A chromEM-staining protocol optimized for cardiac tissue

Elettra Musolino; Christina Pagiatakis; Federica Pierin; Daniele Sabatino; Giovanna Finzi; Rosalba Gornati; Giovanni Bernardini; Roberto Papait

doi:10.3389/fcell.2023.1123114

A chromEM-staining protocol optimized for cardiac tissue

Front Cell Dev Biol. 2023 Jul 5:11:1123114. doi: 10.3389/fcell.2023.1123114. eCollection 2023.

Authors

Elettra Musolino¹, Christina Pagiatakis^{1

2}, Federica Pierin¹, Daniele Sabatino³, Giovanna Finzi³, Rosalba Gornati¹, Giovanni Bernardini¹, Roberto Papait^{1

2}

Affiliations

¹ Department of Biotechnology and Life Sciences, University of Insubria, Varese, Italy.
² Department of Cardiovascular Medicine, Humanitas Research Hospital-IRCCS, Rozzano (MI), Italy.
³ Department of Pathology, ASST Sette Laghi, Varese, Italy.

Abstract

Three-dimensional (3D) chromatin organization has a key role in defining the transcription program of cells during development. Its alteration is the cause of gene expression changes responsible for several diseases. Thus, we need new tools to study this aspect of gene expression regulation. To this end, ChromEM was recently developed: this is an electron-microscopy staining technique that selectively marks nuclear DNA without altering its structure and, thus, allows better visualization of 3D chromatin conformation. However, despite increasingly frequent application of this staining technique on cells, it has not yet been applied to visualize chromatin ultrastructure in tissues. Here, we provide a protocol to carry out ChromEM on myocardial tissue harvested from the left ventricles of C57BL/6J mice and use this in combination with transmission electron microscopy (TEM) to measure some morphological parameters of peripheral heterochromatin in cardiomyocytes. This protocol could also be used, in combination with electron tomography, to study 3D chromatin organization in cardiomyocytes in different aspects of heart pathobiology (e.g., heart development, cardiac aging, and heart failure) as well as help to set-up ChromEM in other tissues.

Keywords: aging; cardiac tissue; chromEM; chromatin structure; heterochromatin.

Grants and funding

This work was supported by the NEWMED project (Metodi e materiali innovativi per la medicina di precisione; ID 1175999) of the Fondazione Regionale Ricerca Biomedica–Lombardia and PRIN project 2020 the link between metabolism and epigenetic in myocardial disease (ID 20205 × 4C9E) of the Italian Ministry of University and Research (MUR) to RP and by the HOTZYMES project (grant agreement n. 829162) under EU’s Horizon 2020 Programme (H2020-FETOPEN) to GB.