Brain microvascular endothelial cells differentiated from a Friedreich's Ataxia patient iPSC are deficient in tight junction protein expression and paracellularly permeable

Kosman J. Daniel ^* Frances M. Smith

• University at Buffalo, Buffalo, United States

Friedreich's Ataxia (FA) is a rare, inherited ataxia resulting from GAA triplet expansions in the first intron of the Frataxin (FXN) gene, which encodes for a mitochondrial protein involved in the incorporation of iron into iron-sulfur clusters. We previously identified decreased abundance of F-actin and tight junction (TJ) proteins co-incident with paracellular permeability in an shRNA-mediated knockdown of FXN immortalized human brain microvascular endothelial cell (BMVEC) model system. Such a premise is lacking in FA literature and therefore we sought to confirm these findings using a patient-derived iPSC. One line each of FA patient iPSCs and an age-and sex-matched apparently healthy iPSCs were differentiated to BMVEC-like cells. We quantified actin glutathionylation, F-actin abundance, TJ expression and organization, and barrier integrity. In the absence of dysregulated F-actin organization, FA iBMVEC lost 50% of ZO-1, 63% Occludin, and 19% of Claudin-5 protein expression with disruption of the bi-cellular organization of the latter two proteins. Functionally, this correlated to barrier hyperpermeability, lack of barrier maturation, and increased flux of the fluorescent tracer Lucifer Yellow. These data show that decreased barrier integrity is a pathophysiologic phenotype of FA brain microvascular endothelial cells. Clinically, this may be a targetable pathway to abrogate brain iron accumulation, neuroinflammation, and neurodegeneration profiles of FA. Additionally, investigation into other barrier systems, such as the blood-nerve barrier, blood-CSF barrier, or cardiac vasculature may inform on extra-neural symptoms experienced by the FA patient.