(E38) Manipulating Epithelial Fluid Secretion Using Equine Rectal Organoids

Abstract: Background– A common mechanism of fluid loss in equine colitis is chloride secretion via the cystic fibrosis transmembrane receptor (CFTR) protein present on epithelial cells. Human rectal biopsies are used to test CFTR function and inhibition. Inhibition of CFTR is a promising target for anti-diarrheal medications, but there are no current CFTR inhibitors for equine diarrhea.
Hypothesis– Rectal organoids will swell in response to CFTR activation and this can be modulated with CFTR inhibitors.
Animals– 5 adult healthy horses (4 mares, 1 gelding; age range: 3-21 years) from NC State Research herds or donated for research.
Methods – Rectal biopsies were obtained under sedation or post-euthanasia (unrelated to study). Rectal crypts were isolated and plated to culture organoids. For all assays, organoids were passaged into 96-well plates and drug application was performed 2-3 days post-passaging. CFTR modulation was performed with the following activators (forskolin, DCEBIO, IBMX) and inhibitors (CFTRinh-172, GlyH-101). Organoid swelling was measured over time with calcein green fluorescence. Drug toxicity 2-3 days post-assay was identified with resazurin dye.
Results– Forskolin induces significant swelling in rectal organoids (Figure 1), consistent with CFTR activation, with no toxicity observed. DCEBIO, IBMX, and CFTRinh-172 minimally affected swelling, with no toxicity observed. GlyH-101 impaired forskolin-induced swelling at the highest dose tested but also exhibited toxicity.
Conclusions and clinical importance– Equine rectal organoids can serve as an in vitro model for chloride and fluid secretion in response to CFTR stimulation. Further experiments are necessary to identify safe and effective CFTR inhibitors for clinical use.