Mast cells reduce survival of myenteric neurons in culture

Abstract

Mast cell–nerve interactions play a key role in intestinal inflammation and irritable bowel disease. Loss of enteric neurons has been reported in inflammatory conditions but the contribution of mast cells in this event is unknown. To study neuronal survival and plasticity of myenteric neurons in contact with mast cells a co-culture system using myenteric neurons from rat small intestine and peritoneal mast cells was set up. Dissociated myenteric neurons were cultured for 4 days before addition of mast cells isolated by peritoneal lavage. Neuronal survival and expression of vasoactive intestinal peptide (VIP) and nitric oxide synthase (NOS) were studied by immunocytochemistry and neuronal cell counting. Myenteric neurons cultured without mast cells were used to study the rate of neuronal survival after the addition of various mast cell mediators, proteinase-activated receptor 2 (PAR 2) agonist, VIP or corticosteroid. A striking mast cell-induced neuronal cell death was found after co-culturing. It was counteracted by the addition of mast cell stabiliser doxantrazole, protease inhibitors, PAR 2 antagonist FSLLRY-amide, corticosteroid or VIP. In myenteric neurons cultured without mast cells the PAR 2 agonist SLIGRL-amide, prostaglandin D 2 and interleukin (IL) 6 reduced neuronal survival while histamine, serotonin, heparin, IL1β and tumour necrosis factor α had no effect; corticosteroid and VIP enhanced neuronal survival. The relative numbers of VIP-, but not NOS-expressing myenteric neurons increased after co-culturing. Mast cell-induced neuronal cell death is suggested to be mediated via PAR 2 activation, IL6 and prostaglandin D 2. Corticosteroid and VIP are neuroprotective and able to prevent cell death of myenteric neurons in co-culture.