In vitro modeling of blood brain barrier dysfunction on a chip to elucidate the pathogenesis of cerebral malaria

Project Summary Brain microvascular endothelial cells (BMECs) which line the vascular network of the central nervous system (CNS) in conjunction with perivascular cells form a specialized barrier termed the blood-brain barrier (BBB) that regulates the dynamic traffic of select molecules into and out of the CNS. Studies of BBB dysfunction have been hampered by an inability to perform direct testing in patients and a lack of in vitro models. Analysis of available putative BMECs from pluripotent stem cell sources reveals that they do not harbor bona fide brain EC molecular signatures. Using a transcription factor-based reprogramming strategy, we demonstrate that durable and functional true endothelial cells with BBB traits may be derived. We will use these verified BMECs in the development of isogenic BBB/on-a-chip devices. Cortical brain organoids derived from patient specific induced pluripotent cells (IPSC) will be vascularized by matched IPSC derived bona fide BMECs in a microfluidic device. To explore the potential of this new technology we will investigate the pathobiology of cerebral malaria a severe disease syndrome that causes neurodisability in 20% of survivors. Red blood cells infected with the Plasmodium falciparum parasite adhere to the brain microvasculature causing dysfunction. We hypothesize that the parasite primes the brain endothelial cells for adherence through their “education” by small extracellular vesicles termed exosomes. This is an established paradigm in cancer biology in which malignant cells educate metastatic “microniches” to prime for and enhance tumor survival. The exosomes mediate local and systemic intercellular communication through the horizontal transfer of information in their proteomes. We recently were able to identify a critical prognosis biomarker for brain metastasis in breast cancer through studies of exosomal cellular uptake and proteomic analysis. We will analyze the proteomes of exosomes from children with cerebral malaria and infuse them into our innovative microfluidic-based BBB model. We will monitor in real time how exosomes trigger BBB dysfunction and subsequently affect IPS-derived brain organoids from patients affected by CM. These studies will provide new insights into disease pathogenesis and potentially identify new prognosis biomarkers.