In Vitro Characterization of the Angiogenic Phenotype and Genotype of the Endothelia Derived From Sporadic Cerebral Cavernous Malformations

Abstract

Cerebral cavernous malformation (CCM) is mainly a disorder of endothelial cells. Although the endothelial function of CCM genes has been characterized in familial CCMs, little attention has been paid to the pathological alterations of the endothelium in sporadic CCMs. We assumed that the endothelia derived from sporadic CCMs present genotypic and/or phenotypic alterations and exhibit unique responses to the pathogenic stimuli. Endothelial cells were prepared from fresh operative specimens of sporadic CCMs with a single lesion (CCM-ECs, n = 20). The expression of VEGF and its receptors and CCM1-3 genes were detected by real-time reverse transcriptase polymerase chain reaction (RT-PCR). Endothelial cell proliferation, migration, sprouting, and tube formation were compared between CCM-ECs and control endothelial cells after different angiogenic stimuli and after silencing CCM1. RT-PCR revealed a highly activated VEGF system in CCM-ECs without significant alteration in CCM1-3 gene expression. Accordingly, CCM-ECs exhibited great growth potential under normal culture conditions and a significantly high proliferation activity in response to various angiogenic stimuli including hypoxia, fetal calf serum, and vascular endothelial growth factor treatment. A considerably higher mobility, spontaneous sprouting and extensive tube-branching were exclusively detected in CCM-ECs. In comparison with control endothelia, CCM-EC resisted apoptotic stimuli and showed distinct responses to activating angiogenesis after silencing CCM1. Distinct genotypic and phenotypic features occur in CCM-EC independently from the deficiency in CCM1-3 gene expression. The distinct responses of CCM-EC to different pathogenic stimuli suggest that CCM-EC is a valuable in vitro model for further study of CCMs.