PB0520 - Atorvastatin treatment inhibits the procoagulant properties of a 3D culture model of the human neointimal layer

Background: Rupture of atherosclerotic plaques exposes the circulation to the neointimal layer of the atherosclerotic plaque, which triggers the unwanted thrombotic responses that lead to stroke and myocardial infarction. Atorvastatin is used to prevent acute cardiovascular events in high-risk patients. Here we have developed a 3D culture model of the human neointimal layer and assessed its responsiveness to atorvastatin treatment.

Aims: To assess the effect of atorvastatin treatment on the prothrombotic properties of the 3D neointimal model.

Methods: THP-1 derived foam cells were cultured within 3D collagen hydrogels by treatment with lipopolysaccharide and IFN-γ. This was followed by treatment with oxidised low-density lipoprotein in the presence or absence of 20 μM atorvastatin. In some experiments the neointimal layer was co-cultured with a medial layer construct in which coronary artery smooth muscle cells were cultured in a collagen hydrogel. Prothrombin time and platelet aggregation onset was measured after exposure of the neointimal models to platelet-poor plasma and washed platelet suspension prepared from blood of healthy, medication-free volunteers. Activity of the extrinsic pathway of coagulation was measured using the fluorogenic substrate SN-17.

Results: Atorvastatin treatment significantly increased the prothrombin time from 155 ± 70s in control samples, to 562 ± 68 s in atorvastatin-treated samples. Atorvastatin treatment significantly reduced the activity of the extrinsic pathway in the neointimal models. The latency of onset of platelet aggregation was also significantly increased in atorvastatin-treated samples (681 ± 140s v 434 ± 129s in control samples). Preliminary experiments in which the neointimal layer was co-cultured with a medial layer, indicated that the neointimal model can elicit smooth muscle cell migration.

Conclusion(s): This 3D culture model is able to replicate the effects of current anti-atherothrombotic therapies, and could be used to develop an in vitro 3D model of human atherosclerosis to replace current animal models.