Background: Enteric glial cells (EGC) participate to gut homeostasis and are involved in intestinal inflammation. In humans, gut inflammation is characterized by EGC-derived S100B protein up-regulation. Nonetheless, it is still unclear if EGC-response is directly involved in, or it represents an epiphenomenon of inflammation itself. Aims: To investigate: 1) the role of EGC-derived S100B protein in mediating inflammation and nitric oxide (NO) production; 2) the enteroglial activation and responses to inflammatory stimuli in mucosa and in isolated myenteric ganglia and 3) the ability of EGC to stimulate immune cells proliferation. Methods: Rectal biopsies from 30 control subjects were stimulated with exogenous S100B and iNOS expression, NO production, lipid peroxidation and p38MAPkinase activation were evaluated in the presence of anti-RAGE neutralizing antibody, SB203580 (inhibitor of p38MAPkinase), TLCK (inhibitor of Nuclear Factor-kappaB transcription), respectively. Mucosal biopsies and isolated myenteric ganglia (obtained from the ileum of 10 surgical specimens) were stimulated with lipopolysaccharides (LPS)+interferon gamma (IFN-γ); a double labeling immunofluorescence using antibodies to S100B or GFAP was performed to identify EGC expressing c-fos in the nucleus, as a marker of cells activation Similarly, S100B mRNA, protein expression and secretion and iNOS expression and NO production were respectively assessed either in the presence, or absence of anti-RAGE antibody. Proliferation of human peripheral blood mononuclear cells (PBMC) isolated from healthy subjects was also checked by incubation with exogenous S100B. Results: In mucosal biopsies, exogenous S100B increased iNOS expression,NOproduction and lipid peroxidation through p38MAPkinase/Nuclear Factor-kappaB activation and via RAGE. Similarly, the addition of LPS/IFN-γ significantly increased S100B mRNA, protein expression and secretion, accompanied with enhanced iNOS expression and NO production. In isolated myenteric ganglia, LPS/IFN-γ incubation resulted in a consistent EGC's activation (c-fos positive nuclei in S100B/GFAP positive cells), together with S100B up-regulation and increased NO production. Interestingly, exogenous S100B significantly increased PBMC proliferation and activation (increased NO and Tumor Necrosis Factor-alpha production) in a RAGE-dependent manner. Conclusions: EGC are activated by inflammatory stimuli and directly participate to NO production through S100B up-regulation. Increased levels of EGC-derived S100B are able to stimulate immune cells proliferation in a RAGE-dependent pathway.

Enteric Glia Stimulates Inflammation-Induced Responses in Human Intestine and Interacts with Immune Cells via S100B Protein

CIRILLO, CARLA;SARNELLI, GIOVANNI;GROSSO, MICHELA;CUOMO, ROSARIO
2009

Abstract

Background: Enteric glial cells (EGC) participate to gut homeostasis and are involved in intestinal inflammation. In humans, gut inflammation is characterized by EGC-derived S100B protein up-regulation. Nonetheless, it is still unclear if EGC-response is directly involved in, or it represents an epiphenomenon of inflammation itself. Aims: To investigate: 1) the role of EGC-derived S100B protein in mediating inflammation and nitric oxide (NO) production; 2) the enteroglial activation and responses to inflammatory stimuli in mucosa and in isolated myenteric ganglia and 3) the ability of EGC to stimulate immune cells proliferation. Methods: Rectal biopsies from 30 control subjects were stimulated with exogenous S100B and iNOS expression, NO production, lipid peroxidation and p38MAPkinase activation were evaluated in the presence of anti-RAGE neutralizing antibody, SB203580 (inhibitor of p38MAPkinase), TLCK (inhibitor of Nuclear Factor-kappaB transcription), respectively. Mucosal biopsies and isolated myenteric ganglia (obtained from the ileum of 10 surgical specimens) were stimulated with lipopolysaccharides (LPS)+interferon gamma (IFN-γ); a double labeling immunofluorescence using antibodies to S100B or GFAP was performed to identify EGC expressing c-fos in the nucleus, as a marker of cells activation Similarly, S100B mRNA, protein expression and secretion and iNOS expression and NO production were respectively assessed either in the presence, or absence of anti-RAGE antibody. Proliferation of human peripheral blood mononuclear cells (PBMC) isolated from healthy subjects was also checked by incubation with exogenous S100B. Results: In mucosal biopsies, exogenous S100B increased iNOS expression,NOproduction and lipid peroxidation through p38MAPkinase/Nuclear Factor-kappaB activation and via RAGE. Similarly, the addition of LPS/IFN-γ significantly increased S100B mRNA, protein expression and secretion, accompanied with enhanced iNOS expression and NO production. In isolated myenteric ganglia, LPS/IFN-γ incubation resulted in a consistent EGC's activation (c-fos positive nuclei in S100B/GFAP positive cells), together with S100B up-regulation and increased NO production. Interestingly, exogenous S100B significantly increased PBMC proliferation and activation (increased NO and Tumor Necrosis Factor-alpha production) in a RAGE-dependent manner. Conclusions: EGC are activated by inflammatory stimuli and directly participate to NO production through S100B up-regulation. Increased levels of EGC-derived S100B are able to stimulate immune cells proliferation in a RAGE-dependent pathway.
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Utilizza questo identificativo per citare o creare un link a questo documento: http://hdl.handle.net/11588/375570
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