CCM2L (cerebral Cavernous Malformation 2 Like) Deletion Aggravates Cerebral Cavernous Malformation Through Map3k3-KLF Signaling Pathway

Background and Purpose: Cerebral cavernous malformation (CCM) is a common cerebrovascular disease. CCMs are major causes of stroke, cerebral hemorrhage, and neurological deficits in young individuals. Loss-of-function mutations in CCM1, CCM2, and CCM3 have been identified to cause CCM in humans. Ccm2-like (Ccm2l) is a paralog of Ccm2 and is predominantly expressed in endothelial cells (ECs). CCM2L (CCM2-like) competes with CCM2 for binding to CCM1 and has been shown to have an antagonistic function to that of CCM2 during vascular development. The role of CCM2L in CCM pathogenesis is unknown. Methods: We isolated brain ECs from the inducible-CCM mouse models for gene expression analysis. Micro-computed tomography imaging was used to analyze CCM lesion burden from the genetic cross of Ccm2l knockout mice (Ccm2l(-/-)) with Ccm1 or Ccm2-deficient mice to determine the role of Ccm2l in CCM pathogenesis. Genetic crosses with Map3k3(fl/fl) mice were used to determine the role of Map3k3 in Ccm2l-facilitated CCM formation. Results: We demonstrated increased Ccm2l expression in brain ECs of Ccm2-deficient mice. Analysis of RNA-seq data from CCM patient samples revealed a trend of increased CCM2L expression and its positive correlation with Kruppel-like factor 2/4 (KLF2/4) expression. Micro-computed tomography revealed that the deletion of Ccm2l in Ccm2-deficient mice increased CCM lesion volume compared with that of controls but had no effect on lesion numbers. Correlating to the increased lesion burden, Klf2/4 mRNA expressions in brain ECs were significantly increased in double knockouts (Ccm2- and Ccm2l-deficient mice) compared with that of controls (Ccm2 deficient). Hemizygous deletion of Map3k3 in ECs relieved CCM lesion burden in the double knockouts. These results suggest that CCM2L regulates the Map3k3-KLF signaling pathway in CCM pathogenesis. Conclusions: Loss of CCM2L aggravates CCM lesion formation in the Ccm2-deficient mouse model through increased Map3k3-KLF signaling. Our data suggest that increased Ccm2l expression is a compensatory mechanism in CCM pathogenesis.