Toward a Long-Term Artificial Lung.

Only a very small portion of end-stage organ failures can be treated by transplantation because of the shortage of donor organs. Although artificial long-term organ support such as ventricular assist devices provide therapeutic options serving as a bridge-to-transplantation or destination therapy for end-stage heart failure, suitable long-term artificial lung systems are still at an early stage of development. Although a short-term use of an extracorporeal lung support is feasible today, the currently available technical solutions do not permit the long-term use of lung replacement systems in terms of an implantable artificial lung. This is currently limited by a variety of factors: biocompatibility problems lead to clot formation within the system, especially in areas with unphysiological flow conditions. In addition, proteins, cells, and fibrin are deposited on the membranes, decreasing gas exchange performance and thus, limiting long-term use. Coordinated basic and translational scientific research to solve these problems is therefore necessary to enable the long-term use and implantation of an artificial lung. Strategies for improving the biocompatibility of foreign surfaces, for new anticoagulation regimes, for optimization of gas and blood flow, and for miniaturization of these systems must be found. These strategies must be validated by in vitro and in vivo tests, which remain to be developed. In addition, the influence of long-term support on the pathophysiology must be considered. These challenges require well-connected interdisciplinary teams from the natural and material sciences, engineering, and medicine, which take the necessary steps toward the development of an artificial implantable lung.

Development, preclinical evaluation and validation of a novel quick vascular closure device for transluminal, cardiac and radiological arterial catheterization.

Following percutaneous coronary intervention, vascular closure devices (VCDs) are increasingly used to reduce time to ambulation, enhance patient comfort, and reduce potential complications compared with traditional manual compression. Newer techniques include complicated, more or less automated suture devices, local application of pads or the use of metal clips and staples. These techniques often have the disadvantage of being time consuming, expensive or not efficient enough. The VCD failure rate in association with vascular complications of 2.0-9.5%, depending on the type of VCD, is still not acceptable. Therefore, the aim of this study is to develop a self-expanding quick vascular closure device (QVCD) made from a bioabsorbable elastic polymer that can be easily applied through the placed introducer sheath. Bioabsorbable block-co-polymers were synthesized and the chemical and mechanical degradation were determined by in vitro tests. The best fitting polymer was selected for further investigation and for microinjection moulding. After comprehensive haemocompatibility analyses in vitro, QVCDs were implanted in arterial vessels following arteriotomy for different time points in sheep to investigate the healing process. The in vivo tests proved that the new QVCD can be safely placed in the arteriotomy hole through the existing sheath instantly sealing the vessel. The degradation time of 14 days found in vitro was sufficient for vessel healing. After 4 weeks, the remaining QVCD material was covered by neointima. Overall, our experiments showed the safety and feasibility of applying this novel QVCD through an existing arterial sheath and hence encourage future work with larger calibers.

Evaluation of platelet adhesion and activation on polymers: Round-robin study to assess inter-center variability.

The regulatory agencies provide recommendations rather than protocols or standard operation procedures for the hemocompatibility evaluation of novel materials e.g. for cardiovascular applications. Thus, there is a lack of specifications with regard to test setups and procedures. As a consequence, laboratories worldwide perform in vitro assays under substantially different test conditions, so that inter-laboratory and inter-study comparisons are impossible. Here, we report about a prospective, randomized and double-blind multicenter trial which demonstrates that standardization of in vitro test protocols allows a reproducible assessment of platelet adhesion and activation from fresh human platelet rich plasma as possible indicators of the thrombogenicity of cardiovascular implants. Standardization of the reported static in vitro setup resulted in a laboratory independent scoring of the following materials: poly(dimethyl siloxane) (PDMS), poly(ethylene terephthalate) (PET) and poly(tetrafluoro ethylene) (PTFE). The results of this in vitro study provide evidence that inter-laboratory and inter-study comparisons can be achieved for the evaluation of the adhesion and activation of platelets on blood-contacting biomaterials by stringent standardization of test protocols.

Surface biofunctionalization of ß-TCP blocks using aptamer 74 for bone tissue engineering.

Successful bone regeneration following oral and maxillofacial surgeries depends on efficient functionalization strategies that allow the recruitment of osteogenic progenitor cells at the tissue/implant interface. We have previously identified aptamer 74, which exhibited a binding affinity for osteogenically induced jaw periosteal cells (JPCs). In the present study, this aptamer was used for the surface biofunctionalization of ß-tricalcium phosphate (ß-TCP) blocks. Atomic force microscopy (AFM) measurements showed increased binding activity of aptamer 74 towards osteogenically induced JPCs compared to untreated controls. The immobilization efficiency of aptamer 74 was analyzed using the QuantiFluor ssDNA assay for 2D surfaces and by amino acid analysis for 3D ß-TCP constructs. Following the successful immobilization of aptamer 74 in 2D culture wells and on 3D constructs, in vitro assays showed no significant differences in cell proliferation compared to unmodified surfaces. Interestingly, JPC mineralization was significantly higher on the 2D surfaces and higher cell adhesion was detected on the 3D constructs with immobilized aptamer. Herein, we report an established, biocompatible ß-TCP matrix with surface immobilization of aptamer 74, which enhances properties such as cell adhesion on 3D constructs and mineralization on 2D surfaces. Further studies need to be performed to improve the immobilization efficiency and to develop a suitable approach for JPC mineralization growing within 3D ß-TCP constructs.

Intrajugular balloon catheter reduces air embolism in vitro and in vivo.

BACKGROUND: Neurosurgical procedures requiring a sitting position may put the patient at risk of a potentially life-threatening air embolism. Transient manual jugular venous compression limits further air entry in this situation. This study presents an alternative technique aimed at reducing the risk of air embolism. METHODS: In an in vitro model, an intrajugular balloon catheter was inserted to demonstrate that this device prevents air embolism. In an in vivo study, this device was bilaterally placed into jugular vessels in pigs. Using an ultrasound technique, blood flow was monitored and jugular venous pressure was recorded before and during cuff inflation. Air was applied proximally to the inflated cuffs to test the hypothesis that this novel device blocks air passage. RESULTS: In vitro, the intrajugular balloon catheter reliably prevented further air entry (n=10). Additionally, accumulated air could be aspirated from an orifice of the catheter (n=10). In vivo, inflation of the catheter balloon completely obstructed venous blood flow (n=8). Bilateral inflation of the cuff significantly increased the proximal jugular venous pressure from 9.8 (2.4) mm Hg to 14.5 (2.5) mm Hg (n=8, P<0.05). Under conditions mimicking an air embolism, air passage across the inflated cuffs was prevented and 78 (20%) (n=6) of the air dose could be aspirated by the proximal orifice of the catheter. CONCLUSIONS: These findings may serve as a starting point for the development of intrajugular balloon catheters designed to reduce the risk of air embolism in patients undergoing neurosurgery in a sitting position.

Preclinical evaluation of the thrombogenicity and endothelialization of bare metal and surface-coated neurovascular stents.

BACKGROUND AND PURPOSE: Stent-assisted coiling is routinely used for the endovascular treatment of complex or wide-neck intracranial aneurysms. However, in-stent thrombosis, thromboembolic events, and ischemic complications remain a major concern associated with stent implants. Therefore, a novel low-profile neurovascular stent with a bare metal surface was investigated with regard to thrombogenicity and endothelialization and compared with the same stent coated with albumin or heparin. MATERIALS AND METHODS: The bare metal and heparin- or albumin-coated stents were loaded in heparin-coated tubing, which was then filled with heparinized human blood (n = 5) and circulated at 150 mL/min and 37°C for 60 minutes. Before and after circulation, measurement of various inflammation and coagulation markers and scanning electron microscopy were performed. Endothelialization of the stents was investigated in an in vitro model including human umbilical vascular endothelial cells. RESULTS: Our results showed that platelet loss and platelet activation and activation of the coagulation cascade, which are induced by the bare metal stents, were significantly reduced by heparin or albumin coating. Adverse effects on erythrocytes, leukocytes, and the complement cascade were not induced by the bare metal or coated stents. Moreover, the bare metal and albumin-coated stents showed good endothelialization properties. CONCLUSIONS: Albumin and heparin coatings clearly improve the thrombogenicity of the stents and thus may represent fundamental progress in the treatment of intracranial aneurysms. Moreover, preclinical evaluation of neurovascular stents under physiologic conditions supports and accelerates the development of more biocompatible neurovascular stents.

Comparison of different in vitro tests for biocompatibility screening of Mg alloys.

Standard cell culture tests according to ISO 10993 have only limited value for the biocompatibility screening of degradable biomaterials such as Mg alloys. The correlation between in vitro and in vivo results is poor. Standard cytotoxicity tests mimic the clinical situation to only a limited extent, since in vivo proteins and macromolecules in the blood and interstitial liquid will influence the corrosion behaviour and, hence, biocompatibility of Mg alloys to a significant extent. We therefore developed a modified cytotoxicity test simulating the in vivo conditions by use of bovine serum as the extraction vehicle instead of the cell culture medium routinely used in standard cytotoxicity testing according to ISO 10993-5. The modified extraction test was applied to eight experimental Mg alloys. Cytotoxicity was assayed by inhibition of cell metabolic activity (XTT test). When extraction of the alloy samples was performed in serum instead of cell culture medium the metabolic activity was significantly less inhibited for six of the eight alloys. The reduction in apparent cytotoxicity under serum extraction conditions was most pronounced for MgZn1 (109% relative metabolic activity with serum extracts vs. 26% in Dulbecco's modified Eagle's medium (DMEM)), for MgY4 (103% in serum vs. 32% in DMEM) and for MgAl3Zn1 (84% vs. 17%), resulting in a completely different cytotoxicity ranking of the tested materials when serum extraction was used. We suppose that this test system has the potential to enhance the predictability of in vivo corrosion behaviour and biocompatibility of Mg-based materials for biodegradable medical devices.

Bioactive coronary stent coating based on layer-by-layer technology for siRNA release.

One procedure to treat stenotic coronary arteries is the percutaneous transluminal coronary angioplasty (PTCA). In recent years, drug-eluting stents (DESs) have demonstrated elaborate ways to improve outcomes of intravascular interventions. To enhance DESs, the idea has evolved to design stents that elute specific small interfering RNA (siRNA) for better vascular wall regeneration. Layer-by-layer (LbL) technology offers the possibility of incorporating siRNA nanoplexes (NPs) to achieve bioactive medical implant coatings. The LbL technique was used to achieve hyaluronic acid/chitosan (HA/Chi) films with incorporated Chi-siRNA NPs. The multilayer growth was monitored by quartz crystal microbalance. The coating on the stents and its thickness were analyzed using fluorescence and scanning electron microscopy. All stents showed a homogeneous coating, and the polyelectrolyte multilayers (PEMs) were not disrupted after ethylene oxide sterilization or expansion. The in vitro uptake of fluorescent-labeled NPs from PEMs in primary human endothelial cells (ECs) was analyzed by flow cytometry for 2, 6 and 9 days. Furthermore, stents coated with HA/Chi and Chi-siRNA NPs were expanded into porcine arteries and showed ex vivo delivery of NPs. The films showed no critical results in terms of hemocompatibility. This study demonstrates that Chi-siRNA NPs can be incorporated into PEMs consisting of HA and Chi. We conclude that the NPs were delivered to ECs under in vitro conditions. Furthermore, under ex vivo conditions, NPs were transferred into porcine artery walls. Due to their good hemocompatibility, they might make an innovative tool for achieving bioactive coatings for coronary stents.

Short-acting P2Y12 blockade to reduce platelet dysfunction and coagulopathy during experimental extracorporeal circulation and hypothermia.

BACKGROUND: Extracorporeal circulation (ECC) and hypothermia are routinely used in cardiac surgery to maintain stable circulatory parameters and to increase the ischaemic tolerance of the patient. However, ECC and hypothermia cause platelet activation and dysfunction possibly followed by a devastating coagulopathy. Stimulation of the adenosinediphosphate (ADP) receptor P(2)Y(12) plays a pivotal role in platelet activation. This experimental study tested P(2)Y(12) receptor blockade as an approach to protect platelets during ECC. METHODS: Human blood was treated with the short-acting P(2)Y(12) blocker cangrelor (1 µM, t(1/2)<5 min) or the P(2)Y(12) inhibitor 2-MeSAMP (100 µM) and circulated in an ex vivo ECC model at normothermia (37°C) and hypothermia (28°C). Before and after circulation, markers of platelet activation and of coagulation (thrombin-antithrombin complex generation) were analysed. During hypothermic ECC in pigs, the effect of reversible P(2)Y(12) blockade on platelet function was evaluated by cangrelor infusion (0.075 µg kg(-1) min(-1)). RESULTS: During ex vivo hypothermic ECC, P(2)Y(12) blockade inhibited platelet granule release (P<0.01), platelet-granulocyte binding (P<0.05), and platelet loss (P<0.001), whereas no effects on platelet-ECC binding, platelet CD42bα expression, glycoprotein IIb/IIIa activation, or thrombin-antithrombin complex generation were observed. During hypothermic ECC in pigs, cangrelor inhibited platelet-fibrinogen binding (P<0.05) and ADP-induced platelet aggregation (P<0.001). Platelet function was rapidly restored after termination of cangrelor infusion. CONCLUSIONS: P(2)Y(12) blockade by cangrelor prevents platelet activation during ECC and hypothermia. Owing to its short half-life, platelet inhibition can be well controlled, thus potentially reducing bleeding complications. This novel pharmacological strategy has the potential to reduce complications associated with ECC and hypothermia.

Formation and photocatalytic decomposition of a pellicle on anatase surfaces.

The acquired dental pellicle plays a critical role in the adhesion and detachment of dental plaque bacteria. It has been reported that titanium dioxide biomaterials decompose single-protein films by photocatalysis. However, it is not known whether this can also be achieved with complex structured pellicle films. This in vitro study investigated in real-time the formation and photocatalytic decomposition of human pellicle at anatase-saliva interfaces. Nanostructured polycrystalline anatase layers were deposited on titanium-coated quartz crystals by magnetron-sputtering, serving as a model for titanium implant surfaces. The quartz crystals were used as acoustic sensors in a quartz-crystal microbalance (QCM) system with dissipation. In situ UV irradiation of pellicle-covered anatase caused a statistically significant decrease of the adsorbed salivary mass. In contrast, photocatalytic decomposition of pellicle could not be observed on reference titanium surfaces. Wettability characterization revealed superhydrophilicity of anatase upon UV irradiation, whereas titanium was unaffected. XPS measurements provide further information concerning the decomposition of the salivary films. The results suggest that the photocatalytic activity of polycrystalline anatase-modified biomaterial surfaces is able to decompose complex structured macromolecular pellicle films. Therefore, this study opens the way to surface modifications supporting therapeutic approaches of biofilm removal.

DEHP and its active metabolites: leaching from different tubing types, impact on proinflammatory cytokines and adhesion molecule expression. Is there a subsumable context?

INTRODUCTION: Di(2-ethylhexyl)phthalate (DEHP) is suspected to be toxic for several reasons. During contact with a lipophilic medium, DEHP leaks from polyvinylchloride (PVC), but its influence on inflammatory reactions remains unknown. We examined specific DEHP leaching out of different tubing types, the possibly modulated liberation of proinflammatory cytokines and the induction of adhesion molecule expression in primary endothelial cells. MATERIALS AND METHODS: Blood samples were circulated in traditional PVC, nodioctyl phthalate (DOP) PVC and heparin-coated PVC tubing within a Chandler loop model. The blood was tested for the concentration of DEHP and its active metabolites as well as the liberation of the proinflammatory cytokines TNFα and IL1ß. Furthermore, we exposed human endothelial cells to circulated blood and analysed them for the expression of the adhesion molecules ICAM-1, VCAM-1 and E-selectin. RESULTS: In contrast to the other tubing, PVC tubing showed significantly elevated DEHP levels, but no alteration was observed concerning a potential up-regulation of the cytokines or activation of the endothelial adhesion molecule receptors. CONCLUSIONS: Our data conclude that there is no correlation between DEHP leaching and the inflammatory response after ECC support, but this study showed that even DEHP-free material is leaching DEHP and its toxic metabolites.

Transfection of short-interfering RNA silences adhesion molecule expression on cardiac microvascular cells.

BACKGROUND: Acute rejection reactions and the development of graft arterial disease are serious limitations after transplantation. Both are connected to the expression of adhesion molecules on the activated microvascular endothelium of the allograft. METHODS: siRNA-mediated silencing of ESELE, ICAM-1, and VCAM-1 on human cardiac microvascular cells (HCMEC) was investigated in order to inhibit leukocyte-endothelial interactions. HCMEC were investigated for the time-dependent expression of ESELE, ICAM-1, and VCAM-1 after TNF-α stimulation and for siRNA mediated suppression using a nonviral transfecting approach. Furthermore, the effects of siRNA transfection on leukocyte binding to the endothelium were analyzed. RESULTS: Transfection with siRNA induced a significant suppression of adhesion molecule expression, regardless of whether there had been a prior single or cocktail transfection of the sequences ( P < 0.05). The quantity of attaching leukocytes was significantly reduced after an equal silencing of adhesion molecules ( P < 0.05). CONCLUSIONS: This investigation demonstrates that liposomal transfection of HCMEC with specific siRNA sequences is capable of both repressing adhesion molecule expression and of reducing subsequent leukocyte-endothelial actions.

Veins are no arteries: even moderate arterial pressure induces significant adhesion molecule expression of vein grafts in an ex vivo circulation model.

AIM: Coronary artery bypass grafting (CABG) is a standard procedure for treatment of coronary heart disease. Eighty percent of all CABGs are performed with venous grafts which then get exposed to an arterial pressure after surgery. This widely used procedure, however, is complicated by the development of alterations in the vein graft wall, leading to a decreased patency rate and graft failure. This study enlightens the influence of an even moderate arterial pressure on the gene expression of adhesion molecules in venous grafts which play a decisive role for the early induction of atherogenesis. METHODS: Segments of porcine vena jugularis and arteria carotis were mounted in a simulated bypass circuit and subjected to pulsatile flow. Vessel segments were examined for adhesion molecule expression with quantitative real-time - polymerase chain reaction (qRT-PCR) and adherence of leukocytes was observed by confocal laser scanning microscopy and scanning electron microscopy. RESULTS: Veins grafts subjected to an even moderate arterial pressure showed a 14-fold increase of ICAM-1 expression already after 4 hours. An arterial pressure of around 100/80 mmHg was enough to stimulate the adhesion molecule expression Furthermore it led to a 9-fold increase of leukocyte adhesion to the venous endothelium, but, in contrast this was not the case in arteries. CONCLUSION: This study showed, that already 100 mmHg upregulates the expression of several adhesion molecules in pig veins followed by increased adhesion of leukocytes. Therefore, our data demonstrate the advantage of arteries for CABG, and that new therapeutic strategies are urgently necessary to protect vein grafts either physically or pharmacologically if arteries are not available for CABG.

Current concepts and new developments for autologous in vivo endothelialisation of biomaterials for intravascular applications.

Circulating endothelial progenitor cells (EPCs) in the peripheral blood of adults represent an auspicious cell source for tissue engineering of an autologous endothelium on blood-contacting implants. Novel materials biofunctionalised with EPC-specific capture molecules represent an intriguing strategy for induction of selective homing of progenitor cells. The trapped EPCs can differentiate into endothelial cells and generate a non-thrombogenic surface on artificial materials. However, the success of this process mainly depends on the use of optimised capture molecules with a high selectivity and affinity. In recent years, various biomedical engineering strategies have emerged for in situ immobilisation of patient's own stem cells on blood contacting materials. The realisation of this in vivo tissue engineering concept and generation of an endothelium on artificial surfaces could exceedingly enhance the performance of not only small calibre vascular grafts and stents, but also, in general all blood-contacting medical devices, such as heart valves, artificial lungs, hearts, kidneys, and ventricular assist devices.

Multifunctional nature of UV-irradiated nanocrystalline anatase thin films for biomedical applications.

Anatase is known to decompose organic material by photocatalysis and to enhance surface wettability once irradiated by ultraviolet (UV) light. In this study, pulse magnetron-sputtered anatase thin films were investigated for their suitability with respect to specific biomedical applications, namely superhydrophilic and biofilm degrading implant surfaces. UV-induced hydrophilicity was quantified by static and dynamic contact angle analysis. Photocatalytic protein decomposition was analyzed by quartz crystal microbalance with dissipation. The surfaces were characterized by X-ray diffraction, atomic force microscopy, scanning electron microscopy and X-ray photoelectron spectroscopy. The radical formation on anatase, responsible for photocatalytic effects, was analyzed by electron spin resonance spectroscopy. Results have shown that the nanocrystalline anatase films, in contrast to reference titanium surfaces, were sensitive to UV irradiation and showed rapid switching towards superhydrophilicity. The observed decrease in carbon adsorbents and the increase in the fraction of surface hydroxyl groups upon UV irradiation might contribute to this hydrophilic behavior. UV irradiation of anatase pre-conditioned with albumin protein layers induces the photocatalytic decomposition of these model biofilms. The observed degradation is mainly caused by hydroxyl radicals. It is concluded that nanocrystalline anatase films offer different functions at implant interfaces, e.g. bedside hydrophilization of anatase-coated implants for improved osseointegration or the in situ decomposition of conditioning films forming the basal layer of biofilms in the oral cavity.

Cellulose pads (Hydrotin-C): a new solid coupling agent.

OBJECTIVES: Hydrotin-C is a bacterial cellulose consisting of 90% of physiological solution. The cellulose pad is smooth and elastic and cannot be torn, and is therefore suitable for use on any local skin condition. The aim of this study was to evaluate the image quality (IQ) and transducer pressure of Hydrotin-C-Pads as a solid coupling agent compared with standard coupling gel. METHODS: To date, transthoracic echocardiography has been performed in 51 patients (ongoing study) using standard coupling gel and Hydrotin-C-Pads. The IQ was categorized as follows: 1 = excellent, 2 = good, 3 = poor, 4 = inadequate. All patients were questioned about the transducer pressure, which was categorized as: 0 = less, 1 = equal, 2 = more. The body mass index (BMI) was documented for all patients. RESULTS: The IQ was very good with Hydrotin-C-Pads. The IQ was better in 84% of all patients. In patients with a BMI of over 25 and in patients with a BMI of over 30, Hydrotin-C-Pads was also better compared with standard gel. The transducer pressure was lower in 33 patients. CONCLUSIONS: Hydrotin-C-Pads can be used as a solid ultrasonic coupling agent. Because of the consistent thickness of the coupling zone, the IQ seems to be better for Hydrotin-C-Pads compared with standard coupling gel. The transducer presser is lower, and these pads are easy to apply and are reusable and reduce the burden of clean-up after treatment.

Aptamer-based isolation and subsequent imaging of mesenchymal stem cells in ischemic myocard by magnetic resonance imaging.

PURPOSE: Mesenchymal stem cells (MSC) seem to be a promising cell source for cellular cardiomyoplasty. We recently developed a new aptamer-based specific selection of MSC to provide "ready to transplant" cells directly after isolation. We evaluated MRI tracking of newly isolated and freshly transplanted MSC in the heart using one short ex vivo selection step combining specific aptamer-based isolation and labeling of the cells. MATERIALS AND METHODS: Bone marrow (BM) was collected from healthy pigs. The animals were euthanized and the heart was placed in a perfusion model. During cold ischemia, immunomagnetic isolation of MSC from the BM by MSC-specific aptamers labeled with Dynabeads was performed within 2 h. For histological identification the cells were additionally stained with PKH26. Approx. 3 x 10(6) of the freshly aptamer-isolated cells were injected into the ramus interventricularis anterior (RIVA) and 5 x 10(5) cells were injected directly into myocardial tissue after damaging the respective area by freezing (cryo-scar). 3 x 10(6) of the aptamer-isolated cells were kept for further characterization (FACS and differentiation assays). 20 h after cell transplantation, MRI of the heart using a clinical 3.0 Tesla whole body scanner (Magnetom Trio, Siemens, Germany) was performed followed by histological examinations. RESULTS: The average yield of sorted cells from 120 ml BM was 7 x 10(6) cells. The cells were cultured and showed MSC-like properties. MRI showed reproducible artifacts within the RIVA-perfusion area and the cryo-scar with surprisingly excellent quality. The histological examination of the biopsies showed PKH26-positive cells within the areas which were positive in the MRI in contrast to the control biopsies. CONCLUSION: Immunomagnetic separation of MSC by specific aptamers linked to magnetic particles is feasible, effective and combines a specific separation and labeling technique to a "one stop shop" strategy.

Endotoxin removal from albumin and saline solutions.

Microporous membranes have been developed which can remove endotoxins selectively from electrolyte and albumin solutions by regioselective adsorption in the membrane matrix and outside surface of the membrane. The membranes were prepared in the form of hollow fibre membranes in a continuous process. By varying the membrane preparation parameters, different pore sizes and adsorption capacities could be realized, thus broadening applications for biological purification. Dynamic adsorption capacities for endotoxin from albumin and saline solution were determined and were found to be in the range of 0.2 and 0.1 microg endotoxin/g membrane, respectively, suggesting different adsorption mechanisms.

Inducible HSP70 levels in thoracic wound fluid indicate myocardial damage after open heart surgery.

AIM: Heat shock proteins (HSP) act as chaperones and protect cellular homeostasis. They are induced by multiple forms of cell stress. Nothing is known about their rise in the drainage fluid after cardiac and thoracic operations. METHODS: Forty-five patients undergoing open heart surgery or major lung resections were studied. Levels of inducible HSP70 were measured daily in thoracic wound fluid and blood with ELISA tests in addition to multiple conventional blood and wound fluid parameters. Data were analyzed using analysis of covariance and Spearman's rank-correlation coefficient. RESULTS: Inducible HSP70 was detectable in all thoracic fluid samples. Mean HSP70-level after open heart surgery was 64 ng/mL and 40 ng/mL in patients without extracorporeal circulation (P=0.015). Drainage HSP70 levels were correlated with white blood cell counts in wound fluid (P<0.0001) and blood CK-MB levels (P<0.0001). Correlation coefficients between white blood cells in blood and local HSP-levels were explicitly lower (P=0.0023). There was a steady decrease in HSP levels in drainage fluid from day 1 to day 4 (P<0.0001). CONCLUSIONS: Inducible HSP70 is present in postoperative thoracic drainage fluid. Drainage levels are higher in patients after open heart surgery and are correlated with blood CK-MB levels, indicating cardiomyocyte damage. Our data further suggest that extracellular HSP70 is released also by white blood cells. Additional studies are needed to examine the role of extracellular HSP70 in wound healing processes and myocardial damage.

Tirofiban (Aggrastat) protects platelets and decreases platelet-granulocyte binding in an extracorporeal circulation model.

OBJECTIVES: Extracorporeal circulation (ECC) induces platelet activation and inflammation with potentially life-threatening organ dysfunction. Short-acting GP IIb/IIIa inhibitors like tirofiban and eptifibatide protect platelets during ECC without increasing bleeding complications and may reduce inflammation. This study investigates anti-thrombotic and anti-inflammatory effects of different platelet inhibitors. METHODS: Control (untreated) and treated (using either 150 ng/mL tirofiban, 2.5 microg/mL eptifibatide, 0.7 microg/mL milrinone, 15 microg/mL dipyridamol, or 300 KIU/mL aprotinin) heparinized blood of healthy volunteers (n = 6) was recirculated in a well-established ECC model (Chandler loop). Percentage of platelet aggregates, P-selectin-expressing (activated) platelets, CD15-positive aggregates (indicating proinflammatory platelet-granulocyte binding), and platelet counts were determined before (baseline) and after 30 minutes recirculation in unstimulated and ADP-stimulated samples using flow cytometry. Statistical analysis was performed using multifactor ANOVA after transforming the data (logarithms for counts and log odds for percentages). Least square means were backtransformed to obtain appropriate means and their 95 % confidence intervals. Multiple post-hoc comparisons were performed by Tukey's HSD test with a global alpha of 5 %. RESULTS: Significant inhibition was observed for: 1) ECC-induced platelet aggregation by tirofiban (unstimulated: 2.2-fold/stimulated: 2.46-fold), eptifibatide (unstimulated: 1.96-fold/stimulated: 2.65-fold), and milrinone (unstimulated: 1.87-fold/stimulated: 1.37-fold); 2) ECC-induced P-selectin expression by tirofiban (unstimulated: 3.95-fold/stimulated: 2.54-fold), and eptifibatide (unstimulated: 5.87-fold/stimulated: 3.28-fold); 3) ECC-induced platelet loss by tirofiban (1.27-fold), and eptifibatide (1.25-fold); 4) ECC-induced platelet-granulocyte binding by tirofiban (unstimulated: 2.25-fold/stimulated: 1.59-fold), but not by eptifibatide. CONCLUSIONS: Amongst the investigated drugs only GP IIb/IIIa inhibitors decreased activation, aggregation, and loss of platelets during ECC but acted differently on platelet-granulocyte interaction. A short-acting GP IIb/IIIa inhibitor with the potential to inhibit platelet activation and platelet-leukocyte interaction should be considered both for platelet protection and inhibition of platelet-mediated inflammation during ECC.