Contributions
Abstract: P388
Type: Poster
Abstract Category: Pathology and pathogenesis of MS - Inflammation and tissue damage
Objective: To characterize molecular mechanisms that mediate the protective function of reactive astrocytes against inflammatory Central Nervous System (CNS) lesion size and severity.
Background: A protective barrier restricts the passage of lymphocytes and soluble proteins from the bloodstream into the CNS parenchyma. This barrier termed the blood brain barrier (BBB), has been characterized as a single layer consisting of tight-junction (TJ) bonds between CNS microvascular endothelial cells. However, recent work deleting the layer of astrocytes encircling the BBB has revealed a protective role for this additional layer, termed the glia limitans (GL), in delimiting lesion size and severity. We hypothesized that astrocytes form TJs themselves at the GL under conditions of inflammation, thereby representing a second line of defense distal to the BBB.
Design and methods: We screened for changes in TJ molecule expression in primary human astrocytes under control and inflammatory conditions, a model of reactive astrogliosis, and positively identified three molecules : claudin-1, claudin-4 and JAM-A. Immunohistochemistry and immune-EM of two in vivo mouse models of inflammatory CNS disease revealed that all three proteins localized to TJ between astrocytic endfeet at the GL. In vitro co-culture assays of reactive astrocytes, in which these genes were silenced using siRNA, and CD3+ T cells, demonstrated that all three proteins regulate the capacity of reactive astrocytes to limit migration of inflammatory cells. An astrocyte-specific claudin-4 knock out mouse was used to characterize lesion pathology and was found to have increased lesion size and severity as well as significantly exacerbated clinical disability in experimental autoimmune encephalomyelitis (EAE).
Results: We report the presence of a second TJ barrier between the blood and the CNS, at the GL, composed of claudin-1, claudin-4 and JAM-A, which are dynamically upregulated in response to inflammatory stimuli. In vitro, claudin-4 knock-down in reactive astrocytes limits their ability to restrict leukocyte dispersion. In vivo, astrocyte-specific loss of claudin-4 leads to increased lesion size in inflammatory models, and worsened clinical disability in EAE.
Conclusions: reactive astrocytes upregulate claudin-1, claudin-4 and JAM-A, leading to the formation of TJs and a second barrier distal to the classical BBB at the GL, which protects against the size and severity of inflammatory CNS lesions.
Disclosure: Candice Chapouly: nothing to disclosure
Abstract: P388
Type: Poster
Abstract Category: Pathology and pathogenesis of MS - Inflammation and tissue damage
Objective: To characterize molecular mechanisms that mediate the protective function of reactive astrocytes against inflammatory Central Nervous System (CNS) lesion size and severity.
Background: A protective barrier restricts the passage of lymphocytes and soluble proteins from the bloodstream into the CNS parenchyma. This barrier termed the blood brain barrier (BBB), has been characterized as a single layer consisting of tight-junction (TJ) bonds between CNS microvascular endothelial cells. However, recent work deleting the layer of astrocytes encircling the BBB has revealed a protective role for this additional layer, termed the glia limitans (GL), in delimiting lesion size and severity. We hypothesized that astrocytes form TJs themselves at the GL under conditions of inflammation, thereby representing a second line of defense distal to the BBB.
Design and methods: We screened for changes in TJ molecule expression in primary human astrocytes under control and inflammatory conditions, a model of reactive astrogliosis, and positively identified three molecules : claudin-1, claudin-4 and JAM-A. Immunohistochemistry and immune-EM of two in vivo mouse models of inflammatory CNS disease revealed that all three proteins localized to TJ between astrocytic endfeet at the GL. In vitro co-culture assays of reactive astrocytes, in which these genes were silenced using siRNA, and CD3+ T cells, demonstrated that all three proteins regulate the capacity of reactive astrocytes to limit migration of inflammatory cells. An astrocyte-specific claudin-4 knock out mouse was used to characterize lesion pathology and was found to have increased lesion size and severity as well as significantly exacerbated clinical disability in experimental autoimmune encephalomyelitis (EAE).
Results: We report the presence of a second TJ barrier between the blood and the CNS, at the GL, composed of claudin-1, claudin-4 and JAM-A, which are dynamically upregulated in response to inflammatory stimuli. In vitro, claudin-4 knock-down in reactive astrocytes limits their ability to restrict leukocyte dispersion. In vivo, astrocyte-specific loss of claudin-4 leads to increased lesion size in inflammatory models, and worsened clinical disability in EAE.
Conclusions: reactive astrocytes upregulate claudin-1, claudin-4 and JAM-A, leading to the formation of TJs and a second barrier distal to the classical BBB at the GL, which protects against the size and severity of inflammatory CNS lesions.
Disclosure: Candice Chapouly: nothing to disclosure