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Human arterial endothelial cells (HAECs) regulate their phenotype by integrating signals encoded in the frictional forces exerted by flowing blood, fluid shear stress (FSS). High laminar FSS promotes establishment of adaptive HAEC phenotype protective against atherosclerosis, whereas low or disturbed FSS cause HAECs to adopt atheroprone phenotypes. A vascular endothelial cadherin (VE cadherin)-based mechanosensory complex allows HAECs to regulate barrier function, cell morphology,/ and gene expression in response to FSS. Previously, we reported that this mechanosensor integrated exchange protein activated by cAMP (EPAC1) and a PDE4D gene derived cyclic nucleotide phosphodiesterase (PDE), but had not identified the PDE4D variant involved. Our hypothesis here was that only one of the two ~100 kDa PDE4D variants expressed in HAECs coordinated these responses. Now, we show one unique PDE4D splice variant, PDE4D7, controls transcriptional responses of HAECs to FSS while another, PDE4D5, does not. Adaptive transcriptional responses of HAECs subjected to laminar FSS in vitro were blunted in cells in which PDE4D7 was silenced, but unaffected in cells with silenced PDE4D5. This work identifies a specific therapeutic target for the treatment or prevention of atherosclerosis and improves our understanding of the role of cAMP signaling in modulating mechanosensory signal transduction in the vascular endothelium. Key words: endothelium, fluid shear stress, nitric oxide synthase, phosphodiesterase, cAMP, atherosclerosis, mechanosensory. Les cellules endotheliales arterielles humaines (CEAH) parviennent a reguler leur phenotype en integrant des signaux codes issus de forces de friction exercees par le flux sanguin et des forces de cisaillement engendrees par les liquides (FCL). Des FCL laminaires elevees favorisent l'etablissement d'un phenotype adaptatif protecteur des CEAH contre l'atherosclerose, tandis que des FCL faibles ou desordonnees font en sorte que les CEAH adoptent des phenotypes enclins a l'atherosclerose. Un complexe de capteur mecanique fonde sur les cadherines de l'endothelium vasculaire (cadherine EV) permet aux CEAH de reguler leur fonction de barriere, la morphologie des cellules et l'expression genique en reaction aux FCL. Precedemment, nous avons rapporte que ce capteur mecanique integrait l'EPAC1 (pour << exchange protein activated by cAMP >>) et une phosphodiesterase (PDE) de nucleotides cycliques issue du gene PDE4D, mais nous n'avions pas etabli quelle variante de PDE4D jouait un role. Ici, notre hypothese voulait qu'uniquement l'une des deux variantes de PDE4D d'environ 100 kDa exprimees dans les CEAH coordonnait ces reactions. Nous montrons maintenant que les reactions transcriptionnelles des CEAH aux FCL sont sous le controle d'un variant d'epissage PDE4D unique, le PDE4D7, mais pas d'un autre, le PDE4D5. Les reactions transcriptionnelles adaptatives des CEAH subissant des FCL laminaires in vitro s'attenuaient dans les cellules ou PDE4D7 etait reprime, mais n'etaient pas affectees dans les cellules ou PDE4D5 etait reprime. Ces travaux designent une cible therapeutique specifique pour le traitement ou la prevention de l'atherosclerose, et permettent d'ameliorer notre comprehension du role de la signalisation par l'AMPc dans la modulation de la transduction des signaux mecanosensoriels dans l'endothelium vasculaire. [Traduit par la Redaction] Mots-cles : endothelium, forces de cisaillement engendrees par les liquides, oxyde nitrique synthase, phosphodiesterase, AMPc, atherosclerose, mecanosensoriel., Introduction Atherosclerosis is a chronic progressive disease involving the buildup of plaque in the arterial wall and leads to coronary artery disease, the principal cause of global mortality (Benjamin et [...]

Numerous cAMP-elevating agents regulate events required for efficient migration of arterial vascular smooth muscle cells (VSMCs). Interestingly, when the impact of cAMP-elevating agents on individual migration-related events is studied, these agents have been shown to have distinct, and sometimes unexpected, effects. For example, although cAMP-elevating agents inhibit overall migration, they promote VSMC adhesion to extracellular matrix proteins and the formation of membrane extensions, which are both events that are essential for and promote migration. Herein, we extend previous observations that identified phosphodiesterase-4D3 (PDE4D3) as an integral component of a PKA/A kinase-anchoring protein (AKAP) complex in cultured/hypertrophied rat cardiac myocytes to the case for nonhypertrophied cardiac myocytes. Moreover, we show that while rat aortic VSMCs also express PDE4D3, this protein is not detected in PKA/ AKAP complexes isolated from these cells. In contrast, we show that another PDE4D splice variant expressed in arterial vascular myocytes, namely, PDE4D8, integrates into PKA/AKAP-based signaling complexes in VSMCs. Consistent with the idea that a PDE4D8/PKA/ AKAP complex regulates specific VSMC functions, PKA and PDFAD8 were each recruited to leading-edge structures in migrating VSMCs, and inhibition of PDE4D8 recruitment to pseudopodia of migrating cells caused localized changes in actin dynamics. Our data are presented in the context that cardiac myocytes and arterial VSMCs may use distinct PDFAD variants to regulate selected pools of targeted PKA activity and that disruption of this complex may allow selective regulation of cAMP-dependent events between these two cardiovascular cell types. migration; vascular smooth muscle cell

An elevated circulating level of the adipocyte-derived satiety hormone leptin is an independent risk factor for cardiovascular disease. Because thrombus formation is a major cause of acute coronary events and leptin was shown previously to facilitate ADP-induced platelet aggregation, we chose to define the signaling events involved in leptin-mediated platelet activation. Using pharmacological, biochemical, and cell biological approaches, we show that leptin-induced platelet activation required activation of a signaling cascade that included the long form of the leptin receptor, three kinases [Janus kinase 2 (JAK2), phosphatidylinositol 3-kinase (PI3K), and protein kinase B (PKB/Akt)], the insulin receptor substrate-1 (IRS-1), and the major human platelet cAMP phosphodiesterase phosphodiesterase 3A (PDE3A). Moreover, we identify a role for an intraplatelet LEPR/JAK2/IRS-1/PI3K/PKB/ PDE3A molecular complex that allows for the selective leptin-mediated activation of platelets. Our data demonstrate that leptin promotes platelet activation, provides a mechanistic basis for the prothromhotic effect of this hormone, and identifies a potentially novel therapeutic avenue to limit obesity-associated cardiovascular disease. cyclic adenosine 3',5'-monophosphate; aggregation; adhesion; signaling

Skeletal muscle insulin resistance is associated with many common age-related diseases, but moderate calorie restriction (CR) can substantially elevate glucose uptake by insulin-stimulated skeletal muscle from both young and old rats. The current study evaluated the isolated epitrochlearis muscle from ~24.5-mo-old rats that were either fed ad libitum (AL) or subjected to CR (consuming ~65% of ad libitum, AL, intake beginning at ~22.5 mo old). Some muscles were also incubated with MK-2206, a potent and selective Akt inhibitor. The most important results were that in isolated muscles, CR vs. AL resulted in 1) greater insulinstimulated glucose uptake 2) that was accompanied by significantly increased insulin-mediated activation of Akt2, as indicated by greater phosphorylation on both Thr309 and Ser474 along with greater Akt2 activity, 3) concomitant with enhanced phosphorylation of several Akt substrates, including an Akt substrate of 160 kDa on Thr642 and Ser588, filamin C on Ser2213 and proline-rich Akt substrate of 40 kDa on Thr246, but not TBC1D1 on Thr596; and 4) each of the CR effects was eliminated by MK-2206. These data provide compelling new evidence linking greater Akt2 activation to the CR-induced elevation of insulin-stimulated glucose uptake by muscle from old animals. [ABSTRACT FROM AUTHOR]