4th Annual BMRP Investigator Meeting - Abstract

Enteric Glia in Intestinal Inflammation

Sebastian Hoff^a, Christiane Hank, Klaus Michel^b, Anne Rühl

Department of Human Biology, Technical University of Munich (Munich, Germany)

Enteric glial cells (EGC) are the predominant cell type in the enteric nervous system (ENS), a network in the gut wall that is capable of controlling all vital gut functions.  The importance of EGC has been demonstrated in a transgenic mouse model where selective ablation of EGC results in epithelial barrier dysfunction with consecutive severe intestinal inflammation and ultimately death of the animals (Bush et al., Cell 1998).  The underlying mechanisms are not yet elucidated.  Therefore, we aimed to study the effects of EGC on epithelial cell function in inflammatory conditions employing a coculture model in which intestinal epithelial cells (IEC; T84 cells) are grown as a mature monolayer in the apical compartment of a transwell filter system with or without EGC in the basolateral compartment.  Inflammatory conditions were mimicked by application of the proinflammatory cytokine tumor necrosis factor (TNF) a  (10ng/mL).  The tightness of the epithelial monolayer was assessed by quantifying both transepithelial electrical resistance (TER) and paracellular flux of fluorescein labeled dextrans. Under unstimulated conditions, coculturing IEC with EGC significantly enhanced TER within 48hrs.  After TNFa stimulation, a marked drop in TER was observed, most likely reflecting tight–junctional alterations because there were no detectable changes in macromolecular permeability at any time-point.  The cytokine-induced drop in TER was prevented when EGC were present in the basolateral compartment during cytokine exposure, suggesting that EGC release (a) soluble mediator(s) to maintain epithelial tight-junctional functions.  Forty-eight hrs after cytokine exposure, all treated IEC displayed a significant rise in TER, which we interpret to reflect counter-regulatory processes after an acute cytokine challenge.  This effect was potentiated in the presence of cytokine-stimulated EGC in the basolateral compartment, indicating that glial mediator release may be cytokine-inducible. Neurotrophin production was measured by qPCR and revealed nerve growth factor (NGF), neurotrophin 3 (NT-3) and glial cell-line derived neurotrophic factor (GDNF) production by basolateral EGC.

Clinically, it is important that disruption of the EGC network has been shown in Crohn’s disease (CD) patients (Cornet et al., PNAS 2001).  However, until now, reliable quantitative markers for human EGC have not been available.  To be able to identify and quantify all EGC in the human gut, we have therefore established an antibody directed against the transcription factor Sox10 as a novel, highly specific pan-glial marker in the ENS.  Employing this marker, we have assembled a comprehensive data set that now constitutes the basis of our ongoing morphometrical studies in human CD tissues.  Overall, we have established the novel glial marker Sox10 to identify and quantify human EGC and we have evidenced glial-epithelial interaction in a unique coculture model.  Currently, we are using these tools to further elucidate the mechanisms by which EGC may modulate the human inflammatory bowel diseases.

^aCo-investigator and Presenter; bPrincipal Investigator