Regulating Neutrophil PAD4/NOX-Dependent Cerebrovasular Thromboinflammation

Area of Science:

• Hematology and Immunology
• Neuroscience and Neurology
• Vascular Biology

Background:

• Neutrophil extracellular trap (NET) production is linked to thromboinflammation in Sickle Cell Disease (SCD), increasing ischemic stroke risk.
• Peptidyl Arginine Deiminase 4 (PAD4) and NADPH oxidase (NOX)-dependent reactive oxygen species (ROS) are critical for NET formation and neutrophil function.
• The precise role of NOX-dependent ROS and NETs in SCD-related cerebral microvascular thrombosis remains unclear.

Purpose of the Study:

• To investigate the role of NOX-dependent ROS and NETs in accelerated cerebral microvascular thrombosis in SCD.
• To evaluate the therapeutic potential of targeting PAD4 and NOX in mitigating thromboinflammation and stroke risk.

Main Methods:

• In vitro studies utilized neutrophils from healthy volunteers and SCD patients to assess the effects of targeting PAD4 and NOX on NET production.
• In vivo studies employed a photoactivation thrombosis model in mice (C57BL/6 and sickle transgenic mice) to examine the impact of PAD4 and NOX inhibition on cerebral microcirculation.
• Real-time fluorescence intravital microscopy was used to assess blood flow dynamics in cerebral microvessels.

Main Results:

• Targeting PAD4 and NOX significantly inhibited ionomycin-dependent H3cit+ neutrophils in human neutrophils.
• In vivo inhibition of PAD4 and NOX prolonged blood flow cessation in cerebral arterioles and venules in mice.
• These findings were replicated in a clinical model of accelerated thromboinflammation, showing attenuation of H3cit+ neutrophil production in SCD patient neutrophils.

Conclusions:

• PAD4 and NOX play a significant role in neutrophil-mediated thromboinflammation.
• Targeting PAD4 and NOX effectively limits pathological H3cit+ neutrophils, suggesting a mechanism for attenuating cerebral thrombosis.
• This study establishes a viable preclinical model for preventing and managing thromboinflammatory complications like ischemic stroke.