β-eudesmol but not atractylodin exerts an inhibitory effect on CFTR-mediated chloride transport in human intestinal epithelial cells

Oriental herbal medicine with the two bioactive constituents, beta-eudesmol (BE) and atractylodin (AT), has been used as a remedy for gastrointestinal disorders. There was no scientific evidence reporting their antidiarrheal effect and underpinning mechanisms. Therefore, we aimed to investigate the anti-secretory activity of these two compounds in vitro. The inhibitory effect of BE and AT on cAMP-induced Cl- secretion was evaluated by Ussing chamber in human intestinal epithelial (T84) cells. Short-circuit current (I-SC) and apical Cl- current (I-cl-) were measured after adding indirect and direct cystic fibrosis transmembrane conductance regulator (CFTR) chloride channel activator. MIT assay was used to determine cellular cytotoxicity. Protein-ligand interaction was investigated by in silico molecular docking analysis. BE, but not AT concentration-dependently (IC50 of similar to 1.05 mu M) reduced cAMP-mediated, CFTRinh-172 inhibitable Cl- secretion as determined by transepithelial I-SC across a monolayer of T84 cells. Potency of CFTR-mediated inhibition by BE did not change with the use of different CFTR activators suggesting a direct blockage of the channel active site(s). Pretreatment with BE completely prevented cAMP-induced I-cl- Furthermore, BE at concentrations up to 200 mu M (24 h) had no effect on T84 cell viability. In silico studies indicated that BE could best dock onto dephosphorylated structure of CFTR at ATP-binding pockets in nucleotide-binding domain (NBD) 2 region. These findings provide the first evidence for the anti-secretory effect of BE involving inhibition of CFTR function. BE represents a promising candidate for the therapeutic or prophylactic intervention of diarrhea resulted from intestinal hypersecretion of Cl-.