Page 1910 - Williams Hematology ( PDFDrive )

1884 Part XII: Hemostasis and Thrombosis Chapter 112: Platelet Morphology, Biochemistry, and Function 1885

may facilitate the transmission of cytoskeletal tension from inside Ca -mobilization. 1777 PI metabolism is also affected, as the activities of
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the platelet to outside, and thus initiate clot retraction. Recombinant, both PLC and PLA are suppressed. 1778 Moreover, PKA also phospho-
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mutated integrin β that cannot be phosphorylated is unable to support rylates Raf kinase on three sites, which inhibits Raf kinase function in
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extensive clot retraction when expressed in a cell line. Other pro- part by inhibiting its binding to the activating protein RasGTP. 1779,1780
teins that bind to the diphosphorylated integrin β cytoplasmic domain Finally, the small G protein, Rap1b, which contributes to integrin α β
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include the SHC adapter proteins, which also become tyrosine phos- activation, 1611 is phosphorylated by PKA, 1781 although it appears that this
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phorylated during platelet aggregation. Therefore, it is possible that phosphorylation event does not inhibit platelet function 1782 and may, in
the SHC adapter proteins may link diphosphorylated integrin β to the fact, contribute to Rap1b activation. 1783
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MAPK pathway. 906,1762 Mice containing mutated integrin β molecules Paradoxically, in contrast to the inhibitory effects of high levels of
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that cannot be phosphorylated exhibit a mild bleeding disorder as evi- PGE , low levels of PGE (<10 M) potentiate agonist-induced plate-
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denced by occasional rebleeding of tail cuts. Moreover platelets derived let aggregation by acting via the EP3 receptor to decrease intraplatelet
from these mice form abnormally loose thrombi when activated by cAMP levels. 1784,1785 Mice lacking the EP3 receptor are protected from
shear forces. 1763 Other integrin β cytoplasmic domain binding proteins AA-induced thrombosis 1786 ; thus it is possible that PGE present in ath-
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have been described, including skelemin, a member of a family of pro- erosclerotic lesions contributes to atherothrombosis.
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teins that regulate myosin, and talin.
Some signaling events that occur downstream of integrin α β Nitric Oxide
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require only integrin clustering, whereas other events require clustering, NO is synthesized from L-arginine by NO synthase in endothelial cells,
ligand binding, and/or platelet aggregation. For example, the tyrosine platelets, and other cells. The formation of NO is enhanced at sites of
kinase Syk becomes activated in response to integrin α β clustering, shear stress and by platelet agonists (e.g., thrombin or ADP), 1787 and it
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independent of cytoskeletal assembly, whereas activation of the tyrosine readily diffuses into platelets. 1788,1789 Similar to PGI or PGE , NO pre-
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kinase FAK requires integrin clustering, ligand binding and cytoskel- treatment of platelets inhibits platelet activation and can reverse platelet
etal assembly. 1764 In studies conducted in cell lines, activation of Syk aggregation soon after initiation. However, NO works not by elevating
downstream of integrin α β leads to phosphorylation of Vav1, a gua- cAMP, but instead by increasing cGMP. 1790 NO synthase activity in
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nine nucleotide exchange factor for Rac, and lamellipodia formation. platelets increases during platelet activation, suggesting that NO pro-
Syk and Vav1 cooperate to activate Jun N-terminal kinase, ERK2, and duction is a normal negative feedback mechanism that limits further
Akt. 1764 These pathways are also likely to be involved in postaggregation platelet aggregation. NO and PGI act together synergistically to inhibit
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events in the platelet. platelet activation. 1791
Proteins other than the well-described integrin α β ligands Elevation in intracellular cGMP levels activates cGMP-dependent
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fibrinogen and VWF also induce signaling events via binding to inte- PKG, whose downstream targets include ERK and the TXA recep-
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grin α β . One such protein is CD40L, a TNF family member that is tor. 1792 In mice, the absence of PKG results in enhanced platelet accu-
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expressed on a variety of cells including activated platelets. Platelets mulation along damaged vessels after ischemic injury, supporting an
are also the major source of a soluble form of CD40L. 1765 In addition important role for PKG in platelet deposition. 1793 VASP, a member of the
to binding to its classical receptor, CD40, CD40L also binds to integrin Pro-rich, actin-regulatory Ena/VASP protein family, is phosphorylated
α β on platelets and induces signaling events 1350 that are required for in response to elevations in either cAMP or cGMP 1794 and both PKA
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normal arterial thrombus formation in mice. 1348 CD40L may also initi- and PKG phosphorylate VASP in vitro. 1795 A role for VASP in inhibition
ate platelet aggregation by binding to CD40 on platelets. 1349 of platelet function was established in studies of VASP-deficient mice:
platelets obtained from the mice display increased P-selectin expression
INHIBITORY PATHWAYS IN PLATELETS and integrin α β activation in response to agonists, 1796 and platelet
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adhesion at sites of vascular injury or atherosclerosis is enhanced in
Prostaglandins VASP-deficient mice. 1797 The enhanced platelet adhesion in VASP-null
Prostaglandins that inhibit platelet activation include PGE (at high mice is not corrected by NO, suggesting that VASP may be a key nega-
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concentrations) and PGI (at low concentrations) (also termed prosta- tive regulator of platelet function in the cGMP-mediated pathways.
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cyclin) (reviewed in Refs. 1766 and 1767). In the vasculature, the endo- Elevation in intracellular cGMP can also increase cAMP levels
thelium produces PGI and PGE , which are important in maintaining via inhibition of PDE activity. 1798 This crosstalk between cGMP and
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vascular patency. 1768 Inhibition is initiated by the binding of these PGs cAMP-dependent pathways may synergize to contribute to the inhib-
to their own specific GPCR. 1769,1770 PG receptor occupancy converts the itory effects of NO on platelet function.
Gα subunit to the GTP bound, active form, which then activates ade-
nylyl cyclase. Adenylyl cyclase catalyzes the formation of cAMP. The CD39 (ATP Diphosphohydrolase; Ecto-ADPase)
exact amount of cAMP present in the cell is also determined by its rate Vascular endothelium regulates platelet function by producing pros-
of breakdown by phosphodiesterases (PDEs). Biochemical studies and tacyclin and NO, as well as by expressing CD39 NTPDase1, a plasma
studies from gene targeted mice support a primary role for PDE3A in membrane-associated ectonucleotide (ATP diphosphohydrolase; ATP-
platelets. 1771–1773 Therefore, agents that inhibit PDE, such as theophylline, Dase; ecto-ADPase; EC 3.6.1.5) that converts extracellular ATP to ADP,
caffeine, and the drug cilostazol, also elevate cAMP levels in platelets and ADP to AMP. 1799–1801 CD39 limits the platelet-activating effects of
and other cells. 1774 cAMP then activates PKA, which phosphorylates ADP released by damaged tissues, RBCs, and activated platelets; further-
specific target proteins. PKA inhibits platelet activation by several path- more, AMP generated by CD39 is degraded by an ecto-5′-nucleotidase
ways. One mechanism involves PKA-dependent phosphorylation of (CD73; EC 3.1.3.5) to adenosine, an inhibitor of ADP-induced platelet
VASP (discussed under “Nitric Oxide” below). A separate mechanism activation, that increases cAMP binding to the A2a adenosine receptor
involves the phosphorylation and inhibition of Gα13, which couples to on platelets. 1802 Adenosine deaminase (EC 3.5.4.4) degrades adenosine
the TXA receptor, thus impairing this activation pathway. 1775 Also, PKA to inosine. CD39 is a Mr 95,000 cell-surface glycoprotein expressed on
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phosphorylates GPIbβ on Ser 166, and negatively regulates the ability of endothelial cells, subsets of activated natural killer (NK) cells, B cells,
GPIb to bind VWF. 1776 In addition, PKA may phosphorylate and inhibit monocytes, and T cells. Small amounts may also be on platelets and
the IP receptor, which would repress agonist-induced intracellular erythrocytes. It is present in the lymphocytes in chronic lymphocytic
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Kaushansky_chapter 112_p1829-1914.indd 1885 17/09/15 3:30 pm