Background:
Moyamoya disease (MMD) is a rare, progressive steno-occlusive cerebrovascular disorder of the internal carotid artery, leading to stroke. Affected arteries exhibit thickened intima with depleted elastic lamina and media, indicating a dysfunction of the vascular smooth muscle cells (VSMCs) and endothelial cells (ECs). However the pathogenesis of the disease is still unclear. We aim to address this gap in knowledge by using patient derived induced pluripotent stem cells (iPSCs), to generate VSMCs and ECs.
Methods:
Peripheral blood mononuclear cells (PBMCs) from controls and MMD patients (n=3 per group) were used for generating iPSCs. VSMC functionality was measured by collagen gel contraction assay and scratch assay. EC proliferative function was assessed by BrDU incorporation assay, and its migration capacity was evaluated by scratch assay and in vitro tube formation. VSMCs and ECs were also exposed to either hydrogen peroxide (H2O2) or normoxia/ hypoxia model (1%O
2
) to investigate how cells respond to these insults. Hypoxia inducible factor 1α (HIF1α) activation was determined using western blot.
Results:
MMD VSMCs trended towards being more contractile and migrating faster than control VSMCs, in response to 10%FBS or SDF1α. On the other hand, MMD ECs migrated slower than control ECs in response to 10%FBS (p=0.0081) or VEGF (p=0.0072). MMD ECs also formed lesser tubes and exhibited fewer branch points when compared to controls. The rate of EC proliferation was similar between both groups. Cell death assays indicate that MMD VSMCs and ECs were more sensitive to the deleterious effects of H2O2 exposure when compared to control cells. Interestingly, MMD VSMCs had elevated HIF1α protein expression in normoxia, which was further increased after hypoxia.
Conclusions:
Our preliminary results indicate that both MMD VSMCs and ECs are dysfunctional and may be related to the elevated basal expression of HIF1α, possibly contributing to MMD pathology. We are currently investigating the interactions between VSMCs and ECs in MMD compared with controls using co-cultures. Ongoing studies also include transcriptome analysis of these differentiated cells, which will advance the understanding of the cellular and molecular mechanisms underlying MMD.
The Application of Induced Pluripotent Stem Cells Against Liver Diseases: An Update and a Review
