Cerebral cavernous malformations (CCMs) are common inherited and sporadic vascular malformations that cause stroke and seizures in younger individuals1. CCMs arise from endothelial cell loss of KRIT1, CCM2, or PDCD10, non-homologous proteins that form an adaptor complex2. How disruption of the CCM complex results in disease remains controversial, with numerous signaling pathways (including Rho3,4, SMAD5 and Wnt/β-catenin6) and processes such as endothelial-mesenchymal transition (EndMT)5 proposed to play causal roles. CCM2 binds MEKK37–11, and we have recently demonstrated that CCM complex regulation of MEKK3 is essential during vertebrate heart development12. Here, we investigate this mechanism in CCM disease pathogenesis. Using a neonatal mouse model of CCM disease, we find that expression of the MEKK3 target genes KLF2 and KLF4, as well as Rho and ADAMTS protease activity, are increased in the endothelial cells of early CCM lesions. In contrast, we find no evidence of EndMT or increased SMAD or Wnt signaling during early CCM formation. Endothelial-specific loss of Mekk3, Klf2, or Klf4 dramatically prevents lesion formation, reverses the increase in Rho activity, and rescues lethality. Consistent with these findings in mice, we demonstrate that endothelial expression of KLF2 and KLF4 is elevated in human familial and sporadic CCM lesions, and that a disease-causing human CCM2 mutation abrogates MEKK3 interaction without affecting CCM complex formation. These studies identify gain of MEKK3 signaling and KLF2/4 function as causal mechanisms for CCM pathogenesis that may be targeted to develop new CCM therapeutics.

Summary Hemodynamic forces play an essential epigenetic role in heart valve development, but how they do so is not known. Here we show that the shear-responsive transcription factor KLF2 is required in endocardial cells to regulate the mesenchymal cell responses that remodel cardiac cushions to mature valves. Endocardial Klf2 deficiency results in defective valve formation associated with loss of Wnt9b expression and reduced canonical WNT signaling in neighboring mesenchymal cells, a phenotype reproduced by endocardial-specific loss of Wnt9b. Studies in zebrafish embryos reveal that wnt9b expression is similarly restricted to the endocardial cells overlying the developing heart valves and dependent upon both hemodynamic shear forces and klf2a expression. These studies identify KLF2-WNT9B signaling as a conserved molecular mechanism by which fluid forces sensed by endothelial cells direct the complex cellular process of heart valve development, and suggest that congenital valve defects may arise due to subtle defects in this mechanotransduction pathway.

Conflict of interest: JDW and MLK are coinventors on a patent, PCTUS1932399, entitled "Medical device for the prevention of thrombosis" that is based on the scientific discoveries described in this article. They are stakeholders in a company, Osciflex, that has an option agreement with the University of Pennsylvania to develop a new device to prevent deep venous thrombosis.

In this Letter we omitted to cite a relevant paper 1 showing that loss of cerebral cavernous malformation (CCM) signalling confers an increase in KLF2 expression in endothelial cells and in the developing zebrafish heart. We regret this oversight. 1. Renz, M. et al. Regulation of β 1 integrin-Klf2-mediated angiogenesis by CCM proteins. Dev. Cell 32, 181-190 (2015).

Vestibular schwannoma is a known cause of progressive sensorineural hearing loss. Treatment options include observation, radiation therapy and surgical resection. Cerebrospinal fluid (CSF) fistula is a known postsurgical complication that can lead to CSF otorrhoea, rhinorrhoea or CSF leakage from the surgical wound. We present a case report of a patient who underwent vestibular schwannoma resection and postoperatively developed CSF rhinorrhoea, which was refractory to multiple attempts at surgical repair. This was successfully treated under endoscopic and fluoroscopic guidance using a biliary cytology brush to disrupt the surface of the eustachian tube followed by injection of n-Butyl cyanoacrylate.