Perspectives from NHLBI Global Health Think Tank Meeting for Late Stage (T4) Translation Research.

Almost three-quarters (74%) of all the noncommunicable disease burden is found within low- and middle-income countries. In September 2014, the National Heart, Lung, and Blood Institute held a Global Health Think Tank meeting to obtain expert advice and recommendations for addressing compelling scientific questions for late stage (T4) research-research that studies implementation strategies for proven effective interventions-to inform and guide the National Heart, Lung, and Blood Institute's global health research and training efforts. Major themes emerged in two broad categories: 1) developing research capacity; and 2) efficiently defining compelling scientific questions within the local context. Compelling scientific questions included how to deliver inexpensive, scalable, and sustainable interventions using alternative health delivery models that leverage existing human capital, technologies and therapeutics, and entrepreneurial strategies. These broad themes provide perspectives that inform an overarching strategy needed to reduce the heart, lung, blood, and sleep disorders disease burden and global health disparities.

Reducing Health Inequities in the U.S.: Recommendations From the NHLBI's Health Inequities Think Tank Meeting.

The National, Heart, Lung, and Blood Institute convened a Think Tank meeting to obtain insight and recommendations regarding the objectives and design of the next generation of research aimed at reducing health inequities in the United States. The panel recommended several specific actions, including: 1) embrace broad and inclusive research themes; 2) develop research platforms that optimize the ability to conduct informative and innovative research, and promote systems science approaches; 3) develop networks of collaborators and stakeholders, and launch transformative studies that can serve as benchmarks; 4) optimize the use of new data sources, platforms, and natural experiments; and 5) develop unique transdisciplinary training programs to build research capacity. Confronting health inequities will require engaging multiple disciplines and sectors (including communities), using systems science, and intervening through combinations of individual, family, provider, health system, and community-targeted approaches. Details of the panel's remarks and recommendations are provided in this report.

Platelet adhesion to dengue-2 virus-infected endothelial cells.

Platelets in circulation normally do not adhere to resting endothelial cells. However, in response to vascular injury they adhere to stimulated endothelium and thereby play an essential role in hemostasis and thrombosis. Infection with dengue-2 virus can cause illness accompanied by thrombocytopenia and hemorrhage. Increased adherence of platelets to stimulated endothelial cells could contribute to the thrombocytopenia. In this study, adherence of radioisotopically labeled platelets to 1) unstimulated, 2) lipopolysaccharide (LPS)-stimulated, and 3) dengue-2 virus-infected human umbilical vein endothelial cells (HUVEC) was measured in an in vitro assay. Primary HUVEC were cultured in 96-well tissue culture plates in the presence or absence of LPS or dengue-2 virus. These cells were co-incubated with 3H-adenine-labeled fresh platelets for 30 min after which the cells were assayed for adherent platelets. Within 30 min there was maximum adherence of platelets to confluent LPS-stimulated HUVEC (36 +/- 4% over controls; P = 0.005). In comparison, there was a significant increase in adherence to dengue-2 infected HUVEC (78 +/- 7%; P < or = 0.001). Additionally, platelet adherence was visualized using fluorescent microscopy. Dengue-2 infection stimulated the HUVEC as monitored by expression of E-selectin. Platelets that adhered to dengue-2 or LPS-stimulated HUVEC were activated as visualized by dual fluorescent probes. These data demonstrate that human platelets adhere to dengue-2 virus-stimulated HUVEC and this interaction could contribute to the thrombocytopenia observed during infection.

Assessment of omega-fatty-acid-supplemented human platelets for potential improvement in long-term storage.

Uptake of omega (omega)-3 fatty acids can influence membrane stability and cell mobility. We investigated the effects of omega-3 and -6 fatty acids on the hemostatic efficacy of human platelets using an in vivo rabbit bleeding model. In vitro assays such as platelet aggregation, vWF bead-mediated ATP release and platelet adhesion to beads (measured by the residual platelet count [RPC] [free platelet count after reacting with the beads]/[baseline platelet count]x 100=%RPC; a high %RPC indicates reduced platelet function) were conducted on platelets treated with 1% fish oil (omega-3); 2% fish oil emulsion or 1% soy oil (omega-6). Oil treatment of platelets reduced the vWF bead-induced ATP release insignificantly. Addition of omega-3 agents reduced physical reactivity (%RPC) with the vWF beads by a factor of 1.2 (oil) and 1.9 (emulsion). The omega-6 oil enhanced reactivity by a factor of 1.7. After washing to remove excess reagent, platelet resuspension was most efficient with the omega-3 emulsion. Platelet function was higher with the omega-3-treated platelets (%RPC=52.3%, omega-3 oil; 63.3%, omega-3 emulsion vs. 85%, omega-6 oil; 82% untreated platelets). Ethyl-palmitate-treated thrombocytopenic rabbits were infused with human platelets. Survival times of the treated platelets, as monitored by flow cytometry (6.2-8.2 h) were comparable to untreated platelets (8.6 h). In the rabbit kidney injury model, blood loss after infusion of the treated platelets was similar to that of saline-infused rabbits (75.3+/-3.4 g). However, platelets washed prior to infusion reduced blood loss to a value comparable to that of fresh platelets (48.3+/-5 g). Furthermore, the presence of the infused platelets at the injury site was clearly visualized using FITC-tagged anti CD42a antibody. Thus, the omega-3-based agents protect the platelets from damage during the washing procedure as demonstrated in vitro by improved platelet resuspension, low %RPC, high stimulus-responsive ATP secretion and a reduction in blood loss in vivo.

Addition of a propyl gallate-based procoagulant to a fibrin bandage improves hemostatic performance in a swine arterial bleeding model.

Fibrin bandages manufactured by Nycomed Austria (TC-S) were modified by the addition of Hemostyptin (HS), a proprietary platelet-activating reagent containing propyl gallate. HS was added as an additional layer to TC-S fibrin bandages and the bandages were tested for hemostatic efficacy in a swine femoral artery bleeding model. Injuries were treated with a TC-S+HS bandage preparation using HS lyophilized onto a bandage surface that was then attached to the fibrin dressing. This preparation qualitatively and quantitatively exhibited more robust blood clotting at the surgical site than the control bandages. TC-S+HS bandages were more effective than control bandages with a difference in blood loss of 251.8+/-66.5 g for TC-S bandage alone, n=12 vs. 121+/-40.7 g, n=13 for the TC-S+HS bandage, P=0.05. Bleeding times were shortened for animals treated with the HS fortified bandages and residual platelets counts in these animals were higher.

Epsilon-amino caproic acid additive decreases fibrin bandage performance in a swine arterial bleeding model.

epsilon -Amino caproic acid (EACA), a lysine analog that inhibits the activity of plasmin, was added to Nycomed (TC-S) fibrin bandages, and the bandages were tested for hemostatic efficacy in a swine femoral artery bleeding model. The blood loss using the bandage with EACA (TC-S+EACA; 728.8+/-132 g, n=5) was much greater as compared to the TC-S bandage alone (TC-S; 237.8+/-47.9 g, n=6, p=.02). The time to "cessation of bleeding" (T(b)) was also increased for animals treated with TC-S+EACA bandages compared to the TC-S controls (33.6+/-10.8 min vs. 9.2+/-2.2 min; p=.05). Although plasma fibrinogen concentration decreased in animals treated with the TC-S+EACA bandage, activated partial thromboplastin times (aPTT) and thrombin times (TT) were decreased. Animals treated with the TC-S control bandage exhibited no changes in fibrinogen, aPTT or TT. Prothrombin times (PT) were unchanged in either group. In conclusion, addition of EACA to the Nycomed product decreased fibrin bandage efficiency.