Morpho-biomolecular investigation for detection of infectious agents in dilated cardiomyopathy and lymphocytic myocarditis

Abstract

Heart failure (HF) remains a major public health problem worldwide, with about 30 to 40% of patients dying after one year of initial diagnosis of the disease, both in Brazil and in the world. Molecular biology has detected viral genomes, Borrelia burgdorferi and Chlamydophila pneumoniae in most cases of lymphocytic myocarditis and dilated cardiomyopathy (DCM). However, the incidence and number of infectious agents have been quite variable, possibly because of technical differences (DNA extraction, primers, temperature, etc.) more than the epidemiological differences. The use of multiple diagnostic techniques (morfobiomolecular research) involving morphology by electron microscopy, antigen detection by immunohistochemistry and DNA by real time PCR or in situ hybridization should provide more reliable results.In previous work, studying Chagas or DCM patients, we identified by electron microscopy and molecular biology organisms with morphology consistent with the agents listed above, as well as double membrane microparticles, coated with compatible pathogenic archaea. We hypothesized that they are inducing inflammation and virulence of co-infecting microorganisms and may in part correspond to the microparticles (MPs) largely found in heart failure (HF) patients sera. Favoring this hypothesis, our work (not yet published) showed in HF chagasic patients sera, larger amount of electron lucent microparticles bigger than 100nm, in association with large amounts of collagenase derived from archaea (AMZ), than in indeterminate group.The objectives of this work are:1. Improve sensitivity and specificity in the diagnosis of infectious agents present in the myocardium of patients with heart failure due to idiopathic dilated cardiomyopathy or suspected idiopathic lymphocytic myocarditis using morphological and biomolecular research.2. Verify if infectious agents detected on biopsy correlates with microparticles with archaeal DNA and / or archaeal collagenase AMZ1 by in situ hybridization and immunohistochemistry in electron microscopy.3. Seek if microparticles with archaeal DNA and enzyme are also present in serum and correlate with the presence in the heart.4. Compare "normal" myocardium from donors with the IML autopsies and correlate with inflammatory cells (lymphocytes, macrophages, HLA-DR/DP/DQ).5. Analyze if the cardiac transplant patient shows increased infectious agents in comparison to what is present in the "normal" heart donor, or in the explanted heart, examining the post-transplant biopsies.6. Analyze cases from item 5 who developed rejection, and if the pulse (high doses of corticosteroids) does regress some of the infectious agents. METHODOLOGY: Myocardium fragments and serum from four groups of individuals will be evaluated: GI - 20 IC patients by CMD, lymphocytic myocarditis or borderline myocarditis submitted to heart transplantation at the Heart Institute of the Clinical Hospital of FMUSP and biopsy from explanted heart. GII - 10 myocardium fragments of normal heart of autopsy from accidental death (IML). GIII - 20 heart biopsies from donors of patients in group I, removed during surgery. GIV - biopsies for monitoring patients from GI group after heart transplantation for up to 6 months. (AU)