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FACS-based Detection of Extracellular ASC Specks from NLRP3 Inflammasomes in Inflammatory Diseases

Overview
Journal Clin Exp Immunol
Publisher Oxford University Press
Specialty Allergy & Immunology
Date 2024 Dec 10
PMID 39657685
Authors
Joanne Topping
Samuel Lara-Reyna
Alice Ibbotson
Heledd Jarosz-Griffiths
Leon Chang
James Poulter
Daniel Peckham
Michael F McDermott
Sinisa Savic
Affiliations
Soon will be listed here.
Abstract

Introduction: The apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC) is crucial for inflammasome assembly and activation of several inflammasomes, including NLRP3. ASC aggregates are detected in human sera post pyroptotic cell death, but their inflammasome origin remains unclear.

Method: This study aimed to develop a method to detect ASC aggregates originating from NLRP3 inflammasomes. Initially, human monocytes, macrophages, and THP-1 ASC reporter cells were employed to validate the detection of ASC/NLRP3-positive events through flow cytometry.

Results: The presence of ASC/NLRP3 specks was confirmed in cell supernatants from monocytes and macrophages treated with LPS and nigericin or ATP. Flow cytometry analysis identified double-positive specks in patient sera from inflammatory conditions when compared with healthy controls. Elevated ASC/NLRP3 specks were observed in conditions such as cryopyrin-associated periodic syndrome and Schnitzler's syndrome.

Conclusion: We validated fluorescence-activated cell sorting as a reliable method for detecting ASC/NLRP3 specks in human sera, with potential diagnostic and monitoring applications in certain systemic autoinflammatory diseases.

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