Background : Steroid refractory acute graft-versus-host-disease (GVHD) after hematopoietic cell transplantation (HCT) is highly morbid with limited treatment options. Murine studies show protection from GVHD with butyrate exposure but direct exposure of stem/progenitor cells to butyrate inhibits colonic stem cell proliferation. Methods : Stool samples were collected weekly in a cohort of HCT recipients (n=210) undergoing allogenic transplant, and underwent 16S rRNA sequencing to determine the number and relative abundance of butyrogens. Dissociated primary human colonoid cell aggregates (200,000 per well) were plated onto collagen IV-coated transwells (0.4 µm pore size, 0.33cm2, PET) in stem cell medium for 24 hours. From 24h onwards, the basal-lateral chamber was switched to differentiation medium; the apical chamber was Hanks Buffered Salt Solution (HBSS), HBSS with 10mM butyrate sodium salt early (24h onwards) or late (72hrs onwards). Trans-epithelial electrical resistance was measured daily.
Results : Retrospective chart review identified 27 recipients who developed acute GVHD of the gut, stratified to be either steroid refractory GVHD (failed to respond to 2mg / kg of methylprednisolone) or responsive. The presence of butyrogens in the gut microbiome after the onset of severe acute GHVD of the gut associated with increased risk of steroid refractory GVHD (Fig 1; p < 0.05). Direct exposure of human colonic stem/progenitor cells to butyrate inhibits the development of trans-epithelial electrical resistance; exposure after differentiation had no inhibition of barrier formation (Fig 2; p < 0.05 by T-test). Conclusion : Butyrogens may help prevent the development of acute GVHD of the gut, but once severe GVHD has developed may inhibit recovery due to the loss of crypt architecture exposing colonic stem cells to microbe-produced butyrate with impaired differentiation and cell replacement.