Clinical Hemorheology and Microcirculation

Blood vessel occlusions of various origin, depending on the duration and extension, result in tissue damage, causing ischemic or ischemia-reperfusion injuries. Necessary surgical clamping of vessels in vascular-, gastrointestinal or parenchymal organ surgery, flap preparation-transplantation in reconstructive surgery, as well as traumatological vascular occlusions, all present special aspects. Ischemia and reperfusion have effects on hemorheological state by numerous ways: besides the local metabolic and micro-environmental changes, by hemodynamic alterations, free-radical and inflammatory pathways, acute phase reactions and coagulation changes. These processes may be harmful for red blood cells, impairing their deformability and influencing their aggregation behavior. However, there are still many unsolved or non-completely answered questions on relation of hemorheology and ischemia-reperfusion. How do various organ (liver, kidney, small intestine) or limb ischemic-reperfusionic processes of different duration and temperature affect the hemorheological factors? What is the expected magnitude and dynamics of these alterations? Where is the border of irreversibility? How can hemorheological investigations be applied to experimental models using laboratory animals in respect of inter-species differences? This paper gives a summary on some of our research data on organ/tissue ischemia-reperfusion, hemorheology and microcirculation, related to surgical research and experimental microsurgery.

Despite of the studies on widespread biological effects of irradiation, surprisingly only little number of papers can be found dealing with its in vivo hemorheological impact. Furthermore, other studies suggested that low-dose irradiation might differ from high-dose in more than linear ways. On Balb/c Jackson female adult mice hematological and hemorheological impacts of total body irradiation were investigated 1 hour following 0.002, 0.005, 0.01, 0.02, 0.05 and 0.1 Gy dose irradiation. In case of 0.01 Gy further groups were analyzed 30 minutes, 2, 4, 6, 24 and 48 h after irradiation. According to the results, it seems that the dose-dependent changes of blood micro-rheological parameters are not linear. The irradiation dose of 0.01 Gy acted as a point of ‘inflexion’, because by this dose we found the most expressed changes in hematological parameters, as well as in red blood cell aggregation, deformability and osmoscan data. The time-dependent changes showed progressive decrease in pH, rise in lactate concentration, further decrease in erythrocyte aggregation index and deformability, with moderate shifting of the optimal osmolarity point and modulation in membrane stability. As conclusion, low-dose total body irradiation may cause micro-rheological changes, being non-linearly correlated with the irradiation dose.

Ischemia-reperfusion-caused hemorheological alterations have been widely studied but the effect of testicular ischemia-reperfusion has not so far. In this study 14 Sprague-Dawley rats were involved. In the ischemia-reperfusion group under general anaesthesia the left testis was explored by opening the scrotum then the deferent duct and vasculature were clamped for 30 minutes. Testicular microcirculation was monitored by laser Doppler flowmetry. The right testis was untouched. In the control group: only anaesthesia was induced. Blood sampling occurred before and after ischemia, at the 60th minute of reperfusion and on the 1st postoperative day for determining hematological parameters (microcell-counter), erythrocyte deformability (slit-flow ektacytometer) and erythrocyte aggregation (light-transmission aggregometer). After the last blood sampling, testicles were removed for histological examination. Hematological parameter changes reflected inflammatory response. Erythrocyte deformability showed a worsening already at the 60th minute of reperfusion compared to base and control values. By the 1st postoperative day further decrease was observed. Erythrocyte aggregation significantly enhanced with great magnitude versus base and control values (p < 0.001). However, conventional histological examinations did not show marked testicular injury. The experienced changes can attract attention to the testicular ischemia-reperfusion causing significant effects on hemorheological parameters, which can lead to further harmful microcirculatory consequences.

Obesity is associated with structural alterations in subcutaneous small resistance arteries. The aim of the present work is to study modifications of perforators vessels of abdominal wall and subcutaneous tissue characteristics in obese patients after massive weight loss. An anatomo-radiologic study was carried out on 15 patients (5M, 10F, mean age 54.9y), who underwent abdominoplasty after massive weight loss. Their pre-operative Computed Tomographic (CT) results of the anterior abdominal wall were compared with CT of 15 normal weighted controls. Anatomo-microscopic and morphometric examinations were conducted on full-thickness specimens of panniculectomy samples. 10 right panniculectomy were sampled from donor cadavers. All the measurements were taken on transverse sections. In patients, at CT the mean luminal diameter (LD) and standard deviation of perforator branches of the deep inferior epigastric artery (DIEA) was 3.7 ± 0.4 mm (control 2.2 ± 0.1 mm; p < 0.05). At microscopic examination, the wall thickness of perforator arteries was 212.7 μ ± 83.9 versus 143.9 ± 32.8 (p < 0.05) deep to the superficial fascia and 120.4 μ ± 74.8 versus 72.3 ± 23.5 (p < 0.05) superficial to it. A thickening of the muscular layer was observable and the tunica media represented 71.4% ± 5.6 of the whole area of the wall (controls 37.1% ± 3.5, p < 0.0001). Our data demonstrate that the major LD of the perforators in patients matches with hypertrophy of the tunica media and we think that the major thickness of perforator walls can facilitate the microsurgical technique in free microsurgical flap reconstruction.

BACKGROUND AND AIM: The relationship between hyperlipidemia and blood and plasma viscosity is not completely clear. While increasing viscosity is often reported with increasing blood lipids, lipid-lowering treatments are often unable to normalize the viscosity values. Aim of this study is to try to clarify the relationship between blood lipids and viscosity. METHODS AND RESULTS: Apparently healthy subjects were enrolled (n = 410). Smokers, diabetics, obese, and hypertriglyceridemic (above 400 mg/dl) were excluded. Blood (at shear rate 225/s) and plasma viscosity were measured at 37°C. Erythrocyte rigidity (Tk) was calculated according to Dintenfass. Blood lipids and glucose were measured by routine methods. Hyperlipidemic subjects (n = 315) had higher values of plasma viscosity (1.44 ± 0.13 vs. 1.40 ± 0.12 cP, p = 0.007), and blood viscosity (4.51 ± 0.54 vs. 4.35 ± 0.55 cP, p = 0.013), compared to normolipidemic subjects (n = 95). In simple correlation analysis, plasma viscosity was directly associated with LDL cholesterol, and inversely with Tk and HDL cholesterol. In multiple regression analysis the association with LDL and HDL was strengthened, though these two variables as a whole accounted for only 5% (adjusted R²) of the variability of plasma viscosity. Blood viscosity was significantly associated with haematocrit, plasma viscosity, Tk and all considered variables but age in simple correlation analysis, but only with haematocrit, plasma viscosity and Tk in multiple regression analysis. CONCLUSIONS: LDL cholesterol and HDL cholesterol influence plasma viscosity, but not blood viscosity. Triglycerides up to values of 400 mg/dl do not seem to have important effects, at least in apparently healthy subjects and at the shear rates used in the present study. The contribution of LDL and HDL cholesterol to plasma viscosity seems however quite limited.

The ability of red blood cells (RBC) to undergo a wide range of deformations while traversing the microvasculature is crucial for adequate perfusion. Interpretation of RBC deformability measurements performed in vitro in the context of microvascular perfusion has been notoriously difficult. This study compares the measurements of RBC deformability performed using micropore filtration and ektacytometry with the RBC ability to perfuse an artificial microvascular network (AMVN). Human RBCs were collected from healthy consenting volunteers, leukoreduced, washed and exposed to graded concentrations (0–0.08%) of glutaraldehyde (a non-specific protein cross-linker) and diamide (a spectrin-specific protein cross-linker) to impair the deformability of RBCs. Samples comprising cells with two different levels of deformability were created by adding non-deformable RBCs (hardened by exposure to 0.08% glutaraldehyde) to the sample of normal healthy RBCs. Ektacytometry indicated a nearly linear decline in RBC deformability with increasing glutaraldehyde concentration. Micropore filtration showed a significant reduction only for concentrations of glutaraldehyde higher than 0.04%. Neither micropore filtration nor ektacytometry measurements could accurately predict the AMVN perfusion. Treatment with diamide reduced RBC deformability as indicated by ektacytometry, but had no significant effect on either micropore filtration or the AMVN perfusion. Both micropore filtration and ektacytometry showed a linear decline in effective RBC deformability with increasing fraction of non-deformable RBCs in the sample. The corresponding decline in the AMVN perfusion plateaued above 50%, reflecting the innate ability of blood flow in the microvasculature to bypass occluded capillaries. Our results suggest that in vitro measurements of RBC deformability performed using either micropore filtration or ektacytometry may not represent the ability of same RBCs to perfuse microvascular networks. Further development of biomimetic tools for measuring RBC deformability (e.g. the AMVN) could enable a more functionally relevant testing of RBC mechanical properties.

The reversible aggregation of red blood cells (RBCs) continues to be of the basic science and clinical interest. Recently it has been reported about a specific binding between fibrinogen and unknown erythrocyte glycoprotein receptors. The aim of this study was to investigate whether the red blood cell aggregation (RBCA) include the cell-cell interaction using the membrane receptors that bind such ligands as fibrinogen or fibronectin. To test this hypothesis the RBCs were incubated with monafram - the drug of the monoclonal antibodies against glycoprotein (GP) IIb/IIIa, with the GPIIb-IIIa receptor antagonist tirofiban, epifibatide and with the fibrinogen inhibiting peptide. It has been found that the RBC incubation with monafram resulted in a marked RBCA decrease mainly in persons with high level of aggregation. Another research session has shown that RBC incubation with fibronectin was accompanied by a significant RBCA rise. The monafram addition to red cell incubation medium resulted in a significant RBCA lowering. The cell incubation with tirofiban and epifibatide issued in RBCA decrease. The similar results were obtained when RBCs were incubated with the fibrinogen inhibiting peptide. Although monafram, tirofiban, eptifibatide and the fibrinogen inhibiting peptide were related to fibrinogen function they didn't inhibit RBCA completely. Therefore, under moderate and low red blood cell aggregation the cell binding is probably related to nonspecific mode. It seems evident that the specific and nonspecific modes of red blood cell aggregate formation could co-exist. Additional theoretical and experimental investigations in this area are needed.