Effects of pathogen reduction technology and storage duration on the ability of cryoprecipitate to rescue induced coagulopathies in vitro

BACKGROUND: Fibrinogen concentrates and cryoprecipitate are currently used for fibrinogen supplementation in bleeding patients with dysfibrinogenemia. Both products provide an abundant source of fibrinogen but take greater than 10 min to prepare for administration. Fibrinogen concentrates lack coagulation factors (i.e., factor VIII [FVIII], factor XIII [FXIII], von Willebrand factor [VWF]) important for robust hemostatic function. Cryoprecipitate products contain these factors but have short shelf lives (\u3c6 h). Pathogen reduction (PR) of cryoprecipitate would provide a shelf-stable immediately available adjunct containing factors important for rescuing hemostatic dysfunction. STUDY DESIGN AND METHODS: Hemostatic adjunct study products were psoralen-treated PR-cryoprecipitated fibrinogen complex (PR-Cryo FC), cryoprecipitate (Cryo), and fibrinogen concentrates (FibCon). PR-Cryo FC and Cryo were stored for 10 days at 20-24°C. Adjuncts were added to coagulopathies (dilutional, 3:7 whole blood [WB]:normal saline; or lytic, WB + 75 ng/ml tissue plasminogen activator), and hemostatic function was assessed by rotational thromboelastometry and thrombin generation. RESULTS: PR of cryoprecipitate did not reduce levels of FVIII, FXIII, or VWF. PR-Cryo FC rescued dilutional coagulopathy similarly to Cryo, while generating significantly more thrombin than FibCon, which also rescued dilutional coagulopathy. Storage out to 10 days at 20-24°C did not diminish the hemostatic function of PR-Cryo FC. DISCUSSION: PR-Cryo FC provides similar and/or improved hemostatic rescue compared to FibCon in dilutional coagulopathies, and this rescue ability is stable over 10 days of storage. In hemorrhaging patients, where every minute delay is associated with a 5% increase in mortality, the immediate availability of PR-Cryo FC has the potential to improve outcomes

Cast Glance Near Infrared Imaging Observations of the Space Shuttle During Hypersonic Re-Entry

High resolution calibrated infrared imagery of the Space Shuttle was obtained during hypervelocity atmospheric entries of the STS-119, STS-125 and STS128 missions and has provided information on the distribution of surface temperature and the state of the airflow over the windward surface of the Orbiter during descent. This data collect was initiated by NASA s Hypersonic Thermodynamic Infrared Measurements (HYTHIRM) team and incorporated the use of air- and land-based optical assets to image the Shuttle during atmospheric re-entry. The HYTHIRM objective is to develop and implement a set of mission planning tools designed to establish confidence in the ability of an existing optical asset to reliably acquire, track and return global quantitative surface temperatures of the Shuttle during entry. On Space Shuttle Discovery s STS-119 mission, NASA flew a specially modified thermal protection system tile and instrumentation package to monitor heating effects from boundary layer transition during re-entry. On STS-119, the windward airflow on the port wing was deliberately disrupted by a four-inch wide and quarter-inch tall protuberance built into the modified tile. In coordination with this flight experiment, a US Navy NP-3D Orion aircraft was flown 28 nautical miles below Discovery and remotely monitored surface temperature of the Orbiter at Mach 8.4 using a long-range infrared optical package referred to as Cast Glance. Approximately two months later, the same Navy Cast Glance aircraft successfully monitored the surface temperatures of the Orbiter Atlantis traveling at approximately Mach 14.3 during its return from the successful Hubble repair mission. In contrast to Discovery, Atlantis was not part of the Boundary Layer Transition (BLT) flight experiment, thus the vehicle was not configured with a protuberance on the port wing. In September 2009, Cast Glance was again successful in capturing infrared imagery and monitoring the surface temperatures on Discovery s next flight, STS-128. Again, NASA flew a specially modified thermal protection system tile and instrumentation package to monitor heating effects from boundary layer transition during re-entry. During this mission, Cast Glance was able to image laminar and turbulent flow phenomenology optimizing data collection for Mach 14.7. The purpose of this paper is to describe key elements associated with STS-119/125/128 mission planning and execution from the perspective of the Cast Glance flight crew that obtained the imagery. The paper will emphasize a human element of experience, expertise and adaptability seamlessly coupled with Cast Glance system and sensor technology required to manually collect the required imagery. Specific topics will include a near infrared (NIR) camera upgrade that was implemented just prior to the missions, how pre-flight radiance modeling was utilized to optimize the IR sensor configuration, communications, the development of aircraft test support positions based upon Shuttle trajectory information, support to contingencies such as Shuttle one orbit wave-offs/west coast diversions and then the Cast Glance perspective during an actual Shuttle imaging mission

The Current Testing Protocols for Biomechanical Evaluation of Lumbar Spinal Implants in Laboratory Setting: A Review of the Literature

In vitro biomechanical investigations have become a routinely employed technique to explore new lumbar instrumentation. One of the most important advantages of such investigations is the low risk present when compared to clinical trials. However, the best use of any experimental data can be made when standard testing protocols are adopted by investigators, thus allowing comparisons among studies. Experimental variables, such as the length of the specimen, operative level, type of loading (e.g., dynamic versus quasistatic), magnitude, and rate of load applied, are among the most common variables controlled during spinal biomechanical testing. Although important efforts have been made to standardize these protocols, high variability can be found in the current literature. The aim of this investigation was to conduct a systematic review of the literature to identify the current trends in the protocols reported for the evaluation of new lumbar spinal implants under laboratory setting

Dose-dependent von Willebrand Factor inhibition by aptamer BB-031 correlates with thrombolysis in a microfluidic model of arterial occlusion

Von Willebrand Factor (VWF) plays a critical role in thrombus formation, stabilization, and propagation. Previous studies have demonstrated that targeted inhibition of VWF induces thrombolysis when administered in vivo in animal models of ischemic stroke. The study objective was to quantify dose-dependent inhibition of VWF-platelet function and its relationship with thrombolysis using BB-031, an aptamer that binds VWF and inhibits its function. VWF:Ac, VWF:RCo, T-TAS, and ristocetin-induced impedance aggregometry were used to assess BB-031-mediated inhibition of VWF. Reductions in original thrombus surface area and new deposition during administration of treatment were measured in a microfluidic model of arterial thrombolysis. Rotational thromboelastometry was used to assess changes in hemostasis. BB-031 induced maximal inhibition at the highest dose (3384 nM) in VWF:Ac, and demonstrated dose-dependent responses in all other assays. BB-031, but not vehicle, induced recanalization in the microfluidic model. Maximal lytic efficacy in the microfluidic model was seen at 1692 nM and not 3384 nM BB-031 when assessed by surface area. Minor changes in ROTEM parameters were seen at 3384 nM BB-031. Targeted VWF inhibition by BB-031 results in clinically measurable impairment of VWF function, and specifically VWF-GPIb function as measured by VWF:Ac. BB-031 also induced thrombolysis as measured in a microfluidic model of occlusion and reperfusion. Moderate correlation between inhibition and lysis was observed. Additional studies are required to further examine off-target effects of BB-031 at high doses, however, these are expected to be above the range of clinical targeted dosing

SIRT1-dependent myoprotective effects of resveratrol on muscle injury induced by compression

© 2015 Sin, Yung, Yip, Chan, Wong, Tam and Siu. Our current understanding on the molecular mechanisms by which sustained compression induces skeletal muscle injury is very limited. This study aimed to test the hypothesis that activation of SIRT1 by the natural antioxidant resveratrol could deactivate apoptotic and catabolic signaling in skeletal muscle exposed to moderate compression. Two cycles of 6-h constant pressure at 100 mmHg was applied to the tibialis region of right, but not left hindlimbs of Sprague Dawley rats pre-treated with DMSO (vehicle control) or resveratrol with/without sirtinol. Skeletal muscle tissues lying underneath and spatially corresponding to the compressed sites were collected for analyses. Resveratrol prevented the compression-induced manifestations of pathohistological damages including elevations of the number of interstitial nuclei and area of interstitial space and ameliorated oxidative damages measured as 4-hydroxy-2-nonenal (4HNE) and nitrotyrosine in skeletal muscle. In parallel, resveratrol augmented the expression level and activity of SIRT1 and phosphorylation levels of Foxo3a and Akt while suppressed the increases in protein abundances of p53, Bax, MAFbx, and ubiquitin, enzymatic activities of caspase 3 and 20S proteasome, and apoptotic DNA fragmentation in the compressed muscle. These favorable myoprotective effects of resveratrol were diminished upon pharmacological blockade of SIRT1 by using sirtinol. These novel data support the hypothesis that the anti-apoptotic and anti-catabolic effects of resveratrol on compression injury in skeletal muscle required the action of SIRT1.Link_to_subscribed_fulltex

Multifactorial resistance to adriamycin: relationship of DNA repair, glutathione transferase activity, drug efflux, and p-glycoprotein in cloned cell lines of adriamycin-sensitive and -resistant P388 Leukemia

Cloned lines of Adriamycin (ADR)-sensitive and -resistant P388 leukemia have been established, including P388/ADR/3 and P388/ADR/7 that are 5- and 10-fold more resistant than the cloned sensitive cell line P388/4 (Cancer Res., 46: 2978, 1986). A time course of ADR-induced DNA double-strand breaks revealed that in sensitive P388/4 cells, evidence of DNA repair was noted 4 h after removal of drug, whereas in resistant clone 3 and 7 cells repair was observed 1 h after drug removal. The earlier onset of DNA repair was statistically significant (p = 0.0154 for clone 3 cells, and p = 0.0009 for clone 7 cells). By contrast, once the repair process was initiated, the rate of repair was similar for all three cell lines. The level of glutathione transferase activity was determined in whole cell extracts. Enzyme activity (mean ± SE) in sensitive cells was 9.49 ± 1.00 nmol/min/mg protein, that in resistant clone 3 cells was 13.36 ± 1.03 nmol/min/mg, and that in clone 7 cells was 13.96 ± 1.44 nmol/min/mg; the 1.44-fold increase in enzyme activity in resistant cells was statistically significant (p = 0.01). Further evidence of induction of glutathione transferase was provided by Northern blot analysis using a 32P-labeled cDNA for an anionic glutathione transferase, which demonstrated approximately a twofold increase in mRNA in resistant clone 7 cells. Western blot analysis with a polyvalent antibody against anionic glutathione transferase also revealed a proportionate increase in gene product in resistant cells. Dose-survival studies showed that ADR-resistant cells were cross-resistant to actinomycin D, daunorubicin, mitoxantrone, colchicine, and etoposide, but not to the alkylating agent melphalan; this finding provided evidence that these cells are multidrug resistant. Using a cDNA probe for P-glycoprotein, a phenotypic marker for multidrug resistance, Northern blot analysis showed an increase in the steady state level of mRNA of approximately twofold in resistant clone 3 and 7 cells. Southern analysis with the same cDNA probe showed no evidence of gene amplification or rearrangement. Western blot analysis with monoclonal C219 antibody demonstrated a distinct increase in P-glycoprotein in resistant cells. Efflux of Adriamycin as measured by the efflux rate constant was identical in all three cell lines. Furthermore, the metabolic inhibitors azide and dinitrophenol did not augment drug uptake in either sensitive or resistant cells. These findings suggest that despite the increase in P-glycoprotein, an active extrusion pump was not operational in these cells. This and previous studies provide unequivocal evidence that resistance to Adriamycin is multifactorial. Decreased drug uptake, decreased formation of DNA single- and double-strand breaks, increased glutathione transferase activity, earlier onset of DNA repair, as well as elevated P-glycoprotein are all characteristic of multifactorial drug resistance

Populations of latent Mycobacterium tuberculosis lack a cell wall: Isolation, visualization, and whole-genome characterization

AbstractObjective/BackgroundMycobacterium tuberculosis (MTB) causes active tuberculosis (TB) in only a small percentage of infected people. In most cases, the infection is clinically latent, where bacilli can persist in human hosts for years without causing disease. Surprisingly, the biology of such persister cells is largely unknown. This study describes the isolation, identification, and whole-genome sequencing (WGS) of latent TB bacilli after 782days (26months) of latency (the ability of MTB bacilli to lie persistent).MethodsThe in vitro double-stress model of latency (oxygen and nutrition) was designed for MTB culture. After 26months of latency, MTB cells that persisted were isolated and investigated under light and atomic force microscopy. Spoligotyping and WGS were performed to verify the identity of the strain.ResultsWe established a culture medium in which MTB bacilli arrest their growth, reduce their size (0.3–0.1μm), lose their acid fastness (85–90%) and change their shape. Spoligopatterns of latent cells were identical to original H37Rv, with differences observed at spacers two and 14. WGS revealed only a few genetic changes relative to the already published H37Rv reference genome. Among these was a large 2064-bp insertion (RvD6), which was originally detected in both H37Ra and CDC1551, but not H37Rv.ConclusionHere, we show cell-wall free cells of MTB bacilli in their latent state, and the biological adaptation of these cells was more phenotypic in nature than genomic. These cell-wall free cells represent a good model for understanding the nature of TB latency

Practice Patterns and Preferences Among Hematopoietic Cell Transplantation Clinicians

Hematopoietic cell transplantation can cure many high-risk diseases but is associated with complexity, cost, and risk. Several areas in transplantation practice were identified in the 2014 Blood and Marrow Transplant Clinical Trials Network State of the Science Symposium (BMT CTN SOSS) as high priorities for further study. We developed a survey for hematopoietic cell transplantation clinicians to identify current practices in BMT CTN SOSS priority areas and to understand, more generally, the variation in approach to transplantation and estimation of transplantation benefit in current medical practice. Of 1439 transplantation clinicians surveyed, 305 responded (20% response rate). Clinicians were well represented by age, experience, geography, and size of practice. We found that several techniques identified in the BMT CTN SOSS, such as maintenance therapy for acute myeloid leukemia or myelodysplastic syndromes after allogeneic transplantation, were already being utilized in practice on and off study, with higher rates of use in higher-volume centers. There was significant variation among clinicians in use of transplantation technologies and approaches to common transplantation scenarios. Appraisals of risks and benefits of transplantation appeared to converge upon similar estimates despite the presentation of different hypothetical scenarios. These results suggest overall equipoise in several BMT CTN SOSS high-priority areas and support the need for better data to inform clinical practice