Clinical Hemorheology and Microcirculation

The efficacy of three weekly interventions with hypervolumetric hemodilution of a new preparation of hydroxyethyl starch (HES 100/0.5, 10%, C2/C6 substitution ratio of 6.5) on pain-free walking distance of patients with peripheral arterial occlusive disease (PAOD) stage IIb on the Fontaine classification was investigated. In addition quantitative data on the pharmacokinetic properties of this HES preparation, and it's impact on hemorheology, hemostasis and homeostasis were shown. Ten patients were included according to a predefined protocol, and treated openly with 500 ml HES 100/0.5 10% on nine occasions over 18 days. Pain-free walking distance, the main outcome measure, showed a mean increase of 82 m (+60%). Hematocrit decreased 4 percentage points on average (5.5 percentage points one hour after interventions). Plasma viscosity dropped 5% on average with significant changes immediately after interventions only in patients whose baseline values had been equal to or above the 2 s reference area. Erythrocyte aggregation decreased by 16% in the course of treatment (8% immediately after interventions), systolic blood pressure by 13%, and total protein by 7%. Complement showed a trend towards lower values (-20%), and creatinine, pH and urine viscosity remained unchanged. Apart from complement changes, all reductions mirrored the dilution effects. As to pharmacokinetics, serum mean molecular weight distribution was very similar to that of the infusion. A minor adverse drug reaction (light, spontaneously disappearing pruritus) was observed in one case.

Geometric distortions and low spatial resolution are current limitations in functional magnetic resonance imaging (fMRI). The aim of this study was to evaluate if application of parallel imaging or significant reduction of voxel size in combination with a new 32-channel head array coil can reduce those drawbacks at 1.5 T for a simple hand motor task. Therefore, maximum t-values (tmax) in different regions of activation, time-dependent signal-to-noise ratios (SNR(t)) as well as distortions within the precentral gyrus were evaluated. Comparing fMRI with and without parallel imaging in 17 healthy subjects revealed significantly reduced geometric distortions in anterior-posterior direction. Using parallel imaging, tmax only showed a mild reduction (7-11%) although SNR(t) was significantly diminished (25%). In 7 healthy subjects high-resolution (2 x 2 x 2 mm3) fMRI was compared with standard fMRI (3 x 3 x 3 mm3) in a 32-channel coil and with high-resolution fMRI in a 12-channel coil. The new coil yielded a clear improvement for tmax (21-32%) and SNR(t) (51%) in comparison with the 12-channel coil. Geometric distortions were smaller due to the smaller voxel size. Therefore, the reduction in tmax (8-16%) and SNR(t) (52%) in the high-resolution experiment seems to be tolerable with this coil. In conclusion, parallel imaging is an alternative to reduce geometric distortions in fMRI at 1.5 T. Using a 32-channel coil, reduction of the voxel size might be the preferable way to improve spatial accuracy.

In cancer patients impaired blood rheology in the presence of coagulation activation may reduce blood flow in the vascular microcirculation that favors thrombosis but may also support tumor progression and metastasis. In 451 patients with gynecological cancer and 177 patients with corresponding benign tumor disease preoperatively, during adjuvant treatment, when venous thrombosis (VT) or cancer progression was diagnosed hematocrit (micro centrifuge), hemoglobin, leukocytes, platelets (Coulter Counter); red blood cell (RBC) aggregation (aggr.) during stasis and low shear conditions (MA 1, Myrenne), plasma viscosity (viscosimeter KSPV 1 Fresenius), and fibrinogen (Multifibren Behring Dade) were investigated. One hundred and twelve healthy women served as controls. Preoperatively, mean plasma viscosity (pv) was significantly higher in cancer patients as compared to patients with the corresponding benign tumor disease (breast cancer: n = 261; pv = 1.32 vs. 1.27 mPa s; p = 0.023; ovarian cancer: n = 68; pv = 1.39 vs. 1.31 mPa s; p < 0.001; endometrial cancer: n = 70; pv = 1.37 vs. 1.25 mPa s; p < 0.001; cervical cancer: n = 52; pv = 1.33 vs. 1.26 mPa s; p = 0.004). RBC aggr. was significantly lower in controls compared to the preoperative values in cancer patients but mean (median) values (RBC aggr. stasis < 21) were within the normal range in all. Preoperatively, plasma viscosity was a significant risk factor for the overall survival in ovarian cancer patients (p = 0.02) and for subsequent thrombosis in ovarian (p = 0.02) and cervical cancer patients (p = 0.007). In the multivariate analysis plasma viscosity was an independent prognostic marker for the overall survival of breast cancer patients (r = 99.45; 95% CI: 7.32-980.2; p < 0.0001). An optimized preoperative cut-off value above 1.40 mPa s (Log-Rank-test) was significantly associated with poor outcome in the Kaplan-Mayer survival estimates, even in node-negative breast cancer. In gynecologic cancer patients the combination of an increase in RBC aggregation and plasma viscosity impairs blood-flow-properties and may induce hypoxia in the microcirculation that favors thrombosis, settlement of tumor-cells and thus metastasis. Improvement of blood fluidity and thus oxygen transfer in the tumor-vascular-microcirculation may increase susceptibility of systemic anti-cancer therapy.

Alterations of blood and plasma viscosity can promote atherosclerosis. The relationship between viscosity and aging is still controversial. The present study evaluated the influence of aging on blood and plasma viscosity in a group of subjects followed for 11.6 years. Forty-five subjects have been evaluated twice 11.6 years apart for hemorheological parameters and coronary heart disease (CHD) risk factors. Plasma viscosity and blood viscosity have been measured with a cone-plate viscometer. Tk has been calculated as index of red blood cell rigidity. CHD risk factors, i.e. obesity, hypertension, hyperlipidemia and diabetes, have been evaluated by routine methods. Hematocrit and plasma viscosity did not change during the study, whereas blood viscosity (shear rate 225/sec: 4.46 ± 0.49 vs. 4.81 ± 0.54 cP, p < 0.0001; shear rate 45/sec: 6.19 ± 0.67 vs. 6.65 ± 0.79 cP, p < 0.0001) and Tk (0.80 ± 0.05 vs. 0.83 ± 0.06, p < 0.005) significantly increased. The percent variation in blood viscosity was not associated with the percent variation in any of the CHD risk factors. Furthermore, the increase in blood viscosity was similar in males and females and in subjects with CHD risk profile worsening or not. The present findings demonstrate that blood viscosity increases with age. This increase seems independent of classical CHD risk factors and is disjoined from haematocrit and plasma viscosity, suggesting a possible direct effect of aging on red blood cells.

To explore the role of contrast-enhanced ultrasound (CEUS) in the early evaluation of microvacularization in patients with aggressive B-cell lymphoma treated by R-CHOP. Fifty-two patients with aggressive B-cell lymphoma underwent combined rituximab-CHOP treatment (CHOP: cyclophosphamide, hydroxydaunomycin, vincristine, prednisonetreatment). Before the treatment and after the first two cycles of R-CHOP, CEUS was performed to assess the microvascularization of tumors. In addition, PET/CT examination was also included in this study before and after the treatment. Ideal cut-off value of CEUS parameters was calculated using receiver-operating characteristic curve (ROC) analysis to predict the treatment outcome. The response to treatment, progression-free survival (PFS) and overall survival (OS) were then compared according to PET/CT and CEUS results. The correlation between CEUS parameters and PET/CT results was investigated based on these analyses. All patients were non-Hodgkin lymphomas (NHL) with CD20 positive. The variations of CEUS parameters (peak intensity and mean intensity) at the mid-term of R-CHOP treatment significantly associated with results of treatment response (P < 0.001), and were also positive correlation with PET/CT results (P = 0.001). The CEUS positive or negative results were determined by cut-off points of peak intensity and mean intensity (4.70 and 4.15 dB). The median time of clinical follow-up was 35.4 months. Kaplan-Meier survival analysis demonstrated that PET/CT and CEUS were both good predictors for PFS and OS rate in these patients. As a microvascular imaging modality, CEUS could be a feasible tool to predict the survival rate of patients with aggressive B-cell lymphoma after R-CHOP treatment.

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.

Unlabelled: PURPOSE, MATERIAL AND METHODS: The aim of this study was to investigate the influence of direct intraarterial application of the contrast agents Iodixanol-270 and Iopentol-150 on the capillary perfusion. This was accomplished through continuous recording of the capillary perfusion in the nailfold capillaries of the right hand before and after a bolus injection of 20 ml of contrast agent into the right axillary artery. Results: After injecting 20 ml of Iodixanol-270, which has a high viscosity compared to the plasma viscosity, a statistically significant decrease in the erythrocyte velocity of 60.8% from 0.439+/-0.273 mm/s to 0.172+/-0.090 mm/s was observed already 10 s after the injection (p = 0.0001). The decreased velocity was maintained until the end of the observation period of 6 min. In contrast to this finding, no change in the erythrocyte velocity was observed after injection of 20 ml of the low-viscous Iopentol-150 (p = 0.1508). Conclusions: The erythrocyte velocity in cutaneous capillaries therefore strongly depends on the viscosity of the contrast agent.

Monocytes are broadly discussed in the literature as cells, which can get properties of endothelial progenitor cells after angiogenic stimulation. Angiogenically stimulated monocytes can be used to promote implant vascularisation. A necessity therefore is that these cells can be stored and used after storage without a loose of their characteristic phenotype. In this study we tested, if freshly thawed cryopreserved human monocytes are positive for the mo/macrophage markers CD14 and CD68 and the endothelial marker CD31 after thawing and following angiogenic stimulation in a VEGF-A(165) enriched (10 ng/ml) angiogenic medium. Thereby the monocytes were tested before and after differentiation towards macrophages. The results revealed that freshly thawed human CD14 positive monocytes are positive for CD14, CD68 and CD31 after angiogenic stimulation. This CD specification was much more intense in the differentiated cells. The differentiation step also resulted in an increased cell count. Both results can be attributed to the method of differentiation, were cell culture bags were used instead of common cell culture dishes. Additionally the differentiation medium (X-VIVO 10+10% FCS) was specifically adapted to the requirements of monocytes/macrophages. The study showed that human CD14 positive monocytes can be thawed after cryopreservation without loss of their monocytes/macrophage phenotype and without loss of their ability to get angiogenically stimulated. To enhance the efficiency of both steps (thawing, angiogenic stimulation) it can be useful to differentiate the thawed cells in cell culture bags by the use of X-VIVO 10 (+10% FCS) before angiogenic stimulation.

RheothRx injection is an intravenous formulation of Poloxamer 188, a non-ionic block copolymer surfactant which is actually used in clinical studies, e.g., during thrombolysis in acute myocardial infarction. We have analysed the influence of RheothRx on plasma and whole blood viscosity of healthy individuals in vitro. For that purpose plasma was incubated with 0, 0.75, 3.75 and 18.75 mg/ml RheothRx and whole blood with a constant hematocrit of 41.4% with 0, 0.4, 2 and 10 mg/ml RheothRx at 37 degrees C. Viscosity was measured with a Couette viscometer at shear rates of 94, 11, 0.9 and 0.1 s-1 as well as with a newly developed oscillating viscometer. In contrast to other studies, we found no favourable effect of RheothRx on plasma and whole blood viscosity. At the highest RheothRx concentration an increase in high and low shear viscosity was observed. Erythrocyte morphology remained unchanged. These data, although on normal blood in vitro, suggest that the positive effects of RheothRx in vivo may not be caused by improved flow properties of blood, but could rely on other mechanisms.

Postsurgical evaluation of bone microcirculation in osseous and osseocutaneus free flaps by contrast enhanced ultrasound (CEUS) with time intensity curve (TIC) analysis and [18F]-positron emission tomography/computed tomography (Fluoride-PET/CT). 10 patients with osseous or osseocutaneus microvascular flaps were evaluated with CEUS. Ultrasound was carried out by an experienced examiner with a high resolution linear probe (6-9 MHz, LOGIQ E9, GE) after intravenous bolus injection of 2.4 ml SonoVue® (BRACCO, Germany). Time intensity curves (TIC) in selected regions of interest (ROI) were analyzed and compared with the evaluation of [18F]-positron emission tomography/computed tomography and the clinical course for at least 2 month. 9/10 patients showed evidence for soft tissue and osseous microcirculation of the transplants in CEUS correlating with the clinical course. The soft tissue parts of the transplant showed a mean value of 84% (range 51-98%) and the bone parts a mean value of 39% (range 11-75%) for the Area under the curve (AuC) compared with the AuC for the anastomotic vessel region. Mean values for time to peak (TTP) were 27,1 sec (range 8.7-52.0 sec) for the anastomotic vessels, 29.3 sec (range 7.9-62.0 sec) for the soft tissue of and 32.0 sec (range 7.4-69.0 sec) for the transplant bone. In 1/10 patients flap failure occurred, after denudation the bone was left as an avascular transplant. AuC showed a mean value of 0.5 % for the bone region compared with the vessel region. Fluoride-PET/CT assessed bone vitality in 7 patients as "good" in 1 patient as "uncertain" and in 1 patient as "poor". CEUS assessment was corresponding with Fluoride-PET/CT in 4 patients, clinical assessment in 6 patients. Fluoride-PET/CT is a valuable tool to make an indirect statement about the perfusion of the transplanted bone and was used as control in this study. CEUS is a new and promising method for the evaluation of microcirculation of buried free microvascular bone grafts and the osseous part of osseocutaneous flaps and may be used for a steady monitoring in the first postoperative days.

Both, skeletal development and fracture healing depend on an orchestrated sequence of cellular growth and differentiation processes. Regional changes in tissue oxygen tension were proposed as key regulators of osteoblast proliferation and phenotype. Hypoxia results in the stabilization of hypoxia-inducible factor-1α (HIF-1α), thus influencing expression of a multitude of genes required for cellular adaptation. In the present study we dissected the effects of HIF-1α on cellular proliferation and gene expression of primary human osteoblasts. Primary human osteoblasts were studied by transfecting siRNA and plasmids coding for human HIF-1α. Gene expression was analyzed by western blot and quantitative PCR. Functional assays were performed to study HIF-1α function, i.e. proliferation and cell cycle analysis. As previously reported exposure to hypoxia led to a stabilization of HIF-1α on protein level and resulted in reduced rates of proliferation and osteocalcin expression. Furthermore, the expression of the proproliferative gene survivin was significantly reduced (p < 0.01). Knock down of HIF-1α attenuated hypoxic downregulation of proliferation (p < 0.05), and osteocalcin (p < 0.05) as well as survivin (p < 0.05) expression significantly. Importantly, the isolated overexpression of HIF-1α impaired proliferative activity and led to significantly reduced rates of expression of osteocalcin (p < 0.05) and survivin (p < 0.01). The present study shows that HIF-1α might reduce proliferation and survivin expression in primary human osteoblasts independently from cellular hypoxia. Furthermore, HIF-1α promoted the loss of the characteristic osteoblastic marker, osteocalcin in vitro. These findings underline the important role of HIF-1α in bone physiology and pathophysiology. Modulating HIF-1α function in hypoxic environments could be of value for future therapeutic approaches.

many studies have investigated between venous stasis, functions of the vascular and perivascular anastomotic structure, venous endothelium and circulating leukocytes. patients with varicose veins (n = 15) and a healthy control group (n = 15). the authors investigated some soluble mediators of monocytes-macrophages, which induce inflammation. They determined interleukin 1beta (IL-1beta) and interleukin 6 (IL-6) and tumor necrosis factor alpha (TNFalpha) levels at rest and after induced venous occlusion (using an inflated cuff to 60 mmHg for 25 minutes). their results revealed elevated baseline production in the former and that induced venous occlusion further augmented the levels of all cytokines in the study series, especially in patients with varicose veins. The authors believe that the study shows functional activation of monocyte-macrophages related to venous stasis as a consequence of venous hypertension. Cell response damages the endothelial structure and may represent an important element in the pathophysiology of chronic venous insufficiency.

It has been suggested that 2,3-diaryl-substituted indole-based cyclooxygenase-2 (COX-2) inhibitors (2,3-diaryl-indole coxibs) do not only appear as potent anti-inflammatory agents but also show the ability to scavenge reactive oxygen species (ROS). This led to the hypothesis that 2,3-diaryl-indole coxibs also may act as potent inhibitors of oxidative modification of low-density lipoprotein (LDL), which is considered a key factor in atherogenesis. The aim of this study was to explore i) the reactivity of a series of new synthesized 2,3-diaryl-indoles with several well characterized LDL oxidation systems and ii) subsequent effects on an inflammatory/atherogenic microenvironment. The results demonstrate that under the present experimental conditions 2,3-diaryl-indoles showed potent ROS scavenging activity and were able to markedly inhibit LDL oxidation. Subsequently, this led to a substantial decrease of modified LDL uptake by scavenger receptors in THP-1 macrophages in vitro and in rats in vivo. Moreover, modified LDL-mediated monocyte/neutrophil adhesion to endothelial cells, macrophage NFκB activation, as well as macrophage and endothelial cell cytokine release was diminished in vitro. The reduction of modified LDL-induced atherogenic effects by antioxidant 2,3-diaryl-indole coxibs may widen the therapeutic window of COX-2 targeted treatment.

It has been shown that a hydroxyethylstarch solution significantly increases the aortic distensibility coefficient (ADC) as compared to other non-hydroxyethylstarch colloid solutions. In order to investigate whether the effect of hydroxyethylstarch on ADC is class-specific, we investigated the effect of two hydroxyethylstarch solutions (HES 200: Elohes and HES 240: Hesteril) on the ADC and compared them with two other colloid solutions: 5% albumin and fluid gelatin (Gelofusin) in a rabbit model of acute isovolumic hemodilution. Twenty-eight male New Zealand white rabbits were anesthetized and randomly allocated to receive (n=7, each): albumin, hydroxyethylstarch-200, hydroxyethylstarch-240 and gelatin for acute isovolumic hemodilution by exchanging 13 ml.kg(-1) body weight of blood with an identical volume of the test solution. Blood viscosity, mean arterial pressure, aortic blood flow and heart rate were measured and ADC was calculated. All groups were comparable with respect to arterial pressure, heart rate and aortic blood flow velocity before and after isovolumic hemodilution. After hemodilution, ADC coefficient remained unchanged as compared with pre-hemodilution values with albumin, hydroxyethylstarch-240 and gelatin, whereas a sustained 3 fold increase was observed with hydroxyethylstarch-200. These results demonstrate that minor physicochemical differences between two hydroxyethylstarch solutions result in measurable differences in ADC and suggest that the clinical effects of colloids should probably be analyzed for each type of colloid and not for classes of colloids (e.g. hydroxyethylstarch or fluid gelatins).

There are several reports demonstrating an involvement of bacterial toxins in the rigidity of red blood cells (RBC). The present study investigates the influence of E. coli F-583-Rd lipid A on RBC deformability under mechanical shear stress. Verapamil (Ca(2+) channel inhibitor), staurosporine (protein kinase inhibitor) and Y-27632 (rho-kinase inhibitor) were used to modify the effect of lipid A on RBC deformability. We also determined if E. coli F-583-Rd Lipid A could induce an increase of intracellular Ca(2+) concentration. For the deformation measurements RBC (10 adult donors) were incubated with E. coli F-583-Rd lipid A (100 μg/ml) and also co-incubated with either verapamil (10(-7) mol/l), staurosporine (10(-7) mol/l) or Y-27632 (10(-7) mol/l). The deformation of the RBC under different shear stresses (0.6-60 Pa) was measured by a shear stress diffractometer (Rheodyne SSD). Intracellular Ca(2+) was determinded by flow cytometry in RBC incubated with Lipid A and labeled with fluorescent Fluo-4/AM which binds intracellular Ca(2+) with high affinity resulting in enhanced green fluorescence intensity. At increasing shear stresses Lipid A induced a significantly lower elongation. Co-incubation of the erythrocytes with verapamil or staurosporine inhibited lipid A induced decrease in elongation while Y-27632 had no effect. Verapamil, Staurosporine and Y-27632 did not influence the elongation response of the cells under control conditions. Lipid A induced a marked increase in fluorescence Fluo-4/AM indicating increased intracellular Ca(2+). These results suggest that E. coli F-583-Rd lipid A is able to influence red blood cell rigidity by a rapid and significant increase of intracellular Ca(2+) concentration. Verapamil and staurosporine abolished the decrease in deformability of Lipid A incubated RBC.

Platelet primary hemostatic function occurs under high shear conditions. Cyclooxygenase inhibitors such as ibuprofen inhibit this platelet aggregation under high shear rates only to a limited extent. This prompted the present study on 10 healthy volunteers treated with 100 mg aspirin for 4 weeks. The platelet function analyser (PFA-100) was used to measure the closure time (CT) of a membrane pore coated with collagen and epinephrine by aggregating platelets under shear rates of 5000-6000 s(-1). A first dose of 100 mg aspirin prolonged the CT above the normal range in 4 of 10 individuals, but the CT for the whole group (153+/-42 s) was not different from baseline (112+/-18 s). After 7 and 28 days of treatment, CTs were >300 s in 8 individuals and the mean values for the group were significantly higher than baseline. However, one subject had an intermediate response and one had an aspirin non-responsiveness, which was not overcome by 300 mg aspirin daily. The CT was normalized in 4 individuals 48 h after the last aspirin dose and in 7 individuals after 72 h, when the mean value for the group became not different from baseline. We conclude that the platelet function measured with the PFA-100 is not inhibited significantly after a single dose of 100 mg aspirin, is thereafter inhibited consistently in the majority, but not all individuals during a 4 week treatment, and returns to normal in 48-72 h. Since large interindividual differences exist, monitoring of platelet inhibition at the beginning of an aspirin treatment should be considered and validated in a prospective study.

Secondary cerebral ischemic injury is a major cause of mortality and disability from subarachnoid hemorrhage (SAH). In this study, the protective effects of nimodipine were investigated. Rat SAH models were divided into a sham-operated group, a saline-controlled, and a nimodipine-treated group by an endovascular piercing method. Nimodipine, 100 microg/kg BW was injected intraperitoneally 30 minutes before operation and was repeated every 6 hours. Dynamic changes in cortical regional cerebral blood flow (rCBF) using a laser Doppler flow-meter probe, and somatosensory evoked potentials (SEP) were estimated. Brain water content, sodium, potassium and calcium contents at different time points were determined. rCBF, latency of SEP, brain water and electrolyte contents did not statistically change in sham-operated rats. In saline-controlled rats, rCBF decreased immediately after SAH, and stabilized at low levels within 24 hours. The latency of SEP delayed gradually after SAH. Brain water and sodium increased, while potassium decreased at 6 hours and 24 hours. Brain calcium content increased significantly from 1 hour to 24 hours after induction of SAH. Extents of alterations of the above parameters caused by SAH in the nimodipine-treated group were less than those in the saline-controlled group, statistically. In conclusion, nimodipine partly prevents a decrease in cerebral blood supply and attenuates secondary cerebral ischemic injury after SAH.

3,4-dihydroxy-phenyl lactic acid (DLA) and salvianolic acid B (SAB) are two major water-soluble components of Salvia miltiorrhiza (SM). Previous works have revealed the ability of DLA and SAB to scavenge oxygen free radicals, inhibiting the expression of adhesion molecules CD11b/CD18 in neutrophil. Cardiotonic pills (CP), which is a traditional Chinese medicine compound preparation containing DLA and SAB, was found to inhibit venular thrombosis induced by photochemical reaction (PR) in rat mesentery. The present study addressed the effect of DLA and SAB on PR-induced thrombosis in rat mesentery by utilizing a microcirculation dynamic viewing system. The result demonstrated that both DLA and SAB delayed thrombus-initiation time, while DLA also prolonged thrombus half-size time. The experiments explored the mechanism underlying that the dihydrorhodamine 123 (DHR) fluorescence in the mesenteric venular walls after PR challenge was diminished by pretreatment with either DLA or SAB, the expression of CD18 in neutrophils elicited by PR was depressed by administration of DLA, while mast cell degranulation in rat mesentery induced by PR was damped by SAB. The antioxidant potential of the two substances is likely to be responsible for their most beneficial effects on thrombosis, through either directly scavenging the peroxides produced and/or indirectly depressing the expression of adhesion molecules in neutrophil.

This study intended to examine the effect of 3,4-dihydroxyphenyl lactic acid (DLA), a major ingredient of Salvia miltiorrhiza, on lipopolysaccahride (LPS) -induced mouse cerebral cortical microcirculatory disturbance. Velocity of red blood cells in, and albumin leakage from venules, and the numbers of leukocytes rolling on, and adherent to the venular wall were determined by an up-right microscope after LPS (5 mg/kg/h) infusion with or without administration of DLA (5 mg/kg/h). Expression of adhesion molecules CD11b/CD18 and L-selectin on neutrophils, plasma concentration of tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) were evaluated by flow cytometry. Concentration of TNF-α in supernatants of LPS-stimulated mononuclear cells was evaluated in vitro by flow cytometry as well. LPS exposure significantly increased the number of rolling and adherent leukocytes as well as albumin leakage, and decreased the velocity of red blood cells in venules. In addition, LPS stimulation apparently increased the expression of CD11b/CD18 on neutrophils, the concentration of plasma TNF-α, and the production of TNF-α from mononuclear cells. Treatment with DLA significantly ameliorated LPS-induced insults in mice, including cerebral microcirculatory disturbance, the expression of CD11b/CD18 on neutrophils, and the increased concentration of plasma TNF-α and the production of TNF-α from mononuclear cells.

Erythrocyte aggregation and elongation index (EI) (deformability) are measured at temperatures ranging from 5 degrees C to 37 degrees C by aggregometer MA1, and diffractometer Rheodyne SSD and the LORCA, respectively. The test samples are prepared from blood obtained from healthy subjects in test tubes containing EDTA as an anticoagulant and prior to measurement are placed in a water bath for 30 min, maintained at respective temperatures. The aggregation process is given in terms of primary aggregation time and indices of aggregation. These show significant to highly significant changes with the increase of temperature compared to that at 5 degrees C. The EI increases with the increase of shear stress but shows significant decrease with the decrease of temperature.

In this study we present a three-dimensional angiogenesis assay in vitro that allows the evaluation of the influence of Poly(lactic-co-glycolic acid) based implants seeded with VEGF-A165 stimulated/activated human CD14+ monocytes on the attraction and migration of human micro vascular endothelial cells (HMVEC-L). Primary HMEC of the capillary bed were cultured on an extracellular matrix generated by bovine corneal endothelial cells (BCEC). The HMEC layer was covered by an agarose gel, upon which a Poly(lactic-co-glycolic acid)/CaP polymer with a Calcium-Phosphate (CaP) nanostructured surface was placed. This scaffold has already been shown to interact with endothelial cells and endothelial progenitor cells respectively in vivo. It was seeded with angiogenically stimulated (VEGF-A165) human CD14+ monocytes, to get a monocyte/macrophage fraction, which can promote angiogenesis, tissue remodelling and tissue repair due to the secretion of growth factors, cytokines, chemokines and enzymes. The study demonstrated that this assay is suitable to test angiogenic effects by stimulated human CD14+ monocytes on human microvascular endothelial cells influenced by Poly(lactic-co-glycolic acid)/CaP scaffolds with a nanostructured CaP surface. The assay can exclude effects on migration caused by gravity and also allows testing in a physiological environment on an extracellular matrix secreted by endothelial cells.

To investigate the extent to which B-flow and B-flow with 3D postprocessing and speckle reduction imaging (SRI) have advantages in appraising the morphology of a high-grade stenosis of the internal carotid artery (ICA) for preinterventional planning and for postinterventional ultrasonographic follow-up. A comparative appraisal of flow with CCDS, power Doppler, B-flow and 3D B-Flow with SRI were carried out prospectively in 50 patients with >70% stenosis according to NASCET criteria in contrast medium-enhanced MRA before and after the intervention. After stenting of the internal carotid artery (ICA), i.a. digital substraction angiography (DSA) served as an additional reference method. In >70% ICA stenosis, simultaneous imaging of the pre-stenotic, intra-stenotic and post-stenotic flow was attained with B-flow in 45/90 cases (90%), with power Doppler in 39/50 cases (78%) and with CCDS in only 31/50 cases (62%). After intervention, a complete detection of flow without overwriting or blooming artifacts was achieved in all 50 patients only by B-flow. The intrastenotic flow (p<0.05) could be better demarcated against the lumen and the vessel wall before the intervention, whereas the flow within the stent could be very much better appraised after the intervention (p<0.01) using 3D postprocessing of B-flow with additional SRI. Re-stenoses with hypoechoic vascular wall changes (3/50 patients) were detected at an early stage using B-flow. B-flow technique with SRI and 3D postprocessing can facilitate the intrastenotic detection of flow in >70% ICA stenosis with fewer flow artifacts. After stenting, the perfused vascular lumen shows less flow artifacts compared with CCDS and power Doppler. In order to elucidate hemodynamic changes, additional Doppler examinations are still necessary.

The results for blood flow in the carotid artery bifurcation on the basis of numerical simulation of Navier-Stokes equations are presented in this study. Four cases of carotid bifurcation are considered: common carotid artery (CCA) bifurcation without stenoses and with one, two and three stenoses are presented too. The results are found by performing numerical simulations considering one pulse wave period based on the finite volume discretization of Navier-Stokes equations. The structures of the flow around the bifurcation are received. Using the numerical simulation the authors can trace the deformation of the pulse wave from common the carotid artery (CCA) to the internal carotid artery (ICA) and external carotid artery (ECA). The axial velocity and wall shear stress (WSS) distribution and contours are presented considering the characteristic time points. The results of the WSS distribution around the bifurcation allow a prediction of the probable sites of stenosis growth.

To investigate the dynamic value of contrast medium-enhanced ultrasonography with Optison for appraisal of the vascularization of hepatic tumors using harmonic imaging, 3D-/power Doppler and B-flow. 60 patients with a mean age of 56 years (range 35-76 years) with 93 liver tumors, including histopathologically proven hepatocellular carcinoma (HCC) [15 cases with 20 lesions], liver metastases of colorectal tumors [17 cases with 33 lesions], metastases of breast cancer [10 cases with 21 lesions] and hemangiomas [10 cases with 19 lesions] were prospectively investigated by means of multislice CT as well as native and contrast medium-enhanced ultrasound using a multifrequency transducer (2.5-4 MHz, Logig 9, GE). B scan was performed with additional color and power Doppler, followed by a bolus injection of 0.5 ml Optison. Tumor vascularization was evaluated with coded harmonic angio (CHA), pulse inversion imaging with power Doppler, 3D power Doppler and in the late phase (>5 min) with B-flow. In 15 cases with HCC, i.a. DSA was performed in addition. The results were also correlated with MRT and histological findings. Compared to spiral-CT/MRT, only 72/93 (77%) of the lesions could be detected in the B scan, 75/93 (81%) with CHA and 93/93 (100%) in the pulse inversion mode. Tumor vascularization was detectable in 43/93 (46%) of lesions with native power Doppler, in 75/93 (81%) of lesions after administering contrast medium in the CHA mode, in 81/93 (87%) of lesions in the pulse inversion mode with power Doppler and in 77/93 (83%) of lesions with contrast-enhanced B-flow. Early arterial and capillary perfusion was best detected with CHA, particularly in 20/20 (100%) of the HCC lesions, allowing a 3D reconstruction. 3D power Doppler was especially useful in investigating the tumor margins. Up to 20 min after contrast medium injection, B-flow was capable of detecting increased metastatic tumor vascularization in 42/54 (78%) of cases and intratumoral perfusion in 17/20 (85%) of HCC cases. All 19 hemangiomas were correctly classified by phase inversion imaging. Contrast medium-enhanced ultrasound investigation of liver tumors with Optison allowed reliable detection of tumor foci and, in most cases, appraisal of tumor vascularization. The time available for evaluation of tumor margin vascularization was substantially longer in B-flow.

Pre-surgical evaluation of the extent of internal carotid artery stenosis (ICA) according to NASCT criteria using digital 3D ultrasound methods. In a prospective study, 25 patients (54-88 years, mean 75) with neurological deficits and the diagnosis of ICA stenosis underwent pre-surgical ultrasound examination using Color Coded Duplex Sonography (CCDS), 3D CCDS, 3D power Doppler, 3D B-flow, contrast enhanced 3D B-flow, and CTA/MRA. Ultrasound was performed by an experienced examiner with a multifrequency linear transducer (6-9 MHz, Logiq 9, GE). After bolus injection of 2.4 ml Sonovue i.v., low mechanical index technique (MI<0.16) was used for contrast enhanced 3D B-flow. As reference method for evaluation of the extent of ICA stenosis each patient underwent CTA (multislice CT, Sensation 16, Siemens) and/or MRA (1.5 T, Symphony Siemens). Indications for surgery (carotid EEA) followed the NASCET criteria. All images were interpreted and evaluated independently by two observers with three measurements of the degree of the ICA stenosis. For assessment of the extent of stenosis a 10%-scale from 50% to 99% was used. Statistical analysis was performed using Spearman Correlation and Wilcoxon Signed Rank Test with a significance threshold of p<0.05. Assessment of the extent of ICA stenosis during surgery and in CTA/MRA displayed a range from 60% to 99% (mean 80%). Non significant differences were found with paired Wilcoxon test only for 3D B-flow with and without contrast medium (p<0.05). Correlation with surgical evaluation regarding the extent of ICA stenosis using Spearman correlation teat was 0.77 for B-scan, 0.90 for 3D CCDS, 0.84 for 3D Power Doppler, 0.91 for B-flow and 0.93 for contrast enhanced 3D B-flow. When circular calcifications were present, contrast enhanced flow detection of 3D B-flow proved to be useful. Visualisation of intrastenotic variances of severe and profound stenosis (70-99%) without blooming and reverberation artefacts was possible only with 3D B-flow. This facilitates the detection of the morphology of plaques ulcers as an embolic source. In correlation with surgery and CTA/MRA, a valid evaluation of the extent and morphology of ICA stenosis using 3D B-flow, with and without contrast medium, is feasible.

Recent studies on the mechanisms of ischemic preconditioning in myocardial tissue have presented convincing evidence that multiple protective pathways converge on inhibition of glycogen synthase kinase-3β (GSK-3β). To directly address the role of GSK-3β in ischemia and reperfusion (I/R) of the lung, a rat model of left lung in situ ischemia was used. The specific non-competitive inhibitor of GSK-3β, TDZD-8, was injected (3 mg/kg, vehicle in controls) 5 min before the left lung hilum was occluded for 60 min. Animals in the ischemia group underwent the same treatment, but without administration of TDZD-8. Lung functional and biochemical parameters were determined at time points 15 min and 60 min reperfusion. Treatment with TDZD-8 improved gas exchange (arterial pO2), but I/R-induced inflammation (plasma interleukin-6, leukocyte invasion) was not affected. The I/R cycle induced a rapid (15 min reperfusion) increase of protein tyrosine phosphorylation, including the activating phosphorylation of focal adhesion kinase at Tyr397, Tyr407, Tyr577, and Tyr861, and the non-receptor kinase Src at Tyr416. The phosphorylation was blocked by the GSK inhibitor. This effect may be related to the reduced plasma level of the strong effector of focal adhesion kinase, transforming growth factor-β1, in the TDZD group. The underlying mechanisms are elusive, but they deserve further investigation, especially in relation to the early increase of lung permeability in this rat model of I/R injury. In conclusion, the results suggest that inhibition of GSK-3β improves rat lung function during an I/R cycle, but only during the early reperfusion phase.

Unlabelled: Exercise training decreases blood viscosity in athletes parallel with metabolic improvements mostly characterized by an increase in insulin sensitivity. Patients with low insulin sensitivity exhibit a host of metabolic disorders that may also benefit from regular training. However, the hemorheologic aspects of training in such subjects are not known and we aimed at characterizing them. Subjects: Thirty-two obese insulin resistant subjects were tested before and after 2 months. Twenty-one of them were trained (3x45 min/wk) at a level defined by exercise calorimetry and corresponding to the power at which lipid oxidation reaches a maximum (LIPOX(max )) and eleven served as controls. The two groups were matched for age and body mass index. There was no weight change in controls while the 2 months training period decreased weight by 2.5 kg (p<0.02). This change was totally explained by a loss in fat mass (-2.7 kg, p<0.02) while fat free mass remained unchanged. Blood rheology was unchanged in the control group while training improved plasma viscosity eta(pl) (before: 1.43+/-0.03 mPa.s; after: 1.35+/-0.03 mPa.s, p<0.02). There was no change in either hematocrit, red cell rigidity or red cell aggregation. The balance of substrates oxidation shifted towards a higher use of lipids (point of crossover where subjects oxidize 70% carbohydrates 30% lipids: before 39.3+/-6.9 watts; after 70.8+/-6 watts, p<0.001; point where lipid oxidation is maximal (LIPOX(max )) before: 16.5+/-1.4 watts; after: 21.4+/-1.3 watts, p<0.001) and V(O(2max )) increased by 74% (p<0.01). Consistent with observations in athletes, the metabolic and ergometric improvements induced by training reduces eta(pl) in sedentary, insulin resistant patients, but at those low levels training does not appear to induce "autohemodilution" (as reflected by hematocrit) neither it improves red cell deformability or aggregation. The reliability of eta(pl) as simple and unexpensive marker of efficiency of training in insulin resistant patients should be further evaluated.

Unlabelled: The concentration of plasma fibrinogen (FIB) is an important factor in the coagulation cascade and also in the determination of blood and plasma viscosity depending on both genetic and acquired factors. The -455G/A polymorphism of the beta-FIB gene is connected to the plasma concentration of FIB but the effect of Leiden mutation on hemorheological parameters is unclear. The two genetic polymorphisms were studied by polymerase chain reaction in healthy subjects and ischemic stroke cohort and the effects on the concentration of plasma FIB, whole blood and plasma viscosity of patients as well. A total of 278 ischemic stroke patients and 173 control subjects were enrolled. Marcro-rheological parameters as plasma FIB concentration, whole blood viscosity (90 sec(-1) shear rate) and plasma viscosity have been measured also in the subgroup of young (age < 50 years) and in a subgroup of non-smoker patients. Results: No significant difference was found in the prevalency of H1/H2 genotype between controls and cases in pooled stroke group OR 0.95 (95% CI: 0.47-1.27), however H2/H2 genotype frequency was increased in young subgroup of patients (OR: 1.66 95% CI: 0.52-5.25). Plasma FIB concentration was increased both in the total cohort (p < 0.05) and in the non-smoker subgroup (p < 0.03) of patients carried H2/H2 as compared to H1/H1 genotype and the prevalence was increased in the group of patients having plasma FIB concentration > 4 g/l (p < 0.05). The whole blood viscosity was elevated in the H2/H2 group as compared to the group carrying wild type (p < 0.03). A tendency of increased plasma viscosity in the group of patients with H2/H2 genotype as compared to wild type was found (p = 0.07). Leiden mutation prevalence showed an increased risk OR: 1.67 (95% CI: 0.75-3.70) in the young patients group as compared to controls. In patients who have had the highest plasma viscosity, higher frequency of Leiden mutation was detected as compared to wild type, in total group (p = 0.01), in young patients (p = 0.03) and in subgroup of non-smoker patients (p = 0.05). Conclusions: Our findings support the notion that the homozigous variant of beta-FIB gene can raise both plasma FIB concentration and whole blood viscosity. Leiden mutation connected to the elevation of plasma viscosity could demonstrate a new pathway of increased thrombophylic potential in ischemic stroke patients.

Genetic polymorphisms in plasminogen activator inhibitor-1 gene-675 4G/5G (PAI-1 4G/5G) are claimed to contribute to an increased risk of venous thromboembolism. Inherited thrombophilia, on the other hand, is associated with the occurrence of spontaneous abortions. The objective of this study was, to explore the significance of genetic polymorphisms of PAI-1 4G/5G with particular emphasis on 4G alleles in pregnant women suffering from venous thromboembolism or early spontaneous abortion, respectively. Therefore genetic PAI-1 4G/5G polymorphisms were studied in 108 pregnant females suffering from venous thromboembolism (n=69) or from spontaneous abortion (<20 week, n=39), respectively. Healthy volunteers (n=238) were taken as controls. The frequencies of 4G alleles (4G/4G or 4G/5G genotypes) of PAI-1 were significantly higher in venous thromboembolism (OR: 3.40, p=0.0088) and slightly higher, but not significantly, in abortions (RR: 2.33; p=0.1162) compared to controls. The incidence of 4G-carriers in females with abortion was 0.68 (-32%) compared to women suffering from venous thromboembolism alone. We conclude from these data, that the occurrence of PAI-1 4G/4G or 4G/5G genotypes, respectively, is clinically significant for the pathogenesis of venous thromboembolism in pregnancy but not for early abortion.

The aim of this study was to analyze the effects of motor rehabilitation, in the form of rhythmic exercise to music, on the rheological characteristics of blood in older women. The study included 30 women (65-80 years of age), and the control group was comprised of 10 women of corresponding age. Women from the experimental group were subjected to a five-month rehabilitation program, in the form of rhythmic exercise performed to music (three 30-minute sessions per week); women from the control group were not involved in any regular physical activity. Blood samples from all the women were examined for hematological, rheological, and biochemical parameters prior to the study and five months thereafter. The rehabilitation program was reflected by a significant improvement of erythrocyte count and hematocrit. Furthermore, an improvement of erythrocyte deformability was observed by lower shear stress levels, while no significant changes were noted by the higher shear stress values. The rehabilitation resulted in a marked decrease of the aggregation amplitude while no significant changes were observed in aggregation index and total aggregation half-time. Additionally, the training regimen was reflected by a significant increase in the plasma viscosity, while no significant changes in fibrinogen levels were noted.

The rheological properties of human leukocytes (WBCs) have been studied using the micropipette aspiration and the filtration technique. Partial micropipette (i.d. 2.8-4.5 microm) aspiration of individual leukocytes under constant aspiration pressure of 8 mm H20 and measurement of the aspirated length as a function of time (creep experiments) according to the Evans model have been carried out and the apparent viscosity mu(app) was estimated. In the filtration experiments, using the Hemorheometer, the Index Rigidity of Leukocytes, ILR, was also estimated. The apparent viscosity mu(app) of normal PMN and MNC was significantly different p < 0.05), while the LYM and PMN had no statistical difference (p < 0.5). The leukocytes of the cell line HL-60 were more rigid than the normal PMN (p < 0.01), while the PMN from patients with type II diabetes mellitus were more rigid than the normal PMN (p < 0.005). The results of IRL showed similar differences among all of the leukocyte subpopulations. Comparison of these findings suggests a possible relationship between ILR and mu(app) which in this case is: ILR = 598 + 0.54 mu(app) (r = 0.986, p-value 0.0003).

A high-dose (7 Gy) whole-body 60Co irradiation for a short period caused disturbances of hematopoietic function. A decrease in the hematocrit of the circulating blood lasted for about 15 days, thus forming an anemic animal model. We studied the influence of high-dose 60Co irradiation on hemorheologic parameters: percentage of reticulocytes, RBC deformability, sedimentation rate and plasma fibrinogen concentration in the rabbit. It was found that the plasma fibrinogen concentration increased to twice more than control level and that percentage of reticulocytes in circulation disappeared immediately after irradiation. The deformation index of RBCs in shear flow decreased from a value of 58% down to a value of 42% in the first two weeks and gradually returned to control levels about 40 days after 60Co irradiation. Our results showed that a short period of high-dose 60Co irradiation caused severe and relatively long-lasting damage of hematopoietic system in animals' body.

Heart rate variability (HRV) and haemorheology adaptations to 12 wk of varied-dose treadmill walking were investigated in women aged 65-74 yr with type 2 diabetes. Subjects were randomly allocated into two groups where exercise frequency and session duration were manipulated (Group 1: 2 × 60 min·wk(-1) or Group 2: 4 × 30 min·wk(-1)), but intensity and accumulated weekly duration of exercise were consistent between groups (100% gas-exchange threshold; 120 min·wk(-1)). Twelve weeks of exercise training significantly improved peak oxygen uptake, time to exhaustion, and gas-exchange threshold (p < 0.05), independent of exercise group. Exercise training did not significantly change glycaemic control or body mass. Red blood cell (RBC) aggregation and RBC deformability significantly decreased (p < 0.05) for both groups. No change in HRV was observed for Group 1, whereas several key indicators of HRV were significantly improved in Group 2 (p < 0.05). The present study was the first to report decreased RBC aggregation following an exercise-only intervention and that exercise training improved RBC aggregation without a concomitant improvement in glycaemic control. The accumulated weekly exercise duration may be the most important training component for the prescription of exercise in older women with type 2 diabetes.

It is not well-established whether the alterations that the hemorheological profile undergoes with aging are an effect of concomitant cardiovascular risk factors or are due to age itself. To clarify this issue, we investigated the effect of age on blood rheology in a population of 927 healthy subjects from eastern Spain aged between 16-85 years, divided into four age groups (<30, 30-44, 45-50, ≥60 years) with and without cardiovascular risk factors. We determined blood viscosity, corrected blood viscosity (BVc), plasma viscosity (PV), erythrocyte aggregation (EA), erythrocyte deformability (EEI60) and fibrinogen, along with glucose and plasma lipids. We found that corrected blood viscosity (p = 0.007), plasma viscosity, erythrocyte aggregation, fibrinogen, glucose, and plasma lipids increased with age (p < 0.001). When subjects with cardiovascular risk factors were excluded, the effect of age on blood rheology persisted for all the cited parameters (p < 0.028). EEI60 increased with age (p = 0.033), and it was attributable to a concomitant increase in mean corpuscular volume (p < 0.001). In the Pearson's correlations, age was related to all the parameters analyzed (P < 0.019). The logistic regression analysis revealed that PV ≥1.30 mPa·s, BVc ≥4.90 mPa·s and EA1 ≥8.3 were associated with age ≥60 years (*p = 0.049, *p = 0.013, *p = 0.045, respectively). These results indicate that, although the presence of cardiovascular risk factors influences rheological properties, aging itself is associated with deterioration of rheological blood behavior, mostly related to inflammatory and lipidic changes.