Enzymatically oxidized lipids are a specific group of biomolecules that function as key signaling mediators and hormones, regulating various cellular and physiological processes from metabolism and cell death to inflammation and the immune response. They are broadly categorized as either polyunsaturated fatty acid (PUFA) containing (free acid oxygenated PUFA “oxylipins”, endocannabinoids, oxidized phospholipids) or cholesterol derivatives (oxysterols, steroid hormones, and bile acids). Their biosynthesis is accomplished by families of enzymes that include lipoxygenases (LOX), cyclooxygenases (COX), cytochrome P450s (CYP), and aldo-keto reductases (AKR). In contrast, non-enzymatically oxidized lipids are produced by uncontrolled oxidation and are broadly considered to be harmful. Here, we provide an overview of the biochemistry and enzymology of LOXs, COXs, CYPs, and AKRs in humans. Next, we present biosynthetic pathways for oxylipins, oxidized phospholipids, oxysterols, bile acids and steroid hormones. Last, we address gaps in knowledge and suggest directions for future work.

Enzymatically oxygenated phospholipids (eoxPL) formed by lipoxygenases (LOX) and cyclooxygenase (COX) in platelets and leukocytes are pro-coagulant in multiple model systems. However, their generation in arterial thrombotic disease, and how their levels are modulated by common therapies is unknown. Here, eoxPL were first characterized in isolated platelets and leukocytes from an arterial vascular disease cohort, a healthy cohort administered low dose aspirin, and from retrieved human arterial thrombi. In both cohorts, aspirin reduced platelet COX-1-derived eoxPL, while elevating diacyl 12-LOX-derived eoxPL in males, through enhanced Lands cycle esterification. Conversely, P2Y12 inhibition reduced 12-LOX-derived eoxPL in leukocytes. Complex aspirin-dependent gender and seasonal effects on platelet eoxPL were seen in healthy subjects. Limb or coronary (STEMI) thrombi showed a platelet eoxPL signature while carotid thrombi had a white cell profile. Mice genetically lacking leukocyte 12/15-LOX, which are deficient in eoxPL, generated smaller carotid thrombi in vivo. In summary, pro-coagulant eoxPL generation is altered in human arterial vascular disease by commonly used cardiovascular therapies. These changes to the phospholipid composition of blood cells in humans at risk of thrombotic events may be clinically significant where the pro-coagulant membrane plays a central understood role in driving elevated thrombotic risk.

Background: Acute coronary syndrome (ACS) is caused by arterial thrombosis and is associated with sustained activation of coagulation. Clotting requires interactions of coagulation factors with aminophospholipids (aPL): phosphatidylserine (PS) and phosphatidylethanolamine (PE) on membrane surfaces. The aPL composition of circulating membranes in coronary disease has not been characterized. Furthermore, the contribution of external-facing aPL to elevated thrombotic risk in ACS is unknown. Methods and results: Thrombin generation was measured on platelet, leukocyte and extracellular vesicles (EV) from patients with ACS (n = 24), stable coronary artery disease (CAD, n = 18), risk factor positive (RF, n = 23) and healthy controls (HC, n = 24). The aPL composition on the surface of EV, platelets and leukocytes was determined using lipidomics. Leukocytes, platelets and EV externalized PE- and PS-containing fatty acids ranging from C16:0-20:4. These included both diacyl and plasmalogen forms, with significant increases stimulated by agonist activation. Thrombin generation on the surface of EV and leukocytes was higher in ACS than HC. Also, thrombin generation was higher for EV from CAD and RF, than HC. EV counts were higher in CAD and ACS compared with HC. Thrombin generation correlated positively with plasma EV counts and membrane surface area. Conclusion: The aPL membrane of EV and leukocytes may contribute to the activation of coagulation in CAD and ACS. Targeting EV formation/clearance and the aPL surface of EV and leukocyte membranes represents a novel anti-thrombotic target in CAD and ACS.