Charcot-Leyden crystal formation is closely associated with eosinophil extracellular trap cell death

Abstract

<jats:title>Abstract</jats:title> <jats:p>Protein crystallization in human tissue rarely occurs. Charcot-Leyden crystals (CLCs) were described in various eosinophilic diseases &amp;gt;150 years ago, but our understanding of CLC formation still remains limited. In this study, we demonstrate that CLCs observed in varied inflamed human tissues are closely associated with eosinophil cell-free granules and nuclear envelope/plasma membrane disintegration with release of filamentous chromatin (extracellular traps), typical morphologies of a regulated pathway of extracellular trap cell death (ETosis). During the process of eosinophil ETosis, eccentrically localized cytoplasmic and perinuclear CLC protein (galectin-10) is homogeneously redistributed in the cytoplasm. Rapid (1-2 minutes) formation of intracytoplasmic CLCs was observed using time-lapse imaging. Plasma membrane rupture enabled the release of both intracellularly formed CLCs and soluble galectin-10 that further contributed to formation of CLCs extracellularly, in parallel with the expulsion of free intact granules and extracellular traps. CLC formation and galectin-10 release were dependent on nicotinamide adenine dinucleotide phosphate oxidase activation. To our knowledge, this is the first demonstration of natural formation of CLCs in association with an active physiological process (ie, ETosis). These results indicate that dynamic changes in intracellular localization and release of galectin-10 contribute to CLC formation in vivo and suggest that CLC/galectin-10 might serve as an indicator of ETosis.</jats:p>

Charcot-Leyden crystals promote neutrophilic inflammation in patients with nasal polyposis

“…1 CLCs are composed of galectin-10 (Gal10), a protein produced by eosinophils, basophils, and some T cells that autocrystalizes when eosinophils are intensely activated and undergo cytolysis associated with extrusion of DNA extracellular traps, a process referred to as EETosis. 2,3 CLCs are abundantly present in mucosa and mucus from patients with chronic rhinosinusitis with nasal polyps (CRSwNP). Recent work provided evidence that CLCs, by analogy to other crystals, such as uric acid and cholesterol crystals, can activate the NLR family pyrin domain containing 3 (NLRP3) inflammasome and cause IL-1b-driven inflammation after uptake by human macrophages in vitro.…”

“…2,5 Recently, EETosis was shown to be at the basis of CLC formation. 2,3 Inspection of tissue from patients with CRSwNP with a type 2 and IL-5-high profile showed that CLCs were often lining the epithelial layer in zones with denuded or abnormal epithelium. 2,3 In line with these observations, we found that CLCs correlated negatively (P < .001, R 5 20.8792, n 5 12; Fig 1, A) with the relative percentage of normal pseudostratified epithelium in polyp tissue from patients with CRSwNP.…”

“…2 Greater fraction of exhaled nitric oxide (FENO) levels correlate with type 2 asthma and better responses to corticosteroids. 3 In this study we investigated whether IL-6 levels, serum sIL-6R levels, FENO values, blood eosinophil counts, and the combination of FENO values or blood eosinophil counts (type 2 asthma) and IL-6 levels (non-type 2 asthma) are associated with clinical and cellular phenotypes in the SARP3 and SARP1-SARP2 cohorts (see the Methods section in this article's Online Repository at www.jacionline.org). The large and comprehensively phenotyped SARP database is ideal to examine the relationship between type 2 eosinophilic and non-type 2 mechanisms in patients with severe and nonsevere asthma.…”

Sialodochitis fibrinosa: Salivary duct obstruction by eosinophil extracellular traps?

“…Thus far, EETosis has been shown to represent cytolytic degranulation that occurs in diverse eosinophil‐associated diseases (Mukherjee, Lacy, & Ueki, 2018). Recent evidence revealed that CLC formation involves EETosis‐mediated crystallisation of galectin‐10, which is abundant in eosinophil cytosol (Ueki, Tokunaga, et al, 2018). Here, we describe a patient with sialodochitis fibrinosa, who underwent treatment by glandular and ductal resection.…”

Protein phase separation and determinants of in cell crystallization

“…Further, in cellulo crystallization has also been associated with several diseases like cataract, hemoglobin C diseases, formation of Charcot‐Leyden crystals (CLCs), Reinke's crystals or mitochondrial myopathies, and more recently in cellulo crystallization was observed as a result of heterologous overexpression of genes in cell lines of bacteria, insect cells, yeast, CHO (Chinese hamster ovary) or HEK (human embryonic kidney) cells . Mostly, these protein crystals were located in different organelles (mitochondria, peroxisomes, lysosomes, or endoplasmic reticulum), as shown in Table but sometimes also within the cytosol or even in the nucleus, as shown in Figure and Table .…”