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Whole blood assay as a model for in vitro evaluation of inflammasome activation and subsequent caspase-mediated interleukin-1 beta release.


ABSTRACT: Processing of pro-interleukin (IL)-1? and IL-18 is regulated by multiprotein complexes, known as inflammasomes. Inflammasome activation results in generation of bioactive IL-1? and IL-18, which can exert potent pro-inflammatory effects. Our aim was to develop a whole blood-based assay to study the inflammasome in vitro and that also can be used as an assay in clinical studies. We show whole blood is a suitable milieu to study inflammasome activation in primary human monocytes. We demonstrated that unprocessed human blood cells can be stimulated to activate the inflammasome by the addition of adenosine 5'-triphosphate (ATP) within a narrow timeframe following lipopolysaccharide (LPS) priming. Stimulation with LPS resulted in IL-1? release; however, addition of ATP is necessary for "full-blown" inflammasome stimulation resulting in high IL-1? and IL-18 release. Intracellular cytokine staining demonstrated monocytes are the major producers of IL-1? in human whole blood cultures, and this was associated with activation of caspase-1/4/5, as detected by a fluorescently labelled caspase-1/4/5 probe. By applying caspase inhibitors, we show that both the canonical inflammasome pathway (via caspase-1) as well as the non-canonical inflammasome pathway (via caspases-4 and 5) can be studied using this whole blood-based model.

SUBMITTER: Tran TAT 

PROVIDER: S-EPMC6453527 | biostudies-literature | 2019

REPOSITORIES: biostudies-literature

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Whole blood assay as a model for in vitro evaluation of inflammasome activation and subsequent caspase-mediated interleukin-1 beta release.

Tran Thi Anh Thu TAT   Grievink Hendrika W HW   Lipinska Katarzyna K   Kluft Cornelis C   Burggraaf Jacobus J   Moerland Matthijs M   Tasev Dimitar D   Malone Karen E KE  

PloS one 20190408 4


Processing of pro-interleukin (IL)-1β and IL-18 is regulated by multiprotein complexes, known as inflammasomes. Inflammasome activation results in generation of bioactive IL-1β and IL-18, which can exert potent pro-inflammatory effects. Our aim was to develop a whole blood-based assay to study the inflammasome in vitro and that also can be used as an assay in clinical studies. We show whole blood is a suitable milieu to study inflammasome activation in primary human monocytes. We demonstrated th  ...[more]

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