Diagnosis of sudden cardiac death due to early myocardial ischemia: An ultrastructural and immunohistochemical study

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Silvia Damiana Visonà *
Donatella Benati
Maria Cristina Monti
Mirco Galiè
Luisa Andrello
Andrea Frontini
Antonio Osculati
(*) Corresponding Author:
Silvia Damiana Visonà | visona.silvia@gmail.com

Abstract

The aim of this post-mortem ultrastructural and immunohistochemical study is to explore the characteristics of acute myocardial ischemia in the context of sudden death, using the combination of two different methods, both more insightful than ordinary histology. Transmission electron microscope and immunohistochemistry, in addition to the traditional histology, were applied to study human heart specimens collected during forensic autopsies. The whole series was sub-grouped into cases (n=17) and controls (n=10). The control group consisted of unnatural death with a short agonal period (immediately lethal injuries). Heart samples of the two cohorts of subjects were prepared for electron microscopy. On the other hand, each specimen, formalin fixed and paraffin embedded, was stained with haematoxylin and eosin and immunoreacted with the following primary antibodies: antiFibronectin, antiConnexin-43, anti npCx43 (dephosphorylated form of Connexin43), antiZonula occludens-1. Immunopositivity of each marker in the myocardium was semi-quantitatively graded. Electron microscopy revealed a number of interesting differences between acute myocardial ischemia and controls, regarding the morphology of nucleus, mitochondria and intercellular junctions. By immunohistochemistry, fibronectin was found to be markedly increased in the extracellular matrix of the acute myocardial ischemia cases, with a remarkable difference in respect of controls. Connexin 43 staining disclosed a slightly increase in the cytoplasm of acute myocardial ischemia cases with respect to the controls, whereas no relevant differences were seen between cases and controls at intercellular junctions. Dephosphorylated form of Cx43 showed an evident difference of staining in cases compared to controls and overall this difference more evident in the cytoplasm. Zonula occludens 1, described as an important marker for functional modification of cardiac muscle fibers, resulted negative or very weak in the vast majority of both cases and controls. The present study attempts to simultaneously apply electron microscopy and immunohistochemistry, in order to figure out the morphological changes that might lead to pathological processes underlying the sudden, unexpected death due to acute myocardial ischemia, and consequently to find useful diagnostic markers of very early ischemic injury. Both methods showed significant differences between acute myocardial ischemia and controls, regarding, overall nuclei, mitochondria, and intercellular junctions. 


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Author Biographies

Silvia Damiana Visonà, University of Pavia

Department of Public Health, Experimental and Forensic Medicine, Unit of Legal Medicine and Forensic Sciences, University of Pavia, Italy, Via Forlanini 12, 27100 Pavia, Italy

Donatella Benati, University of Verona

Department of Neurosciences, Biomedicine and Movement Sciences, University of Verona, Italy. Strada Le Grazie 8, 37134 Verona, Italy

Maria Cristina Monti, University of Pavia

Department of Public Health, Experimental and Forensic Medicine, Unit of Biostatistics and Clinical Epidemiology

Mirco Galiè, University of Verona

Department of Neurosciences, Biomedicine and Movement Sciences, University of Verona, Italy. Strada Le Grazie 8, 37134 Verona, Italy

Luisa Andrello, Service of Legal Medicine, Canton Ticino

Service of Legal Medicine, Canton Ticino, Bellinzona, Switzerland. Via Guisan 3, Bellinzona, Switzerland.

Andrea Frontini, University of Pavia

Department of Public Health, Experimental and Forensic Medicine, Unit of Normal Human Anatomy, University of Pavia, Italy, Via Forlanini 12, 27100 Pavia, Italy

Antonio Osculati, University of Pavia

Department of Public Health, Experimental and Forensic Medicine, Unit of Legal Medicine and Forensic Sciences, University of Pavia, Italy, Via Forlanini 12, 27100 Pavia, Italy.