(Provided by the applicant) Abstract: In this proposal, we describe a plan to build artificial platelets from biological components. The vision is based on the belief that the critical accumulation of our knowledge about individual biomolecules will enable us to integrate them into a system in a meaningful way to create cellular devices. We have identified platelets as a tractable target of such emulated biological system. Our design strategy necessitates an understanding of the functionalities of natural platelets so that the artificial platelets can confer the essential functions of natural platelets. Platelets are the first cellular responder to injured vasculature. It binds to the exposd collagen that is otherwise protected by endothelial cells. Platelets couple binding to activation via the expression of several cell surface receptors. Importantly, activated platelets attain predominantly negatively charged membranes by increasing phosphatidylserine (PS) on the outer lipid leaflet. Activated platelets then trigger a series of biochemical events known as the coagulation cascade that results in the formation of cross-linked fibrin network that forms a plug to block the injured vasculature. The artificial platelets will mimic the essential functions of natural platelets. They will be made as lipid vesicles that have defined lipid and protein composition. Microfluidic jetting, a technique recently developed that is akin to blowing bubble from a soap film, will be used to make the vesicles. This technique allows the ability to create asymmetric lipid bilayer with PS on the inner leaflet as well as incorporation of membrane proteins. The vesicles will be decorated with anti-collagen antibody that will bind to the exposed collagen during vascular injury. Our platelet activation strategy will utilize the large mechanosensitive channel (MscL) that is gated by membrane tension. When the artificial platelets attaches to the substrate, shear flow will activate MscL, this would then causes calcium in the blood serum to enter the vesicles. Scramblase will be part of the artificial platelet such that upon calcium entry, it becomes activated and exposes PS to the outer lipid leaflet. When PS is exposed, factors V and X become activated and subsequently stimulate thrombin activity. Thrombin then cleaves fibrinogen into fibrin, which assembles into a durable and cross-linked network. At each stage of our artificial platelet assembly, the design strategy can be tested and validated in a microfluidic platform that mimics the injured vasculature. In summary, the proposed research describes our vision for how synthetic biology can potentially bring about novel cellular devices that would have tremendous benefits in medical settings. Public Health Relevance: Platelets are the first cellular responder to injured vasculature, but it is constantly under shortage in the blood bank due to its short shelf life, susceptibility to contamination, and challenges in storage condition. Using biological components, we propose to build artificial platelets that mimic the functionalities of natural platelets. We envision such cellular devices will bring tremendous benefits in medical settings.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
NIH Director’s New Innovator Awards (DP2)
Project #
1DP2HL117748-01
Application #
8335509
Study Section
Special Emphasis Panel (ZGM1-NDIA-C (01))
Program Officer
Link, Rebecca P
Project Start
2012-09-30
Project End
2017-08-31
Budget Start
2012-09-30
Budget End
2017-08-31
Support Year
1
Fiscal Year
2012
Total Cost
$2,332,500
Indirect Cost
$832,500
Name
University of Michigan Ann Arbor
Department
Engineering (All Types)
Type
Schools of Engineering
DUNS #
073133571
City
Ann Arbor
State
MI
Country
United States
Zip Code
48109
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Coyne, Christopher W; Patel, Karan; Heureaux, Johanna et al. (2014) Lipid bilayer vesicle generation using microfluidic jetting. J Vis Exp :e51510