Untangling the Pathophysiology of Cerebral Cavernous Malformations

Neuroscience and Neurobiology; Pathology; Molecular and Cellular Neuroscience; Cellular and Molecular Physiology; Bioimaging and Biomedical Optics

We will study calcium signaling in endothelial cells derived from cerebral cavernous malformations under various shear stress conditions to instigate the development of new therapeutic strategies.

Research Interests
  • Neuroimaging Data Analysis
  • Calcium signaling mechanisms
  • Genetic approaches
  • Neurodegenerative Disease
  • Vascular malformations
  • Multi-modal brain MRI to study different aspects of neurodegenerative diseases
  • Focused Ultrasound
  • Cerebrovascular disease
  • Mouse models of disease
  • 2photon

Cerebral cavernous malformations (CCM) are vascular abnormalities of the central nervous system with a prevalence of 0.4–0.5% in the general population. They are caused by mutations in one of the 3 genes, KRIT1, CCM2 or PDCD10. They initially have the appearance of a small mulberry, but gradually develop into malformations that vary in size from 2 millimeters to several centimeters in diameter. CCMs may leak blood, leading to brain or spinal hemorrhages that can generate a wide range neurological symptoms, including severe headaches and seizures. Women and men are equally affected, and patients typically present during the 2nd to 5th decades of life, although lesions have been also reported in infants and children. The risk of CCM hemorrhage varies between 0.25 to 4.5% per patient per year, necessitating in long-term follow-up at a significant cost to the health care system. The primary treatment option for symptomatic CCM is surgical removal, which often presents high risk to the patient. Better understanding of the CCM pathophysiology and new therapeutic approaches are urgently needed.

Murine models have been developed for all three loci. These mice faithfully recapitulate the hallmarks of the human disease. We have established chronic CCM models of Krit1 and Ccm2 etiology, by postnatally deleting the causative genes in the endothelial cells using a tamoxifen inducible Cre/lox system. Our CCM mice have a lifespan of 5 months or more, while developing a rich repertoire of CCM lesions. In this model, the lesions are first detectable with T2-SPACE or SWI MRI sequence at one month of age, and the lesion load progressively increases over the next 3-4 months. Having the chronic animal model established is the first necessary step for subsequent in-depth investigation of the pathology and experimental therapies.

Desired outcomes

This trio of Cavalier Investigators is well positioned to address an outstanding question concerning intracellular calcium activity in Krit1 mutant endothelial cells due to blood pressure and shear stress. Although the CCM mutations occur in endothelia of all blood vessels in the brain, cavernous malformations only develop in low flow veins.  The physiologic factor of laminar shear stress has been implicated in mitigating the onset of CCM pathology, but mechanistic understanding is lacking.  Nothing is presently known about the patterns of calcium signals in CCM mutant endothelial cells, let alone how these signals change after increasing flow and shear stress. The knowledge of these intracellular responses may provide clues for therapeutic interventions. For example, altered calcium signaling patterns may affect cellular motility of CCM endothelia, which are known to undergo endothelial-to-mesenchymal transition, contributing to CCM lesion heterogeneity and slow, benign expansion. To address these questions, the Tvrdik lab has established primary cell cultures of brain microvascular endothelial cells (BMEC) from Krit1 conditional (floxed) mice harboring the tamoxifen-inducible Pdgfb-CreER allele. These primary cell cultures represent normal endothelial cells but can be easily converted to Krit1 mutant cells by adding hydroxytamoxifen to the culture media. This flexible and precisely controlled in vitro system will be exposed to varying laminar shear stress conditions as directed by the Helmke laboratory who has decades-long experience with applying and evaluating laminar shear stress to endothelial cells. The Sonkusare lab will complete the experimental setup by providing calcium imaging technology and calcium signal analysis in the cell culture settings. We anticipate that this project will generate pioneering insight into intracellular activity in the mutant CCM cells under various shear stress conditions. The research environment in this group will be highly conducive to training of students and research scientists interested in vascular physiology and pathophysiology. It will also generate preliminary data for grant applications to NIH/NINDS.