Neutrophils are critical to the inflammatory process and tissue homeostasis. The lobulated neutrophil nucleus has a unique membrane composition, yielding a deformable structure that is highly responsive to the environment and allows for efficient migration to the site of inflammation. Furthermore, neutrophils possess an arsenal of antimicrobial defense mechanisms, including the expulsion of neutrophil extracellular traps (NETs), composed of nuclear DNA studded with cellular and antimicrobial proteins. However, dysregulated or excessive neutrophil activation is implicated in many malignant and inflammatory diseases, highlighting the need to understand neutrophil biology. To date, thorough study of neutrophil morphology and especially the nucleus has been greatly limited by the resolution limits inherent in microscopy and thus requires expensive and highly specialized super-resolution equipment. Therefore, we have implemented expansion microscopy for imaging of neutrophils, as it is cost- and time-effective and can easily be performed using conventional equipment. We used this novel technique to characterize the distribution of chromatin, histone H1, and nucleophosmin (NPM1) in both unstimulated neutrophils and those undergoing NETosis. We also labeled lamins B1 and B2 as nuclear envelope markers and myeloperoxidase (MPO) as a neutrophil cytoplasmic protein. The gain in resolution was calculated by assessing the colocalization of histone H1 and chromatin using Pearson’s coefficient. Line scan analysis of NPM1 distribution throughout the nucleus showed an increase in detail. In conclusion, expansion microscopy is a novel tool to study neutrophil nuclear dynamics and image neutrophil proteins at super-resolution using conventional microscopes. This method can easily be adapted to other (immune) cell types and is thus a valuable tool to analyze cellular morphology.

ESDR377 - Exploring neutrophil nuclear dynamics and neutrophil extracellular traps using expansion microscopy

The scientific meeting of the European Society for Dermatological Research (ESDR) is held yearly, usually in September. With an international attendance of more than 1000 skin scientists from around the globe, the four-day event includes short oral presentations from peer-reviewed abstracts, keynote lectures from international experts, sponsored symposia, satellite programs, educational events as well as an important focus on posters, networking and building collaborations amongst scientists. For more information about this year’s meeting visit our website.