Abstract
S1-1 Potential application of dynamic contrast enhanced ultrasound in predicting microvascular invasion of hepatocellular carcinoma
*Yi Dong1, Yijie Qiu1, Daohui Yang1, Dan Zuo1, Qi Zhang1, Wen-Ping Wang1, Ernst Michael Jung2
1Zhongshan Hospital, Fudan University, 2Department of Radiology, University Medical Center Regensburg, Germany
S2-1 Vascular toxicity in Cardio-Oncology
*Shohei Moriyama, Michinari Hieda
Department of Hematology, Oncology and Cardiovascular Medicine, Kyushu University Hospital
Cardio-Oncology is a new field focusing on the mechanisms, early detection, and appropriate treatment of cardiovascular toxicity related to cancer itself and cancer therapy. While the development of novel anti-cancer therapy has been improved, prognosis in patients with cancer, cardiotoxicity, as adverse effects of cancer therapy, has been increasing. Venous thromboembolism (VTE) is frequently detected in patients with cancer. Furthermore, the incidence of VTE increases with anti-cancer treatment, especially with new generation anti-cancer agents such as a vascular endothelial growth factor (VEGF) inhibitor. Interestingly, VEGF inhibitor also acts on the micro artery and provokes hypertension. In addition to the above events, anti-cancer drugs could evoke arterial thrombosis, coronary artery vasospasm, and pulmonary artery hypertension as a venous adverse event. In the Cardio-Oncology field, the novel insights and future prospects of vascular toxicity will be highlighted.
S2-2 Catheter treatment of leg arterial atherosclerosis: Endovascular treatment for peripheral arterial disease
Eiji Karashima
Shimonoseki City Hospital
Peripheral artery disease (PAD) is a common circulatory problem characterized by reduced blood flow to the limbs. This may cause symptoms such as leg pain when walking, ischemic rest pain, arterial insufficiency ulcers, and gangrene. The main reason for leg artery narrowing is atherosclerosis. The population of PAD has been increasing due to an aging society and improved diagnosis. Surgical treatment was a mainstream in the treatment of symptomatic PAD. However, endovascular treatment (EVT) using catheter techniques has been recently developed and widely accepted as an effective therapeutic option for symptomatic PAD patients. To improve clinical results of EVT, devices such as wires, balloons, and stents have been employed. However, treating long-term occluded arterial lesions is one of the challenging situations for EVT. Such EVT cases showing long-occluded arterial lesions are presented herein. Compared to coronary artery disease, PAD is not as well-studied. Indeed, PAD is hardly recognized as a reason for arterial insufficiency induced ulcers or gangrene. In addition, when legs were diagnosed with PAD, amputation was often difficult to avoid. Early diagnosis of PAD is indeed one of the best ways to improve the outcome of these patients. In this presentation, current situations of PAD and EVT are reviewed.
S2-3 Advances in diagnosis and treatment of pulmonary hypertension
Kohtaro Abe
Department of Cardiovascular Medicine, Kyushu University Hospital
Pulmonary hypertension is defined as a resting mean pulmonary artery pressure of 20 mmHg or above. Pulmonary arterial hypertension (PAH, group 1) is characterized by obstructed pulmonary arteries or arterioles arising from vasoconstriction and vascular remodeling. Progress has been made in identifying the causes of PAH and approving new drug therapies. Obstructive pulmonary vascular remodelling in PAH increases right ventricular afterload. In some patients, maladaptive changes in the right ventricle, including ischemia and fibrosis, reduce right ventricular function and cause right ventricular failure. Patients with PAH have dyspnea, reduced exercise capacity, exertional syncope, and premature death from right ventricular failure. PAH targeted therapies (prostaglandins, phosphodiesterase-5 inhibitors, endothelin receptor antagonists, and soluble guanylate cyclase stimulators used alone or in combination), improve functional capacity and hemodynamics and reduce the occurrence of acute heart failure. The epidemiology, pathogenesis, diagnosis, and treatment of PAH are presented.
S2-4 Myocardial pathological changes in patients with epilepsy and psychiatric disorders
*Marin Takaso, Misa Tojo, Masahito Hitosugi
Department of Legal Medicine, Shiga University of Medical Science
The incidence of sudden death in epilepsy patients is reported to be 24 times higher than that in normal subjects. In addition, schizophrenia patients are said to have a short life expectancy and a high number of sudden cardiac deaths. We examined the pathological changes of the heart and myocardium in patients with epilepsy (Epi) and psychiatric disorders (Psy). From forensic autopsies performed at Shiga University of Medical Science from 2014 to 2020, cases of sudden cardiac death (SCD) and cases of exogenous death (ED) with a history of Epi and Psy were extracted. As control groups, those without a history of Epi or Psy were extracted. In each case, body height and weight, heart weight, left ventricular thickness, right ventricular thickness were examined by autopsy records. Microscopically, the thickness of cardiomyocyte, nuclear diameter, and nuclear density were examined. Eighty-five subjects were studied (65 male, 20 female) with a mean decedent age of 43.4. Fourteen had a history of Epi (5 of SCD, 9 of ED), 23 had a history of Psy (9 of SCD, 14 of ED), and 48 were control (27 of SCD, 21 of ED). Mean heart weight and the thickness of left heart ventricle were significantly smaller in SCDs with Epi than the control. Mean nuclear diameter of the cardiomyocyte was smaller in SCDs with Epi or Psy than the control. The heart weight, thicknesses of the left ventricle, and nuclear diameter were smaller in Epi or Psy patients suffering from SCD than control. These morphological changes may affect the occurrence of SCD.
S3-1 Calcium signaling in red cells induced by mechanical stress and flow
Lars Kaestner
Saarland University
Within recent years it has been shown that healthy mammalian red cells, albeit missing a translational machinery, have a tremendous and sophisticated signaling capacity. The second messenger calcium plays an important role in the mechanism of spatio-temporal signaling in red cells. Calcium plays a pivotal role in translating mechanical stress and flow into cellular responses. This holds true for normal physiological responses as well as for pathophysiological situations. Established and recent findings are presented on how mechanical stress and flow drives calcium signaling in red cells, which molecular players are involved and what the cellular consequences are.
S3-2 Shear conditioning attenuates the effects of superoxide in red blood cells: Role of generation and signaling of nitrogen species in different cell subpopulations
*Marijke Grau1, Lennart Kuck2, Thomas Dietz1, Michael J Simmonds2
1German Sport University Cologne, Institute of Cardiovascular Research and Sports Medicine, 2Griffith University Mechanobiology Research Laboratory
Red blood cell (RBC) deformability appears to be enhanced by physiological shear stress, although classic studies demonstrate cell mechanics are negatively impacted by free radical exposure. It remains unknown whether determinants of RBC deformability – e.g. the activation of RBC-nitric oxide synthase (NOS) and resultant intracellular NO generation – are sensitive to free radicals, which might explain, in part, the negative effects of these species. Further, whether all RBC subpopulations of varying density and age are equivalently affected remains unresolved. A series of experiments examined whether intracellular superoxide (O2−) generation by phenazine methosulfate (PMS), and/or specific mechanical stress, altered RBC deformability and the PI3K/Akt kinase and RBC-NOS signaling in RBC that were differentiated into “young” and “old” subpopulations. Generated O2− decreased activation of both PI3/Akt kinase and RBC-NOS. This observation was accompanied by decreased RBC deformability. Shear conditioning significantly improved cellular deformability, even in RBCs previously exposed to O2−. Old RBCs exhibited lower RBC deformability compared to young RBCs, although both sub-populations were negatively impacted by O2− exposure. Shear conditioning did not affect intracellular reactive species, although intriguingly, it increased RBC deformability and reduced RBC-NOS activation in old RBCs. The co-application of O2− and shear exposure improved cellular deformability in older cells previously exposed to O2−, but not in younger cells. It thus appears that shear stress might in part counteract the deleterious effects of reactive species. But differences in the complex interplay of mechanical stress and redox metabolism in circulating RBCs were found between different RBC subpopulations.
S3-3 Signaling in erythroid cells: Role in physiology and contribution to pathological manifestations
Wassim El Nemer
EFS
Human red blood cells (RBCs) are enucleated biconcave discs produced in the bone marrow through a developmental process called erythropoiesis. The main role of RBCs in the circulation is to ensure gas exchange, oxygenating the tissues and carrying carbon dioxide out of the organism. Under normal physiological conditions, RBCs spend an average of 120 days in the circulation, but this is dramatically shortened under pathological conditions such as hemolytic anemia. Erythropoiesis is a finely controlled differentiation process that is divided into early and terminal phases in which hematopoietic stem cells undergo several cell divisions and differentiate into orthochromatic erythroblasts that enucleate giving rise to reticulocytes. Reticulocytes are young immature red cells with a convoluted shape that exit the bone marrow and mature in the circulation undergoing several structural and functional changes. Cell signaling plays an important role during erythroid differentiation, regulating critical processes, such as cellular interactions of differentiating erythroblasts within the erythroid niche, nucleus and organelle polarization and expulsion, as well as reticulocyte maturation. This presentation gives an overview of the signaling pathways involved in erythroid differentiation and reticulocyte maturation and the impact of their dysregulation on circulating red cells under pathological conditions.
S3-4 Impact of oxidative stress and decreased NO bioavailability on eryptosis and red blood cell microparticles in SCA: Consequences on endothelial cells and vascular function
*Elie Nader1,2, Marc Romana3,4, Nicolas Guillot1,2, Romain Fort5, Emeric Stauffer1,2,6, Nathalie Lemonne7, Yohann Garnier3,4, Sarah Skinner1,2, Maryse Etienne-Julan7, Mélanie Robert1,2,8, Alexandra Gauthier1,2,9, Giovanna Cannas5, Sophie Antoine-Jonville3, Benoit Tressières10, Marie-Dominique Hardy-Dessources3,4, Yves Bertrand9, Céline Renoux1,2,11, Philippe Joly1,2,11, Marijke Grau12, Philippe Connes1,2
1Laboratoire Interuniversitaire de Biologie de la Motricité (LIBM) EA7424, Team “Vascular Biology and Red Blood Cell”, Université Claude Bernard Lyon, Université de Lyon, Lyon, France, 2Laboratoire d’Excellence du Globule Rouge (Labex GR-Ex), PRES Sorbonne, Paris, France, 3Université des Antilles, Pointe-à-Pitre, France, 4Université de Paris, Paris, France, 5Département de Médecine Interne, Hôpital Edouard Herriot, Hospices Civils de Lyon, Lyon, France, 6Centre de Médecine du Sommeil et des Maladies Respiratoires, Hospices Civils de Lyon, Hôpital de la Croix Rousse, Lyon, France, 7Unité Transversale de la Drépanocytose, Hôpital de Pointe-à-Pitre, Hôpital Ricou, Guadeloupe, France, 8Erytech Pharma, Lyon, France, 9Institut d’Hématologie et d’Oncologie Pédiatrique, Hospices Civils de Lyon, Lyon, France, 10Centre Investigation Clinique Antilles Guyane, 1424 Inserm, Academic Hospital of Pointe-à-Pitre, Guadeloupe, France, 11Laboratoire de Biochimie et de Biologie Moléculaire, UF de Biochimie des Pathologies érythrocytaires, Centre de Biologie et de Pathologie Est, Hospices Civils de Lyon, Lyon, France, 12Molecular and Cellular Sport Medicine, Deutsche Sporthochschule Köln, Köln, Germany
Sickle Cell Anemia (SCA) is caused by a mutation in the beta-globin gene that leads the production of abnormal hemoglobin called hemoglobin S (HbS). HbS can polymerize in deoxygenated conditions, causing red blood cell (RBC) sickling. Patients with SCA suffer from chronic hemolytic anemia and repeated vasoocclusive crises. Accumulating evidence suggests that chronic hemolysis, enhanced oxidative stress and decreased bioavailability of nitric oxide (NO) are at the origin of the vascular dysfunction in SCA. Oxidative stress and decreased NO bio-availability have been shown, in healthy RBCs to promote eryptosis. RBC membrane blebbing during eryptosis could lead to the release of microparticles (MPs) into the blood circulation which could promote vascular dysfunction through the activation of TLR4 by heme. However, the role of oxidative stress and NO bioavailability in eryptosis and MPs release in SCA, as well as the impact of RBC-MPs in vascular dysfunction in SCA are unclear.
Markers of eryptosis and oxidative stress, plasma RBC-MPs concentration and arterial stiffness were compared between SCA and healthy (AA) individuals. In-vitro experiments were performed to test: (1) the effects of oxidative stress and NO on eryptosis, RBC deformability and RBC-MP genesis; (2) the effects of RBC-MPs, directly isolated from SCA and AA blood, on human aortic endothelial cell (HAEC) inflammatory phenotype and TLR4 pathway. Eryptosis, RBC-MPs, oxidative stress and arterial stiffness were increased in SCA. NAC increased RBC deformability and decreased eryptosis and RBC-MPs release, while cumene did the opposite. SNP increased RBC deformability and limited eryptosis, but had no effect on RBC-MPs. Arterial stiffness was correlated with RBC-MPs concentration in SCA. RBC-MPs isolated directly from SCA blood increased adhesion molecules expression (I-CAM1 and E-Selectin) and the production of cytokines by HAEC compared to those isolated from AA blood. TLR4 inhibition with TAK-242 alleviated these effects. Increased oxidative stress in SCA could promote eryptosis and the release of RBC-MPs that are potentially involved in macrovascular dysfunction in SCA. RBC-MPs could exert their deleterious effects on endothelial cells by activating TLR4 through the heme they carry.
S3-5 Phosphoproteomic changes in red blood cell membranes by Adenylyl cyclase/Protein kinase A signaling pathway and their roles on the mechanical stress responses of red blood cells
*Elif Ugurel1,2, Neslihan Cilek1,2, Evrim Goksel1,2, Ozlem Yalcin1,2
1Koc University School of Medicine Department of Physiology, 2Koc University Research Center for Translational Medicine
The mechanical stress responses of red blood cells (RBC) is modulated by the phosphorylation status of the cytoskeletal proteins that regulates the interactions of integral transmembrane complexes. Proteomic studies revealed that receptor-related signaling molecules and regulatory proteins involved in signaling cascades are present in RBCs. In this study, we investigated the roles of Adenylyl cyclase (AC)/Protein kinase A (PKA) signaling mechanism in the modulation of RBC mechanical stress responses. We implemented the inhibitors of AC (SQ22536), PKA (H89) and phosphodiesterase (PDE) (Pentoxifylline) to whole blood samples. We applied a physiologically relevant shear stress (SS) at 5 Pa for 300 s by a capillary tubing system and evaluated RBC deformability by LORRCA MaxSis. The inhibition of signaling molecules significantly deteriorated mechanical stress responses of RBCs (p < 0.05). We also examined the phosphorylative changes of RBC membrane proteins by SS inducement and pathway inhibition. Tyrosine and serine phosphorylation of RBC membrane proteins slightly increased by the SS inducement. Tyrosine phosphorylation significantly increased by the inhibition of AC and PDE (p < 0.05). Serine phosphorylation significantly decreased by the inhibition of PDE (p < 0.05). AC is the core element of this signaling pathway and PDE works as a negative feed-back mechanism which could all have potential roles in the SS-induced improvement of deformability. The mechanical stress responses of RBCs could be regulated by the alterations in the phosphorylative status of membrane proteins through the signaling molecules of AC/PKA pathway. The modulation of this pathway may yield new clinical and therapeutic approaches to circulatory disorders.
S4-1 Microfluidic assays to investigate the role of red blood cell-derived extracellular vesicles in sickle cell disease
*Ran An An, Umut Gurkan
Case Western Reserve University
Sickle cell disease (SCD) is associated with hemolysis, vaso-occlusion, and endothelial dysfunction, together contributing to acute pain and chronic and cumulative organ damage. Extracellular vesicles (EVs), spontaneously or post-traumatically released by red and white blood cells, are vehicles for exchange of biological material between cells, and may serve as surrogate markers for the activation state of the parent cell. Upon deoxygenation, sickle hemoglobin (HbS) containing red blood cells (RBCs) experience elevated intravascular hemolysis, release damage associated molecular patterns including cell-free hemoglobin, heme, and generate EVs (REVs) during intravascular hemolysis. REVs exhibit phosphatidylserine expression, transport heme, can promote coagulation and microvascular inflammation. In this work, we utilized microfluidic systems to perform in-vitro studies to assess endothelial activation following exposure to RBC-derived EVs in SCD. Here, we analyzed the short-term impact of exposure to SS REV on pulmonary microvascular endothelial cell activation. Compared with AA REVs, SS REVs promoted microvascular endothelial cell activation at 2 hours indicated by increased vWF expression. Under microfluidic conditions, we found abnormal RBC adhesion to REV-exposed endothelium only in SRBCs on to SREV-activated endothelium. The adhesion profile varied from subject to subject, and associated with markers of hemolysis, as well as concomitant clinical diagnosis of deep vein thrombosis. Our results emphasize the critical contribution made by REVs to the pathophysiology of SCD, in short term, by triggering EC activation and abnormal RBC adhesion. These findings may help to better understand short term pathophysiological mechanism of SCD and thereby the development of new treatment strategies.
S4-2 Hemorheology and pathophysiology of COVID-19 induced thrombosis predicted by Vein-Chip
Navaneeth Krishna Rajeeva Pandian, *Abhishek Jain
Texas A&M University
There is a serious limitation of experimental models that can improve our limited knowledge of the mechanisms that regulate endotheliopathy and venous thrombosis (VT) clinically observed frequently amongst the most severe COVID-19 patients. Also, while observation and study of VT in humans are difficult due to the deep-lying nature of the deep veins in which VT develops, lab animal models do not include the venous valves, which are the sites of unique hemodynamics and thrombus development in humans. We develop a Vein-chip microfluidic platform that includes venous valve architecture, endothelial cells (ECs), and whole blood flow, which can include the three factors of Virchow’s triad – endothelial inflammation, stasis of blood flow, and coagulable nature of blood. Our in silico and in vitro observations with Vein-Chip reveal that incompetent valves and thrombosis changes the blood flow pattern in and around the venous valves. We show that healthy endothelium at the venous valve cusps adapt to the complex flow patterns and have an anti-thrombotic phenotype compared to the venous lumen. But exposure of the lumen to living and replicating SARS-CoV-2 virus and inflammatory cytokines found in COVID-19 patient samples inflames the lumen and the valve endothelium becomes prothrombotic. Interestingly, when we directed our investigation to analyze the ACE2 expression on these cells, as ACE2 is the functional receptor of the SARS-CoV-2 virus, we found that ACE2 expression was poor under a static culture, but increased dramatically when venous ECs were exposed to shear stress within the vein-chip. This data supports our hypothesis that ACE2 expression (and therefore, SARS-CoV2 entry into the endothelium) is dependent on venous hemodynamics and the Vein-Chip model is a highly dissectible platform that will help us to unravel the molecular mechanisms that lead to VT and its treatment strategies for COVID-19 and beyond.
S5-1 The importance of blood rheology in left ventricular assist device therapy
*Mohammed Chowdhury1, Valmiki Maharaj2, Arianne Agdamag2, Blair Edmiston3, Bellony Nzemenoh3, Victoria Charpentier4, Tamas Alexy2
1North Central Heart, Sioux Falls, SD, USA, 2Department of Medicine, Division of Cardiology, University of Minnesota, MN, USA, 3Department of Medicine, University of Minnesota, MN, USA, 4University of Minnesota Medical School, Minneapolis, MN, USA
Heart failure remains an epidemic of the 21st century with almost 1 million new cases diagnosed annually in the US alone. Alongside this, the number of patients reaching end stage disease (Stage D) continues to rise. As the number of heart transplants remains limited due to organ availability, an increasing number of patients are supported with a continuous flow Left Ventricular Assist Device (LVAD). Thorough understanding of computational fluid dynamic principles combined with technological advancements have revolutionized LVAD design and led to the development of smaller and increasingly more hemocompatible pumps. These ultimately translate to improved patient outcomes, including survival and quality of life. The newest pump design is based on centrifugal technology, utilizes a fully magnetically levitated impeller, and provides hemodynamic support for up to 10 liters per minute. Larger gaps within the housing as well as lack of friction improve efficiency and reduce complications. This presentation aims to review the evolution of continuous flow LVADs from the hemorheological viewpoint and how it contributes to significantly improved patient outcomes.
S5-2 Hemo-compatibility related adverse events with left ventricular assist device (LVAD) support: past, present, and future
*Valmiki Maharaj1, Mohammed Chowdhury2, Arianne Agdamag1, Blair Edmiston3, Bellony Nzemenoh3, Victoria Charpentier4, Tamas Alexy1
1Department of Medicine, Division of Cardiology, University of Minnesota, MN, USA, 2North Central Heart, Sioux Falls, SD, USA, 3Department of Medicine, University of Minnesota, MN, USA, 4University of Minnesota Medical School, Minneapolis, MN, USA
Hemo-compatibility is the study of the thrombotic response of blood when in contact with a foreign material or device. Despite advances in design and technology with the use of titanium alloys, hemocompatibility related adverse events remain frequent in patients requiring left ventricular assist device (LVAD) support. These complications include hemolysis, gastrointestinal bleeding, pump thrombosis, ischemic and hemorrhagic stroke. Although the majority are non-fatal, these adverse events can have a significant negative effect on quality of life. Therefore, reducing hemo-compatibility related complications has been one of the driving forces of new pump development. We will provide an overview of relevant design changes over several LVAD generations and summarize the newest clinical trial data, including our experience, with the most recent devices available on the market. We will also pay special attention reviewing how design changes at the bench led to a profound improvement in patient outcomes and overall clinical experience.
S5-3 Analysis of the HeartMate 3 pump characteristics under continuous and pulsatile flow operation – An in vitro study
*Jo Pauls1,2, Nicole Bartnikowski2,3, E-Peng Seah2, Clayton Semenzin2, Martin Mapley2
1Griffith University – School of Engineering and Built Environment, 2Innovative Cardiovascular Engineering and Technology Laboratory, Critical Care Research Group, The Prince Charles Hospital, 3Queensland University of Technology – Science and Engineering Faculty
S5-4 Re-evaluation of blood trauma from a sublethal perspective
Michael J Simmonds1,2
1Menzies Health Institute Queensland, 2Griffith University, Australia
S6-1 Micro-tomographic visualization of tissue rheological properties by mechanical stimulation using optical coherence tomography
*Daisuke Furukawa1, Souichi Saeki2
1Akita Prefectural University, Faculty of Systems Science and Technology, 2Meijo University, Graduate School of Science and Technology
The “wrinkles” and “sagging” of the skin are related to the metabolic function of the skin. Related to the decrease in metabolic function, the collagen and elastin fibers inside the dermis decrease and the function of skin viscoelasticity is impaired. The purpose of this study is to detect the viscoelastic properties of the skin by mechanical stimulation using optical coherence tomography (OCT). In this paper, tomographic images were detected from creep recovery experiments using the negative pressure suction method, and the strain rate was micro-transformed into a tomographic visualization based on the image correlation method. The strain rate was detected using the cheek as the measurement target, and the strain rate in the compressive direction in the boundary layer between the epidermis and dermis was visualized by microtomography. This suggests that an integrated evaluation of the mechanical properties and circulatory function of the skin may lead to effective skin care and anti-aging.
S6-2 Three-dimensional detection of hemodynamic changes in skin microcirculation by optical coherence tomography-angiography
Yu Nakamichi
Sanyo-Onoda City University
Skin has various functions to maintain our beauty and health conditions. The skin function has been found to be related to hemodynamic changes in the skin microcirculation. For examples, Sawane and colleagues showed that skin aging is caused by destabilization in structures of blood vessel and lymphatic (J Soc Chem Jpn 46(3), 188–196, 2012), and Nakagomi and colleagues suggested that responses in skin microcirculation induced by ultraviolet B light irradiation are related to formations of erythema (Trans JSMBE 48(1), 42–49, 2010). In this way, detection of hemodynamic changes in the skin microcirculation provides insights into various skin functions. In the skin, which consists of dermis, epidermis, and subcutaneous tissue, however, there is no established technique to three-dimensionally evaluate hemodynamic changes in the microcirculation. In this study, a technique to three-dimensionally detect hemodynamic changes in the skin microcirculation is proposed by means of optical coherence tomography-angiography (OCTA). OCTA is a functional extension of optical coherence tomography (OCT), which noninvasively visualizes biological tissues with micro-scale spatial resolution. OCTA can detect three-dimensional vascular networks by quantifying temporal variation in OCT signals that is proportional to blood flow velocities. Therefore, the proposed technique is capable of three-dimensionally evaluating hemodynamic changes in vessel structures and blood flow velocities. In this presentation, results are shown for validation experiments where hemodynamic changes induced by an alcohol stimulus to the skin were detected with the proposed technique, and an investigation conducted of the utility of the proposed technique.
S6-3 Investigation the extensional effects on the viscosity distribution of bile in the cystic duct
*Ngoc Minh Nguyen1, Hiromichi Obara2
1Department of Mechanical Engineering, Thuyloi University, 175 Tay Son, Dong Da, Ha Noi, Vietnam, 2Department of Mechanical Systems Engineering, Tokyo Metropolitan University, 1-1 Minami Osawa, Hachioji, Tokyo, Japan
Bile flow is thought to play an important role in the pathophysiological genesis of gallstone formation. In this paper, we considered the second and the third invariant in the flow equation governing generalized Newtonian and viscoelastic fluids. It is found that the stretching properties of bile lead to enhancement of its viscosity and resistance to flow. This promotes increased viscosity of bile at or near the spiral valves of Heister that contributes to gallstone formation and sludge formation in the cystic duct.
S6-4 Quantitative evaluation of flowing blood with the electrical parameters based on the Hanai mixture equation
*Yusuke Nakajima1, Daisuke Kawashima1, Ryubu Shoji1, Katsuhiro Matsuura2, Masahiro Takei1
1Division of Mechanical Engineering, Chiba University, 2Department of Veterinary Surgery, Tokyo University of Agriculture and Technology
S6-5 Development of butterfly type artificial atrioventricular valve with anisotropic valvular cusps by using collagenous connective tissue membrane “Biosheet®” similar to in vivo tissue architecture
*Yota Sekido1, Yasuhide Nakayama2, Tsutomu Tajikawa3
1Kansai University, Graduate School of Science and Engineering, 2Biotube Co., Ltd, 3Kansai University, Faculty of Engineering Science
We have been developing heart valve-like tissue (Biovalve®) similar to in vivo tissue architecture (iBTA®). In our previous study, we have fabricated a novel prosthetic valve by using a tissue membrane (Biosheet®) by iBTA as valve cusps with a valvular root stent of which the shape was similar to the annuloplasty ring of the mitral valve. The purpose of this study was to develop a compatible prosthetic atrioventricular valve that is both a simple structure and easily implanted. To achieve the higher valve performance, the valve cusps with mechanical anisotropy were prescribed and its effect was investigated. Because it takes 1–2 months to obtain Biosheets from doner animals, a model experiment employed a thinner polyurethane sheet instead of a Biosheet with the same mechanical properties. Experiments were conducted with varied cusp mechanical parameters. Valve performance was measured using our fabricated in vitro flow simulator under physiological conditions similar to those in humans. The anisotropic valve cusps were fabricated from a combination of polyurethane sheets and shape memory alloy wires. For experimental parameters, the direction of anisotropy and the amount of valve cusp protrusion over the stent were varied. In addition, valve cusps without anisotropy were fabricated to compare the valve performance with those with anisotropy. Based on ISO 5840, the temporal waveforms of left atrial pressure, left ventricular pressure, and mitral flow rate were measured, and the regurgitation rate (Rf), mean pressure drop (Δp), and effective orifice area (EOA) were calculated to evaluate the performance of the valve.
S7-1 Investigation of hemorheological and hematological properties of blood in stented mice
*Despoina Kokkinidou1,2, Konstantinos Kapnisis2, Efstathios Kaliviotis1, Andreas Anayiotos2
1Biorheology Laboratory, Dept. of Mechanical Engineering and Material Science and Engineering, Cyprus University of Technology, Cyprus, 2BioLISYS Laboratory, Dept. of Mechanical Engineering and Material Science and Engineering, Cyprus University of Technology, Cyprus
S7-2 Estimation of whole blood coagulation using image processing techniques
*Marinos Louka1, Antonios Inglezakis2, Constantinos Loizou2, Savvas Psarelis3, Elena Nikiphorou4, Efstathios Kaliviotis1
1Cyprus University of Technology, 2EMBIO Diagnostics Ltd, Nicosia, Cyprus, 3Ministry of Health Cyprus, Nicosia, Cyprus, 4King’s College London, King’s College Hospital, London, UK
Blood coagulation is an essential mechanism for preventing the uncontrollable loss of blood. In certain cases however inherent, or even iatrogenic malfunction of this mechanism occurs, and medical intervention with tight monitoring of blood coagulation is required. In this work whole blood coagulation was studied by monitoring the structural changes occurring in the fluid, as the clotting progresses in time. Finger-prick blood samples where acquired from healthy volunteers, placed directly on purpose made testing chips, without the use of reagents to trigger the coagulation process. The testing chips were constructed using glass slides and adhesive tapes. Images were captured for a period of 35 minutes via a microscopy-camera setup. Specialized algorithms were applied for the processing and analysis of the images, in order to extract relevant information. Color, grey, and binary conversions of the images were analyzed by both a statistical and a Boolean logic approach. The adopted methods and techniques in this study allowed to identify the erythrocytes/aggregates, the plasma and clot regions, and their changes as coagulation progresses in the sample. Coagulation was characterized by developing appropriate indices. The results, illustrate a good sensitivity of the coagulation indices, and indicate a significant potential for a robust characterization of blood coagulation in an inexpensive set-up. The extracted coagulation indices are also compared to data produced by using hemorheological and mechanical methods for the characterization of blood coagulation.
S7-3 Capillary blood flow on a chip: Influence of hemorheological factors
*Dimitris Pasias Pasias, Andreas Passos, Georgios Constantinides, Loukas Koutsokeras, Stavroula Balabani, Efstathios Kaliviotis
Cyprus University of Technology
Capillary driven blood flows have gained popularity for Lab-on-Chip applications, due to the simplicity and low cost of their set-up. A blood drop was placed in the entrance of the chip and transported to various locations for analytical purposes. In this study various hemorheological factors (red blood cell (RBC) deformability, aggregation and concentration) were altered in blood samples in order to investigate their effects on the capillary driven flow of blood, in a hydrophilic microchannel. RBC aggregation was induced by utilizing Dextran suspensions, RBC deformability was altered by heat-treating the samples, and two RBC concentrations were produced at 40 and 45%. The velocity of blood in the channel was derived by tracing the meniscus of the flowing samples, using image-based tracking techniques. The velocity of the meniscus was correlated with the mean relative viscosity of the samples, as well as with RBC aggregation and deformability indices. The results confirmed the negative impact of the increasing viscosity and reduced RBC deformability in the velocity behavior of the samples. RBC aggregation intensity also had a negative influence in the meniscus velocity, however, this was not the case for flow of fluids with reduced deformability, which appeared to have benefited from the condition. The effective shear rate derived for the duration of the flow, appeared very intense at the early stages of the flow and persisted sufficiently strong until the flow reached the end of the channel.
S7-4 Erythrocyte sedimentation rate measurements in a high aspect ratio microfluidic channel
*Andreas Passos1, Antonis Nikolaidis1, Charalampos Vryonidis1, Konstantinos Loizou2, Antonis Inglezakis2, Efstathios Kaliviotis1
1Dept. of Mechanical Engineering and Material Science and Engineering, Cyprus University of Technology, Cyprus, 2EMBIO Diagnostics Ltd, Nicosia, Cyprus
Erythrocyte sedimentation rate (ESR) is a useful tool for assisting in the monitoring of various pathological conditions and in predicting responsiveness to treatment. The traditional ESR testing methods require blood volume larger than 1 ml which in some cases is considered as non-efficient. In the present work a novel technique is presented that enables capillary driven microfluidics and image processing algorithms to measure ESR. Blood samples of <25 ul volume at physiological haematocrits (30–50%) were introduced in surface-tension driven, superhydrophilic, high-aspect ratio microchannels (channel dimensions: ∼100 um height, ∼3 mm mean width and ∼25 mm length). After the application of the sample, the channel was re-oriented so that the direction of the width was aligned with gravity. The red blood cell (RBC) sedimentation layer was captured by acquiring images for 30 min and 0.2 Hz sampling rate using a USB camera. The ESR was estimated using automated image processing techniques measuring the RBC layer width in time. The ESR of the samples were also measured and compared with commercial ESR measuring instruments. The results showed that the current approach can provide ESR information from a microscale setup, and that a correlation exists with ESR measurements performed in standard industrial devices. The technique used in the present study produces high resolution ESR results, well correlated with the commercial instruments and conventional measuring techniques, providing a quick and inexpensive tool with potential to be self-administered by patients for home use.
S7-5 Influence of hemorheological parameters on the local velocity characteristics of blood in a super-hydrophilic channel
*Efstathios Kaliviotis1, Dimitris Pasias1, Andreas Passos1, Loukas Koutsokeras1, Georgios Constantinides1, Stavroula Balabani2
1Cyprus University of Technology, 2University College London
The local characteristics of the flow of blood are of fundamental and physiological interest, as they are essential for a better understanding of the fluid mechanics and dynamics as well as for clarifying various physiological/pathological processes. In this work the local flow of blood in a surface-tension driven, super-hydrophilic configuration, is studied in order to assess the influence of specific hemorheological factors. The microchannels were constructed in-house using surface-treated glass slides, and xurography techniques. Hemorheological parameters included erythrocyte aggregation (whole blood and Dextran suspensions), erythrocyte deformability (altered by heat-treatment) and concentration. The local velocity field in the channel was derived by micro-PIV techniques. Images were acquired from a camera/microscopy set-up and processed by specialized algorithms. Velocity profiles were calculated, from the produced vector fields, and characterized by a bluntness parameter and mean velocity values. The results showed that the mean velocity of blood in the channel is indeed affected by the increasing sample viscosity and the reduced erythrocyte deformability as expected. The elevated RBC aggregation also had a negative influence in the mean velocity of the samples, however it appeared to influence more the shape of velocity profiles (blunter) in the normal compared to the heat-treated samples. The local and bulk shearing conditions in the channel appeared to be elevated reaching magnitudes above 200 s−1 near the walls, and indicating that erythrocyte aggregation was kept at low levels.
S8-1 Role of Piezo1 in red blood cell sickling
*Elie Nader1,2,3, Aline Hatem4, Robin Bertot1, Philippe Joly1,2,3, Camille Boisson1,2,3, Guillaume Bouyer4, Nicolas Guillot1,2,3, Alexandra Gauthier1,2,3, Solène Poutrel1,2,3, Céline Renoux1,2,3, Nicola Conran5, Flavia Costa5, Yves Bertrand3, Stéphane Égée4, Philippe Connes1,2,3
1Laboratoire Interuniversitaire de Biologie de la Motricité (LIBM) EA7424, Team Vascular Biology and Red Blood Cell, Université Claude Bernard Lyon 1, Université de Lyon, Lyon, France, 2Laboratoire d’Excellence du Globule Rouge (Labex GR-Ex), PRES Sorbonne, Paris, France, 3Reference Center in Sickle cell disease, Thalassemia and rare red blood cell and erythropoiesis diseases, Hospices Civils de Lyon, Lyon, France, 4UMR 8227 CNRS-Sorbonne Université, Station Biologique, Place Georges Teissier, BP 74, 29682 Roscoff Cedex, France, 5Hematology Center, University of Campinas – UNICAMP, Cidade Universitária, Campinas-SP, Brazil
S8-2 Shear induced red blood cell nitric oxide production is increased in sickle cell disease
*Jon A Detterich1,2, Silvie Suriany1, Honglei Liu1, Pinar Ulker3, G Esteban Fernandez1, Matthew Borzage1, Rosalinda Wenby2, Herbert J Meiselman2, Henry J Forman4, Thomas D Coates1
1Children’s Hospital of Los Angeles, 2Keck School of Medicine, University of Southern California, 3Department of Physiology, Akdeniz University, 4Department of Gerontology, University of Southern California
Sickle cell anemia (SCA) is a beta-globin gene mutation resulting in decreased red blood cell (RBC) deformability due to polymerization of deoxygenated hemoglobin. This leads to abnormal mechanical properties of RBC that causes them to lyse under physiologic conditions. Prior work has demonstrated that NO∙ influences normal RBC hydration and deformability. NO∙ is produced at a basal rate that increases under shear stress. Nevertheless, the effect of hemoglobin S on RBC nitric oxide production is unknown. We aimed to assess the basal and shear-mediated production of NO∙ in RBC from SCA patients and control (CTRL) subjects. RBCs loaded with a fluorescent NO∙ detector, DAF-FM (4-Amino-5methylamino-2 ′ ,7 ′ -difluorofluorescein diacetate), were imaged in microflow channels over 30-min without shear stress, followed by a 30-min period under 0.5 Pa shear stress. We utilized a nonspecific nitric oxide synthase (NOS) blockade and carbon monoxide (CO) saturation of hemoglobin to assess the contribution of NOS and hemoglobin, respectively, to NO∙ production. Quantification of DAFFM fluorescence intensity in individual RBC showed an increase in NO∙ in SCA RBC at the start of the basal period; however, both SCA and CTRL RBC increased NO∙ by a similar quantity under shear. The discoid CTRL and SCA RBC elongated in the direction of flow. A subpopulation of sickle-shaped RBC exhibited lower basal NO∙ production compared to discoid RBC from SCA, and they became more circular while under shear. Also, there were bright inclusions present that Both CO and NOS inhibition caused a decrease in basal NO∙ production. Shear-mediated NO∙ production was decreased by CO in all RBC but was decreased by NOS blockade only in SCA. In conclusion, total NO∙ production is increased and shear mediated NO∙ production is preserved in SCA RBC in a NOS-dependent manner. Sickle shaped RBC with inclusions have higher NO∙ production and they become more circular rather than elongated with shear.
S8-3 Contribution of red blood cells to pulmonary arterial hypertension pathogenesis: NOS activity and vessel responses
*Pinar Ulker1, Ibrahim Basarici2, Nur Özen1, Ece Kilavuz1, Fatih Kisak1, Filiz Basrali1, Nazmi Yaras3, Sadi Koksoy4, Mukadder Levent Celik5, Leyla Abueid1, Ahmet Yildirim1
1Department of Physiology, Medical Faculty, Akdeniz University, Antalya, Turkey, 2Department of Cardiology, Medical Faculty, Akdeniz University, Antalya, Turkey, 3Department of Biophysics, Medical Faculty, AkdenizUniversity, Antalya, Turkey, 4Department of Medical Microbiology, Medical Faculty, AkdenizUniversity, Antalya, Turkey, 5Department of Internal Medicine, University of Health Sciences Antalya Training and Research Hospital, Antalya, Turkey
S8-4 Impaired mechanotransduction in diamide-treated erythrocytes
*Lennart Kuck1, Jason N. Peart2, Michael J. Simmonds1
1Biorheology Research Laboratory, Menzies Health Institute, Griffith University Gold Coast, Queensland, Australia, 2School of Medical Science, Griffith University Gold Coast, Queensland, Australia
S9-1 Deformability measurement of RBCs flowing in capillary channels using a coflowing channels-based pressure sensor
*Yang Jun Kang1, Sami Serhrouchni2, Anna Bogdanova2, Sung-Sik Lee3
1Chosun University, 2University of Zürich, 3ETH Zürich
In this study, a simple method is suggested for measuring RBC deformability in capillary channels in which blood is supplied with a constant pressure source. A microfluidic device was composed of capillary channels for inducing RBC clogging, and a coflowing channel for measuring pressure. A reference fluid was supplied at a constant flow rate. Both fluids (blood, and reference fluid) flowed and formed parallel streams. The interface in the coflowing streams acted as a pressure sensor, which was able to monitor RBC clogging in the pillar channels. The interface was varied depending on the degree of RBC deformability. To evaluate the present method, RBCs with differing degrees of deformability were prepared by suspending normal RBCs in a glutaraldehyde solution. From the experimental results, the present method had the ability to sufficiently measure minor differences in RBC deformability. Furthermore, the present method was employed to quantify RBC deformability with respect to differences in density of RBCs separated by the PercollTM gradient method. In conclusion, the present method was successfully applied to detect differences in RBC deformability under microfluidic platforms.
S9-2 Numerical study of local parabolic rouleaux formation analyzed by axial and radial shear rates
*Cheong-Ah Lee1, Dong-Guk Paeng1,2
1Jeju National University, 2University of Virginia
Red blood cell (RBC) aggregation is an essential rheological determinant that resists blood flow at a lower shear rate. Recently, we have introduced two components of shear rate, namely, the radial and axial shear rates, to understand local parabolic rouleaux formation under sinusoidal pulsatile flow. These were further applied to the Womersley flow with the elastic tube wall and the stenotic artery in this study. A 2D RBC model was used to simulate RBC motion driven by interactional and hydrodynamic forces based on the depletion theory of the RBC mechanics. Based on computational fluid dynamics (CFD) simulation, velocity, axial shear rate, and radial shear rate fields were obtained to calculate the RBC motion. The simulation results from both the Womersley flow with an elastic tube and the stenotic arteries demonstrated that specific ranges of the axial shear rates strongly affected parabolic rouleaux formation. In addition, local RBC aggregation was clearly observed when the axial shear rate was increased. Although the range of the axial shear rate was relatively small (−32 s−1 to 32 s−1) compared with one of the radial shear rate (from 10 s−1 to 100 s−1) at the center of the tube under Womersley flow, rouleaux formation, and its parabolic shape were locally influenced by the axial shear rate and its ratio to radial shear rate. These principal findings may provide a crucial role in understanding the hemorheological characteristics of pulsatile flow in the arteries.
S9-3 Total volume ratio (TVR): A new parameter to evaluate the risk of aneurysm rupture
*Jinmu Jung1, Ui Yun Lee1, Hyosung Kwak2, Dongwhan Lee1
1Division of Mechanical Design Engineering, College of Engineering, Jeonbuk National University, Jeonju, South Korea, 2Department of Radiology, Jeonbuk National University Hospital, Jeonju, South Korea
S9-4 Measurement of platelet adhesion by using correlation mapping
Eunseop Yeom
Pusan National University
Platelets can be activated by the surrounding blood flow where a blood vessel is narrowed as a result of atherosclerosis. Numerous studies have been conducted to identify the relation between platelet activation and thrombus formation. To measure platelet adhesion, this study proposes an image analysis technique. Blood samples were delivered in a microfluidic channel and then platelets were activated by a stenotic micro-channel with 90% severity. By applying the proposed correlation mapping, which visualizes decorrelation of the streaming blood flow, the area of adhered platelets (APlatelet) was estimated without labeling platelets. In order to evaluate the performance of correlation mapping on the detection of platelet adhesion, the effect of tile size was investigated by calculating 2D correlation coefficients with binary images obtained by manual labeling and the correlation mapping method with different sizes of the square tile ranging from 3 to 50 pixels. A maximum 2D correlation coefficient was observed with an optimum tile size of 5 × 5 pixels. As the area of the platelet adhesion increased, the platelets pluged the channel and there was only a small amount of blood flow. This image analysis could provide new insights for better understanding of the interactions between platelet aggregation and blood flows under various physiological conditions.
S9-5 Thrombus formation through upstream activation and downstream adhesion of platelets in a microfluidic system
*Sehyun Shin1, SeonYoung Kim2, ByoungKwon Lee3, ChaeSeung Lim4
1Korea University, 2Rheomeditech. Inc., 3Gangnam Severance Hospital, Yeonsei University, 4Guro Hospital, Korea University
Activation of platelets tends to cause adhesion and aggregation of platelets to the vascular wall. In the circulatory environment, activation and adhesion may not occur at the same site since platelet activation requires time. Thus, we hypothesize that platelet activation at upstream and adhesion at downstream would be relevant in assessing the increased thrombotic risk associated with hyper-responsive vulnerable blood patients. In this study, we designed a highly integrated microfluidic flow system to mimic the hemodynamic environment of the vasculature with an upstream activation of platelets and downstream adhesion of platelets. Platelets can be activated by either agonists or shear stress generated with a rotational disk. The degree of platelet adhesion was monitored by the migration distance (MD) of blood through the microchannel until it was blocked. The degree of pre-activation was widely varied with incubation time, agonist concentration and shear stress levels, which directly affected downstream adhesion of preactivated platelets. There was a linear relation between degree of activation and that of adhesion. However, excessive activation, rather, degraded downstream adhesion of platelets due to upstream aggregation and platelet lysis. These results suggested that platelet adhesion was strongly dependent on the general conditions of upstream activation of platelets. This study implies that downstream thrombus formation may be caused by upstream platelet activation rather than on-site activation.
S10-1 Hemorheological alterations in patients with chronic cerebrovascular disease
*Peter Kenyeres1, Kinga Totsimon1, Alexandra Nagy3, Barbara Sandor1, Katalin Biro1, Laszlo Szapary2, Kalman Toth1, Zsolt Marton1
11st Department of Medicine, University of Pecs, Medical School, Pecs, Hungary, 2Department of Neurology, University of Pecs, Medical School, Pecs, Hungary, 3Department of Behavioral Sciences, University of Pecs, Medical School, Pecs, Hungary
S10-2 Novel predictors of future vascular events in post-stroke patients
*Diana Schrick1, Erzsebet Ezer1, Margit Tokes-Fuzesi2, Tihamer Molnar1
1Department of Anaesthesiology and Intensive Therapy, University of Pecs, Medical School, Pecs, Hungary, 2Department of Laboratory Medicine, University of Pecs, Medical School, Pecs, Hungary
S10-3 Hemorheological investigations in critically ill patients
*Zsolt Marton, Zsofia Eszter Szabo, Kinga Totsimon, Kalman Toth, Peter Kenyeres
1st Department of Medicine, University of Pecs, Medical School, Pecs, Hungary
S10-4 Maternal hemorheological changes in early-onset preeclampsia
*Beata Csiszar1,2, Gergely Galos1,2, Peter Kenyeres1,2, Kalman Toth1,2, Barbara Sandor1,2
11st Department of Medicine, University of Pecs, Medical School, Pecs, Hungary, 2Szentagothai Research Centre, Pécs, Hungary
S10-5 The French paradox – From a rheological point of view
*Andras Toth1,2, Barbara Sandor2, Judit Papp2,3, Miklos Rabai2, Peter Kenyeres2, Istvan Juricskay2, Kalman Toth2
1Department of Medical Imaging, University of Pecs, Medical School, Pecs, Hungary, 21st Department of Medicine, University of Pecs, Medical School, Pecs, Hungary, 3Hungarian Defense Forces Medical Centre, Budapest, Hungary
S10-6 Hemorheological, hematological and histological examination, and 3D flow simulation of arterio-venous fistulas or loop-shaped venous grafts in the rat
*Balazs Szabo1, Adam Varga1, Barbara Barath1, Souleiman Ghanem1, Orsolya Matolay2, Gyorgy Trencseny3, Levente Kiss-Papai5, Balazs Gasz5, Lajos Daroczi4, Norbert Nemeth1
1Department of Operative Techniques and Surgical Research, University of Debrecen, Faculty of Medicine, Debrecen, Hungary, 2Department of Pathology, Faculty of medicine, University of Debrecen, Debrecen, Hungary, 3Division of Nuclear Medicine, Department of Medical Imaging, Faculty of medicine, University of Debrecen, Debrecen, Hungary, 4Institute of Physics, Department of Solid State Physics, Faculty of Physics, University of Debrecen, Debrecen, Hungary, 5Department of Surgical Research and Techniques, Faculty of Medicine, University of Pecs, Hungary
S11-1 Association of sulfur concentration in erythrocytes with heart geometry parameters and blood pressure
*Maria Fornal1, Janusz Lekki2, Jarosław Krolczyk1, Barbara Wizner1, Tomasz Grodzicki1
1Jagiellonian University Medical College, Krakow, Poland, 2Institute of Nuclear Physics PAN, Krakow, Poland
S11-2 Results of blood research relating to rheology, morphology and biochemistry of blood in a man living 50 days in extremely low temperatures
*Zbigniew Joseph Dabrowski1, Aneta Teleglow1, Anna Marchewka1, Maria Fornal2
1Academy of the Physical Education in Cracow, Poland, 2Collegium Medicum, Jagiellonian University, Krakow, Poland
Regular exposure to a cold factor, such as cold water swimming or ice swimming and cold-air, results in an increased tolerance to cold due to numerous adaptive mechanisms in humans. Due to the lack of scientific reports on the effects of extremely low outdoor temperatures on the functioning of the human circulatory system, the aim of this study is to evaluate complete blood count and rheological and biochemical blood indices in multiple Guinness world record holder Valerjan Romanovski, who was exposed to extremely cold environment from below −25° for 50 days in Rovaniemi (a city in northern Finland). Valerjan Romanovski proved that humans can function in extremely cold temperatures. Blood from the subject was collected before and after the expedition. The subject was found to have abnormalities for the following blood indices: testosterone increase by 60.14%, RBC decrease by 4.01%, HGB decrease by 3.47%, WBC decrease by 21.53%, neutrocytes decrease by 17.31%, PDW increase by 5.31%, AspAT increase by 52.81%, ALAT increase by 68.75%, CK increase by 8.61%, total cholesterol decrease by 5.88%, HDL increase by 28.18%. Percentage changes in other complete blood count and biochemical indices were within standard limits. Long-term exposure of the subject (50 days) to extreme cold stress had no noticeable negative effect on daily functioning.
S11-3 Interactions of β-carotene with red blood cells – its effect on their stability and functioning
*Joanna Fiedor1, Mateusz Przetocki1, Aleksander Siniarski2,3, Grzegorz Gajos2,3, Nika Spiridis4, Kinga Freindl4, Kvetoslava Burda1
1AGH University of Science and Technology, Faculty of Physics and Applied Computer Science, Krakow, Poland, 2Jagiellonian University Medical College, Krakow, Poland, 3The John Paul II Hospital, Krakow, Poland, 4Polish Academy of Sciences, Krakow, Poland
S11-4 Correlations between hemorheological parameters in a group of qualified healthy blood donors compared to those in a group of cardiovascular patients
Anna Marcinkowska-Gapinska
Department of Biophysics, Karol Marcinkowski University of Medical Sciences in Poznan, Poland
S11-5 Can nanoparticles be responsible for the development of hypertension?
*Kvetoslava Burda1, Joanna Fiedor1, Magdalena Peter1, Mateusz Przetocki1, Jaroslaw Kiecana2,3, Aleksander Siniarski2,3, Grzegorz Gajos2,3, Nika Spiridis4
1AGH-University of Science and Technology, Krakow, Poland, 2The John Paul II Hospital, Krakow, Poland, 3Jagiellonian University, Medical College, Krakow, Poland, 4Jerzy Haber Institute of Catalysis and Surface Chemistry, Polish Academy of Sciences, Krakow, Poland
Nanoparticles (NPs) due to their peculiar physical and chemical properties are in the center of interest in new technologies. For example, they are used as carries, implants, contrast agents applicable in medicine. The vast range of their uses in our everyday life induces especially high exposure of humans to their action. Therefore it is very important to identify potential risk related to NPs penetrating our environment, in particular to those manufactured and diversified by humans. This knowledge is desired because of the predicted lack of our organism’s capability to tolerate and respond to NPs.
NPs reaching the bloodstream can circulate and cause inflammatory reactions, alterations of central nervous system or cardiovascular system. Their first targets are the blood components. Erythrocytes are exposed to nanoparticles throughout their life span. This means that red blood cells (RBCs) can undergo constant biochemical and morphological changes due to circulated NPs. It has been already postulated that RBC rheological alterations associated with NPs may cause circulatory diseases in macro- and micro-circulation systems. However, results available in the scientific reports are limited and very often provide contradictory opinions. Increased risks of non-accidental, circulatory mortality were reported even at very low concentrations of ambient nanoparticles with an average diameter<2.5 μm.
Based on our preliminary studies, we can postulate that NPs (including the most common in our environment) interactions with RBCs could be responsible for acute occurrence of primary hypertension in young people.
All procedures involving the use of human blood were carried out in accordance with the recommendations of The World Medical Association and approved by the local Bioethical Commission of Physicians (317/KBL/OIL/2019).
S12-1 Intricate journey of micro- and nano-carriers for drug delivery in the blood stream
Dmitry Fedosov
Forschungszentrum Juelich GmbH
Drug delivery by various micro- and nano-carriers offers the possibility of controlled transport of pharmaceuticals to targeted sites (e.g., cancerous tissue). The adhesion of micro- and nanocarriers in blood flow is strongly affected by their distribution within the vessel cross-section. To investigate the adhesion potential of carriers of different sizes, we employ mesoscopic hydrodynamic simulations of blood flow in order to predict margination of carriers or their migration properties toward vessel walls. The margination of carriers is studied for a wide range of hematocrit values and flow rates, and various sizes, ranging from about hundred nanometers to several micrometers, are considered. Our results show that margination strongly depends on the thickness of the available free space close to the wall, the so-called red blood cell-free layer (RBC-FL), in comparison to the carrier size. The carriers with a few micrometers in size are comparable with the RBC-FL thickness and marginate better than their submicrometer counterparts. Deformable carriers, in general, show worse margination properties than rigid particles. Particle margination is also found to be most pronounced in small channels with a characteristic size comparable to blood capillaries. As a conclusion, micron-size particles seem to be favorable for vascular drug delivery in comparison to sub-micron carriers.
S12-2 Physical mechanism of erythrocyte sedimentation: Experiments and gel-model
*Alexis Darras1, Anil Kumar Dasanna2, Thomas John1, Gerhard Gompper2, Lars Kaestner1, Dmitry A. Fedosov2, Christian Wagner1
1Experimental Physics, Saarland University, 66123 Saarbruecken, Germany, 2Theoretical Physics of Living Matter, Institute of Biological Information Processing and Institute for Advanced Simulation, Forschungszentrum Jülich, 52425 Jülich, Germany
Erythrocyte sedimentation rate (ESR) is a physical parameter which is often checked in medical diagnosis. It is indeed well known that in the case of inflammation, the increase in fibrinogen and other proteins induces a higher ESR. Until now, researchers thought that the increase of fibrinogen accelerates the ESR by creating larger aggregates of red blood cells (RBC). Fibrinogen is indeed an aggregation supporting agent of RBCs, and bigger aggregates tend to sediment faster in Stokes regime. However, modeling the ESR measurements with this hypothesis is challenging and often requires physical assumptions specific to this system. Besides, modern colloidal science has shown that attractive particles can form percolating aggregates, as wide as the container. The sedimentation of those colloids then follows a so-called “colloidal gel collapse” regime. Here, we show that RBCs actually follow the same behavior. We present detailed measurements of experimental ESR curves, and original microscopic pictures supporting this claim. We also demonstrate that such assumption naturally leads to efficient colloidal modeling for the sedimentation curve of RBCs.
S12-3 The erythrocyte sedimentation rate as a diagnostic biomarker for neuroacanthocytosis syndromes
Alexis Darras1, Kevin Peikert2,3, Antonia Rabe1,4, François Yaya1,5, Greta Simionato1,6, Thomas John1, Anil Kumar Dasanna7, Semen Bavalyy7, Jürgen Geisel8, Andreas Hermann2,3,9,10, Dmitry A. Fedosov7, Adrian Danek11, Christian Wagner1,12, *Lars Kaestner1,4
1Experimental Physics, Saarland University, 66123 Saarbruecken, Germany, 2Translational Neurodegeneration Section “Albrecht-Kossel”, Department of Neurology, University Medical Center Rostock, University of Rostock, Rostock, Germany, 3Neurodegenerative Diseases, Department of Neurology, Technische Universität Dresden, Dresden, Germany, 4Theoretical Medicine and Biosciences, Saarland University, 66424 Homburg, Germany, 5Laboratoire Interdisciplinaire de Physique, UMR 5588, 38402 Saint Martin d’Hères, France, 6Institute for Clinical and Experimental Surgery, Saarland University, 66424 Homburg, Germany, 7Institute of Biological Information Processing and Institute for Advanced Simulation, Forschungszentrum Jülich, 52425 Jülich, Germany, 8Central Clinical Laboratory, Saarland University, 66424 Homburg, Germany, 9DZNE, German Center for Neurodegenerative Diseases, Research Site Rostock/Greifswald, Rostock, Germany, 10Center for Transdisciplinary Neurosciences Rostock (CTNR), University Medical Center Rostock, University of Rostock, Rostock, Germany, 11Neurologische Klinik und Poliklinik, Ludwig-Maximilians-Universität, 81366 Munich, Germany, 12Physics and Materials Science Research Unit, University of Luxembourg, Luxembourg City, Luxembourg
S12-4 Investigating the red blood cell (dis)aggregation mechanism by means of optical tweezers
*Francois Yaya1,2, Olivera Korculanin3,4, Mehrnaz Babaki3,4, Pavlik Lettinga3,4, Christian Wagner1, Kisung Lee5
1Experimental Physics, University of Saarland, Saarbrücken, Germany, 2Laboratoire Interdisciplinaire de Physique (LIPhy), CNRS and University of Grenoble, Grenoble, France, 3Biomacromolecular Systems and Processes (IBI-4), Forschungszentrum Jülich GmbH, Jülich, Germany, 4Laboratory for Soft Matter and Biophysics, KU Leuven, Leuven, Belgium, 5Center for Soft and Living Matter, Institute for Basic Science, Ulsan, South Korea
Red blood cells (RBCs) are one of the main components of blood. They aggregate to form rouleaux, and disaggregate under high shear rates. Two models have been proposed to describe this phenomenon, namely: cross-bridging and depletion. While experiments in model solution using a polysaccharide (Dextran) have attempted to show evidence in favor of each model, the mechanism of RBCs aggregation still remains unclear. In our experiments, RBCs are in presence of Dextran or fd-virus, a long range depletant borrowed from the field of colloidal physics. We use holographic optical tweezers, to bring RBCs together with a chosen contact area in the two model solutions. The spontaneous aggregation force F a and the disaggregation force F d have been measured along with recording of the whole dynamics of the process. Our results show that the aggregation and disaggregation mechanisms differ in terms of forces and that the dynamics of disaggregation in Dextran is different from fd-virus. Therefore, we conclude that the models to describe the (dis)aggregation appear to be more complex and may not be mutually exclusive.
S13-1 Nanoparticle-mediated delivery of nucleic acids in primary human endothelial cells
*Manfred Gossen1,2, Skadi Lau1,2, Hanieh Moradian1,2,3, Marc Behl1, Andreas Lendlein1,2,3
1Institute of Active Polymers, Helmholtz-Zentrum Hereon, Teltow, Germany, 2Berlin-Brandenburg Center for Regenerative Therapies (BCRT), Berlin, Germany, 3Institute of Biochemistry and Biology, University of Potsdam, 14476 Potsdam, Germany
S13-2 Long-term stabilization of three-dimensional perfusable microvascular networks in microfluidic devices
Ho-Ying Wan1, Jack Chun Hin Chen2, Qinru Xiao2, Christy Wingtung Wong1, Yi-Ping Megan Ho2, Roger D. Kamm3, Sebastian Beyer2, Anna Maria Blocki1
1Institute for Tissue Engineering and Regenerative Medicine, The Chinese University of Hong Kong, 2Department of Biomedical Engineering, Faculty of Engineering, The Chinese University of Hong Kong, 3Department of Biology and Mechanical Engineering, Massachusetts Institute of Technology
Microvasculature supply oxygen and nutrient, hence, they are important for many biological processes, such as wound healing. Engineering microvasculature in vitro provide a chance to study these processes in detail. Currently, in vitro engineered microvasculature involve culturing perfusable microvascular networks (MVNs) in a microfluidic system, which provides a robust microenvironment, allows tight control over important cellular parameters and facilitates microvasculature forming in vitro. However, MVNs often retract immediately after formed and cannot retain function for a long period.
To engineer stable MVNs, we introduce macromolecular crowding (MMC), which provide biophysical stimulus into MVNs cultures, and co-culture of umbilical vein endothelial cells (HUVECs), which from inner lining of MVNs and mesenchymal stem cells (MSCs), which are known to give rise to pericytes. MVNs exhibited hollow lumina that are perfusble and outlined by basement membrane within 2 days of culture. While MVN cultured without MMC regress immediately after being formed, MVNs treated with MMC were stable up to 10 days. Quantification of major basement membrane components, Collagen IV and laminin α5 suggested better basement membrane formation and acumination in MVN supplemented with MMC. Reduced contractivity in 3D co-culture of HUVEC and MSCs were confimed by a functional contractility assay.
To conclude, current data suggested MMC facilitate formation of basement membrane around MVNs and reduce cell contractility that results in microvessel retraction. Hence, stable and functional MVNs model can be engineered in vitro, that provide close resemblance of physiological microvasculature environment and biological processes related to microvasculature.
S13-4 Assessment of leukocyte activation in the intestinal microcirculation in a novel model of CNS injury-induced immunodepression
*Bashir Bietar, Christian Lehmann
Dalhousie University
S13-5 Experimental cannabinoid receptor 2 modulation for the treatment of interstitial cystitis
*Geraint Christopher Berger1, Juan Zhou1, Melanie Kelly2,1,4, Christian Lehmann1,2,3,5
1Department of Anesthesia, Pain Management and Perioperative Medicine, Dalhousie University, 2Department of Pharmacology, Dalhousie University, 3Department of Microbiology & Immunology, Dalhousie University, 4Department of Ophthalmology & Visual Sciences, Dalhousie University, 5Department of Physiology and Biophysics, Dalhousie University
S14-2 Retrospective study of COVID-19-associated coagulopathy in hospitalized patients at Tokai University Hospital
*Masayuki Oki1, Hideki Yanagi1, Masahiro Kamono1, Saki Manabe1, Akiko Taoda1, Ayumi Tsuda1, Satoshi Abe1, Takako Kobayashi1, Koichiro Asano2, Yoshihide Nakagawa1, Yasuhiro Kanatani3, Hideki Ozawa1, Shinya Goto2
1Department of General Medicine, Tokai University School of Medicine, 2Department of Internal Medicine, 3Department of Pharmacology
*Interquartile range are shown within [ ].
S14-3 COVID-19 and thrombosis: The importance of endothelial function
*Shinichi Goto1,2,3, Shinya Goto3
1Brigham and Women’s Hospital, Harvard Medical School, 2Keio University School of Medicine, 3Tokai University School of Medicine
Starting around late 2019, corona virus disease 2019 (COVID-19) caused by severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) virus has rapidly evolved as a large threat to the public health world-wide. Growing evidence has suggested that COVID-19 not only causes severe pneumonia but also increases the risk of thrombosis which is associated with a high risk of mortality. Studies have revealed that the thrombosis caused by COVID-19 is characterized by the damage to the endothelial cells.
In the first part, we will first summarize the basic mechanism of thrombosis caused by endothelial injury and discuss recent evidence on how COVID-19-induced thrombosis is related. We will focus on the damage to the endothelial cells, caused by SARS-CoV-2 infection, that leads to increase in factor VIII and von Willebrand factor (VWF) levels. The similarity and difference between classical thrombotic thrombocytopenic purpura will also be summarized.
In the second part, the informatics and simulation-based approach that could potentially help the research for thrombosis in SARS-CoV-2 infection will be discussed. The machine-learning approach will help integrate high-dimensional data, which is difficult for human brain to understand, to a model that could predict thrombotic risks. The simulation-based approach will help understand the quantitative effect of thrombotic factors/treatments in a complex system comprising the thrombosis/hemostasis system.
S15-1 A novel integrated biomarker for screening diabetic kidney diseases: Critical shear stress of RBCs
*Sehyun Shin1, Junsung Moon2, Jimi Choi3, Sin-Gon Kim3, Kyu Jang Won2
1Korea University, 2Yeungnam University Hospital, 3Korea University Anam Hospital
Many studies have demonstrated that an alteration in hemorheological properties is correlated with diabetic microcirculatory diseases. However, most of these studies have been limited to animal studies or used a small number of clinical samples, due to a lack of point-of-care (POC) devices to measure such properties within clinical environments. Owing to our development of the microfluidic device (RheoScan system) as a POC device, one can easily measure RBC deformability and critical shear stress at any clinical environments. With the RheoScan system, we have conducted extensive clinical studies for more than 10-years to investigate whether any hemorheological properties can be used as diagnostic indexes for diabetic complications including retinopathy as well as nephropathy. After all, we found that the critical shear stress, as a measure of RBC aggregation force, was strongly correlated with two independent diagnostic indexes of albumin to urine creatinine ratio (uACR) and glomerular filtration rate (GFR) for chronic kidney disease (CKD). With the heat map of CKD using ACR and GFR, CSS can show the high sensitivity and specificity for orange zone (moderately) as well as red zone (severely). Therefore, CSS could be a novel integrated biomarker for screening diabetic kidney diseases considering both ACR and GFR.
S15-2 Clinical microfluidic biomarker assays for red cell health and blood rheology
Umut Gurkan
Case Western Reserve University
Alterations in red cell biophysical properties and blood rheology have been associated with numerous hematologic and circulatory disorders. Recent advances in microfluidic biomarker assays enable effective assessment of these biophysical and rheological properties in normoxia or physiological hypoxia in a clinically meaningful way. There are emerging targeted or curative therapies that aim to improve red cell pathophysiology, especially in the context of inherited hemoglobin disorders, such as sickle cell disease. Red cell pathophysiology can be therapeutically targeted and the improvements in membrane and cellular biophysics and blood rheology can now be feasibly assessed via new microfluidic biomarker assays. Recent advances provide a new hope and novel treatment options for major red cell ailments, including inherited hemoglobin disorders, membrane disorders, and other pathologies of the red cell, such as malaria. In this talk, I will describe the significant biophysical and rheological aspects of red blood cell physiology and pathophysiology in relation to recent advances in microfluidic biomarker assays and emerging targeted or curative intent therapies.
S15-3 Concurrent assessment of deformability and adhesiveness of sickle red blood cells by measuring perfusion of an adhesive artificial microvascular network
Madeleine Lu1, Celeste Kanne2,3, Riley Reddington1, Dalia Lezzar1, Vivien Sheehan2,3, *Sergey Shevkoplyas1
1Department of Biomedical Engineering, University of Houston, Houston, TX, USA, 2Aflac Cancer and Blood Disorders Center, Children’s Healthcare of Atlanta, Atlanta, GA, USA, 3Department of Pediatrics, Emory University School of Medicine, Atlanta, GA, USA
Biomarker development is a key clinical research need in sickle cell disease (SCD). Hemorheological parameters are excellent candidates as abnormal red blood cell (RBC) rheology plays a critical role in SCD pathophysiology. Here we describe a microfluidic device capable of evaluating RBC deformability and adhesiveness concurrently, by measuring their effect on perfusion of an artificial microvascular network (AMVN) that combines microchannels small enough to require RBC deformation, and laminin (LN) coating on channel walls to model intravascular adhesion. Each AMVN device consists of three identical capillary networks, which can be coated with LN (adhesive) or left uncoated (non-adhesive) independently. The perfusion rate for sickle RBCs in the LN-coated networks (0.18 ± 0.02 nL/s) was significantly slower than in non-adhesive networks (0.20 ± 0.02 nL/s), and both were significantly slower than the perfusion rate for normal RBCs in the LN-coated networks (0.22 ± 0.01 nL/s). Importantly, there was no overlap between the ranges of perfusion rates obtained for sickle and normal RBC samples in the LN-coated networks. Interestingly, treatment with poloxamer 188 decreased the perfusion rate for sickle RBCs in LN-coated networks in a dose-dependent manner, contrary to previous studies with conventional assays, but in agreement with the latest clinical trial which showed no clinical benefit. Overall, these findings suggest the potential utility of the adhesive AMVN device for evaluating the effect of novel curative and palliative therapies on the hemorheological status of SCD patients during clinical trials and in post-market clinical practice.
S15-4 Usefulness of oxygen gradient ektacytometry in sickle cell disease
*Philippe Connes1,2, Camille Boisson1,2,3, Minke Rab4,5, Elie Nader1,2, Céline Renoux1,2,3, Philippe Joly1,2,3, Romain Fort1,2,6, Alexandra Gauthier1,2,7, Yves Bertrand7, Richard van Wijk4, Vivien Sheehan8, Eduard van Beers5
1Laboratoire Interuniversitaire de Biologie de la Motricite (LIBM) EA7424, Team Vascular Biology and Red Blood Cell, Universite Claude Bernard Lyon 1, Universite de Lyon, France, 2Laboratoire d’Excellence du Globule Rouge (Labex GR-Ex), PRES Sorbonne, Paris, France, 3Laboratoire de Biochimie et de Biologie Moleculaire, Lyon, 4Central Diagnostic Laboratory – Research, University Medical Center Utrecht, Utrecht University, Utrecht, The Netherlands, 5Van Creveldkliniek, University Medical Center Utrecht, Utrecht University, Utrecht, The Netherlands, 6Departement de Meedecine Interne, Hopital Edouard Herriot, Hospices Civils de Lyon, 7Institut d’Hematologie et d’Oncologie Pediatrique, Hospices Civils de Lyon, Lyon, 8Department of Pediatrics, Division of Hematology/Oncology, Baylor College of Medicine; Houston Texas, USA
Sickle cell disease (SCA) is caused by a mutation in the beta-globin gene leading to the production of an abnormal hemoglobin (Hb), called HbS. The main pathophysiological feature of HbS is that it polymerizes when deoxygenated, leading to a mechanical distortion of red blood cells (RBCs): i.e., sickling. Sickled RBCs are very fragile, which explains enhanced hemolysis and anemia. In addition, these RBCs are poorly deformable and play a role in several complications, such as acute vaso-occlusive crises. The recently developed technique oxygen gradient ektacytometry (Oxygenscan, LORRCA, Mechatronics, The Netherland) allows for a more comprehensive functional characterization of the rheological behavior of SCA RBCs over a range of oxygen tensions. Several parameters can be derived: (1) EImax; the RBC deformability at normoxia; (2) EImin; the lowest RBC deformability reached upon deoxygenation; (3) Point of Sickling (PoS); the pO2 at which RBC deformability decreases below 5% of EImax during deoxygenation, which reflects the beginning of sickling; (4) Delta EI; the difference between EImin and EImax. In this presentation, we will show how the oxygenscan parameters are affected by the treatment (transfusion, hydroxyurea), percentage of foetal Hb and genetic factors in SCA patients. We will provide results on the comparisons of oxygen parameters between SCA patients and other sickle cell genotypes. We will also show some associations with several indicators of clinical severity and present the evolution of oxygenscan parameters during acute complications. Altogether, our data suggest that oxygenscan would be useful to characterize SCA clincal phenotypes and should stimulate further works to identify new therapeutical strategies.
S15-5 A novel microfluidics-based point of care technique for viscoelastic hemostatic assay
*Ozlem Yalcin1, Ahmet Can Erten2, Berfin Irmak Torun3, Fatma Oz3
1Koc University, School of Medicine, Koç University, Research Center for Translational Medicine (KUTTAM), Istanbul, Turkey, 2Department of Electronics and Communication Engineering, Istanbul Technical University, Istanbul, Turkey, 3Koç University, Graduate School of Biomedical Sciences and Engineering, Istanbul, Turkey
Blood coagulation assessment plays a crucial role in studying the risk of bleeding during extensive surgery and diagnosis of underlying coagulopathy. In this study, we have investigated the blood coagulation monitoring potential of a novel microfluidic device, termed microTEM. with embedded micropillars. By tracking the movement of micropillars with a CCD camera, which are facing the forces due to the clotting blood within the microfluidic channel, microTEM can detect coagulation signals through micropillars’ movements within shear stresses from 2.3 up to 11.5 dyne/cm2 while clotting blood is moving inside the microchannel. ROTEM® Delta device and its test reagents (INTEM, EXTEM) were used as a gold standard system for comparison with the microTEM results. The effect of hematocrit level (10, 20, 30 and 40%) on coagulation parameters in both the ROTEM Delta and the microTEM devices were examined. The effect of varying calcium concentration on the coagulation time was investigated in this study as well. The citrated blood samples were treated with calcium chloride solutions at 25, 40, 50, 75 and 100 mM concentrations, and the coagulation times were subsequently recorded by the microTEM and visual observation of the same samples. The microTEM parameters showed strong positive correlations with the ROTEM parameters and visual observation of the samples (r = 0.87, r = 0.96, p < 0.001, respectively). This study demonstrates a new method and device for blood coagulation monitoring based on the displacement of micropillars due to dragging force of the clotting blood.
S16-1 Competition between red blood cell aggregation and breakup: Depletion force due to filamentous viruses vs. shear flow
*Olivera Korculanin1,2, Tatiana Kochetkova1, Pavlik Minne Paul Lettinga1,2
1Biomacromolecular Systems and Processes (IBI-4), Forschungszentrum Juelich GmbH, Germany, 2Laboratory for Soft Matter and Biophysics, KU Leuven, Belgium
Human blood is a shear-thinning fluid with a complex response that strongly depends on the red blood cell’s (RBC’s) ability to form aggregates, called rouleaux. Despite numerous investigations, microscopic understanding of the break up of RBC aggregates has not been fully elucidated. Here, we present a study of breaking up aggregates consisting of two RBCs during shear flow. For the first time, we employ the filamentous fd bacteriophage as a rod-like depletant agent with a very long range interaction force. The depletion interaction between the cells is fine-tuned through mixtures of RBCs in PBS at different rod concentrations. We visualize the structures while shearing by combining a home-build counter-rotating cone-plate shear cell with fast microscopy imaging. The non-equilibrium phase diagram of shear rates versus depletant concentrations shows regions for different flow responses and separation stages of the RBCs doublets. With increasing interaction forces, the full-contact flow states dominate, such as rolling and tumbling. It is seen that the RBC doublets start to undergo separation only in the tumbling motion during a critical angle range that the normal of the doublets makes with the flow direction. However, at sufficiently high shear rates, the critical range becomes too short time-wise that the cells continue to tumble without separating.
S16-2 Sphingosine-1-phosphate and Adenosine affect the oxygen dependence of erythrocyte metabolism
Francesco Misiti
Cassino and Lazio Meridionale University
Human erythrocyte metabolism (glycolysis and Pentose Pathway) is modulated by the cell oxygenation state. Among other mechanisms, competition of deoxyhemoglobin and some glycolytic enzymes for the cytoplasmic domain of band 3 is involved in modulation. This metabolic modulation is connected to variations in intracellular NADPH and ATP levels as a function of the oxygenation state of the cell, and, consequently, it has physiologic relevance. The present study investigates the effect of sphingosine-1phosphate (S1P) and adenosine exposure on this metabolic modulation. We show that both S1P and adenosine promote deoxygenated Hb anchoring to the membrane; this event enhances the release of membrane-bound glycolytic enzymes to the cytosol, inducing glycolysis with respect to Pentose Pathway (PPP). As a result, metabolic differences between erythrocytes incubated at high and low oxygen saturation increase following exposure to both sphingosine-1-phosphate (S1P) and adenosine. Our findings suggest a novel signaling network regulating the oxygen-dependent metabolic modulation.
S16-3 The role of macromolecular depletion on the adhesion of red blood cells with a reduced sialic acid content
*Björn Neu1, Huimin Teo2, Zhengwen Zhang2
1Rhine-Waal University of Applied Sciences, 2Nanyang Technological University
Senescent red blood cell (RBC) adhesion to other surfaces is governed by the interplay of specific and non-specific forces. To elucidate the role of sialic acid in cell adhesion, desialylated RBCs were treated with various concentrations of neuraminidase and the adhesion to albumin-coated glass surface induced by dextran solution was quantified by quantitative interference reflection microscopy. It is observed that membrane undulation is markedly suppressed with increasing molecular weight and concentration of dextran. The interpretation of our results in terms of the depletion model appears appropriate, in that cell adhesion depends on the balance of polymer depletion force vs. the electrostatic repulsive force and undulation force. It is concluded that the membrane glycocalyx, especially the sialic acid, plays a significant role in polymer depletion induced cell–substrate interaction. This study thus suggests that depletion interaction can control cell–cell interactions in complex environments (e.g., in vivo), and indicates that considering the interplay of all plasma constituents as well as the cellular properties might be important to understand the removal of senescent red blood cells from the circulation.
S16-4 The Mizar®: A novel, fully-automated aggregometer
*Lennart Kuck1, Francesco A. Frappa2, Michael J. Simmonds1
1Biorheology Research Laboratory, Menzies Health Institute Queensland, Australia, 2Alcor Scientific Inc., Rhode Island, USA
S17-1 Development of an experimental microfluidic device and methodology for assessing microrheological properties of blood
*Nadia Mladenova Antonova1, Khristo Khristov2, Anika Svilenova Alexandrova3, Alexei Vasilievich Muravyov4
1Dept. Biomechanics, Institute of Mechanics at the Bulgarian Academy of Sciences, Sofia, Bulgaria, 2Institute of Physical Chemistry at the Bulgarian Academy of Sciences, Sofia, Bulgaria, 3Institute of Mechanics at the Bulgarian Academy of Sciences, Sofia, Bulgaria, 4Yaroslavl State Pedagogical University Ushinskii, Yaroslavl, Russia
S17-2 Comparative study of the microrheological properties of blood in patients with type 2 diabetes mellitus, using viscometry and microfluidic flow analysis
*Anika Svilenova Aleksandrova-Watanabe1, Nadia Mladenova Antonova1, Alexey Vasilievich Muravyov2, Khristo Ivanov Khistov3, Irena Vasileva Velcheva4
1Dept. of Biomechanics, Institute of Mechanics, Bulgarian Academy of Sciences, Sofia, Bulgaria, 2Dept. of Medical and Biological Foundations of Sports, Yaroslavl State Pedagogical University named after K. D. Ushinsky, Yaroslavl, Russia, 3Dept. of Interfaces and Colloids, Institute of Physical Chemistry, Bulgarian Academy of Sciences, Sofia, Bulgaria, 4Clinic of Nervous Diseases, Uni Hospital, Panagyurishte, Bulgaria
S17-3 Microrheological responses of red blood cells (RBCs) to gasotransmitters in persons with different levels of oxygen supply to the body
*Alexei Vasilievich Muravyov1, Pavel Valentinovich Mikhailov1, Irina Alexandrovna Tikhomirova1, Roman Sergeevich Ostroumov1, Victor Vasilievich Zinchuk2
1State Pedagogical University, Yaroslavl, 2State Medical University, Grodno, Belarus
The aim was to investigate the RBC microrheological responses to GTs in individuals with different levels of oxygen supply.
The reported study was funded by RFBR and BRFBR, project number 20-515-00019 (No. M20P-428 – БРФФИ.
S17-4 Effect of gasotransmitters (NO and H2S) on hemorheology and blood clotting
*Irina Alexandrovna Tikhomirova1, Elena Petrovna Petrochenko1, Yulia Viktorovna Malysheva1, Alexei Vasiljevich Muravyov1, Alexander Sergeevich Petrochenko2
1Yaroslavl State Pedagogical University, 2Yaroslavl State Medical University
S17-5 Interaction Forces of pairs of RBCs and their relation to aggregation parameters under normal and pathological conditions
*Alexander V. Priezzhev1, Andrei E. Lugovtsov1, Alexey N. Semenov1, Larissa I. Dyachuk2, Petr B. Ermolinskiy1
1Physics Department of Lomonosov Moscow State University, 2Medical Research and Education Centre of Lomonosov Moscow State University
The interaction of red blood cells (RBC) with each other leading to the formation of aggregates is one of the most important mechanisms for regulating blood fluidity in the vessels. Such interaction has a significant effect on the microcirculation and rheology of blood in the body. We measured the kinetics and the forces of interaction of different pairs of RBCs resulting in the formation of paired aggregates in vitro, using double-channel laser tweezers (LT). Also, we measured the aggregation parameters of RBC such as aggregation index, characteristic time of aggregation and hydrodynamic strength of aggregates in whole blood samples using laser aggregometry techniques based on diffuse light scattering. Overall we tested the RBC in blood samples of more than 240 individuals including 18 healthy volunteers and more than 220 patients of Lomonosov Moscow State University medical centre suffering from arterial hypertension and/or type 2 diabetes mellitus. The correlation analysis and quantitative comparison of the results allowed for concluding that the aggregation of RBC is enhanced in these pathologic conditions in somewhat different ways. The interaction forces and aggregation time of single cells measured with LT are interrelated with parameters of RBC aggregation obtained on ensembles of RBC in whole blood by laser aggregometry. These results are important both for understanding the fundamental mechanisms of RBC aggregation, and for assessing the differences in the parameters of their aggregation kinetics under normal and pathological conditions.
The study was supported by Russian Foundation for Basic Research (grant No. 19-52-51015).
S17-6 Microrheological responses of RBCs after age (density) separation
*Petr Ermolinskiy1, Andrei Lugovtsov1, François Yaya2,5, Lars Kaestner2,3, Christian Wagner2,4, Alexander Priezzhev1
1Physics Department, Lomonosov Moscow State University, 119991 Moscow, Russia, 2Experimental Physics, Saarland University, 66123 Saarbrücken, Germany, 3Theoretical Medicine and Biosciences, Saarland University, 66123 Saarbrücken, Germany, 4Physics and Materials Science Research Unit, University of Luxembourg, L-1511 Luxembourg, Luxembourg, 5Laboratoire Interdisciplinaire de Physique, UMR 5588 CNRS and University Grenoble–Alpes, 38058 Grenoble, France
Red blood cell (RBC) spontaneous aggregation process strongly influences the microcirculation of blood in vessels and capillaries. RBC aggregation can alter due to many factors including the composition of blood plasma, temperature of blood and many others. Aggregation mechanisms are still not very clear, as well as the influence of different factors. We studied the effect of RBC in vivo aging on the RBC aggregation properties in vitro at the cellular level using laser tweezers and laser aggregometry (LA) based on diffuse light scattering. RBC were separated in four different age fractions using Percoll density gradient centrifugation. We measured the aggregation forces (AF) between different pairs of RBC suspended in autologous plasma of blood samples freshly drawn from 5 healthy individuals. The results show an enhancement of RBC aggregation in cell doublets with age: the youngest cells exhibit AF = 2.4 ± 0.2 pN (M ± SE), whilst the oldest ones – AF = 3.8 ± 0.1 pN (M ± SE). Also, the hydrodynamic strength of aggregates (critical shear stress (CSS)) was measured using LA for different age fractions. A statistically significant increase in CSS was measured during RBC aging. The obtained data allow us to better understand the contribution of the cellular factors to RBC aggregation. This is important both for the development of future clinical applications and for understanding the interrelation between RBC in vivo aging and RBC aggregation properties.
The study was supported by Russian Foundation for Basic Research (grant No.19-52-51015).
S19-1 Disturbances in skin temperature oscillations and blood rheological and electrical properties in patients with Diabetes mellitus type 2
*Nadia Mladenova Antonova1, Vasilka Krumova Paskova2, Irena Vasileva Velcheva3, Sergey Yurievich Podtaev4
1Dept. Biomechanics, Institute of Mechanics at the Bulgarian Academy of Sciences, Sofia, Bulgaria, 2Institute of Mechanics at the Bulgarian Academy of Sciences, Sofia, Bulgaria, 3Uni Hospital, Panagyurishte, Bulgaria, 4Institute of Continuous Media Mechanics, RAS, Perm, Russia
S19-2 Functional states of PMN in pregnant women with hypertension assessed with a chemiluminescent method – Preliminary data
*B. Bechev1, M. Magrisso2, S. Stoeff1, Sv. Jovtchev1, S. Miteva1, S. Alexandrov1, J. Ivanov1, M. Pencheva3, D. Koleva4, I. Buteva4, M. Vretenarska5, N. Nikolova6 and V. Iliev6,7
1Dept Medical Physics and Biophysics and 3Dept Biology of Medical University Sofia, Bulgaria, 2Omrad Electronics LTD, Beer Sheva, Israel, 4OGW/MHAT “Nadezhda” Sofia, 52nd MHAT Sofia, Nephrology Ward, 6MC Vitclinic, 7Military Medical Academy of Sofia
S19-3 Participation of polymorphonuclear leukocytes in initiation and evolvement of pathologies induced by SARS-Cov-2 virus
*B. Bechev1, S. Stoeff1 and K. Kavaldzhieva2
1Dept Medical Physics and Biophysics, 2Dept Biology of Medical University Sofia, Bulgaria
PMN and development of Covid-19 disease: For Covid-19 we postulate 3 possible scenarios: asymptomatic course, moderate and severe (with hyperinflammation). The possible scenario depends mainly on opposing processes of virus replication and its destruction/neutralization by the immune system, its capacity (including the influx of newly synthesized cells or depots) and damage of vascular endothelium. The participation of PMN in the implementation of the three scenarios is discussed.
Interrelation between the PMN activities and COVID-19 microcirculatory problems: The neutrophil-to-lymphocyte ratio (NLR) and dynamic lowering of most routine blood parameters seem to be critical markers in severe COVID-19. Hypersegmented and large deformable activated neutrophils, size increase of monocytes, RBC alterations (shape and “rouleaux” formation) and physical phenotype changes of RBC (smaller and less deformable) are likely the boost of vascular microthrombosis, ischemia and multiorgan failure. Hyperproduction of cytokines and chemokines orchestrate neutrophils, monocytes, and macrophages mobilization and adhesion to endothelial cells with release of reactive oxygen species that damages the endothelial barrier. The resulting pro-adhesive and prothrombotic effects stimulate further adhesion of leukocytes and platelets to the vascular endothelium, causing vascular micro-thrombosis, capillary plugging and greater impairment of capillary flow.
S20-1 Search for efficient diagnosis and therapy of resistant BRAF mutated melanoma using biophysical methods
*Tomasz Kobiela1, Anna Sobiepanek1, Swamy Kasarla1, Weronika Prorok1, Tomasz Gambin2
1Warsaw University of Technology, Faculty of Chemistry, 2Warsaw University of Technology, Faculty of Electronics and Information Technology
Melanoma, which originates from melanocytes, after entering the metastatic stage causes the highest mortality among skin malignant tumors, resulting from the lack of effective therapy due to common resistance to the applied drugs [1]. In the view of increasing worldwide prevalence, the intensive search for successful therapy pattern is undertaken with the use of combined strategy. This includes the combination of recognized anti-cancer drugs with compounds acting on various vital cellular signaling pathways, particularly on the energy metabolism. In melanoma, the mutation of the proto-oncogene BRaf cytoplasmic serine – threonine kinase (BRAF) gene is the most common (over 50% of patients), which is why most of the applied drugs are directed at inhibiting that signaling pathway. For the optimization of the choice of possible compounds, we performed the modified SynGeNet drug combination prediction study [2]. Experimental validation of effective combinations was done by monitoring the interaction of specific lectins with cellular surface glycans typical for various stages of melanoma progression in realtime experiments using quartz crystal microbalance with the dissipation monitoring and atomic force microscopy. Evaluation of the modification of the glycosylation process of metastatic melanoma cells as result of the applied combination of compounds could reveal potential usability in malignant melanoma treatment.
[1] Sobiepanek, A. et al. (2021). European Biophysics Journal, 50, 523.
[2] Regan-Fendt, K. E. et al. (2019). Npj Systems Biology and Applications, 5(1), 6.
S20-2 Rheological properties of biological materials
*Joanna Zemla1, Claude Verdier2, Malgorzata Lekka1
1Institute of Nuclear Physics Polish Academy of Sciences, PL-31342 Krakow, Poland, 2Laboratoire Interdisciplinaire de Physique, Université Grenoble Alpes, CNRS, Grenoble, F-38000, France
The role of the rheological properties at the cellular and macroscopic scale with regards to normal, diseased, and treated samples will be reviewed. We will discuss the role of viscosity in the overall mechanical properties of soft materials. One of the questions is whether we can find different properties at the macroscopic level than the ones found for individual cells. Various techniques used for measuring micro- and macro-rheological properties will be described. Chosen constitutive laws will be presented, and finally, applications of theoretical models will be discussed in chosen situations of interest.
S20-3 Nanomechanical assessment of cancer cells and solid tumors as a mechanical biomarker
Andreas Stylianou1,2
1European University Cyprus, 2University of Cyprus
Cancer progression is closely related to changes in mechano-cellular phenotype and in the structure and mechanical properties of the tumor microenvironment in a complex and not well-understood manner. During cancer progression cancer cells, as well as cancer-associated fibroblasts, become softer. On the other hand, desmoplasia, a tumour-associated fibrotic reaction, is responsible for tumor stiffening. Desmoplasia poses a major barrier to effective drug delivery and has been associated with a poor prognosis. Tumor stiffening can cause blood vessel inefficiency and hypo-perfusion, and as a result, it poses major physiological barriers to the systemic delivery of drugs. Consequently, there is an urgent need for the development of novel biomarkers, that characterize the mechanical state of a particular tumor so as to support the development of novel therapeutic strategies that target the tumor mechanical microenvironment. Atomic Force Microscopy (AFM) arises as a unique tool for assessing the nanomechanical properties of cancer cells and solid tumors. The identification of unique AFM-based NanoMechanical FingerPrints can lead to the development of novel mechanical biomarkers for treatment perdition and monitoring.
S21-1 Cell age sensitivity of red cells to mechanical stresses and calcium load
Lennart Kuck1, Jason N. Peart2, Oliver Todd1, *Michael J. Simmonds1
1Biorheology Research Laboratory, Menzies Health Institute, Griffith University Gold Coast, Queensland, Australia, 2School of Medical Science, Griffith University Gold Coast, Queensland, Australia
S21-2 Senescence and red cell rheology
*Edgar O’Rear1, James Buerck1, Phillip Coghill2, Ahmed El Banayosy3, Hendra Setiadi3
1University of Oklahoma, 2VADovations, Inc., 3INTEGRIS Baptist Medical Center
The senescent cell phenotype exists in tissues and is associated with pathologies of the elderly. In the absence of a nucleus, senescent red blood cells have distinctly different properties from senescent cells of other tissues. These properties, namely loss of volume, reduced deformability and binding of immunoglobulin IgG to senescent cell antigen, contribute to their removal from the circulation by the spleen. Shortened circulatory life spans reported for red cells of patients with prosthetic heart valves and those on mechanical circulatory support devices (MCSDS) suggest accelerated ageing by shear stress. Fresh human erythrocytes were obtained from the blood of healthy volunteers, resuspended in Ringer’s/albumin, and exposed to supraphysiologic shear in a microfluidics channel. After exposure to shear, cells were incubated in autologous plasma for possible binding of IgG. Interrogation of the cells by flow cytometry was carried out with labelled mouse monoclonal anti-human IgG and anti-Glycophorin A (CD235a). Average percentage of CD235a+ RBCs with IgG increased steadily from a control value of 1.0% to 15.9% (p < 0.001) after a single exposure at 100,000 s−1 for durations ranging from 0 to 15 ms. Flow cytometry results were suggestive of greater binding (p = .07), a result confirmed by image analysis of confocal microscopy images. Erythrocyte microparticle (ErMP) concentrations in the size range 0.5–1.0 μm went from 8800 to 85,000 (p = 0.001). Production of ErMPs by MCSDs in mock flow loop experiments with fresh human blood increased four-fold and eight-fold after two hours for the HeartMate II and CentriMag, respectively. The percentage of IgG-bound RBCs increased from 3–5% (CentriMag) and 2.5–7% (HeartMate II). These results support the concept of shear-induced ageing of erythrocytes.
S21-3 Asymmetrical erythrocyte morphology to detect sublethal damage
*Nobuo Watanabe1,2, Antony P. McNamee3, Jarod T. Horibin3,4, John F. Fraser5, Masataka Inoue2, Masaya Hakozaki2, Fukuta Matsuzawa2, Michael J. Simmonds3
1Biofluid Science and Engineering Laboratory, Dept. of Bio-Science and Engineering, College of Systems Engineering and Science, Shibaura Institute of Technology, Saitama, Japan, 2Biofluid Science and Engineering Laboratory, Systems Engineering and Science, Graduate School of Engineering and Science, Shibaura Institute of Technology, Saitama, Japan, 3Biorheology Research Laboratory, Menzies Health Institute Queensland, Griffith University, Gold Coast, Australia, 4Perth Blood Institute, West Perth, Perth, Australia, 5Critical Care Research Group, University of Queensland & The Prince Charles Hospital, Brisbane, Australia
The unique viscoelastic properties of erythrocytes facilitate dramatic morphological changes in response to the surrounding environment. The stability of erythrocyte shape in flow, and subsequent shape recovery is intrinsically linked to physical properties of the cytosol, cytoskeleton, and membrane – all of which are highly sensitive to supraphysiological shears that are intrinsic to mechanical circulatory support. Given the need to minimize blood trauma within artificial organs, we aimed to investigate whether subtle morphological changes to erythrocytes in flow could indicate sublethal blood damage.
Healthy human blood was acquired from donors through venipuncture of a prominent vein in the antecubital region. Blood was 200 times diluted in a solution of polyvinylpyrrolidone (PVP) in phosphate buffered saline with a viscosity of 30 ± 1.5 mPas and pH of 7.4 ± 0.05 at 37 °C. Resultant erythrocyte-PVP suspensions were then exposed to discrete shears ranging 10–60 Pa for up to 300 s exposure duration. Following accumulated exposure to supraphysiological shear stress (60 Pa × 40 s, 40 Pa × 120 s), asymmetrical ellipses were detected in small quantities of erythrocyte subpopulations. The majority of erythrocytes, however, maintained their normal morphology up to 60 Pa × 300 s.
These findings indicate that asymmetrical morphology may be a sensitive marker to delineate the onset of sublethal blood damage. It is likely that these subpopulations of abnormal erythrocytes would impact conventional indirect deformability measurement techniques (such as ektacytometry and filterability).
S21-4 Effects of mechanical heart valves on circulating blood in patients with valvular heart diseases
*Toru Maruyama1, Michinari Hieda1, Aya Sato2, Takehiko Fujino2
1Kyushu University, 2Institute of Rheological Function of Foods, Co. Ltd.
S21-5 A structured mechanical risk sensitivity assessment system using red cell deformability and fragmentation parameters
*Ozlem Yalcin1,2, Elif Ugurel2, Polat Goktas1, Evrim Goksel1,3, Neslihan Cilek1,3, Dila Atar1
1Koç University, School of Medicine, Istanbul, Turkey, 2Koç University, Research Center for Translational Medicine (KUTTAM), Istanbul, Turkey, 3Koç University, Graduate School of Biomedical Sciences and Engineering, Istanbul, Turkey
Supra-physiological shear stresses (SS) induce irreversible impairments of red blood cell (RBC) deformability, overstretching of RBC membrane and fragmentation of RBCs causing free hemoglobin to be released into plasma, which may lead to anemia and thrombotic complications. In this study, we investigated RBC mechanical damage in oxidative stress (OS) and metabolic depletion (MD) models by applying SS (>100 Pa for 300 s) by an ektacytometer (LORRCA MaxSis) and assessed RBC deformability. Next, we examined hemolysis and measured RBC volume and count by Multisizer 3 Coulter Counter to evaluate RBC fragmentation. RBC deformability was significantly impaired at all applied SS levels in OS (p < 0.05) and in the range of 40–100 Pa in MD (p < 0.01). Hemolysis was detected at 90–100 Pa SS levels in MD and at all applied SS levels in OS. Supra-physiological SS increased RBC volume in both damage models and control group. The number of fragmented cells increased at 100 Pa SS in the control and MD and at all SS levels in OS, which is accompanied by hemolysis. Fragmentation sensitivity index (FSI) increased at 50–100 Pa SS in the control, 100 Pa SS in MD and at all SS levels in OS. Therefore, we propose a novel index (FSI) to evaluate the susceptibility of RBCs experiencing a mechanical trauma before they undergo fragmentation. The assessment of RBC fragmentation would provide an effective and accurate method for detecting sub-hemolytic RBC damage in patients supported by mechanical circulatory assist devices and in pathological conditions where RBCs are more prone to fragmentation.
S21-6 Measurements of erythrocyte deformation in shear and extensional flows
*M Keith Sharp1, Mohammad M Faghih2
1University of Louisville, 2US Food and Drug Administration
Fluid stress-induced hemolysis is a persistent concern during the development of cardiovascular devices and treatment protocols. To accelerate device development cycles, a long-standing goal has been to formulate a hemolysis prediction algorithm that can be universally applied to the wide range of flows that can be encountered in such devices. A number of models, including extensions of the popular powerlaw model, have correlated hemolysis to a von-Mises-like scalar stress that weighs shear stresses more highly than extensional stresses by a factor of 31∕2. This scaling, borrowed from Hookean solid mechanics, was hypothesized to also apply to the fluid–solid interaction between erythrocytes and surrounding plasma. Experiments were performed to test this hypothesis. The deformation of cells in extensional flow in a hyperbolic contraction was compared to that in shear flow between flat plates. Microscopic images were captured in each channel using a spark lamp to stop the motion. A dilute suspension of cells was forced through each channel using a custom-built high-power syringe pump. Fluid stress up to about 100 Pa was achieved with this setup by adjusting the pump speed and by increasing the viscosity of the suspending medium, which also maintained laminar flow. In the contraction, cells were imaged near the centerline, where extensional stress dominates. The aspect ratios of the cells were measured with image-based digital calipers. Aged (banked) blood samples from two human donors were tested. These experiments showed that extensional stress caused more deformation than did shear stress, which suggests that it is more hemolytic. It was also found that suspending medium viscosity significantly impacted aspect ratio for shear flow, but not for extensional flow. Curves of aspect ratio versus stress for extensional flow could be shifted to match that for shear flow at the suspending medium viscosity (4.2 cP) closest to that of whole blood by applying a factor of approximately 34. Therefore, the von-Mises-like scalar stress, which weighs shear stress more strongly than extensional stress, does not faithfully scale cell deformation, and a revised scalar stress with the weightings found in these experiments is recommended.
S21-7 In silico simulation of hemodynamics and blood cell mechanics inside human vasculature
*Senol Piskin1, Aya Ahmed Faeek Elgebaly2
1Department of Mechanical Engineering, College of Engineering, Istinye University, Istanbul, Turkey, 2Department of Biomedical Engineering, Faculty of Electrical and Electronics Engineering, Yildiz Technical University, Istanbul, Turkey
Blood perfusion is the process of delivering blood to the capillary bed of organs or tissues. This process includes nutrient and oxygen transportation through the capillaries. Red blood cells (RBCs) are the main actors of this process and carry and deliver oxygen to peripheral tissues through different microcirculatory regions. The circulatory system consists of the heart, arteries, arterioles, capillaries, veins, lungs, organs, and lymphatic system. The blood flow dynamics and vessel sizes vary significantly among these components. Also, the deformability of the vessels is quite different and can change due to ageing or diseases such as atherosclerosis or diabetes. Blood flow coupled with vessel deformation is the main factor that maintains the stress and strain mechanics on the RBCs (and even white blood cells). The behavior of RBCs can be observed in vitro by applying a predetermined shear strain or shear stress. But this does not represent the real physiological state in most cases. The stress/strain level changes in vivo are substantially based on the vessel size and anatomy, material behavior, flow dynamics, and even the location of the cell inside the vessel (e.g., distance from the center or position inside the flow profile). Turbulent flow makes it even more diverse due to vorticities and eddies. All these complexities can be captured with high precision using computational models and simulations instead of physical experiments. In this presentation, we will present our previous studies on computational simulation of cell mechanics using computational fluid dynamics (CFD) tools inside real human vasculature. Furthermore, we will discuss some of the current literature and our project plans regarding continuum mechanics and particle dynamics.
S22-1 Motion of molecular motors reflecting rheological properties in cells
Takayuki Ariga
Yamaguchi University
Various biomolecular machines are working in cells. In particular, molecular motors play a crucial role in converting free energy from chemical reactions into mechanical work. Recently, the molecular mechanisms of the molecular motors have been clarified in detail by the progress of single-molecule observations and manipulation techniques. However, these studies were mainly conducted in vitro, i.e., in a laboratory environment, and thus their findings on the mechanism did not reflect the rheological properties of the cells in which the motors work.
We investigated the energetics of a molecular motor, kinesin-1, which transports vesicles in cells. Quantification of the dissipation by measuring fluctuations and responses of the kinesin movement indicates that kinesin is unexpectedly less efficient. On the other hand, measurement of fluctuations and responses in cells revealed that the intracellular environment is actively (nonthermally) fluctuating. However, such intracellular active fluctuations have not been reflected in the previous studies on molecular motors. From this, it was hypothesized that active fluctuations in cells affect kinesin movement and efficiency.
To investigate the effect of intracellular active fluctuations on kinesin, we developed a measurement system that artificially applies active fluctuations observed in cells to single-molecule kinesin as external force fluctuations. The change of kinesin velocity was measured by applying the external force fluctuations, indicating that kinesin accelerates due to the active fluctuations. In addition, the acceleration was reproduced by a mathematical model analysis. The universality of the theory behind the model suggests that the acceleration occurs in general biomolecular machines.
In this symposium, we will introduce our current finding on the effect of active fluctuations on the kinesin movements and discuss the effects of rheological properties of cells on individual molecular machines.
S22-2 Glassy cytoplasm driven by non-thermal forces
*Kenji Nishizawa1,2, Daisuke Mizuno3
1IBDM, 2CNRS, 3Department of Physics, Kyushu University
Cell interiors are crowded with various soft objects which are driven out of equilibrium with metabolic activity. The effects of “crowding” and “activity” on the cell mechanics present an intriguing challenge which is investigated here by performing microrheology experiments in cell extracts and in living cells.
For that purpose, we prepared several models of cytoplasm which lack cytoskeletons (E. coli, Xenopus eggs, and HeLa cells). Viscosity of cell extracts showed super-exponential increase with the protein concentrations. Furthermore, viscoelastic properties of cytoplasm in the living cultured cells (HeLa cells) were measured by optical-trap-based microrheology under 3-dimensional feedback of a piezo-mechanical sample stage [1]. Volume fraction of intracellular macromolecules was changed in the range of (0.2–0.6 g/ml) by adjusting the osmotic pressure. We found that the viscosity in living cells showed purely exponential increase with respect to the macromolecule concentration.
The Angell plot of our data clarified that the cell extract and living cytoplasm is a fragile and a strong glass former, respectively. Glasses are fragile when their components cannot individually relax local frustrations in their arrangements. In living cytoplasm (a strong glass former), non-thermal forces are generated by e.g., motor proteins as we observed the violation of fluctuation-dissipation theorem. These non-thermal forces can activate the local structural relaxations. We speculate that such active mixing can erase the origin of cytoplasmic fragility (dynamic heterogeneity), and thus convert fragile cytoplasm to strong glass formers [2]. By comparing fluctuations of a probe measured by Passive MR and a response of the probe measured by Active MR, we quantify the non-thermal forces in living cells.
[1] K. Nishizawa, M. Bremerich, H. Ayade, C. F. Schmidt, T. Ariga and D. Mizuno Science Advances 3, e1700318 (2017).
[2] K. Nishizawa, K. Fujiwara, M. Ikenaga, N. Nakajo, M. Yanagisawa and D. Mizuno Scientific Reports 7, 15143 (2017).
S22-3 Metabolism-dependent active diffusion in living cells
*Yujiro Sugino1, Kenji Nishizawa2, Daisuke Mizuno1
1Department of Physics, Kyushu University, 2IBDM-CNRS
The interior of cells is highly crowded with biomacromolecules that perform life-sustaining chemical reactions, collectively referred to as metabolism. In many metabolic processes, mechano-enzymes (e.g., motor proteins) generate forces by changing their shapes. Cells are therefore driven far from equilibrium by the mechanical energy derived from metabolism. Living cytoplasm maintains some fluidity whereas metabolism-deficient cytoplasm is glassy at the physiologically-relevant concentrations [1]. However, the link between metabolism and rheology in living cytoplasm is elusive.
In this study, we aim to clarify whether the living cytoplasm is fluidized in a metabolism dependent manner, by conducting microrheology experiments in ordinary and ATP-depleted living cells. By comparing spontaneous fluctuation of probe particles and their response to the applied force, the probe’s nonthermal fluctuations were quantified as the violation of the fluctuation-dissipation theorem (FDT). It was found that the probe’s non-thermal motions generally lose their memory at long time scales; the probe particles fluctuate as if they were diffusing. We discuss that the metabolism-driven forces yield microscopic structural relaxations and induce slow fluctuations which we refer to as “active diffusion”. Consistent with the idea, the active diffusion was clearly associated with the metabolism. Following the recent non-equilibrium formula, other quantities such as the non-equilibrium dissipations and force power were also derived from the FDT violation. However, they did not show a clear-cut relation with metabolism. We discuss the microscopic mechanism of active diffusion and explain why the active diffusion is a good measure for the cell metabolism.
[1] K. Nishizawa, et al., D. Mizuno, Scientific Reports, 7 (2017).
S22-4 Microrheology of a concentrated emulsion as a model cytoplasm
*Shono Inokuchi, Ryosuke Matsuoka, Daisuke Mizuno
Kyushu University
Recently, it has been shown that the cytoplasm in vitro and in vivo are glassy and exhibit anomalous viscous loss; the imaginary part of the dynamic shear modulus G (𝜔) = G ′ (𝜔) + G ′′ (𝜔) varies as the square root of frequency as G ′′ (𝜔) ∝ 𝜔0.5. Such anomalous behavior has been predicted in a theoretical model for a concentrated suspension of droplets, e.g., emulsions, foams, and microgels. Cytoplasm may then be similar to these suspensions, considering biomolecules in cells are condensed into phase-separated liquid-like droplets. However, direct experimental evidence is still lacking whether such suspensions show the predicted anomalous behavior at all. In this study, we measured G (𝜔) of the concentrated emulsions using the high-bandwidth microrheology technique, and indeed found that the emulsions show G ′′ (𝜔) ∝ 𝜔0.5 in wide range of frequencies. Besides, the frequency range that conforms to the theoretical prediction is extended towards high frequencies as the volume fraction increases or the interfacial tension decreases. By contrast, concentrated solutions of a globular biomolecule, BSA, show G ′′ (𝜔) ∝ 𝜔1. Note that the fundamental difference between BSA and liquid-like droplets may be their surface roughness. While the interfaces of liquid-like droplets are slippery, that of biomolecules are not. When they are jammed, droplets can still slip relative to each other, causing the observed anomalous relaxations. These results therefore suggest that the cytoplasm is crowded with the condensed droplets, but not with individual biomolecules.
S22-5 Non-equilibrium fluctuations in cells report on driving forces and organelle mechanics
*Kengo Nishi1,2, Sufi Raja1, An Pham1, Fred C MacKintosh3, Christoph F Schmidt1
1Duke University, 2UNC Chapel Hill, 3Rice University
Cells are complex active materials. Dispersed motor proteins drive in more or less organized manners transport, organelle dynamics, shape changes and cell movements. Forces and local material properties are difficult to measure. We have here developed a method that takes advantage of motor-generated forces deforming rod shaped elastic objects, to measure both the driving forces and the elastic properties of the rods. Examples of such rods are microtubules, intermediate filaments or externally introduced probes, such as carbon nanotubes. Their shapes are determined by the active forces, the response characteristics of the cytoplasm and the material properties of the rods. If the response of the cytoplasm is measured independently by microrheology, the other two quantities can be determined. We present a theory describing the non-equilibrium dynamics of a probe filament embedded in the cytoplasm and show how the method can be applied in HeLa cells.
S23-1 The Global Thrombosis Test
Diana Adrienne Gorog
University of Hertfordshire & Imperial College, London
The Global Thrombosis Test is a fully automated, highly physiological point-of-care test that assesses platelet reactivity, coagulation and endogenous fibrinolysis. It utilises native, non-anticoagulated blood and therefore is not subject to interference by anticoagulants. Blood introduced into the instrument is subjected to high shear flow rates, similar to that encountered in an artery at the site of significant stenosis. The instrument measures time taken to form an occlusive platelet thrombus, and in the next phase of the test, measures time taken for restart of flow due to native endogenous fibrinolysis. This is the only available test to assess endogenous fibrinolysis/thrombolysis in native whole blood.
Impaired lysis, as assessed with this technique, is a novel independent risk factor for major adverse cardiovascular events (MACE) in a number of settings. Among patients with acute coronary syndrome, impaired endogenous fibrinolysis was detected in 20% and was strongly predictive of recurrent MACE after adjustment for conventional risk factors (HR 8.03, 95% CI 4.28-15.03, P < 0.001). Amongst patients with end stage renal failure, impaired fibrinolysis occurred in 42% of patients, and was strongly associated MACE (HR 4.25, 95% CI 1.58–11.46, P = 0.004). Antiplatelet therapy appears not to alter endogenous fibrinolysis but non-vitamin K antagonist oral anticoagulants may enhance fibrinolysis. Ethnic differences have been observed between Asian and Western individuals with atrial fibrillation, showing more effective endogenous fibrinolysis in Japanese than in white Europeans on anticoagulation, which may explain the propensity to bleeding in East Asian populations on anticoagulation. This talk will summarise this technique and discuss the clinical studies supporting its use.
S23-2 Thromboelastography: Viscoelastic properties of clot formation and their clinical impact in ASCVD patients
Young-Hoon Jeong
Gyeongsang National University Changwon Hospital
Hemostasis is a finely tuned process composed of ∼80 tightly coupled biochemical reactions involving both cellular elements and plasma proteins (procoagulant, anticoagulant, and fibrinolysis). However, routinely used coagulation tests are mostly plasma-based and ignore the interaction of cellular elements and endogenous anticoagulant factors. Global hemostatic assays that reflect the interaction between these factors better represent hemostatic capacity and may assist us in differentiating the mechanism(s) associated with clot generation and associated clinical outcomes. The TEG Global Hemostasis Analyzer provides information on changes in the shear-induced elastic modulus as activated blood goes from a liquid state to a clot state in a low shear environment. This analysis provides information on thrombin generation by the coagulation, the interaction between fibrinogen/fibrin, and platelets to form a platelet – fibrin clot, the contribution of platelet count and function, and fibrinogen level and activity to clot strength, and the stability of the clot. Since this assay can identify the hypercoagulable state in patients with acute coronary syndrome and stroke, use of this test has been expanded to clinical settings such as interventional cardiology and neurology.
S23-3 Clinical trial with microfluidic platelet function assays (Anysis-200): Comparison with turbidity-based drug response assay (Verify-NOW)
*Byoung Kwon Lee1, Miney Cho1, Sehyun Shin2
1Cardiology, Department of Internal Medicine, Gangnam Severance Hospital, Yonsei University, Seoul, Korea, 2Department of Mechanical Engineering, Korea University, Seoul, Korea
S23-4 T-TAS 01: A novel flow-based system for hemostasis monitoring
Jeffrey Dahlen
Fujimori Kogyo Co., Ltd.
The T-TAS 01 System is a novel, easy-to-use, flow-based system that has been developed to measure hemostatic function under physiological flow conditions. The system uses three different assay chips to measure various aspects of hemostatic function. The T-TAS 01 PL assay passes whole blood through a collagen-coated microcapillary bed at physiological arterial shear stress to specifically measures the platelet thrombus formation process (primary hemostasis). The PL assay can be used for the assessment of overall primary hemostatic function in patients with unexplained bleeding, to assess the effects of antiplatelet therapy, and to evaluate primary hemostatic function in patients with primary hemostasis abnormalities. Because the PL assay does not use soluble platelet activation pathway-specific agonists, it provides an overall assessment of primary hemostatic function and can allow for the direct comparison of the effects of various antiplatelet therapy regimens. The T-TAS 01 AR assay measures platelet dominant fibrin-rich thrombus formation by passing whole blood through a capillary channel coated with collagen and tissue thromboplastin (combined primary and secondary hemostasis). The AR assay may be helpful in assessing bleeding risk and can provide a phenotypic assessment of the effect of direct oral anticoagulants, either with or without concomitant antiplatelet therapy. The T-TAS 01 HD assay also passes whole blood through a capillary channel coated with collagen and tissue thromboplastin and is the first assay that is able to measure thrombogenicity in patients with thrombocytopenia. The HD assay may be helpful for assessing bleeding risk and can provide a functional measurement of the improvement in thrombogenicity following platelet transfusion. The combination of the three assays allows the T-TAS 01 System to function as a complete measurement system for assessing primary hemostatic function.
S24-1 Identification of leader cells in cell migration by filopodia using computer vision
*Baasansuren Otgon1, Ganbat Danaa2, Toshiro Ohashi3
1Graduate School of Engineering, Hokkaido University, Japan, 2Open Education Center, Mongolian University of Science and Technology, Mongolia, 3Faculty of Engineering, Hokkaido University, Japan
It is generally known that collective cell migration has a crucial role during wound healing, angiogenesis and cancer metastasis. Recent studies have highlighted a type of cells, called leader cells, that forms at the leading edge and regulate the rate and the direction of the collective migration. In contrast, there is another type of cells, called follower cells, that simply follow the leader cells to migrate in a group of cells. Therefore, the identification of leader cells using modern advanced technology is vital to study the metastasis of cancer cells and the treatment of cancer. In this study, we aimed to identify leader cells by examining the structure of the plasma membrane of cells and quantitated the number of filopodia (finger-like shaped, actin-rich projections). It was hypothesized that leader cells may have a greater number of filopodia compared to follower cells. In order to characterize the filopodia of the leader cells the number of filopodia in leader and follower cells were counted by utilizing specifically designed software to identify filopodia. Using fluorescent microscopic images of both cell types we further analyzed the expression of a specific protein, cdc42, which could be highly active in the filopodia. The results showed that the number of filopodia counted in the leader cells were significantly higher than that of the follower cells. These findings indicate that the number of filopodia could be used to identify leader cells.
S24-2 Intracellular tension of osteoblast in collagen gel elicits osteocyte alignment under uniaxially-fixed boundary condition
*Jeonghyun Kim1, Keiichi Ishikawa2, Junko Sunaga2, Taiji Adachi2
1Nagoya University, 2Kyoto University
A well-aligned osteocyte network in the bone matrix is known to contribute to its mechanosensing function to modulate bone remodeling. In this study, we focused on the mechanical environment for the osteocyte in the unmineralized bone matrix, osteoid. To reconstruct this in vivo unmineralized bone matrix, we established a novel collagen culture model exerting a uniaxial boundary condition. In the uniaxial boundary condition, we fixed two opposite sides of a square collagen matrix with PDMS. As a control group, a biaxial boundary condition sample was prepared by fixing all the borders in the square collagen matrix. We then subcultured mouse osteoblast-like MC3T3-E1 cells in the uniaxially fixed boundary condition and consequently evoked the cellular alignment along the uniaxial boundary condition on the surface of the collagen matrix within 24-hours. Using blebbistatin, we showed the involvement of intracellular tension in the cellular alignment against the uniaxial mechanical boundary condition. After 4-days of culture, the cellular alignment was further distributed in the parallel direction to the uniaxial boundary condition. More than half of the cells migrated inside the matrix after 10-days of culture while the migrated cells rendered the cellular alignment inside the collagen. As a result of real-time PCR, mRNA expressions of osteoblast and osteocyte markers were up-regulated in the 10-day culture model compared to the 4-day culture model. The results altogether represented that the uniaxially-fixed boundary condition in the collagen matrix successfully induced the cellular alignment with induction of osteogenesis. Our novel model is expected to apply for the osteocyte differentiation and network studies.
S24-3 Emulating endothelial dysfunction by mimicking the microenvironment of early atherosclerotic lesions within a microfluidic chip
*Bomi Gweon1, Yujin Shin2
1Sejong University, 2Hanyang University
It is generally known that endothelial dysfunction is closely related to vascular diseases, including atherosclerosis. Many recent studies have emphasized that the microenvironment of endothelial cells (ECs) may be one of the key factors in endothelial dysfunction. Our study tried to replicate the pathophysiological phenotype and function of ECs within microchannels by mimicking the microenvironment of early atherosclerotic lesions. We focused on the extracellular matrices, such as the elevated deposition of fibronectin and the stiffer vessel walls observed in atherosclerotic plaque regions. Thus, two different environmental factors were incorporated into our microfluidic channel: fibronectin and a stiff endothelial substrate. As a result, we demonstrated that endothelial integrity on FN coated microchannels is likely to be undermined exhibiting a random orientation in response to the applied fluid flow, notable disruption of vascular endothelial cadherins (VE-cadherins), and higher endothelial permeability as opposed to that on microchannels coated with collagen, the atheroresistant vascular model.
S24-4 Enhancement and stabilization of sprouting angiogenesis by curvature-oriented behaviors of mesenchymal stem cells
*Takanori Sano1, Jun-Ichi Kawabe2, Yukiko T. Matsunaga1
1Institute of Industrial Science, The University of Tokyo, 2Asahikawa Medical University
Sprouting angiogenesis is one of the essential mechanisms for morphogenetic phenomena such as organ development, wound healing, and tissue regeneration. In regenerative medicine, therapeutic angiogenesis is a clinical solution for recovery of ischemic limbs. Mesenchymal stem cells (MSCs) have been clinically used owing to their angiogenic effects. MSCs are thought to progress angiogenesis by differentiation into endothelial cells (ECs) or via cell–cell communication using multiple protein–protein interactions (PPIs) between ECs and MSCs. However, the mechanisms how MSCs contact ECs and keep the sprouting angiogenesis active are still unclear. We show a series of EC-MSC crosstalk analyses using human umbilical vein endothelial cells (HUVECs) and MSCs derived from subcutaneous adipose tissue on a 3D in vitro model, microvessel-on-a-chip, which allows cell-to-tissue level research. On the chip, the microvessel of HUVECs was cultured for 10 days in collagen matrix where MSCs were embedded. Immunofluorescence imaging by confocal laser microscope unveiled that MSCs smoothened the surface of the microvessel and elongated the angiogenic sprouts by binding to specific microstructures of the microvessel. Three-dimensional modeling of HUVEC-MSC intersections revealed that MSCs were selectively located around protrusions or roots of angiogenic sprouts, whose Gaussian curvature was excessively high or low, respectively. Mechanistic analysis of comprehensive gene expression with a publicly available dataset suggested a few possible PPIs for EC-MSC crosstalk, including para- or juxtacrine signal transduction. Combining cellular and molecular level analyses implicated that MSCs are mechanosensitive to concave–convex surfaces on scaffold structures and that the curvature-oriented behaviors of MSCs contribute to stable sprouting angiogenesis and capillary maturation.
S24-5 Mechanism driving hydrostatic pressure-induced endothelial tube formation
Daisuke Yoshino
Tokyo University of Agriculture and Technology
Blood vessels are tightly linked with physiological and pathological events in the living body. Endothelial cells (ECs), which are constantly exposed to hemodynamic forces, change their function and morphology to maintain vascular homeostasis. Hydrostatic pressure, in particular, has been shown to elicit biophysical and biochemical responses leading to EC-mediated homeostasis. Hydrostatic pressure has a two-sided effect: it causes cell death and cell proliferation in vascular endothelial cells. However, the relationship between endothelial responses and hydrostatic pressure remains to be elucidated. Here, we propose a mechanism through which hydrostatic pressure promotes tubulogenesis, which is a crucial factor for the physiological and pathological events. We show that 50 mmHg pressure exposure transiently activates the Ras-extracellular signal regulated kinase (ERK) pathway in ECs, inducing endothelial tubulogenic responses. Activation of protein kinase C via alpha-1 adrenergic receptor and serotonin 2A receptor is essential to the pressure-induced transient activation of the Ras-ERK pathway. Pressure-dependent water efflux through water channel aquaporin 1 is a trigger for the activation of these signaling pathways. Our findings are part of the answer to the long-standing question as to how ECs sense hydrostatic pressure and convert it to intracellular signaling. Our approach could provide a basis for mechanopathology/mechanotherapy for endothelial tubulogenesis-related diseases and human health.
S25-1 Numerical analysis of the inertial migration of red blood cells in a channel
*Naoki Takeishi1, Hiroshi Yamashita1,2, Naoto Yokoyama3, Seki Masako1,2, Shigeo Wada1
1Osaka University, 2Kansai University, 3Tokyo Denki University
Particles flowing in a channel experience the drag force driving particles along their streamlines, and the shear-induced inertial lift force driving particles away from the channel center, the so-called inertial migration. This phenomenon is useful in cell sorting or separation techniques. However, much is still unknown about the relationship between cell membrane deformation and equilibrium radial position of the cell. We present a numerical analysis of a single red blood cell (RBC) in a channel. The flow is assumed as inertia. The problem is modeled as a fluid-membrane interaction in a straight circular channel, where the RBC is modeled as a biconcave capsule whose membrane follows the isotropic hyperelastic constitutive law. The internal and external fluids are modeled as an incompressible, Newtonian viscous fluid. Simulations are performed for a wide range of Reynolds number (Re), capillary number (Ca), which is ratio between the viscous shear force and membrane elastic force, and channel sizes to quantify those effects on lateral movement of RBC. Our numerical results showed that the radial equilibrium position depends on Re and Ca, and channel sizes. The radial equilibrium position is also affected by steady orientation of the cell, which is related to the initial orientation angle.
S25-2 Droplet breakup limits in simple shear flows
*Mohamed Shoieb Abdelgawad, Marco Edoardo Rosti
Okinawa Institute of Science and Technology
Deformation and breakup of droplets are paramount to numerous scientific and technological processes such as combustion systems, emulsion, enhanced oil recovery, and biological cell systems. In a dilute emulsion, where the interaction between droplets is negligible, the dynamic of a single droplet provides valuable information about the emulsion behaviour. The deformation and breakup of an initially spherical droplet suspended in a carrier fluid subjected to a shear flow by means of direct numerical simulation was studied. We initially considered a Newtonian system where the effect of capillary number, viscosity ratio between the droplet phase and the carrier phase, Reynolds number, and the degree of confinement is investigated. In addition, the simple inelastic power-law models are used for shear thickening and shear thinning fluids with different power indices. Based on the simulation results, a new capillary number was defined that governs the droplet breakup limits.
S25-3 Swelling and hemolytic behavior of human red blood cells in hypotonic fluid
*Ryoko Otomo, Ryuta Minami, Kiyoshi Bando
Kansai University
The swelling and hemolysis behavior of human red blood cells (RBCs) in hypotonic fluid has been investigated experimentally and numerically. In the experiments, a hypotonic solution is added to the RBC suspension, and then RBCs were observed to swell. By using the results of two-dimensional observation from the direction in which the concave shape can be seen, the time change in the volume of a swelling RBC was estimated. The volume became the maximum when the shape of an RBC approached a sphere, and then the cell hemolyzed after a slight contraction. Furthermore, the amount of hemoglobin in the cell was calculated based on the light absorption, which was measured by the brightness values in the image of the experimental results. It was suggested that a certain amount of hemoglobin was maintained in a RBC, but it suddenly decreased after the swelling, implying hemolysis. It was found that there was a slight difference in time between the maximum volume and the onset of hemolysis. As a method to elucidate the relationship between volume change and hemolytic behavior observed in the experiments, a one-dimensional simulation, a method similar to Ge et al. (2018), was considered. The volume change wss calculated from the osmotic pressure difference between the inside and outside of an RBC by the Kedem-Katchalsky equation. Under the same conditions as in the experiments, the calculation results reproduced the cell swelling, slight membrane stretching, and hemolysis. By comparison to the experimental results, we confirmed the validity of our calculations in terms of the volume change and discuss the hemolytic behavior of the RBC from complicated osmotic pressure changes inside and outside the cell.
S25-4 Spectral change of a stress-responsive fluorescent molecule caused by the hydrodynamic stress field of microchannel flow
*Reiko Kuriyama1, Waka Yamamoto1, Hidetsugu Kitakado2, Shohei Saito2, Kazuya Tatsumi1, Kazuyoshi Nakabe1
1Department of Mechanical Engineering and Science, Kyoto University, 2Graduate School of Science, Kyoto University
The present study measures the change in fluorescence spectrum of a stress-responsive fluorescent molecule which undergoes flow-induced conformational change over the microchannel elongational flow. Using this fluorescent molecule, we aim at developing a technique for visualizing the hydrodynamic stress field in channel flow and at the channel wall. The fluorescent molecule used as the stress probe is called FLAP (flexible and aromatic photofunctional systems), which is composed of two rigid fluorescent aromatic rings fused with a flexible eight-membered ring (cyclooctatetraene). FLAP flexibly changes its conformation between V-shape and planar shape in response to the local viscosity of the surrounding medium and external mechanical stimulus and emits blue or green fluorescence depending on its conformation. This unique fluorescence characteristic of FLAP enables us to monitor the viscosity in the curing process of epoxy resins and visualize the tensile force distribution during polymer deformation. In the present study, the response of the FLAP molecule to hydrodynamic stress was experimentally evaluated by measuring fluorescence spectrum of a FLAP solution in microchannel flow. A FLAP molecule whose molecular ends were modified with PEG (polyethylene glycol) chains was used. The FLAP-PEG molecule was added to a dimethyl sulfoxide/glycerol mixture and supplied to a sudden contraction part of the microchannel at a constant flow rate by using a syringe pump. The solution was excited by ultraviolet light (∼365 nm) and fluorescence spectra were obtained under a microscope at different flow rates in the range of 2.23–446 μL/min. This flow rate corresponds to the maximum elongational stress of approximately 100–5000 Pa at 40 μm upstream of the entrance of the contraction part. The result showed that the intensity ratio of green (494 nm) to blue (529 nm) fluorescence increased monotonically with the flow rate, which indicates that the number of FLAP molecules in planar shape increased with increasing elongational stress. Also presented are some results of two-dimensional fluorescence imaging of the contraction part.
S25-5 Segregation in shear-thickening materials
*Alessandro Monti, Marco Edoardo Rosti
Okinawa Institute of Science and Technology (OIST)
Segregation of polydispersed particles is a widely studied phenomenon for its ubiquitous occurrence (e.g., red blood cells and platelets in a blood flow or particulate material filled and stored in hoppers). In this work, we numerically investigate the appearance of the phenomena of segregation in shear-thickening suspensions in a non-uniform shear flow. We studied the behaviour of bidispersed (with volume ratio equal to 27), inertialess, non-Brownian, quasi-rigid particles densely suspended (volume fraction of 50%) in shear flow with a wavy disturbance. An efficient, in-house built, DEM software has been used to address the numerical investigation and to understand the influence on the shear-thickening behaviour.
S26-1 Important physical regulatory roles of erythrocytes on platelet adhesion under blood flow conditions
*Noriko Tamura1,2, Kazuya Shimizu3, Seiji Shiozaki2, Kazuyasu Sugiyama4, Masamitsu Nakayama2, Shinichi Goto2, Shu Takagi3, Shinya Goto2
1Department of Health and Nutrition, Niigata University of Health and Welfare, 2Department of Medicine (Cardiology), Research Center for Metabolic Disease, Tokai University School of Medicine and Tokai University Graduate School of Medicine, 3Graduate School of Engineering, The University of Tokyo, 4Department of Mechanical Science and Bioengineering, Osaka University School of Engineering Science
S26-2 Physical interactions between platelets and erythrocytes play an important role for initial platelet adhesion mediated by the interaction of glycoprotein 1b with von Willebrand factor
*Shinichi Goto1,2,3, Noriko Tamura4, Kazuya Shimizu5, Masamitsu Nakayama3, Shu Takagi5, Shinya Goto3
1Brigham and Women’s Hospital, Harvard Medical School, 2Keio University School of Medicine, 3Tokai University School of Medicine, 4Niigata University of Health and Welfare, 5The University of Tokyo
S26-3 Water–ethanol separation with tip charged carbon nanotubes
*Noriko Ono, Eiji Yamamoto, Kenji Yasuoka
Keio University
Water–ethanol separation is one of the most important processes for producing pure bioethanol. This separation is a difficult process due to their close boiling points. The most general method for the water–ethanol complete separation is azeotropic distillation. However, this distillation consumes large amounts of energy which is equal to nearly half of the energy on ethanol combustion. Therefore, improving the water–ethanol separation process is an important issue. Carbon nanotubes (CNTs) are one of the most likely materials to apply to separation membranes due to their nano pores. Recent investigations have demonstrated that water molecules in CNTs under an axial electric field forms specific helical structures and exclude alcohol molecules from CNTs using molecular dynamics (MD) simulation. Therefore, the combination of CNTs and axial electric fields can be used for water–alcohol separation. However, it is not easy to apply strong electric fields in nanoscale spaces. Hence, in this study, we used and regarded ionized functional groups at the tip of CNTs as electric field sources and investigated water–ethanol separation performance of tip functionalized (tip charged) CNTs using MD simulation.
S27-1 Inertial focusing of red blood cells suspended in blood plasma flowing through square tubes
*Masako Sugihara-Seki1,2, Saori Tanaka1
1Kansai University, 2Osaka University
Particles and biological cells suspended in laminar tube flows are known to migrate laterally due to the lift force generated by the effects of inertia, particle deformability, medium viscoelasticity, and so on. As a result, they are often observed to pass through specific locations in the downstream cross-section of the tube. In particular, rigid spherical particles in a Newtonian fluid flowing through square tubes are focused on four points located near the center of tube faces under the effect of inertia. In the present study dilute suspensions of human red blood cells (RBCs) in human blood plasma were injected into capillary square tubes, and the distribution of RBCs over the tube cross-section was observed longitudinally along the tube centerline at small distances upstream of the outlet to obtain an enface view. RBCs were found to be focused around the tube centerline at low flow rates, due the effect of RBC deformability, whereas an increase in flow rates induced the RBC focusing off-center near four points located on the diagonal of the cross-section. Additional experiments using hardened RBCs and various suspending media indicated that the RBC focusing on the diagonal could be caused by a combined effect of deformability of RBCs and viscoelasticity of plasma as well as inertia.
S27-2 Role of fluid dynamics in optical trapping
Tetsuro Tsuji
Kyoto University
Ever since the pioneering work of A. Ashkin on optical trapping, a focused laser has been a useful tool to control the motion of micro- and nanoparticles suspended in a fluid. To better understand particle motion in optical-trapping experiments, the effects of the surrounding fluid should be considered. In this talk, based on the author’s recent experimental works and the analysis using simple models, some typical cases of optical-trapping experiments, in which the effects of surrounding fluid are significant, will be introduced.
S27-3 Deformable particle suspensions
Marco Edoardo Rosti
Okinawa Institute of Science and Technology
We consider a model non-Newtonian fluid consisting of a suspension of deformable particles in a Newtonian solvent. Einstein showed in his pioneering work that the relative increase in effective viscosity is a linear function of the particle volume fraction for dilute suspensions of rigid particles. We focus here on the effect of elasticity, i.e., visco-elastic deformable particles. To tackle the problem at hand, we perform three-dimensional Direct Numerical Simulation of a plane Couette flow with a suspension of neutrally buoyant deformable viscous hyper-elastic particles. The problem is solved in a fully Eulerian methodology. We show that elasticity produces a shear-thinning effect in elastic suspensions (in comparison to rigid ones) and that it can be understood in terms of a reduction of the effective volume fraction of the suspension. Particle migration and the effect of inertia will be discussed.
S27-4 On-chip manipulation for revealing novel aspects of red blood cell mechanics
Hiroaki Ito
Chiba University
Microfluidic platforms have been intensively developed along with various techniques in microfabrication and have been applied to the quantification of the mechanical properties of small and soft viscoelastic materials such as living cells. Red blood cells (RBCs) are the typical targets for the deformability measurement by using a microchannel because they are abundant (1/3 of all the cells in the body) and various types of deformation they experience in blood vessels are physiologically important for normal blood flow. To reveal the cell mechanics under various types of deformation, we have developed an on-chip precise feedback manipulation system by combining a high-speed camera and an actuator. With this system, we controlled the period of cell deformation at a narrow constriction designed in a microchannel and analyzed the cell response upon long-time loading. Surprisingly, their response clearly shows the loading-time-dependent nonlinear viscoelasticity, which has been inaccessible under microfluidic experiments due to the high-speed but low-information-content measurements. The loading-time-dependent viscoelasticity was quantified as ratios between viscoelastic coefficients, and it therefore lacks the unit of force. To obtain the full viscoelastic information with the unit of force in a microchannel, we have recently developed another type of on-chip manipulation, which enables the onchip dynamic viscoelasticity measurement. We will also report the details of such recent developments in the talk.
S27-5 Measurement of near-wall microparticle motion under the influence of radiation pressure of an evanescent field
*Miyu Inoue, Reiko Kuriyama, Kazuya Tatsumi, Kazuyoshi Nakabe
Kyoto University
In this study, microparticle motions induced by the radiation pressure of an evanescent field were measured in a fluid for the purpose of developing a technique that can manipulate particles near the wall surface. When a dielectric particle is located in an evanescent field generated at the interface between the fluid and the wall, optical forces (called scattering and gradient forces) act on the particle in the tangential and normal directions of the interface. These forces are generated by the interaction between the electromagnetic field of the evanescent field and the particle. By using this phenomenon, particles can be manipulated selectively and collectively in the near-wall region without direct contact. In the present experiment, the evanescent field was generated by the total internal reflection of a laser beam at the interface between a borosilicate cover glass and an aqueous buffer solution, in which polystyrene microparticles (1–10 μm in diameter) were suspended. We measured the velocity of microparticles in the target spot of the evanescent field to estimate the radiation force acting on the particles and confirmed that the particles near the glass surface were driven at the speed of ∼1 μm/s in the traveling direction of the evanescent wave. Converting the particle velocity into the force acting on the particle along the tangential direction of the evanescent wave using the Stokes’s law, the force applied to the particle is approximately 0.19 pN. This value is on the same order as the theoretically obtained one. The effects of the laser light intensity, incident angle of total internal reflection, and particle size on the particle motion were evaluated. The results showed that the average particle velocity in the evanescent field increased with laser light intensity and decreased with increasing incident angle from the critical angle, which qualitatively agreed with the theoretical prediction. In addition, we found some dependence of the particle velocity on particle size. These results indicate that the radiation force applied to the microparticles in the evanescent field can be controlled by changing the optical conditions and particle specifications.
S28-1 Protein disintegration as a possible mode of protein dissociation between GP1bα and VWF in blood flow condition: Insights from steered molecular dynamic simulation
*Shinichi Goto1,2,3, Masamitsu Nakayama2, Shu Takagi4, Shinya Goto2
1Brigham and Women’s Hospital, Harvard Medical School, 2Tokai University School of Medicine, 3Keio University School of Medicine, 4Graduate School of Engineering, The University of Tokyo
S28-2 salt bridge formation between A1 Domain of von Willebrand factor and Platelet Glycoprotein (GP) Ibα by molecular dynamics simulations
*Masamitsu Nakayama, Shinichi Goto, Shinya Goto
Tokai University School of Medicine
S28-3 Finite element analysis of blood clots through visco-hyperelastic constitutive theories
*Koichiro Tashiro1,2, Yasuhiro Shobayashi2, Iku Ota1, Atsushi Hotta1
1Department of Mechanical Engineering, Keio University, 2Biomedical Solutions Inc.
Mechanical thrombectomy has become the standard treatment for patients with an acute ischemic stroke worldwide. As a thrombectomy device removes blood clots by its mechanical force, the interaction between the clot and the device significantly affects the thrombectomy performance. It is, therefore, important to analyze the interaction for the evaluation of the thrombectomy performance. A finite element method (FEM) is a powerful tool to visualize the mechanical interactions. We studied the validity of the viscoelastic theory and the numerical simulation by assessing their ability to reproduce our experimentally-obtained mechanical characteristics of clots. Soft and hard types of clot analogues were produced by changing the amount of fibrinogen. Tensile tests were conducted, and 7 types of hyperelastic models were applied to the curve fitting of the experimental data, revealing that the Mooney-Rivlin model reproduced the hyperelastic characteristics of the clots well. Additionally, the rheological models were applied to the stress relaxation data, confirming that the 3-chain Maxwell model could accurately fit the experimental viscoelastic data. The FEM in a three dimensional model with an implicit integration scheme was implemented using the identified material parameters. The FEM results matched very well with the experimental data for the both types of clot analogues, reproducing the important characteristic of blood clots, including the mechanical hysteresis during unloading, the stress dependence on the strain rate, and the time-dependent stress-decrease in the stress relaxation test. The results indicate that the presented theory and the computation method are capable of predicting the actual visco-hyperelastic characteristics of clots.
S28-4 Newly developed drug-eluting stent (DES) system for cardiovascular diseases: Hybrid nano-coating technology
*Terumitsu Hasebe1,2, Shunto Maegawa1,3, Kenta Bito1,3, Yutaka Okamoto3, Shunsuke Kamei1, Shota Yamamoto1,3, Kosuke Tomita1, Satoshi Suda1, Kazunobu Hashida1, Tomohiro Matsumoto1, Yoko Usami4,1, Yasutaka Baba4,1, Yutaka Imai1, Atsushi Hotta3
1Tokai University Hachioji Hospital, Tokai University School of Medicine, 2Keio University Hospital Clinical & Translational Research Center, 3Keio University Faculty of Science and Technology, 4Saitama Medical University International Medical Center