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

Real time visualization and characterization of platelet deposition under flow onto clinically relevant opaque surfaces

<p>Although the thrombogenic nature of the surfaces of cardiovascular devices is an important aspect of blood biocompatibility, few studies have examined platelet deposition onto opaque materials used for these devices in real time. This is particularly true for the metallic surfaces used in current ventricular assist devices (VADs). Using hemoglobin depleted red blood cells (RBC ghosts) and long working distance optics to visualize platelet deposition, we sought to perform such an evaluation. Fluorescently labeled platelets mixed with human RBC ghosts were perfused across six opaque materials (a titanium alloy (Ti6Al4V), silicon carbide (SiC), alumina (Al₂O₃), 2-methacryloyloxyethyl phosphorylcholine polymer coated Ti6Al4V (MPC-Ti6Al4V), yttria partially stabilized zirconia (YZTP), and zirconia toughened alumina (ZTA)) for 5 min at wall shear rates of 400 and 1000 s⁻¹. Ti6Al4V had significantly increased platelet deposition relative to MPC-Ti6Al4V, Al₂O₃, YZTP, and ZTA at both wall shear rates (<em>p</em> < 0.01). For all test surfaces, increasing the wall shear rate produced a trend of decreased platelet adhesion. The described system can be a utilized as a tool for comparative analysis of candidate blood-contacting materials with acute blood contact.</p

Cell free layer width

<p>Cell-free region (CFL) normalized to baseline for control (black, N=6 animals) and DRP treated animals (gray, N=5 animals) presented as mean ± SEM. In the control group there is a significant increase in CFL at 15, 25 and 35 minutes of 20.2 ± 0.02% (SEM), 20.1 ± 0.01% and 21.1 ± 0.01%, respectively. After administration of DRP there is a significant reduction in CFL compared to baseline, at 5 min CFL is 87.8 ± 0.02% of baseline and 83.6 ± 0.01%, 92.0 ± 0.01%, and 86.5 ± 0.01% at 15, 25 and 35 minutes, respectively. * P<0.05 from corresponding baseline dimensions before administration of DRP or saline, † P<0.05 compared to DRP at same moment in time.</p

Parametric study of blade tip clearance, flow rate, and impeller speed on blood damage in rotary blood pump.

Phenomenological studies on mechanical hemolysis in rotary blood pumps have provided empirical relationships that predict hemoglobin release as an exponential function of shear rate and time. However, these relations are not universally valid in all flow circumstances, particularly in small gap clearances. The experiments in this study were conducted at multiple operating points based on flow rate, impeller speed, and tip gap clearance. Fresh bovine red blood cells were resuspended in phosphate-buffered saline at about 30% hematocrit, and circulated for 30 min in a centrifugal blood pump with a variable tip gap, designed specifically for these studies. Blood damage indices were found to increase with increased impeller speed or decreased flow rate. The hemolysis index for 50-microm tip gap was found to be less than 200-microm gap, despite increased shear rate. This is explained by a cell screening effect that prevents cells from entering the smaller gap. It is suggested that these parameters should be reflected in the hemolysis model not only for the design, but for the practical use of rotary blood pumps, and that further investigation is needed to explore other possible factors contributing to hemolysis.</p

An arteriole with the video intensity profile corresponding to a single horizontal line of pixels

<p>A. Portion of a 43.9 µm arteriole used in the analysis. RBCs, appearing dark, are located on the left side of this panel. The vessel wall and muscle tissue are located on the right side of the image. B. The video intensity profile which corresponds to a single line of pixels indicated by the dotted line in the left panel. All 200 lines of the image were analyzed based on their video-intensity profile. The inner vessel wall, RBC column edge, and maximal and minimal pixel value within the vascular lumen are indicated by arrows.</p

Microchannel bifurcation setup

<p>A. Microchannel experimental set-up. The feed reservoir and loop are positioned on the right. Blood enters from the top and as it traverses through the feed channel, a fraction is aspirated into a 50 µm parent branch which passes straight through the plastic mold to the discharge port which is connected the top syringe pump. The parent branch gives rise to a 50 µm daughter branch which is connected to the syringe pump on the bottom. Black arrows indicate the direction of blood flow. B. A microscopic view (10x magnification) of 50 µm bifurcation showing smooth, continuous contours. C. Photo of microchannel showing feed loop, parent branch, and daughter branch.</p

Data analysis

<p>An example of a section of an artery before data analysis (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0077252#pone-0077252-g004" target="_blank">Figure 4A</a>) and after tracing the RBC column and vessel wall over a length of 200 pixels (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0077252#pone-0077252-g004" target="_blank">Figure 4B</a>). The difference between vessel wall and RBC column is defined as the CFL.</p

Conversion to methemoglobin by oxidative treatment of HSC culture increases the magnetic cell fractional concentration.

<p>MM histograms (A) and dot plots (B) of HSC culture before and after the oxidative treatment used for conversion of Hb to paramagnetic metHb and used to determine the fractional concentration of maturing RBCs in culture.</p

Magnetic separation of HSC cultures improves viscoelastic properties of the cells.

<p>Deformability of cells in positive fraction (A) and negative fraction (B). Note increased frequency of dark, elongated objects, associated with hemoglobin containing deformable cells characteristic of mature RBC, in panel A.</p

Donor blood oxyHb RBC settling velocity as a control for HSC culture analysis.

<p>The blood settling velocity histogram and its cumulative frequency distribution are shown. The cut-off settling velocity was set at 0.995 cumulative frequency for the subsequent HSC culture analysis.</p