Aim/background: Atherosclerosis is a chronic inflammatory disease. NLRP3-inflammasome activation and the subsequent release of IL-1b have been shown to impact its development. Parts of the multiproteincomplex of NLRP3 such as ASC specks can be released in the extracellular space and have been implicated to further act as a pro-inflammatory danger signal. We aim to investigate if patients with atherosclerosis show elevated counts of extracellular ASC specks in blood and evaluate their role in human atherosclerotic plaques. 

Methods and results: We first primed THP-1 cells with LPS, activated their inflammasome with Nigericin and isolated the resulting ASC-specks using density gradient centriguation. Isolated ASC-specks were detected with flow cytometry using a primary, fluorochrome-conjugated antibody. By infusing a defined amount of isolated, GFP-labeled ASC-specks in human blood, we established a method to reliably detect extracellular ASC-specks in the serum of patients. We then collected blood samples from patients who received coronary angiography, no matter if they were previously diagnosed with atherosclerosis or if healthy, and quantified the extracellular ASC-specks with flow cytometry. Interestingly, extracellular ASC-specks were detectable in both patients with and without cardiovascular disease. In our current, small cohort of patient (n=45), we found no correlation with severity of atherosclerosis and risk factors (e.g. aHT, Diabetes, Cholesterol, hsCRP). We plan on validating these findings in a larger cohort.

We next asked if extracellular ASC specks could be detected by florescence microscopy in the plaque of patients undergoing carotid endarterectomy. After establishing the method in paraffin embedded samples, we are now evaluating if the amount of extracellular ASC specks correlates with histological parameters like lesion size, thickness of the fibrous cap or composition of inflammatory cells.

Discussion: We showed that extracellular ASC specks are detectable in the blood of patients undergoing coronary catheterization. Despite no obvious differences in ASC speck count between patients in a small cohort, their role should be investigated further to better understand the chronic inflammatory processes underlying cardiovascular diseases.