Abstract-Upon vascular injury, platelets are activated by adhesion to adhesive proteins, such as von Willebrand factor and collagen, or by soluble platelet agonists, such as ADP, thrombin, and thromboxane A 2 . These adhesive proteins and soluble agonists induce signal transduction via their respective receptors. The various receptor-specific platelet activation signaling pathways converge into common signaling events that stimulate platelet shape change and granule secretion and ultimately induce the "inside-out" signaling process leading to activation of the ligand-binding function of integrin ␣ IIb ␤ 3 . Ligand binding to integrin ␣ IIb ␤ 3 mediates platelet adhesion and aggregation and triggers "outside-in" signaling, resulting in platelet spreading, additional granule secretion, stabilization of platelet adhesion and aggregation, and clot retraction. It has become increasingly evident that agonist-induced platelet activation signals also cross talk with integrin outside-in signals to regulate platelet responses. Platelet activation involves a series of rapid positive feedback loops that greatly amplify initial activation signals and enable robust platelet recruitment and thrombus stabilization. Recent studies have provided novel insight into the molecular mechanisms of these processes.

Aim Although it is established that peri-implantitis is a bacterially induced disease, little is known about the bacterial profile of peri-implant communities in health and disease. The purpose of the present investigation was to examine the microbial signatures of the peri-implant microbiome in health and disease. Materials and methods Subgingival and submucosal plaque samples were collected from forty subjects with periodontitis, peri-implantitis, periodontal and peri-implant health and analyzed using 16S pyrosequencing. Results Peri-implant biofilms demonstrated significantly lower diversity than subgingival biofilms in both health and disease, however, several species, including previously unsuspected and unknown organisms, were unique to this niche. The predominant species in peri-implant communities belonged to the genera Butyrivibrio, Campylobacter, Eubacterium, Prevotella, Selenomonas, Streptococcus, Actinomyces, Leptotrichia, Propionibacterium, Peptococcus, Lactococcus and Treponema. Peri-implant disease was associated with lower levels of Prevotella and Leptotrichia and higher levels of Actinomyces, Peptococcus, Campylobacter, non-mutans Streptococcus, Butyrivibrio, and Streptococcus mutans than healthy implants. These communities also demonstrated lower levels of Prevotella, non-mutans Streptococcus, Lactobacillus, Selenomonas, Leptotrichia, Actinomyces and higher levels of Peptococcus, Mycoplasma, Eubacterium, Campylobacter, Butyrivibrio, Streptococcus mutans, and Treponema when compared to periodontitis-associated biofilms. Conclusion The peri-implant microbiome differs significantly from the periodontal community in both health and disease. Peri-implantitis is a microbially heterogeneous infection with predominantly gram-negative species, and is less complex than periodontitis.

Integrins are critical in thrombosis and hemostasis1. Antagonists of the platelet integrin αIIbβ3 are potent anti-thrombotic drugs, but also have the life-threatening adverse effect of bleeding2,3. It is thus desirable to develop new antagonists that do not cause bleeding. Integrins transmit signals bidirectionally4,5. Inside-out signaling activates integrins via a talin-dependent mechanism6,7. Integrin ligation mediates thrombus formation and outside-in signaling8,9, which requires Gα13 and greatly expands thrombi. Here we show that Gα13 and talin bind to mutually exclusive, but distinct sites within the integrin β3 cytoplasmic domain in opposing waves. The first talin binding wave mediates inside-out signaling and also “ligand-induced integrin activation”, but is not required for outside-in signaling. Integrin ligation induces transient talin dissociation and Gα13 binding to an ExE motif, which selectively mediates outside-in signaling and platelet spreading. The second talin binding wave is associated with clot retraction. An ExE motif-based inhibitor of Gα13-integrin interaction selectively abolishes outside-in signaling without affecting integrin ligation, and suppresses occlusive arterial thrombosis without affecting bleeding time. Thus, we have discovered a novel mechanism for the directional switch of integrin signaling and, based on this mechanism, we designed a potent new anti-thrombotic that does not cause bleeding.

The Akt family of serine/threonine kinases includes Akt1, Akt2, and Akt3 isoforms. Prior studies have reported that Akt1 and Akt2, but not Akt3, are expressed in platelets. Here, we show that Akt3 is expressed in substantial amounts in platelets. Akt3 ؊/؊ mouse platelets selectively exhibit impaired platelet aggregation and secretion in response to low concentrations of thrombin receptor agonists and thromboxane A 2 (TXA 2 ), but not collagen or VWF. In contrast, platelets from Akt1 ؊/؊ or Akt2 ؊/؊ mice are defective in platelet activation induced by thrombin, TXA 2 , and VWF, but only Akt1 ؊/؊ platelets show significant defects in response to collagen, indicating differences among Akt isoforms. Akt3 ؊/؊ platelets exhibit a significant reduction in thrombin-induced phosphorylation of glycogen synthase kinase 3␤ (GSK-3␤) at Ser9, which is known to inhibit GSK-3␤ function. Thus, Akt3 is important in inhibiting GSK-3␤. Accordingly, treatment of Akt3 ؊/؊ platelets with a GSK-3␤ inhibitor rescued the defect of Akt3 ؊/؊ platelets in thrombin-induced aggregation, suggesting that negatively regulating GSK-3␤ may be a mechanism by which Akt3 promotes platelet activation. Importantly, Akt3 ؊/؊ mice showed retardation in FeCl 3 -induced carotid artery thrombosis in vivo. IntroductionPlatelets are critical for hemostasis, but under pathologic conditions, are also important in thrombosis. 1 Platelet activation is initiated at sites of vascular injury on exposure to soluble agonists such as thrombin, ADP, and thromboxane A 2 , and adhesion to subendothelial matrix proteins, such as von Willebrand factor and collagen. 2 These adhesive proteins and agonists stimulate an intracellular signal transduction cascade leading to transformation of the major platelet adhesion receptor, integrin ␣ IIb ␤ 3 from its resting to active state (inside-out signaling), which allows the integrin to bind fibrinogen, and therefore mediate platelet aggregation. 3,4 Activated platelets secrete proaggregating factors and adhesive glycoproteins from granules, which further cause stabilization and amplification of aggregation, leading to thrombus formation. Ligand binding to the activated integrin ␣ IIb ␤ 3 also transmits "outside-in" signals, which are critically important in stable platelet adhesion, spreading, and clot retraction. 3,5,6 Elucidation of the signaling pathways regulating platelet activation is essential for the identification of novel anti-thrombotic targets for the prevention of thrombosis, a major cause of heart attack and stroke. It is established that phosphoinositide 3-kinases (PI3K) play important roles in platelet activation. [7][8][9][10][11][12] Akt (also known as Protein Kinase B or PKB), the most well known effector of PI3K, is activated downstream of PI3K during platelet activation. 9,13,14 Akt is a family of serine/threonine kinases with 3 isoforms: Akt1, Akt2, and Akt3 (for reviews see Manning et al 15 and Bhaskar et al 16 ). Akt isoforms are 80% homologous in their protein sequences. However, knockout mouse models of Akt...

This study examined the extent to which maternal attention-deficit/hyperactivity disorder (ADHD) symptoms predict improvement in child behavior following brief behavioral parent training. Change in parenting was examined as a potential mediator of the negative relationship between maternal ADHD symptoms and improvement in child behavior. Seventy mothers of 6–10 year old children with ADHD underwent a comprehensive assessment of adult ADHD prior to participating in an abbreviated parent training program. Before and after treatment, parenting was assessed via maternal reports and observations and child disruptive behavior was measured via maternal report. Controlling for pre-treatment levels, maternal ADHD symptomatology predicted post-treatment child disruptive behavior problems. The relation between maternal ADHD symptomatology and improvement in child behavior was mediated by change in observed maternal negative parenting. This study replicated findings linking maternal ADHD symptoms with attenuated child improvement following parent training, and is the first to demonstrate that negative parenting at least partially explains this relationship. Innovative approaches combining evidence-based treatment for adult ADHD with parent training may therefore be necessary for families in which both the mother and child have ADHD. Larger-scale studies using a full evidence-based parent training program are needed to replicate these findings.

Objective Approximately 15–20 percent of young children can be classified as having a behaviorally inhibited (BI) temperament. Stable BI predicts the development of later anxiety disorders (particularly social anxiety), but not all inhibited children develop anxiety. Parenting characterized by inappropriate warmth/sensitivity and/or intrusive control predicts the stability of BI and moderates risk for anxiety among high-BI children. For these reasons, we developed and examined the preliminary efficacy of the Turtle Program: a multi-modal early intervention for inhibited preschool-aged children. Method Forty inhibited children between the ages of 42–60 months and their parent(s) were randomized to either the Turtle Program (n = 18) or a waitlist control condition (WLC; n = 22). Participants randomized to the Turtle Program condition received 8 weeks of concurrent parent and child group treatment. Participants were assessed at baseline and post-treatment with multi-source assessments, including parent and teacher report measures of child anxiety, diagnostic interviews, and observations of parenting behavior. Results The Turtle Program resulted in significant beneficial effects relative to the WLC condition on maternal-reported anxiety symptoms of medium to large magnitude; large effects on parent-reported BI; medium to large effects on teacher-rated school anxiety symptoms; and medium effects on observed maternal positive affect/sensitivity. Conclusions This study provides encouraging preliminary support for the Turtle Program for young behaviorally inhibited children. Importantly, the effects of the Turtle Program generalized to the school setting. Future studies should examine whether this early intervention program improves long-term developmental outcomes for these at-risk children.

Objective More than 50% of mothers of children with attention-deficit/hyperactivity disorder (ADHD) have a lifetime history of major depressive disorder (MDD). Maternal depressive symptoms are associated with impaired parenting and predict adverse developmental and treatment outcomes for children with ADHD. For these reasons, we developed and examined the preliminary efficacy of an integrated treatment targeting parenting and depressive symptoms for mothers of children with ADHD. This integrated intervention incorporated elements of 2 evidence-based treatments: behavioral parent training (BPT) and cognitive behavioral depression treatment. Method Ninety-eight mothers with at least mild depressive symptoms were randomized to receive either standard BPT (n = 51) or the integrated parenting intervention for ADHD (IPI-A; n = 47). Participants were assessed at baseline, posttreatment, and 3- to 6-month follow-up on measures of (a) self-reported maternal depressive symptoms, (b) observed positive and negative parenting, and (c) observed and mother-reported child disruptive behavior and mother-reported child and family impairment. Results The IPI-A produced effects of small to moderate magnitude relative to BPT on maternal depressive symptoms, observed negative parenting, observed child deviance, and child impairment at posttreatment and on maternal depressive symptoms, child disruptive behavior, child impairment and family functioning at follow-up. Contrary to expectations, the BPT group demonstrated moderate to large effects relative to IPI-A on observed positive parenting at follow-up. Conclusions This treatment development study provides encouraging preliminary support for the integrated intervention targeting parenting and depressive symptoms in mothers of children with ADHD. Future studies should examine whether this integrated intervention improves long-term developmental outcomes for children with ADHD.

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