The research interest in our laboratory is to understand how blood clotting factors in circulation function together to stop bleeding when a blood vessel is injured. Under normal physiological conditions, clotting factors circulate in blood as inert molecules with no activity. This is because the signal that can trigger the activation of clotting factors (a receptor called tissue factor) is not exposed to circulating blood but rather located beneath the cells that line the blood vessels. These cells, called endothelial cells, also have clot-inhibiting factors embedded on their surfaces, ensuring that blood remains fluid in circulation. However, when there is an injury to a blood vessel the hidden tissue factor gets exposed. Tissue factor can endow high activity for an otherwise inactive clotting factor, named factor VII. Active factor VII initiates the clotting cascade by the activation of additional factors, culminating in generation of thrombin. Thrombin can aggregate platelets and also cleave fibrinogen to form blood clots at the site of the injury. We study how clotting factors function by cloning their genes into expression vectors and preparing high quantities of recombinant clotting factors in bacterial or animal cells. We then try to understand how these factors (more than a dozen) work together to generate the right amount of blood clot to close-off the wound site. We also introduce specific mutations, known to cause bleeding or thrombosis in humans, into these genes to understand how these mutations cause life-threatening bleeding or thrombotic diseases. Understanding these questions can lead to novel strategies to treat these patients.
Following sealing the wound site on the injured vessel, thrombin generation must be stopped and activities of all clotting factors (active enzymes) must be neutralized to ensure that they will not generate unnecessary clots to occlude blood vessels which can cause heart attack or stroke. We are investigating how two natural pathways can stop thrombin generation. The first pathway is called protein C pathway, which is initiated by thrombin itself when it binds to the vascular cell surface receptor called thrombomodulin (TM). TM is present at high levels on the surface of all endothelial cells that line blood vessels. Thrombin has a very high affinity for TM and upon complex formation with the receptor it can no longer generate blood clots, but instead can rapidly convert the inactive plasma protein C to activated protein C (APC). APC is a potent anticoagulant enzyme that enzymatically degrades two cofactors (Va and VIIIa) that are essential for thrombin generation. In the second pathway, a series of plasma inhibitors neutralize the enzymatic activity of clotting enzymes. A major inhibitor in this pathway is called antithrombin (AT). The inhibitory activity of AT is greatly enhanced when it binds to heparin-like molecules that are attached to selected vessel wall receptors. This is the basis for the worldwide use of heparin, as the primary therapeutic drug in cardiovascular medicine. We have made significant progress as to how AT and heparin function as antithrombotic drugs.
Finally, another major project in our laboratory is to understand how clotting enzymes bind to specific vascular cell surface receptors to activate intracellular signaling pathways. Clotting enzymes can communicate environmental cues in circulation to cells and tissues below the vessel wall through this mechanism. Clotting enzymes can modulate inflammatory responses during injury and infection. We and others have demonstrated that signaling by both APC and AT plays key protective roles in the vasculature during injury and infection. A deficiency in the component of either APC or AT pathway is associated with pathogenesis of cardiovascular and inflammatory diseases including atherosclerosis, severe sepsis and cancer.
Coagulation and inflammation are closely linked pathways. Both pathways can be activated when there is an injury to the blood vessel and/or a foreign substance (bacteria or virus) enters circulation. The molecular players in both pathways have extensive interactions in order to full-fill their physiological functions (i.e., stop bleeding and mount inflammatory responses). Thus, in addition to clotting fibrinogen to fibrin clots during coagulation, the proteases of the coagulation cascade also interact with specific vascular cell surface receptors to modulate inflammatory responses. Similarly, inflammatory cytokines secreted during inflammation, not only activate immune cells to fight infection but they also play pivotal roles in regulating coagulation reactions. Two natural anticoagulant pathways, namely protein C and antithrombin (AT) pathways are known to regulate both coagulation and inflammatory reactions. The current focus of our laboratory is on understanding the mechanisms through which these two pathways operate at the molecular level. The protein C pathway is initiated when thrombin binds to endothelial cell surface receptor thrombomodulin (TM) to activate protein C (APC). Several years ago, OMRF investigator, Dr. Charles Esmon, who has discovered TM, also discovered that another receptor called endothelial protein C receptor (EPCR) significantly accelerates the protein C activation reaction. Dr. Esmon’s group found that the N-terminal Gla-domain of both protein C and APC binds EPCR with a similar equilibrium dissociation constant of approximately 30 nM. APC can dissociate from EPCR and form a complex with the vitamin K-dependent cofactor, protein S, to rapidly degrade cofactors Va and VIIIa on negatively charged phospholipids that have become exposed on injured cells and/or activated platelets. Interestingly, when APC remains associated with EPCR, the resulting protease-receptor complex activates the G-protein coupled receptor, protease-activated receptor 1 (PAR1), thereby eliciting physiologically significant cytoprotective and antiinflammatory responses in the vasculature. This finding was initially controversial since thrombin can activate PAR1 with an efficiency that is greater than three orders of magnitude higher than that of APC and that the activation of PAR1 by thrombin initiates potent proinflammatory responses in endothelial cells. During the last few years, our laboratory has developed innovative approaches to investigate these apparent PAR1-depependent paradoxical signaling responses by APC and thrombin. We have discovered that all three receptors involved (TM, EPCR and PAR1) in protein C activation and signaling are co-localized within caveolin-1 containing membrane lipid-rafts of endothelial cells and that when EPCR is occupied by either protein C or APC, the activation of PAR1 by either APC or thrombin is cytoprotective independent of the protease activating PAR1. Our results have indicated that the ligand occupancy and association of receptors with caveolin-1 regulates the PAR1-dependent signaling specificity of coagulation proteases. We have also discovered that proinflammatory reactions are dramatically up-regulated by polyphosphates, secreted by activated platelets, through interaction with nuclear proteins and inducing signaling through pathogen-associated pattern recognition receptors; toll-like receptors (TLR) 2 and 4 and the receptor for advanced glycation end products (RAGE), thereby activating mTOR/Wnt/β-catenin signaling pathways in vascular endothelial cells. We have demonstrated that coagulation proteases modulate these signaling pathways through activation of PAR1. These questions are currently being investigated by our laboratory.
In another project, we investigate mechanisms through which antithrombin regulates both anticoagulant and antiinflammatory pathways. Antithrombin (AT), as a major member of the serine protease inhibitor (serpin) family of protease inhibitors, neutralizes the enzymatic activity of all clotting proteases by directly binding their active-sites, thereby trapping them in the form of inactive complexes. The inhibitory activity of AT is greatly enhanced when it binds to the heparin-like molecules that are attached to selected vascular cell surface receptor proteins that are collectively referred to as heparan sulfate proteoglycans (HSPGs). This is the basis for the widespread use of heparin, as the primary anticoagulant drug in the treatment of thrombotic complications. We have made significant contributions to the mechanism by which AT and heparin function as anticoagulants. Recent results have indicated that the interaction of an exposed basic helix (D-helix) of AT with specific vascular HSPGs initiates antiinflammatory signaling responses that are required for the regulation of vascular integrity in health and disease. It has been hypothesized that the interaction of AT with HSPGs is also required for the conformational activation of AT, facilitating its interaction with the active-site pocket of coagulation proteases. Mutations in D-helix interfering with the AT interaction with vascular HSPGs are associated with increased risk of thrombosis. Thus, AT, similar to APC, interfaces the coagulation cascade with inflammatory pathways. The D-helix-dependent antiinflammatory signaling mechanism of AT is poorly understood. We and others have demonstrated that the interaction of AT with vascular HSPGs leads to prostacyclin (PGI2) production and down-regulation of expression of genes regulated by the NF-κB signaling pathway. We also demonstrated that interaction with the 3-O-Sufate containing HSPG named Syndecan 4 is required for the antiinflammatory signaling function of AT. Interestingly, Syndecan 4 also functions as a co-receptor for the basic fibroblast growth factor (FGF2) receptor. The mechanism by which the occupancy of Syndecan 4 by two different ligands (AT and FGF2) elicits distinct intracellular signaling responses in vascular endothelial cells is not understood and this is an active area of investigation in our laboratory.
Our laboratory is employing cellular and animal models to understand mechanisms through which the components of the two protein C and antithrombin anticoagulant pathways described above regulate coagulation and inflammation. A deficiency in the component of either pathway can be associated with pathogenesis of cardiovascular and inflammatory diseases including atherosclerosis, severe sepsis and cancer.
M.A., Harvard University, Cambridge, MA, 1986
Ph.D. Boston University, Boston, MA, 1989
Postdoc, Howard Hughes Medical Institute, Oklahoma City, OK, 1994
Member of Hemostasis and Thrombosis study section (2007-2011)
Ad hoc member of numerous NHLBI Special Emphasis Panel, RFA and PPG study sections
Reviewer of grants for national and international funding organizations
Reviewer for major scientific journals
J Thromb Haemost Advisory Board (2009-Pesent)
Enzyme Research Editorial Board Member (2014-Present)
Science Advances Associate Editor (2017-Present)
American Society of Hematology
International Society of Thrombosis and Hemostasis
Joined OMRF Scientific Staff in 2017
Roy RV, Ardeshirylajimi A, Dinarvand P, Yang L, Rezaie AR. Occupancy of human EPCR by protein C induces β-arrestin-2 biased PAR1 signaling by both APC and thrombin. Blood. 2016 Aug 25. pii: blood-2016-06-720581. [Epub ahead of print]
Hassanian SM, Ardeshirylajimi A, Dinarvand P, Rezaie AR. Inorganic polyphosphate promotes cyclin D1 synthesis through activation of mTOR/Wnt/β-catenin signaling in endothelial cells. J Thromb Haemost. 2016 Oct 5. doi: 10.1111/jth.13477. [Epub ahead of print]
Ding Q, Yang L, Dinarvand P, Wang X, Rezaie AR. Protein C Thr315Ala variant results in gain of function but manifests as type II deficiency in diagnostic assays. Blood. 2015 Apr 9;125(15):2428-34.
Dinarvand P, Hassanian SM, Qureshi SH, Manithody C, Eissenberg JC, Yang L, Rezaie AR. Polyphosphate amplifies proinflammatory responses of nuclear proteins through interaction with receptor for advanced glycation end products and P2Y1 purinergic receptor. Blood. 2014 Feb 6;123(6):935-45
Yang L, Rezaie AR. Residues of the 39-loop restrict the plasma inhibitor specificity of factor IXa. J Biol Chem. 2013 May 3;288(18):12692-8
Costa R, Morrison A, Wang J, Manithody C, Li J, Rezaie AR. Activated protein C modulates cardiac metabolism and augments autophagy in the ischemic heart. J Thromb Haemost. 2012 Sep;10(9):1736-44.
Bae JS, Rezaie AR. Activated protein C inhibits high mobility group box 1 signaling in endothelial cells. Blood. 2011 Oct 6;118(14):3952-9
Rezaie AR, Bae JS, Manithody C, Qureshi SH, Yang L. Protein Z-dependent protease inhibitor binds to the C-terminal domain of protein Z. J Biol Chem. 2008 Jul 18;283(29):19922-6.
Bae JS, Yang L, Manithody C, Rezaie AR. The ligand occupancy of endothelial protein C receptor switches the protease-activated receptor 1-dependent signaling specificity of thrombin from a permeability-enhancing to a barrier-protective response in endothelial cells. Blood. 2007 Dec 1;110(12):3909-16.
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Cardiovascular Biology Research Program, MS 45
Oklahoma Medical Research Foundation
825 N.E. 13th Street
Oklahoma City, OK 73104
Phone: (405) 271-4711
Lab: (405) 271-4743
Fax: (405) 271-3137