Neuro-immune-epithelial pathways involving substance P may contribute to mucosal pathology in gastro-oesophageal reflux disease

Introduction Substance P (SP) is contained in nerve fibres innervating the oesophageal mucosa and is released into the mucosa during oesophageal acid perfusion. However, the downstream effects of this peripheral SP are not known. We investigated the expression of SP receptors in the oesophageal mucosa of healthy controls and GORD patients, localisation of mast cells in relation to SP+ nerve fibres, and effects of SP exposure on oesophageal epithelial cells. Methods Distal oesophageal biopsies were collected from healthy controls (HC; N = 10), functional heartburn (FH; N = 10), non-erosive reflux disease (NERD; N = 14), and erosive reflux disease (ERD; N = 13) patients. IHC was used to determine the cellular expression of the SP receptor, NK1R, on E-Cadherin+, CD3+, and tryptase+ cells. NE-1 oesophageal epithelial cells were incubated with SP to investigate inflammatory pathway activation. The proportion and density of mast cells expressing an SP receptor, MRGPRX2, was analysed, and the distance between SP+ nerve fibres and mast cells was determined. Results NK1R was expressed on epithelial cells and, rarely, on CD3+ T cells. NK1R fluorescence intensity was significantly higher in NERD and ERD oesophageal mucosa compared with HCs. In NE-1 cells, exposure to SP induced NF-κB phosphorylation, ameliorated by NK1R inhibition, and release of IL-6 and IL-8. In NERD and ERD mucosa, the density of MRGPRX2+ mast cells was significantly higher than HCs. Across all subjects, 25 SP+ nerves were detected; 32% were located within 10 µm of a mast cell, and 84% within 50 µm. Conclusion Expressions of SP receptors NK1R and MRGPRX2 are upregulated in NERD and ERD oesophageal mucosa, on epithelial cells and mast cells, respectively. Mast cells are also closely apposed to SP+ nerve fibres in the oesophageal mucosa. Furthermore, SP directly induced inflammatory pathway activation and release of cytokines in oesophageal epithelial cells in vitro . Communication between SP+ nerve fibres, epithelial cells, and mast cells may be of pathological significance in GORD.