Atherosclerosis is characterised by disturbed blood flow due to abnormal lipid and cell accumulation within arterial layers. Modified cholesterol forms such as oxidised low-density lipoprotein (oxLDL) enter cells altering their phenotype, triggering over exuberant repair and arterial occlusion, myocardial infarction or stroke. We hypothesised that oxLDL enters vascular wall cells and induces IL-1β secretion, potentially via a caspase-1/NLRP3 mechanism. Human coronary artery endothelial cells (HCAEC) and smooth muscle cells (VSMC), isolated from different donors, were cultured and stimulated (primed) with pro-inflammatory cytokines TNFα and IL-1α (10 ng/ml each, for 48 hours), followed by incubation with human oxLDL (10-50ug/ml) for - up to 6 hours. Inhibitors of Caspase-1 (YVAD), NLRP3 (MCC950) and Gasdermin D (Disulfiram) were added 1h before oxLDL. Cell lysates and culture supernatants were collected and analysed for IL-1β using ELISA. Microscopy imaging showed oxLDL entered stimulated cells and formed particles. OxLDL at 20 and 50 ug/ml induced the maximum release of IL-1β from stimulated HCASMCs and HCAECs respectively compared to control. Inhibition of either NLRP3, Caspase-1 or Gasdermin D significantly reduced the release of IL-1β (4-fold, p<0.0001; 14-fold, p<0.0001, 1.5-fold, p<0.0003 respectively) in HCAEC. In contrast, in HCASMCs, only Caspase-1 inhibition reduced release of IL-1β (2.1-fold, P<0.0001). HCAECs and HCASMCs elicited release of IL-1β in response to the same stimulus via different mechanisms. In HCAECs, released IL-1β potentially exits via a GSDMD-induced membrane pore. These data suggest that caspase-1 or gasdermin D inhibition are likely to be effective vessel wall cell specific strategies for the reduction of atherosclerosis.

This paper was prepared for presentation at the 8th Abu Dhabi International Petroleum Exhibition and Conference held in Abu Dhabi, U.A.E., 11-14 October 1998.