Differences in human and sheep platelet adherence, aggregation and activation induced by glass beads in a modified chandler loop-system.

In human cardiovascular research, sheep in particular are used as a large animal model in addition to pigs. In these animals, medical products, developed and tested for human medical purposes, are almost exclusively used in interventional studies. Therefore, the extent to which platelets from human and ovine blood differ in terms of adherence, aggregation and activation after a 4- or 8-minutes exposure to glass was investigated. Testing was performed with platelet-rich plasma (PRP) and a modified chandler loop-system, with 4- and 8-minute blood-material exposure times corresponding to 20 and 40 test cycles, respectively, through the entire silicone tube loop of the test system.In sheep and human PRP, contact with the silicone tubing resulted in a decrease in platelet count after 4 minutes and 20 test cycles, respectively. Four more minutes (20 additional test cycles) caused a further decrease of the platelet count only in sheep PRP. When the silicon tube was partly filled with glass beads, these effects were more pronounced and stronger in sheep then in human PRP.The mean platelet volume, which was used as parameter for platelet aggregation, did not change over time in human PRP without glass exposure. With glass exposure in human and sheep PRP the mean platelet volume increased within 40 test cycles, but this increase was stronger in sheep than in human PRP.Regarding activation behavior, the activation markers CD62P and CD63 were detectable only in < 30% (sheep) and < 45% (human) of platelets, whereas after 8 min of glass exposure, the proportion of CD62P+ and CD63+ cells was more increased than before only in sheep. These results indicate that ovine platelets adhere more strongly to glass and show stronger aggregation behavior after glass contact than human platelets, but that ovine and human platelets differ only slightly in activability by glass.

Differences in human and minipig platelet number, volume and activation induced by borosilicate glass beads in a modified chandler loop-system.

The pig is the most widely used large animal model in Europe, with cardiovascular research being one of the main areas of application. Adequate refinement of interventional studies in this field, meeting the requirements of Russel and Burchs' 3 R concept, can only be performed if blood-contacting medical devices are hemocompatible. Because most medical devices for cardiovascular interventional procedures are developed for humans they are tested mostly for compatibility with human blood. The aim of this study was therefore to determine whether there are differences in behavior of porcine and human platelets when they come into contact with glass, which was used as an exemplary thrombogenic material. For this purpose changes of platelet count, platelet volume and platelet expression of the activation markers CD61, CD62P and CD63 were measured using a modified chandler loop-system simulating the fluidic effects of the blood flow. Minipig and human platelets showed significant differences in number and volume, but not in activation after 4-8 min exposure to glass.

Cutaneous microcirculation in patients with peripheral arterial occlusive disease: Comparison of capillary blood circulation in the nail fold of finger and toe.

In patients with peripheral arterial occlusive disease (PAOD) a restricted circulation in cutaneous microvessels has been reported. In this study the velocity of erythrocytes (very) in finger nailfold capillaries - a vascular area without upstream macroangiopathy - and also in toe nailfold capillaries - a post-stenotic area -was investigated using capillary microscopy in apparently healthy subjects and patients with PAOD. Already in finger nailfold capillaries very of patients with PAOD under resting conditions was significantly lower than in capillaries of healthy subjects. This was also true for the circulation in toe capillaries. In addition, the erythrocyte velocities under resting conditions in the toe capillaries were significantly lower than in the finger capillaries. Similar results were found for the duration and the maximum velocity of postocclusive hyperemia. It is concluded that the resting blood flow in the skin microcirculation is impaired in PAOD patients, both under resting conditions and during postocclusive hyperemia in finger as well in toe nailfold capillaries.

Pododermal angioarchitecture in the equine hoof wall: A light and scanning electron microscopic study of the wall proper.

BACKGROUND: Blood supply is an important factor for the normal function of the equine hoof, but earlier studies present conflicting data on functional characteristics of its angioarchitecture. OBJECTIVE: Emphasis was laid on demonstration of the microvascularisation in the different hoof wall regions, aiming at assessment of specialised vascular structures, e.g. vascular sphincter mechanisms and arteriovenous anastomoses. METHODS: The angioarchitecture of the adult pododerma in the equine hoof wall was examined by scanning electron microscopy of micro-corrosion casts assisted by exemplary histological and immuno-histochemical characterisation of the pododermal vasculature. RESULTS: The microvasculature of the lamellae and terminal papillae in all hoof wall regions was described in detail. Focal dilations and microvascular sphincters were a common feature. In contrast to former investigations, true arteriovenous anastomoses were detected at the base of the primary lamellae and the terminal papillae only, while thoroughfare channels proved a regular element within the microvasculature of the wall proper. Bicuspid venous valves were detected as regular feature. For the first time, the alpha-smooth muscle actin-reactivity of the microvascularisation in the hoof wall was systematically assessed, verifying its specialised vasomotor devices. CONCLUSIONS: The vasculature of the hoof wall displays specific angio-adaptations to high pressure and tensile load.

Growth-related micromorphological characteristics of the porcine femoral artery.

BACKGROUND: In cardiovascular research small pigs breeds like Göttingen® minipigs (GM) are established animal models, but systematic data about the micromorphology of the GM vasculature at different ages are scarce. OBJECTIVE: The study was aimed at gaining knowledge about the micromorphology of the femoral artery (FA) from German Landrace pigs (DL) and GM during the period of growth over a body weight range of 10-40 kg. METHODS: FA samples from DL aged two or three months were compared to GM ones, aged 18 or 40 months using transmitted light microscopy. RESULTS: All FA samples showed typical characteristics of muscular arteries. Growth was associated with increased vessel wall thickness. In the GM this resulted in a slight decrease of the luminal diameter (LD), while in the DL pigs, an increase of the LD and smooth muscle cell content (10%) with decreased elastic fiber content (10%) has been detected. In contrast, within the 22 months lasting growth period of the GM, the tunica media content of smooth muscle cells and elastic fibers remained stable. CONCLUSIONS: FA maturation strongly depends on the pig breed and age. It can be different from what is described in humans.

Albumin solder covalently bound to a polymer membrane: New approach to improve binding strength in laser tissue soldering in-vitro.

Laser tissue soldering (LTS) based on indocyanine green (ICG)-mediated heat-denaturation of proteins might be a promising alternative technique for micro-suturing, but up to now the problem of too weak shear strength of the solder welds in comparison to sutures is not solved. Earlier reports gave promising results showing that solder supported by carrier materials can enhance the cohesive strength of the liquid solder. In these studies, the solder was applied to the carriers by dip coating. Higher reliability of the connection between the solder and the carrier material is expected when the solder is bound covalently to the carrier material. In the present study a poly(ether imide) (PEI) membrane served as carrier material and ICG-supplemented albumin as solder substrate. The latter was covalently coupled to the carrier membrane under physiological conditions to prevent structural protein changes. As laser source a diode continuous-wave laser emitting at 808 nm with intensities between 250 mW and 1500 mW was utilized. The albumin functionalized carrier membrane was placed onto the tunica media of explanted pig thoracic aortae forming an overlapping area of approximately 0.5×0.5 cm2. All tests were performed in a dry state to prevent laser light absorption by water. Infrared spectroscopy, spectro-photometrical determination of the secondary and primary amine groups after acid orange II staining, contact angle measurements, and atomic force microscopy proved the successful functionalization of the PEI membrane with albumin. A laser power of 450 mW LTS could generate a membrane-blood vessel connection which was characterized by a shear strength of 0.08±0.002 MPa, corresponding to 15% of the tensile strength of the native blood vessel. Theoretically, an overlapping zone of 4.1 mm around the entire circumference of the blood vessel could have provided shear strength of the PEI membrane-blood vessel compound identical to the tensile strength of the native blood vessel. These in-vitro results confirmed the beneficial effects of solder reinforcement by carrier membranes, and suggest LTS with covalently bound solders on PEI substrates for further studies in animal models.

Effects of Tacrolimus or Sirolimus on the adhesion of vascular wall cells: Controlled in-vitro comparison study.

In drug eluting stents the cytostatic drugs Sirolimus or Tacrolimus are used to inhibit blood vessel restenosis by limiting the proliferation of smooth muscle cells. However, the cytostatic activity of both drugs was shown to be not cell specific and could also affect the stent endothelialisation, respectively. Currently, only limited in vitro data are available about the impact of Sirolimus and Tacrolimus on endothelial cell proliferation over a broad concentration range. To answer this question the following study was performed.Commercially obtained HUVEC were expanded with DMEM cell culture medium (GIBCO, Germany) supplemented with 5 vol% fetal calf serum on non-coated regular polystyrene-based 24-multiwell plates. For drug testings 2×104 cells/cm2 were seeded and grown for 24 h until 30-40% of the multiwell surfaces were covered and then exposed to Sirolimus (1.0×10-11 - 1.0×10-5 mol/l) or Tacrolimus (2.0×10-8 - 6.2×10-5 mol/l), both dissolved in DMSO. 12, 24 and 48 h after adding the drugs cell numbers per area were quantified by counting the cells in six wells with four fields of view per well, representing 0.6 mm2, using a confocal laser microscope.After 48 h of cell growth in the drug-free cell culture medium, the HUVEC number increased from 2.0×104 to 3.55×104 cells/cm2 (mean cell doubling time: 53.6 h, n = 6). At lower concentrations (≤2.0×10-6 mol/l) Tacrolimus reduced the number of adherent HUVEC significantly less than Sirolimus (p < 0.05). However, at higher concentrations (≥2.07×10-5 mol/l) the effect of Tacrolimus on the number of adherent endothelial cells was significantly greater than that of Sirolimus (p < 0.05). At the highest concentration applied (6.22×10-5 mol/l), Tacrolimus induced detachment of all HUVECs within 12 h after drug application. The number of adherent HUVEC decreased only slightly (about 9%) after Sirolimus application at the highest concentration (1.09×10-5 mol/l).These data show that in a non-flow model the cytostatic drug Tacrolimus reduced the number of adherent endothelial cells less than Sirolimus, as long as the drug concentration did not surpass 10-6 mol/l. At the limits of solubility, Sirolimus (1×10-5 mol/l) reduced the number of adherent endothelial cells less than Tacrolimus (6×10-5 mol/l), which induced detachment of endothelial cells.

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.

Organotypic soft-tissue co-cultures: Morphological changes in microvascular endothelial tubes after incubation with iodinated contrast media.

INTRODUCTION: Clinical complications like thrombosis or anaphylaxis have been described to go along with the intra-venous or intra-arterial injection of iodinated contrast media (CM). It has been suggested that the administration of CM affects rheological parameters and thereby causes reduced blood velocity in microvessels. In vitro studies revealed significant buckling of endothelial cells after exposure to CM reducing the lumen of vessels. The aim of this study was to test the influence of CM on three-dimensional microvascular tubules with open lumina within an organotypic soft-tissue co-culture assay in vitro. This model, which is based on the co-culture of endothelial cells and fibroblasts, allows the analysis and quantitation of different parameters of microvascular endothelial capillary structures. MATERIAL AND METHODS: Human dermal fibroblasts and human dermal microvascular endothelial cells were co-cultured for 10 days. Fibroblasts were adapted to the endothelial cell medium before co-culture and allowed to proliferate as well as produce extracellular matrix. The co-cultures were exposed to three different CM, i.e., Iomeprol (Imeron 400MCT), Iodixanol (Visipaque 320) or Iohexol (Accupaque 350) for 1.5 minutes or 5.0 minutes, respectively. For this, a mixture of CM and cell culture medium in a ratio of 30% CM by volume was prepared. After fixation in methanol/acetone, the endothelial cells were immunolabeled with the endothelial marker anti-CD31 and the tubular structures were assessed morphometrically. RESULTS: In the organotypic soft-tissue co-cultures with fibroblasts, the endothelial cells developed three-dimensional capillary-like structures which expanded via sprouting branches. After incubation with the different CM, the numbers of endothelial tubes (p = 0.001) and their lengths (p = 0.003) were significantly lower after the 5 minutes incubation time, when compared to the 1.5 minutes incubation time. The tubular diameters were significantly reduced after 5 minutes (p < 0.001), when compared to the 1.5 minutes incubation duration. Interestingly, Iomeprol and Iodixanol induced an elongation of the tubular branches during incubation duration of 1.5 minutes (p = 0.015). However, after 5 minutes incubation, the tubular branches were drastically shorter in the presence of Iomeprol and Iodixanol than the tubular branches of the control (p = 0.007). SUMMARY AND CONCLUSION: All CM exerted a negative effect on the parameters of in vitro blood vessel development.

Tissue reaction induced by implanted venous access ports in adult patients after infection of the implantation site.

Implantable long-term central venous port systems (CVPS) are widely used as a permanent means of accessing the vascular system for intravenous delivery of drugs, parenteral nutrition, blood transfusion, and blood sampling. These systems allow easy and repetitive puncture without causing much damage to the vessels. However, the body foreign surface of CVPS induces an inflammatory response with varying intensity (depending on the implant materials) that leads to formation of a fibrous tissue capsule around the implant. This study was designed to investigate the influence of bacterial infection on the tissue reaction induced by implanted CVPS in adult patients. 20 patients (9 women, 11 men, 58 ± 14 yrs of age) were included in this study. These patients received explantation of a polysulfone based CVPS (ChemoSite™, Covidien, Mansfield, USA) due to port related infections (patients with bacterial infections at the implantation site: group A, 5 men, 1 women) or to other reasons such as termination of treatment, thrombosis, or CVPS dysfunction (patients without bacterial infections, group B, 6 men, 8 women) 299.9 ± 261.2 days after CVPS implantation. A sample of the encapsulating tissue covering the CVPS together with surrounding tissue (at least 1 × 1 cm2) was placed in a small container with fixing agent, a buffered neutral 4% formalin solution (pH 7). Histological sections of the samples were prepared for light microscopic analysis after paraffin embedding. Sections of 3 µm were cut and stained with haematoxylin and eosin, Weigert's elastic stain, and Heidenhain's azan stain. There was no difference in thickness, collagen and elastin content, or cell and capillary density of the fibrous capsule between both groups. Due to the wound healing reaction involving angiogenesis and fibroblast activation cell density and number of capillaries in the capsule tissue of all patients showed a positive correlation (r = 0.45, p < 0.05). However, the study demonstrated that at the end of the foreign body reaction the artificial tissue layer which covers the CVPS after implantation due to foreign body reaction shows only low reactivity towards infections.

Influence of iodine-containing radiographic contrast media on the phenotype of erythrocytes from different laboratory animal species.

It is well known that clinically relevant concentrations of iodine-containing radiographic contrast media (CM) induce morphological changes in human erythrocytes. However, there are only few reports about CM effects on erythrocytes of animals (e.g. mice, rats, rabbits, and pigs). Thus, two conventional iodine-containing CM (iodixanol, Visipaque™ 320; iomeprol, Iomeprol™ 350) were tested for their effects on the morphology of erythrocytes from these. After venous blood sampling and blood centrifugation, the autologous plasma was supplemented with 40 vol% CM. Then, a defined number of erythrocytes was incubated in this CM-supplemented plasma for 5 min at body temperature (37°C). Subsequently, 10 µL of the cell suspension were transferred to a purified glass slide and the number of discocytes, echinocytes, and acanthocytes was counted within a total number of 100 erythrocytes (40 fold primary magnification, transmitted light mode). Shape changes of the erythrocytes from all animal species strongly depended on the type of CM and compared to the effects which have already been described for human erythrocytes. Incubation in both CM resulted in morphological changes of the erythrocytes. Incubation in a iodixanol/plasma mixture induced the lowest echinocyte or acanthocyte formation. Porcine erythrocytes showed a much more distinct shape change than those of other animal species and humans. These results suggest erythrocytes from mice, rats, and rabbits are a suitable model system for a model system for human erythrocytes when CM effects on the cellular shape of erythrocytes have to be tested. The distinct deformation of the pig erythrocytes could be due to differences in the pig erythrocyte membrane or the physical and chemical constitution of pig erythrocytes.

Influence of a silicon (Si14)-based coating substrate for biomaterials on fibroblast growth and human C5a.

Despite considerable efforts in biomaterial development there is still a lack on substrates for cardiovascular tissue engineering approaches which allow the establishment of a tight a functional endothelial layer on their surface to provide hemocompatibility. The study aimed to test the biocompatibility of a silicon (Si14)-based coating substrate (Supershine Medicare, Permanon) which was designed to resist temperatures from -40°C up to 300°C and which allows the use of established heat-inducing sterilization techniques respectively. By X-ray photoelectron spectroscopy it could be validated that this substrate is able to establish a 40-50 nm thick layer of silica, oxygen and carbon without including any further elements from the substrate on an exemplary selection of materials (silicone, soda-lime-silica glass, stainless steel). Analysis of the LDH-release, the cell activity/proliferation (MTS assay) and the cell phenotype after growing 3T3 cells with extracts of the coated materials did not indicate any signs of cytotoxicity. Additionally by measuring the C5a release after exposure of the coated materials with human serum it could be demonstrated, that the coating had no impact on the activation of the complement system. These results generally suggest the tested substrate as a promising candidate for the coating of materials which are aimed to be used in cardiovascular tissue engineering approaches.

The effect of prone versus supine positioning of Goettingen minipigs on lung density as viewed by computed tomography.

Minipigs are frequently used for scientific research as they are easy to handle and the dimensions of their vascular system do not change after 20 months of age. Although surgical interventions under anaesthesia are often performed in the supine position the effects of this positioning on lung functionality in minipigs have not been systematically described. This study aimed to analyse the influence of supine positioning on the macrostructure of the lung and the pulmonary density by the use of computed tomography imaging in pre-adult Göttingen minipigs. Twelve pre-adult female minipigs were used in the study and lung density was investigated in both the prone and the supine positions. The time between the scans in prone and supine positions was less than 5 minutes (296 ± 6 sec). In the prone position lung density did not differ between the dorsal and ventral part of the lung (-641 ± 72 Hounsfield units [HU]). However in the supine position there was a ventrodorsal gradient of decreasing density (ventral part of the lung: -497 ± 106 HU, dorsal part of the lung: -723 ± 51 HU). The changes in lung density were not accompanied by changes in lung volume (829 ± 191 ml). These results suggest an influence of the body position on the ventilation/perfusion (V(A)/Q) matching of the lung which could possibly result in lowered lung oxygenation as well as in an increased heart activity in the supine position. Additionally, due to the steep course of the vena cava caudalis from the caval foramen in the diaphragm across to the heart (in contrast to the more shallow course in the prone position) the activity of the heart necessary to pump the venous blood to the right atrium has to be higher in the supine position than in the prone position. In pigs the capacity of the heart to increase frequency is limited due to a diastolic/systolic (D/S) ratio <1. Supine positioning may possibly increase their risk of cardiovascular complications.

Influence of radiographic contrast media (Iodixanol and Iomeprol) on the endothelin-1 release from human arterial and venous endothelial cells cultured on an extracellular matrix.

Various radiographic contrast media (RCM) are available for visualization of blood vessels in interventional cardiology which can vary widely in their physicochemical properties thereby influencing different functions of blood cells. In the in vitro study described here the influence of two RCMs on arterial as well as on venous endothelial cells was compared to control cultures and examined under statical culture conditions, thus eliminating the influence of RCM viscosity almost completely. The supplementation of the culture medium with RCM (30% v/v) resulted in clearly different reactions of the endothelial cells exposed. Exposition to Iodixanol supplemented culture medium was followed by endothelin-1 release from venous endothelial cells which was equivalent to the endothelin-1 release from venous control cultures. Compared to control cultures, venous endothelial cells exposed to culture medium supplemented with Iomeprol displayed a completely different reaction, the increase in endothelin-1 secretion was missing completely after a 12 hours exposure. Following a 12 hours exposure to both RCMs there were no longer endothelial cells adherent, neither in venous nor in arterial endothelial cell cultures. The study showed that not the wall shear stress was responsible for the differing effects visible after 1.5 min, 5 min, and 12 hours exposure to culture media supplemented with RCM but differences in chemotoxicity of the RCM applied.

Behaviour of fibroblasts on water born acrylonitrile-based copolymers containing different cationic and anionic moieties.

The chemical composition of a substrate can influence the adhesion, viability and proliferation of cells seeded on the substrate. The aim of this work was to investigate the influence of different cationic or anionic moieties in acrylonitrile-based copolymers on the interaction with fibroblasts. A series of ten different types of acrylonitrile-based copolymers with a random sequence structure was prepared using a water born synthesis process to exclude potential residues of organic solvents. As charged comonomers cationic methacrylic acid-2-aminoethylester hydrochloride (AEMA), N-3-amino-propyl-methacrylamide hydrochloride (APMA) and anionic 2-methyl-2-propene-1-sulfonic acid sodium salt (NaMAS) were utilized. By application of a specific sintering procedure the copolymer materials were processed into transparent disks for conducting cell tests in direct contact. The copolymers were analyzed with respect to their composition and surface properties. Cytotoxicity tests of the polymer extracts, as well as of the disks were performed with L929 mouse fibroblasts. All copolymers showed no cytotoxic effects. Furthermore, for higher molar ratios of AEMA an increase in cell growth could be observed, which might be a hint that higher charge densities are favorable for the proliferation of L929 cells.

Influence of systemic hypothermia on the myocardial oxygen tension during extracorporeal circulation: comparative study in German Landrace pigs.

During extracorporeal circulation (ECC) controlled hypothermia is a common method of myocardial protection due to a reduction of the myocardial oxygen consumption. Although the beneficial aspects of hypothermia on the myocardial metabolism have been widely demonstrated the effect of hypothermia on the myocardial oxygen tension (PmyO2) is unclear. For this reason the PmyO2 of German Landrace pigs (male, three months of age) during ECC was analysed under mild hypothermia (32°C, n = 6 pigs) and under normothermia (n = 10 pigs, control group) within a time period of 23 min (1400 sec). Flexible invasive Clark type microcatheters were used to measure the PmyO2 in the beating heart. During normothermal ECC a continuous PmyO2 increase from 36.5 ± 15.8 mmHg to 52.6 ± 27.2 mmHg (+44.1%) after 1400 sec was measured (p = 0.02). In contrast, mild hypothermia caused a continuous PmyO2 decrease from initially 46.9 ± 17.5 mmHg to 36.7 ± 20.8 mmHg (-21.8%, p < 0.013) in the test period. Electrocardiography revealed no signs of ischemia or arrhythmia during normo- and hypothermic ECC. It seems obvious that mild hypothermia results in a reduction of the oxygen transfer to the myocardial cells and that this effect outweighs the beneficial effects of hypothermia in the myocardium which are related to reduced oxygen consumption. However, in mild hypothermia oxygen supply to the myocardium remained sufficient for normal myocardial function.

Age- and gender-dependent changes of bovine myocardium architecture.

Growth, gravidity and lactation put high demands on the performance of the myocardium. The aim of this study, which was performed in 40 female and 20 male bovines ranging from 1 to 4.5 years old, was to determine gross and microscopic morphometric data of bovine myocardium to establish a comparative measure of myocardial growth during juvenile development. During the developmental stage of young adulthood, age-related increases in female myocardial characteristics included cardiac mass, left and right ventricular mass and the ratio of cardiac mass to loose connective tissue. Age-related decreases were observed in the number of myocyte nuclei per mm(2) and the thickness of the right ventricular wall. Sex differences in these parameters were found between 2-year-old bulls (N = 20) and 2-year-old heifers (N = 10), with males having heavier hearts, thicker ventricular walls, less myocytes in the left ventricle and less connective tissue in both ventricles. Age and sex had no influence on the ratio of capillaries to myocytes, estimated at 0.98 in the adult bovine. Capillary density does not change during juvenile development, but cross-sectional capillary area does adapt to myocyte cross-sectional area, accounting for this relatively constant ratio.

Do radiographic contrast media (Iodixanol or Iomeprol) induce a perturbation of human arterial and/or venous endothial cells in vitro on extracellular matrix?

After intra-arterial administration of several radiographic contrast media (RCM) a disorder of the downstream microcirculation with regard to blood flow velocity in microvessels and to tissue oxygen partial pressure in the myocardium of the pig heart was described. Iodixanol did not induce such a microcirculatory disorder in the myocardium of the beating heart of pigs. Whether the morphological changes reported in venous endothelial cells after incubation in culture media supplemented with RCM in vitro coincide with a serious endothelial cell dysfunction is not known. In this study we wanted to get information on possible states of dysfunction or perturbation of venous and arterial ECs through the release of prostacyclin, which was shown to follow the perturbation of ECs. Functionally confluent venous endothelial cells on extracellular matrix secreted great amounts of prostacyclin in reaction to the RCMs indicating a clear perturbation of the ECs. This was not the case in arterial EC cultures. The prostacyclin release from arterial ECs exposed to Iodixanol was more than 10-fold higher than that from arterial ECs exposed to Iomeprol. This could be one of the important factors contributing to the undisturbed myocardial microcirculation after injection of Iodixanol despite a slight echinocyte formation.

Viability, morphology and function of primary endothelial cells on poly(n-butyl acrylate) networks having elastic moduli comparable to arteries.

Soft hydrophobic poly(n-butyl acrylate) networks (cPnBA) were developed as entropy elastic substrates for passive mechanical stimulation of cells, where the elastic modulus of the cPnBAs could be systematically adjusted by variation of the cross-link density. The networks were synthesized by thermally-induced radical polymerization from n-butyl acrylate, with poly(propylene glycol) dimethacrylate (PPGDMA) acting as cross-linker, whereby the purity of the cPnBAs was confirmed by(1) H-NMR spectroscopy and gas chromatography. In this work two cPnBA polymer networks with an elastic modulus around 200 kPa and 1 MPa were investigated having an elastic modulus similar to that of arteries. Both cPnBAs exhibited an almost smooth surface with a surface roughness (R q) in the wet state ranging from 17 to 37 nm and a similar zetapotential, indicating an almost identical chemical composition within the topmost surface layer in terms of functional groups. In contrast, wettability of the samples was found to be different with an advancing angle ( advancing) of 123 ± 3.8° for cPnBA0250, while for cPnBA1100 significantly lower values for advancing (111 ± 3.8°) were obtained. First in vitro tests were performed with primary endothelial cells (HUVEC) to study its effects on vascular cell functions. Within the time period of cultivation (72 h), the cells on the cPnBA samples reached subconfluence and showed a viability rate of almost 100%. Although cell density differed after 72 h with more cells on cPnBA0250 than on cPnBA1100, both materials showed no significant effect on the cell morphology, the cellular LDH-release, which was used as marker for the integrity of the cell membrane, and the organisation of the VE-cadherin. However, lower cell density and less actin stress fibre formation on cPnBA1100 might indicate that cell-material interaction was weaker on cPnBA1100 than on cPnBA0250. The secretion of the vasoactive cytokines prostacyclin (PGI2) and thromboxane A2 (TXA2) was low compared to previously reported values. However, the anti-thrombogenic ratio of PGI2/TXA2 - which is balanced under physiological conditions - with much higher PGI2 compared to TXA2 (up to 17.6-fold after 72 h for cPnBA1100) suggests that this material might be effective to preventing thrombosis.