Membrane technology applied to donor plasmapheresis

Washed red blood cells (RBCs) are indicated for immunoglobulin (Ig)A-deficient recipients when RBCs from IgA-deficient donors are not available. Canadian Blood Services recently began using the automated ACP 215 cell processor (Haemonetics Corporation) for RBC washing, and its suitability to produce IgA-deficient RBCs was investigated. RBCs produced from whole blood donations by the buffy coat (BC) and whole blood filtration (WBF) methods were washed using the ACP 215 or the COBE 2991 cell processors and IgA and total protein levels were assessed. A double-wash procedure using the ACP 215 was developed, tested, and validated by assessing hemolysis, hematocrit, recovery, and other in vitro quality variables in RBCs stored after washing, with and without irradiation. A single wash using the ACP 215 did not meet Canadian Standards Association recommendations for washing with more than 2 L of solution and could not consistently reduce IgA to levels suitable for IgA-deficient recipients (24/26 BC RBCs and 0/9 WBF RBCs had IgA levels < 0.05 mg/dL). Using a second wash sequence, all BC and WBF units were washed with more than 2 L and had levels of IgA of less than 0.05 mg/dL. During 7 days' postwash storage, with and without irradiation, double-washed RBCs met quality control criteria, except for the failure of one RBC unit for inadequate (69%) postwash recovery. Using the ACP 215, a double-wash procedure for the production of components for IgA-deficient recipients from either BC or WBF RBCs was developed and validated.

The technique of leukocyte filtration has been introduced into cardiac surgery to reduce leukocyte mediated reperfusion injury. When autologous whole blood (WB) is used as filtrate, it is more likely to be activated by the filter material than are banked red blood cell concentrates (RCC), because of its richness in plasma components. This study was designed to compare the activation of plasma components during leukocyte filtration from WB (n = 10) taken from the heart-lung machine, with RCC (n = 10) obtained from the blood bank. Leukocyte filters made from either cellulose acetate or polyester were used. Blood samples were taken simultaneously from the inlet and outlet of the filter after filtration of either 700 ml of WB or 350 ml of RCC. Results indicated that the complement cascade was activated, as reflected by the increase of C3a and C5a during filtration of WB by filters made from cellulose acetate. In contrast, there was no significant increase of C3a and C5a during filtration of RCC. The clotting system, indicated by fibrinopeptide A, and the fibrinolytic system, indicated by fibrinogen degradation products, were not activated during leukocyte filtration. These data suggest that it is the WB taken from the heart-lung machine rather than the RCC from the blood bank that is being activated during leukocyte filtration. Thus, careful selection of filter material is important for leukocyte filtration of autologous whole blood during cardiac surgery.

Patient-Specific Adhesion of Sickle Red Blood Cells in Shear-Gradient Microscale Flow

The aim of the present work is to study the transfer processes in a cross-flow filtration cell, in order to determine the conditions for stable and efficient operation of a side-stream filtration module, integrated with a bioreactor. The current interest in membrane integrated bioreactors is connected with the pursuit of energy and cost efficiency in a wide area of industrial applications, including wastewater treatment, food industry, pharmaceutical industry, and fuel production. A numerical CFD model is employed, based on previous experience with experimental concentration of antioxidants, such as polyphenols and flavonoids from extracts of natural products by nanofiltration. The geometry under investigation is a 3D model of the experimental flat-sheet cell with tangential orientation of the feed inlet. The swirling turbulent flow in the feed channel is favourable for reducing the concentration polarization layer on the membrane surface and preventing fouling. The main factors, affecting the filtration process, are the shear stress distribution and the concentration profiles in the vicinity of the membrane surface. The CFD models of mass transfer in cross-flow nanofiltration are scarce and there are none for the reference experimental filtration cell. The present CFD simulation reveals the concentration distribution in the feed channel. It complements previous data for the flow pattern with new knowledge on the mass transfer there, directed to understanding and control of the concentration polarization phenomenon. The numerical study uses the tools of ANSYS Fluent R13, based on the finite volume method for solving the Reynolds-Averaged Navier-Stokes (RANS) equations. The obtained results are analysed in rapport with available experimental data.

In an effort to improve performances of forward osmosis (FO) systems, several innovative draw spacers have been proposed. However, the small pressure generally applied on the feed side of the process is expected to result in the membrane bending towards the draw side, and in the gradual occlusion of the channel. This phenomenon potentially presents detrimental effects on process performance, including pressure drop and external concentration polarization (ECP) in the draw channel. A flat sheet FO system with a dot-spacer draw channel geometry was characterized to determine the degree of draw channel occlusion resulting from feed pressurization, and the resulting implications on flow performance. First, tensile testing was performed on the FO membrane to derive a Young’s modulus, used to assess the membrane stretching, and the resulting draw channel characteristics under a range of moderate feed pressures. Membrane apex reached up to 67% of the membrane channel height when transmembrane pressure (TMP) of 1.4 bar was applied. The new FO channels considerations were then processed by computational fluid dynamics model (computational fluid dynamics (CFD) by ANSYS Fluent v19.1) and validated against previously obtained experimental data. Further simulations were conducted to better assess velocity profiles, Reynolds number and shear rate. Reynolds number on the membrane surface (draw side) increased by 20% and shear rate increased by 90% when occlusion changed from 0 to 70%, impacting concentration polarisation (CP) on the membrane surface and therefore FO performance. This paper shows that FO draw channel occlusion is expected to have a significant impact on fluid hydrodynamics when the membrane is not appropriately supported in the draw side.

Background: Uncontrolled bleeding is a leading cause of death on the battlefield. We demonstrated previously that supernatant fluid from stored non-leukoreduced (NLR) red blood cells (RBC) inhibited aggregation of platelets in whole blood from healthy donors as measured by impedance aggregometry and decreased GPIIb/IIIa expression. In the current study, we examine stored NLR RBC supernatant fluid inhibition of platelet aggregation in a physiological model of arterial flow. Methods: Fresh citrated blood (500 ul) was mixed with supernate from stored NLR RBC (median age 44 days, 250 ul) and tyrodes buffer (250 ul) for impedance aggregometry. Flow model experiments utilized blood prepared with calcein-labeled platelet rich plasma, RBCs (40% hematocrit), and supernate (25% total volume). Arterial simulated flow (shear rate 1000/sec) over collagen I was carried out in a 75μm x 350μm x 5mm microchamber (Bioflux). GPIIb/IIIa expression was assessed by flow cytometry. Results: Supernates from nine stored NLR RBC units decreased platelet aggregation amplitude in fresh blood from eleven donors by an average of 40±12% vs Tyrode’s buffer controls while one supernate did not exhibit significant inhibition. In concordance with previously reported results, supernates decreased GPIIb/IIIa expression on platelets in whole blood by 53±1.2%. Linear regression analysis of a single flowed experiment showed a 60.5% decrease in aggregation rate (NLR RBC supernate vs HEPES buffered saline control). Inhibition of the rate of aggregation observed in the flow model vs aggregation amplitude by impedance correlated well (sup1: 85.1% vs. 52%; sup 2: 46.6% vs. 41%). Conclusions: Stored NLR RBC units significantly inhibit collagen-stimulated aggregation of platelets in the presence of high (arterial) and low (stirring) shear rates, and reduce expression of the fibrinogen-binding glycoprotein, IIb/IIIa, on the platelet surface. The substantial decrease in platelet aggregation in a physiologically relevant model of whole blood flow suggests massive transfusion of NLR RBC units may decrease the hemostatic capacity of platelets in combat casualties and contribute to uncontrolled hemorrhage. [apc1]unclear

We examined the role of erythrocyte (red blood cell; RBC) aggregation and deformability, neutrophil (polymorphonuclear neutrophil; PMN) deformability, whole-blood viscosity, and platelet-neutrophil interactions on cell filtration in subjects who were critically ill with sepsis (CIS), critically ill noninfected subjects (CINS), and healthy controls (C). We assessed cell deformability by filtration through filters of 5-microm pore size. Whole blood, RBC, PMN, and combinations of PMN and RBC were studied. Viscometry was done on isolated RBC. Platelet-PMN interactions were assessed with monoclonal antibodies to CD41 and activated CD63 platelet receptors, and to CD66b PMN receptors. Filtration pressure (Pi) for CIS was significantly greater than for C and CINS at both high and low PMN and RBC concentrations. Viscometry confirmed decreases in RBC deformability and demonstrated significant increases in RBC aggregation in CIS. Increments in Pi were significantly greater with PMN and PMN-RBC combinations suspended in platelet rich plasma (PRP) than in platelet poor plasma (PPP) for CIS as compared with CINS or C. Flow cytometry confirmed significantly greater platelet activation in CIS than in CINS or C (mean fluorescence: 39 +/- 9 lfu versus 18.7 +/- 4.0 lfu and 17.1 +/- 2.3 lfu, respectively) and greater platelet-PMN aggregation (mean fluorescence: 44.7 +/- 3.6 lfu versus 23 +/- 4.1 lfu, respectively) in CIS than in C. We conclude that decreased filtration of whole blood in CIS is related to decreases in RBC and PMN deformability, increases in RBC aggregation, and increased platelet-PMN interactions. Of these, the formation of platelet-PMN aggregates appeares to have the greatest effect in impairing cell filtration. These rheologic abnormalities may contribute to impaired microvascular blood flow in patients with sepsis.

Prestorage filtration of blood components appears to be an effective method to reduce leukocyte-induced adverse reactions and other complications. To determine whether it is better to filter whole blood before component separation, we compared the efficiency of in-line filtration of whole blood with that of postseparation filtration. Blood was collected from normal, healthy donors into either regular triple-bag containers or into whole-blood integral-filter container systems. We then compared the in vitro storage values of leukocyte-depleted red blood cell concentrates (RBCC) kept at 4 degrees C, and plasma frozen for 1 year with nonfiltered blood components as control. All counts of white blood cells after filtration were < 1 x 10(6) per unit. For almost all storage parameters no significant differences were found between leukocyte-reduced RBCC and control units. The plasma fibrinopeptide A values below 30 ng/ml prior to freezing indicate that filtration does not activate the coagulation factors. Furthermore, the filtration did not influence either the biological values or the coagulation factors of plasma units. Whole blood filtration prior to component preparation seems to offer a useful alternative technique for obtaining leukocyte-reduced RBCC and plasma.

Artificial Surface Effect on Red Blood Cells and Platelets in Laminar Shear Flow

Considering the heterogeneity of cell populations and other critcal problems in mass cultured senescent human fibroblasts, we proposed several new approaches for studying true cellular aging, as follows. 1) To establish a correlation among aging indexes at the individual cell level, we demonstrated a relationship among 3H-thymidine incorporation activity, nuclear size, cell volume, and DNA contents at individual cell level. 2) To fractionate homogenous cell populations and examine the relationship between their life spans and aging indexes, we separated human cells into relatively homogenous populations by the sedimentation velocity method, and found that life span of fractionated cells was almost identical among various fractions, irrespective of their great differences in cell volume and 3H-thymidine incorporation indexes. This suggests that some aging indexes, such as cell volumes and 3H-thymidine incorporation activity, are reversible cell properties, and are not specific properties at the individual cell level. 3) To find age-specific cell properties in fractionated senescent cell populations, we performed a quantitative analysis with an image analyzer on electromicroscopic pictures of fractionated small and large cell populations from young and senescent cultures. We discovered that the differences between young and senescent cultures in nuclear size, mitochonria size and number of lysosomes are due solely to the presence of large cells in senescent cultures, and that an increase in the constricted endoplasmic reticulum is a common phenotype of fractionated senescent cell populations. This suggests that there is some loss of function in endoplasmic reticulum or changes in the membrane system. 4) To examine age-specific changes in membrane system at the individual cell level, we found that the amount of absorption of concanavalin A-bound red blood cells to fibroblast surfaces increases linearly with in vitro passage. Further examination of individual cells indicated that this passage-related change in surface membrane is not dependent on cell cycle phase, surface area, or metabolic age, but certainly is dependent upon division age.

A novel experimental approach based on electrical properties of red blood cell (RBC) suspensions was applied to study the effects of the size and morphology of RBC aggregates on the transient cross‐stream hematocrit distribution in suspensions flowing through a square cross‐section flow channel. The information about the effective size of RBC aggregates and their morphology is extracted from the capacitance (C) and conductance (G) recorded during RBC aggregation, whereas a slower process of particle migration is manifested by delayed long‐term changes in the conductance. Migration‐induced changes in the conductance measured at low shear rates (≤3.1 s−1) for suspensions of RBCs in a strongly aggregating medium reveal an increase to a maximum followed by a decrease to the stationary level. The ascending branch of G(t) curves reflects the aggregate migration in the direction of decreasing shear rate. A further RBC aggregation in the region of lower shear stresses leads to the formation of RBC networks and results in the transformation of the rheological behavior of suspensions from the thinning to the thickening. It is suggested that the descending branches of the G(t) curves recorded at low shear rates reflect an adjustment of the Hct distribution to a new state caused by a partial dispersion of RBC networks. For suspensions of non‐aggregating RBCs it is found that depending on whether the shear rate is higher or lower compared with the prior value, individual RBCs migrate either toward the centerline of the flow or in the opposite direction.

Effect of Renal Replacement Therapy on Viscosity in End-Stage Renal Disease Patients

The cell shape and orientation of red blood cells (RBCs) can be influenced by shear rate and osmolarity. Changes in cell shape and cell orientation can be linked to changes in the optical behavior of the cells. The optical parameters, absorption coefficient &mu;_a, scattering coefficient &mu;_s, and effective scattering phase function of blood in the spectral range from 250 nm to 1100 nm were investigated dependent on shear rate and osmolarity. Integrating sphere measurements of light transmittance and reflectance in combination with inverse Monte-Carlo simulations were carried out for different wall shear rates between 0 and 1000 s^-1 and osmolarity variations from 225 to 400 mosmol/l. Changes in shear rate and osmolarity could be shown to have significant influences on the optical parameters which can in part be explained by changes in the complex refractive index, cell shape and organization. Spherical forms of RBCs induced by low osmolarity show reduced scattering effects compared to normal RBC biconcave disks shape. Spinocytes, induced by high osmolarity, show the highest scattering effects. Randomly oriented cells exhibited maximum &mu;_a and &mu;_s values whereas cell alignment and elongation at high shear rates led to an asymptotical decrease. Moreover a relationship exists between the observed effects and the hemoglobin absorption. It could be shown that 10% changes in osmolarity have a drastic influence on the optical parameters which is of the same order as they appear for 10% Hct and oxygen saturation changes. Flow induced variations of about 10% have less effect on the optical parameters.

Higher baseline whole blood viscosity is associated with chronic SCD complications

The rate of breakage of duplex DNA molecules by laminar flow through a capillary has been studied. For λb2b5c DNA (molecular wt., M = 25 × 106) the point at which breakage occurs is normally distributed around the center of the molecule with a standard deviation of 12.5% of the molecular length. At constant shear stress or shear rate, the breakage rate is independent of ionic strength. Thus, shear induced local denaturation is not a rate limiting, preliminary step in breakage. In experiments at constant temperature with varying solvent viscosity (controlled by added sucrose) the breakage rate is a function of shear rate, not of shear stress. The rate of opening of hydrogenbonded circles into linear molecules by hydrodynamic shear is also shown to be a function of shear rate and not of shear stress. The breakage rate at constant shear rate is not greatly dependent on temperature. The shear rate required to achieve breakage is inversely proportional to M1,2. The breakage rate constant, k varies as a very high power of the shear rate; at 25°C, d In k/d In Gm ∼ 15; at 10°C, d In k/d In Gm ∼ 26, where Gm is the maximum shear rate at the capillary wall. The unexpected result that breakage rate is mainly dependent on shear rate, not shear stress, supports a model in which the DNA molecule is distorted with a driving force which depends on the hydrodynamic shear stress, ηG, but the rate limiting step is segment diffusion into a highly extended configuration. The characteristic time to achieve this configuration is proportional to solvent viscosity, η, hence the breakage rate is dependent on ηG/η or G, the shear rate.