The NLRP3 inflammasome is assembled in macrophages and monocytes in response to inflammatory and danger stimuli. The atypical nature of the NLRP3 complex impedes detection of NLRP3 inflammasome formation by conventional biochemical and cell biology methods. In situ proximity ligation assay (PLA) provides an alternative method of detection, localization, and quantification of protein-protein interactions in tissue and cell samples. Two primary antibodies raised in different species detect the two proteins of interest. When the proteins are in close proximity, secondary antibodies conjugated with specific DNA probes hybridize with linking oligonucleotides to form a DNA bridge between the two proteins. Amplification of the DNA bridge then facilitates detection by microscopy using fluorescence probes. Here, we describe application of in situ PLA to detect NLRP3 inflammasome assembly in mouse bone marrow-derived macrophages and human monocyte cell line THP1.
Download full-text PDF |
Source |
|---|---|
| http://dx.doi.org/10.1007/978-1-4939-3566-6_12 | DOI Listing |
Publication Analysis
Top Keywords
Similar Publications
Dynamin-Related Protein 1 Orchestrates Inflammatory Responses in Periodontal Macrophages via Interaction With Hexokinase 1.
J Clin Periodontol
January 2025
Department of Oral Implantology, School and Hospital of Stomatology, China Medical University, Liaoning Provincial Key Laboratory of Oral Diseases, Shenyang, China.
Aim: To explore the potential roles of mitochondrial dysfunction in the initiation of inflammation in periodontal macrophages and to determine the mechanism underlying the involvement of dynamin-related protein 1 (Drp1) in macrophage inflammatory responses through its interaction with hexokinase 1 (HK1).
Materials And Methods: Gingival tissues were collected from patients diagnosed with periodontitis or from healthy volunteers. Drp1 tetramer formation and phosphorylation were analysed using western blot.
NLRP3 inflammasome-modulated angiogenic function of EPC via PI3K/ Akt/mTOR pathway in diabetic myocardial infarction.
Cardiovasc Diabetol
January 2025
Department of Clinical Pharmacy, School of Preclinical Medicine and Clinical Pharmacy, China Pharmaceutical University, 24 Tong jia Lane, Nanjing, 210009, People's Republic of China.
Background: Inflammatory diseases impair the reparative properties of endothelial progenitor cells (EPC); however, the involvement of diabetes in EPC dysfunction associated with myocardial infarction (MI) remains unknown.
Methods: A model was established combining high-fat diet (HFD)/streptozotocin (STZ)-induced diabetic mice with myocardial infarction. The therapeutic effects of transplanted wild-type EPC, Nlrp3 knockout EPC, and Nlrp3 overexpression EPC were evaluated.
Helicobacter pylori infection promotes M1 macrophage polarization and gastric inflammation by activation of NLRP3 inflammasome via TNF/TNFR1 axis.
Cell Commun Signal
January 2025
Department of Gastroenterology, Jiangxi Provincial Key Laboratory of Digestive Diseases, Jiangxi Clinical Research Center for Gastroenterology, Digestive Disease Hospital, The First Affiliated Hospital, Jiangxi Medical College, Nanchang University, Nanchang, Jiangxi, China.
Background: Macrophages play a crucial role in chronic gastritis induced by the pathogenic Helicobacter pylori (H. pylori) infection. NLRP3 inflammasome has emerged as an important component of inflammatory processes.
View Article and Find Full Text PDFCombination therapies and other therapeutic approaches targeting the NLRP3 inflammasome and neuroinflammatory pathways: a promising approach for traumatic brain injury.
Immunopharmacol Immunotoxicol
January 2025
Tobacco and Health Research Center, Endocrinology and Metabolism Research Center, Molecular Medicine Research Center, Hormozgan Health Institute, Hormozgan University of Medical Sciences, Bandar Abbas, Iran.
Traumatic brain injury (TBI) precipitates a neuroinflammatory cascade, with the NLRP3 inflammasome emerging as a critical mediator. This review scrutinizes the complex activation pathways of the NLRP3 inflammasome by underscoring the intricate interplay between calcium signaling, mitochondrial disturbances, redox imbalances, lysosomal integrity, and autophagy. It is hypothesized that a combination therapy approach-integrating NF-κB pathway inhibitors with NLRP3 inflammasome antagonists-holds the potential to synergistically dampen the inflammatory storm associated with TBI.
View Article and Find Full Text PDFRunx2-NLRP3 Axis Orchestrates Matrix Stiffness-evoked Vascular Smooth Muscle Cell Inflammation.
Am J Physiol Cell Physiol
January 2025
Department of Physiology and Pathophysiology, School of Basic Medical Sciences, State Key Laboratory of Vascular Homeostasis and Remodeling, Peking University, Beijing 100191, China.
Arterial stiffening is a hallmark of chronic kidney disease (CKD) related cardiovascular events and is primarily attributed to the elevated matrix stiffness. Stiffened arteries are accompanied by low-grade inflammation, but the causal effects of matrix stiffness on inflammation remain unknown. For analysis of the relationship between arterial stiffness and vascular inflammation, pulse wave velocity (PWV) and aortic inflammatory markers were analyzed in an adenine-induced mouse model of CKD in chronological order.
View Article and Find Full Text PDFWant AI Summaries of new PubMed Abstracts delivered to your In-box?
Enter search terms and have AI summaries delivered each week - change queries or unsubscribe any time!