gms | German Medical Science

Objective: The Moyamoya disease (MMD), a rare cerebrovascular disorder, is characterized by an aberrant network of collateral circulation vessels leading to hemorrhagic and ischemic strokes. MMD progression has been attributed to some growth and angiogenic factors influencing circulating endothelial progenitors, smooth muscle and endothelial cells (ECs). However the molecular background of fragile vessel development in MMD still needs to be elucidated. Here we propose a well described cell culture model of the blood-brain barrier, a murine microvascular brain EC line to examine the properties of ECs incubated with MMD sera.

Method: We compared the endothelial sealing of cultures incubated with serum from healthy controls, patients with arteriosclerosis and MMD using molecular and electrophysiological tools. First we tested the transendothelial electrical resistance besides protein and mRNA status of EC barrier maintaining components claudin-3 and -5, occludin, ZO-1 and VE-cadherin in brain ECs. We measured intra- and extra-cellular levels of angiopoietin-2 (Ang-2), a barrier destabilizing angiogenic factor and its antagonist Ang-1 in culture supernatants. Than we assessed the intracellular protein levels of VEGF-A and MMP-9, both so far described as serum markers in MMD. Finally the MMP-9 activity in sera samples was shown by gelatinase zymography assays.

Results: Incubation with MMD patient sera resulted in a firm increase of endothelial monolayer permeability compared to controls. The protein and transcript status of claudin-3 and -5, occludin, ZO-1 and VE-cadherin was clearly decreased upon MMD sera. We also observed an induction of intra- and extracellular levels of Ang-2 following reduction of Ang-1 in MMD sera treated brain ECs. Further the distinct up-regulation of VEGF-A and MMP-9 was shown in cultures incubated with MMD sera compared to the controls and was accompanied with a higher MMP-9 activity in MMD sera themselves.

Conclusions: The pathological processes and the role of the endothelium on a molecular level in MMD still need to be clarified in detail. Hence we introduce a brain EC line as an in vitro tool providing a new insight into molecular mechanisms affecting the endothelial barrier sealing in fragile MMD vessels. We propose the enhancement of endothelial plasticity induced by an imbalance of angiogenic factors and of proteins of the junctional complex may play a key role in the development of occlusive lesions and edema in MMD.