Shear rate gradients promote a bi-phasic thrombus formation on weak adhesive proteins, such as fibrinogen in a VWF-dependent manner.

Blood flow profoundly varies throughout the vascular tree due to its pulsatile nature and to the complex vessel geometry. While thrombus formation has been extensively studied in vitro under steady flow, and in vivo under normal blood flow conditions, the impact of complex hemodynamics such as flow acceleration found in stenosed arteries has gained increased appreciation. We investigated the effect of flow acceleration, characterized by shear rate gradients, on the function of platelets adhering to fibrinogen, a plasma protein which plays a key role in hemostais and thrombosis. While we confirmed that under steady flow, fibrinogen only supports single platelet adhesion, we observed that under shear rate gradients, this surface becomes highly thrombogenic, supporting efficient platelet aggregation leading to occlusive thrombus formation. This shear rate gradient-driven thrombosis is biphasic with an initial step of slow platelet recruitment supported by direct plasma VWF adsorption to immobilized fibrinogen and followed by a second phase of explosive thrombosis initiated by VWF fiber formation on platelet monolayers. In vivo experiments confirmed that shear rate gradients accelerate thrombosis in a VWF-dependent manner. Together, this study characterizes a process of plasma VWF-dependent accelerated thrombosis on immobilized fibrinogen in the presence of shear rate gradients.

Adipokines expression profile in liver, adipose tissue and muscle during chicken embryo development.

In broiler chickens, the intense genetic selection for rapid growth has resulted in an increase in growth rate and fat deposition. Adipose tissue is now recognized as an important endocrine organ that secretes a variety of factors including adipokines. However, the expression pattern of these adipokines is unclear in chicken embryo development. In the present study, we determined the expression profile of three novel adipokines, NAMPT, RARRES2 and ADIPOQ, and their cognate receptors in metabolic tissues (liver, muscles and adipose tissue) of chicken embryo/chicks from 15 days of incubation (E15) to hatching (D0). From E15 to hatching, embryos gradually gained weight and started to develop subcutaneous adipose tissue at E15. We conducted western blot and RT-qPCR tests and found that ADIPOQ expression increased over time and was positively correlated with adipose tissue weight. In addition, NAMPT expression increased only in muscles. By using a new homemade chicken RARRES2 specific antibody we showed that RARRES2 protein levels increased specifically at hatching in adipose tissue, liver and pectoralis major and this was associated with an increase in the weight of embryo. Taken together, these results support a potential involvement of adipokines in metabolic regulation during chicken embryo development.

Anatomic correlates of deep brain stimulation electrode impedance.

BACKGROUND: The location of the optimal target for deep brain stimulation (DBS) of the subthalamic nucleus (STN) remains controversial. Electrode impedance affects tissue activation by DBS and has been found to vary by contact number, but no studies have examined association between impedance and anatomic location. OBJECTIVES: To evaluate the relationship between electrode impedance and anatomic contact location, and to assess the clinical significance of impedance. METHODS: We gathered retrospective impedance data from 101 electrodes in 73 patients with Parkinson's disease. We determined contact location using microelectrode recording (MER) and high-field 7T MRI, and assessed the relationship between impedance and contact location. RESULTS: For contact location as assessed via MER, impedance was significantly higher for contacts in STN, at baseline (111 Ω vs STN border, p=0.03; 169 Ω vs white matter, p<0.001) and over time (90 Ω vs STN border, p<0.001; 54 Ω vs white matter, p<0.001). Over time, impedance was lowest in contacts situated at STN border (p=0.03). Impedance did not vary by contact location as assessed via imaging. Location determination was 75% consistent between MER and imaging. Impedance was inversely related to absolute symptom reduction during stimulation (-2.5 motor portion of the Unified Parkinson's Disease Rating Scale (mUPDRS) points per 1000 Ω, p=0.01). CONCLUSIONS: In the vicinity of DBS electrodes chronically implanted in STN, impedance is lower at the rostral STN border and in white matter, than in STN. This finding suggests that current reaches white matter fibres more readily than neuronal cell bodies in STN, which may help explain anatomic variation in stimulation efficacy.

Dehydration of blood platelets by zeodration: in vitro characterization and hemostatic properties in vivo.

BACKGROUND: Platelets (PLTs) are currently stored at room temperature (RT) for 5 to 7 days. So far, there exists no validated method for the preparation and long-term storage of dehydrated PLTs suitable for transfusion after rehydration. In this study, a desiccation process, zeodration, was applied to PLTs. STUDY DESIGN AND METHODS: A complete procedure of dehydration at RT by zeodration was employed. Zeodrated human and mouse PLTs were characterized in vitro. Zeodrated mouse PLTs were transfused into clopidogrel-treated mice to evaluate their hemostatic properties. RESULTS: The optimal conditions for dehydration of PLTs at RT in a laboratory scale zeodrator were defined as 145 mbar and 20.2 ± 1.5 °C. The recovery rate was 85 ± 2% and the dryness of zeodrated PLTs (Z_PLTs) indicated that they were sufficiently stable for long-term storage. Rehydrated Z_PLTs were round, were not aggregated, and expressed the glycoproteins required for PLT function. Z_PLTs agglutinated in the presence of von Willebrand factor (VWF) and aggregated in response to thrombin or collagen independently of an active metabolism. In a flow system, Z_PLTs could adhere to VWF and form aggregates on a collagen matrix. Thrombin was generated at the surface of Z_PLTs as efficiently as on fresh PLTs. In clopidogrel-treated mice, which exhibited a severely prolonged bleeding time, continuous perfusion of Z_PLTs restored normal hemostasis. CONCLUSION: Zeodration represents a new strategy to prepare PLTs with partly preserved aggregative properties after storage and rehydration. Z_PLTs have potential hemostatic properties provided it is possible to improve their transfusion efficacy.

Integrin α6ß1 is the main receptor for vascular laminins and plays a role in platelet adhesion, activation, and arterial thrombosis.

BACKGROUND: Laminins are major components of basement membranes, well located to interact with platelets upon vascular injury. Laminin-111 (α1ß1γ1) is known to support platelet adhesion but is absent from most blood vessels, which contain isoforms with the α2, α4, or α5 chain. Whether vascular laminins support platelet adhesion and activation and the significance of these interactions in hemostasis and thrombosis remain unknown. METHODS AND RESULTS: Using an in vitro flow assay, we show that laminin-411 (α4ß1γ1), laminin-511 (α5ß1γ1), and laminin-521 (α5ß2γ1), but not laminin-211 (α2ß1γ1), allow efficient platelet adhesion and activation across a wide range of arterial wall shear rates. Adhesion was critically dependent on integrin α6ß1 and the glycoprotein Ib-IX complex, which binds to plasmatic von Willebrand factor adsorbed on laminins. Glycoprotein VI did not participate in the adhesive process but mediated platelet activation induced by α5-containing laminins. To address the significance of platelet/laminin interactions in vivo, we developed a platelet-specific knockout of integrin α6. Platelets from these mice failed to adhere to laminin-411, laminin-511, and laminin-521 but responded normally to a series of agonists. α6ß1-Deficient mice presented a marked decrease in arterial thrombosis in 3 models of injury of the carotid, aorta, and mesenteric arterioles. The tail bleeding time and blood loss remained unaltered, indicating normal hemostasis. CONCLUSIONS: This study reveals an unsuspected important contribution of laminins to thrombus formation in vivo and suggests that targeting their main receptor, integrin α6ß1, could represent an alternative antithrombotic strategy with a potentially low bleeding risk.

Targeting platelet GPIbß reduces platelet adhesion, GPIb signaling and thrombin generation and prevents arterial thrombosis.

OBJECTIVE: The glycoprotein (GP) Ib-V-IX complex regulates the adhesion, activation, and procoagulant activity of platelets. We previously reported that RAM.1, a rat monoclonal antibody directed against the extracellular domain of mouse GPIbß, diminished adhesion of platelets and chinese hamster ovary cells transfected with the human GPIb-IX complex to von Willebrand factor under flow conditions. Here, we further evaluated the functional importance of GPIbß by studying the impact of RAM.1 on GPIb-mediated platelet responses and in vitro and in vivo thrombus formation. APPROACH AND RESULTS: We show that RAM.1 dramatically reduced GPIb-mediated filopodia extension of chinese hamster ovary GPIb-IX cells after adhesion to von Willebrand factor. RAM.1 also reduced filopodia extension and GPIb-mediated Ca(2+) signaling after adhesion of mouse platelets to von Willebrand factor. RAM.1 inhibited thrombin generation in platelet-rich plasma without impairing phosphatidylserine exposure. In addition, RAM.1 reduced thrombus formation after perfusion of mouse whole blood over collagen in a shear-dependent manner. This effect was confirmed in vivo, because injection of F(ab)'2 fragments of RAM.1 diminished thrombus formation induced by laser beam injury of mesenteric arterioles and forceps injury of the abdominal aorta. In contrast, RAM.1 F(ab)'2 did not prolong the tail-bleeding time or increase the volume of blood lost. CONCLUSIONS: These findings are the first evidence that targeting a subunit other than GPIbα can lead to an antithrombotic effect via the GPIb-V-IX complex. This could represent an alternative way to reduce thrombus formation with a minor impact on hemostasis.

Post-Implementation Outcomes of a Remote Patient Monitoring Program After Emergency Department Discharge.

To better communicate and improve post-visit outcomes, a remote patient monitoring (RPM) program was implemented for patients discharged from emergency departments (ED) across 10 hospitals. The solution was offered to patients at the time of ED discharge and staffed by a group of care coordinators to respond to questions/urgent needs. Of 107,477 consecutive patients offered RPM, 28,425 patients (26.4%) engaged with the program. Activated patients with RPM were less likely to return to the ED within 90 days of their index visit [19.8% compared to 23.6%, p<.001]. While activation rates were modest, we observed fewer return visits to the ED in patients using RPM, with a 16.2% lower hazard of returning in the next year. Future research is needed to understand methods to improve RPM activation, any causal effects of RPM activation on return ED visits, and external validation of these findings.

Preserved functional and biochemical characteristics of platelet components prepared with amotosalen and ultraviolet A for pathogen inactivation.

BACKGROUND: Platelet concentrate (PC) functionality decreases during storage. This is referred to as the storage lesion. Pathogen inactivation may accelerate or induce lesions, potentially accounting for reduced viability. Our aim was to characterize functional and biochemical properties of platelets (PLTs) from photochemically treated buffy-coat PCs (PCT-PCs) compared to those from conventional PCs. STUDY DESIGN AND METHODS: Four PCT-PCs and four conventional PCs were stored for 6.5 days and PLT function and proteomic profiles were examined at various time points during storage. To evaluate their intrinsic properties, samples of stored PLTs were taken, washed, and suspended in Tyrode's buffer before testing. RESULTS: PLT counts and morphology were conserved although a slight increase in the PLT volume was observed after PCT. Glycoprotein (GP) IIbIIIa, IaIIa, and VI expression remained stable while GPIbα declined similarly in both types of PCs. A steep decrease (50%) in GPV occurred on Day 1.5 in PCT-PCs and Day 2.5 in control PCs. For both PCT- and control PCs, P-selectin expression and activated GPIIbIIIa remained low during storage. PCT- and control PCs were fully responsive to aggregation agonists up to Day 4.5 and exhibited similar perfusion functionality. Mitochondrial membrane potential and annexin A5 binding of PCT-PCs and control PCs were comparable. Two-dimensional differential in-gel electrophoresis and mass spectrometry profiles for 1882 protein spots revealed only three proteins selectively changed in PCT-PCs compared to control-PCs. CONCLUSION: Washed treated and untreated PCs have similar functional, morphologic, and proteomic characteristics provided that PLTs are suspended in an appropriate medium during testing.

Alcohol dependence is related to overall internalizing psychopathology load rather than to particular internalizing disorders: evidence from a national sample.

BACKGROUND: Alcohol dependence is more prevalent among those with any one of several anxiety or depressive ("internalizing") disorders than among those in the general population. However, because internalizing disorders are highly intercorrelated, it is ambiguous whether alcohol dependence is related to internalizing psychopathology components that are: (i) unique to a particular internalizing disorder ("specific"); versus (ii) shared across a number of internalizing disorders ("general"). To clarify this ambiguity, we employed structural equation and logistic models to decompose the specific versus general components of internalizing psychopathology and then related these components separately to alcohol dependence. METHODS: The data were based on face-to-face interviews of U.S. community residents collected in the 2001 to 2002 National Epidemiologic Survey on Alcohol and Related Conditions (NESARC; N = 43,093). RESULTS: Both analytic approaches demonstrated that increases in the general internalizing psychopathology load are accompanied by increases in the prevalence of alcohol dependence. Once the general internalizing psychopathology load is accounted for, knowing whether a particular internalizing disorder is present or absent provides little additional information regarding the prevalence of alcohol dependence. CONCLUSIONS: The components of internalizing psychopathology that are associated with alcohol dependence are shared and cumulative among common anxiety and depressive disorders. These findings have the potential to influence clinical and scientific conceptualizations of the association between alcohol dependence and internalizing psychopathology.

Meta-analysis of supplemental treatment for depressive and anxiety disorders in patients being treated for alcohol dependence.

Approximately half of those receiving treatment for an alcohol use disorder (AUD) also suffer with an anxiety or depressive (internalizing) disorder. Because all internalizing disorders mark a poor alcohol treatment outcome, it seems reasonable to supplement AUD treatment with a psychiatric intervention when these disorders co-occur with AUD. However, this conclusion may be faulty given that the various possible interrelationships between AUD and internalizing disorders do not uniformly imply a high therapeutic yield from this approach. Unfortunately, the studies conducted to date have been too few and too small to resolve this important clinical issue with confidence. Therefore, we used a meta-analytic method to synthesize the effects from published randomized controlled trials examining the impact of supplementing AUD treatment with a psychiatric treatment for co-occurring internalizing disorder (N = 15). We found a pooled effect size (d) of .32 for internalizing outcomes and .22 for a composite of alcohol outcomes; however, the alcohol outcomes effect sizes were greater than this for some specific outcome domains. Subgroups that differed in terms of internalizing outcomes included treatment type (medication vs. cognitive behavioral therapy) and treatment focus (anxiety vs. depression). There was also a trend for the studies with better internalizing disorder outcomes to have better alcohol outcomes. These results indicate that clinical outcomes (both psychiatric and alcohol-related) could be somewhat improved by supplementing AUD treatment with psychiatric treatment for co-occurring internalizing disorder.

Urge to gamble in a simulated gambling environment.

Cue reactivity, while increasingly recognized as a central feature of drug and alcohol addiction, is not well studied in gambling. We evaluated the urge to gamble in a simulated casino environment among frequent gamblers who alternated between cycles in which they observed others playing ten hands of Blackjack (first, third and fifth cycle) and cycles in which they played ten hands of Blackjack themselves (second and fourth cycle). The played cycles served as a manipulation for the observed cycles in terms of "priming" (having previously gambled in the environment vs. not) and "anticipation" (expecting more opportunities to gamble in the environment vs. not) and, thus, allowed these conditions: observed cycle 1 = anticipation (+) and prime (-); observed cycle 2 = anticipation (+) and prime (+); and observed cycle 3 = anticipation (-) and prime (+). Subjects' urge to gamble was greater in the gambling environment than in a neutral setting and both positive anticipation and positive priming increased cue reactivity within the gambling environment. The frequency of gambling outside of the study did not affect cue reactivity. However, a preference for Blackjack (vs. other types of gambling) and observing winning (vs. losing) hands were both associated with stronger cue reactivity in the study. These findings contribute to our understanding of pathological gambling.

Fibrillar cellular fibronectin supports efficient platelet aggregation and procoagulant activity.

The ability of cellular fibronectin, found in the vessel wall in a fibrillar conformation, to regulate platelet functions and trigger thrombus formation remains largely unknown. In this study, we evaluated how parietal cellular fibronectin can modulate platelet responses under flow conditions. A fibrillar network was formed by mechanically stretching immobilised dimeric cellular fibronectin. Perfusion of anticoagulated whole blood over this surface resulted in efficient platelet adhesion and thrombus growth. The initial steps of platelet adhesion and activation, as evidenced by filopodia extension and an increase in intracellular calcium levels (419 ± 29 nmol/l), were dependent on integrins α5ß1 and αIIbß3. Subsequent thrombus growth was mediated by these integrins together with the GPIb-V-IX complex, GPVI and Toll-like receptor 4. The involvement of Toll-like receptor 4 could be conveyed via its binding to the EDA region of cellular fibronectin. Upon thrombus formation, the platelets became procoagulant and generated fibrin as revealed by video-microscopy. This work provides evidence that fibrillar cellular fibronectin is a strong thrombogenic surface which supports efficient platelet adhesion, activation, aggregation and procoagulant activity through the interplay of a series of receptors including integrins α5ß1 and αIIbß3, the GPIb-V-IX complex, GPVI and Toll-like receptor 4.

Hybrid cognitive behavioral therapy versus relaxation training for co-occurring anxiety and alcohol disorder: a randomized clinical trial.

OBJECTIVE: Treatment for alcohol use disorder (AUD) is far less effective for those with a co-occurring anxiety disorder. Surprisingly, adding an independent anxiety treatment to AUD treatment does not substantially improve the poor alcohol outcomes of these patients. This may reflect the lack of attention from independent treatments to the dynamic interaction of anxiety symptoms with alcohol use and drinking motivation. On the basis of this view, we assembled a cognitive behavioral therapy (CBT) program designed to both reduce anxiety symptoms and weaken the links between the experience of anxiety and the motivation to drink. METHOD: 344 patients undergoing residential AUD treatment with current social phobia, generalized anxiety disorder, or panic disorder were randomly assigned to receive either the CBT or an active comparison treatment, progressive muscle relaxation training (PMRT). Assessments took place immediately following treatment and 4 months later (n = 247). RESULTS: As predicted, the CBT group demonstrated significantly better alcohol outcomes 4 months following treatment than did the PMRT group. Although both groups experienced a substantial degree of anxiety reduction following treatment, there were no significant group differences immediately after treatment and only a slight advantage for the CBT group 4 months after treatment. CONCLUSIONS: These findings suggest that specific interventions aimed at weakening the association between the experience of anxiety and drinking motivation play an important role in improving the alcohol outcomes of these difficult-to-treat patients beyond that of anxiety reduction alone.

Greater elevation in risk for nicotine dependence per pack of cigarettes smoked among those with an anxiety disorder.

OBJECTIVE: Recent work shows that the time from the initial use of nicotine, cannabis, and alcohol to the onset of dependence on these substances is shorter ("telescoped") in anxiety-disordered individuals. Previously, we hypothesized that telescoping may result from a shared neurobiology underlying both anxiety disorders and dependence. This hypothesis implies that telescoping occurs because individuals with an anxiety disorder transition to dependence with less overall drug exposure ("dependence susceptibility"). To investigate this further, we examined an estimate of the amount smoked (rather than the time transpired) from smoking initiation milestones to the onset of nicotine dependence in those with and without an anxiety disorder. METHOD: We used the subset of respondents in the National Epidemiologic Survey on Alcohol and Related Conditions (NESARC) Wave 1 who reported having smoked at least 100 cigarettes (N = 18,013). All data were based on face-to-face interviews. RESULTS: Individuals with any anxiety disorder transitioned to nicotine dependence after smoking fewer total cigarettes than did individuals with no anxiety disorder. Furthermore, those with more than one anxiety disorder transitioned to nicotine dependence after smoking fewer cigarettes than did those with one anxiety disorder only. Several potentially confounding covariates were controlled for in these analyses. CONCLUSIONS: Dependence susceptibility is a novel concept with the potential to inform theoretical accounts of and prevention strategies for substance dependence among those with an anxiety disorder. In addition to nicotine, our theory and past data suggest that dependence susceptibility for other addictive substances (e.g., alcohol) also would be found among those with an anxiety disorder.

Vulnerability to the rapid ("telescoped") development of alcohol dependence in individuals with anxiety disorder.

OBJECTIVE: The frequent co-occurrence of alcohol dependence and anxiety disorder is a long-standing clinical conundrum. An underdeveloped perspective on this issue concerns the impact of a co-occurring anxiety disorder on the sequence and developmental course of alcohol-related milestones. Extrapolating from the body of basic science indicating overlap in the neurobiological processes associated with both anxiety disorder and alcohol dependence-particularly those involving the hypothalamic-pituitary-adrenal axis and elements of the amygdala- we hypothesized that anxiety-disordered individuals are vulnerable to the rapid development of alcohol dependence. Specifically, we predicted that the time from pre-dependence alcohol milestones (e.g., age at which regular drinking began) to post-dependence alcohol milestones would be briefer ("telescoped") among those with an anxiety disorder. METHOD: Seventy-eight individuals with alcohol dependence who had recently begun a chemical dependency treatment program underwent a diagnostic interview to determine the presence of current anxiety disorders and to establish the age at which several alcohol use and dependence milestones were first achieved. RESULTS: We found that, compared with others in the sample, anxiety-disordered individuals transitioned significantly more quickly from the time they first began drinking regularly and first began getting drunk regularly to the onset of alcohol dependence, as well as from most pre-dependence alcohol milestones to the point at which their alcohol dependence was most severe. CONCLUSIONS: Individuals with anxiety disorders transition from regular drinking to alcohol dependence more rapidly than do individuals without anxiety disorders. These findings contribute to an improved understanding of the etiology of comorbidity and suggest novel directions for future research.

The prevalence and clinical implications of self-medication among individuals with anxiety disorders.

Alcohol dependence (AD) is more likely to occur among individuals with rather than without an anxiety disorder. Self-medication theory (SMT) holds that drinking behavior is negatively reinforced when alcohol temporarily reduces anxiety and that the resulting escalation of drinking increases the risk for AD. We set out to empirically scrutinize SMT using the National Epidemiologic Survey on Alcohol and Related Conditions (NESARC) dataset. We found that only a minority (about 20%) of anxiety disordered individuals endorsed drinking to control anxiety symptoms. This minority drank more alcohol, had a higher cross-sectional rate of AD, and was at higher risk for developing new AD over four years compared to anxiety disordered non-self-medicators and individuals with no anxiety disorder. Consistent with SMT, increased prospective risk for AD among self-medicators is partially mediated by an increased level of alcohol use. Understanding the processes that promote and inhibit self-medication should be a priority for anxiety disorder researchers.

Cognitive-behavioral therapy for panic disorder in patients being treated for alcohol dependence: Moderating effects of alcohol outcome expectancies.

Anxiety disorders commonly co-occur with alcohol use disorders and reliably mark a poor response to substance abuse treatment. However, treating a co-occurring anxiety disorder does not reliably improve substance abuse treatment outcomes. Failure to account for individual differences in the functional dynamic between anxiety symptoms and drinking behavior might impede the progress and clarity of this research program. For example, while both theory and research point to the moderating role of tension-reduction alcohol outcome expectancies (TR-AOEs) in the association between anxiety symptoms and alcohol use, relevant treatment studies have not typically modeled TR-AOE effects. We examined the impact of a hybrid cognitive-behavioral therapy (H-CBT) treatment for panic disorder (independent variable) on response to a community-based alcohol dependence treatment program (dependent variable) in patients with higher vs. lower TR-AOEs (moderator). The H-CBT treatment was generally effective in relieving participants' panic symptoms relative to controls. However, TR-AOEs interacted with study cohort (H-CBT vs. control) in predicting response to substance abuse treatment. As expected, the H-CBT was most effective in improving alcohol use outcomes among those with the highest TR-AOEs. The study's primary methodological limitations are related to the quasi-experimental design employed.