Erythrocyte enrichment in hematopoietic progenitor cell cultures based on magnetic susceptibility of the hemoglobin

Using novel media formulations, it has been demonstrated that human placenta and umbilical cord blood-derived CD34+ cells can be expanded and differentiated into erythroid cells with high efficiency. However, obtaining mature and functional erythrocytes from the immature cell cultures with high purity and in an efficient manner remains a significant challenge. A distinguishing feature of a reticulocyte and maturing erythrocyte is the increasing concentration of hemoglobin and decreasing cell volume that results in increased cell magnetophoretic mobility (MM) when exposed to high magnetic fields and gradients, under anoxic conditions. Taking advantage of these initial observations, we studied a noninvasive (label-free) magnetic separation and analysis process to enrich and identify cultured functional erythrocytes. In addition to the magnetic cell separation and cell motion analysis in the magnetic field, the cell cultures were characterized for cell sedimentation rate, cell volume distributions using differential interference microscopy, immunophenotyping (glycophorin A), hemoglobin concentration and shear-induced deformability (elongation index, EI, by ektacytometry) to test for mature erythrocyte attributes. A commercial, packed column high-gradient magnetic separator (HGMS) was used for magnetic separation. The magnetically enriched fraction comprised 80% of the maturing cells (predominantly reticulocytes) that showed near 70% overlap of EI with the reference cord blood-derived RBC and over 50% overlap with the adult donor RBCs. The results demonstrate feasibility of label-free magnetic enrichment of erythrocyte fraction of CD34+ progenitor-derived cultures based on the presence of paramagnetic hemoglobin in the maturing erythrocytes. © 2012 Jin et al

Возможности импульсной осциллометрии в диагностике обструкции дыхательных путей легкой степени выраженности

The aim of this study was to investigate a role of impulse oscillometry (IO) for detection of mild bronchial obstruction. Methods. The study involved 87 patients with different respiratory diseases. Bronchial obstruction according to spirometry results was found in 50 patients (the study group). The control group patients did not demonstrate any abnormalities in spirometry, body plethysmography and lung diffusing test. Results. An abnormal increase in frequency-dependent resistive component of respiratory impedance (Rrs) at the oscillation frequency of 5Hrz and 10 Hrz [both the relative oscillation frequency (Rrs5–Rrs20)/Rrs20 and the absolute oscillation frequency (Rrs5–Rrs20)] was found in patients with mild bronchial obstruction. The resonance frequency (fres) shifted towards higher frequency; reactance area (AX) increase and expiratory airflow (DXrs5) limitation were also found in those patients. Rrs5, Rrs20, and the reactive resistance (Xrs5) were within the normal range. Conclusion. Rrs5 и Xrs5 could detect mild bronchial obstruction only in 32 % of the cases. AX is more useful parameter as it increased in 64% of the patients. The severity of bronchial obstruction diagnosed with IO or spirometry was not identical in some cases.Нередко начальным проявлением заболеваний легких являются функциональные нарушения, а именно – нарушение проходимости бронхов. Выявление обструктивного типа вентиляционных расстройств на ранних стадиях является важной клинической задачей, т. к. позволяет своевременно диагностировать болезнь и начать лечение, улучшив тем самым прогноз заболевания. Одним из самых доступных методов, позволяющих выявлять бронхиальную обструкцию, является спирометрия. Необходимое условие проведения спирометрии – правильное выполнение дыхательных маневров, однако при этом требуется хорошая кооперация пациента с медицинским персоналом, что не всегда выполнимо. Импульсная осциллометрия (ИОМ) отличается тем, что измерения осуществляются при спокойном дыхании, активного участия пациента не требуется. Однако остается много неизученных и спорных вопросов, касающихся интерпретации результатов ИОМ. Целью данного исследования явилось изучение возможности ИОМ при диагностике обструкции дыхательных путей легкой степени выраженности. Материалы и методы. Обследованы пациенты (n = 87) с разнообразной бронхолегочной патологией, при этом выявлены вентиляционные нарушения обструктивного типа легкой степени (n = 50). В группе сравнения (n = 37) нарушений по данным спирометрии, бодиплетизмографии и диффузионного теста не отмечено. Результаты. У больных с легкой степенью бронхиальной обструкции, установленной по данным спирометрии, наблюдалось патологическое увеличение частотной зависимости резистивного компонента дыхательного импеданса (Rrs) при частоте осцилляций 5 и 20 Гц (как относительной (Rrs5–Rrs20) / Rrs20, так и абсолютной (Rrs5–Rrs20)), смещение резонансной частоты (fres) в область высоких частот, увеличение площади реактанса (АХ) и экспираторное ограничение потока (DXrs5) при сохранении в пределах нормальных значений Rrs5, Rrs20 и реактивного сопротивления (Xrs5). Заключение. Показано, что при использовании базовых параметров Rrs5 и Xrs5 обструкция легкой степени выраженности, установленная при помощи традиционных функциональных методов, наблюдалась только в 32 % случаев. АХ явился более информативным методом, поскольку изменялся значительно чаще – его увеличение зарегистрировано у 64 % пациентов. Выраженность обструктивных нарушений по данным ИОМ не всегда совпадало с выраженностью нарушений, определяемых при помощи спирометрии

A Quasi-Mechanistic Mathematical Representation for Blood Viscosity

Blood viscosity is a crucial element for any computation of flow fields in the vasculature or blood-wetted devices. Although blood is comprised of multiple elements, and its viscosity can vary widely depending on several factors, in practical applications, it is commonly assumed to be a homogeneous, Newtonian fluid with a nominal viscosity typically of 3.5 cP. Two quasi-mechanistic models for viscosity are presented here, built on the foundation of the Krieger model of suspensions, in which dependencies on shear rate, hematocrit, and plasma protein concentrations are explicitly represented. A 3-parameter Asymptotic Krieger model (AKM) exhibited excellent agreement with published Couette experiments over four decades of shear rate (0–1000 s-1, root mean square (RMS) error = 0.21 cP). A 5-parameter Modified Krieger Model (MKM5) also demonstrated a very good fit to the data (RMS error = 1.74 cP). These models avoid discontinuities exhibited by previous models with respect to hematocrit and shear rate. In summary, the quasi-mechanistic, Modified-Krieger Model presented here offers a reasonable compromise in complexity to provide flexibility to account for several factors that affect viscosity in practical applications, while assuring accuracy and stability

Drag-reducing polymers diminish near-wall concentration of platelets in microchannel blood flow.

The accumulation of platelets near the blood vessel wall or artificial surface is an important factor in the cascade of events responsible for coagulation and/or thrombosis. In small blood vessels and flow channels this phenomenon has been attributed to the blood phase separation that creates a red blood cell (RBC)-poor layer near the wall. We hypothesized that blood soluble drag-reducing polymers (DRP), which were previously shown to lessen the near-wall RBC depletion layer in small channels, may consequently reduce the near-wall platelet excess. This study investigated the effects of DRP on the lateral distribution of platelet-sized fluorescent particles (diam. = 2 μm, 2.5 × 10⁸/ml) in a glass square microchannel (width and depth = 100 μm). RBC suspensions in PBS were mixed with particles and driven through the microchannel at flow rates of 6-18 ml/h with and without added DRP (10 ppm of PEO, MW = 4500 kDa). Microscopic flow visualization revealed an elevated concentration of particles in the near-wall region for the control samples at all tested flow rates (between 2.4 ± 0.8 times at 6 ml/h and 3.3 ± 0.3 times at 18 ml/h). The addition of a minute concentration of DRP virtually eliminated the near-wall particle excess, effectively resulting in their even distribution across the channel, suggesting a potentially significant role of DRP in managing and mitigating thrombosis.</p

Drag-reducing hyaluronic acid increases survival in profoundly hemorrhaged rats

We tested the hypothesis that the infusion of a small volume of a drag-reducing polymer (DRP) solution can prolong survival in rats subjected to lethal hemorrhagic shock (HS; shed 51% of estimated blood volume) in the absence of complete resuscitation with fluids or blood. In this set of experiments, we used a newly designed mixture of hyaluronic acid (molecular weight, â\u88¼2.0 x 10 6 d; 0.4 mg/mL) and polyethylene oxide (molecular weight, â\u88¼4 x 10 6 d; 0.05 mg/mL) dissolved in sterile phosphate-buffered saline. Anesthetized rats were subjected to a volume-controlled HS. During the first 20 min, blood (21.7 mL/kg) was withdrawn. During the next 40 min, additional blood (14 mL/kg) was withdrawn, and during the final 20 min, saline vehicle or saline + DRP (2.8 mLVkg) was simultaneously infused. The survival rate of the rats treated with the hyaluronic acid/polyethylene oxide was significantly higher (P< 0.01). The mean survival times for control and DRP-treated animals were 100.4 ± 9.5 vs. 154.8 ± 7.0 min (P < 0.001). MAP was higher (P < 0.005) and skin perfusion was significantly improved in the DRP-treated group after the end of the DRP infusion. These results support the use of nanomolar concentrations of DRP to prolong survival in rats after lethal HS in the absence of fluid resuscitation. The DRP formulation studied here warrants further evaluation for the amelioration of critical illness associated with profound shock when access to resuscitation fluids may not be possible or delayed. © 2009 by the Shock Society

New insights into the microvascular mechanisms of drag reducing polymers: effect on the cell-free layer.

Drag-reducing polymers (DRPs) significantly increase blood flow, tissue perfusion, and tissue oxygenation in various animal models. In rectangular channel microfluidic systems, DRPs were found to significantly reduce the near-wall cell-free layer (CFL) as well as modify traffic of red blood cells (RBC) into microchannel branches. In the current study we further investigated the mechanism by which DRP enhances microvascular perfusion. We studied the effect of various concentrations of DRP on RBC distribution in more relevant round microchannels and the effect of DRP on CFL in the rat cremaster muscle in vivo. In round microchannels hematocrit was measured in parent and daughter branch at baseline and after addition of DRP. At DRP concentrations of 5 and 10 ppm, the plasma skimming effect in the daughter branch was eliminated, as parent and daughter branch hematocrit were equivalent, compared to a significantly lowered hematocrit in the daughter branch without DRPs. In anesthetized rats (N=11) CFL was measured in the cremaster muscle tissue in arterioles with a diameter of 32.6 ± 1.7 µm. In the control group (saline, N=6) there was a significant increase in CFL in time compared to corresponding baseline. Addition of DRP at 1 ppm (N=5) reduced CFL significantly compared to corresponding baseline and the control group. After DRP administration the CFL reduced to about 85% of baseline at 5, 15, 25 and 35 minutes after DRP infusion was complete. These in vivo and in vitro findings demonstrate that DRPs induce a reduction in CFL width and plasma skimming in the microvasculature. This may lead to an increase of RBC flux into the capillary bed, and thus explain previous observations of a DRP mediated enhancement of capillary perfusion