Abstract
Kaempferol is a flavonol identified as the most potent activator of chloride (Cl(-)) secretion among other flavonoids in airway epithelial cells. This study aimed to investigate the cellular mechanisms by which kaempferol stimulates Cl(-) secretion in the T84 human colon carcinoma cell line by Ussing chambers and voltage clamp technique. Bilateral addition of kaempferol (1-100 µM) increased short-circuit current (I (sc) ) in a concentration-dependent manner. Ion substitution of Cl(-) or CFTR inhibitors NPPB and glibenclamide or a Na(+)/K(+)/2Cl(-) cotransporter inhibitor bumetanide attenuated kaempferol-induced I (sc) response. In permeabilized monolayers, selective channel inhibitors CFTRinh-172 and CaCCinh-A01 inhibited kaempferol-induced apical Cl(-) current (I (Cl) ), and K(+) blockers BaCl(2) and clotrimazole inhibited basolateral K(+) current (I (Kb) ). The kaempferol-induced I (Cl) showed no additive effects with forskolin or 8cpt-cAMP. The kaempferol-induced I (Cl) was mostly abolished by protein kinase A inhibitor H89, but not by tyrosine kinase inhibitors, AG490 and tyrphostin A23, or tyrosine phosphatase inhibitor vanadate. Treatment with kaempferol for 24 h increased the expression of CFTR protein as determined by the Western blot analysis. These results demonstrated that kaempferol activates Cl(-) secretion across T84 cells by activating the apical Cl(-) current and basolateral K(+) current. The mechanisms may involve the cAMP/PKA pathway and CFTR expression. Taken together, these findings reveal the beneficial effects of kaempferol to increase fluid secretion which can be used to treat constipation.