Does Manual Abdominal Pressure During Colonoscopy Put Endoscopy Staff and Patients at Risk? Experiences of Endoscopy Nurses and Technicians.

Endoscopy staff suffer work-related musculoskeletal disorders at a rate greater than or comparable to nurses and technicians in other subspecialities, which may be attributable to the widespread use of manual pressure and repositioning during colonoscopy. In addition to negatively impacting staff health and job performance, colonoscopy-related musculoskeletal disorder injuries may also signal potential risks to patient safety. To assess the prevalence of staff injury and perceived patient harm relating to the use of manual pressure and repositioning techniques during colonoscopy, 185 attendees of a recent national meeting of the Society of Gastroenterology Nurses and Associates were asked to recall experiencing injuries to themselves or observing injuries to other staff or patients during colonoscopy. A majority of respondents (84.9%, n = 157) reported either experiencing or observing staff injury, whereas 25.9% ( n = 48) reported observing patient complications. Among respondents who perform manual repositioning and apply manual pressure during colonoscopy (57.3%, n = 106), 85.8% ( n = 91) reported experiencing musculoskeletal disorders from performing these tasks; 81.1% ( n = 150) reported no awareness of colonoscopy-specific ergonomics policies at their facility. Results highlight the relationship between the physical job requirements of endoscopy nurses and technicians, staff musculoskeletal disorders, and patient complications, and suggest that implementation of staff safety protocols may benefit patients as well as endoscopy staff.

Clinical translation of the assets of biomedical engineering - a retrospective analysis with looks to the future.

Introduction: Biomedical-engineering (BME) plays a major role in modern medicine. Many BME-based assets have been brought to clinical translation in the twentieth century, but translation currently stagnates. Here, we compare the impact of past and present scientific, economic and societal climates on the translation of BME-based assets, in order to provide the BME-community with incentives to address current stagnation.Areas covered: In the twentieth century, W.J. Kolff brought kidney dialysis, the total artificial heart, artificial vision and limbs to clinical application. This success raises the question whether Kolff and other past giants of clinical translation had special mind-sets, or whether their problem selection, their training, or governmental and regulatory control played roles. Retrospective analysis divides the impact of BME-based assets to clinical application into three periods: 1900-1970: rapid translation from bench-to-bedside, 1970-1990: new diseases and increased governmental control, and the current translational crisis from 1990 onward.Expert opinion: Academic and societal changes can be discerned that are concurrent with BME's translational success: mono-disciplinary versus multi-disciplinary training, academic reward systems based on individual achievements versus team achievements with strong leadership, increased governmental and regulatory control, and industrial involvement. From this, recommendations can be derived for accelerating clinical translation of BME-assets.

Detachment and successive re-attachment of multiple, reversibly-binding tethers result in irreversible bacterial adhesion to surfaces.

Bacterial adhesion to surfaces occurs ubiquitously and is initially reversible, though becoming more irreversible within minutes after first contact with a surface. We here demonstrate for eight bacterial strains comprising four species, that bacteria adhere irreversibly to surfaces through multiple, reversibly-binding tethers that detach and successively re-attach, but not collectively detach to cause detachment of an entire bacterium. Arguments build on combining analyses of confined Brownian-motion of bacteria adhering to glass and their AFM force-distance curves and include the following observations: (1) force-distance curves showed detachment events indicative of multiple binding tethers, (2) vibration amplitudes of adhering bacteria parallel to a surface decreased with increasing adhesion-forces acting perpendicular to the surface, (3) nanoscopic displacements of bacteria with relatively long autocorrelation times up to several seconds, in absence of microscopic displacement, (4) increases in Mean-Squared-Displacement over prolonged time periods according to tα with 0 < α ≪ 1, indicative of confined displacement. Analysis of simulated position-maps of adhering particles using a new, in silico model confirmed that adhesion to surfaces is irreversible through detachment and successive re-attachment of reversibly-binding tethers. This makes bacterial adhesion mechanistically comparable with the irreversible adsorption of high-molecular-weight proteins to surfaces, mediated by multiple, reversibly-binding molecular segments.

Computational modeling of thrombotic microparticle deposition in nonparallel flow regimes.

Thrombotic microparticles (MPs) released from cells and platelets in response to various stimuli are present in elevated numbers in various disease states that increase the risk for thrombotic events. In order to understand how particles of this size may localize in nonparallel flow regimes and increase thrombotic risk, a computational analysis of flow and MP deposition was performed for 3 deg of stenosis at moderate Reynolds number (20

A computational analysis of an in vitro vessel wall injury model.

Implantation of vascular grafts or stents causes significant injury to the vessel wall. Activated coagulation factors, such as FXa are generated at the injury site. The size of the injury and the flow conditions influence the transport of these activated factors. A simulation model has been developed to evaluate surface bound coagulation inhibitors on medical devices. Tissue factor pathway inhibitor (TFPI) isa potent inhibitor of FXa in vivo. This physiologically relevant in vitro model studies the mechanism by which immobilized rTFPI effectively inhibits TF initiated thrombosis.Computational fluid dynamics was used to develop the model and validated by experiments performed in a parallel plate flow chamber. The lower plate was divided into two zones. The first zone represents fibroblasts that catalyze FXto FXa by TF-FVIIa. The second represents passively absorbed surface bound rTFPI. The efficacy of rTFPI in inhibiting FXa from the injury site was studied under both venous and moderate arterial shear rates. This model extends a previous model to include a more physiologic model of vessel wall injury in which FXa generation by TF:VIIa occurs at the wall upstream of the TFPI coated surface. The previous model estimated a uniform inlet FXa concentration of 20 nM (20% conversion of the approximate physiologic concentration of 100 nM) to the parallel plate chamber.

Evaluating surface bound rTFPI through an in vitro model of vessel wall injury.

INTRODUCTION: Injury to the surrounding vessel wall is one of the major reasons for failure of implantable medical devices. The surgical procedure itself or the altered flow conditions after implantation can cause damage to the vessel wall. This damage exposes tissue factor (TF), the initiator of the extrinsic pathway of coagulation. One approach to combat thrombosis is to use an anticoagulant on the surface of the device. The primary aim of this study is to develop a simplified physiologically relevant in vitro model of vessel wall injury to study the mechanisms by which immobilized recombinant tissue factor pathway inhibitor (rTFPI) effectively inhibits TF initiated thrombosis. MATERIALS AND METHODS: A two well chamber slide was used for the study. Fibroblasts were cultured on the upstream portion of the slide. Fibroblast cells stimulated with TNF-α acted as a source of surface TF. The downstream portion of the slide was coated with rTFPI. A mixture of FX, FVIIa and calcium was perfused over the slides to generate FXa. Effluent collected at the outlet was used to analyze the inhibition of this surface generated FXa by the rTFPI present downstream. RESULTS AND CONCLUSIONS: Different shear rates and rTFPI densities were used to study this effect. In most cases rTFPI inhibited FXa generated upstream as a function of the wall shear rate and rTFPI dosage (surface density). This study shows the effectiveness of the surface bound inhibitor when FXa is generated from an upstream injury site and the bulk of FXa is near the wall.

Cryptococcus neoformans causes lipid peroxidation; therefore it is a potential inducer of atherogenesis.

Certain viral and bacterial species play a role in the development of atherosclerosis. Our hypothesis was that yeasts, such as Cryptococcus neoformans, also may be a cause of lipid peroxidation (LPO), which can lead to atherosclerosis. Rabbits were inoculated with heat-killed C neoformans several times during the first 6 wk in 10 wk observation. These tests were done at regular intervals: (i) blood cell count, (ii) the nitroblue tetrazolium (NBT) test on isolated neutrophils to determine their super oxide anion production and (iii) LPO of plasma. The histopathology of the lungs also was evaluated. In the inoculated rabbits we found an increase in the number of neutrophils in blood with an elevated NBT reduction, an increase in lipid peroxidation of plasma and bronchopneumonia with various types of inflammatory cells. Our findings suggest that the ability of C. neoformans to induce LPO in human asymptomatic carriers should be studied. Moreover the cryptococcal rabbit model we use rapidly induces LPO and may be of value in the assessment of therapy for atherosclerosis.

Monocyte-derived macrophage microparticles impart tissue factor activity to biomaterial surfaces.

The initiation of coagulation on biomaterials is attributed to the contact pathway of coagulation. However, recent discoveries of blood-borne tissue factor (TF) activity suggest that the TF pathway of coagulation may contribute to thrombosis on biomaterials. To evaluate the role of TF bearing microparticles to biomaterial thrombogenicity, the adhesion of monocyte-derived macrophage microparticles (MMPs) to bare, bovine serum albumin (BSA) blocked, and plasma-coated materials was examined. MMP suspensions consisted of 20-37% TF positive particles that exhibited TF activity. Data from static experiments with polyethylene (PE), polydimethylsiloxane (PDMS), polystyrene (PS), and glass knitted and woven Dacron(R) grafts showed that MMPs adhered to uncoated, and plasma coated surfaces supported TF activity, whereas surfaces blocked with BSA supported less activity. Flow studies were performed on plasma-coated glass and tissue culture-treated polystyrene (TCPS) as a model system to demonstrate deposition and firm adhesion of microparticles under physiologic flow conditions. MMPs deposited and imparted TF activity to plasma-coated glass at wall shear rates of 100, 400, and 1200 sec(-1). Deposition on TCPS was comparable to glass at 100 sec(-1), but virtually nonexistent at the two higher shear rates after a 1 h perfusion, implying material and shear dependent adhesion. The localization of procoagulant MMPs to biomaterial surfaces could lead to an increased risk of thrombosis on cardiovascular implants beyond that anticipated by the contact pathway alone.

Construction and characterization of a thrombin-resistant designer FGF-based collagen binding domain angiogen.

Humans demonstrate limited spontaneous endothelialization of prosthetic bypass grafts. However the local application of growth factors to prosthetic grafts or to injured blood vessels can provide an immediate effect on endothelialization. Novel chimeric proteins combining potent angiogens with extracellular matrix binding domains may localize to exposed matrices and provide sustained activity to promote endothelial regeneration after vascular interventions. We have ligated a thrombin-resistant mutant of fibroblast growth factor (FGF)-1 (R136K) with a collagen binding domain (CBD) in order to direct this growth factor to sites of exposed vascular collagen or selected bioengineered scaffolds. While FGF-1 and R136K are readily attracted to a variety of matrix proteins, R136K-CBD demonstrated selective and avid binding to collagen approximately 4x that of FGF-1 or R136K alone (P<0.05). The molecular stability of R136K-CBD was superior to FGF-1 and R136K. Its chemotactic activity was superior to R136K and FGF-1 (11+/-1% vs. 6+/-2% and 4+/-1%; P<0.01). Its angiogenic activity was similar to R136K and significantly greater than control by day 2 (P<0.01). After day 3, FGF-1-treated endothelial cell's (EC) sprouts had regressed back to levels insignificant compared to the control group (P=0.17), while both R136K and R136K-CBD continued to demonstrate greater sprout lengthening as compared to control (P<0.0002). The mitogenic activity of all growth factors was greater than control groups (20% PBS); in all comparisons (P<0.0001). This dual functioning angiogen provides proof of concept for the application of designer angiogens to matrix binding proteins to intelligently promote endothelial regeneration of selected matrices.

Computational modeling of factor Xa inhibition by immobilized tissue factor pathway inhibitor.

Coating surfaces of implanted devices with anticoagulants can reduce thrombosis and studies using a recombinant form of endogenous tissue factor pathway inhibitor (rTFPI) are promising. The anticoagulant function of immobilized rTFPI is thought to occur primarily by its inhibition of plasma clotting factor Xa (FXa); however the kinetics of this reaction at a surface are as yet unknown. To better understand the surface inhibition reaction under flow conditions, a theoretical model was developed delineating the roles of mass transport and reaction kinetics for an in vitro parallel plate device used in prior experimental studies [Hall et al., J. Biomech. Eng. 120:484-490, 1998]. As a first approximation, the kinetics of inhibition of FXa by rTFPI reported for static, homogeneous systems was considered. The unsteady convection-diffusion equation was solved for different wall-shear rates and inlet concentrations of FXa using the computational fluid dynamics software CFD-ACE (ESI Software Group). The results show that the heterogeneous inhibition reaction is diffusion controlled prior to saturation of the rTFPI. The experimental results compare favorably with the model at the lower shear rates (100-400 s(-1)). At higher shear rates (>400 s(-1)) the theoretical results follow the same trend as the experimental results but show a greater inhibition of FXa, implying an effect of flow or shear on the inhibition reaction.

Factor Xa inhibition by immobilized recombinant tissue factor pathway inhibitor.

The recombinant form of the endogenous anticoagulant, tissue factor pathway inhibitor (rTFPI), is a potent inhibitor of Factor Xa (FXa) and the tissue factor-factor VIIa (TF:VIIa) complex. Surface-immobilized rTFPI reduces the thrombogenicity and intimal hyperplasia associated with synthetic vascular grafts in animal models and specifically reduces fibrin deposition on collagen-impregnated Dacron grafts from native blood in an in vitro flow model. The FXa inhibitory capacity of rTFPI in the bulk phase has been demonstrated in static systems and immobilized rTFPI reduces fibrin deposition in whole blood in vitro and animal studies; however, the specific mode of this anticoagulation has not been studied. Therefore, a comparison was made between the FXa binding capacity of two forms of immobilized rTFPI, i.e., passively adsorbed and covalently bound. The rTFPI-coated surfaces were evaluated using a parallel-plate flow reactor and comparing the amount of FXa exiting the flow chamber after exposure to an rTFPI-coated versus an uncoated plate. The results demonstrate that adsorbed rTFPI exhibits increased binding capacity (1.5-3.6 times) the expected stoichiometry via interactions with the C-terminus, whereas covalently-bound rTFPI interacts with FXa in a 1:1 stoichiometry. Thus, the results imply that specific FXa inhibition is a key component of the anticoagulant effect of rTFPI-coated surfaces and that passive adsorption of rTFPI to glass surfaces produces a more effective coating than covalent binding of rTFPI.

Trypanosoma lewisi-induced immunosuppression: the effects on alveolar macrophage activities against Cryptococcus neoformans.

The immunosuppressive effect of Trypanosoma lewisi infection on alveolar macrophage (AM) activities against Cryptococcus neoformans was studied in an animal model. Two groups of rats were treated with T. lewisi and killed after 4 (4d-rats) and 7 days (7d-rats), respectively. A third group not given T. lewisi, served as control. AM were challenged in vitro with C. neoformans. Phagocytosis was assessed with a fluorescence method. Superoxide anion production was evaluated with the nitroblue tetrazolium (NBT) test. The survival of cryptococci was estimated by counting colony-forming units. The numbers of detached AM from culture plates were determined using a Bürker chamber. The NBT response, adhesion to plate surface and killing activity, but not the phagocytosis of AM from 4d-rats were significantly impaired compared to control or 7d-rats. Thus, T. lewisi causes transitory immunosuppressive effects on AM activities. This rapid T. lewisi immunosuppression model may be useful to study new approaches to anticryptococcal therapy.

Polymer cushions functionalized with lipid molecules.

This Letter reports a novel approach to the fabrication of a biomimicking surface by modification of an end-functionalizable smooth polymer cushion constructed via chemoselective ligation with a phospholipid-like molecule containing oxyamine groups. The mobility of a phospholipid bilayer formed by vesicle fusion on the phospholipid-like molecule terminated polymer film was characterized by fluorescence recovery after bleaching. Platelet adhesion, as one measure of biocompatibility of the film was also studied and compared to other surfaces such as polyethylene or poly(dimethylsiloxane). The results show that the end-functionalized smooth polymer cushion has potential as a biocompatible platform to reconstitute membrane proteins.

In vitro flow evaluation of recombinant tissue factor pathway inhibitor immobilized on collagen impregnated Dacron.

Recombinant tissue factor pathway inhibitor (rTFPI) was immobilized on collagen impregnated (CI) knitted Dacron surfaces and its resistance to fibrin deposition evaluated following exposure to nonanticoagulated whole blood. Recombinant TFPI readily adsorbed to the CI Dacron surface and maintained its inhibitory activity. Under static conditions, rTFPI treated CI Dacron showed little fibrin deposition when compared with untreated surfaces. Treated samples exposed to flowing native blood at wall shear rates of 100 or 200 sec(-1) also demonstrated reduced fibrin deposition (up to 56%) compared with untreated samples. To assess the relative roles of the contact and tissue factor pathways in fibrin formation on artificial grafts, flow studies were performed with whole blood containing corn trypsin inhibitor, a potent inhibitor of FXIIa and contact activation. Corn trypsin inhibitor reduced fibrin deposition on untreated CI Dacron by 40%. Immobilized rTFPI alone, or corn trypsin inhibitor in combination with immobilized rTFPI, reduced fibrin deposition by 58% and 61%, respectively. These data suggest that immobilized rTFPI slows fibrin deposition on the vascular graft material by inhibiting both the contact pathway and blood borne tissue factor procoagulant activity arising from either the alternatively spliced form of tissue factor or from tissue factor containing microparticles.

Localization of tissue factor pathway inhibitor to lipid rafts is not required for inhibition of factor VIIa/tissue factor activity.

Tissue factor pathway inhibitor (TFPI) abrogates coagulation initiated by the factor VIIa/tissue factor catalytic complex. While the gene structure of TFPI suggests that it is a secreted protein, a large pool of TFPI is associated with the vascular endothelium through its affinity for a glycosylphosphati-dylinositol (GPI)-linked membrane protein. Inhibition of tissue factor by TFPI coincides with the translocation of quaternary complexes containing tissue factor, factor VIIa, factor Xa, and TFPI to detergent-insoluble plasma membrane domains rich in cholesterol, sphingomyelin, and GPI-linked proteins known as lipid rafts and caveolae. It is not known if localization of TFPI to these membrane domains is required for its inhibition of tissue factor procoagulant activity. We generated chimeric TFPI molecules linked directly to the plasma membrane via a GPI anchor or hydrophobic transmembrane domain and expressed these in HEK293 cells that produce tissue factor but not endogenous TFPI. The GPI-anchored chimera was exclusively enriched in detergent-insoluble membrane fractions while the transmembrane molecule was not. Transfectants expressing equal levels of the GPI-linked or transmembrane TFPI displayed equal anticoagulant potency as assessed by tissue factor-mediated conversion of factor X to factor Xa. Disruption of lipid rafts with cyclodextrin likewise had no effect on the inhibitory activity of the transmembrane or GPI-linked TFPI chimeras in HEK293 cells, nor on endogenous TFPI expressed by ECV304 cells. Thus, we conclude that the GPI anchor and membrane localization to lipid rafts does not enhance inhibition of factor VIIa/tissue factor by cell-surface associated TFPI.

Characterization of the association of tissue factor pathway inhibitor with human placenta.

OBJECTIVE: Tissue factor pathway inhibitor (TFPI) is an endothelial-associated inhibitor of blood coagulation. Because the mechanism for attachment of TFPI to endothelium is not clear, we investigated its association with human placenta. METHODS AND RESULTS: Western blots demonstrate that treatment with phosphatidylinositol-specific phospholipase C (PIPLC) removes more placental TFPI than either PBS or heparin, a finding confirmed by immunohistochemistry. The amounts of heparin-releasable and PIPLC-releasable TFPI activity on placental endothelium were measured in placentas from 5 individuals. PIPLC removes >10-fold more TFPI activity from the placental fragments than 10 U/mL heparin and >100-fold more than 1 U/mL heparin. Pretreatment of the placental fragments with PIPLC increases the amount of heparin-releasable TFPI by approximately 3-fold. An antibody specific for the C-terminal region of TFPI recognizes PIPLC-releasable TFPI in Western blots. CONCLUSIONS: GPI-anchored TFPI is the predominant form on placental endothelium. Heparin-releasable TFPI likely represents only a small portion of the total TFPI on endothelium that remains attached to cell-surface glycosaminoglycans after cleavage of the GPI anchor by endogenous enzymes. The predominance of GPI-anchored TFPI suggests that heparin infusion does not significantly redistribute TFPI within the vasculature. The intact C-terminus in GPI-anchored TFPI indicates it is not directly attached to a GPI anchor. Rather, it most likely associates with endothelium by binding to a GPI-anchored protein.