The serum- and glucocorticoid-inducible kinase 1 (SGK1) influences platelet calcium signaling and function by regulation of Orai1 expression in megakaryocytes

Abstract

IntroductionPlatelet adhesion, activation, and aggregation are essential for primary hemostasis at sites of vascular injury but are also critically important for the development of acute thrombotic occlusion at regions of atherosclerotic plaque rupture, the major pathophysiologic mechanism underlying myocardial infarction and ischemic stroke. 1 Platelet activation is triggered by various agonists, including subendothelial collagen, ADP released from activated platelets, thrombin generated by the coagulation cascade, or the collagen receptor glycoprotein VI (GPVI)-specific agonists convulxin (CVX) and collagen-related peptide (CRP). 2 The agonists lead to platelet granule release, integrin ␣ IIb ␤ 3 activation, phosphatidylserine exposure, aggregation, and thrombus formation. 2 All those platelet responses depend on an increase of cytosolic Ca 2ϩ concentration ([Ca 2ϩ ] i ), 3,4 which is accomplished by inositol-1,4,5-triphosphatemediated Ca 2ϩ release from intracellular stores triggering subsequent stimulation of store-operated Ca 2ϩ entry (SOCE) across the plasma membrane. 5 Two key players in platelet SOCE have recently been identified: The 4-transmembrane-spanning poreforming calcium release-activated channel moiety Orai1, which mediates entry of extracellular Ca 2ϩ , and stromal interaction molecule 1 (STIM1), an Orai1 regulating Ca 2ϩ sensor expressed predominantly in the endoplasmic reticulum. [6][7][8] Regulators of Orai1 in other cell types include receptor for activated protein kinase C-1, 9 reactive oxygen species, 10 and lipid rafts. 11 However, regulation of Orai1 in platelets is poorly understood. Platelet activation has been shown to be regulated in vitro and in vivo by the PI3K/Akt signaling cascade. 12,13 Interference with PI3K signaling has previously been shown to compromise Ca 2ϩ influx into platelets. 14,15 Signaling molecules regulated by PI3K signaling include the serum-and glucocorticoid-inducible kinase 1 (SGK1), a kinase belonging to the AGC family of serine/threonine protein kinases. 16,17 SGK1 has originally been cloned as a glucocorticoidsensitive gene but later shown to be regulated by a variety of hormones and other triggers, including thrombin, growth factors IGF-1 and TGF-␤, oxidative stress, and ischemia. 17 SGK1 has previously been reported to regulate a wide variety of carriers and ion channels, including the epithelial Ca 2ϩ channels TRPV5 and TRPV6. 17 Most recently, SGK1 has been shown to be critically important for the Ca 2ϩ entry into mast cells after activation of the IgE receptor, 18 an effect mediated by regulation of Orai1. 19 Furthermore, SGK1 participates in the regulation of renal tubular Na ϩ reabsorption, salt appetite, and thus blood pressure. 17 A gain-of-function SGK1 gene variant, the combined presence of single nucleotide polymorphism in intron 6 (rs1743966) and in exon 8 (rs1057293), is associated with enhanced blood pressure. 20 Submitted June 9, 2011; accepted August 28, 2011. Prepublished online as Blood First Edition paper, October 26, 2011; DOI 10.1182...

Intracellular cyclophilin A is an important Ca2+ regulator in platelets and critically involved in arterial thrombus formation

“…11 It is known that the inhibition of PPIase and chaperone activity of cyclophilins by CsA is able to impair protein maturation and folding followed by reduced level of functional expression. 38 To ensure that impaired Ca 2ϩ mobilization in CyPAdeficient platelets is not caused by reduced expression level of either STIM1 or Orai1 as observed in sgk1 Ϫ/Ϫ mice, 46 we confirmed unaltered expression levels of STIM1 and Orai1 in CyPAdeficient platelets (supplemental Figure 4). To determine whether the enzymatic activity of CyPA has any regulatory role in STIM1 activity, we analyzed colocalization of STIM1 and CyPA by immunoprecipitation experiments.…”

“…10,13,46 STIM1 deficiency reduced Ca 2ϩ efflux from intracellular stores and SOCE in a manner similar to that observed in Cypa Ϫ/Ϫ platelets. 11 It is known that the inhibition of PPIase and chaperone activity of cyclophilins by CsA is able to impair protein maturation and folding followed by reduced level of functional expression.…”

Indoxyl sulfate induces platelet hyperactivity and contributes to chronic kidney disease–associated thrombosis in mice

“…22 After anesthesia, mouse blood was collected from the inferior vena cava with acid citrate dextrose (ACD) (65 mM trisodium citrate, 70 mM citric acid, and 100 mM dextrose; pH 6.5) as the anticoagulant. To detect the expression of P-selectin and activated integrin GPIIb/ IIIa on platelet surface, ACD-whole blood was diluted in modified Tyrode buffer (137 mM NaCl, 2.7 mM KCl, 12 mM NaHCO 3 , 0.4 mM NaH 2 PO 4 , 5 mM N-2-hydroxyethylpiperazine-N9-2-ethanesulfonic acid, 5.6 mM glucose, and 0.35% bovine serum albumin; pH 7.4).…”

SGK1 inhibitor reverses hyperglycemia partly through decreasing glucose absorption

“…Various functions of SGK1 have been identified (Salker et al 2011, Borst et al 2012, Das et al 2012, Hall et al 2012. Compelling evidence indicates a role for SGK1 in the development of metabolic syndrome (Lang et al 2009, Lang & Voelkl 2013.…”