A chromEM-staining protocol optimized for cardiac tissue.

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.

Aerosolization risk during endoscopic transnasal surgery: a prospective qualitative and quantitative microscopic analysis of particles spreading in the operating room.

OBJECTIVE: The coronavirus disease 2019 (COVID-19) pandemic represents the greatest public health emergency of this century. The primary mode of viral transmission is droplet transmission through direct contact with large droplets generated during breathing, talking, coughing, and sneezing. However, the virus can also demonstrate airborne transmission through smaller droplets (< 5 µm in diameter) generated during various medical procedures, collectively termed aerosol-generating procedures. The aim of this study was to analyze droplet contamination of healthcare workers and splatter patterns in the operating theater that resulted from endoscopic transnasal procedures in noninfected patients. METHODS: A prospective nonrandomized microscopic evaluation of contaminants generated during 10 endoscopic transnasal procedures performed from May 14 to June 11, 2020, in the same operating theater was carried out. A dilution of monosodium fluorescein, repeatedly instilled through nasal irrigation, was used as a marker of contaminants generated during surgical procedures. Contaminants were collected on detectors worn by healthcare workers and placed in standard points in the operating theater. Analysis of number, dimensions, and characteristics of contaminants was carried out with fluorescence microscopy. RESULTS: A total of 70 samples collected from 10 surgical procedures were analyzed. Liquid droplets and solid-tissue fragments were identified as contaminants on all detectors analyzed. All healthcare workers appeared to have been exposed to a significant number of contaminants. A significant degree of contamination was observed in every site of the operating room. The mean (range) diameter of liquid droplets was 4.1 (1.0-26.6) µm and that of solid fragments was 23.6 (3.5-263.3) µm. CONCLUSIONS: Endoscopic endonasal surgery is associated with the generation of large amounts of contaminants, some of which measure less than 5 µm. All healthcare workers in the surgical room are exposed to a significant and similar risk of contamination; therefore, adequate personal protective equipment should be employed when performing endoscopic endonasal surgical procedures.

Complex karyotype in a case of cutaneous lymphangiosarcoma associated with chronic lymphedema of the lower limb.

Lymphangiosarcoma is a rare malignant neoplasm of endothelial cells. The term is used to describe an angiosarcoma associated with chronic lymphedema. The skin of the head and neck region is the most common site of origin. Rather few cytogenetic studies on lymphangiosarcoma are reported in the literature. We here describe a case of an 87-year-old woman, with a history of recurring lymphangitis and with an ulcerated nodular lesion of the leg. The histological diagnosis was a malignant neoplasm of vascular origin, with the morphological and immunohistochemical features of a lymphangiosarcoma. A series of antibodies (CD31, CD34, vimentin, podoplanin and HHV-8), conventional and molecular cytogenetic and Spectral Karyotyping (SKY-FISH) analyses were used to study this case. The immunohistochemical evaluation revealed that the neoplasm was positive for vimentin, CD31, CD34 and podoplanin and negative for HHV-8. The proliferation rate (Ki-67) was about 70%. Karyotype was defined using conventional cytogenetic and SKY-FISH. In addition, high-level of amplification was observed with MYC split signal probe. The morphological and immunohistochemical evaluations supported the diagnosis of lymphangiosarcoma. Moreover, the cytogenetic and molecular findings contributed towards accurately defining the karyotypic aberrations of this rare sarcoma.