VALIDATION AND CHARACTERIZATION OF AN IN VITRO MODEL FOR VASCULAR SMOOTH MUSCLE CELLS AND ENDOTHELIAL CELLS WITH MARFAN SYNDROME

Marfan Syndrome (MFS) is a heritable connective tissue disorder caused by a mutation to the fibrillin-1 encoding FBN1 gene that causes abnormalities in the skeletal, ocular, and cardiovascular system. The most clinically lethal symptom is the dilation, aneurysm, and rupture of the aorta. Current treatment involves the administration of β blockers in order to reduce the shear stress on the aortic wall and slow the vessel’s rate of dilation. Once the vessel is dilated beyond the point of safety, surgical intervention must be performed to prevent aneurysm and rupture and even this does not preclude the need for further prophylactic surgeries. A greater understanding of the disease is needed to develop treatments which prevent the underlying pathology of the aortic wall. Mouse models have provided much needed insight into the mechanistic underpinnings of MFS, implicating transforming growth factor beta (TGF-β) signaling in disease progression. However, human studies have not shown the same beneficial effects of TGF-β antagonists as are observed in the mouse model, highlighting the need for a humanized model of MFS. This study aims to validate and characterize such a model by using human induced pluripotent stem cells (iPSCs) to derive vascular smooth muscle cells (vSMCs) and endothelial cells (ECs) with the MFS phenotype. Human iPSCs with MFS were differentiated into vSMCs and ECs. Immunofluorescence (IF) staining and confocal imaging were used to confirm the presence of vSMC protein markers and EC markers and validate the success of each differentiation. The same techniques were used to confirm the MFS phenotype by quantification of fibrillin, as a decrease in extracellular fibrillin is characteristic of vascular cells with the MFS phenotype. A live/dead analysis was performed on the vSMCs which confirmed the increased apoptotic activity of the MFS phenotype. Finally, an enzyme-linked immunosorbent assay (ELISA) was used to quantify the heightened presence of TGF-β in the vSMCs. further confirming the MFS phenotype. Observations made in this study indicate that this is a valid model for the progression of MFS in vascular cells and future studies may use this as a basis for investigating the role of TGF-β signaling in MFS