Child Maltreatment and Neural Networks Underlying Emotion Regulation

This is a project funded by NIMH (R01-MH103291) that seeks to identify mechanisms linking child maltreatment to child and adolescent psychopathology, with a particular focus on the role of emotion regulatio

Roles and interactions among protease-activated receptors and P2ry12 in hemostasis and thrombosis

Toward understanding their redundancies and interactions in hemostasis and thrombosis, we examined the roles of thrombin receptors (protease-activated receptors, PARs) and the ADP receptor P2RY12 (purinergic receptor P2Y G protein-coupled 12) in human and mouse platelets ex vivo and in mouse models. Par3−/− and Par4+/− mouse platelets showed partially decreased responses to thrombin, resembling those in PAR1 antagonist-treated human platelets. P2ry12+/− mouse platelets showed partially decreased responses to ADP, resembling those in clopidogrel-treated human platelets. Par3−/− mice showed nearly complete protection against carotid artery thrombosis caused by low FeCl3 injury. Par4+/− and P2ry12+/− mice showed partial protection. Increasing FeCl3 injury abolished such protection; combining partial attenuation of thrombin and ADP signaling, as in Par3−/−:P2ry12+/− mice, restored it. Par4−/− mice, which lack platelet thrombin responses, showed still better protection. Our data suggest that (i) the level of thrombin driving platelet activation and carotid thrombosis was low at low levels of arterial injury and increased along with the contribution of thrombin-independent pathways of platelet activation with increasing levels of injury; (ii) although P2ry12 acts downstream of PARs to amplify platelet responses to thrombin ex vivo, P2ry12 functioned in thrombin/PAR-independent pathways in our in vivo models; and (iii) P2ry12 signaling was more important than PAR signaling in hemostasis models; the converse was noted for arterial thrombosis models. These results make predictions being tested by ongoing human trials and suggest hypotheses for new antithrombotic strategies

Selectin-Like Kinetics and Biomechanics Promote Rapid Platelet Adhesion in Flow: The GPIbα-vWF Tether Bond

AbstractThe ability of platelets to tether to and translocate on injured vascular endothelium relies on the interaction between the platelet glycoprotein receptor Ib α (GPIbα) and the A1 domain of von Willebrand factor (vWF-A1). To date, limited information exists on the kinetics that govern platelet interactions with vWF in hemodynamic flow. We now report that the GPIbα-vWF-A1 tether bond displays similar kinetic attributes as the selectins including: 1) the requirement for a critical level of hydrodynamic flow to initiate adhesion, 2) short-lived tethering events at sites of vascular injury in vivo, and 3) a fast intrinsic dissociation rate constant, koff0 (3.45±0.37s−1). Values for koff, as determined by pause time analysis of transient capture/release events, were also found to vary exponentially (4.2±0.8s−1 to 7.3±0.4s−1) as a function of the force applied to the bond (from 36 to 217 pN). The biological importance of rapid bond dissociation in platelet adhesion is demonstrated by kinetic characterization of the A1 domain mutation, I546V that is associated with type 2B von Willebrand disease (vWD), a bleeding disorder that is due to the spontaneous binding of plasma vWF to circulating platelets. This mutation resulted in a loss of the shear threshold phenomenon, a approximately sixfold reduction in koff, but no significant alteration in the ability of the tether bond to resist shear-induced forces. Thus, flow dependent adhesion and rapid and force-dependent kinetic properties are the predominant features of the GPIbα–vWF-A1 tether bond that in part may explain the preferential binding of platelets to vWF at sites of vascular injury, the lack of spontaneous platelet aggregation in circulating blood, and a mechanism to limit thrombus formation

Performance of a Point of Care Test for Detecting IgM and IgG Antibodies Against SARS-CoV-2 and Seroprevalence in Blood Donors and Health Care Workers in Panama

Novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the etiologic agent of the ongoing coronavirus disease 2019 (COVID-19) pandemic, which has reached 28 million cases worldwide in 1 year. The serological detection of antibodies against the virus will play a pivotal role in complementing molecular tests to improve diagnostic accuracy, contact tracing, vaccine efficacy testing, and seroprevalence surveillance. Here, we aimed first to evaluate a lateral flow assay's ability to identify specific IgM and IgG antibodies against SARS-CoV-2 and second, to report the seroprevalence estimates of these antibodies among health care workers and healthy volunteer blood donors in Panama. We recruited study participants between April 30th and July 7th, 2020. For the test validation and performance evaluation, we analyzed serum samples from participants with clinical symptoms and confirmed positive RT-PCR for SARS-CoV-2, and a set of pre-pandemic serum samples. We used two by two table analysis to determine the test positive and negative percentage agreement as well as the Kappa agreement value with a 95% confidence interval. Then, we used the lateral flow assay to determine seroprevalence among serum samples from COVID-19 patients, potentially exposed health care workers, and healthy volunteer donors. Our results show this assay reached a positive percent agreement of 97.2% (95% CI 84.2–100.0%) for detecting both IgM and IgG. The assay showed a Kappa of 0.898 (95%CI 0.811–0.985) and 0.918 (95% CI 0.839–0.997) for IgM and IgG, respectively. The evaluation of serum samples from hospitalized COVID-19 patients indicates a correlation between test sensitivity and the number of days since symptom onset; the highest positive percent agreement [87% (95% CI 67.0–96.3%)] was observed at ≥15 days post-symptom onset (PSO). We found an overall antibody seroprevalence of 11.6% (95% CI 8.5–15.8%) among both health care workers and healthy blood donors. Our findings suggest this lateral flow assay could contribute significantly to implementing seroprevalence testing in locations with active community transmission of SARS-CoV-2.</jats:p

Genetic elimination of prothrombin in adult mice is not compatible with survival and results in spontaneous hemorrhagic events in both heart and brain

Mice carrying a conditional prothrombin knockout allele (fIIlox) were established to develop an experimental setting for exploring the importance of thrombin in the maintenance of vascular integrity, the inflammatory response, and disease processes in adult animals. In the absence of Cre-mediated recombination, homozygous fIIlox/lox mice or compound heterozygous mice carrying one fIIlox allele and one constitutive-null allele were viable. Young adults exhibited neither spontaneous bleeding events nor diminished reproductive success. However, the induction of Cre recombinase in fIIlox mice using the poly I:C-inducible Mx1-Cre system resulted in the rapid and near-complete recombination of the fIIlox allele within the liver, the loss of circulating prothrombin, and profound derangements in coagulation function. Consistent with the notion that thrombin regulates coagulation and inflammatory pathways, an additional early consequence of reducing prothrombin was impaired antimicrobial function in mice challenged with Staphylococcus aureus peritonitis. However, life expectancy in unchallenged adults genetically depleted of prothrombin was very short (∼5-7 days). The loss of viability was associated with the development of severe hemorrhagic events within multiple tissues, particularly in the heart and brain. Unlike the constitutive loss of either clotting or platelet function alone, the conditional loss of prothrombin is uniformly not compatible with maintenance of hemostasis or long-term survival