Endothelial basement membranes constitute barriers to extravasating leukocytes during inflammation, a process where laminin isoforms define sites of leukocyte exit; however, how this occurs is poorly understood. In addition to a direct effect on leukocyte transmigration, we show that laminin 511 affects endothelial barrier function by stabilizing VE-cadherin at junctions and downregulating expression of CD99L2, correlating with reduced neutrophil extravasation. Binding of endothelial cells to laminin 511, but not laminin 411 or non-endothelial laminin 111, enhanced transendothelial cell electrical resistance (TEER) and inhibited neutrophil transmigration. Data suggest that endothelial adhesion to laminin 511 via β1 and β3 integrins mediates RhoA-induced VE-cadherin localization to cell-cell borders, and while CD99L2 downregulation requires integrin β1, it is RhoA-independent. Our data demonstrate that molecular information provided by basement membrane laminin 511 affects leukocyte extravasation both directly and indirectly by modulating endothelial barrier properties.

Enteric glial cells (EGC) modulate motility, maintain gut homeostasis, and contribute to neuroinflammation in intestinal diseases and motility disorders. Damage induces a reactive glial phenotype known as "gliosis", but the molecular identity of the inducing mechanism and triggers of "enteric gliosis" are poorly understood. We tested the hypothesis that surgical trauma during intestinal surgery triggers ATP release that drives enteric gliosis and inflammation leading to impaired motility in postoperative ileus (POI). ATP activation of a p38-dependent MAPK pathway triggers cytokine release and a gliosis phenotype in murine (and human) EGCs. Receptor antagonism and genetic depletion studies revealed P2X2 as the relevant ATP receptor and pharmacological screenings identified ambroxol as a novel P2X2 antagonist. Ambroxol prevented ATPinduced enteric gliosis, inflammation, and protected against dysmotility, while abrogating enteric gliosis in human intestine exposed to surgical trauma. We identified a novel pathogenic P2X2dependent pathway of ATP-induced enteric gliosis, inflammation and dysmotility in humans and mice. Interventions that block enteric glial P2X2 receptors during trauma may represent a novel therapy in treating POI and immune-driven intestinal motility disorders.

Molecular mechanisms that determine lesion localization or phenotype variation in multiple sclerosis are mostly unidentified. Although transmigration of activated encephalitogenic T cells across the bloodbrain barrier (BBB) is a crucial step in the disease pathogenesis of CNS autoimmunity, the consequences on brain endothelial barrier integrity upon interaction with such T cells and subsequent lesion formation and distribution are largely unknown. We made use of a transgenic spontaneous mouse model of CNS autoimmunity characterized by inflammatory demyelinating lesions confined to optic nerves and spinal cord (OSE mice). Genetic ablation of a single immune-regulatory molecule in this model [i.e., B7-homolog 1 (B7-H1, PD-L1)] not only significantly increased incidence of spontaneous CNS autoimmunity and aggravated disease course, especially in the later stages of disease, but also importantly resulted in encephalitogenic T-cell infiltration and lesion formation in normally unaffected brain regions, such as the cerebrum and cerebellum. Interestingly, B7-H1 ablation on myelin oligodendrocyte glycoprotein-specific CD4 + T cells, but not on antigen-presenting cells, amplified T-cell effector functions, such as IFN-γ and granzyme B production. Therefore, these T cells were rendered more capable of eliciting cell contact-dependent brain endothelial cell dysfunction and increased barrier permeability in an in vitro model of the BBB. Our findings suggest that a single immuneregulatory molecule on T cells can be ultimately responsible for localized BBB breakdown, and thus substantial changes in lesion topography in the context of CNS autoimmunity.neuroinflammation | multiple sclerosis | spontaneous EAE | CNS lesion distribution | blood-brain barrier M ultiple sclerosis (MS) is the most common chronic inflammatory demyelinating disease of the CNS. Disease pathogenesis is initiated by peripheral activation of autoimmune T lymphocytes by yet unknown mechanisms, followed by T-cell expansion and subsequent migration across the complex structure of the blood-brain barrier (BBB). Within the CNS, entry of this first wave of T cells elicits recruitment of other immune cells, which together evoke a local inflammatory process ultimately resulting in demyelination, as well as axonal and neuronal damage (1). Several histological MS subtypes have been described with regard to lesion distribution and cellular composition (2). The reasons underlying distinct lesion development at different anatomical sites still remain however largely elusive: Some authors have proposed that the nature and expression pattern of the target autoantigen might play a role (3, 4). Others have observed an influence of the HLA complex and its role in shaping antigen presentation, thus suggesting that T-cell antigen specificity might impact the location of inflammation (5). Additionally, T-cell polarization into distinct T helper subtypes, as well as their expression pattern of chemokine receptors and adhesion molecules, has been implicated in determining the lo...

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