Yasmin Friedmann

The CentriMag Centrifugal Blood Pump as a Benchmark for In Vitro Testing of Hemocompatibility in Implantable Ventricular Assist Devices

Implantable ventricular assist devices (VADs) have proven efficient in advanced heart failure patients as a bridge-to-transplant or destination therapy. However, VAD usage often leads to infection, bleeding, and thrombosis, side effects attributable to the damage to blood cells and plasma proteins. Measuring hemolysis alone does not provide sufficient information to understand total blood damage, and research exploring the impact of currently available pumps on a wider range of blood cell types and plasma proteins such as von Willebrand factor (vWF) is required to further our understanding of safer pump design. The extracorporeal CentriMag (Thoratec Corporation, Pleasanton, CA, USA) has a hemolysis profile within published standards of normalized index of hemolysis levels of less than 0.01 g/100 L at 100 mm Hg but the effect on leukocytes, vWF multimers, and platelets is unknown. Here, the CentriMag was tested using bovine blood (n = 15) under constant hemodynamic conditions in comparison with a static control for total blood cell counts, hemolysis, leukocyte death, vWF multimers, microparticles, platelet activation, and apoptosis. The CentriMag decreased the levels of healthy leukocytes (P < 0.006), induced leukocyte microparticles (P < 10−5), and the level of high molecular weight of vWF multimers was significantly reduced in the CentriMag (P < 10−5) all compared with the static treatment after 6 h in vitro testing. Despite the leukocyte damage, microparticle formation, and cleavage of vWF multimers, these results show that the CentriMag is a hemocompatible pump which could be used as a standard in blood damage assays to inform the design of new implantable blood pumps.

A Model of Void Formation

We introduce a simple model for the formation of voids. In this model the underdensity of galaxies in voids is the product of two factors. The first arises from a gravitational expansion of the negative density perturbation. The second is due to biasing: galaxies are less likely to form in an underdense region. One feature of the model is an upper cutoff in void sizes. We calculate the volume-filling factor of characteristic voids for different cold dark matter (CDM) models and find that our formation model points to ΛCDM models as preferred models of the power spectrum. A natural consequence of our model is that the underdensity of the dark matter inside voids is smaller than the galaxy underdensity.

The Effect of Shear Stress on the Size, Structure, and Function of Human von Willebrand Factor

Clinical outcomes from ventricular assist devices (VADs) have improved significantly during recent decades, but bleeding episodes remain a common complication of long-term VAD usage. Greater understanding of the effect of the shear stress in the VAD on platelet aggregation, which is influenced by the functional activity of high molecular weight (HMW) von Willebrand factor (vWF), could provide insight into these bleeding complications. However, because VAD shear rates are difficult to assess, there is a need for a model that enables controlled shear rates to first establish the relationship between shear rates and vWF damage. Secondly, if such a dependency exists, then it is relevant to establish a rapid and quantitative assay that can be used routinely for the safety assessment of new VADs in development. Therefore, the purpose of this study was to exert vWF to controlled levels of shear using a rheometer, and flow cytometry was used to investigate the shear-dependent effect on the functional activity of vWF. Human platelet-poor plasma (PPP) was subjected to different shear rate levels ranging from 0 to 8000/s for a period of 6 h using a rheometer. A simple and rapid flow cytometric assay was used to determine platelet aggregation in the presence of ristocetin cofactor as a readout for vWF activity. Platelet aggregates were visualized by confocal microscopy. Multimers of vWF were detected using gel electrophoresis and immunoblotting. The longer PPP was exposed to high shear, the greater the loss of HMW vWF multimers, and the lower the functional activity of vWF for platelet aggregation. Confocal microscopy revealed for the first time that platelet aggregates were smaller and more dispersed in postsheared PPP compared with nonsheared PPP. The loss of HMW vWF in postsheared PPP was demonstrated by immunoblotting. Smaller vWF platelet aggregates formed in response to shear stress might be a cause of bleeding in patients implanted with VADs. The methodological approaches used herein could be useful in the design of safer VADs and other blood handling devices. In particular, we have demonstrated a correlation between the loss of HMW vWF, analyzed by immunoblotting, with platelet aggregation, assessed by flow cytometry. This suggests that flow cytometry could replace conventional immunoblotting as a simple and rapid routine test for HMW vWF loss during in vitro testing of devices.

Fluctuation Analysis of the Far-Infrared Background - Information from the Confusion

We investigate the extent to which one can use a P(D) analysis to extract number counts of unclustered sources from maps of the far-infrared background. Such maps currently available, and those expected to emerge in the near future, are dominated by confusion noise resulting from poor resolution. We simulate background maps with an underlying two-slope model for N(S), and we find that, in an experiment of FIRBACK type, we can extract the high-flux slope with an error of a few per cent, whereas other parameters are not so well constrained. However, we find that, in a SIRTF-type experiment, all parameters of this N(S) model can be extracted with errors of only few per cent.

Recombination-Mediated Host Adaptation by Avian Staphylococcus aureus

Staphylococcus aureus are globally disseminated among farmed chickens causing skeletal muscle infections, dermatitis, and septicaemia. The emergence of poultry-associated lineages has involved zoonotic transmission from humans to chickens but questions remain about the specific adaptations that promote proliferation of chicken pathogens. We characterized genetic variation in a population of genome-sequenced S. aureus isolates of poultry and human origin. Genealogical analysis identified a dominant poultry-associated sequence cluster within the CC5 clonal complex. Poultry and human CC5 isolates were significantly distinct from each other and more recombination events were detected in the poultry isolates. We identified 44 recombination events in 33 genes along the branch extending to the poultry-specific CC5 cluster, and 47 genes were found more often in CC5 poultry isolates compared with those from humans. Many of these gene sequences were common in chicken isolates from other clonal complexes suggesting horizontal gene transfer among poultry associated lineages. Consistent with functional predictions for putative poultry-associated genes, poultry isolates showed enhanced growth at 42 °C and greater erythrocyte lysis on chicken blood agar in comparison with human isolates. By combining phenotype information with evolutionary analyses of staphylococcal genomes, we provide evidence of adaptation, following a human-to-poultry host transition. This has important implications for the emergence and dissemination of new pathogenic clones associated with modern agriculture.

Quantification methods for human and large animal leukocytes using DNA dyes by flow cytometry.

Ovine and bovine blood is used heavily within the development of blood‐handling medical devices, such as heart pumps (left ventricular assist devices, LVADs), for which blood cell damage needs to be monitored during in vitro testing. Hematology analyzers provide cell counts but no information about cell viability. The anthraquinone DNA dyes CyTRAK Orange™ and DRAQ7™ have practical and spectral properties rendering them suitable for multicolor assays. Compared to other DNA dyes such as Vybrant Dyecycle, CyTRAK Orange enables a faster staining protocol and does not require incubation at +37°C. Compared to traditional viability dyes such as propidium iodide and 7AAD, DRAQ7s unique spectral profile of excitation in both blue and red lasers and far‐red emission enables identification of dual positive dead cell events and frees up detectors for use with other reagents. CyTRAK Orange and DRAQ7 could be used in combination with absolute counting bead standards to provide cell counts and viability but the combination of these dyes has previously only been used for microscopy on rodent cells. The purpose of this study was to evaluate the use of these dyes in combination in large animal blood samples for flow cytometry. A viability and cell counting protocol for bovine, ovine, and human leukocytes using CyTRAK Orange and DRAQ7 was prepared. Four different counting bead standards were evaluated using the Navios and FACSAria cytometers and compared to counts obtained from hematology analyzers. CyTRAK Orange successfully detected CD45+ leukocytes in all species. The DRAQ7 single‐stained dead cell counts correlated well with the CyTRAK Orange/DRAQ7 double‐stained dead cell counts in human and bovine blood, but not in ovine blood, which could be related to the blood source. In conclusion, for human and bovine blood, this method works well for viability counts using different flow cytometers and bead standards.

Intra-Pelvic Pressure Changes after Pelvic Fracture: A Cadaveric Study Quantifying the Effect of a Pelvic Binder and Limb Bandaging Over a Bolster

INTRODUCTION Unstable pelvic fractures can be life-threatening due to catastrophic haemorrhage. Non-invasive methods of reducing and stabilising these injuries include pelvic binder application and also lower limb bandaging over a knee-flexion bolster. Both of these methods help close the pelvic ring and should tamponade bleeding. This study aimed to quantify the intra-pelvic pressure changes that occurred with 3 different manoeuvres: lower limb bandaging over a bolster; a Trauma Pelvic Orthotic Device (T-POD) pelvic binder, and a combination of both. METHODS Following a pilot study with 2 soft embalmed cadavers, a formal study with 6 unembalmed cadavers was performed. For each specimen an unstable pelvic injury was created (OA/OTA 61-C1) by dividing the pelvic ring anteriorly and posteriorly. A 3-4cm manometric water-filled balloon was placed in the retropubic space and connected to a 50ml syringe and water manometer via a 3-way tap. A baseline pressure of 8cmH2O (equating to the average central venous pressure) was used for each cadaver. Steady intra-pelvic pressures (more reliably reflecting the pressures achieved following an intervention) were used in the subsequent statistical analysis, using R statistical language and Rstudio. Paired t-test or Wilcoxons rank sum test were used (depending on the normality of the dataset) to determine the impact of each intervention on the intra-pelvic pressure. RESULTS The mean steady intra-pelvic pressures were significantly greater than the baseline pressure for each intervention. The binder and limb bandaging over a bolster alone increased the mean steady pelvic pressures significantly to 24 (SE=5) (p<0.036) and 15.5 (SE=2) (p<0.02)cmH2O respectively. Combining these interventions further increased the mean steady pressure to 31 (SE=7)cmH2O. However, this was not significantly greater than pressures for each of the individual interventions. DISCUSSION Both lower limb bandaging over a bolster and pelvic binder application significantly increased intra-pelvic pressure above the baseline pressure. This was further increased through combining these interventions, which could be useful clinically to augment haemorrhage control in these fractures. CONCLUSION Lower-limb bandaging over a bolster, and pelvic binder application, both significantly increased intra-pelvic pressures, and were greatest in combination. These findings support the use of these techniques to facilitate non-surgical haemorrhage control.

Evaluation of Four Veterinary Hematology Analyzers for Bovine and Ovine Blood Counts for In Vitro Testing of Medical Devices

Small affordable automated hematology analyzers that produce rapid and accurate complete blood cell counts are a valuable tool to researchers developing blood-handling medical devices, such as ventricular assist devices, for in vitro safety assessments. In such studies, it is common to use the blood of large animals such as cattle and sheep. However, the commercially available instruments have not been evaluated for their ability to measure the blood counts of these animals. In this study, we compare, for the first time, four veterinary analyzers for blood counts on bovine and ovine blood samples. We look at ease of use, repeatability and agreement with a view to inform researchers of the benefits of these instruments in routine measurement of ovine and bovine bloods during in vitro testing. Complete blood cell counts and a three-part differential (granulocytes, monocytes, and lymphocytes) were measured by each of the instruments, and the results compared to those obtained from two additional analyzers used in a reference laboratory. Repeatability and agreement were evaluated using the Bland-Altman method; bias and 95% limits of agreement between the instruments, and between the instruments and two reference instruments, were used to evaluate instrument performance. In summary, there are advantages and disadvantages with all instruments. Of the four instruments tested, the repeatability and agreement was fairly similar for all instruments except one instrument which cannot be recommended for bovine or ovine blood counts.

Impact of hand washing on bacteraemia rates

Stone and colleagues’ results are consistent with mathematical models of hospital transmission of meticillin resistant Staphylococcus aureus (MRSA).1 This can be shown using Cooper and colleagues’ model, whereby transmission within hospital is described by an effective contact rate parameter between susceptible and infectious people. Improved hygiene conditions reduce the probability that contact between people results …