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Role of Quercetin in Modulating Chloride Transport in the Intestine

View Article: PubMed Central - PubMed

ABSTRACT

Epithelial chloride channels provide the pathways for fluid secretion in the intestine. Cystic fibrosis transmembrane conductance regulator (CFTR) and calcium-activated chloride channels (CaCCs) are the main chloride channels in the luminal membrane of enterocytes. These transmembrane proteins play important roles in many physiological processes. In this study, we have identified a flavonoid quercetin as a modulator of CaCC chloride channel activity. Fluorescence quenching assay showed that quercetin activated Cl− transport in a dose-dependent manner, with EC50 ~37 μM. Short-circuit current analysis confirmed that quercetin activated CaCC-mediated Cl− currents in HT-29 cells that can be abolished by CaCCinh-A01. Ex vivo studies indicated that application of quercetin to mouse ileum and colon on serosal side resulted in activation of CFTR and CaCC-mediated Cl− currents. Notably, we found that quercetin exhibited inhibitory effect against ANO1 chloride channel activity in ANO1-expressing FRT cells and decreased mouse intestinal motility. Quercetin-stimulated short-circuit currents in mouse ileum was multi-component, which included elevation of Ca2+ concentration through L-type calcium channel and activation of basolateral NKCC, Na+/K+-ATPase, and K+ channels. In vivo studies further revealed that quercetin promoted fluid secretion in mouse ileum. The modulatory effect of quercetin on CaCC chloirde channels may therefore represent a potential therapeutic strategy for treating CaCC-related diseases like constipation, secretory diarrhea and hypertension. The inverse effects of quercetin on CaCCs provided evidence that ANO1 and intestinal epithelial CaCCs are different calcium-activated chloride channels.

No MeSH data available.


Activation of CaCC chloride channel activity by quercetin. (A) Activation of CaCC -mediated Cl− current by apical application of quercetin without or with subsequent addition of 30 μM CaCCinh-A01. (B) Representative trace of short-circuit current activated by basolateral administration of quercetin without or with subsequent addition of 30 μM CaCCinh-A01. The histograms show the magnitudes of short-circuit current obtained from the corresponding traces (n = 4).
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Figure 2: Activation of CaCC chloride channel activity by quercetin. (A) Activation of CaCC -mediated Cl− current by apical application of quercetin without or with subsequent addition of 30 μM CaCCinh-A01. (B) Representative trace of short-circuit current activated by basolateral administration of quercetin without or with subsequent addition of 30 μM CaCCinh-A01. The histograms show the magnitudes of short-circuit current obtained from the corresponding traces (n = 4).

Mentions: To confirm the activation of CaCC by quercetin, short-circuit current was further measured in HT-29 cells. Since quercetin also acts as a CFTR chloride channel activator (Pyle et al., 2010; Zhang et al., 2011), 20 μM CFTRinh-172 was added to the bath solution before administration of quercetin to eliminate the influence of CFTR-mediated Cl− current. The result showed that quercetin in the apical side of HT-29 monolayers activated the short-circuit currents in a dose-dependent manner. The activation effect could be abolished by the CaCC-specific inhibitor CaCCinh-A01 (30 μM) (Figure 2A). In addition, basolateral application of quercetin also activated CaCC-mediated short-circuit current, although this was much less potent than that produced by apical side application (Figure 2B). These results suggested that quercetin can activate both CFTR and CaCC mediated Cl− transport in HT-29 cells.


Role of Quercetin in Modulating Chloride Transport in the Intestine
Activation of CaCC chloride channel activity by quercetin. (A) Activation of CaCC -mediated Cl− current by apical application of quercetin without or with subsequent addition of 30 μM CaCCinh-A01. (B) Representative trace of short-circuit current activated by basolateral administration of quercetin without or with subsequent addition of 30 μM CaCCinh-A01. The histograms show the magnitudes of short-circuit current obtained from the corresponding traces (n = 4).
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Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC5120089&req=5

Figure 2: Activation of CaCC chloride channel activity by quercetin. (A) Activation of CaCC -mediated Cl− current by apical application of quercetin without or with subsequent addition of 30 μM CaCCinh-A01. (B) Representative trace of short-circuit current activated by basolateral administration of quercetin without or with subsequent addition of 30 μM CaCCinh-A01. The histograms show the magnitudes of short-circuit current obtained from the corresponding traces (n = 4).
Mentions: To confirm the activation of CaCC by quercetin, short-circuit current was further measured in HT-29 cells. Since quercetin also acts as a CFTR chloride channel activator (Pyle et al., 2010; Zhang et al., 2011), 20 μM CFTRinh-172 was added to the bath solution before administration of quercetin to eliminate the influence of CFTR-mediated Cl− current. The result showed that quercetin in the apical side of HT-29 monolayers activated the short-circuit currents in a dose-dependent manner. The activation effect could be abolished by the CaCC-specific inhibitor CaCCinh-A01 (30 μM) (Figure 2A). In addition, basolateral application of quercetin also activated CaCC-mediated short-circuit current, although this was much less potent than that produced by apical side application (Figure 2B). These results suggested that quercetin can activate both CFTR and CaCC mediated Cl− transport in HT-29 cells.

View Article: PubMed Central - PubMed

ABSTRACT

Epithelial chloride channels provide the pathways for fluid secretion in the intestine. Cystic fibrosis transmembrane conductance regulator (CFTR) and calcium-activated chloride channels (CaCCs) are the main chloride channels in the luminal membrane of enterocytes. These transmembrane proteins play important roles in many physiological processes. In this study, we have identified a flavonoid quercetin as a modulator of CaCC chloride channel activity. Fluorescence quenching assay showed that quercetin activated Cl− transport in a dose-dependent manner, with EC50 ~37 μM. Short-circuit current analysis confirmed that quercetin activated CaCC-mediated Cl− currents in HT-29 cells that can be abolished by CaCCinh-A01. Ex vivo studies indicated that application of quercetin to mouse ileum and colon on serosal side resulted in activation of CFTR and CaCC-mediated Cl− currents. Notably, we found that quercetin exhibited inhibitory effect against ANO1 chloride channel activity in ANO1-expressing FRT cells and decreased mouse intestinal motility. Quercetin-stimulated short-circuit currents in mouse ileum was multi-component, which included elevation of Ca2+ concentration through L-type calcium channel and activation of basolateral NKCC, Na+/K+-ATPase, and K+ channels. In vivo studies further revealed that quercetin promoted fluid secretion in mouse ileum. The modulatory effect of quercetin on CaCC chloirde channels may therefore represent a potential therapeutic strategy for treating CaCC-related diseases like constipation, secretory diarrhea and hypertension. The inverse effects of quercetin on CaCCs provided evidence that ANO1 and intestinal epithelial CaCCs are different calcium-activated chloride channels.

No MeSH data available.