Background:
Moyamoya disease (MMD) is a rare, progressive steno-occlusive cerebrovascular disorder of the internal carotid artery, leading to stroke. Affected arteries exhibited 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. Functional properties of differentiated ECs and VSMCs in normoxia/hypoxia model (1%O
2
) were assessed for cell proliferation by BrDU incorporation, migration by scratch assays and apoptosis by exposure to hydrogen peroxide (H2O2).
In vitro
angiogenic tube formation was assessed with ECs alone, as well as ECs and VSMCs as a co-culture. Hypoxia inducible factor 1α (HIF1α) and Intercellular adhesion molecule-1 (ICAM-1) activation was determined using qPCR and western blot in VSMCs.
Results:
Functional proliferative assays showed that MMD ECs proliferated faster than control ECs. Migration assays showed that MMD ECs migrate slower in response to VEGF after hypoxia. MMD ECs were found to be more sensitive to insults such as H2O2 treatment and exhibited more apoptosis. In contrast, MMD VSMCs proliferate and migrate similar to controls, but exhibited elevated levels of HIF1α, ICAM-1 mRNA and protein expression. MMD and control ECs showed similar levels of tube formation in single cultures, however, when co-culturing with VSMCs, MMD VSMCs failed to support EC tubes beyond 24 h, resulting in tube destabilization.
Conclusions:
Our preliminary results indicate that both MMD VSMCs and ECs are dysfunctional and may be related to the elevated expression of HIF1α and ICAM-1, possibly contributing to MMD pathology. Current ongoing studies include investigating the interactions between MMD VSMCs and ECs using co-cultures, as well as transcriptome analysis of these differentiated cells, which will provide important insights into the cellular and molecular mechanisms underlying MMD.
Functional vascular smooth muscle cells derived from human induced pluripotent stem cells via mesenchymal stem cell intermediates
