Micropore Filtration Research Articles

On-chip circulating tumor cells isolation based on membrane filtration and immuno-magnetic bead clump capture

Isolating rare circulating tumor cells (CTCs) from blood is critical for the downstream analysis that is important in cancer-related research, diagnosis, and medicine, and efforts are ongoing to increase the efficiency and purity of CTC isolation in microfluidics. Reported in this paper is a two-stage integrated microfluidic chip for coarse-to-fine CTC isolation from whole blood. First, blood cells are removed by filtration using a micropore-array membrane, then CTCs and other cells that are trapped in the micropores are peeled off the membrane by a novel release method based on air–liquid interfacial tension, which significantly increases the recovery rate of CTCs. The second stage is CTC capture based on an on-chip dense immuno-magnetic-bead clump, which offers high capture efficiency and purity. Both the micropore filtration and immuno-magnetic-bead capture are validated and optimized experimentally. Overall, the integrated microfluidic chip can realize a recovery rate of 85.5% and a purity of 37.8% for rare cancer cells spiked in whole blood.

Red Blood Cell Morphology in Diabetic Patients: A Case-control Study

Introduction: According to earlier studies, patients suffering from diabetes frequently experience haematological changes. India is the diabetes capital, with 69.1 million residents suffering from the disease. Aim: To study Red Blood Cell (RBC) morphological changes and grading on percentage of cells that differ in size and shape from normal erythrocytes in diabetic patients. Materials and Methods: The case-control study was conducted at Division of Central Clinical Laboratory in Department of Pathology in coordination with Department of General Medicine at Jawaharlal Nehru Medical College, Wardha, Maharashtra, India, from October 2019 to October 2021. Total 90 known cases of diabetes mellitus, aged between 18-80 years and 90 subjects (control group) with normal blood glucose levels were included for the study. Demographic details, anthropometric, parameters i.e, Fasting Plasma Glucose (FPG), Random Blood Sugar (RBS), Glycated haemoglobin (HbA1c) and morphology of blood smear were assessed. Results: The mean age of study group was 54.98±2.31 years and control group was 50.45±17.94 years (p-value= 0.0004). The mean random blood sugar of study group was 163.92±91.31 mg/dL and of control group was 96.92±22.31 mg/dL (p-value=0.0001). Mean HbA1c for the study group was 7.73±1.71%, while in control group was 5.24±0.62% (p-value=0.0001). In the study group, 9 (10%) cases showed a slight anisocytosis (1+) and 3 (3.3%) cases showed moderate anisocytosis (2+), 2 (2.2%) cases showed marked anisocytosis (3+) (variation in size of RBCs with respect to normal) in respect to Mean Corpuscle Volume (MCV), and 11 (12.2%) cases showed MCV <80 fL, 2 (2.2%) cases showed MCV >80 fL, while majority of the cases; 77 (85.5%) had normal sized RBCs, had MCV between 80-99 fL. Conclusion: The findings of the study suggest that there is need for some simple and effective techniques like ectocytometry, micropore filtration for routine haematological tests in type 2 diabetes mellitus.

Estimation of membrane bending modulus of stiffness tuned human red blood cells from micropore filtration studies.

Human red blood cells (RBCs) need to deform in order to pass through capillaries in human vasculature with diameter smaller than that of the RBC. An altered RBC cell membrane stiffness (CMS), thereby, is likely to have consequences on their flow rate. RBC CMS is known to be affected by several commonly encountered disease conditions. This study was carried out to investigate whether an increase in RBC CMS, to the extent seen in such commonly encountered medical conditions, affects the RBC flow rate through channels with diameters comparable to that of the RBC. To do this, we use RBCs extracted from a healthy individual with no known medical conditions and treated with various concentrations of Bovine Serum Albumin (BSA). We study their flow through polycarbonate membranes with pores of diameter 5μm and 8μm which are smaller than and comparable to the RBC diameter respectively. The studies are carried out at constant hematocrit and volumetric flow rate. We find that when the diameter of the capillary is smaller than that of the RBC, the flow rate of the RBCs is lowered as the concentration of BSA is increased while the reverse is true when the diameter is comparable to that of the RBC. We confirm that this is a consequence of altered CMS of the RBCs from their reorientation dynamics in an Optical Tweezer. We find that a treatment with 0.50mg/ml BSA mimics the situation for RBCs extracted from a healthy individual while concentrations higher than 0.50mg/ml elevate the RBC CMS across a range expected for individuals with a condition of hyperglycemia. Using a simple theoretical model of the RBC deformation process at the entry of a narrow channel, we extract the RBC membrane bending modulus from their flow rate.

空气细颗粒物PM2.5水溶成分对人黑素细胞系PIG1增殖、迁移和黑素合成的影响

Objective To evaluate the effect of water-soluble components of atmospheric fine particulate matter PM2.5 on proliferation, migration, tyrosinase activity and melanin content of a human melanocyte line PIG1. Methods PM2.5 was collected during haze weather in heating seasons, and processed into suspensions. PIG1 melanocytes were cultured and divided into 5 experimental groups and 1 control group. PIG1 melanocytes in the 5 experimental groups were treated with 10, 20, 50, 100 and 200 mg/L PM2.5 suspensions respectively for 48 hours, while cells in the control group were not treated with PM2.5 suspensions. In cell migration assay, there was only 1 experimental group treated with 10 mg/L PM2.5 suspensions. After treatment, methyl thiazol tetrazolium (MTT) assay, micropore filtration assay, DOPA oxidase assay and NaOH lysis method were performed to determine the cell proliferation rate, migration rate, tyrosinase activity and melanin content respectively. Statistical analysis was carried out by using t test for comparison of means of two samples, one-way analysis of variance for means of multiple samples, Student-Newman-Keuls (SNK) -q test for multiple comparisons, and linear correlation analysis for analysis of correlations. Results Compared with the control group ([100 ± 1.41]%) , the proliferation rate of PIG1 cells significantly decreased in the 20-, 50-, 100- and 200-mg/L PM2.5 groups ([93.41 ± 2.13]%, [88.31 ± 1.3557]%, [79.75 ± 1.89]%, [69.83 ± 2.50]% respectively, all P < 0.05) . Linear correlation analysis showed that the proliferation rate and tyrosinase activity of PIG1 cells decreased with the increase in PM2.5 concentrations (r = -0.98, -0.93, respectively, both P < 0.01) . After the treatment with 10 mg/L PM2.5, the migration rate of PIG1 cells significantly decreased (66.23% ± 1.11%) compared with the control group ([76.86 ± 1.81]%, t = 7.55, P < 0.01) . With the increase in PM2.5 concentrations (50-200 mg/L) , the melanin content of PIG1 cells gradually decreased (r = -0.97, P < 0.01) . Conclusion Atmospheric fine particulate matter PM2.5 can affect the normal functions of melanocytes by inhibiting their proliferation and migration, and reducing their tyrosinase activity and melanin content. Key words: Melanocytes; Particulate matter; Cell proliferation; Melanins; Cell migration assays; PM2.5

Simultaneous determination of fifteen ultraviolet filters in sunscreen cosmetics by high performance liquid chromatography

An analytical method for the simultaneous determination of fifteen ultraviolet (UV) filters in sunscreen cosmetics was developed by high performance liquid chromatography (HPLC). Various cosmetic samples including lotions, emulsions, creams and waxes were firstly mixed thoroughly by adding tetrahydrofuran containing 2 g/L ammonium hydroxide followed by vortex and oscillation. If the wax samples were still homogenized incompletely, a ultrasonic oscillation protocol with heating to 50 °C was adopted. The homogeneous samples were then ultrasonically extracted by 80% aqueous methanol solution. After centrifugation and micropore filtration, the extracted components were separated on an XTerra MS C18 column with gradient elution by water containing 0. 1% (v/v) formic acid and methanol containing 0. 1% (v/v) formic acid, determined by a diode array detector (DAD) at 280 nm and 311 nm, and quantified using external standard method. Optimization for the pretreatment conditions of different matrices of cosmetics including sample dispersion solvents, extraction solvents and time were carried out. This developed method showed good linearity (r2 ≥ 0.999 1) and the limits of quantification (LOQs) were between 1.2 and 5.1 µg/g. The recoveries of the fifteen compounds ranged from 84.2% to 100.7% at three spiked levels with the relative standard deviations (RSDs) between 0.9% and 9.5%. The analytes were well separated and this analytical method proved to be sensitive and accurate, so it can be used in the practical determination of the fifteen UV filters in commercial sunscreen cosmetics.

Sequential Blood Filtration for Extracorporeal Circulation: Initial Results from a Proof-of-Concept Prototype

Micropore filters are used during extracorporeal circulation to prevent gaseous and solid particles from entering the patient’s systemic circulation. Although these devices improve patient safety, limitations in current designs have prompted the development of a new concept in micropore filtration. A prototype of the new design was made using 40-μm filter screens and compared against four commercially available filters for performance in pressure loss and gross air handling. Pre- and postfilter bubble counts for 5- and 10-mL bolus injections in an ex vivo test circuit were recorded using a Doppler ultrasound bubble counter. Statistical analysis of results for bubble volume reduction between test filters was performed with one-way repeated-measures analysis of variance using Bonferroni post hoc tests. Changes in filter performance with changes in microbubble load were also assessed with dependent t tests using the 5- and 10-mL bolus injections as the paired sample for each filter. Significance was set at p &lt; .05. All filters in the test group were comparable in pressure loss performance, showing a range of 26–33 mmHg at a flow rate of 6 L/min. In gross air-handling studies, the prototype showed improved bubble volume reduction, reaching statistical significance with three of the four commercial filters. All test filters showed decreased performance in bubble volume reduction when the microbubble load was increased. Findings from this research support the underpinning theories of a sequential arterial-line filter design and suggest that improvements in microbubble filtration may be possible using this technique.

The relationship between red blood cell deformability metrics and perfusion of an artificial microvascular network

The ability of red blood cells (RBC) to undergo a wide range of deformations while traversing the microvasculature is crucial for adequate perfusion. Interpretation of RBC deformability measurements performed in vitro in the context of microvascular perfusion has been notoriously difficult. This study compares the measurements of RBC deformability performed using micropore filtration and ektacytometry with the RBC ability to perfuse an artificial microvascular network (AMVN). Human RBCs were collected from healthy consenting volunteers, leukoreduced, washed and exposed to graded concentrations (0-0.08%) of glutaraldehyde (a non-specific protein cross-linker) and diamide (a spectrin-specific protein cross-linker) to impair the deformability of RBCs. Samples comprising cells with two different levels of deformability were created by adding non-deformable RBCs (hardened by exposure to 0.08% glutaraldehyde) to the sample of normal healthy RBCs. Ektacytometry indicated a nearly linear decline in RBC deformability with increasing glutaraldehyde concentration. Micropore filtration showed a significant reduction only for concentrations of glutaraldehyde higher than 0.04%. Neither micropore filtration nor ektacytometry measurements could accurately predict the AMVN perfusion. Treatment with diamide reduced RBC deformability as indicated by ektacytometry, but had no significant effect on either micropore filtration or the AMVN perfusion. Both micropore filtration and ektacytometry showed a linear decline in effective RBC deformability with increasing fraction of non-deformable RBCs in the sample. The corresponding decline in the AMVN perfusion plateaued above 50%, reflecting the innate ability of blood flow in the microvasculature to bypass occluded capillaries. Our results suggest that in vitro measurements of RBC deformability performed using either micropore filtration or ektacytometry may not represent the ability of same RBCs to perfuse microvascular networks. Further development of biomimetic tools for measuring RBC deformability (e.g. the AMVN) could enable a more functionally relevant testing of RBC mechanical properties.

Technical feasibility study of an onshore ballast water treatment system

To fulfill the requirements of Guidelines for approval of ballast water management system (G8), a set of onshore ballast water treatment equipment utilizing micro-pore ceramic filtration (MPCF) and UV radiation (MPCF&UV) system was designed and set up with a maximum flow rate of 80 m3·h−1. Technical feasibilities of MPCF&UV system were evaluated in three areas: removal efficiencies of indicator organism and oceanic bacteria, perdurability of a ceramic filter, and application on native seawater. The results showed that no indicator organism (Dunaliella) or oceanic bacteria was detected after treatment of 20 L MPCF and UV radiation at 1.3×104 μW·s·cm−2. A 20 L ceramic filter can run continuously for 5.3 h at the flow rate of 15 m3·h−1 before its pressure drop up to 0.195 MPa. The removal percentage of total plankton amounts were 91.9% at a flow rate of 70 m3·h−1 by 80 L MPCF and UV radiation at 1.3×104 μW·s·cm−2.

Three-Dimensional Endothelium-Lined Microfabricated Channels as in Vitro assays for Sickle Cell Vaso-Occlusion

Vaso-occlusion in sickle cell disease is fundamentally biophysical in nature, involving a complex set of cellular interactions. Originally attributed solely to the entrapment of abnormally rigid sickle red cells (RBCs) in the microcirculation, this process is now known to involve the decreased deformability of white blood cells (WBCs) and increased endothelial adhesion to different cell types (sickle RBCs, reticulocytes, WBCs, platelets). These biophysical interactions, which are also mediated biochemically by a variety of soluble factors (coagulation proteins, inflammatory mediators, reactive oxygen species, free hemoglobin, etc.), then ultimately lead to microvascular obstruction. However, in vitro experimental approaches that measure the vaso-occlusive properties of sickle blood cells have been unable to separate the contributions of decreased cell deformability and cell-cell adhesion in a single assay. Historically, cell deformability is measured using techniques such as micropipette aspiration, micropore filtration, and ektacytometry, whereas adhesive interactions between the endothelium, blood cells, and soluble factors are assessed using endothelial-lined flow chamber assays. No existing technique effectively evaluates both cell deformability and cell adhesion simultaneously, which is required to comprehensively study sickle cell vaso-occlusion.In the current study, we present an “endothelialized” microfluidic system that simultaneously integrates cell deformability and cell adhesion under physiologic microvascular flow conditions to investigate the underlying biophysical mechanisms of sickle cell vaso-occlusion. Briefly, a layer of human umbilical vein endothelial cells (HUVECs) was cultured along the inner walls of microfabricated microchannels made of the biocompatible and optically transparent polymer polydimethylsiloxane, or PDMS (Figure). Standard lab-on-chip photolithography techniques were utilized to design and create the microchannels, which geometrically emulate a branching microvasculature network with the smallest lumen size approximately 15 μm in diameter. Then, the inner lining of the microchannels were coated with fibronectin and seeded with HUVECs, which grew to a confluent monolayer within 4–5 days and encompassed the entire inner surface in all three dimensions.Whole blood or cell suspensions can then be perfused into the system, and cells can be visualized under flow using standard immunofluorescence microscopy techniques, which allows for identification of specific cellular subpopulations. Pressure and flow rate can be tightly controlled, and cell transit time, time to cell aggregation or obstruction of the system can be recorded using automated image processing software. The effect of different cell types and biologic modifiers (i.e. cytokines, coagulation factors, etc) on in vitro vaso-occlusion can be systematically analyzed and quantified. Furthermore, adhesion-blocking antibodies and drugs that increase cell deformability can be used to quantify the vaso-occlusive effect of adhesion versus cell rigidity. As this system physically constrains cells three-dimensionally in lumens the size of the human microvasculature, physical cellular interactions that are likely to occur in vivo and the endothelial cell response to these processes can also be directly evaluated with live cell fluorescence, immunofluorescence or western blotting.Overall, this system will provide a quantitative and controlled approach to investigate the underlying complex biophysical processes that govern sickle cell vaso-occlusion. In addition, this platform will also serve as a useful complimentary technology to in vivo experiments using sickle cell mouse models. [Display omitted]

Micro-pore Ceramic Filter and UV System as Ballast Water Treatment: Preliminary Study on Chlorella Removal and Bacteria Inactivation

AbstractA combination system of micro-pore ceramic filtration and UV radiation was built up for ballast water treatment. Two treatment methods, single micro-pore ceramic filtration and micro-pore filtration and UV radiation, were studied in the research, respectively. Chlorella removal rates and total oceanic bacteria inactivation rates were measured at different flow rates (0.6 ~ 4.7 m

Sampling of gaseous trace constituents from air on active carbon

Next to the collection of particulate matter from air by micropore filtration, gaseous trace constituents may be sampled by an additional or separate active carbon cartridge. Either reversible or irreversible adsorption results. Important respective applications are the assay 222Rn (emanation) and the collection of mercury vapor. Formulation of both cases and their consequences for routine application are given here.

Endotoxin binding to erythrocyte membrane and erythrocyte deformability in human sepsis and in vitro

Several studies have shown that lipopolysaccharide and lipid A impair red blood cell deformability and. However, it is unclear whether impaired red blood cell deformability is associated with binding of lipopolysaccharide to the red blood cell membrane. Analysis of hydroxymyristic acid content in red blood cell membranes and red blood cell deformation in patients with Gram-negative septicemia and after incubation of red blood cells from healthy adults with 100 microg of lipid A or 1 mg of lipopolysaccharide per milliliter of red blood cell in buffer solution and in whole blood. Hydroxymyristic acid is a fatty acid of the lipid A part of lipopolysaccharide in most Gram-negative bacteria. University research laboratories. Ten healthy adults and four patients with clinical and laboratory signs of septicemia. Blood sampling. Red blood cell deformation was measured with a laser-diffraction shearing device (Rheodyn) and a computerized micropore filtration system (CTA). Lipopolysaccharide and lipid A binding to red blood cell membranes was studied by measuring the amide-linked hydroxymyristic acid by gas chromatography. The detection rates of hydroxymyristic acid were 82% for lipopolysaccharide and 79% for lipid A in buffer solution. In membranes of washed red blood cell, the detection rates of lipopolysaccharide and lipid A were 0.26 +/- 0.03% (2.6 +/- 0.3 microg/mL) and 1.3 +/- 0.5% (1.3 +/- 0.5 microg/mL), and in red blood cell membranes of whole blood the detection rates were 2.6% (25.5 microg/mL) and 4.1% (4.1 microg/mL), respectively. The lipopolysaccharide content in red blood cell membranes of septic patients ranged from 47 to 103 microg/mL of red blood cell. Red blood cell deformation in the Rheodyn and in the CTA were not influenced by lipopolysaccharide incubated with washed red blood cells. In the Rheodyn, red blood cell deformation was significantly decreased by 18% after lipid A incubation in washed red blood cells, by 26% after lipopolysaccharide incubation in whole blood, and by 31% in septic patients. Similar effects were observed when we used the CTA. Red blood cell deformation is decreased in septic patients, after incubation of washed red blood cells with lipid A and of whole blood with lipopolysaccharide. Lipopolysaccharide did not influence red blood cell deformation after incubation with washed red blood cells. The decrease of red blood cell deformation was related to the amount of hydroxymyristic acid measured in red blood cell membranes, suggesting that endotoxin binding directly affects mechanical properties of red blood cells.

Influence of Storage on Red Blood Cell Rheological Properties

Background. It is known that the age of transfused blood is a risk factor for the development of multiple organ failure in surgical patients. However, the character of hemorrheological changes in stored blood as well as the time when they appear remains disputable. We assumed that blood storage was accompanied by a progressive decrease of RBC deformability and rheological disorders. The degree of rheological disturbances should be directly proportional to the number of RBC with altered geometry.Materials and methods. Nine packages of RBC kept in adenine saline solution were examined from the 5th to the 42nd day of storage. RBC deformability index (DI) was determined by micropore filtration technique. RBC shape was estimated by means of scanning electron microscopy. Blood clotting time was measured by Sonoclot coagulation analyzer.Results. Significant alterations of RBC shape started at the second week of storage and progressed during the rest of the storage period. RBC shape changes were accompanied by progressive decrease in DI and increase in hemolysis and acidosis. The correlation index between the percentage of abnormally shaped RBC and DI was −0.81 (P = 0.0258). Blood clotting progressively decreased after 2 weeks of storage, probably due to the exhaustion of some procoagulant plasma factors.Conclusions. Serious hemorrheological disorders, including the decrease in RBC deformability secondary to shape abnormalities, acidosis, and the decrease of blood clotting, start already at the second week of storage and progress up to the end of the storage period. Transfusion of packed RBC older than 7 days may contribute to hemorrheological disorders in critically ill patients.

The Effects of Aged Garlic Extract on Lipid Peroxidation and the Deformability of Erythrocytes

The effects of aged garlic extract (AGE) on lipid peroxidative damage and the deformability of erythrocytes were evaluated in rats. The deformability of erythrocytes was measured using the micropore filtration method. AGE significantly prevented the decrease of erythrocyte deformability induced by lipid peroxidation in a dose-dependent manner. The addition of AGE significantly inhibited an increase in thiobarbituric acid-reactive substances (TBARS) and hemolysis rate and prevented the loss of intraerythrocytic ATP and 2,3-diphosphoglycerate (2,3-DPG) in oxidized erythrocytes. Moreover, AGE significantly suppressed not only the hemolysis rate induced by peroxidation but also hemolysis due to nonperoxidation. These results suggest the possibility that AGE improves microcirculation and rheological blood properties and preserves the structure and function of erythrocytes not only through an antioxidant process, but also via the glycolysis pathway and membrane stabilization of erythrocytes.

Synthesis and chromatographic characterization of [Tc-99m]technetium-humic acid species

A natural moor soil humic acid (HA) was labeled with Tc-99m via reduction of pertechnetate with stannous chloride. The humic acid species obtained were characterized by thin layer chromatography (TLC), gel permeation chromatography (GPC), sequential chromatographic analysis (SCA), paper electrophoresis and micropore filtration. Labeling was found to take place in all ranges of molecular weight. Due to the complex humic acid composition and the formation of hydroxo species the labeling yields strongly depend on the separation conditions, ranging from 42% to 80%. The pH-dependent distribution of mobile and immobile species was determined by SCA for HTcO4, HA and for Tc-HA-species obtained after labeling. Geochemically relevant properties of HA remain unchanged after the labeling procedure, only macroscopic amounts of the reducing agent Sn2+give rise to a shift to lower molecular weight species.

Hemodialysis Neutropenia Correlates with a Decreased Filterability and an Increase in the Number of Cytoplasmic Actin Filaments in Peripheral Blood Neutrophils, Which Is Preceded by a Decrease in the Number of Surface Expression of L-Selectin

In order to clarify the precise cellular mechanism of hemodialysis neutropenia, we examined the changes in the viscoelasticity of peripheral blood neutrophils using both the micropore and the microchannel filtration methods, and the changes in the neutrophil surface expression of Mac-1, L-selectin and sialyl Lewis X and the cytoplasmic expression of the actin filaments using a flow cytometric analysis during a dialysis session. Five patients with chronic renal failure were selected who showed a nadir leukocyte count in peripheral blood at 30 min after the initiation of the dialysis session. The neutrophil count also reached a nadir at 30 min and thereafter returned to almost the predialysis level by 180 min. Both the micropore filtration time and the microchannel passage time, which reflect the viscoelasticity of the peripheral blood neutrophils, correlated inversely with the neutrophil count. At the nadir of neutropenia, the neutrophils were observed to have become both adhesive and viscoelastic. The actin filaments in the neutrophil cytoplasm gradually increased in number from the start of dialysis, reaching a peak level at 30 min, and thereafter decreasing to predialysis levels. The Mac-1 expression continuously increased up from 30 min until the end of dialysis. The L-selectin expression first decreased at 15 min, but thereafter returned to predialysis levels within 60 min. The SLe^x expression did not change throughout the course of the session. These results thus indicated the neutrophil counts during a dialysis session to inversely correlate with the viscoelasticity of the neutrophils expressed by the micropore filtration time or microchannel passage time, which possibly depends on the contents of cytoplasmic actin filaments. In addition, the shedding of L-selectin from neutrophil surface may also be involved in the first step of hemodialysis neutropenia.

Increased rigidity and priming of polymorphonuclear leukocytes in sepsis.

It has been proposed that abnormal mechanical properties may contribute to capillary retention of polymorphonuclear leukocytes (PMN) in sepsis, leading to the development of organ dysfunction. The present study was designed to determine whether PMN rigidity is increased in severe sepsis, and whether changes in the rheologic behavior of PMN correlate with the clinical course in sepsis. Eighteen adults with severe sepsis were studied over a period of 14 d; 11 survived and seven died. PMN deformation behavior was investigated via micropore filtration, using the cell transit analyzer. On Day 0, PMN rigidity was 2.5-fold greater for sepsis patients than for five normal controls (p < 0.001). PMN rigidity progressively improved over the 14 d study period for patients who recovered, but not for those who died; clinical indicators correlated with PMN rigidity. Patient PMN also exhibited a 5-fold greater increase in rigidity in response to formyl-methionylleucylphenylalanine (fMLP) than did control PMN. Both the increased rigidity and enhanced response to fMLP could be simulated in vitro by incubation of normal PMN with tumor necrosis factor-alpha (TNF-alpha). We conclude that circulating PMN are more rigid in severe sepsis, and are "primed" for an augmented response to chemotactic stimuli. These findings support the hypothesis that cytokine-mediated increases of PMN rigidity may lead to sequestration of these cells in capillaries and to the consequent impairment of microvascular perfusion in sepsis.

Red blood cell deformability in sepsis.

The microcirculatory disturbances in sepsis have prompted micropore bulk-filtration studies of red blood cell (RBC) mechanical behavior (i.e., deformability). However, these prior reports may not solely reflect RBC behavior because of possible white blood cell (WBC) occlusion of the filter pores. The present study was designed to examine RBC mechanical alterations in human and experimental sepsis using techniques that are not affected by WBC artifacts. RBC were obtained from adult patients with sepsis and from healthy control donors. RBC were also obtained from Swiss-albino rats in which experimental sepsis was induced via cecal ligation-puncture. Red cell mechanical behavior was tested using a computerized micropore filtration system (CTA) and a laser-diffraction shearing device (LORCA); the latter provides the extent of RBC deformation at various stresses and the time constant for RBC shape recovery. Salient findings include: (1) for human RBC, significantly decreased deformability at fluid shear stresses < 5 Pa (LORCA) yet no differences from control with the CTA; (2) for rat RBC in experimental sepsis, significant decreases of deformability and shape-recovery time constant (LORCA) but no differences with the CTA. We conclude that RBC deformability is reduced in sepsis but that micropore bulk-filtration methods may not be appropriate for detecting these changes.

Autotransfusion after total knee arthroplasty: Effects on blood cells, plasma chemistry and whole blood rheology

Postoperative drain blood was collected and reinfused using the ConstaVac system (Stryker, Kalamazoo, MI) in 30 patients after total knee arthroplasty. Of the total 1.1-L volume of postoperative bleeding, 60% was reinfused. No clinical complications were observed. Differences between venous blood and drain blood and between venous blood and drain blood after separate incubation were studied with respect to acidic and inflammatory effects on blood cells, plasma chemistry, and whole blood rheology. In drain blood, leukocyte and platelet counts were reduced ( P < .001), probably as a result of consumption in the wound. Acidic incubation occurs in the drain container because of production of lactate from glucose, with a minimum pH at 5 hours of 7.2. The low pH caused slight but significant erythrocyte swelling ( P < .01). The complement C3d indicated leukocyte activation, although of modest magnitude. Despite incubation and complement activation, maximum erythrocyte hemolysis after 24 hours of incubation was less than 1%. Drain blood showed a lower resistance against micropore filtration than venous blood ( P < .001), mainly because of the reduced number of leukocytes, and remained unchanged with incubation. Although the autotransfusion system can be improved with respect to blood quality, filtered drain blood should be considered acceptable for reinfusion.

Peripheral blood neutrophil rheology measured by micropore filtration reflects Behçet's disease activity well

Activated neutrophils take a long time to pass through a narrow lumen like a micropore, and are supposed to play a deteriorating effect on microcirculation. Although the activation of neutrophils has been demonstrated in Behçet's disease, nobody analyzes the clinical activity of the disease by means of the rheological measure of neutrophils activity. Using a micropore (pore diameter 5 μm) filtration technique, we measured the filtration time of peripheral blood neutrophils, as a rheological measure of their activity, in order to determine the clinical activity of Behçet's disease. Twenty-one patients with Behçet's disease and 14 healthy control individuals were enrolled in the study. Symptoms and signs exhibited in the patients led us to distinguish the Behçet's disease into inactive and active cases. The latter were further differentiated into cases with absent symptoms and with present symptoms. Neutrophil filtration times were 11.5 ± 4.8 s in the active cases with present symptoms, which were significantly ( P < 0.05) larger than those (7.4 ± 1.9 s) in the active cases with absent symptoms. The latter filtration times were further significantly ( P < 0.001) larger than values (3.7 ± 1.3 s) in the inactive cases and also those (4.8 ± 1.2 s) in control subjects. Furthermore, increases in the filtration time obtained immediately after the exposure of cells to the chemotactic peptide formyl-methionyl-leucyl-phenylalanine (FMLP; 10 nM) were significantly ( P < 0.01) larger in the active cases with present symptoms than those in the active cases with absent symptoms. The latter were also larger, but not significantly, than those in the inactive cases, and were significantly ( P < 0.01) larger than those in control subjects. The present results demonstrate that the micropore filtration method reflects well the rheological activity of neutrophils as well as the clinical status of Behçet's disease. This method is much better than the measurement of O 2 − production to differentiate between active cases with absent symptoms and inactive patients or even control individuals. Furthermore, it is more sensitive and useful than laboratory data like the CRP value or the number of peripheral blood neutrophils.