Posts in Category: Vascular Biology

Bioresorbable Scaffold: The Emerging Reality and Future Directions

Bioresorbable Scaffold: The Emerging Reality and Future Directions

Yohei Sotomi, Yoshinobu Onuma, Carlos Collet, Erhan Tenekecioglu, Renu Virmani, Neal S. Kleiman, Patrick W. Serruys

Strut thickness and platelet activation. The thick protruding strut disrupts the laminar flow and induces flow disturbances, and thereby endothelial shear stress (ESS) microgradients (upper panel). The shear microgradients can induce the formation of stabilized discoid platelet aggregates, the size of which is directly regulated by the magnitude and spatial distribution of the gradient.72,73 Shear microgradient–dependent platelet aggregation requires 3 principal features: shear acceleration phase, peak shear phase, and shear deceleration phase. During shear acceleration, platelets in the central regions of blood flow exposed to laminar flow (constant physiological shear) are suddenly accelerated through the shear microgradient. During the peak shear phase, a proportion of the discoid platelets that are accelerated into the peak shear zone adhere to exposed thrombogenic surfaces through platelet membrane glycoprotein (GP) Ib/IX/V. Exposure of these platelets to elevated hemodynamic drag leads to the extrusion of thin filamentous membrane tethers. Membrane tether formation initiates discoid platelet adhesion with the thrombogenic surface and also facilitates the recruitment of discoid platelets into the downstream deceleration zone. During the shear deceleration phase, platelets transitioning into the flow deceleration zone experience decreasing hemodynamic drag forces. Reduced shear within this zone progressively favors the formation of integrin αIIbβ3 adhesion contacts. Integrin αIIbβ3 engagement is associated with low-frequency calcium spikes that trigger tether restructuring, leading to the stabilization of discoid platelet aggregates. Ongoing discoid platelet recruitment drives the propagation of the thrombus in the downstream deceleration zone, which may in turn amplify the shear microgradient and promote further platelet aggregation. Thus, the shear microgradients caused by the thick struts induce platelet aggregation, formation of microthrombi with potential embolization, and micromyocardial necrosis (so-called a nidus of thrombus). The magnitude of flow disturbance depends on the degree of protrusion of the strut into the lumen. Therefore, thin struts could be a potential solution for the less flow disturbance and thus less thrombogenic status (lower panel). There is another cascade of von Willebrand factor (VWF)/GPIb activation, namely agglutination-elicited GPIb signaling.73 In contrast to shear stress–induced GPIb-elicited signaling, agglutination-elicited GPIb signaling that activates integrin αIIbβ3 requires thromboxane A2 (TXA2). Agglutination-elicited TXA2 production is independent of Ca2+ influx and mobilization of internal Ca2+ stores. [Powerpoint File]

Bioresorbable Scaffold: The Emerging Reality and Future Directions

Bioresorbable Scaffold: The Emerging Reality and Future Directions

Yohei Sotomi, Yoshinobu Onuma, Carlos Collet, Erhan Tenekecioglu, Renu Virmani, Neal S. Kleiman, Patrick W. Serruys

Design and optical coherence tomography (OCT) appearance of first and next generation bioresorbable scaffolds (BRSs). [Powerpoint File]

Circulating Noncoding RNAs as Biomarkers of Cardiovascular Disease and Injury

Circulating Noncoding RNAs as Biomarkers of Cardiovascular Disease and Injury

Janika Viereck, Thomas Thum

Biogenesis and function of microRNAs (miRNAs). MiRNAs are transcribed from longer precursors with protein or other noncoding gene sequences and further processed via 2 endonucleolytic processing steps. Mature miRNAs associate with Argonaute proteins (Ago2) forming the RNA-induced silencing complex (miRISC). Within this complex, miRNAs recognize their target sequence and block their expression by translational repression or degradation. MiRNAs can be released or actively secreted into the extracellular space and circulatory system stabilized in vesicles or proteinous binding partners. DGCR8, DiGeorge syndrome critical region 8; TRBP, transactivation-responsive RNA-binding protein. [Powerpoint File]

Circulating Noncoding RNAs as Biomarkers of Cardiovascular Disease and Injury

Circulating Noncoding RNAs as Biomarkers of Cardiovascular Disease and Injury

Janika Viereck, Thomas Thum

Molecular mechanism of long noncoding RNA (lncRNA) activities. LncRNAs regulate the expression of genes in the nucleus by interacting directly with DNA recruiting chromatin modifying complexes and various transcriptional regulators. Cytoplasmatic noncoding transcripts act as sponges for other transcripts like microRNAs (miRNAs) or for proteins, serve as templates for small peptide synthesis or regulate messenger RNA (mRNA) degradation and translation. These transcripts enter the bloodstream bound to proteinous carriers or incorporated into extracellular vesicles that leads to a stabilization of transcripts. Ago2, Argonaute protein 2. [Powerpoint File]

MicroRNA Biomarkers and Platelet Reactivity: The Clot Thickens

MicroRNA Biomarkers and Platelet Reactivity: The Clot Thickens

Nicholas Sunderland, Philipp Skroblin, Temo Barwari, Rachael P. Huntley, Ruifang Lu, Abhishek Joshi, Ruth C. Lovering, Manuel Mayr

MiRNAs and platelet reactivity. Association of miR-126 and miR-223 levels in platelet-poor plasma (PPP) to the VerifyNow P2Y12 aggregation test (A) and the vasodilator-stimulated phosphoprotein (VASP) phosphorylation assay (B) in patients on dual antiplatelet therapy for 30 days postacute coronary syndrome. Note that fewer samples were measured with the VerifyNow (n=39) compared with the VASP assay (n=123). PRU denotes P2Y12 reaction units (x axis). Higher PRU values reflect higher P2Y12-mediated platelet reactivity. [Powerpoint File]

MicroRNA Biomarkers and Platelet Reactivity: The Clot Thickens

MicroRNA Biomarkers and Platelet Reactivity: The Clot Thickens

Nicholas Sunderland, Philipp Skroblin, Temo Barwari, Rachael P. Huntley, Ruifang Lu, Abhishek Joshi, Ruth C. Lovering, Manuel Mayr

Choice of samples. A, Workflow for the preparation of platelet-poor plasma (PPP) from platelet-rich plasma (PRP). Addition of prostacyclin inhibits platelet activation during centrifugation. B, Concerns with regards to miRNA measurements in the different samples range from a contamination with leukocytes in PRP, proteolytic activity in serum to residual platelets in plasma. PRP and serum should reflect platelet miRNA content; PPP platelet miRNA release; plasma samples will reflect extracellular miRNA content but may additionally reflect either, platelet miRNA content or release. This depends on the amount of residual platelets in the plasma samples or platelet activation during plasma preparation, in particular during centrifugation. [Powerpoint File]

Pleiotropic Effects of Statins on the Cardiovascular System

Pleiotropic Effects of Statins on the Cardiovascular System

Adam Oesterle, Ulrich Laufs, James K. Liao

Regulation of the Rho GTPase cycle. Rho cycles between an inactive, cytoplasmic, GDP bound form and after geranylgeranylation is translocated to the plasma membrane and activated when it is bound to GTP. Inhibition of mevalonate synthesis by statins decreases geranylgeranyl (GG) pyrophosphate and prevents the geranylgeranylation of Rho and therefore its activation of Rho kinase (ROCK). ROCK mediates the downstream effects of Rho and has effect on endothelial cells, inflammatory cells, fibroblasts, cardiomyocytes, and vascular smooth muscle cells (SMC) that promote atherosclerosis and cardiac remodeling and may be responsible for the pleiotropic effects of statins. CRD indicates cysteine-rich domain; eNOS, endothelial nitric oxide synthase; GAP, GTPase-activating proteins; GDI, guanine nucleotide dissociation inhibitors; GEF, guanine nucleotide exchange factors; PI3K, phosphatidylinositol 3-kinase; and RBD, Rho-binding domain. Illustration Credit: Ben Smith. [Powerpoint File]

Protein Interactions at Endothelial Junctions and Signaling Mechanisms Regulating Endothelial Permeability

Protein Interactions at Endothelial Junctions and Signaling Mechanisms Regulating Endothelial Permeability

Yulia A. Komarova, Kevin Kruse, Dolly Mehta, Asrar B. Malik

Mechanotransduction at adherens junctions (AJs). The mechanosensory complex in endothelial cells is composed of vascular endothelial (VE)-cadherin, platelet endothelial cell adhesion molecule (PECAM)-1, and vascular endothelial growth factor receptor (VEGFR) 2. Mechanosensing of shear stress occurs through PECAM-1–dependent activation of Fyn, which in turn facilitates VEGFR2-mediated signaling in a ligand-independent manner and activates phosphatidylinositol-4,5-bisphosphate 3-kinase (PI3K). PI3K activates both Rac1 and endothelial nitric oxide synthase (eNOS) signaling pathways. Rac1 relieves tension at AJs, whereas NO concomitantly promotes vasorelaxation of smooth muscle cells. PECAM-1–dependent sensing of shear stress also promotes α2β1 integrin signaling and consequently activation of protein kinase A (PKA) in atheroresistant regions. PKA phosphorylates RhoA and decreases RhoA-dependent cellular stiffness allowing the endothelial cell to align in the direction of blood flow. GPCR indicates G-protein–coupled receptor; and sGC, soluble guanylyl cyclase. [Powerpoint File]

Protein Interactions at Endothelial Junctions and Signaling Mechanisms Regulating Endothelial Permeability

Protein Interactions at Endothelial Junctions and Signaling Mechanisms Regulating Endothelial Permeability

Yulia A. Komarova, Kevin Kruse, Dolly Mehta, Asrar B. Malik

Role of actomyosin apparatus in stabilizing adherens junctions (AJs). A, Domain structure of nonmuscle myosin-II (NM-II). The NM-II consists of a globular head domain containing both actin-binding and motor domains, essential light chains (ELCs), regulatory light chains (RLCs), and heavy chains. NM-II possesses a head-to-tail interaction in the absence of phosphorylation. Phosphorylation of regulatory light chain at Thr18/Ser19 by myosin light chain kinase (MLCK) unfolds the molecule, enabling assembly of antiparallel filaments through interactions between their rod domains. Activation of Rho-associated kinase (ROCK), which inhibits phosphatase activity of myosin light chain phosphatase (MLCP) in a phosphorylation-dependent manner, also favors RLC phosphorylation. NM-II filaments bind to actin filaments, which slide along each other, and cause a cell contraction. Modified from Vicente-Manzanares et al415 with permission of the publisher. Copyright ©2009, Nature Publishing Group. B, Proposed mechanism of regulation of NM-II activity at AJs in confluent endothelium. NM-II regulates attachment of the vascular endothelial (VE)-cadherin adhesion complex to the actin cytoskeleton, thereby generating mechanical tension required for binding of α-catenin to both β-catenin and f-actin. NM-II phosphorylation is controlled by MLCK and MLCP activities. In the model, we propose that Src and Cdc42 pathways cooperate in regulating NM-II activity at AJs. Cdc42 facilitates activation of NM-II through myotonic dystrophy kinase–related Cdc42-binding kinase (MRCK)–dependent phosphorylation of MLCP, whereas Src phosphorylates MLCK at sites of VE-cadherin adhesion. CaM indicates calmodulin; GAP, GTPase-activating protein; and GEF, guanine nucleotide exchange factor. [Powerpoint File]

Protein Interactions at Endothelial Junctions and Signaling Mechanisms Regulating Endothelial Permeability

Protein Interactions at Endothelial Junctions and Signaling Mechanisms Regulating Endothelial Permeability

Yulia A. Komarova, Kevin Kruse, Dolly Mehta, Asrar B. Malik

Composition of interendothelial junctions. A, Schematic representation of interendothelial junctions composed of tight junctions (TJs), adherens junctions (AJs), and gap junctions (GJs). TJs are mediated by adhesion proteins such as claudins, occludin, and junctional adhesion molecules (JAMs), whereas the zona occludin proteins (ZO-1, ZO-2, and ZO-3) connect adhesion molecules to the actin cytoskeleton. AJs are composed of vascular endothelial (VE)-cadherin and associated β- and p120-catenins. α-catenin binds β-catenin to connect AJs to the actin cytoskeleton. GJs are composed of 2 connexin hexamers forming hemichannels. B, VE-cadherin mediates adhesion by trans-dimerization of tryptophan 2 and tryptophan 4 residues in a hydrophobic pocket of the opposing VE-cadherin molecule. Ribbon presentations of VE-cadherin trans-dimer. C, Strand swap dimerization occurs through the insertion of a tryptophan residue into the hydrophobic binding pocket of the opposing cadherin. D, Trans-dimerization (between endothelial cell [EC] 1 and EC1 of opposing cadherins) orients VE-cadherin molecules and facilitates cis interactions (between EC1 and EC2 of neighboring cadherins). Panels B, C, and D are modified from Brasch et al115 with permission of the publisher. Copyright ©2012, Elsevier. [Powerpoint File]

Inflammatory Disequilibrium in Stroke

Inflammatory Disequilibrium in Stroke

Danica Petrovic-Djergovic*, Sascha N. Goonewardena*, David J. Pinsky

Inflammatory disequilibrium after stroke. After brain ischemia, circulating cells such as neutrophils, monocytes (innate immune cells), T cells (adaptive immune cells) interact with platelets, causing sludging and cessation of cerebral blood flow and tissue hypoxia. Acute ischemia refers to the early phases (minutes to hours) and delayed refers to late phases of the ischemic process (hours to days). Endothelial cell activation promotes immune cell transmigration into the injured brain parenchyma. In acute phase of brain ischemia, infiltrating immune cells (neutrophils, monocytes, dendritic cells, and T cells) and resident brain cells (microglia and astrocytes) are activated and promote tissue injury. Activated monocytes adhere to activated endothelium and after transmigration into the brain tissue transform into the blood-borne macrophages. Macrophages generate reactive oxygen intermediates, secrete proinflammatory cytokines, upregulate costimulatory molecules (CD80 shown here), and create a prothrombotic environment. Similarly, microglia migrates toward the lesion site early on after the insult and secretes proinflammatory mediators that cause additional injury; however, microglia promotes tissue repair and remodeling through debris phagocytosis. Astrocytes can secrete both proinflammatory (CXC-chemokine ligand [CXCL10], monocyte chemoattractant protein-1 [MCP-1]) as well anti-inflammatory (interleukin [IL]-10, transforming growth factor-β [TGF-β]) chemokines/cytokines to promote injury or repair, respectively; IL-23 produced by macrophages/microglia activate γδT-cells, which contributes to tissue injury via secretion of IL-17; Naïve T-cells (CD4/CD8) contribute to brain injury in an antigen unspecific manner, possibly via interferon-γ (INF-γ), reactive oxygen species (ROS), and perforin. Other T cells such as T-regulatory cells (T-regs) induce brain tissue repair via IL-10 secretion, inhibition of the effector T-cells response, neurogenesis, and central nervous system (CNS) tissue antigen tolerization. In the delayed phase after brain ischemia, antigen-presenting cells (APC), that is, macrophages, microglia, astrocytes, and dendritic cells (not depicted in the schematic), present CNS antigens to CD4+ or CD8+ T cells in antigen-specific manner in context of major histocompatibility complex (MHC) class I or II. Purinergic mediators such as ATP and adenosine act as an extracellular danger signals differentially influencing immune responses in stroke. Ectoenzymes, cluster of differentiation 39 (CD39), and cluster of differentiation 73 (CD73) act in tandem to dissipate proinflammatory and generate anti-inflammatory mediators. Brain resident cells and certain immune cells express these ectoenzymes. ICAM indicates intercellular adhesion molecule-1; MMP, matrix metalloproteinases; mo-MФs, monocytes–macrophages; and NET, neutrophil extracellular trap. [Powerpoint File]

Resolution of Acute Inflammation and the Role of Resolvins in Immunity, Thrombosis, and Vascular Biology

Resolution of Acute Inflammation and the Role of Resolvins in Immunity, Thrombosis, and Vascular Biology

Brian E. Sansbury, Matthew Spite

Biological actions of resolvins and other specialized proresolving lipid mediators (SPMs) related to cardiovascular inflammation. During acute inflammation or infection, SPMs are generated during leukocyte–endothelial interactions and directly block polymorphonuclear neutrophil (PMN) chemotaxis. By directly targeting the endothelial cells (ECs) of the vasculature (eg, postcapillary venules), SPMs also prevent the capture, rolling, and adhesion of PMN to the endothelial monolayer and block their extravasation across the endothelium. In addition, they stimulate production of nitric oxide (NO) and prostacyclin (PGI2) from EC. In the tissue, SPMs promote the protective actions of leukocytes by enhancing the phagocytosis of bacteria and cellular debris and macrophage efferocytosis of apoptotic PMN. In the context of atherosclerosis, SPMs decrease EC production of leukocyte chemoattractants and adhesion molecules, potentially halting further recruitment of leukocytes to the vessel wall. They also potently prevent platelet aggregation and thrombosis. In the subendothelium, enhanced macrophage efferocytosis and promotion of a resolving macrophage phenotype contributes to a more stable plaque with decreased necrosis. During pathological inward remodeling (ie, restenosis), SPMs combat neointimal hyperplasia by inhibiting vascular smooth muscle cell (VSMC) proliferation and migration while simultaneously preventing monocyte adhesion and inflammatory signaling. IL indicates interleukin; and MCP, monocyte chemoattractant protein. [Powerpoint File]

Resolution of Acute Inflammation and the Role of Resolvins in Immunity, Thrombosis, and Vascular Biology

Resolution of Acute Inflammation and the Role of Resolvins in Immunity, Thrombosis, and Vascular Biology

Brian E. Sansbury, Matthew Spite

The coordinated temporal events of self-limited acute inflammation. The ideal outcome of an acute inflammatory response is complete resolution. The inflammatory response can be divided into 2 general phases: initiation and resolution. Critical to progressing from initiation to resolution is the temporal switch in lipid mediators that are biosynthesized by leukocytes in the tissue, a process known as lipid mediator class switching. The earliest stage of the inflammatory response is marked by tissue edema caused by increased blood flow and microvascular permeability and is mediated by the release of proinflammatory lipid mediators including the cysteinyl leukotrienes and prostaglandins. Polymorphonuclear neutrophils (PMN) infiltrate in response to lipid mediators including leukotriene B4 and engulf and degrade pathogens. Subsequently, PMN undergo apoptosis and also switch from releasing proinflammatory mediators to proresolving mediators (eg, resolvins) that signal the clearance of apoptotic cells by macrophages (MΦ) in an anti-inflammatory process termed efferocytosis. In addition to promoting efferocytosis, proresolving lipid mediators halt further PMN recruitment and stimulate a proresolving macrophage phenotype that is important for tissue repair. [Powerpoint File]

Systems Analysis of Thrombus Formation

Systems Analysis of Thrombus Formation

Scott L. Diamond

Systems biology of thrombosis. The computer simulation of clotting requires a multiscale and integrated description of platelet signaling and adhesion, coagulation kinetics, and hemodyamics. Platelet signaling is driven by soluble activators (ADP, thromboxane A2 [TXA2], thrombin), soluble inhibitors (NO, prostacyclin [PGI2] and insoluble activators (collagen) to drive intracellular calcium mobilization. Calcium mobilization occurs rapidly through IP3-mediated release and store-operated calcium entry (STIM1-Orai1). Dense platelet deposits in clots result in significant ADP and thromboxane and thrombin-driven signaling, often targeted by inhibition of P2Y12, cyclooxygenase (COX)-1, and PAR1, respectively. During coagulation, the generation of thrombin (FIIa) is primarily driven by TF/FVIIa (extrinsic tenase) via subsequent engagement of the FIXa/VIIIa (intrinsic tenase) and FXaVa (prothrombinase). Thrombin has significant regulatory control on its own production through activation of FVIIIa, FVa, FXIa, as well as regulation of fibrin through activation of FXIIIa that crosslinks fibrin. Local hemodynamics (inset; reprinted from Taylor et al3 with permission of the publisher. Copyright ©2013, Elsevier.) can be determined by computational fluid dynamics to define locations with at-risk plaque burden, stenosis, and high wall shear stress. Full systems biology models of platelet activation and coagulation in a patient-specific flow field are directed at simulation of acute coronary syndromes (bottom). AC indicates adenylate cyclase; AP, antiplasmin; APC, activated protein C; ATIII, antithrombin III; catG, cathepsin G; CTI, corn trypsin inhibitor; DTS, dense tubular system; EC, endothelial cell; FDP, fibrin degradation product; GC, guanylate cyclase; GPVI, glycoprotein VI; HMWK, high molecular weight kininogen; HNE, human neutrophil elastase; IP, prostacyclin receptor; MG, macroglobulin; NO, nitric oxide; P2Y purinergic receptor type 2; PAI, plasminogen activator inhibitor; PAR, protease activated receptor; PC, protein C; PLC, phospholipase; PMCA, plasma membrane calcium ATPase; PN2, proteonexin 2; PS, protein S; TM, thrombomodulin; TP, thromboxane receptor; tPA, tissue-type plasminogen activator; and uPA, urokinase plasminogen activator. [Powerpoint File]

Animal Models of Thrombosis From Zebrafish to Nonhuman Primates: Use in the Elucidation of New Pathologic Pathways and the Development of Antithrombotic Drugs

Animal Models of Thrombosis From Zebrafish to Nonhuman Primates: Use in the Elucidation of New Pathologic Pathways and the Development of Antithrombotic Drugs

Pudur Jagadeeswaran, Brian C. Cooley, Peter L. Gross, Nigel Mackman

Formation of an occlusive thrombus. After vessel injury platelets rapidly adhere to collagen and deposited von Willebrand Factor. The adhered platelets are activated by primary and secondary activators that lead to platelet aggregation mediated by various ligands, including fibrinogen. In parallel to platelet activation, the clotting system is activated by exposure of tissue factor (TF) in the vessel wall. In addition, factor XII may contribute to the activation of coagulation. Thrombin is the central protease of the coagulation cascade and cleaves fibrinogen to fibrin monomers that are crosslinked into a network by factor XIIIa. There is cross talk between the platelet and coagulation cascades. For instance, activated platelets provide a thrombogenic surface for the assembly of various coagulation protease complexes and thrombin is a potent activator of platelets by cleavage of protease activated receptors. Formation of an occlusive thrombosis will block blood flow. [Powerpoint File]