Reduction of erythrocyte deformability in Krushinskiĭ-Molodkina rats in acute hemorrhagic cerebrovascular circulation disorders

The ability of erythrocytes to change their shapes in the shear flow under acute strokes of hemorrhagic type in rats of the Krushinsky-Molodkina line was studied. The rigidity of membranes and the internal viscosity of erythrocytes were investigated by the laser diffraction method. The method consists in obtaining diffraction images from a thin layer of a dilute suspension of erythrocytes moving in the shear flow and subsequent computer processing of these images. It was shown that strokes of hemorrhagical type in rats of the Krushinsky-Molodkina line cause a reduction in the ability of erythrocytes to change theirs shapes.     

Shear rate and orientation of erythrocytes in pulmonary microvessels of bullfrogs

Wall shear rates in arterioles and capillaries in the surface of exposed bullfrog lung were estimated to be 436 and 975 sec-1, respectively, at the intra-lung pressure at which a maximum flow velocity was observed. These high shear rates will probably permit an orientation of erythrocytes in a high degree. An orientation of erythrocytes was confirmed at a wall shear rate smaller than 16 sec-1 by means of a microscope connected with a video camera system. Erythrocytes kept their long axis along the direction of the overall blood flow, even when the blood flow transiently stopped flowing during the diastolic phase. The orientation of erythrocytes in such a high degree will be effective to reduce the blood flow resistance in pulmonary microvessels.     

肠淋巴液引流对失血性休克大鼠红细胞流变性的影响

目的:观察肠淋巴液引流对失血性休克大鼠红细胞流变性指标以及血液黏度的作用。方法:Wistar雄性大鼠均分为假休克组、休克组(复制失血性休克模型)、引流组(复制失血性休克模型,自低血压1 h引流休克肠淋巴液)。在低血压3 h或相应时间,经腹主动脉取血,检测红细胞参数、红细胞电泳、红细胞沉降率(ESR)以及血液黏度,计算红细胞聚集指数、红细胞变形指数。结果:与假休克组比较,休克组红细胞数量、红细胞比积(HCT)、血红蛋白(Hb)、平均红细胞血红蛋白浓度(MCHC)、红细胞电泳率与迁移率、红细胞变形指数、全血黏度、全血低切与高切相对黏度和还原黏度显著降低,休克组平均红细胞体积、红细胞电泳时间、ESR、血沉方程K值与校正K值、红细胞聚集性指数、血浆黏度显著升高;引流组MCHC、红细胞电泳率与迁移率、全血黏度、全血低切与高切还原黏度均显著降低,引流组红细胞体积分布宽度(RDW-SD)显著增加。同时,引流组HCT、RDW-SD、红细胞变形指数、全血黏度、全血低切与高切相对黏度显著高于休克组;ESR、血沉方程K值与校正K值、红细胞聚集性指数、血浆黏度显著低于休克组。结论:休克肠淋巴液引流可改善失血性休克大鼠红细胞流变行为,从而改善血液流变性。     

Microscopic investigation of erythrocyte deformation dynamics

The understanding of erythrocyte deformation under conditions of high shear stress and short exposure time is central to the study of hemorheology and hemolysis within prosthetic blood contacting devices. A combined computational and experimental microscopic study was conducted to investigate the erythrocyte deformation and its relation to transient stress fields. A microfluidic channel system with small channels fabricated using polydimethylsiloxane on the order of 100 mum was designed to generate transient stress fields through which the erythrocytes were forced to flow. The shear stress fields were analyzed by three-dimensional computational fluid dynamics. Microscopic images of deforming erythrocytes were experimentally recorded to obtain the changes in cell morphology over a wide range of fluid dynamic stresses. The erythrocyte elongation index (EI) increased from 0 to 0.54 with increasing shear stress up to 123 Pa. In this shear stress range, erythrocytes behaved like fluid droplets, and deformed and flowed following the surrounding fluid. Cells exposed to shear stress beyond 123 Pa (up to 5170 Pa) did not exhibit additional elongation beyond EI=0.54. Two-stage deformation of erythrocytes in response to shear stress was observed: an initial linear elongation with increasing shear stress and a plateau beyond a critical shear stress.     

Dynamic adhesion of eryptotic erythrocytes to endothelial cells via CXCL16/SR-PSOX

Suicidal death of erythrocytes, or eryptosis, is characterized by cell shrinkage and cell membrane scrambling leading to phosphatidylserine exposure at the cell surface. Eryptosis is triggered by increase of cytosolic Ca2+ activity, which may result from treatment with the Ca2+ ionophore ionomycin or from energy depletion by removal of glucose. The present study tested the hypothesis that phosphatidylserine exposure at the erythrocyte surface fosters adherence to endothelial cells of the vascular wall under flow conditions at arterial shear rates and that binding of eryptotic cells to endothelial cells is mediated by the transmembrane CXC chemokine ligand 16 (CXCL16). To this end, human erythrocytes were exposed to energy depletion (for 48 h) or treated with the Ca2+ ionophore ionomycin (1 μM for 30 min). Phosphatidylserine exposure was quantified utilizing annexin-V binding, cell volume was estimated from forward scatter in FACS analysis, and erythrocyte adhesion to human vascular endothelial cells (HUVEC) was determined in a flow chamber model. As a result, both, ionomycin and glucose depletion, triggered eryptosis and enhanced the percentage of erythrocytes adhering to HUVEC under flow conditions at arterial shear rates. The adhesion was significantly blunted in the presence of erythrocyte phosphatidylserine-coating annexin-V (5 μl/ml), of a neutralizing antibody against endothelial CXCL16 (4 μg/ml), and following silencing of endothelial CXCL16 with small interfering RNA. The present observations demonstrate that eryptotic erythrocytes adhere to endothelial cells of the vascular wall in part by interaction of phosphatidylserine exposed at the erythrocyte surface with endothelial CXCL16.     

Increased hemorrhagic transformation and altered infarct size and localization after experimental stroke in a rat model type 2 diabetes

Background  

Interruption of flow through of cerebral blood vessels results in acute ischemic stroke. Subsequent breakdown of the blood brain barrier increases cerebral injury by the development of vasogenic edema and secondary hemorrhage known as hemorrhagic transformation (HT). Diabetes is a risk factor for stroke as well as poor outcome of stroke. The current study tested the hypothesis that diabetes-induced changes in the cerebral vasculature increase the risk of HT and augment ischemic injury.     

出血性卒中与MTHFR C677T基因多态性的相关性研究

摘要 目的：探讨与分析出血性卒中与亚甲基四氢叶酸还原酶（MTHFR）C677T基因多态性的相关性。方法：2020年2月到2021年4月选择在本地区诊治的H型高血压患者220例作为研究对象,检测所有患者的MTHFR C677T基因多态性状况,检测血清同型半胱氨酸、叶酸、维生素B12含量。随访判定患者的出血性卒中状况并进行相关性分析。结果：随访调查1年,220例患者中出现出血性卒中20例（出血性卒中组）,占比9.1 %。出血性卒中组的血清同型半胱氨酸含量明显高于非出血性卒中组,血清维生素B12、叶酸明显低于非出血性卒中组（P<0.05）。两组的MTHFR C677T基因型分布均符合Hardy-Weinberg遗传平衡,出血性卒中组的TT基因型、等位基因T占比分别为70.0 %、80.0 %,都显著高于非出血性卒中组的24.0 %、35.0 %（P<0.05）。Spearman相关系数分析显示H型高血压患者的血清同型半胱氨酸、叶酸、维生素B12含量、TT基因型、等位基因T都与出血性卒中存在相关性（P<0.05）。多元回归分析显示血清同型半胱氨酸、叶酸、维生素B12含量、TT基因型、等位基因T都为导致H型高血压患者出血性卒中发生的重要因素（P<0.05）。结论：H型高血压在随访过程中容易发生出血性卒中,也伴随有血清同型半胱氨酸、维生素B12、叶酸含量异常,MTHFR C677T的T基因型、等位基因T与出血性卒中存在相关性,也是导致出血性卒中发生的重要危险因素。     

Tank treading of optically trapped red blood cells in shear flow

Tank-treading (TT) motion is established in optically trapped, live red blood cells (RBCs) held in shear flow and is systematically investigated under varying shear rates, temperature (affecting membrane viscosity), osmolarity (resulting in changes in RBC shape and cytoplasmic viscosity), and viscosity of the suspending medium. TT frequency is measured as a function of membrane and cytoplasmic viscosity, the former being four times more effective in altering TT frequency. TT frequency increases as membrane viscosity decreases, by as much as 10% over temperature changes of only 4°C at a shear rate of ∼43 s⁻¹. A threshold shear rate (1.5 ± 0.3 s⁻¹) is observed below which the TT frequency drops to zero. TT motion is also observed in shape-engineered (spherical) RBCs and those with cholesterol-depleted membranes. The TT threshold is less evident in both cases but the TT frequency increases in the latter cells. Our findings indicate that TT motion is pervasive even in folded and deformed erythrocytes, conditions that occur when such erythrocytes flow through narrow capillaries.     

Deformation of erythrocytes under shear: A small-angle light scattering study

In this study, sharp small-angle light scattering (SALS) images of erythrocytes under increasing shear stresses in a Couette flow were obtained, and accurate measurements of the angular positions of the two first minima and maxima have been carried out. The deformed cells were assumed to be three-axis ellipsoids of constant volume for all shear stresses. Application of the Physical Optics Approximation (POA) then permitted the determination of the cell dimensions as a function of the applied shear stress. Our results agree with determinations obtained by other methods.     

Use of spin label and the flow-induced ESR spectral difference for studying erythrocyte deformation

ESR spin-labeling method is expanded to measure the macroscopic visco-elastic properties of erythrocytes. A suspension of erythrocytes with an incorporated fatty acid spin label was forced to flow through a flat ESR sample cells, and the ESR spectral change caused by the shear flow was utilized to assess the cell deformability. Chemical cross-linking or heat denaturation of membrane proteins to make the cells less deformable without any morphological change was found to reduce the relative spectral difference (delta h/h). This result indicates that the spectral difference is related to the cell deformation that accompanies the orientation of the cells in the shear flow. In addition, the average decay time (tau av) for the spectral difference observed when the flow was abruptly interrupted became shorter with an increase in the degree of cross-linking or heat-denaturation abruptly interrupted became shorter with an increase in the degree of cross-linking or heat-denaturation at 49 degrees C. Since the observed tau av is much shorter than the expected rotational correlation time for the erythrocyte, the decay is attributed to the deformation recovery process. It is demonstrated that the measurements of both delta h/h and tau av by ESR spectroscopy give qualitative information on the viscosity and the elasticity of the cell membrane system.     

Neuroprotective Mechanisms of Melatonin in Hemorrhagic Stroke

Hemorrhagic stroke which consists of subarachnoid hemorrhage and intracerebral hemorrhage is a dominant cause of death and disability worldwide. Although great efforts have been made, the physiological mechanisms of these diseases are not fully understood and effective pharmacological interventions are still lacking. Melatonin (N-acetyl-5-methoxytryptamine), a neurohormone produced by the pineal gland, is a broad-spectrum antioxidant and potent free radical scavenger. More importantly, there is extensive evidence demonstrating that melatonin confers neuroprotective effects in experimental models of hemorrhagic stroke. Multiple molecular mechanisms such as antioxidant, anti-apoptosis, and anti-inflammation, contribute to melatonin-mediated neuroprotection against brain injury after hemorrhagic stroke. This review article aims to summarize current knowledge regarding the beneficial effects of melatonin in experimental models of hemorrhagic stroke and explores the underlying mechanisms. We propose that melatonin is a promising neuroprotective candidate that is worthy of further evaluation for its potential therapeutic applications in hemorrhagic stroke.     

Experimental measurement of dynamic fluid shear stress on the aortic surface of the aortic valve leaflet

Aortic valve (AV) calcification is a highly prevalent disease with serious impact on mortality and morbidity. Although exact causes and mechanisms of AV calcification are unclear, previous studies suggest that mechanical forces play a role. Since calcium deposits occur almost exclusively on the aortic surfaces of AV leaflets, it has been hypothesized that adverse patterns of fluid shear stress on the aortic surface of AV leaflets promote calcification. The current study characterizes AV leaflet aortic surface fluid shear stresses using Laser Doppler velocimetry and an in vitro pulsatile flow loop. The valve model used was a native porcine valve mounted on a suturing ring and preserved using 0.15% glutaraldehyde solution. This valve model was inserted in a mounting chamber with sinus geometries, which is made of clear acrylic to provide optical access for measurements. To understand the effects of hemodynamics on fluid shear stress, shear stress was measured across a range of conditions: varying stroke volumes at the same heart rate and varying heart rates at the same stroke volume. Systolic shear stress magnitude was found to be much higher than diastolic shear stress magnitude due to the stronger flow in the sinuses during systole, reaching up to 20 dyn/cm² at mid-systole. Upon increasing stroke volume, fluid shear stresses increased due to stronger sinus fluid motion. Upon increasing heart rate, fluid shear stresses decreased due to reduced systolic duration that restricted the formation of strong sinus flow. Significant changes in the shear stress waveform were observed at 90 beats/min, most likely due to altered leaflet dynamics at this higher heart rate. Overall, this study represents the most well-resolved shear stress measurements to date across a range of conditions on the aortic side of the AV. The data presented can be used for further investigation to understand AV biological response to shear stresses.     

Comparison of the effects of human erythrocyte ghosts and intact erythrocytes on platelet interactions with subendothelium in flowing blood

We investigated whether ghosts behaved similarly to intact erythrocytes to maintain regular primary hemostasis under flow conditions. To this end we performed perfusion experiments with whole blood in which erythrocytes were replaced by pink ghosts, and platelet interaction with the subendothelial surface of a damaged vessel was morphometrically evaluated. The same objective was sought by means of studies with a platelet function analyzer (PFA-100(TM) instrument). Perfusions performed with control blood reconstituted with intact erythrocytes gave rise to 0.4+/-0.2% contact but not spread platelets, 10.8+/-3.4% adhering and spread platelets, 16.3+/-4.6% platelets in thrombi, with 27.5+/-7.4% of the surface covered. Even though the average diameter of the ghosts was smaller than that of intact erythrocytes (5.3 microm vs. 7.7 microm), the values obtained in perfusions performed with ghosts were similar to those of the erythrocyte controls. Studies performed with the PFA-100(TM) analyzer were consistent with those observed in perfusion studies. The viscosity of control blood was compared with that of blood reconstituted with ghosts. At shear rates lower than 450 s(-1), the viscosity of the ghost samples was higher than that of the controls, but the difference progressively decreased as shear rate increased up to 750 s(-1) (3.61+/-0.15 and 3.71+/-0.17 cP, respectively). In conclusion, the results of our study showed that ghosts behaved similarly to intact erythrocytes in maintaining a normal platelet interaction with digested subendothelium, under conditions of moderate shear rate and constant hematocrit (40%). The rheological activity of ghosts, bodies that are metabolically less active, was sufficient for them to satisfactorily act as substitutes for intact erythrocytes in our system.     

Variation in mortality of ischemic and hemorrhagic strokes in relation to high temperature

Outdoor temperature has been reported to have a significant influence on the seasonal variations of stroke mortality, but few studies have investigated the effect of high temperature on the mortality of ischemic and hemorrhagic strokes. The main study goal was to examine the effect of temperature, particularly high temperature, on ischemic and hemorrhagic strokes. We investigated the association between outdoor temperature and stroke mortality in four metropolitan cities in Korea during 1992–2007. We used time series analysis of the age-adjusted mortality rate for ischemic and hemorrhagic stroke deaths by using generalized additive and generalized linear models, and estimated the percentage change of mortality rate associated with a 1°C increase of mean temperature. The temperature-responses for the hemorrhagic and ischemic stroke mortality differed, particularly in the range of high temperature. The estimated percentage change of ischemic stroke mortality above a threshold temperature was 5.4 % (95 % CI, 3.9–6.9 %) in Seoul, 4.1 % (95 % CI, 1.6–6.6 %) in Incheon, 2.3 % (?0.2 to 5.0 %) in Daegu and 3.6 % (0.7–6.6 %) in Busan, after controlling for daily mean humidity, mean air pressure, day of the week, season, and year. Additional adjustment of air pollution concentrations in the model did not change the effects. Hemorrhagic stroke mortality risk significantly decreased with increasing temperature without a threshold in the four cities after adjusting for confounders. These findings suggest that the mortality of hemorrhagic and ischemic strokes show different patterns in relation to outdoor temperature. High temperature was harmful for ischemic stroke but not for hemorrhagic stroke. The risk of high temperature to ischemic stroke did not differ by age or gender.     

Spin label study of erythrocyte deformability. Ca2+-induced loss of deformability and the effects of stomatocytogenic reagents on the deformability loss in human erythrocytes in shear flow

The Ca2+-induced loss of deformability in human erythrocytes and the recovery of the lost deformability by stomatocytogenic reagents were investigated by means of a new flow electron paramagnetic resonance (EPR) spin label method, which provides information on deformation and orientation characteristics of spin labeled erythrocytes in shear flow. The Ca2+-induced loss of deformability is attributed mainly to the increase in intracellular viscosity resulting from efflux of intracellular potassium ions and water (Gardos effect). Partial recovery of the lost deformability is demonstrated in the presence of stomatocytogenic reagents, such as chlorpromazine, trifluoperazine, W-7, and calmidazolium (R24571). The recovery can not be explained solely by suppression of the Gardos effect due to the reagents. Incorporation of an optimal amount of the reagents into the membrane appears to compensate for the membrane modification due to Ca2+ ions to restore a part of the lost deformability.     

Shear-induced alignment of self-associated hemoglobin in human erythrocytes: small angle neutron scattering studies

Small angle neutron scattering (SANS) was performed on suspensions of actively metabolising human erythrocytes in the constant shear field induced by a Couette cell. The SANS pattern recorded on a two-dimensional detector was a function of the shear rate; at zero shear, the SANS pattern had radial symmetry around the direction of the beam. The radial average of the SANS pattern consisted of a broad intensity maximum superimposed on a decay. The intensity maximum at q = 0.1 Å^-1 was attributed to isotropically oriented self-associated complexes of the tetrameric oxygen transport protein hemoglobin inside the erythrocytes. A flow curve of the cell suspension was used to identify at what shear rate a suspension of uniaxially oriented ellipsoidal cells is produced. The radial symmetry of the SANS patterns persisted until the shear rate was sufficient to produce a suspension of uniaxially oriented ellipsoidal cells. Again, an intensity maximum was present in directions parallel and orthogonal to the shear axis, but this intensity maximum was superimposed upon quite different intensity decays in each direction from that of the primary neutron beam. The angular range of the SANS instrument was limited, however the results from shear-induced structural changes is consistent with a model that involves hemoglobin complexes that are aligned with respect to the plasma membranes of the elongated cells.     

Effect of β-adrenergic stimulation of trout erythrocytes on blood viscosity

1. Both the viscosity of trout blood and its dependence on shear rate are reduced when erythrocyte β-adrenoceptors are stimulated.2. This suggests that catecholamines released during stress may decrease the resistance of nucleated erythrocytes to blood flow.