Peng Kai Ong

The cell-free layer in microvascular blood flow.

This review discusses the cell-free layer at the wall of the microcirculatory vessels with emphasis on the influence of blood rheological parameters on its formation as well as its possible effects on microvascular functions. Understanding cell-free layer characteristics in the microcirculation has been of increased interest for its possible influence on physiological function. The availability of better imaging and measurement techniques in recent years has created an excellent opportunity for researchers to examine the cell-free layer in much greater detail than possible previously. The most recent approach enables capturing spatial and temporal information of cell-free layer characteristics which can provide new insights to how cell-free layer width and its variations can impact physiological function. Cell-free layer formation is dependent on diameter of the vessel, flow rate, and rheological properties of blood including hematocrit, red blood cell aggregation, and deformability. Specifically in microvessels, its formation can also be affected by the presence of the glycocalyx layer that lines the luminal surface of the vessel wall. As the cell-free layer is omnipresent in microcirculatory vessels, its formation at the vessel wall could affect function throughout the microvascular network by altering wall shear stress, local hematocrit, flow distribution, effective viscosity and nitric oxide production as well as its scavenging by red blood cells.

Effect of erythrocyte aggregation and flow rate on cell-free layer formation in arterioles

Formation of a cell-free layer is an important dynamic feature of microcirculatory blood flow, which can be influenced by rheological parameters, such as red blood cell aggregation and flow rate. In this study, we investigate the effect of these two rheological parameters on cell-free layer characteristics in the arterioles (20-60 mum inner diameter). For the first time, we provide here the detailed temporal information of the arteriolar cell-free layer in various rheological conditions to better describe the characteristics of the layer variation. The rat cremaster muscle was used to visualize arteriolar flows, and the extent of aggregation was raised by dextran 500 infusion to levels seen in normal human blood. Our results show that cell-free layer formation in the arterioles is enhanced by a combination of flow reduction and red blood cell aggregation. A positive relation (P < 0.005) was found between mean cell-free layer widths and their corresponding SDs for all conditions. An analysis of the frequency and magnitudes of cell-free layer variation from their mean value revealed that the layer deviated with significantly larger magnitudes into the red blood cell core after flow reduction and dextran infusion (P < 0.05). In accordance, the disparity of cell-free layer width distribution found in opposite radial directions from its mean became greater with aggregation in reduced flow conditions. This study shows that the cell-free layer width in arterioles is dependent on both flow rate and red blood cell aggregability, and that the temporal variations in width are asymmetric with a greater excursion into the red blood cell core than toward the vessel wall.

Spatio-temporal variations in cell-free layer formation near bifurcations of small arterioles

Blood flow partitioning at an arteriolar bifurcation could lead to spatio-temporal variations in cell-free layer formation in the upstream and downstream vessels of the bifurcation. To investigate this effect, we quantitatively analyzed characteristics of the cell-free layer in the vicinity of an arteriolar bifurcation in the rat cremaster muscle in normal physiological flow conditions. To simulate hemorheological relevance to humans, red blood cell aggregation was elevated by infusion of Dextran 500 to levels seen in humans in normal states. Spatial variations of the layer width were observed in both the parent and larger daughter vessels. A more pronounced attenuation of the layer width was generally observed in the parent vessel at its wall adjacent to the side branch than at its opposite wall. A thicker layer width was consistently found at the opposite than adjacent wall of the larger daughter vessel. Accordingly, large asymmetries of the layer widths could be developed on opposite sides of the larger daughter vessel, which were significantly greater (P<0.01) than those observed in the parent vessel. A positive correlation was generally observed between mean layer widths in the downstream vessel and on the side of the parent vessel from which bulk of the flow enters the downstream vessel. The fraction of the downstream layer formation constituted by the side branch decreased with increasing flow fraction in this vessel. These findings confirmed the modulation of the cell-free layer formation near an arteriolar bifurcation, implicated by flow separation at the bifurcation.

Cell-Free Layer Formation in Small Arterioles at Pathological Levels of Erythrocyte Aggregation

Please cite this paper as: Ong, Jain, Namgung, Woo and Kim (2011). Cell‐Free Layer Formation in Small Arterioles at Pathological Levels of Erythrocyte Aggregation. Microcirculation 18(7), 541–551.

Temporal variations of the cell-free layer width may enhance NO bioavailability in small arterioles: Effects of erythrocyte aggregation

Recently, we have shown that temporal variations in the cell-free layer width can potentially enhance nitric oxide (NO) bioavailability in small arterioles. Since the layer width variations can be augmented by red blood cell aggregation, we tested the hypothesis that an increase in the layer width variations due to red blood cell aggregation could provide an underlying mechanism to improve NO bioavailability in the endothelium and promote vasodilatory effects. Utilizing cell-free layer width data acquired from arterioles of the rat cremaster muscle before and after dextran infusion in reduced flow conditions (wall shear stress=0.13-0.24Pa), our computational model predicted exponential enhancements of NO bioavailability in the endothelium and soluble guanylyl cyclase (sGC) activation in the smooth muscle layer with increasing temporal variability of the layer width. These effects were mediated primarily by the transient responses of wall shear stress and NO production rate to the layer width variations. The temporal variations in the layer width were significantly enhanced (P<0.05) by aggregation, leading to significant improvements (P<0.05) in NO bioavailability and sGC activation. As a result, the significant reduction (P<0.05) of sGC activation due to the increased width of the layer after aggregation induction was diminished by the opposing effect of the layer variations. These findings highlighted the possible enhancement of NO bioavailability and vascular tone in the arteriole by the augmented layer width variations due to the aggregation.

A comparative study of histogram-based thresholding methods for the determination of cell-free layer width in small blood vessels

We have recently proposed a computer-based method utilizing a thresholding algorithm (the Otsu method) to provide a convenient way of measuring the cell-free layer width in vivo and in vitro. However, this method does not seem to be a universal method that can be applied to all microvascular studies. Thus, we examined four different histogram-based thresholding algorithms (Otsu, intermode, minimum and second peak) to provide a technical suggestion on the selection of a suitable thresholding algorithm for the cell-free layer measurement. All the measurements were taken in microvascular flows in the rat cremaster muscle recorded with a high-speed camera. The width of the cell-free layer manually measured was compared with that determined by the automated method utilizing the four thresholding algorithms. With our experimental system, results showed that the cell-free layer width determined by the minimum algorithm was in best accordance with the manual measurement. We concluded that the accuracy of the automated methods for determination of the cell-free layer width would depend on the image quality, in particular on the contrast between the red blood cell core and background, which might differ due to the different microscopic setup. Therefore, one may need to examine several appropriate thresholding methods when selecting the best suitable algorithm for the experimental conditions.

Effect of Erythrocyte Aggregation on Spatiotemporal Variations in Cell‐Free Layer Formation Near on Arteriolar Bifurcation

To investigate how red blood cell aggregation could modulate the spatial variations in cell‐free layer formation in the vicinity of an arteriolar bifurcation.

An automated method for cell-free layer width determination in small arterioles

Histogram-based thresholding techniques utilized for cell-free layer width measurement in arteriolar flow may produce an overestimation of the layer width since they do not consider faint shaded regions near the vessel wall as part of the erythrocyte column. To address this problem, we developed a new method for detecting the boundary of the erythrocyte column based on an edge detection algorithm. This automated method (grayscale method) provides local detections of the inner vessel wall as well as the boundary between the cell-free layer and the erythrocyte column without binarization of grayscale images. The cell-free layer width measurements using the grayscale method and existing techniques (minimum method and Otsus method) were compared with those determined manually in arteriolar flows of the rat cremaster muscle. In the absence of the shaded regions, values obtained by the grayscale method and minimum method were statistically in good agreement with the manual method but not in the case of Otsus method. When the faint shaded regions were present, the grayscale method appeared to produce more accurate results than the minimum method and Otsus method.

Effects of cell-free layer formation on NO/O2 bioavailability in small arterioles.

We developed a new time-dependent computational model for coupled NO/O(2) transport in small arterioles that incorporates potential physiological responses (temporal changes in NO scavenging rate and O(2) partial pressure in blood lumen and NO production rate in endothelium) to the temporal cell-free layer width variations. Two relations between wall shear stress (WSS) and NO production rate based on the linear and sigmoidal functions were considered in this simulation study. The cell-free layer data used for the simulation were acquired from arteriolar flows (D=48.3 ± 1.9 μm) in the rat cremaster muscles under normal flow conditions (WSS=3.4-5.6 Pa). For both cases of linear and sigmoidal relations, temporal layer width variations were found to be capable of significantly enhancing NO bioavailability and this effect was more pronounced in the latter (P<0.0005) than the former (P<0.005). In contrast, O(2) bioavailability in the arteriolar wall was not considerably altered by the temporal layer width variations, irrespective of the relation. Prominent enhancement (P<0.005) of soluble guanylyl cyclase (sGC) activation in the smooth muscle by the temporal layer width variations were predicted for both relations. The extent of sGC activation was generally lower (P<0.01) in the case of the sigmoidal relation than that of the linear relation, suggesting a lesser tendency for arterioles to dilate with the former.

Effect of low molecular weight dextrans on erythrocyte aggregation

Abstract