Bone morphogenetic proteins: Novel mediators of atherothrombosis

Abstract

Atherosclerosis is a major cause of morbidity and mortality in western world. Bone morphogenetic proteins are secreted regulatory proteins that regulate various processes throughout human body. Recent studies indicate their presence in atherosclerotic plaques and emerging role in plaque development. Since monocytes are the key effector cells in atherosclerosis, the aim of this thesis was to investigate the actions of BMP-2 and -7 on monocyte thrombogenicity and motility. In our first paper we investigated the effects of BMP-2 on TF expression in human mononuclear cells (MNCs). We showed that BMP-2 induced phosphorylation of Smad 1/5/8, thus activating the canonical BMP signaling pathway. Though BMP-2 had no effect on the baseline TF expression, it was able to significantly reduce LPS-induced TF expression. When MNCs were pretreated with BMP-2 prior to LPS stimulation, a marked decrease in phosphorylation of ERK1/2, JNK and p38 was observed. BMP-2 also blocked the activation of AP-1 transcription factor, as was shown by use of AP-1 or NFkB sensitive luciferase constructs. This study shows that BMP-2 reduces LPS-induced TF expression in human MNCs by reducing activation of ERK1/2, JNK and p38 as well as blockade of AP-1 transcription factor. In our second and third papers we investigated the signaling pathways behind the ability of BMP-7 to induce TF in human MNCs. We showed that BMP-7 upregulates both TF protein levels, surface presentation and procoagulant activity as well as mRNA levels. BMP-7 was able to induce phosphorylation of ERK1/2, JNK and p38, signaling kinases essential in regulation of TF gene expression. Using luciferase constructs driven by either wildtype or mutated F3 gene promoters we showed that intact NFkB binding site on the F3 promoter is necessary for BMP-7-induced TF expression. Experiments with NFkB inhibitor, JSH-23, supported this finding. In our forth paper we present a novel function for BMP-7 – regulation of monocyte motility. Human monocytes pretreated with BMP-7 crawl for longer distances, attach to endothelium more readily and migrate faster through endothelial monolayers and show higher levels of active beta2 integrins on the cell surface. The observed effects were dependent on the activation of Akt/FAK signaling pathway and can be blocked by either natural BMP antagonist Noggin or synthetic BMP type 1 receptor inhibitor, Dorsomorphin.