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Updated: Mar 28, 2026
Evaluation of Caspase Activation to Assess Innate Immune Cell Death
Published on: January 20, 2023
Impact of caspase-8 and PKA in regulating neutrophil-derived microparticle generation
Emily F Midura1, Priya S Prakash1, Bobby L Johnson1
1Division of Research, Department of Surgery, University of Cincinnati College of Medicine, 231 Albert Sabin Way ML 0558, Cincinnati, OH 45267, USA.
Abstract:
The morbidity and mortality from sepsis continues to remain high despite extensive research into understanding this complex immunologic process. Further, while source control and antibiotic therapy have improved patient outcomes, many immunologically based therapies have fallen short. Microparticles (MPs) are intact vesicles that serve as mediators of intercellular communication as well as markers of inflammation in various disease processes. We have previously demonstrated that MPs can be produced at the infected foci during sepsis, are predominantly of neutrophil derivation (NDMPs) and can modulate immune cells. In this study, we sought to elucidate the molecular mechanisms underlying NDMP generation. Using thioglycolate (TGA) to recruit and activate neutrophils, we first determined that intra-peritoneal TGA increase NDMP accumulation. We next utilized TGA-elicited neutrophils in vitro to investigate signaling intermediates involved in NDMP production, including the intrinsic and extrinsic caspase pathways, cAMP dependent PKA and Epac activation as well as the role myosin light chain kinase (MLCK) as a final mediator of NDMP release. We observed that NDMP generation was dependent on the extrinsic caspase apoptotic pathway (caspase 3 and caspase 8), cAMP activation of PKA but not of Epac, and on activation of MLCK. Altogether, these data contribute to an overall framework depicting the molecular mechanisms that regulate NDMP generation.
Insights
Neutrophil-derived microparticles (NDMPs) play a role in sepsis inflammation. This study identified key molecular pathways, including caspase and MLCK activation, that regulate NDMP generation, offering potential therapeutic targets.
Area of Science:
- Immunology
- Cell Biology
- Molecular Medicine
Background:
- Sepsis remains a leading cause of morbidity and mortality despite advances in treatment.
- Microparticles (MPs), particularly neutrophil-derived MPs (NDMPs), are implicated in inflammatory processes and intercellular communication during sepsis.
- Previous research has shown NDMPs can modulate immune cells, but the mechanisms of their generation are not fully understood.
Purpose of the Study:
- To elucidate the molecular mechanisms regulating the generation of neutrophil-derived microparticles (NDMPs).
- To identify key signaling pathways involved in NDMP production.
Main Methods:
- Neutrophils were recruited and activated using thioglycolate (TGA) in vivo and in vitro.
- Investigated the roles of caspase pathways (intrinsic and extrinsic), cAMP-dependent PKA and Epac activation, and myosin light chain kinase (MLCK) in NDMP release.
Main Results:
- Intra-peritoneal TGA administration increased NDMP accumulation.
- NDMP generation was dependent on the extrinsic caspase apoptotic pathway (caspase 3 and 8).
- NDMP production required cAMP activation of PKA (protein kinase A) and MLCK activation, but not Epac.
Conclusions:
- The study delineates a molecular framework for NDMP generation.
- Identified specific signaling pathways (extrinsic caspase, PKA, MLCK) crucial for NDMP release.
- Findings provide insights into the inflammatory mechanisms of sepsis and potential targets for immunomodulatory therapies.
