MIcrophysiological systems to model vascular malformations

PROJECT SUMMARYCOVID19 is caused by SARS-CoV-2, a novel member of the human coronavirus family that includes the closelyrelated SARS-CoV (SARS) and MERS-CoV (MERS) viruses. SARS-CoV-2 viral spike protein binds to humanangiotensin converting enzyme-2 (ACE2) on the surface of cells and is then primed by the serine/threonineprotease TMPRS22, whereupon the entire complex is internalized by the target cell. ACE2 is expressed bymultiple cell types of the body, including lung and gut epithelium (likely the primary sites of initial infection) andvascular endothelial cells (EC) of multiple organs. In addition to the well-described pneumonia-like diseasecharacterized by compromised lung function with subsequent depressed pO2 levels in blood, patients often alsoshow signs of multi-organ involvement, which can include gut, kidney, liver, heart, and brain. Most recently,numerous pediatric patients have been showing signs of Kawasaki disease, a systemic vascular inflammation.Over 30% of COVID19 ICU patients also show signs of thrombosis and 25% suffer venous embolism. Cerebralischemia, likely due to clot formation, has also been reported. What is not clear is whether this multi-organinvolvement is due to secondary infection of these tissues or whether these are all a consequence of systemichyperinflammation. The sequence of events that could drive systemic hyperinflammation stems from the SARS-CoV-2 mechanism of infection. Angiotensin II (AngII) is an important vasoconstrictor and under normalphysiological conditions its level is closely controlled through rapid degradation by ACE2, however, SARS-CoV-2 entry into cells clears ACE2 from the cell surface, potentially prolonging the action of AngII. Primaryconsequences of this would be two-fold: prolonged vasoconstriction in the lung (exacerbating poor oxygenationof the blood by the already compromised lungs) and a shift toward a pro-inflammatory state, as it is wellestablished that AngII can drive local vascular inflammation, in large part through the induction of IL-6 in EC andsmooth muscle cells (SMC). IL-6 is one of the major drivers of systemic hyperinflammation, and in its mostsevere form, a so-called "cytokine storm". IL-6 is also strongly correlated with thrombosis, likely throughupregulation of tissue factor on EC and macrophages, and by downregulation of thrombomodulin on EC.Through the parent award we have generated Vascularized Micro-Organs (VMOs), comprised of perfusedhuman vasculature and a surrounding stroma, and have further developed these into Vascularized Micro-Brains(VMBs) incorporating a Blood-Brain Barrier, and Vascularized Micro-Livers (VMLs). Additional cells, includingmacrophages and SMC have also been incorporated into the VMO. Using these platforms we will address threehypotheses: 1) That the VMO can be used to assess the ability of convalescent serum, soluble ACE2 or smallmolecule inhibitors to block entry of a SARS-CoV-2 pseudotyped virus; 2) That sustained AngII expression cantrigger a hyperinflammatory response from EC and macrophages; and 3) That hyperinflammation cancompromise peripheral and BBB vasculature as well as liver function.