**4. Electroanatomic mapping-guided endomyocardial biopsy in patients with ventricular arrhythmias**

There is growing evidence that 3D-EAM-guided endomyocardial biopsy represent a new important tool to improve the diagnosis and the treatment of patients with arrhythmias, as well as the knowledge of the pathological substrates and mechanisms ubderlying arrhythmic syndromes. Since 2006 we performed 66 3D-EAM-guided endomyocardial biopsy procedures. In all cases the biopsies where drawn from the right ventricle, with biventricular mapping and 3D-EAM-guided biopsy being performed in 10 patients. Although data on a direct comparison between 3D-EAM-guide and conventional technique are not yet available, the 3D-EAM-guided approach seems to significantly improve the diagnostic sensitivity and specificity of endomyocardial biopsy. In fact, when applying immunohistochemistry and molecular biology techniques, a definite histologic diagnosis was obtained in all cases. The new technique was firstly adopted in a series of 30 patients with a noninvasive diagnosis of ARVC according to current diagnostic criteria (Pieroni et al., 2009). Twenty-nine (97%) of 30 patients presented an abnormal voltage map. Histology and immunohistochemistry confirmed the diagnosis of ARVC in 15 patients, while showed an active myocarditis in the remaining 15 patients (Figure 5). Patients with ARVC were not distinguishable on the basis of clinical features, presence and severity of structural and functional right ventricular abnormalities and three-dimensional 3D-EAM findings.

Fig. 5. 3D-EAM-guided EMB, in a patient with bigeminism due to ventricular ectopic beats with LBBB morphology and inferior axis (A). 3D-EAM shows low voltages in the outflow tract and free wall (B). Endomyocardial biopsies drawn from free wall (arrow) shows active myocarditis (C).

**4. Electroanatomic mapping-guided endomyocardial biopsy in patients with** 

There is growing evidence that 3D-EAM-guided endomyocardial biopsy represent a new important tool to improve the diagnosis and the treatment of patients with arrhythmias, as well as the knowledge of the pathological substrates and mechanisms ubderlying arrhythmic syndromes. Since 2006 we performed 66 3D-EAM-guided endomyocardial biopsy procedures. In all cases the biopsies where drawn from the right ventricle, with biventricular mapping and 3D-EAM-guided biopsy being performed in 10 patients. Although data on a direct comparison between 3D-EAM-guide and conventional technique are not yet available, the 3D-EAM-guided approach seems to significantly improve the diagnostic sensitivity and specificity of endomyocardial biopsy. In fact, when applying immunohistochemistry and molecular biology techniques, a definite histologic diagnosis was obtained in all cases. The new technique was firstly adopted in a series of 30 patients with a noninvasive diagnosis of ARVC according to current diagnostic criteria (Pieroni et al., 2009). Twenty-nine (97%) of 30 patients presented an abnormal voltage map. Histology and immunohistochemistry confirmed the diagnosis of ARVC in 15 patients, while showed an active myocarditis in the remaining 15 patients (Figure 5). Patients with ARVC were not distinguishable on the basis of clinical features, presence and severity of structural and

functional right ventricular abnormalities and three-dimensional 3D-EAM findings.

Fig. 5. 3D-EAM-guided EMB, in a patient with bigeminism due to ventricular ectopic beats with LBBB morphology and inferior axis (A). 3D-EAM shows low voltages in the outflow tract and free wall (B). Endomyocardial biopsies drawn from free wall (arrow) shows active

**ventricular arrhythmias** 

myocarditis (C).

On the basis of clinical and histological features, a cardioverter defibrillator was implanted in 13 patients with biopsy-proven ARVC and in 1 patient only with myocarditis. At a mean follow-up of 21±8 months, 7 (47%) patients with ARVC experienced recurrence of symptomatic sustained ventricular arrhythmias with appropriate defibrillator intervention in all cases. All patients with myocarditis remained asymptomatic and free from arrhythmic events. Our study was the first to demonstrate that 3D-EAM-guided EMB may allow obtaining a differential diagnosis in patients with otherwise undistiguishable clinical, arrhythmic and imaging features. In addition our study clearly showed that patients with similar arrhythmic presentation may have a dramatically different prognosis in terms of arrhythmias' recurrence according to the underlying disorders, as only 53% of biopsyproven ARVC patients remained free from arrhythmias at a mean follow-up of 21±8 months years, while no patients with biopsy-proven myocarditis experienced major arrhythmic events during the same time (Figure 6).

Fig. 6. Kaplan-Meier analysis of arrhythmic event-free survival depending on the histological findings (Modified from Pieroni et al. 2009).

In a more recent study (Pieroni et al unpublished data) we performed 3D-EAM-guided EMB in a series of elite competitive athletes presenting with sustained ventricular arrhythmias. We studied 13 consecutive competitive athletes with evidence of sustained ventricular arrhythmias within the previous six months on 12-lead ECG, 24-hour Holter ECG or ECG exercise testing and who were judged as having a structurally normal heart after a thorough non-invasive evaluation, including signal-averaged electrocardiogram, transthoracic echocardiogram and CMR. Depending on the presumed site of arrhythmias origin according to 12-lead ECG criteria, patients underwent right or left ventricular 3D-EAM and 3D-EAM-guided endomyocardial biopsy. Twelve (92%) patients presented at least 1 lowvoltage region at 3D-EAM, while the histologic diagnosis was active myocarditis in 7 patients, and of arrhythmogenic RV cardiomyopathy in 5. In one patient the histological evidence of contraction-band necrosis allowed to unmask caffeine and ephedrine abuse.

The identification of the underlying histological substrate in patients with ventricular arrhythmias may be relevant also when radiofrequency catheter ablation (RFCA) is

Myocarditis Presenting with Ventricular Arrhythmias:

targets for therapy.

Role of Electroanatomical Mapping-Guided Endomyocardial Biopsy in Differential Diagnosis 381

Immunosuppression is clearly recommended essentially for the treatment of eosinophilic, granulomatous, giant-cell myocarditis and lymphocytic myocarditis associated with connective tissue diseases or with the rejection of a transplanted heart. With regard to idiopathic lymphocytic myocarditis, beyond acute phase where a spontaneous resolution has been reported in up to 40% of the cases, many patients with idiopathic myocarditis and chronic heart failure are likely to benefit from immunosuppression. An up-regulation of HLA antigens in the myocardial tissue of patients with lymphocytic myocarditis has been proposed as a marker of susceptibility to beneficial effects of immunosuppression (Wojnicz et al., 2001). Recently the data from a randomized double-blind, placebo-controlled study designed to evaluate the efficacy of immunosuppression in virus-negative inflammatory cardiomyopathy have been reported (Frustaci et al., 2009). Eighty-five patients with myocarditis and chronic (>6 months) heart failure unresponsive to conventional therapy and no evidence of myocardial viral genomes were randomized to receive either prednisone and azathioprine (43 patients, Group 1) or placebo (42 patients, Group 2) in addition to conventional therapy for heart failure. Primary outcome was the 6 months improvement in left-ventricular function. Group 1 showed a significant improvement of left-ventricular ejection fraction and a significant decrease in left-ventricular dimensions and volumes compared with baseline. None of Group 2 patients showed improvement of ejection fraction, that significantly worsened compared with baseline. No major adverse reaction was registered as a result of immunosuppression. These data confirmed the efficacy of immunosuppression in virus-negative inflammatory cardiomyopathy. Lack of response in 12% of cases suggested the presence of not screened viruses or mechanisms of damage and inflammation not susceptible to immunosuppression. Antiviral therapy is a logical strategy advocated for the treatment of myocarditis in patients with evidence of viral presence in myocardial tissue. In a small phase II study 22 patients with myocarditis and biopsy-proven viral persistence (15 enteroviral and 7 adenoviral) were treated with interferon-beta 18x106 IU/week for 24 weeks. Interferon-b treatment resulted in complete elimination of both enteroviral and adenoviral genomes and hemodynamic improvement, as shown by improvement of EF and amelioration of heart failure symptoms. Importantly, treatment with interferon-beta was safe and well-tolerated, with no adverse cardiac effects, and flulike side effects could be efficiently eliminated by non-steroidal anti-inflammatory drugs (Kuhl et al., 2003). After these encouraging results, a prospective placebo-controlled randomized multicenter study, the Betaferon® in Chronic Viral Cardiomyopathy (BICC) trial, was initiated in 2002 and presented at the 2008 American Heart Association scientific sessions. The primary endpoint was virus elimination or reduction of virus load. At 24-week follow-up, compared to placebo, virus elimination and/or viral load reduction were significantly higher in the interferon group. Moreover, interferon-beta showed a good safety profile and was associated with beneficial effects on clinical secondary endpoints, such as NYHA functional class and quality of life, as assessed by Minnesota questionnaire, although echocardiographic and hemodynamic parameters were not statistically significantly different. This interesting phase II study showed proof of concept for targeted therapy based on molecular diagnosis in inflammatory cardiomyopathy. Future directions in the field of antiviral therapy include preventing direct viral damage and inhibition of viral proliferation by preventing the interaction of viruses with their cellular receptor and their consequent signalling amplification systems, such as the tyrosine kinase p56lck, phosphatase CD45 and downstream ERK1/2 (Liu et al., 2000). Therefore, additional research is required to further understand the mechanisms of viral heart disease and consequently identify new potential

considered as a possible therapeutic strategy. In fact there is growing evidence that in patients with ARVC, RFCA is effective in abolishing ventricular tachycardias, but the rate or recurrence of arrhthmias with the same or a different morphology may be very high even when an epicardial approach is adopted (Dalal et al., 2007). On the contrary several reports have demonstrated that RFCA may be effective in eliminating ventricular arrhythmias in myocardial inflammatory disorders, including Chagas cardiomyopathy and sarcoidosis (Sosa et al., 1999; Sarabanda et al., 2005; Jefic et al., 2009; Henz et al., 2009). Recent reports and also our experience suggest that endocardial ablation cannot be sufficient in some patients and a combined endo-epicardial approach is required for definitive results. (See below, Clinical implications of myocarditis diagnosis)
