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Dynamics of Blood Flow and Thrombus Formation in a Multi-Bypass Microfluidic Ladder Network

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Abstract

The reaction dynamics of a complex mixture of cells and proteins, such as blood, in branched circulatory networks within the human microvasculature or extravascular therapeutic devices such as extracorporeal oxygenation machine (ECMO) remains ill-defined. In this report we utilize a multi-bypass microfluidics ladder network design with dimensions mimicking venules to study patterns of blood platelet aggregation and fibrin formation under complex shear. Complex blood fluid dynamics within multi-bypass networks under flow were modeled using COMSOL. Red blood cells and platelets were assumed to be non-interacting spherical particles transported by the bulk fluid flow, and convection of the activated coagulation factor II, thrombin, was assumed to be governed by mass transfer. This model served as the basis for predicting formation of local shear rate gradients, stagnation points and recirculation zones as dictated by the bypass geometry. Based on the insights from these models, we were able to predict the patterns of blood clot formation at specific locations in the device. Our experimental data was then used to adjust the model to account for the dynamical presence of thrombus formation in the biorheology of blood flow. The model predictions were then compared to results from experiments using recalcified whole human blood. Microfluidic devices were coated with the extracellular matrix protein, fibrillar collagen, and the initiator of the extrinsic pathway of coagulation, tissue factor. Blood was perfused through the devices at a flow rate of 2 µL/min, translating to physiologically relevant initial shear rates of 300 and 700 s−1 for main channels and bypasses, respectively. Using fluorescent and light microscopy, we observed distinct flow and thrombus formation patterns near channel intersections at bypass points, within recirculation zones and at stagnation points. Findings from this proof-of-principle ladder network model suggest a specific correlation between microvascular geometry and thrombus formation dynamics under shear. This model holds potential for use as an integrative approach to identify regions susceptible to intravascular thrombus formation within the microvasculature as well as extravascular devices such as ECMO.

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Abbreviations

PDMS:

Polydimethylsiloxane

RBCs:

Red blood cells

TF:

Tissue factor

VWF:

Von Willebrand factor

DiOC6 :

3,3′-Dihexyloxacarbocyanine iodide

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Acknowledgments

We thank Dr. András Gruber for insightful comments and Chantal Wiesenekker for technical assistance. This work was supported by West Virginia University startup funds awarded to J. Maddala and by grants from the National Institutes of Health (R01HL101972, R01GM116184, R44HL126235). O.J.T. McCarty is an American Heart Association Established Investigator (13EIA12630000).

Conflict of interest

J. Zilberman-Rudenko, J.L. Sylman, H.H.S. Lakshman, O.J.T. McCarty and J. Maddala declare no competing financial interests.

Human and Animal Rights and Informed Consent

All procedures followed were in accordance with the ethical standards of the responsible committee on human experimentation (institutional and national) and with the Helsinki Declaration of 1975, as revised in 2000 (5). Informed consent was obtained from all patients for being included in the study. All human subject research was carried out in accordance with institutional guidelines approved by the Oregon Health & Science University Institutional Review Board. No animal studies were carried out by the authors for this article.

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Correspondence to Jeevan Maddala.

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Associate Editor Michael R. King oversaw the review of this article.

O. J. T. McCarty and J. Maddala are co-senior authors.

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Supplemental Video 1

Initial blood flow dynamics within the ladder microfluidic device. Recalcified whole human blood was perfused at a 2 µL/min flow rate through a PDMS-ladder network coated with collagen and tissue factor; real-time video of the blood flow was recorded differential using interference contrast, DIC, microscopy. Supplementary material 1 (WMV 4603 kb)

Supplemental Video 2

Blood flow dynamics within the ladder microfluidic device after 10 min of blood perfusion. Recalcified whole human blood was perfused at a 2 µL/min flow rate through a PDMS-ladder network coated with collagen and tissue factor; real-time video of the blood flow was recorded differential using interference contrast, DIC, microscopy at the 10 min time point. Supplementary material 2 (WMV 447 kb)

Supplemental Fig. 1

Quantification of the spatial distribution of thrombus formation. DiOC6-labeled whole human blood was perfused at a 2 µL/min flow rate through a PDMS-ladder network coated with collagen and tissue factor; images of thrombus formation were recorded using differential interference contrast, DIC microscopy. The thrombus surface area was quantified for 5 experiments using ImageJ. Supplementary material 3 (TIFF 976 kb)

Supplementary material 4 (PDF 110 kb)

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Zilberman-Rudenko, J., Sylman, J.L., Lakshmanan, H.H.S. et al. Dynamics of Blood Flow and Thrombus Formation in a Multi-Bypass Microfluidic Ladder Network. Cel. Mol. Bioeng. 10, 16–29 (2017). https://doi.org/10.1007/s12195-016-0470-7

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