Inflammation and neutrophil extracellular traps in cerebral cavernous malformation

Abstract

Cerebral Cavernous Malformation (CCM) is a brain vascular disease with various neurological symptoms. In this study, we describe the inflammatory profile in CCM and show for the first time the formation of neutrophil extracellular traps (NETs) in rodents and humans with CCM. Through RNA-seq analysis of cerebellum endothelial cells from wild-type mice and mice with an endothelial cell-specific ablation of the Ccm3 gene (Ccm3iECKO), we show that endothelial cells from Ccm3iECKO mice have an increased expression of inflammation-related genes. These genes encode proinflammatory cytokines and chemokines, as well as adhesion molecules, which promote recruitment of inflammatory and immune cells. Similarly, immunoassays showed elevated levels of these cytokines and chemokines in the cerebellum of the Ccm3iECKO mice. Consistently, both flow cytometry and immunofluorescence analysis showed infiltration of different subsets of leukocytes into the CCM lesions. Neutrophils, which are known to fight against infection through different strategies, including the formation of NETs, represented the leukocyte subset within the most pronounced increase in CCM. Here, we detected elevated levels of NETs in the blood and the deposition of NETs in the cerebral cavernomas of Ccm3iECKO mice. Degradation of NETs by DNase I treatment improved the vascular barrier. The deposition of NETs in the cavernomas of patients with CCM confirms the clinical relevance of NETs in CCM.

Contact-dependent signaling triggers tumor-like proliferation of CCM3 knockout endothelial cells in co-culture with wild-type cells

“…The results of our in vitro model are also consistent with recently published analyses in Ccm3 iECKO mice. Using an acute, “fast progression” and a chronic, “slow progression” model of CCM, the authors demonstrated that the expression of Ccl2 , Cxcl2 , Cx3cl1 and other immune-related genes was significantly upregulated in brain microvascular ECs of Ccm3 iECKO mice [ 48 ]. In general, the proportion of cells with a Ccm3 deletion is relatively low (< 15%) in the chronic model [ 48 ].…”

“…Using an acute, “fast progression” and a chronic, “slow progression” model of CCM, the authors demonstrated that the expression of Ccl2 , Cxcl2 , Cx3cl1 and other immune-related genes was significantly upregulated in brain microvascular ECs of Ccm3 iECKO mice [ 48 ]. In general, the proportion of cells with a Ccm3 deletion is relatively low (< 15%) in the chronic model [ 48 ]. In the acute model, it is much higher (> 80%) but cells with Ccm3 wild-type alleles are still present [ 17 ].…”

Neuroinflammation plays a critical role in cerebral cavernous malformation disease

“…Indeed, an increase in astrocyte CX3CL1 signaling may play an important role in recruiting both microglia and peripheral inflammatory cells into CCM lesions via the receptor CX3CR1 60 . Moreover, the increase of astrocyte MCP1 ( Ccl2 ), a chemoattractant for monocytes and activated T cells, may synergize with CX3CL1 to recruit the innate and adaptive immune response 58, 61 as well as to induce disassembly of tight junctions in brain endothelial cells during CCM disease 8, 62 . We observed that neuroinflammatory astrocytes induced in CCMs expressed high mRNA levels of CD74, a cell-surface form of the MHC class-II invariant chain.…”

Cerebral Cavernous Malformation: Immune and Inflammatory Perspectives

“…Despite the limitation of small sample size and the uncertainty of whether the mAbs would react with other antibodies, these data have given us a novel insight into the immunopathogenesis of CCM. In another piece of research, the infiltration of different subsets of leukocytes in CCM was revealed ( 72 ). Through RNA-seq analysis, increased inflammation-related genes were observed in the endothelial cells from CCM mouse model ( 72 ).…”

“…In another piece of research, the infiltration of different subsets of leukocytes in CCM was revealed ( 72 ). Through RNA-seq analysis, increased inflammation-related genes were observed in the endothelial cells from CCM mouse model ( 72 ). Notably, the cytokines which perform the roles of inflammatory and immune cells’ recruitment, involving proinflammatory cytokines, chemokines, and adhesion molecules, are closely related to the differentially expressed genes.…”