Engineering Microvasculature for the Study of Thrombosis and Hemostasis

Engineering microvasculature for the study of thrombosis and hemostasis

Ying Zheng

Department of Bioengineering, University of Washington, Seattle, WA.

The microvasculature defines the biological and physical characteristics of the surrounding tissue environment, and plays a role in the initiation and progression of many pathologies. Recapitulation of the microvasculature and an organ-specific vascular niche in vitro can provide a platform to study complex vascular phenomena, and organ-specific functions. In this talk, I will first introduce our approaches on microvascular engineering and describe the structure and functions of these engineered microvessels. Three examples will follow to describe our recent work on the application of microvessels for the studies of thrombosis, hemostasis, and organ specific microvascular events. In the first example, engineered microvessels of complex geometry were generated to examine the pathological responses to endothelial stimulation. Our most striking finding was the capacity of endothelial-secreted von Willebrand factor (VWF) to assemble into thick bundles or complex meshes, depending on the vessel geometry and flow characteristics. Assembly was greatest in vessels of diameter ≤ 300 µm, with strong flow acceleration, and with sharp turns. VWF bundles and webs bound platelets, leukocytes, and erythrocytes, as well as blood proteins such as fibrinogen. This work uncovered the biophysical requirements for initiating microvascular thrombosis and suggested mechanisms for the onset and progression of microvascular diseases. In the second example, engineered microvessels were treated with platelet-rich plasma, plasma, and platelet lysate to understand the influence of platelets on the vessel wall integrity and endothelial cell maturation. Platelets were observed to bind to the vessel wall defects, where sub-endothelial matrix were exposed. Endothelial cells were stimulated initially, changed their polarity, and interacted with platelets. After 24 hours, platelets were found underneath endothelium integrate with the sub-endothelial matrices. Some platelets formed junctions or even fused with endothelial cells. The vessels exhibited larger diameter and better barrier functions with lower permeability of fluorscence molecules. The comparison studies have been made for different platelet-relevant substances to reveal the mechanisms of platelets repairing vessel barrier functions. In the last example, a modified microvascular system was generated to describe a marrow microvascular niche. A thrombopoietic microvascular niche by embedding megakaryocytes (Megs), platelet progenitors, in the matrix around microvessels. Megs migrated and differentiated towards the microvessel with time, penetrated the endothelial cells, and released the platelets into the circulation. When marrow specific stromal cells were embedded in the matrix, they also interacted with microvessels through physical contact as well as chemical modification. They changed marrow microenvironment, consequently led to selective homing of hematopoietic cells including monocytes and HSCs from mcirovessels towards the matrices. Our work demonstrates that complex microvascular environment can be recapitulated in vitro to depict the complex vascular events in vivo, and suggests the broad potential of our system to study vascular biology and pathophysiology in both a systemic and organ-specific context.