Aging is interrelated with changes in red blood cell parameters and functionality. In this article, we focus on red blood cells (RBCs) and provide a review of the known changes associated with the characterization of RBC deformability in aging and related pathologies. The biophysical parameters complement the commonly used biochemical parameters and may contribute to a better understanding of the aging process. The power of the deformability measurement approach is well established in clinical settings. Measuring RBCs' deformability has the advantage of relative simplicity, and it reflects the complex effects developing in erythrocytes during aging. However, aging and related pathological conditions also promote heterogeneity of RBC features and have a certain impact on the variance in erythrocyte cell properties. The possible applications of deformability as an early biophysical biomarker of pathological states are discussed, and modulating PIEZO1 as a therapeutic target is suggested. The changes in RBCs' shape can serve as a proxy for deformability evaluation, leveraging single-cell analysis with imaging flow cytometry and artificial intelligence algorithms. The characterization of biophysical parameters of RBCs is in progress in humans and will provide a better understanding of the complex dynamics of aging.

The development of accurate blood-mimicking fluids (BMFs) is essential for in vitro studies of blood contacting medical devices. Experimental data typically relies on single-phase glycerin/water solutions as substitutes to visualize simplified blood flow. These models are accurate only at high shear rates, limiting their applicability at lower shear rates. In this study, we investigated three potential BMFs, each composed of poly(sodium acrylate-co-acrylamide) hydrogel microparticles (beads) as artificial erythrocytes. Microbeads were produced using microfluidic systems (MFS) and were suspended in three plasma-like solutions: 10% and 50% (v/v) glycerol/water solutions and a Dextran40/CaCl2 solution. The BMFs were evaluated for their rheological and mechanical properties, including particle elasticity, sedimentation behavior, and shear flow analysis, to assess their suitability for mimicking blood. Rheometric measurements were performed at room temperature using a plate-plate configuration, measuring viscosity and shear stress for shear rates of 5-500 s-1. Atomic force microscopy (AFM) measurements were conducted to assess their mechanical response. The Dextran40/CaCl2-based BMF was identified as the most promising, demonstrating rheological and mechanical properties that closely align with those of human blood. This research offers a refined approach to developing blood analogs that better simulate the mechanical response and flow characteristics of blood for the validation and development of blood contacting medical devices.

The organization and dynamics of the spectrin-actin membrane cytoskeleton play a crucial role in determining the mechanical properties of red blood cells (RBC). RBC are subjected to various forces that induce deformation during blood microcirculation. Such forces also regulate membrane tension, leading to Piezo1 channel activation, which is functionally linked to RBC dehydration through calcium influx and subsequent activation of Gardos channels, ultimately resulting in variations in RBC volume. In this study, we investigated how actin instability affects Piezo1 channel gating, in relation to RBC deformation and mechanical properties, using micropipette aspiration and optical tweezers. Actin instability, induced by 0.5 μM Cytochalasin-D (Cyt-D), led to a 22% reduction in the activation pressure. Additionally, we observed a decreasing trend in Young's modulus, membrane tension, and viscosity. By measuring the time required for cell shape recovery after deformation in an optical trap, we found that Cyt-D-treated RBC took approximately 14% longer to recover compared to untreated cells. The bimodal imaging feature of our experimental approach allowed us to simultaneously measure and correlate activation pressure with mechanical properties at the single-cell level. A significant correlation was found between these parameters in both treated and untreated RBC. Our findings demonstrate the influence of actin instability on both Piezo1 activation and RBC mechanics. These results offer new insights into the interplay between F-actin and Piezo1 in RBC mechanobiology.

Cell elasticity serves as a crucial physical biomarker that reflects changes in cellular structures and physiological states, providing key insights into cell behaviors. It links mechanical properties to biological function, highlighting its importance for understanding cell health and advancing biomedical research. Microfluidic technologies, with their capabilities for precise manipulation and high-throughput analysis, have significantly advanced the measurement of cell elasticity and elasticity-based cell sorting. This paper presents a comprehensive overview of advanced microsystems for assessing cell elasticity, discussing their advantages and limitations. The biomedical applications of elasticity-based sorting are highlighted, including cell classification, clinical diagnosis, drug screening, and stem cell differentiation prediction. The paper addresses the current challenges in the field, such as limited measurement efficiency and scalability, and explores future research directions, including the development of automated, high-throughput systems and the integration of elasticity measurements into practical biomedical applications. These advancements aim to deepen our understanding of cellular mechanics, improve diagnostic precision, and foster the development of novel therapeutic strategies. Ultimately, this work emphasizes the potential of cell elasticity as a key parameter in advancing disease diagnosis and therapeutic research.

Hereditary spherocytosis is a common hemolytic anemia with different severity. The causes of hereditary spherocytosis are mutations in genes that encode red blood cell (RBC) membrane and cytoskeletal proteins, including ankyrin-1, Band 3 (or AE1), α spectrin, β spectrin, and protein 4.2. Molecular defects in these proteins decrease membrane integrity, leading to vesiculation, decreased membrane surface area, and reduced deformability of the cells. Eventually, this leads to the trapping the abnormal RBCs (spherocytes) in the spleen. In most severe cases, splenectomy may be necessary to prevent general RBC collapse during the passage of RBCs through the narrow slits of venous sinuses in the spleen. The clinical benefit of splenectomy results from elimination the primary site of RBC damage and destruction. Splenectomy is a curative approach but can cause complications and should be undertaken after examination by various laboratory approaches. Splenectomy does not correct most genetically determined membrane abnormalities in erythrocytes in patients with hereditary spherocytosis. The transformation of biconcave erythrocytes into spherocytes continues, although to a lesser degree than before surgery. Nevertheless, splenectomy increases the lifespan of red cells, significantly reducing the severity of anemia and improving many physiological signs of HS.

Stomatocyte-discocyte-echinocyte (SDE) transformations in human red blood cells (RBCs) have significant influences on blood dynamics and related disorders. The mechanical properties of the RBC membrane, such as shear modulus and bending elasticity, play crucial roles in determining RBC shapes. Recent biophysical findings reveal that building a comprehensive model capable of describing SDE shape transformations is a challenging problem. Based on dissipative particle dynamics, this study develops a two-component RBC model considering the detachment between the lipid bilayer and cytoskeleton, as well as the cytoskeletal reorganization during echinocyte formation. This model is validated by comparing RBCs' geometric shape and the apparent membrane tension with previous experimental measurements. Results indicate that a complete SDE sequence represented by six typical shapes can be obtained by modulating the model's mechanical and geometric parameters. Furthermore, a phase diagram based on reduced variables is obtained using principal-component analysis, demonstrating the phase transformations among SDE shapes. Our result suggests that the transformation from discocyte to stomatocyte is primarily influenced by dimensionless bending rigidity, whereas, during echinocyte formation, three key variables, i.e., dimensionless bending rigidity, targeting cytoskeleton shrinkage ratio, and connecting pattern, have joint impacts on the formation of spicules or bumps and the development of the cytoskeletal framework. The present two-component RBC model and the associated findings provide a perspective for a deeper understanding of the SDE transformation mechanism. This framework offers new insights into biological science and potential applications in the field of biomedical engineering.

In physiological conditions, red blood cells (RBCs) demonstrate remarkable deformability, allowing them to undergo considerable deformation when passing through the microcirculation. However, this deformability is compromised in Type 1 diabetes mellitus (T1DM) and related pathological conditions. This study aims to investigate the biomechanical properties of RBCs in T1DM patients, focusing on identifying significant mechanical alterations associated with microvascular complications (MCs). We conducted a case-control study involving 38 T1DM subjects recruited from the Diabetes Care Unit at Fondazione Policlinico Gemelli Hospital, comprising 22 without MCs (control group) and 16 with MCs (pathological group). Atomic Force Microscopy was employed to assess RBC biomechanical properties in a liquid environment. We observed significant RBC stiffening in individuals with MCs, particularly during large indentations that mimic microcirculatory deformations. Univariate analysis unveiled significant differences in RBC stiffness (median difference 0.0006 N/m, p = 0.012) and RBC counts (median difference -0.39 × 1012/L, p = 0.009) between the MC and control groups. Bivariate logistic regression further demonstrated that combining these parameters could effectively discriminate between MC and non-MC conditions, achieving an AUC of 0.82 (95% CI: 0.67-0.97). These findings reveal the potential of RBC biomechanical properties as diagnostic and monitoring tools in diabetes research. Exploring RBC mechanical alterations may lead to the development of novel biomarkers, which, in combination with clinical markers, could facilitate the early diagnosis of diabetes-related complications.

Hemoglobin and red blood cells (erythrocytes) have been studied extensively from the perspective of life and biomedical sciences. However, no analysis of hemoglobin and red blood cells from the perspective of chemical thermodynamics has been reported in the literature. Such an analysis would provide an insight into their structure and turnover from the aspect of biothermodynamics and bioenergetics. In this paper, a biothermodynamic analysis was made of hemoglobin and red blood cells. Molecular formulas, empirical formulas, biosynthesis reactions, and thermodynamic properties of formation and biosynthesis were determined for the alpha chain, beta chain, heme B, hemoglobin and red blood cells. Empirical formulas and thermodynamic properties of hemoglobin were compared to those of other biological macromolecules, which include proteins and nucleic acids. Moreover, the energetic requirements of biosynthesis of hemoglobin and red blood cells were analyzed. Based on this, a discussion was made of the specific structure of red blood cells (i.e. no nuclei nor organelles) and its role as an evolutionary adaptation for more energetically efficient biosynthesis needed for the turnover of red blood cells.

The clearance of senescent and altered red blood cells (RBCs) in the red pulp of the human spleen involves sequential processes of prefiltration, filtration, and postfiltration. While prior work has elucidated the mechanisms underlying the first two processes, biomechanical processes driving the postfiltration phagocytosis of RBCs retained at interendothelial slits (IES) are still poorly understood. We present here a unique computational model of macrophages to study the role of cell biomechanics in modulating the kinetics of phagocytosis of aged and diseased RBCs retained in the spleen. After validating the macrophage model using in vitro phagocytosis experiments, we employ it to probe the mechanisms underlying the kinetics of phagocytosis of mechanically altered RBCs, such as heated RBCs and abnormal RBCs in hereditary spherocytosis (HS) and sickle cell disease (SCD). Our simulations show pronounced deformation of the flexible and healthy RBCs in contrast to minimal shape changes in altered RBCs. Simulations also show that less deformable RBCs are engulfed faster and at lower adhesive strength than flexible RBCs, consistent with our experimental measurements. This efficient sensing and engulfment by macrophages of stiff RBCs retained at IES are expected to temper splenic congestion, a common pathogenic process in malaria, HS, and SCD. Altogether, our combined computational and in vitro experimental studies suggest that mechanical alterations of retained RBCs may suffice to enhance their phagocytosis, thereby adapting the kinetics of their elimination to the kinetics of their mechanical retention, an equilibrium essential for adequately cleaning the splenic filter to preserve its function.

Biological and living organisms sense and process information from their surroundings, typically having access only to a subset of external observables for a limited amount of time. In this Letter, we uncover how biological systems can exploit these accessible degrees of freedom to transduce information from the inaccessible ones with a limited energy budget. We find that optimal transduction strategies may boost information harvesting over the ideal case in which all degrees of freedom are known, even when only finite-time trajectories are observed, at the price of higher dissipation. We apply our results to red blood cells, inferring the implemented transduction strategy from membrane flickering data and shedding light on the connection between mechanical stress and transduction efficiency. Our framework offers novel insights into the adaptive strategies of biological systems under nonequilibrium conditions.

Vascular diseases are widespread, and sometimes such life-threatening medical disorders cause abnormal blood flow, blood particle damage, changes to flow dynamics, restricted blood flow, and other adverse effects. The study of vascular flow is crucial in clinical practice because it can shed light on the causes of stenosis, aneurysm, blood cancer, and many other such diseases, and guide the development of novel treatments and interventions. Microfluidics and computational fluid dynamics (CFDs) are two of the most promising new tools for investigating these phenomena. When compared to conventional experimental methods, microfluidics offers many benefits, including lower costs, smaller sample quantities, and increased control over fluid flow and parameters. In this paper, we address the strengths and weaknesses of computational and experimental approaches utilizing microfluidic devices to investigate the rheological properties of blood, the forces of action causing diseases related to cardiology, provide an overview of the models and methodologies of experiments, and the fabrication of devices utilized in these types of research, and portray the results achieved and their applications. We also discuss how these results can inform clinical practice and where future research should go. Overall, it provides insights into why a combination of both CFDs, and experimental methods can give even more detailed information on disease mechanisms recreated on a microfluidic platform, replicating the original biological system and aiding in developing the device or chip itself.

A series of successes in RNA interference (RNAi) therapies for liver diseases using lipid nanoparticles and N-acetylgalactosamine have heralded a current era of RNA therapeutics. However, alternative delivery strategies are required to take RNAi out of the comfort zone of hepatocytes. Here we report SIRPα IgV/anti-CD47 siRNA (vS-siCD47) conjugates that selectively and persistently disrupt the antiphagocytic CD47/SIRPα axis in solid tumors. Conjugation of the SIRPα IgV domain protein to siRNAs enables tumor dash through CD47-mediated erythrocyte piggyback, primarily blocking the physical interaction between CD47 on cancer cells and SIRPα on phagocytes. After internalization of the vS-siCD47 conjugates within cancer cells, the detached free-standing anti-CD47 siRNAs subsequently attack CD47 through the RNAi mechanism. The dual-action approach of the vS-siCD47 conjugate effectively overcomes the "don't eat me" barrier and stimulates phagocyte-mediated tumor destruction, demonstrating a highly selective and potent CD47-blocking immunotherapy. This delivery strategy, employing IgV domain protein-siRNA conjugates with a dual mode of target suppression, holds promise for expanding RNAi applications beyond hepatocytes and advancing RNAi-based cancer immunotherapies for solid tumors.

Existing hemolysis algorithms are often constructed for laminar flows that expose red blood cells (RBCs) to a constant rate of shear. It remains an open question whether such models are applicable to turbulent flows, where there is a significant variation in shear rate along cell trajectories. To evaluate the effect of turbulence on hemolysis, we perform cell-resolved simulations of isolated RBCs in turbulent channel flow atRe τ = 180 and 360 and compare them against the results obtained from laminar flow simulations at an equivalent wall shear stress. The RBCs are modeled as isolated cells in an unbounded domain with the viscosity of the bulk fluid used for the surrounding fluid. This comparison shows that, while the laminar flow generally induces greater stretch in the cell in a time-averaged sense, cells experience an overall larger deformation in turbulence. This difference is attributed to extreme events in turbulence that occasionally create bursts of high shear conditions, which, consequently, induce a large deformation in the cells. Associating damage with the most extreme deformation regimes, we observe that, in the worst case, the turbulent flow can produce deformation in the cell that is higher than the absolute maximum value in the analogous laminar case approximately 14% of the time. Additionally, theRe τ = 180 universally induced greater deformation in the cells than theRe τ = 360 case, suggesting that increasing the range of scales in the flow does not necessarily yield greater deformation when all other parameters are kept constant. A strong direct correlation ( R > 0.8 ) between shear rate and deformation metrics was observed in turbulence. The correlation against Q -criterion is inverse and weaker ( R ≈ - 0.26 ) , but once the shear contribution is subtracted, it improves in terms of areal dilatation ( R ≈ - 0.6 ) .

Red blood cells (RBC), the primary carriers of oxygen in the body, play a crucial role across several biomedical applications, while also being an essential model system of a deformable object in the microfluidics and soft matter fields. However, RBC behavior in viscoelastic liquids, which holds promise in enhancing microfluidic diagnostic applications, remains poorly studied. We here show that using viscoelastic polymer solutions as a suspending carrier causes changes in the clustering and shape of flowing RBC in microfluidic flows when compared to a standard Newtonian suspending liquid. Additionally, when the local RBC concentration increases to a point where hydrodynamic interactions take place, we observe the formation of equally-spaced RBC structures, resembling the viscoelasticity-driven ordered particles observed previously in the literature, thus providing the first experimental evidence of viscoelasticity-driven cell ordering. The observed RBC ordering, unaffected by polymer molecular architecture, persists as long as the surrounding medium exhibits shear-thinning, viscoelastic properties. Complementary numerical simulations reveal that viscoelasticity-induced repulsion between RBCs leads to equidistant structures, with shear-thinning modulating this effect. Our results open the way for the development of new biomedical technologies based on the use of viscoelastic liquids while also clarifying fundamental aspects related to multibody hydrodynamic interactions in viscoelastic microfluidic flows.

Red blood cells possess a singular mechanobiology, enabling efficient navigation through capillaries smaller than their own size. Their plasma membrane exhibits non-equilibrium shape fluctuation, often reported as enhanced flickering activity. Such active membrane motion is propelled by motor proteins that mediate interactions between the spectrin skeleton and the lipid bilayer. However, modulating the flickering in living red blood cells without permanently altering their mechanical properties represents a significant challenge. In this study, we developed holographic optical tweezers to generate a force field distributed along the equatorial membrane contour of individual red blood cells. In free-standing red blood cells, we observed heterogeneous flickering activity, attributed to localized membrane kickers. By employing holographic optical forces, these active kickers can be selectively halted under minimal invasion. Our findings shed light on the dynamics of membrane flickering and established a manipulation tool that could open new avenues for investigating mechanotransduction processes in living cells.

Biomechanical attributes have emerged as novel markers, providing a reliable means to characterize cellular and subcellular fractions. Numerous studies have identified correlations between these factors and patients' medical status. However, the absence of a thorough overview impedes their applicability in contemporary state-of-the-art therapeutic strategies. In this context, we provide a comprehensive analysis of the dimensions, configuration, rigidity, density, and electrical characteristics of normal and abnormal circulating cells. Subsequently, the discussion broadens to encompass subcellular bioparticles, such as extracellular vesicles (EVs) enriched either from blood cells or other tissues. Notably, cell sizes vary significantly, from 2 μm for platelets to 25 μm for circulating tumor cells (CTCs), enabling the development of size-based separation techniques, such as microfiltration, for specific diagnostic and therapeutic applications. Although cellular density is relatively constant among different circulating bioparticles, it allows for reliable density gradient centrifugation to isolate cells without altering their native state. Additionally, variations in EV surface charges (-6.3 to -45 mV) offer opportunities for electrophoretic and electrostatic separation methods. The distinctive mechanical properties of abnormal cells, compared to their normal counterparts, present an exceptional opportunity for diverse medical and biotechnological approaches. This review also aims to provide a holistic view of the current understanding of popular techniques in this domain that transcend conventional boundaries, focusing on early harvesting of malignant cells from body fluids, designing effective therapeutic options, cell targeting, and resonating with tissue and genetic engineering principles.

Type 2 diabetes mellitus (T2DM) is a prevalent and debilitating disease with numerous health risks, including cardiovascular diseases, kidney dysfunction, and nerve damage. One important aspect of T2DM is its association with the abnormal morphology of red blood cells (RBCs), which leads to increased blood viscosity and impaired blood flow. Therefore, evaluating the mechanical properties of RBCs is crucial for understanding the role of T2DM in cellular deformability. This provides valuable insights into disease progression and potential diagnostic applications. In this study, we developed an open micro-electro-fluidic (OMEF) biochip technology based on dielectrophoresis (DEP) to assess the deformability of RBCs in T2DM. The biochip facilitates high-throughput single-cell RBC stretching experiments, enabling quantitative measurements of the cell size, strain, stretch factor, and post-stretching relaxation time. Our results confirm the significant impact of T2DM on the deformability of RBCs. Compared to their healthy counterparts, diabetic RBCs exhibit ∼27% increased size and ∼29% reduced stretch factor, suggesting potential biomarkers for monitoring T2DM. The observed dynamic behaviors emphasize the contrast between the mechanical characteristics, where healthy RBCs demonstrate notable elasticity and diabetic RBCs exhibit plastic behavior. These differences highlight the significance of mechanical characteristics in understanding the implications for RBCs in T2DM. With its ∼90% sensitivity and rapid readout (ultimately within a few minutes), the OMEF biochip holds potential as an effective point-of-care diagnostic tool for evaluating the deformability of RBCs in individuals with T2DM and tracking disease progression.

Malaria is one of the serious health concerns worldwide as it remains a clinical challenge due to the complex life cycle of the malaria parasite and the morphological changes it undergoes during infection. The malaria parasite multiplies rapidly and spreads in the population by changing its alternative hosts. These various morphological stages of the parasite in the human host cause clinical symptoms (anemia, fever, and coma). These symptoms arise due to the preprogrammed biology of the parasite in response to the human pathophysiological response. Thus, complete elimination becomes one of the major health challenges. Although malaria vaccine(s) are available in the market, they still contain to cause high morbidity and mortality. Therefore, an approach for eradication is needed through the exploration of novel molecular targets by tracking the epidemiological changes the parasite adopts. This review focuses on the various novel molecular targets.

Pathological processes often change the mechanical properties of cells. Increased rigidity could be a marker of cellular malfunction. Erythrocytes are a type of cell that deforms to squeeze through tiny capillaries; changes in their rigidity can dramatically affect their functionality. Furthermore, differences in the homeostatic elasticity of the cell can be used as a tool for diagnosis and even for choosing the adequate treatment for some illnesses. More accurate types of equipment needed to study biomechanical phenomena at the single-cell level are very costly and thus out of reach for many laboratories around the world. This study presents a simple and low-cost technique to study the rigidity of red blood cells (RBCs) through the application of electric fields in a hand-made microfluidic chamber that uses a capacitor principle. As RBCs are deformed with the application of voltage, cells are observed under a light microscope. From mechanical force vs. deformation data, the elastic constant of the cells is determined. The results obtained with the capacitor-based method were compared with those obtained using optical tweezers, finding good agreement. In addition, P. falciparum-infected erythrocytes were tested with the electric field applicator. Our technique provides a simple means of testing the mechanical properties of individual cells.

The ability to change shape is essential for the proper functioning of red blood cells (RBCs) within the microvasculature. The shape of RBCs significantly influences blood flow and has been employed in microfluidic lab-on-a-chip devices, serving as a diagnostic biomarker for specific pathologies and enabling the assessment of RBC deformability. While external flow conditions, such as the vessel size and the flow velocity, are known to impact microscale RBC flow, our comprehensive understanding of how their shape-adapting ability is influenced by channel confinement in biomedical applications remains incomplete. This study explores the impact of various rectangular and square channels, each with different confinement and aspect ratios, on the in vitro RBC flow behavior and characteristic shapes. We demonstrate that rectangular microchannels, with a height similar to the RBC diameter in combination with a confinement ratio exceeding 0.9, are required to generate distinctive well-defined croissant and slipper-like RBC shapes. These shapes are characterized by their equilibrium positions in the channel cross section, and we observe a strong elongation of both stable shapes in response to the shear rate across the different channels. Less confined channel configurations lead to the emergence of unstable other shape types that display rich shape dynamics. Our work establishes an experimental framework to understand the influence of channel size on the single-cell flow behavior of RBCs, providing valuable insights for the design of biomicrofluidic single-cell analysis applications.

Out-of-plane fluctuations, also known as stochastic displacements, of biological membranes play a crucial role in regulating many essential life processes within cells and organelles. Despite the availability of various methods for quantifying membrane dynamics, accurately quantifying complex membrane systems with rapid and tiny fluctuations, such as mitochondria, remains a challenge. In this work, we present a methodology that combines metal/graphene-induced energy transfer (MIET/GIET) with fluorescence correlation spectroscopy (FCS) to quantify out-of-plane fluctuations of membranes with simultaneous spatiotemporal resolution of approximately one nanometer and one microsecond. To validate the technique and spatiotemporal resolution, we measure bending undulations of model membranes. Furthermore, we demonstrate the versatility and applicability of MIET/GIET-FCS for studying diverse membrane systems, including the widely studied fluctuating membrane system of human red blood cells, as well as two unexplored membrane systems with tiny fluctuations, a pore-spanning membrane, and mitochondrial inner/outer membranes.

Red blood cells (RBCs) undergo a progressive morphological transformation from smooth biconcave discocytes into rounder echinocytes with spicules on their surface during cold storage. The echinocytic morphology impacts RBCs' ability to flow through narrow sections of the circulation and therefore transfusion of RBC units with a high echinocytic content are thought to have a reduced efficiency. We use an optical tweezers-based technique where we directly trap and measure linear stiffness of RBCs under stress without the use of attached spherical probe particles or microfluidic flow to induce shear. We study RBC deformability with over 50 days of storage performing multiple stretches in blood plasma (serum with cold agglutinins removed to eliminate clotting). In particular, we find that discocytes and echinocytes do not show significant changes in linear stiffness in the small strain limit (∼ 20 % change in length) up to day 30 of the storage period, but do find differences between repeated stretches. By day 50 the linear stiffness of discocytes had increased to approximately that measured for echinocytes throughout the entire period of measurements. These changes in stiffness corresponded to recorded morphological changes in the discocytes as they underwent storage lesion. We believe our holographic trapping and direct measurement technique has applications to directly control and quantify forces that stretch other types of cells without the use of attached probes.

Contraction of blood clots plays an important role in blood clotting, a natural process that restores hemostasis and regulates thrombosis in the body. Upon injury, a chain of events culminate in the formation of a soft plug of cells and fibrin fibers attaching to wound edges. Platelets become activated and apply contractile forces to shrink the overall clot size, modify clot structure, and mechanically stabilize the clot. Impaired blood clot contraction results in unhealthy volumetric, mechanical, and structural properties of blood clots associated with a range of severe medical conditions for patients with bleeding and thrombotic disorders. Due to the inherent mechanical complexity of blood clots and a confluence of multiple interdependent factors governing clot contraction, the mechanics and dynamics of clot contraction and the interactions with red blood cells (RBCs) remain elusive. Using an experimentally informed, physics-based mesoscale computational model, we probe the dynamic interactions among platelets, fibrin polymers, and RBCs, and examine the properties of contracted blood clots. Our simulations confirm that RBCs strongly affect clot contraction. We find that RBC retention and compaction in thrombi can be solely a result of mechanistic contraction of fibrin mesh due to platelet activity. Retention of RBCs hinders clot contraction and reduces clot contractility. Expulsion of RBCs located closer to clot outer surface results in the development of a dense fibrin shell in thrombus clots commonly observed in experiments. Our simulations identify the essential parameters and interactions that control blood clot contraction process, highlighting its dependence on platelet concentration and the initial clot size. Furthermore, our computational model can serve as a useful tool in clinically relevant studies of hemostasis and thrombosis disorders, and post thrombotic clot lysis, deformation, and breaking.

Hemolytic-uremic syndrome (HUS), mostly secondary to infectious diseases, is a common cause of acute kidney injury in children. It is characterized by progressive acute kidney failure due to severe thrombotic microangiopathy, associated with nonimmune, Coombs-negative hemolytic anemia and thrombocytopenia. HUS is caused mostly by Shiga toxin-producing E. Coli, and to a lesser extent by Streptococcus pneumonia. In Streptococcus pneumonia HUS (pHUS), bacterial neuraminidase A exposes masked O-glycan sugar residues on erythrocytes, known as the T antigen, triggering a complement cascade causing thrombotic microangiopathy. Atypical HUS (aHUS) is a life-threatening genetic form of the disease, whose molecular mechanism is only partly understood. Through genetic studies, we demonstrate a novel X-linked form of aHUS that is caused by a de-novo missense mutation in C1GALT1C1:c.266 C > T,p.(T89I), encoding a T-synthase chaperone essential for the proper formation and incorporation of the T antigen on erythrocytes. We demonstrate the presence of exposed T antigen on the surface of mutant erythrocytes, causing aHUS in a mechanism similar to that suggested in pHUS. Our findings suggest that both aHUS caused by mutated C1GALT1C1 and pHUS are mediated by the lectin-complement-pathway, not comprehensively studied in aHUS. We thus delineate a shared molecular basis of aHUS and pHUS, highlighting possible therapeutic opportunities.

The protozoa, Toxoplasma gondii and Plasmodium spp., are preeminent members of the Apicomplexa parasitic phylum in large part due to their public health and economic impact. Hence, they serve as model unicellular eukaryotes with which to explore the repertoire of molecular and cellular strategies that specific developmental morphotypes deploy to timely adjust to their host(s) in order to perpetuate. In particular, host tissue- and cell-invasive morphotypes termed zoites alternate extracellular and intracellular lifestyles, thereby sensing and reacting to a wealth of host-derived biomechanical cues over their partnership. In the recent years, biophysical tools especially related to real time force measurement have been introduced, teaching us how creative are these microbes to shape a unique motility system that powers fast gliding through a variety of extracellular matrices, across cellular barriers, in vascular systems or into host cells. Equally performant was this toolkit to start illuminating how parasites manipulate their hosting cell adhesive and rheological properties to their advantage. In this review, besides highlighting major discoveries along the way, we discuss the most promising development, synergy, and multimodal integration in active noninvasive force microscopy methods. These should in the near future unlock current limitations and allow capturing, from molecules to tissues, the many biomechanical and biophysical interplays over the dynamic host and microbe partnership.

The flow dynamics of red blood cells in vivo in blood capillaries and in vitro in microfluidic channels is complex. Cells can obtain different shapes such as discoid, parachute, slipper-like shapes and various intermediate states depending on flow conditions and their viscoelastic properties. We use artificial intelligence based analysis of red blood cells (RBCs) in an oscillating microchannel to distinguish healthy red blood cells from red blood cells treated with formaldehyde to chemically modify their viscoelastic behavior. We used TensorFlow to train and validate a deep learning model and achieved a testing accuracy of over 97%. This method is a first step to a non-invasive, label-free characterization of diseased red blood cells and will be useful for diagnostic purposes in haematology labs. This method provides quantitative data on the number of affected cells based on single cell classification.

Neurodegenerative disorders (NDDs) are complex, multifactorial disorders with significant social and economic impact in today's society. NDDs are predicted to become the second-most common cause of death in the next few decades due to an increase in life expectancy but also to a lack of early diagnosis and mainly symptomatic treatment. Despite recent advances in diagnostic and therapeutic methods, there are yet no reliable biomarkers identifying the complex pathways contributing to these pathologies. The development of new approaches for early diagnosis and new therapies, together with the identification of non-invasive and more cost-effective diagnostic biomarkers, is one of the main trends in NDD biomedical research. Here we summarize data on peripheral biomarkers, biofluids (cerebrospinal fluid and blood plasma), and peripheral blood cells (platelets (PLTs) and red blood cells (RBCs)), reported so far for the three most common NDDs-Alzheimer's disease (AD), Parkinson's disease (PD), and amyotrophic lateral sclerosis (ALS). PLTs and RBCs, beyond their primary physiological functions, are increasingly recognized as valuable sources of biomarkers for NDDs. Special attention is given to the morphological and nanomechanical signatures of PLTs and RBCs as biophysical markers for the three pathologies. Modifications of the surface nanostructure and morphometric and nanomechanical signatures of PLTs and RBCs from patients with AD, PD, and ALS have been revealed by atomic force microscopy (AFM). AFM is currently experiencing rapid and widespread adoption in biomedicine and clinical medicine, in particular for early diagnostics of various medical conditions. AFM is a unique instrument without an analog, allowing the generation of three-dimensional cell images with extremely high spatial resolution at near-atomic scale, which are complemented by insights into the mechanical properties of cells and subcellular structures. Data demonstrate that AFM can distinguish between the three pathologies and the normal, healthy state. The specific PLT and RBC signatures can serve as biomarkers in combination with the currently used diagnostic tools. We highlight the strong correlation of the morphological and nanomechanical signatures between RBCs and PLTs in PD, ALS, and AD.

Blood image intensity has been used to detect erythrocyte sedimentation rate (ESR). However, it does not give information on the biophysical properties of blood samples under continuous ESR. In this study, to quantify mechanical variations of blood under continuous ESR, blood shear stress and blood image intensity were obtained by analyzing blood flows in the capillary channel. A blood sample is loaded into a driving syringe to demonstrate the proposed method. The blood flow rate is set in a periodic on-off pattern. A blood sample is then supplied into a capillary chip, and microscopic blood images are captured at specific intervals. Blood shear stress is quantified from the interface of the bloodstream in the coflowing channel. τ0 is defined as the maximum shear stress obtained at the first period. Simultaneously, ESRτ is then obtained by analyzing temporal variations of blood shear stress for every on period. AII is evaluated by analyzing the temporal variation of blood image intensity for every off period. According to the experimental results, a shorter period of T = 4 min and no air cavity contributes to the high sensitivity of the two indices (ESRτ and AII). The τ0 exhibits substantial differences with respect to hematocrits (i.e., 30-50%) as well as diluents. The ESRτ and AII showed a reciprocal relationship with each other. Three suggested properties represented substantial differences for suspended blood samples (i.e., hardened red blood cells, different concentrations of dextran solution, and fibrinogen). In conclusion, the present method can detect variations in blood samples under continuous ESR effectively.

"Artificial intelligence" and "big data" increasingly take the step from just being interesting concepts to being relevant or even part of our lives. This general statement holds also true for transfusion medicine. Besides all advancements in transfusion medicine, there is not yet an established red blood cell quality measure, which is generally applied.

We highlight the usefulness of big data in transfusion medicine. Furthermore, we emphasize in the example of quality control of red blood cell units the application of artificial intelligence.

A variety of concepts making use of big data and artificial intelligence are readily available but still await to be implemented into any clinical routine. For the quality control of red blood cell units, clinical validation is still required.

Blood elasticity is quantified using a single compliance model by analyzing pulsatile blood flow. However, one compliance coefficient is influenced substantially by the microfluidic system (i.e., soft microfluidic channels and flexible tubing). The novelty of the present method comes from the assessment of two distinct compliance coefficients, one for the sample and one for the microfluidic system. With two compliance coefficients, the viscoelasticity measurement can be disentangled from the influence of the measurement device. In this study, a coflowing microfluidic channel was used to estimate blood viscoelasticity. Two compliance coefficients were suggested to denote the effects of the polydimethylsiloxane (PDMS) channel and flexible tubing (C1), as well as those of the RBC (red blood cell) elasticity (C2), in a microfluidic system. On the basis of the fluidic circuit modeling technique, a governing equation for the interface in the coflowing was derived, and its analytical solution was obtained by solving the second-order differential equation. Using the analytic solution, two compliance coefficients were obtained via a nonlinear curve fitting technique. According to the experimental results, C2/C1 is estimated to be approximately 10.9-20.4 with respect to channel depth (h = 4, 10, and 20 µm). The PDMS channel depth contributed simultaneously to the increase in the two compliance coefficients, whereas the outlet tubing caused a decrease in C1. The two compliance coefficients and blood viscosity varied substantially with respect to homogeneous hardened RBCs or heterogeneous hardened RBCs. In conclusion, the proposed method can be used to effectively detect changes in blood or microfluidic systems. In future studies, the present method can contribute to the detection of subpopulations of RBCs in the patient's blood.

The invasion of a merozoite into an erythrocyte by membrane wrapping is a hallmark of malaria pathogenesis. The invasion involves biomechanical interactions whereby the merozoite exerts actomyosin-based forces to push itself into and through the erythrocyte membrane while concurrently inducing biochemical damage to the erythrocyte membrane. Whereas the biochemical damage process has been investigated, the detailed mechanistic understanding of the invasion mechanics remains limited. Thus, the current study aimed to develop a mathematical model describing the mechanical factors involved in the merozoite invasion into an erythrocyte and explore the invasion mechanics. A shell theory model was developed comprising constitutive, equilibrium and governing equations of the deformable erythrocyte membrane to predict membrane mechanics during the wrapping of an entire non-deformable ellipsoidal merozoite. Predicted parameters include principal erythrocyte membrane deformations and stresses, wrapping and indentation forces, and indentation work. The numerical investigations considered two limits for the erythrocyte membrane deformation during wrapping (4% and 51% areal strain) and erythrocyte membrane phosphorylation (decrease of membrane elastic modulus from 1 to 0.5 kPa). For an intact erythrocyte, the maximum indentation force was 1 and 8.5 pN, and the indentation work was 1.92 × 10^-18 and 1.40 × 10^-17 J for 4% and 51% areal membrane strain. Phosphorylation damage in the erythrocyte membrane reduced the required indentation work by 50% to 0.97 × 10^-18 and 0.70 × 10^-17 J for 4% and 51% areal strain. The current study demonstrated the developed model's feasibility to provide new knowledge on the physical mechanisms of the merozoite invasion process that contribute to the invasion efficiency towards the discovery of new invasion-blocking anti-malaria drugs.

Cellular mechanical property analysis reflecting the physiological and pathological states of cells plays a crucial role in assessing the quality of stored blood. However, its complex equipment needs, operation difficulty, and clogging issues hinder automated and rapid biomechanical testing. Here, we propose a promising biosensor assisted by magnetically actuated hydrogel stamping to fulfill it. The flexible magnetic actuator triggers the collective deformation of multiple cells in the light-cured hydrogel, and it allows for on-demand bioforce stimulation with the advantages of portability, cost-effectiveness, and simplicity of operation. The magnetically manipulated cell deformation processes are captured by the integrated miniaturized optical imaging system, and the cellular mechanical property parameters are extracted from the captured images for real-time analysis and intelligent sensing. In this work, 30 clinical blood samples with different storage durations (<14 days and >14 days) were tested. A deviation of 3.3% in the differentiation of blood storage durations by this system compared to physician annotation demonstrated its feasibility. This system should broaden the application of cellular mechanical assays in diverse clinical settings.

Microgravity is proven to impact a wide range of human physiology, from stimulating stem cell differentiation to confounding cell health in bones, skeletal muscles, and blood cells. The research in this arena is progressively intensified by the increasing promises of human spaceflights. Considering the limited access to spaceflight, ground-based microgravity-simulating platforms have been indispensable for microgravity-biology research. However, they are generally complex, costly, hard to replicate and reconfigure - hampering the broad adoption of microgravity biology and astrobiology. To address these limitations, we developed a low-cost reconfigurable 3D-printed microscope coined EuniceScope to allow the democratization of astrobiology, especially for educational use. EuniceScope is a compact 2D clinostat system integrated with a modularized brightfield microscope, built upon 3D-printed toolbox. We demonstrated that this compact system offers plausible imaging quality and microgravity-simulating performance. Its high degree of reconfigurability thus holds great promise in the wide dissemination of microgravity-cell-biology research in the broader community, including Science, technology, engineering, and mathematics (STEM) educational and scientific community in the future.

The aim of this study was to analyze the selected hematological and rheological indices in female rowers during the competitive season. The study included 10 female rowers (aged 21.2 ± 2.6) and the control group consisted of 10 woman of corresponding age (non-athletes). The examination of athletes took place two times: at the beginning of the season during high endurance low intensity training period in January (baseline) and at the end of the competitive season in October (after). Blood samples taken from all woman were analyzed for hematological and rheological parameters. The training period of rowers during the 10 months resulted in decrease in red blood cell count and RBC deformability, in contrast to an improvement in some rheological functions such a decrease in fibrinogen concentration, plasma viscosity and aggregation index. The training program practice in rowing modulated some hematological and rheological indices. Some of them positively influenced the cardiovascular system and reduced potential risks connected with hard training and dehydration, but others may have followed from overtraining or not enough relaxation time between training units.

Biological milieus are highly crowded and heterogeneous systems where organization of macromolecules within nanodomains (e.g. membraneless compartments) is vital to the regulation of metabolic processes. There is an increasing interest in understanding the effects that such packed environments have on different biochemical and biological processes. In this context, the redox biochemistry and redox signaling fields are moving towards investigating oxidative processes under conditions that exhibit these key features of biological systems in order to solve existing paradigms including those related to the generation and transmission of specific redox signals within and between cells in both normal physiology and under conditions of oxidative stress. This review outlines the effects that crowding, nanodomain formation and altered local viscosities can have on biochemical processes involving proteins, and then discusses some of the reactions and pathways involving proteins and oxidants that may, or are known to, be modulated by these factors. We postulate that knowledge of protein modification processes (e.g. kinetics, pathways and product formation) under conditions that mimic biological milieus, will provide a better understanding of the response of cells to endogenous and exogenous stressors, and their role in ageing, signaling, health and disease.

The complex spreading dynamics of blood on paper matrix is likely to be quantitatively altered with variations in the fractional occupancy of red blood cells in the whole blood (haematocrit). Here, we presented an apparently surprising observation that a finite volume blood drop undergoes a universal time-dependent spreading on a filter paper strip that is virtually invariant with its hematocrit level within physiologically healthy regime, though distinctively distinguishable from the spreading laws of blood plasma and water.

Our hypothesis was ascertained by performing controlled wicking experiments on filter papers of different grades. Spreading of human blood samples of different haematocrit levels ranging between 15% and 51% and the plasma separated from therein were traced by combined high-speed imaging and microscopy. These experiments were complemented with a semi-analytical theory to decipher the key physics of interest.

Our results unveiled the exclusive influence of the obstructing cellular aggregates in the randomly distributed hierarchically structured porous pathways and deciphered the role of the networked structures of the various plasma proteins that induced hindered diffusion. The resulting universal signatures of spontaneous dynamic spreading, delving centrally on the fractional reduction in the interlaced porous passages, provide novel design basis for paper-microfluidic kits in medical diagnostics and beyond.

Red blood cells (RBCs) are the most numerous cells in the body and perform gas exchange between all tissues. During the infusion of cancer chemotherapeutic (CT) agents, blood cells are the first ones to encounter aggressive cytostatics. Erythrocyte dysfunction caused by direct cytotoxic damage might be a part of the problem of chemotherapy-induced anemia-one of the most frequent side effects. The aim of the current study is to evaluate the functional status of RBCs exposed to mono and combinations of widely used commercial pharmaceutical CT drugs with different action mechanisms: paclitaxel, carboplatin, cyclophosphamide, and doxorubicin, in vitro. Using laser diffraction, flow cytometry, and confocal microscopy, we show that paclitaxel, having a directed effect on cytoskeleton proteins, by itself and in combination with carboplatin, caused the most marked abnormalities-loss of control of volume regulation, resistance to osmotic load, and stomatocytosis. Direct simulations of RBCs' microcirculation in microfluidic channels showed both the appearance of a subpopulation of cells with impaired velocity (slow damaged cells) and an increased number of cases of occlusions. In contrast to paclitaxel, such drugs as carboplatin, cyclophosphamide, and doxorubicin, whose main target in cancer cells is DNA, showed significantly less cytotoxicity to erythrocytes in short-term exposure. However, the combination of drugs had an additive effect. While the obtained results should be confirmed in in vivo models, one can envisioned that such data could be used for minimizing anemia side effects during cancer chemotherapy.

On-chip study of blood flow has emerged as a powerful tool to assess the contribution of each component of blood to its overall function. Blood has indeed many functions, from gas and nutrient transport to immune response and thermal regulation. Red blood cells play a central role therein, in particular through their specific mechanical properties, which directly influence pressure regulation, oxygen perfusion, or platelet and white cell segregation toward endothelial walls. As the bloom of in-vitro studies has led to the apparition of various storage and sample preparation protocols, we address the question of the robustness of the results involving cell mechanical behavior against this diversity. The effects of three conservation media (EDTA, citrate, and glucose-albumin-sodium-phosphate) and storage time on the red blood cell mechanical behavior are assessed under different flow conditions: cell deformability by ektacytometry, shape recovery of cells flowing out of a microfluidic constriction, and cell-flipping dynamics under shear flow. The impact of buffer solutions (phosphate-buffered saline and density-matched suspension using iodixanol/Optiprep) are also studied by investigating individual cell-flipping dynamics, relative viscosity of cell suspensions, and cell structuration under Poiseuille flow. Our results reveal that storing blood samples up to 7 days after withdrawal and suspending them in adequate density-matched buffer solutions has, in most experiments, a moderate effect on the overall mechanical response, with a possible rapid evolution in the first 3 days after sample collection.

The mechanical properties of red blood cells (RBCs) play key roles in their biological functions in microcirculation. In particular, RBCs must deform significantly to travel through microcapillaries with sizes comparable with or even smaller than their own. Although the dynamics of RBCs in microcapillaries have received considerable attention, the effect of membrane viscoelasticity has been largely overlooked. In this work, we present a computational study based on the boundary integral method and thin-shell mechanics to examine how membrane viscoelasticity influences the dynamics of RBCs flowing through straight and constricted microcapillaries. Our results reveal that the cell with a viscoelastic membrane undergoes substantially different motion and deformation compared with results based on a purely elastic membrane model. Comparisons with experimental data also suggest the importance of accounting for membrane viscoelasticity to properly capture the transient dynamics of an RBC flowing through a microcapillary. Taken together, these findings demonstrate the significant effects of membrane viscoelasticity on RBC dynamics in different microcapillary environments. The computational framework also lays the groundwork for more accurate quantitative modeling of the mechanical response of RBCs in their mechanotransduction process in subsequent investigations.

Gas, especially oxygen, transport in the microcirculation is a complex phenomenon, however, of critical importance for maintaining normal biological functions, and the cytoplasm fluid in red blood cells (RBCs) is the major vehicle for transporting oxygen from lungs to tissues via the circulatory system. Existing theoretical and numerical studies have neglected the cytoplasm convection effect by treating RBCs as rigid particles undergoing a constant translation velocity. As a consequence, the influence and mechanism of the cytoplasm flow on oxygen transport are still not clear in microcirculation research. In this study, we consider a tank-treading capsule in shear flow, which is generated with two parallel plates moving in opposite directions: the top plate of a higher oxygen pressure (P_O2) representing the RBC core in the central region of a microvessel and the bottom plate of a lower P_O2 representing the microvessel wall. Numerical simulations are conducted to investigate the individual and combined effects of cytoplasm convection and oxygen-hemoglobin (O₂-Hb) reaction on the oxygen transport efficiency across the tank-treading capsule, and different P_O2 situations and shear rates are also tested. Due to the lower oxygen diffusivity in cytoplasm, the presence of the capsule reduces the oxygen transfer flux across the gap by 7.34 % in the pure diffusion system where the flow convection and O₂-Hb reaction are both neglected. Including the flow convection or the O₂-Hb reaction has little influence on the oxygen flux; however, when they act together as in real microcirculation situations, the enhancement in oxygen transport could be significant, especially in the low P_O2 and high shear rate situations. In particular, with the respective P_O2 at 60 and 30 mmHg on the top and bottom plates and a 400 s^-1 shear rate, the oxygen flux reduction is only 0.02 %, suggesting that the cytoplasm convection can improve the oxygen transport across RBCs considerably. The simulation results are scrutinized to explore the underlying mechanism for the enhancement, and a new nondimensional parameter is introduced to characterize the importance of cytoplasm convection in oxygen transport. These simulation results, discussion and analysis could be helpful for a better understanding of the complex oxygen transport process and therefor valuable for relevant studies.

Double-layered channels of sinusoid lumen and Disse space separated by fenestrated liver sinusoidal endothelial cells (LSECs) endow the unique mechanical environment of the liver sinusoid network, which further guarantees its biological function. It is also known that this mechanical environment changes dramatically under liver fibrosis and cirrhosis, including the reduced plasma penetration and metabolite exchange between the two flow channels and the reduced Disse space deformability. The squeezing of leukocytes through narrow sinusoid lumen also affects the mechanical environment of liver sinusoid. To date, the detailed flow-field profile of liver sinusoid is still far from clear due to experimental limitations. It also remains elusive whether and how the varied physical properties of the pathological liver sinusoid regulate the fluid flow characteristics. Here a numerical model based on the immersed boundary method was established, and the effects of Disse space and leukocyte elasticities, endothelium permeability, and sinusoidal stenosis degree on fluid flow as well as leukocyte trafficking were specified upon a mimic liver sinusoid structure. Results showed that endothelium permeability dominantly controlled the plasma penetration velocity across the endothelium, whereas leukocyte squeezing promoted local penetration and significantly regulated wall shear stress on hepatocytes, which was strongly related to the Disse space and leukocyte deformability. Permeability and elasticity cooperatively regulated the process of leukocytes trafficking through the liver sinusoid, especially for stiffer leukocytes. This study will offer new insights into deeper understanding of the elaborate mechanical features of liver sinusoid and corresponding biological function.

The use of Intraoperative Cell Salvage (ICS) is currently limited in oncological surgeries, due to safety concerns associated with the ability of existing devices to successfully remove circulating tumour cells. In this work, we present the first stages towards the creation of an alternative platform to current cell savers, based on the extremely selective immunoaffinity membrane chromatography principle. Non-woven membranes were produced via electrospinning using poly(vinyl alcohol) (PVA), and further heat treated at 180 °C to prevent their dissolution in aqueous environments and preserve their fibrous morphology. The effects of the PVA degree of hydrolysis (DH) (98 % vs 99 %), method of electrospinning (needleless DC vs AC), and heat treatment duration (1-8 h) were investigated. All heat treated supports maintained their cytocompatibility, whilst tensile tests indicated that the 99 % hydrolysed DC electrospun mats were stronger compared to their 98 % DH counterparts. Although, and at the described conditions, AC electrospinning produced fibres with more than double the diameter compared to those from DC electrospinning, it was not chosen for subsequent experiments because it is still under development. Evidence of unimpeded passage of SY5Y neuroblastoma cells and undiluted defibrinated sheep's blood in flow-through filtration experiments confirmed the successful creation of 3D networks with minimum resistance to mass transfer and lack of non-specific cell binding to the base material, paving the way for the development of novel, highly selective ICS devices for tumour surgeries.

Red blood cells (RBCs) are characterized by a remarkable elasticity, which allows them to undergo very large deformation when passing through small vessels and capillaries. This extreme deformability is altered in various clinical conditions, suggesting that the analysis of red blood cell (RBC) mechanics has potential applications in the search for non-invasive and cost-effective blood biomarkers. Here, we provide a comparative study of the mechanical response of RBCs in patients with Alzheimer's disease (AD) and healthy subjects. For this purpose, RBC viscoelastic response was investigated using atomic force microscopy (AFM) in the force spectroscopy mode. Two types of analyses were performed: (i) a conventional analysis of AFM force-distance (FD) curves, which allowed us to retrieve the apparent Young's modulus, E; and (ii) a more in-depth analysis of time-dependent relaxation curves in the framework of the standard linear solid (SLS) model, which allowed us to estimate cell viscosity and elasticity, independently. Our data demonstrate that, while conventional analysis of AFM FD curves fails in distinguishing the two groups, the mechanical parameters obtained with the SLS model show a very good classification ability. The diagnostic performance of mechanical parameters was assessed using receiving operator characteristic (ROC) curves, showing very large areas under the curves (AUC) for selected biomarkers (AUC > 0.9). Taken all together, the data presented here demonstrate that RBC mechanics are significantly altered in AD, also highlighting the key role played by viscous forces. These RBC abnormalities in AD, which include both a modified elasticity and viscosity, could be considered a potential source of plasmatic biomarkers in the field of liquid biopsy to be used in combination with more established indicators of the pathology.

In this article, extensive three-dimensional simulations are conducted for tank-treading (TT) red blood cells (RBCs) in shear flow with different cell viscous properties and flow conditions. Apart from recent numerical studies on TT RBCs, this research considers the uncertainty in cytoplasm viscosity, covers a more complete range of shear flow situations of available experiments, and examines the TT behaviors in more details. Key TT characteristics, including the rotation frequency, deformation index, and inclination angle, are compared with available experimental results of similar shear flow conditions. Fairly good simulation-experiment agreements for these parameters can be obtained by adjusting the membrane viscosity values; however, different rheological relationships between the membrane viscosity and the flow shear rate are noted for these comparisons: shear thinning from the TT frequency, Newtonian from the inclination angle, and shear thickening from the cell deformation. Previous studies claimed a shear-thinning membrane viscosity model based on the TT frequency results; however, such a conclusion seems premature from our results and more carefully designed and better controlled investigations are required for the RBC membrane rheology. In addition, our simulation results reveal complicate RBC TT features and such information could be helpful for a better understanding of in vivo and in vitro RBC dynamics.

In this paper, a 3D-printed multi-modal device was designed and fabricated to simultaneously detect nitric oxide (NO) and adenosine triphosphate (ATP) in red blood cell suspensions prepared from whole blood. Once a sample was injected into the device, NO was first detected (via amperometry) using a three-electrode, dual-opposed, electrode configuration with a platinum-black/Nafion coated gold working electrode. After in-line amperometric detection of NO, ATP was detected via a chemiluminescence reaction, with a luciferin/luciferase solution continuously pumped into an integrated mixing T and the resulting light being measured with a PMT underneath the channel. The device was optimized for mixing/reaction conditions, limits of detection (40 nM for NO and 30 nM for ATP), and sensitivity. This device was used to determine the basal (normoxic) levels of NO and ATP in red blood cells, as well as an increase in concentration of both analytes under hypoxic conditions. Finally, the effect of storing red blood cells in a commonly used storage solution was also investigated by monitoring the production of NO and ATP over a three-week storage time.

Spectrins are large, evolutionarily well-conserved proteins that form highly organized scaffolds on the inner surface of eukaryotic cells. Their organization in different cell types or cellular compartments helps cells withstand mechanical challenges with unique strategies depending on the cell type. This Review discusses our understanding of the mechanical properties of spectrins, their very distinct organization in red blood cells and neurons as two examples, and the contribution of the scaffolds they form to the mechanical properties of these cells.