Investigation of the molecular mechanisms of vascular endothelial dysfunction in Hutchinson-Gilford progeria syndrome through in vitro 2D and 3D models

Hutchinson-Gilford progeria syndrome (HGPS) is a rare genetic disorder with features of accelerated aging. Predominantly, HGPS is caused by a de novo point mutation in the LMNA gene (c.1824C > T; p.G608G) resulting in progerin, a toxic lamin A protein variant. Children with HGPS typically die from coronary artery diseases or strokes at an average age of 14.6 years. Endothelial dysfunction is a known driver of cardiovascular pathogenesis; however, it is currently unknown how progerin antagonizes endothelial function in HGPS. In this study, I used human iPSC-derived endothelial cell (iPSC-EC) models that cultured under both static and fluidic culture conditions. HGPS iPSC-ECs show reduced endothelial nitric oxide synthase (eNOS) expression and activity compared to healthy controls and concomitant decreases in intracellular nitric oxide (NO) level, which result in deficits in capillary-like microvascular network formation. In addition, expression of matrix metalloproteinase 9 (MMP-9) was reduced in HGPS iPSC-ECs while expression of tissue inhibitor metalloproteinases 1 and 2 (TIMP1 and TIMP2) were upregulated relative to healthy controls. Moreover, I used an adenine base editor (ABE7.10max-VRQR) to correct the pathogenic c.1824C > T allele in HGPS iPSC-ECs. Remarkably, ABE7.10max-VRQR correction of the HGPS mutation significantly reduced progerin expression to a basal level, rescued nuclear blebbing, increased intracellular NO level, normalized TIMPs , and restored angiogenic competence in HGPS iPSC-ECs. Furthermore, to elucidate the effects of progerin on endothelial cells and vascular remodeling, in collaboration with Dr. Truskey’s lab at Duke university, we developed tissue-engineered blood vessels (TEBVs) using iPSC-ECs and smooth muscle cells (iPSC-SMCs) from normal and HGPS patients. Relative to normal TEBVs, HGPS TEBVs showed reduced function and exhibited markers of cardiovascular disease associated with endothelium, including a reduction in both vasoconstriction and vasodilation with increased inflammation markers, VCAM-1 and E-selectin protein. Hence, the TEBV model has identified a role of the endothelium in HGPS. Together, the results of the study provide molecular insights of endothelial dysfunction in HGPS and suggest that ABE7.10max-VRQR could be a promising therapeutic approach for correcting HGPS-related cardiovascular phenotypes