Attenuation of Atherosclerosis with PAR4 Deficiency: Differential Platelet Outcomes in apoE-/-vs. Ldlr-/-Mice

Abstract

ABSTRACTObjectiveCardiovascular disease (CVD) is a significant burden globally and, despite current therapeutics, remains the leading cause of death. Platelet inhibitors are of interest in CVD treatment to reduce thrombus formation post-plaque rupture as well their contribution to inflammation throughout the progression of atherosclerosis. Protease activated receptor 4 (PAR4) is a receptor highly expressed by platelets, strongly activated by thrombin, and plays a vital role in platelet activation and aggregation. However, the role of PAR4Approach and ResultsMice on a low-density lipoprotein receptor-deficient (Ldlr-/-) background were bred withPar4deficient (Par4-/-) mice to createLdlr-/-/Par4+/+andLdlr-/-/Par4-/-cousin lines. Mice were fed high fat (42%) and cholesterol (0.2%) ‘Western’ diet for 12 weeks for all studies. Bone marrow transplant (BMT) studies were conducted by irradiatingLdlr-/-/Par4+/+andLdlr-/-/Par4-/-mice with 550 rads (2x, 4 hours apart) and then repopulated withPar4+/+orPar4-/-bone marrow. To determine if the effects of thrombin were mediated solely by PAR4, the thrombin inhibitor dabigatran was added to the ‘Western’ diet.Ldlr-/-/Par4-/-given dabigatran did not further decrease their atherosclerotic burden. Differences between apolipoprotein E deficient (apoE-/-) andLdlr-/-platelets were assessed for changes in reactivity. We observed higher PAR4 abundance in arteries with atherosclerosis in human and mice versus healthy controls. PAR4 deficiency attenuated atherosclerosis in the aortic sinus and root versus proficient controls. BMT studies demonstrated this effect was due to hematopoietic cells, most likely platelets. PAR4 appeared to be acting independent of PAR1, as there werer no changes with addition of dabigatran to PAR4 deficient mice.apoE-/-platelets are hyperreactive compared toLdlr-/-platelets.ConclusionsHematopoietic-derived PAR4, most likely platelets, plays a vital role in the development and progression of atherosclerosis. Specific targeting of PAR4 may be a potential therapeutic target for CVD.HighlightsDeficiency of protease-activated receptor 4 attenuates the development of diet-induced atherosclerosis in aLdlr-/-mouse model.PAR4 deficiency in hematopoietic cells is atheroprotective.PAR4 deficiency accounts for the majority of thrombin-induced atherosclerosis in aLdlr-/-mouse model.The examination of platelet-specific proteins and platelet activation should be carefully considered before using theapoE-/-orLdlr-/-mouse models of atherosclerosis.