Caspase-Driven Microvascular Inflammation and Hypoperfusion in Intravascular Hemolysis: Roles of Leukocyte and Endothelial Activation

Intravascular hemolysis (IVH), a pathological process associated with various conditions, triggers vascular responses; however, the molecular and cellular mechanisms driving this process remain unclear. To explore the role of NLRP3 inflammasome- and caspase-dependent pathways in IVH-induced inflammation, we used in vivo models of acute and chronic IVH, alongside hemestimulation of endothelial cells, thereby isolating this disease mechanism from its etiological causes. Acute IVH induced immediate inflammatory responses in C57BL/6 mice, marked by the release of pro-inflammatory molecules, including IL-1{beta}, within just 15 minutes and NLRP3-dependent caspase activation in circulating leukocytes. Chronic IVH processes in mice elevated liver monocyte-derived macrophage caspase activity and NLRP3 protein expression. In turn, acute IVH impaired cutaneous microvascular blood flow and perfusion, and induced microvascular leukocyte recruitment, which were both caspase-1-dependent. Acute IVH induced time-dependent CD11b-integrin-subunit presentation on leukocytes, while heme stimulation augmented endothelial cell adhesion molecule expression, potentially promoting leukocyte recruitment. This endothelial activation was associated with reactive oxygen species generation, which promoted caspase-1 activation and was key to adhesion molecule upregulation. Our findings highlight a role for inflammasome-/caspase-dependent pathways in hemolytic inflammation, contributing to microvascular leukocyte recruitment and particularly to cutaneous hypoperfusion, a consequence that could facilitate the progression of skin lesions. Targeting caspase-dependent pathways and their downstream effects in disorders that display IVH may offer therapeutic potential for maintaining endothelial integrity, reducing leukocyte activation, and mitigating ischemic injury. NEW & NOTEWORTHYThis study identifies caspase-1 as a driver of the vascular inflammation and hypoperfusion induced by intravascular hemolysis (IVH). Using in vivo models and heme-stimulated endothelial cells, we show that hemolysis rapidly induces caspase-1-dependent endothelial-leukocyte recruitment, microvascular dysfunction, and also IL-1{beta} release. Oxidative stress promotes heme-induced endothelial caspase-1 activation and adhesion molecule expression, potentially amplifying vascular dysfunction. These findings provide insight into IVH-driven pathology in hemolytic disorders, including sickle cell disease.