magnetic resonance imaging (CMRI) for detection of cardiac amyloidosis compared with endomyocardial biopsy (EMB) in a clinical routine setting. BACKGROUND For the clinical workup of heart failure with restrictive filling, pattern cardiac amyloidosis is an important differential diagnosis that is difficult to verify with current noninvasive techniques, especially in the presence of myocardial hypertrophy. METHODS A total of 33 consecutive patients underwent both CMRI and EMB for workup of heart failure with restrictive filling pattern in combination with myocardial hypertrophy (n = 24) and/or clinical conditions often associated with cardiac amyloidosis (n = 18). RESULTS Cardiac amyloidosis was detected by EMB in 15 of the 33 patients. In patients with biopsy-proven cardiac amyloidosis, CMRI revealed a distinct pattern of late gadolinium enhancement, which was distributed over the entire subendocardial circumference, extending in various degrees into the neighboring myocardium. This pattern was found in 12 of the 15 patients diagnosed with cardiac amyloidosis by EMB, compared with only 1 individual in the group of 18 patients diagnosed with other myocardial diseases. Consequently, using this pattern as a diagnostic criterion, the sensitivity of CMRI for diagnosing cardiac amyloidosis was 80%, yielding a specificity of 94%. The positive predictive value was 92%, and the negative predictive value was 85%. CONCLUSIONS In patients with biopsy-proven cardiac amyloidosis, late gadolinium enhancement frequently occurs in a peculiar pattern. On the basis of the gold standard, EMB, noninvasive CMRI can be used to diagnose or rule out cardiac amyloidosis with good sensitivity and excellent specificity in a clinical routine setting.

Detection and Typing of Human Papillomavirus by E6 Nested Multiplex PCR

ABSTRACT A nested multiplex PCR (NMPCR) assay that combines degenerate E6/E7 consensus primers and type-specific primers was evaluated for the detection and typing of human papillomavirus (HPV) genotypes 6/11, 16, 18, 31, 33, 35, 39, 42, 43, 44, 45, 51, 52, 56, 58, 59, 66, and 68 using HPV DNA-containing plasmids and cervical scrapes (n = 1,525). The performance of the NMPCR assay relative to that of conventional PCR with MY09-MY11 and GP5+-GP6+ primers, and nested PCR with these two primer sets (MY/GP) was evaluated in 495 cervical scrapes with corresponding histologic and cytologic findings. HPV prevalence rates determined with the NMPCR assay were 34.7% (102 of 294) in the absence of cervical intraepithelial neoplasia (CIN 0), 94.2% (113 of 120) in the presence of mild or moderate dysplasia (CIN I/II), and 97.8% (44 of 45) in the presence of severe dysplasia (CIN III). The combination of all four HPV detection methods applied in the study was taken as “gold standard”: in all three morphological subgroups the NMPCR assay had significantly (P < 0.0001) higher sensitivities than the MY09-MY11 and GP5+-GP6+ assays and sensitivities comparable or equal to those of the MY/GP assay. All 18 HPV genotypes investigated were detected among the clinical samples. The ratio of high- to low-risk HPV genotypes increased from 4:1 (80 of 103) in CIN 0 to 19:1 (149 of 157) in CIN I to III. Multiple infections were detected in 47.9% (124 of 259) of the patients. In conclusion, the novel NMPCR method is a sensitive and useful tool for HPV DNA detection, especially when exact HPV genotyping and the identification of multiple HPV infections are required.

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.

Molecular Typing of Enteroviruses: Current Status and Future Requirements

SUMMARY Human enteroviruses have traditionally been typed according to neutralization serotype. This procedure is limited by the difficulty in culturing some enteroviruses, the availability of antisera for serotyping, and the cost and technical complexity of serotyping procedures. Furthermore, the impact of information derived from enterovirus serotyping is generally perceived to be low. Enteroviruses are now increasingly being detected by PCR rather than by culture. Classical typing methods will therefore no longer be possible in most instances. An alternative means of enterovirus typing, employing PCR in conjunction with molecular genetic techniques such as nucleotide sequencing or nucleic acid hybridization, would complement molecular diagnosis, may overcome some of the problems associated with serotyping, and would provide additional information regarding the epidemiology and biological properties of enteroviruses. We argue the case for developing a molecular typing system, discuss the genetic basis of such a system, review the literature describing attempts to identify or classify enteroviruses by molecular methods, and suggest ways in which the goal of molecular typing may be realized.

Enteroviral RNA Replication in the Myocardium of Patients With Left Ventricular Dysfunction and Clinically Suspected Myocarditis

BACKGROUND Previous studies dealing with the detection of enteroviral RNA in human endomyocardial biopsies have not differentiated between latent persistence of the enteroviral genome and active viral replication. Enteroviruses that are considered important factors for the development of myocarditis have a single-strand RNA genome of positive polarity that is transcribed by a virus-encoded RNA polymerase into a minus-strand mRNA during active viral replication. The synthesis of multiple copies of minus-strand enteroviral RNA therefore occurs only at sites of active viral replication but not in tissues with mere persistence of the viral genome. METHODS AND RESULTS We investigated enteroviral RNA replication versus enteroviral RNA persistence in endomyocardial biopsies of 45 patients with left ventricular dysfunction and clinically suspected myocarditis. Using reverse-transcriptase polymerase chain reaction in conjunction with Southern blot hybridization, we established a highly sensitive assay to specifically detect plus-strand versus minus-strand enteroviral RNA in the biopsies. Plus-strand enteroviral RNA was detected in endomyocardial biopsies of 18 (40%) of 45 patients, whereas minus-strand RNA as an indication of active enteroviral RNA replication was detected in only 10 (56%) of these 18 plus-strand-positive patients. Enteroviral RNA was not found in biopsies of the control group (n=26). CONCLUSIONS These data demonstrate that a significant fraction of patients with left ventricular dysfunction and clinically suspected myocarditis had active enteroviral RNA replication in their myocardium (22%). Differentiation between patients with active viral replication and latent viral persistence should be particularly important in future studies evaluating different therapeutic strategies. In addition, molecular genetic detection of enteroviral genome and differentiation between replicating versus persistent viruses is possible in a single endomyocardial biopsy.