Atypical hemolytic uremic syndrome (aHUS) is a life-threatening disease that causes microvascular thrombosis, especially in the kidneys, with a global mortality rate of 25%, and a frequent progression to end-stage renal disease (ESRD)1,2. While aHUS is known to be associated with uncontrolled activation of the alternative pathway (AP) of complement, the underlying pathophysiology of how AP activation causes endothelial injury and thrombosis remains unclear, which largely limits the development of additional therapies for aHUS. Even with the currently most effective drug, eculizumab, many still suffer from acute flare ups, progression to ESRD, and extra-renal manifestations, which are thought to be associated with unregulated endothelial activation. The proposed work aims to develop the first in vitro model of aHUS that incorporates all of the major components thereof, including complement and other plasma proteins, platelets, red blood cells, intact endothelium, extracellular matrix with physiologic biophysical properties, and shear stress, to not only improve our understanding of the pathophysiology of this feared disease but also to explore new therapeutics that may improve better clinical outcomes for aHUS patients. Here we hypothesize that a novel hydrogel-based ?endothelialized? microvasculature-on-a-chip that we recently invented recapitulates the in vivo microvascular microenvironment and exhibits long-term (>2 months) microvasculature functionality. This can be used as an in vitro model for aHUS, as our microvasculature-on-a-chip system enables the decoupling of the complex cellular and molecular factors (including complement and regulators of complement activation) involved in the pathophysiology of aHUS as well as quantitatively characterizes how these factors interact and lead to endothelial injury or activation. These factors will be perfused into the engineered microvasculature, and biomarkers of endothelial dysfunction, such as endothelial permeability, reduced nitric oxide and increased reactive oxygen species, will be monitored in real time. In addition, the system will allow assessment of how these factors synergistically lead to thrombi formation in the engineered microvasculature in the context of aHUS through quantifying fibrin formation and platelet aggregate/microthrombi size. As eculizumab only blocks the formation of the membrane attack complex in the terminal pathway of complement activation, this microvasculature-on-a-chip model will enable the testing of other novel treatment strategies to promote endothelial repair. Successful completion of this study will result in the development of a novel and robust model for aHUS, gain a better understanding of endothelial damage in aHUS and more broadly how the complement system interacts with hemostasis and thrombosis, and provide insights into developing rational pharmacological approaches in the management of aHUS and other thrombotic microangiopathies.

Public Health Relevance

Atypical hemolytic uremic syndrome (aHUS) is a chronic and rare genetic disease that affects complement system proteins that are part of our immune system. Patients with aHUS suffer from anemia, low platelet count and kidney injury, though it can affect many other parts of the body. Patients if untreated develop chronic kidney disease, severe anemia and death. This research is intending to seek answers on how aHUS affects the lining of blood vessels and causes microscopic blood clots, using novel tools such as engineered blood vessels (hydrogel-based microvasculature-on-a-chip). This proposal has great potential to not only advance our understanding of aHUS but also aid in development of newer treatment strategies.

National Institute of Health (NIH)
National Institute of Biomedical Imaging and Bioengineering (NIBIB)
Exploratory/Developmental Grants (R21)
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Hemostasis and Thrombosis Study Section (HT)
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Rampulla, David
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Emory University
Schools of Medicine
United States
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