Introduction

Disorders of gut-brain interaction (DGBIs), such as functional dyspepsia and irritable bowel syndrome, are chronic gastrointestinal disorders with recurrent and bothersome abdominal symptoms. Although incompletely understood, immune activation, particularly involving eosinophils and mast cells, and visceral hypersensitivity have been implicated in their pathophysiology.

Aims & Methods

Using the normoglycemic Biobreeding diabetes-prone (BBDP-N) rat, we aim to 1) characterise the immune activation in jejunal and colonic mucosa and to 2) study the effect of budesonide, a local anti-inflammatory corticosteroid, on gut physiology and sensitivity.
90-day-old BBDP-N and Biobreeding diabetes-resistant (control) rats underwent a 28-day treatment with budesonide (0.5 mg/kg/day) or vehicle. Eosinophils and mast cells were identified by staining jejunal and colonic sections with Chromotrope 2R and anti-mast cell protease 2 antibody, respectively. To assess their activity, eosinophil peroxidase (EPO) and β-hexosaminidase levels were measured. Jejunal and colonic segments were mounted in modified Ussing chambers, and permeability was assessed by the transmucosal passage of a fluorescently labelled 20 and 4kDa dextran molecule (FITC-Dx4/20), and the transepithelial electrical resistance (TEER). Sensitivity to isobaric colorectal distension was assessed(1): the ∆ area under the curve (AUC) of the visceromotor response (VMR) to wall tension was calculated at baseline and after treatment. Depending on the normality of the data, an unpaired t-test or Mann-Whitney test was performed.

Results

Comparing vehicle-treated groups, the BBDP-N rats had higher numbers of jejunal eosinophils (1366.0±201.3 vs. 561.7±55.3 eosinophils/mm²; p<0.01) and mast cells (1151.0±136.7 vs. 549.1±44.8 mast cells/mm²; p<0.001), whereas only colonic eosinophil numbers were higher (222.7±40.4 vs. 128.9±14.5 eosinophils/mm²; p=0.03). Additionally, EPO levels were higher in the BBDP-N rats in both regions (jejunum: 59.4±9.8 vs. 4.4±3.0 µg/g tissue; p<0.01 & colon: 82.0±23.6 vs. 1.6±1.1 µg/g tissue; p<0.01), with no difference in mast cell activation. Following budesonide treatment, eosinophil (jejunum, BBDP-N: 167.3±53.6 vs. 1366.0±201.3 eosinophils/mm²; p<0.001 & control: 220.1±43.7 vs. 561.7±55.3 eosinophils/mm²; p<0.01, colon: BBDP-N: 58.2±11.9 vs. 222.7±40.4 eosinophils/mm²; p<0.01 & control: 78.1±14.4 vs. 128.9±14.5 eosinophils/mm²; p=0.01) and mast cell (jejunum: BBDP-N: 418.8±51.0 vs. 1151.0±136.7 mast cells/mm²; p<0.001 & control: 305.4±67.6 vs. 549.1±44.8 mast cells/mm²; p=0.01, colon: BBDP-N: 146.5±19.1 vs. 262.6±37.5 mast cells/mm²; p<0.01 & control: 137.4±17.8 vs. 244.9±29.4 mast cells/mm²; p<0.01) levels decreased in both groups in both regions. Colonic eosinophil activation was also reduced in BBDP-N rats (21.5±9.7 vs. 82.0±23.6 µg/g tissue; p=0.05), with no effect on mast cell activation. Jejunal permeability, assessed by 20kDa passage was increased in vehicle-treated BBDP-N rats (258.0±51.5 vs. 132.0±25.1 pmol/cm²; p=0.03). Budesonide treatment significantly increased colonic TEER in BBDP-N rats (73.2±4.3 vs. 38.3±4.1 Ω.cm²; p<0.001). Colonic sensitivity to distention was significantly reduced in BBDP-N following the treatment (-20.1±17.2 vs. 207.0±85.5 mV/s x mmHg.cm; p=0.02).

Conclusion

In conclusion, our work shows the importance of eosinophil and mast cell predominant immune activation in visceral hypersensitivity in a well-characterised preclinical model of DGBI. Anti-inflammatory treatments should be re-evaluated in DGBI with intestinal eosinophil and mast cell infiltration.

References

1: Meleine M et al. Neurogastroenterology and motility, 2019.