Axial and Nonaxial Migration of Red Blood Cells in a Microtube.

Human red blood cells (RBCs) are subjected to high viscous shear stress, especially during microcirculation, resulting in stable deformed shapes such as parachute or slipper shape. Those unique deformed RBC shapes, accompanied with axial or nonaxial migration, cannot be fully described according to traditional knowledge about lateral movement of deformable spherical particles. Although several experimental and numerical studies have investigated RBC behavior in microchannels with similar diameters as RBCs, the detailed mechanical characteristics of RBC lateral movement-in particular, regarding the relationship between stable deformed shapes, equilibrium radial RBC position, and membrane load-has not yet been fully described. Thus, we numerically investigated the behavior of single RBCs with radii of 4 µm in a circular microchannel with diameters of 15 µm. Flow was assumed to be almost inertialess. The problem was characterized by the capillary number, which is the ratio between fluid viscous force and membrane elastic force. The power (or energy dissipation) associated with membrane deformations was introduced to quantify the state of membrane loads. Simulations were performed with different capillary numbers, viscosity ratios of the internal to external fluids of RBCs, and initial RBC centroid positions. Our numerical results demonstrated that axial or nonaxial migration of RBC depended on the stable deformed RBC shapes, and the equilibrium radial position of the RBC centroid correlated well with energy expenditure associated with membrane deformations.

Margination of Platelet-Sized Particles in the Red Blood Cell Suspension Flow through Square Microchannels.

In the blood flow through microvessels, platelets show high concentrations near the vessel wall. This phenomenon is called margination of platelets and is closely associated with hemostasis and thrombosis. In the present study, we conducted in vitro experiments using platelet-sized fluorescent particles as platelet substitutes to investigate the cross-sectional distribution of these particles in the red blood cell suspension flowing through microchannels with a square cross section. Fluorescence observations were performed to measure the transverse distribution of particles at various heights from the bottom face with the use of a confocal laser scanning microscope system. In downstream cross sections of the channel, particles showed focusing near the four corners rather than uniform margination along the entire circumference of the cross section. The focusing of particles near the corners was more enhanced for higher hematocrits. On the other hand, particles in circular channel flows showed nearly axisymmetric uniform accumulation adjacent to the channel wall. The present result suggests that the segregation of suspended particles in the flow of multicomponent suspensions could have such heterogeneous 2D features of particle distribution in the cross section of channels, especially for rectangular channels often used in microfluidics.

Pattern Transition on Inertial Focusing of Neutrally Buoyant Particles Suspended in Rectangular Duct Flows.

The continuous separation and filtration of particles immersed in fluid flows are important interests in various applications. Although the inertial focusing of particles suspended in a duct flow is promising in microfluidics, predicting the focusing positions depending on the parameters, such as the shape of the duct cross-section and the Reynolds number (Re) has not been achieved owing to the diversity of the inertial-focusing phenomena. In this study, we aimed to elucidate the variation of the inertial focusing depending on Re in rectangular duct flows. We performed a numerical simulation of the lift force exerted on a spherical particle flowing in a rectangular duct and determined the lift-force map within the duct cross-section over a wide range of Re. We estimated the particle trajectories based on the lift map and Stokes drag, and identified the particle-focusing points appeared in the cross-section. For an aspect ratio of the duct cross-section of 2, we found that the blockage ratio changes transition structure of particle focusing. For blockage ratios smaller than 0.3, particles focus near the centres of the long sides of the cross-section at low Re and near the centres of both the long and short sides at relatively higher Re. This transition is expressed as a subcritical pitchfork bifurcation. For blockage ratio larger than 0.3, another focusing pattern appears between these two focusing regimes, where particles are focused on the centres of the long sides and at intermediate positions near the corners. Thus, there are three regimes; the transition between adjacent regimes at lower Re is found to be expressed as a saddle-node bifurcation and the other transition as a supercritical pitchfork bifurcation.

Effect of the Propionylation Method on the Deformability under Thermal Pressure of Block-Shaped Wood.

Converting wood waste into thermoplastic materials is an attractive means of increasing its utilization because complex three-dimensional molded products can easily be obtained by press molding wood with thermoplasticity. Chemical modification, especially esterification, is a promising method for imparting thermoplasticity to wood. In this study, we produced multiple propionylated wood specimens using several propionylation methods and elucidated the factors affecting the deformability of the wood. Regardless of the method, all of the propionylated wood samples showed deformability in the tangential direction. However, in the longitudinal direction, not only the degree of propionylation but also the propionylation method had a significant influence on the deformability. The flow in the tangential direction occurred under a relatively low pressure, whereas the flow in the longitudinal direction occurred under higher pressure. The chemical composition and motility of each sample were evaluated using solid-state NMR measurements. With some propionylation methods, decomposition of the cellulose main chain occurred during the reaction, which had a dominant effect on the deformability of the wood in the longitudinal direction. These results indicate that the deformability of wood can be controlled by the appropriate selection of a propionylation method and its treatment conditions.

Rapid Benzylation of Wood Powder without Heating.

Converting wood waste into thermoplastic materials is an attractive means of increasing its utilization. A promising method for imparting thermoplasticity to wood is chemical modification, in which the hydroxyl groups in wood are substituted with benzyl groups. In the common method, wood powder is first treated with a highly concentrated aqueous NaOH solution, and then reacted with a benzylation reagent by heating for a long time under stirring. In this study, a 50% aqueous tetra-n-butylphosphonium hydroxide solution was used for the pretreatment of wood powder. This modified alkaline treatment enhanced the efficiency of the subsequent benzylation reaction, which could be conducted without heating over a shorter time. The effects of various conditions on the efficiency of the benzylation reaction were evaluated. Both the alkali pretreatment and the subsequent benzylation required only ~5-10 min of stirring without heating to obtain benzylated wood with a similar degree of benzylation as that achieved by the common method. The chemical structure of the benzylated wood powder was characterized by Fourier-transform infrared and solid-state NMR spectroscopies, and its thermal softening characteristics were evaluated by thermomechanical analysis. Finally, a translucent film could be obtained by hot-pressing the benzylated wood powder.

Development of margination of platelet-sized particles in red blood cell suspensions flowing through Y-shaped bifurcating microchannels.

BACKGROUND: In the blood flow through microvessels, platelets exhibit enhanced concentrations in the layer free of red blood cells (cell-free layer) adjacent to the vessel wall. The motion of platelets in the cell-free layer plays an essential role in their interaction with the vessel wall, and hence it affects their functions of hemostasis and thrombosis. OBJECTIVE: We aimed to estimate the diffusivity of platelet-sized particles in the transverse direction (the direction of vorticity) across the channel width in the cell-free layer by in vitro experiments for the microchannel flow of red blood cell (RBC) suspensions containing platelet-sized particles. METHODS: Fluorescence microscope observations were performed to measure the transverse distribution of spherical particles immersed in RBC suspensions flowing through a Y-shaped bifurcating microchannel. We examined the development of the particle concentration profiles along the flow direction in the daughter channels, starting from asymmetric distributions with low concentrations on the inner side of the bifurcation at the inlet of the daughter channels. RESULTS: In daughter channels of 40 µm width, reconstruction of particle margination revealed that a symmetric concentration profile was attained in ∼30 mm from the bifurcation, independent of flow rate. CONCLUSIONS: We presented experimental evidence of particle margination developing in a bifurcating flow channel where the diffusivity of 2.9-µm diameter particles was estimated to be ∼40 µm2/s at a shear rate of 1000 s-1 and hematocrit of 0.2.

Visualization of solute diffusion into cell walls in solution-impregnated wood under varying relative humidity using time-of-flight secondary ion mass spectrometry.

The purpose of the present study is to clarify the diffusion of non-volatile substances into cell walls during the conditioning procedure under varying relative humidities (RH). In this paper, wood blocks were impregnated using an aqueous solution of melamine formaldehyde (MF), and they were subsequently conditioned under RHs of 11, 43, and 75%. The solute that diffused into the cell walls was visualized using time-of-flight secondary ion mass spectrometry (TOF-SIMS). The volumetric relative swelling of the samples during the conditioning procedure was calculated. The results showed increased cell wall swelling at higher RH, which may have been caused by higher MF diffusion into the cell walls and/or higher moisture content. Cryo-TOF-SIMS measurements showed that more cell cavities were unfilled with MF at higher RH, indicating that most of the MF diffused from the cell cavities into the cell walls. The relative intensity of MF in the cell walls of the cured samples was evaluated from dry-TOF-SIMS images, which showed a higher relative intensity of MF in the cell walls at higher RH. With the ability to visualize and semi-quantitatively evaluate the solute in cell walls, TOF-SIMS will serve as a powerful tool for future studies of solute diffusion mechanisms in solution-impregnated wood.

Cross-sectional distributions of normal and abnormal red blood cells in capillary tubes determined by a new technique.

BACKGROUND: In the microcirculation, red blood cells (RBCs) were observed to be confined to an axial stream surrounded by a marginal RBC depleted layer. This axial accumulation of RBCs is considered to arise from the RBC deformability. OBJECTIVE: To quantitatively evaluate the effect of RBC deformability on their axial accumulation at a flow condition comparable to that in arterioles by developing a new observation system for accurate measurements of radial RBC positions in the cross section of capillary tubes. METHODS: The cross-sectional distributions of normal and hardened RBCs as well as softened RBCs suspended in capillary tube flows were measured with high spatial resolution. A new observation system was developed in which enface views of the cross-section of the tube were obtained at small distances upstream of the outlet at various longitudinal positions in the tube. RESULTS: The radial positions of individual RBCs were detected within 1 µm accuracy. It was found that normal and softened RBCs rapidly migrated away from the wall towards the tube axis, whereas glutaraldehyde-hardened RBCs were dispersed widely over the tube cross-section, depending on the concentration of glutaraldehyde solution. CONCLUSIONS: The newly devised observation system revealed quantitatively the essential role of RBC deformability in their axial accumulation.

Molecular and genealogical analysis of grain dormancy in Japanese wheat varieties, with specific focus on MOTHER OF FT AND TFL1 on chromosome 3A.

In the wheat (Triticum aestivum L.) cultivar 'Zenkoujikomugi', a single nucleotide polymorphism (SNP) in the promoter of MOTHER OF FT AND TFL1 on chromosome 3A (MFT-3A) causes an increase in the level of gene expression, resulting in strong grain dormancy. We used a DNA marker to detect the 'Zenkoujikomugi'-type (Zen-type) SNP and examined the genotype of MFT-3A in Japanese wheat varieties, and we found that 169 of 324 varieties carry the Zen-type SNP. In Japanese commercial varieties, the frequency of the Zen-type SNP was remarkably high in the southern part of Japan, but low in the northern part. To examine the relationship between MFT-3A genotype and grain dormancy, we performed a germination assay in three wheat-growing seasons. On average, the varieties carrying the Zen-type SNP showed stronger grain dormancy than the varieties carrying the non-Zen-type SNP. Among commercial cultivars, 'Iwainodaichi' (Kyushu), 'Junreikomugi' (Kinki-Chugoku-Shikoku), 'Kinuhime' (Kanto-Tokai), 'Nebarigoshi' (Tohoku-Hokuriku), and 'Kitamoe' (Hokkaido) showed the strongest grain dormancy in each geographical group, and all these varieties, except for 'Kitamoe', were found to carry the Zen-type SNP. In recent years, the number of varieties carrying the Zen-type SNP has increased in the Tohoku-Hokuriku region, but not in the Hokkaido region.

Venular valves and retrograde perfusion.

Forced retrograde perfusion through the venous system with arterial blood can provide adequate oxygen to peripheral tissues at rest through veno-capillary networks which is the basis for surgical restoration of blood flow by distal vein arterialization (DVA). To be successful such surgery requires disruption of valve leaflets in the veins, which can be accomplished easily in the larger vessels. However, the smallest veins (venules) of less than 100 µm in diameter, also possess valves, are distributed widely throughout all tissues and are too fine for any effective surgical interference. Thus venular valves cannot be disrupted or dissected with presently available technology. Nevertheless, clinical observations suggest that retrograde peripheral blood flow is rapidly established after DVA surgery. There is as yet no rational explanation for this phenomenon. In the present study, using Laplace's law, we attempt to elucidate the mechanical properties of venules and their valves. We speculate that the remarkably thin venular walls (and especially those of the smaller vessels which have the thinnest walls), are capable of considerable, rapid distension when subjected to increased hemostatic pressure. The increase in diameter of venules in response to the increased blood pressure renders their valve leaflets incompetent, so that the valves themselves cannot close the vessel lumen. In addition, the thin bicuspid leaflets may also be forced open retrogradely by the increased blood pressure.

Distribution of photoperiod-insensitive allele Ppd-A1a and its effect on heading time in Japanese wheat cultivars.

The Ppd-A1 genotype of 240 Japanese wheat cultivars and 40 foreign cultivars was determined using a PCR-based method. Among Japanese cultivars, only 12 cultivars, all of which were Hokkaido winter wheat, carried the Ppd-A1a allele, while this allele was not found in Hokkaido spring wheat cultivars or Tohoku-Kyushu cultivars. Cultivars with a photoperiod-insensitive allele headed 6.9-9.8 days earlier in Kanto and 2.5 days earlier in Hokkaido than photoperiod-sensitive cultivars. The lower effect of photoperiod-insensitive alleles observed in Hokkaido could be due to the longer day-length at the spike formation stage compared with that in Kanto. Pedigree analysis showed that 'Purple Straw' and 'Tohoku 118' were donors of Ppd-A1a and Ppd-D1a in Hokkaido wheat cultivars, respectively. Wheat cultivars recently developed in Hokkaido carry photoperiod-insensitive alleles at a high frequency. For efficient utilization of Ppd-1 alleles in the Hokkaido wheat-breeding program, the effect of Ppd-1 on growth pattern and grain yield should be investigated. Ppd-A1a may be useful as a unique gene source for fine tuning the heading time in the Tohoku-Kyushu region since the effect of Ppd-A1a on photoperiod insensitivity appears to differ from the effect of Ppd-B1a and Ppd-D1a.

Transition of planar Couette flow at infinite Reynolds numbers.

An outline of the state space of planar Couette flow at high Reynolds numbers (Re<10^{5}) is investigated via a variety of efficient numerical techniques. It is verified from nonlinear analysis that the lower branch of the hairpin vortex state (HVS) asymptotically approaches the primary (laminar) state with increasing Re. It is also predicted that the lower branch of the HVS at high Re belongs to the stability boundary that initiates a transition to turbulence, and that one of the unstable manifolds of the lower branch of HVS lies on the boundary. These facts suggest HVS may provide a criterion to estimate a minimum perturbation arising transition to turbulent states at the infinite Re limit.

Segregation by size difference in binary suspensions of fluid droplets in channel flow.

In channel flow of multicomponent suspensions, segregation behavior of suspended components perpendicular to the flow direction is often observed, which is considered to be caused by the differential properties of the lateral migration depending on their shape, size, flexibility, and other characteristics. In the present study, we investigate the effect of size differences between suspended components on the segregation behavior, by a two-dimensional numerical simulation for binary dispersed suspensions of fluid droplets of two different sizes subjected to a plane Poiseuille channel flow. The small and large droplets are assumed to have equal surface tensions and equal viscosity ratios of internal to external fluids. The time evolutions of the lateral positions of large and small droplets relative to the channel centerline were computed by changing the area fraction of the small droplets in a mixture with a constant total area fraction. The large droplets are found to migrate closer to the channel centerline and the small droplets are found to migrate closer to the channel wall compared to the corresponding lateral positions in mono-dispersed suspensions at the same area fractions, although the mean lateral positions of the large and small droplets in mono-dispersed suspension are comparable. This segregation behavior as well as the margination of small droplets are enhanced when the size difference between large and small droplets is increased and the area fraction of large droplets is increased. These results may arise from higher tendencies for the large droplets to approach the channel centerline compared to the small droplets, which consequently expel small droplets from the central region toward the channel walls.

Isolation of a wheat (Triticum aestivum L.) mutant in ABA 8'-hydroxylase gene: effect of reduced ABA catabolism on germination inhibition under field condition.

Pre-harvest sprouting, the germination of mature seeds on the mother plant under moist condition, is a serious problem in cereals. To investigate the effect of reduced abscisic acid (ABA) catabolism on germination in hexaploid wheat (Triticum aestivum L.), we cloned the wheat ABA 8'-hydroxyase gene which was highly expressed during seed development (TaABA8'OH1) and screened for mutations that lead to reduced ABA catabolism. In a screen for natural variation, one insertion mutation in exon 5 of TaABA8'OH1 on the D genome (TaABA8'OH1-D) was identified in Japanese cultivars including 'Tamaizumi'. However, a single mutation in TaABA8'OH1-D had no clear effect on germination inhibition in double haploid lines. In a screen for a mutation, one deletion mutant lacking the entire TaABA8'OH1 on the A genome (TaABA8'OH1-A), TM1833, was identified from gamma-ray irradiation lines of 'Tamaizumi'. TM1833 (a double mutant in TaABA8'OH1-A and TaABA8'OH1-D) showed lower TaABA8'OH1 expression, higher ABA content in embryos during seed development under field condition and lower germination than those in 'Tamaizumi' (a single mutant in TaABA8'OH1-D). These results indicate that reduced ABA catabolism through mutations in TaABA8'OH1 may be effective in germination inhibition in field-grown wheat.

Charge effects on the hindered transport of macromolecules across the endothelial surface glycocalyx layer.

A fluid mechanical and electrostatic model for the transport of solute molecules across the vascular endothelial surface glycocalyx layer (EGL) was developed to study the charge effect on the diffusive and convective transport of the solutes. The solute was assumed to be a spherical particle with a constant surface charge density, and the EGL was represented as an array of periodically arranged circular cylinders of like charge, with a constant surface charge density. By combining the fluid mechanical analyses for the flow around a solute suspended in an electrolyte solution and the electrostatic analyses for the free energy of the interaction between the solute and cylinders based on a mean field theory, we estimated the transport coefficients of the solute across the EGL. Both of diffusive and convective transports are reduced compared to those for an uncharged system, due to the stronger exclusion of the solute that results from the repulsive electrostatic interaction. The model prediction for the reflection coefficient for serum albumin agreed well with experimental observations if the charge density in the EGL is ranged from approximately -10 to -30 mEq/l.

Distribution of photoperiod-insensitive alleles Ppd-B1a and Ppd-D1a and their effect on heading time in Japanese wheat cultivars.

The genotypes of photoperiod response genes Ppd-B1 and Ppd-D1 in Japanese wheat cultivars were determined by a PCR-based method, and heading times were compared among genotypes. Most of the Japanese wheat cultivars, except those from the Hokkaido region, carried the photoperiod-insensitive allele Ppd-D1a, and heading was accelerated 10.3 days compared with the Ppd-D1b genotype. Early cultivars with Ppd-D1a may have been selected to avoid damage from preharvest rain. In the Hokkaido region, Ppd-D1a frequency was lower and heading date was late regardless of Ppd-D1 genotype, suggesting another genetic mechanism for late heading in Hokkaido cultivars. In this study, only 11 cultivars proved to carry Ppd-B1a, and all of them carried another photoperiod-insensitive allele, Ppd-D1a. The Ppd-B1a/Ppd-D1a genotype headed 6.7 days earlier than the Ppd-B1b/Ppd-D1a genotype, indicating a significant effect of Ppd-B1a in the genetic background with Ppd-D1a. Early-maturity breeding in Japan is believed to be accelerated by the introduction of the Ppd-B1a allele into medium-heading cultivars carrying Ppd-D1a. Pedigree analysis showed that Ppd-B1a in three extra-early commercial cultivars was inherited from 'Shiroboro 21' by early-heading Chugoku lines bred at the Chugoku Agriculture Experimental Station.

Galectin-9 ameliorates immune complex-induced arthritis by regulating Fc gamma R expression on macrophages.

Galectin-9 up-regulated Fc gamma RIIb expression of mouse peritoneal macrophages in vitro but down-regulated Fc gamma RIII expression. Galectin-9-treated macrophages stimulated with immune complexes (IC) produced less TNFalpha and IL-1 beta but more IL-10 than PBS-treated macrophages. Macrophage enhancing effects on IC-induced C5a and neutrophil chemotactic activity were also diminished for galectin-9-treated macrophages. In galectin-9-treated mice, the severity of IC-induced arthritis was reduced, as were pro-inflammatory cytokine levels in inflamed joints and serum C5a. Fc gamma RIIb expression of macrophages from galectin-9-treated mice was up-regulated, whereas Fc gamma RIII expression was down-regulated. Macrophages from galectin-9-treated mice produced less TNFalpha and IL-1 beta but more IL-10 than PBS-treated mice. Disease severity of galectin-9-transgenic mice was milder than wild-type mice, whereas that of galectin-9-deficient mice was exaggerated. Furthermore, macrophage Fc gamma RIIb expression in galectin-9-deficient mice was down-regulated, while Fc gamma RIII expression was up-regulated. These results suggest that galectin-9 suppresses IC-induced inflammation partly by regulating Fc gamma R expression on macrophages.

Galectin-9 accelerates transforming growth factor beta3-induced differentiation of human mesenchymal stem cells to chondrocytes.

Galectin-9 (Gal-9), a beta-galactoside binding lectin, plays a crucial role in innate and adaptive immunity. In the rat collagen-induced arthritis model, administration of Gal-9 induced repair of existing cartilage injury even when joints were already swollen with cartilage destruction. We thus attempted to explore the role of Gal-9 in chondrocyte differentiation utilizing human mesenchymal stem cell (MSC) pellet cultures. During chondrogenesis induced by transforming growth factor beta3 (TGFbeta3), MSCs strongly expressed endogenous Gal-9. Expression of Gal-9 peaked on day 14 and the neutralization of endogenous Gal-9 resulted in the reduced chondrogenesis, indicating possible involvement of Gal-9 in TGFbeta-mediated chondrogenesis. In pellets, addition of Gal-9 significantly enhanced TGFbeta3-induced chondrogenesis, as evidenced by increasing proteoglycan content, but not cell proliferation. In the absence of Gal-9, collagen expression by MSCs switched from type I to type II on 28 days after stimulation with TGFbeta3. When MSCs were co-stimulated with Gal-9, the class switch occurred on day 21. In addition, Gal-9 synergistically enhanced TGFbeta3-induced phosphorylation of Smad2, though Gal-9 did not itself induce detectable Smad2 phosphorylation. These results suggest that Gal-9 has a beneficial effect on cartilage repair in injured joints by induction of differentiation of MSCs into chondrocytes.

Galectin-9 suppresses the generation of Th17, promotes the induction of regulatory T cells, and regulates experimental autoimmune arthritis.

The effects of galectin-9 on a mouse collagen-induced arthritis (CIA) model were assessed to clarify whether galectin-9 suppresses CIA by regulating T cell immune responses. Galectin-9 suppressed CIA in a dose-dependent manner, and such suppression was observed even when treatment was started on 7 days after the booster, indicating its preventive and therapeutic effects. Galectin-9 induced the decreased levels of pro-inflammatory cytokines, IL-17, IL-12, and IFNgamma in the joint. Galectin-9 induced the decreased number of CD4(+) TIM-3(+) T cells in peripheral blood. Galectin-9-deficient mice became susceptible to CIA may be by increased number of CD4(+) TIM-3(+) T cells and decreased number of Treg cells. We further found that galectin-9 induces differentiation of naive T cells to Treg cells, and it suppresses differentiation to Th17 cells in vitro. The present results suggested that galectin-9 ameliorates CIA by suppressing the generation of Th17, promoting the induction of regulatory T cells.

Mechanotransduction in leukocyte activation: a review.

We review recent evidence which suggests that leukocytes in the circulation and in the tissue may readily respond to physiological levels of fluid shear stress in the range between about 1 and 10 dyn/cm 2, a range that is below the level to achieve a significant passive, viscoelastic response. The response of activated neutrophilic leukocytes to fluid shear consists of a rapid retraction of lamellipodia with membrane detachment from integrin binding sites. In contrast, a subgroup of non-activated neutrophils may project pseudopods after exposure to fluid shear stress. The evidence suggests that G-protein coupled receptor downregulation by fluid shear with concomitant downregulation of Rac-related small GTPases and depolymerization of F-actin serves to retract the lamellipodia in conjunction with proteolytic cleavage of beta 2 integrin to facilitate membrane detachment. Furthermore, there exists a mechanism to up- and down-regulate the fluid shear-response, which involves nitric oxide and the second messenger cyclic guanosine monophosphate (cGMP). Many physiological activities of circulating leukocytes are under the influence of fluid shear stress, including transendothelial migration of lymphocytes. We describe a disease model with chronic hypertension that suffers from an attenuated fluid shear-response with far reaching implications for microvascular blood flow.