Hyperbaric oxygen enhances collagen III formation in wound of ZDF rat.

Diabetic foot ulcer (DFU) is a serious complication of diabetes and hyperbaric oxygen therapy (HBOT) is also considered in comprehensive treatment. The evidence supporting the use of HBOT in DFU treatment is controversial. The aim of this work was to introduce a DFU model in ZDF rat by creating a wound on the back of an animal and to investigate the effect of HBOT on the defect by macroscopic evaluation, quantitative histological evaluation of collagen (types I and III), evaluation of angiogenesis and determination of interleukin 6 (IL6) levels in the plasma. The study included 10 rats in the control group (CONT) and 10 in the HBOT group, who underwent HBOT in standard clinical regimen. Histological evaluation was performed on the 18th day after induction of defect. The results show that HBOT did not affect the macroscopic size of the defect nor IL6 plasma levels. A volume fraction of type I collagen was slightly increased by HBOT without reaching statistical significance (1.35+/-0.49 and 1.94+/-0.67 %, CONT and HBOT, respectively). In contrast, the collagen type III volume fraction was ~120 % higher in HBOT wounds (1.41+/-0.81 %) than in CONT ones (0.63+/-0.37 %; p=0.046). In addition, the ratio of the volume fraction of both collagens in the wound ((I+III)w) to the volume fraction of both collagens in the adjacent healthy skin ((I+III)h) was ~65 % higher in rats subjected to HBOT (8.9+/-3.07 vs. 5.38+/-1.86 %, HBOT and CONT, respectively; p=0.028). Vessels density (number per 1 mm2) was found to be higher in CONT vs. HBOT (206.5+/-41.8 and 124+/-28.2, respectively, p<0.001). Our study suggests that HBOT promotes collagen III formation and decreases the number of newly formed vessels at the early phases of healing.

Animal models of liver diseases and their application in experimental surgery.

Both acute and chronic liver diseases are frequent and potentially lethal conditions. Development of new therapeutic strategies and drugs depends on understanding of liver injury pathogenesis and progression, which can be studied on suitable animal models. Due to the complexity of liver injury, the understanding of underlying mechanisms of liver diseases and their treatment has been limited by the lack of satisfactory animal models. SO far, a wide variety of animals has been used to mimic human liver disease, however, none of the models include all its clinical aspects seen in humans. Rodents, namely rats and mice, represent the largest group of liver disease models despite their limited resemblance to human. On the other hand, large animal models like pigs, previously used mostly in acute liver failure modeling, are now playing an important role in studying various acute and chronic liver diseases. Although significant progress has been made, the research in hepatology should continue to establish animal models anatomically and physiologically as close to human as possible to allow for translation of the experimental results to human medicine. This review presents various approaches to the study of acute and chronic liver diseases in animal models, with special emphasis on large animal models and their role in experimental surgery.

Evaluation of later morphologic alterations in renal artery wall and renal nerves in response to catheter-based renal denervation in sheep: comparison of the single-point and multiple-point ablation catheters.

This study evaluated the subacute morphologic alterations in renal artery wall and renal nerves in response to catheter-based renal denervation (RDN) in sheep and also compared the efficiency of single-point and multiple-point ablation catheters. Effect of each ablation catheter approved for the clinical use (Symplicity Flex(TM), Medtronic, Inc., or EnligHTN(TM), St. Jude Medical, INC.) was compared to intact contralateral renal artery in 12 sheep by histopathology and immunohistochemistry evaluation after a 10-day period post-RDN procedure. The safety was verified by extensive evaluation of kidney morphology. Vascular wall lesions and nerve injuries were more pronounced in those animals treated with multi-point EnligHTN catheter when compared with animals treated with single-point Symplicity Flex catheter. However, neither RDN procedure led to complete renal nerve ablation. Both systems, tested in the present study, provided only incomplete renal nerve ablation in sheep. Moreover, no appreciable progression of the nerve disintegration in subacute phase post-RDN procedure was observed. This study further supports the notion that the effectiveness remains fully dependent on anatomical inter-individual variability of the sympathetic nerve plexus accompanying the renal artery. Therefore, new systems providing deeper penetrance to targeted perivascular structure would be more efficient.

Distribution of Connective Tissue in the Male and Female Porcine Liver: Histological Mapping and Recommendations for Sampling.

The pig is a large animal model that is often used in experimental medicine. The aim of this study was to assess, in normal pig livers, sexual dimorphism in the normal fraction of hepatic interlobular and intralobular connective tissue (CT) in six hepatic lobes and in three macroscopical regions of interest (ROIs) with different positions relative to the liver vasculature. Using stereological point grids, the fractions of CT were quantified in histological sections stained with aniline blue and nuclear fast red. Samples (415 tissue blocks) were collected from healthy piglets, representing paracaval, paraportal and peripheral ROIs. There was considerable variability in the CT fraction at all sampling levels. In males the mean fraction of interlobular CT was 4.7 ± 2.4% (mean ± SD) and ranged from 0% to 11.4%. In females the mean fraction of the interlobular CT was 3.6 ± 2.2% and ranged from 0% to 12.3%. The mean fraction of intralobular (perisinusoidal summed with pericentral) CT was <0.2% in both sexes. The interlobular CT represented >99.8% of the total hepatic CT and the fractions were highly correlated (Spearman r = 0.998, P <0.05). The smallest CT fraction was observed in the left medial lobe and in the paracaval ROI and the largest CT fraction was detected in the quadrate lobe and in the peripheral ROI. For planning experiments involving the histological quantification of liver fibrosis and requiring comparison between the liver lobes, these data facilitate the power analysis for sample size needed to detect the expected relative increase or decrease in the fraction of CT.

[Sinusoidal obstruction syndrome induced by monocrotaline in a large animal experiment - a pilot study]. / Sinusoidální obstrukcní syndrom indukovaný monokrotalinem v experimentu na velkém zvíreti - pilotní studie.

INTRODUCTION: Sinusoidal obstruction syndrome (SOS) is a disease which is caused by toxic injury to hepatic sinusoids. This syndrome is most frequently caused by myeloablative radiochemotherapy in patients before hematopoietic stem cells transplantation and also by oxaliplatin mainly in patients with colorectal liver metastases. The aim of this study was to establish a large animal model of SOS, which would enable further study of this disease and facilitate translation of experimental outcomes into human medicine. METHODS: A total of 27 domestic pigs (Prestice Black-Pied pig) were involved in this study (12 females). A group with a higher dose of monocrotaline (180 mg/kg) included 5 animals, and the remaining 22 pigs formed another group with a lower dose (36 mg/kg). Monocrotaline was administered via the portal vein and one week after the administration, partial hepatectomy of the left lateral liver lobe was performed. The animals were followed up for 3 weeks after monocrotaline administration. Regular ultrasound examinations were performed as well as examination of biochemical markers of liver and kidney functions and histological examination of liver parenchyma samples. RESULTS: The features of toxic liver injury which we observed in case of all animals were comparable with macroscopic and microscopic appearance of SOS. We recorded AST, ALT, bilirubin and ammonia elevation after monocrotaline administration. Echogenicity on ultrasound images of injured liver parenchyma was higher compared to echogenicity of healthy parenchyma. All the five animals from the first group with a higher monocrotaline dose had died before partial hepatectomy (1st-3rd day after monocrotaline administration). Death before partial hepatectomy occurred in 3 cases (6th and 7th day after monocrotaline administration) in the second group of 22 animals with a lower dose of monocrotaline. Death after partial hepatectomy occurred in 8 cases (7th-17th day after moncrotaline administration) in the same group. 11 animals survived the entire experimental period. The cause of death (in both groups) was metabolic failure in 10 animals and exsanguination in 4 animals, both due to severe hepatopathy. Death of 2 animals was not associated with monocrotaline intoxication (strangulation of small intestine, gastrectasis). CONCLUSIONS: We established a large animal model of SOS induced by monocrotaline administration (36 mg/kg via portal vein). This model can contribute to research of therapeutic modalities for this disease or to evaluation of surgical treatment of patients with SOS.Key words: sinusoidal obstruction syndrome monocrotaline oxaliplatin hepatotoxicity experimental model.

[Experimental promotion of liver regeneration after portal vein branch ligation]. / Moznosti experimentálního ovlivnení regenerace jaterního parenchymu v souvislosti s ligací vetví portální zíly.

INTRODUCTION: Portal vein embolization or ligation (PVE/PVL) is part of most multi-stage liver procedures in the case of low future liver remnant volume (FLRV). PVE initiates compensatory hypertrophy of non-occluded liver parenchyma. This hypertrophy is stimulated by an increased volume of portal blood in the non-occluded veins. PVE results in adequate FLRV growth necessary for resection only in 63-96% patients. The aim of this publication is to summarize the possibilities of influencing liver regeneration after PVE/PVL in an experiment using cytokines (TNF-α, IL-6), a monoclonal antibody against TGF-ß1 (MAB TGF-ß1) and mesenchymal stem cells (MSC). METHODS: The experimental model of PVE/PVL was chosen as best compatible for potential use in human medicine. 9 (control group), 9 (TNF-α group), 8 (IL-6 group), 6 (MSC group) and 7 piglets (MAB TGF-ß1 group) were enrolled in individual studies. We performed laparotomy with PVL of the right-sided liver lobes under general anaesthesia. The following amounts of substances were applied in the non-occluded portal vein branches immediately after PVL: physiological solution (control group), recombinant porcine TNF-α (5 µg/kg), recombinant porcine IL-6 (0.5 µg/kg) and MSC (8.75, 14.0, 17.0, 17.5, 43.0 and 61.0 x 106 MSC). MAB TGF-ß1 was applied 24 hours after PVL (40 µg/kg). Biochemical parameters were analysed repeatedly and FLRV ultrasound assessments were performed in the postoperative period. The experiments were ended on postoperative day 14 by sacryfiing the animals under general anaesthesia. Liver samples of hypertrophic and atrophic liver parenchyma were analysed. RESULTS: Repeated ultrasound assessments of the effects of MSC, TNF-α, IL-6 and MAB TGF-ß1 compared with the physiological solution in the control group demonstrated statistically significant acceleration of FLRV growth in the experimental groups. For MSC, maximum growth was observed between postoperative days 3 and 7, on day 7 for TNF-α, between days 3 and 7 for MAB TGF-ß1 and on day 7 for IL-6. Serum levels of AST and ALT increased after PVL and MSC whereas other biochemical parameters showed no statistically significant differences. We identified individual MSC using immunohistochemistry in the hypertrophic tissue of the MSC group. A statistically significant difference was observed in the number of binucleated hepatocytes, with their increased concentration in the IL-6 group. CONCLUSION: Application of IL-6, TNF-α, MAB TGF-ß1 and MSC seems to provide suitable stimulation for achieving faster FLRV growth. Nevertheless, many controversial questions still remain to be answered with respect to the mechanism of their respective effects.Key words: liver regeneration portal vein embolization large animal experiment mesenchymal stem cells cytokines.

The combination of nanofibrous and microfibrous materials for enhancement of cell infiltration and in vivo bone tissue formation.

Fibrous scaffolds are desired in tissue engineering applications for their ability to mimic extracellular matrix. In this study we compared fibrous scaffolds prepared from polycaprolactone using three different fabrication methods, electrospinning (ES), electro-blowing and melt-blown combined with ES. Scaffolds differed in morphology, fiber diameters and pore sizes. Mesenchymal stem cell adhesion, proliferation and osteogenic differentiation on scaffolds was evaluated. The most promising scaffold was shown to be melt-blown in combination with ES which combined properties of both technologies. Microfibers enabled good cell infiltration and nanofibers enhanced cell adhesion. This scaffold was used for further testing in critical sized defects in rabbits. New bone tissue formation occurred from the side of the treated defects, compared to a control group where only fat tissue was present. Polycaprolactone fibrous scaffold prepared using a combination of melt-blown and ES technology seems to be promising for bone regeneration. The practical application of results is connected with enormous production capacity and low cost of materials produced by melt-blown technology, compared to other bone scaffold fabrication methods.

Early morphologic alterations in renal artery wall and renal nerves in response to catheter-based renal denervation procedure in sheep: difference between single-point and multiple-point ablation catheters.

Renal sympathetic hyperactivity is critically involved in hypertension pathophysiology; renal denervation (RDN) presents a novel strategy for treatment of resistant hypertension cases. This study assessed effects of two RDN systems to detect acute intravascular, vascular and peri-vascular changes in the renal artery, and renal nerve alterations, in the sheep. The procedures using a single-point or multi-point ablation catheters, Symplicity Flex(TM), Medtronic versus EnligHTN(TM), St. Jude Medical were compared; the intact contralateral kidneys served as controls. Histopathological and immunohistochemical assessments were performed 48 h after RDN procedures; the kidney and suprarenal gland morphology was also evaluated. Special staining methods were applied for histologic analysis, to adequately score the injury of renal artery and adjacent renal nerves. These were more pronounced in the animals treated with the multi-point compared with the single-point catheter. However, neither RDN procedure led to complete renal nerve ablation. Forty-eight hours after the procedure no significant changes in plasma and renal tissue catecholamines were detected. The morphologic changes elicited by application of both RDN systems appeared to be dependent on individual anatomical variability of renal nerves in the sheep. Similar variability in humans may limit the therapeutic effectiveness of RDN procedures used in patients with resistant hypertension.

How many Layers has the Adventitia? - Structure of the Arterial Tunica Externa Revisited.

Tunica adventitia or tunica externa is the outer layer of the blood vessel wall. It consists of connective tissue with vasa and nervi vasorum and plays a key role in vascular health. The aim of our study was to compare the wall layers beyond tunica media in arteries of different type and location. The following arteries of pig, dog and cat were processed histologically and analysed by light microscopy: aorta ascendens, arcus aortae, aorta thoracica, aorta abdominalis, arteria (a.) femoralis, a. tibialis cranialis, a. carotis communis, a. lingualis, a. basilaris, a. cerebralis media, a. testicularis and aa. jejunales. We found two layers of connective tissue outside the media: (1) a compact layer with many elastic fibres in muscular and few in elastic arteries and (2) an outer layer of loose connective tissue. The compact layer was missing in aorta ascendens, arcus aortae and intracranial vessels. Adventitial stripping removed only the loose connective tissue layer. In spite of the still present compact layer, stripped arteries were very flimsy. We suggest using the term 'tunica externa' for the compact connective tissue layer and 'tunica adventitia' for the outermost loose connective tissue layer as in other organs. The presence of the tunica externa differs between species, arteries and arterial side, as well as the removability of tunica adventitia and tunica externa by anatomical dissection.

Porcine liver vascular bed in Biodur E20 corrosion casts.

BACKGROUND: Pigs are frequently used as animal models in experimental medicine. To identify processes of vascular development or regression, vascular elements must be recognised and quantified in a three-dimensional (3D) arrangement. Vascular corrosion casts enable the creation of 3D replicas of vascular trees. The aim of our study was to identify suitable casting media and optimise the protocol for porcine liver vascular corrosion casting. MATERIALS AND METHODS: Mercox II® (Ladd Research, Williston, Vermont, USA) and Biodur E20® Plus (Biodur Products, Heidelberg, Germany) were tested in 4 porcine livers. The resins (volume approximately 700 mL) were injected via the portal vein. Corrosion casts were examined by macro-computed tomography, micro-computed tomography and scanning electron microscopy. RESULTS: For hepatectomies, the operating protocol was optimised to avoid gas or blood clot embolisation. We present a protocol for porcine liver vascular bed casting based on corrosion specimens prepared using Biodur E20® epoxy resin. CONCLUSIONS: Only Biodur E20®Plus appeared to be suitable for high-volume vascular corrosion casting due to its optimal permeability, sufficient processing time and minimum fragility. Biodur E20® Plus is slightly elastic, radio-opaque and alcohol-resistant. These properties make this acrylic resin suitable for not only vascular research but also teaching purposes.

Is the amount of coronary perivascular fat related to atherosclerosis?

Interesting and stimulating data about the effect of the perivascular adipose tissue size on atherogenesis are based mainly on CT findings. We studied this topic by directly analyzing perivascular adipose tissue in explanted hearts from patients undergoing transplantation. Ninety-six consecutive patients were included, including 58 with atherosclerotic coronary heart disease (CHD) and 38 with dilation cardiomyopathy (DCMP). The area of perivascular fat, area of the coronary artery wall, and ratio of CD68-positive macrophages within the perivascular fat and within the vascular wall were quantified by immunohistochemistry. There was no significant difference in the perivascular adipose tissue size between the two groups. Nevertheless, there was a significantly higher number of macrophages in the coronary arterial wall of CHD patients. In addition, we found a close relationship between the ratio of macrophages in the arterial wall and adjacent perivascular adipose tissue in the CHD group, but not in the DCMP group. According to our data interaction between macrophages in the arterial wall and macrophages in surrounding adipose tissue could be more important mechanism of atherogenesis than the size of this tissue itself.

Collagen/hydroxyapatite scaffold enriched with polycaprolactone nanofibers, thrombocyte-rich solution and mesenchymal stem cells promotes regeneration in large bone defect in vivo.

A three-dimensional scaffold of type I collagen and hydroxyapatite enriched with polycaprolactone nanofibers (Coll/HA/PCL), autologous mesenchymal stem cells (MSCs) in osteogenic media, and thrombocyte-rich solution (TRS) was an optimal implant for bone regeneration in vivo in white rabbits. Nanofibers optimized the viscoelastic properties of the Coll/HA scaffold for bone regeneration. MSCs and TRS in the composite scaffold improved bone regeneration. Three types of Coll/HA/PCL scaffold were prepared: an MSC-enriched scaffold, a TRS-enriched scaffold, and a scaffold enriched with both MSCs and TRS. These scaffolds were implanted into femoral condyle defects 6 mm in diameter and 10-mm deep. Untreated defects were used as a control. Macroscopic and histological analyses of the regenerated tissue from all groups were performed 12 weeks after implantation. The highest volume and most uniform distribution of newly formed bone occurred in defects treated with scaffolds enriched with both MSCs and TRS compared with that in defects treated with scaffolds enriched by either component alone. The modulus of elasticity in compressive testing was significantly higher in the Coll/HA/PCL scaffold than those without nanofibers. The composite Coll scaffold functionalized with PCL nanofibers and enriched with MSCs and TRS appears to be a novel treatment for bone defects.

Focal histopathological progression of porcine experimental abdominal aortic aneurysm is mitigated by atorvastatin.

AIM: Observational studies in human patients and animal experiments suggested that statins have a potential in slowing the growth of small abdominal aortic aneurysms (AAA). Our aim was to quantify histological postoperative changes of AAA in porcine experimental model of AAA with and without administration of atorvastatin. METHODS: The AAA was induced by intraaortic infusion of porcine pancreatic elastase and subrenal application of plastic cuff. The AAA statin group (N.=14) received atorvastatin 1 mg/kg daily for 28 days, the other AAA group (N.=13) did not. The aortic diameter was measured by ultrasonography. Aortic samples were described using eleven quantitative histological parameters and compared with healthy aortae. RESULTS: There was no difference in aortic diameter between the AAA with statin when compared to AAA without statin. Administration of atorvastatin led to a better postoperative histological condition of the aortic elastin network, preservation of contractile phenotype of vascular smooth muscle, a higher density of vasa vasorum, it prevented thickening of intima and media. The increase in wall thickness in AAA without atorvastatin has not been accompanied by a proportional increase in number of vasa vasorum. CONCLUSION: The effects of atorvastatin seem to prevent the histopathological progression of AAA.

A preliminary study into the correlation of stiffness of the laminar junction of the equine hoof with the length density of its secondary lamellae.

REASON FOR PERFORMING STUDY: The relationship between mechanical behaviour and microscopic structure of the laminar junction of equine hooves under testing conditions requires elucidation. OBJECTIVES: To determine mechanical parameters and 2D length density of profiles of secondary lamellae of the laminar junction in the dermal region and to assess possible correlations. METHODS: Specimens (25 samples in total) of the laminar junction were taken from front, quarter and heel parts from 3 equine hooves and exposed to a uniaxial tensile test until rupture to obtain Young's moduli of elasticity, ultimate stress and strain. Neighbouring specimens to those used for the biomechanical experiment were processed histologically to assess the length density of laminar junction basement membrane using stereological grids. RESULTS: The estimated median (interquartile range) length density of the laminar junction basement membrane was 0.024 (0.020-0.027)/µm. Young's modulus of elasticity was 0.15 (0.11-0.35) MPa in the small deformation region, and 7.58 (6.14-8.68) MPa in the linear region was. The ultimate stress was 1.67 (1.41-2.67) MPa, and the ultimate strain was 0.50 (0.38-0.70). The Young's modulus of elasticity in the region of small deformations has a moderate correlation with the length density of the laminar junction basement membrane. CONCLUSIONS: As with most soft biological tissues, the laminar junction has a nonlinear mechanical behaviour. Within the range of small deformations, which correspond to physiological loading of the laminar junction, a higher length density of the laminar junction basement membrane is correlated with a higher resistance of the laminar junction against high stresses transmitted from the distal phalanx to the hoof wall. POTENTIAL RELEVANCE: The condition of the laminar junction apparatus may be easily quantified as the length density of profiles of secondary dermal lamellae. This quantification provides a simple tool that could be used for comparing the proneness of the various parts of the laminar junction to initial stages of laminitis.

[Pharmacological influencing of abdominal aortic aneurysm model - animal experimental study, primary results]. / Farmakologické ovlivnení modelového aneuryzmatu brisní aorty - experiment na zvíreti, prvotní výsledky.

INTRODUCTION: The aim of our work was to influence growth and histological changes in the wall of an experimentally induced aneurysm of the abdominal aorta in a large laboratory animal (domestic pig) by administering atorvastatin (3-hydroxy-3-methylglutaryl-coenzyme A reductase inhibitor). MATERIAL AND METHODS: Within the scope of the above mentioned experimental work, we compared the growth rate of the aneurysm between the group influenced by statin administration (n=14) and the group without any pharmacological treatment (n=13). We also compared histological changes in the structure of the aortic wall in both groups with aneurysm and the wall of the aorta without aneurysm (n=6). RESULTS: During the 4-week follow-up, we did not prove a statistically significant difference in the growth rate of aneurysms between the above mentioned groups. The histological structure of the aneurysm walls, however, differed between the two groups. The structure of the wall in the group of animals influenced by statin administration resembled the structure of the aortic wall without aneurysm. CONCLUSION: The results presented demonstrate that statins do influence the composition of the aortic wall. In our opinion, the administration of statins could lead to changes resulting in a more stable aneurysmatic wall. We believe that patients with smaller aneurysms who are not indicated for surgery or endovascular treatment could be treated with statins. Stabilization of the aneurysmal wall could slow down the growth of the aneurysm and prevent its rupture.

The contribution of vascular smooth muscle, elastin and collagen on the passive mechanics of porcine carotid arteries.

The main components responsible for the mechanical behavior of the arterial wall are collagen, elastin, and smooth muscle cells (SMCs) in the medial layer. We determined the structural and mechanical changes in porcine carotid arteries after administration of Triton® X-100, elastase, and collagenase using the inflation-deflation test. The arteries were intraluminarly pressurized from 0 to 200 mmHg, and the outer diameter of the artery was measured. The pressure-strain elastic modulus was determined based on the pressure/diameter ratio. The intima-media thickness, wall thickness, thickness of the tunica adventitia layer, and the area fractions of SMCs, elastin, and collagen within the arterial wall (A(A)(SMC/elastin/collagen, wall)) were measured using stereological methods. The relative changes in the relevant components of the treated samples were as follows: the decrease in A(A)(SMC, wall) after administration of Triton® X-100 was 11% ± 7%, the decrease in A(A)(elastin, wall) after administration of elastase was 40% ± 22%, and the decrease in A(A)(collagen, wall) after the application of collagenase was 51% ± 22%. The Triton® X-100 treatment led to a decrease in the SMC content that was associated with enlargement of the arterial wall (outer diameter) for pressures up to 120 mmHg, and with mechanical stiffening of the arterial wall at higher pressures. Elastase led to a decrease in the elastin content that was associated with enlargement of the arterial wall, but not with stiffening or softening. Collagenase led to a decrease in collagen content that was associated with a change in the stiffness of the arterial wall, although the exact contribution of mechanical loading and the duration of treatment (enlargement) could not be quantified.

Angiogenic markers in canine lymphoma tissues do not predict survival times in chemotherapy treated dogs.

Angiogenesis, which is essential for malignancies to progress, depends on various signalling proteins including vascular endothelial growth factor (VEGF) and vascular endothelial growth factor receptors 1 and 2 (VEGFR-1 and VEGFR-2). Microvessel density (MVD) is frequently used to evaluate angiogenesis. This study assessed the relationship between expression of VEGF, VEGFR-1 and VEGFR-2, MVD and the survival time in dogs with lymphoma. VEGF, VEGFR-1 and VEGFR-2 expression was evaluated immunohistochemically and microvessel profiles were counted in 34 lymphoma samples. Seventy-nine percent of the samples showed high VEGF expression and 62% were highly positive for VEGFR-1; VEGFR-2 immunoreactivity was mostly negative. Dogs treated with chemotherapy had a median survival time of 266days, but no significant relationships were found between overall survival time, MVD and expression of VEGF, VEGFR-1 or VEGFR-2. In this study, VEGF its receptors and the MVD were no prognostic factors in dogs with lymphoma.

Optimized conditions for mesenchymal stem cells to differentiate into osteoblasts on a collagen/hydroxyapatite matrix.

Collagen/hydroxyapatite (HA) composite scaffolds are known to be suitable scaffolds for seeding with mesenchymal stem cells (MSCs) differentiated into osteoblasts and for the in vitro production of artificial bones. However, the optimal collagen/HA ratio remains unclear. Our study confirmed that a higher collagen content increased scaffold stiffness but that a greater stiffness was not sufficient for bone tissue formation, a complex process evidently also dependent on scaffold porosity. We found that the scaffold pore diameter was dependent on the concentration of collagen and HA and that it could play a key role in cell seeding. In conclusion, the optimal scaffold for new bone formation and cell proliferation was found to be a composite scaffold formed from 50 wt % HA in 0.5 wt % collagen I solution.

[Effect of biological glues on vascular wall in an experimental model of aortic dissection]. / Vliv biologických lepidel na cévní stenu v modelu disekce aorty v experimentu.

INTRODUCTION: The use of biological glues and their application between the two dissection layers and into the anastomosis region is a common integral part of surgical management of thoracic aortic dissections. AIM: The aim of the experimental study was to assess and evaluate histopathological changes of vascular wall following deposition of the following three types of glue--GRF, Tissucol, Bioglue, based on qualitative and quantitative parameters. The secondary aim of the study was to assess dynamics of these changes depending on the glue effect duration and to formulate expected behaviour of the vascular wall during the time beyond the experimental period. METHODOLOGY: The dissection model was performed with pigs of the same gender and age, assigned to four groups. Different glues were used to close artificial infrarenal aortic dissections in Group 1-3, while direct suturing and no glue was used to close false lumini in Group 4. Samples of the dissected aorta were then collected at Month 1, 6 and 12 and then histologically examined. RESULTS: Upon assessment of the whole group of qualitative and quantitative parameters, the most significant changes in the smooth muscle histological picture were observed with the use GRF glue. The smooth muscle changes following the Bioglue application and, in particular, Tissucol glue application, are similar to changes observed in Group 4, where no glue was used. CONCLUSION: Based on the results, the authors present a hypothesis that, in a long-time horizont, vascular wall destructions, eventually redissections, are likely to occur more frequently in patients, in whom GRF glue is used.

[Plasmatic levels of proinflammatory cytokines in abdominal aortic aneurysms]. / Plazmatické hladiny prozánetlivých cytokinu u aneuryzmatu abdominální aorty.

INTRODUCTION: Inflammation within the abdominal aortic wall is generally considered a very significant ethiopathogenic factor in the development of abdominal aortic aneurysms. Proinflammatory cytokines are important mediators of inflammation within the abdominal aortic wall. AIM: The aim of the study was to research, whether plasmatic levels of certain proinflammatory cytokines, which can commonly be evaluated (TNFalpha, IL-1, -2, -6 a -8), play a significant role in the development of AAA. METHOD: The prospective non-randomized study included 345 patients with AAAs. The patients were assigned to 5 subgroups based on their symptoms and AAA diameters. The first subgroup included patients with symptomatic AAAs, including AAA ruptures (N = 69), the second subgroup included subjects with asymptomatic AAAs (N = 276) with AAA diameters up to 5 cm (N = 72), the third subgroup included 5 cm (N = 72), the fourth included 5-8 cm (N = 192) and the fifth subgroup included subjects with AAA diameters of more than 8 cm (N = 81). The mean age of patients was 74.1 +/- 7.8 years (56-84 y.o.a.). The male to female ratio was 5:1. The control group included 30 healthy volunteer subjects of similar age and male to female rates, who had no clinical signs of arterial disorders. Plasmatic levels of cytokines were evaluated from venous blood samples using ELISA (Bender, Austria) testing. Statistical assessment of the results was performed using ANOVA and Wilcoxon tests with Spearman's correlation. P values < 0.05 were considered significant. RESULTS: Plasmatic concentrations of proinflammatory cytokines were found to be statistically significantly higher in patients with AAAs compared to those in healthy volunteers. Plasmatic IL8 levels were significantly decreasing proportionally to decreasing AAA diameters (p < 0.05). TNFalpha levels were found to be significantly low in symptomatic patients with AAA ruptures (p < 0.05). CONCLUSION: The study confirmed the significance of proinflammatory cytokines levels monitoring in AAA patients. The authors showed that, for instance IL8 activity and to a certain extent TNFalpha activity, is the highest in small and developing AAAs. These findings would be significant for customized medication therapy aimed at blocking the effects of these factors on the inflammatory process within the AAA wall.