Functional characterization and localization of AQP3 in the human colon

American Journal of Physiology-Gastrointestinal and Liver Physi

Baba

Ushiki

et al. 2011

Ikarashi N, Baba K, Ushiki T, Kon R, Mimura A, Toda T, Ishii M, Ochiai W, Sugiyama K. The laxative effect of bisacodyl is attributable to decreased aquaporin-3 expression in the colon induced by increased PGE 2 secretion from macrophages. Am J Physiol Gastrointest Liver Physiol 301: G887-G895, 2011. First published August 25, 2011; doi:10.1152/ajpgi.00286.2011.-The purpose of this study was to investigate the role of aquaporin3 (AQP3) in the colon in the laxative effect of bisacodyl. After oral administration of bisacodyl to rats, AQP3, macrophages, cyclooxygenase 2 (COX2), and prostaglandin E 2 (PGE2) were examined in the colon. The mechanism by which bisacodyl decreases the expression of AQP3 was examined using HT-29 and Raw264.7 cells. When diarrhea occurred, a significant increase in the expression of PGE 2 and a decrease in AQP3 expression were observed. Immunostaining showed COX2 expression only in macrophages. The PGE 2 concentration increased significantly 30 min after the addition of bisacodyl to Raw264.7 cells. Thirty minutes after PGE 2 addition to HT-29 cells, the AQP3 expression level decreased to 40% of the control. When pretreated with indomethacin, bisacodyl did not induce an increase in the colon PGE 2 level, a decrease in the AQP3 expression level, or diarrhea. The results suggest that bisacodyl may decrease the expression of AQP3 in the colon, which inhibits water transfer from the luminal to the vascular side and leads to a laxative effect. This study also showed that direct activation of colon macrophages by bisacodyl increases the secretion of PGE 2, which acts as a paracrine factor and decreases AQP3 expression in colon mucosal epithelial cells. aquaporin-3; prostaglandin E 2; bisacodyl; cyclooxygenase IN RECENT YEARS, IT HAS BECOME increasingly clear that aquaporins (AQPs), water channels, are involved in water transport in the intestinal tract (20). There are currently 13 known types of AQPs in humans, AQP0 through AQP12, which are expressed in a variety of tissues (15). Several AQPs are expressed in the intestinal tract, and at least the following eight types are known to exist there: AQP1, AQP2, AQP3, AQP4, AQP7, AQP8, AQP9, and AQP10 (5, 7, 17, 23). The main AQPs expressed in the colon are AQP1, AQP2, AQP3, AQP4, and AQP8 (5,16,23). Of these, extensive research has been conducted on AQP3, which is considered to play an important role in the colon, especially regarding water transfer (13, 34). We have found that the administration of the osmotic laxative magnesium sulfate (MgSO 4 ) to rats increases the expression of AQP3 in the colon, and an increase in the expression of AQP3 plays an essential role in the laxative effect of MgSO 4 (11,12).Bisacodyl is classified as a stimulant laxative and is widely used to treat constipation. Bisacodyl increases the production of prostaglandin E 2 (PGE 2 ) in intestinal epithelial cells and inhibits the activity of Na ϩ -K ϩ -ATPase, and, as a result, the osmotic pressure in the intestinal tract increases. It is believed that this increase in osmoti...

Section: Resultsmentioning

confidence: 99%

The laxative effect of bisacodyl is attributable to decreased aquaporin-3 expression in the colon induced by increased PGE₂secretion from macrophages

American Journal of Physiology-Gastrointestinal and Liver Physi

Baba

Ushiki

et al. 2011

“…The second band was observed at approximately 50 kDa and represented the glycosylated form of AQP3. 13,14) Glycosylation has no influence on water permeability. [15][16][17] Therefore, in this study, both of these bands were accounted for when analyzing the total protein expression level of AQP3.…”

Section: Effects Of Gypsum Fibrosum or Caso 4 On The Expressionmentioning

confidence: 99%

Elucidating the Mechanism by Which <i>Gypsum fibrosum</i>, a Traditional Chinese Medicine, Maintains Cutaneous Water Content

Biological & Pharmaceutical Bulletin

Ogiue

Toyoda

et al. 2013

Aquaporin-3 (AQP3) plays an important role in maintaining the normal water content of the skin. Previously, we revealed that the expression of cutaneous AQP3 increased following oral administration of Gypsum fibrosum (main component: CaSO 4 ) to mice. The purpose of this study is to elucidate the mechanism by which Gypsum fibrosum increases the expression of cutaneous AQP3 in a keratinocyte cell line. Gypsum fibrosum or CaSO 4 was added to keratinocytes, and the expression level of AQP3, the Ca concentration, the activity of protein kinase C (PKC), and the degrees of phosphorylation of both extracellular signal-regulated kinase (ERK) and cAMP response element binding protein (CREB) were measured. The mRNA and protein expression levels of AQP3 increased significantly 6 h-post addition of Gypsum fibrosum. In keratinocytes treated with Gypsum fibrosum, increases in the concentration of intracellular Ca, PKC activity, and the phosphorylation of ERK and CREB were observed. Pre-treatment with GF109203X, a PKC inhibitor, suppressed the mRNA expression levels of AQP3. Similarly to treatment with Gypsum fibrosum, the addition of CaSO 4 led to the same observations in keratinocytes. It is hypothesized that Gypsum fibrosum causes an increase in the intracellular Ca concentration, PKC activity, and the phosphorylation levels of ERK and CREB, resulting in increased AQP3 expression in keratinocytes. In addition, it is possible that the effect of Gypsum fibrosum is attributable to CaSO 4 , based on the results demonstrating that the mechanisms of action of Gypsum fibrosum and CaSO 4 were nearly identical.

“…These are believed to represent non-glycosylated AQP3 and glycosylated AQP3, respectively. 15,16) The sum of these bands was analyzed as the protein expression level of AQP3. The protein expression levels of AQP3 in the colon increased over time following the administration of MgSO 4 , and at 8 h after administration, Rat faecal samples were collected at various times for up to 8 h beginning immediately after the administration of MgSO 4 , and faecal water content was measured.…”

Section: Changes In Mrna and Protein Expression Levels Of Aqp3 In Thementioning

confidence: 99%

Effects of Magnesium Sulphate Administration on Aquaporin 3 in Rat Gastrointestinal Tract

Biological & Pharmaceutical Bulletin

Ushiki

Mochizuki

et al. 2011

In human, aquaporins (AQPs) are expressed in a variety of tissues, and there are currently 13 known types of AQPs: AQP0 through AQP12.1) Several members of the AQP family are expressed in the intestinal tract, and at least 8 types are known to exist there: AQP1, AQP2, AQP3, AQP4, AQP7, AQP8, AQP9, and AQP10. 2-4)The main types expressed in mucosal epithelial cells in the colon are AQP3, AQP4 and AQP8.5) It has been reported that vasoactive intestinal polypeptide (VIP), a gastrointestinal hormone, is the causative agent in Verner-Morrison syndrome, a disease associated with diarrhoea 6) ; that intravenous administration of VIP in healthy adults causes diarrhoea 7) ; that serum VIP concentrations are elevated in rats with colitis 8) ; and that VIP increases mRNA and protein expression levels of AQP3 in HT-29 cells, which are human colonic epithelial cells.9) It has also been reported that following resection of the small bowel in rats, there is an increase in the mRNA expression of AQP3 in the colon as diarrhoea occurs.9) Based on these findings, AQP3 appears to play a particularly important role in water transport in the colon. Despite suggestions of an apparent relationship between the occurrence of diarrhoea and AQP3 expression levels, the manner in which variations in AQP3 expression are related to the mechanism of action of laxatives has yet to be elucidated.It is believed that commonly used osmotic laxatives, such as magnesium sulphate (MgSO 4 ) and magnesium oxide (MgO), induce diarrhoea by causing an increase in the osmotic pressure in the intestinal tract, 10) but no details are known about the relationship between these osmotic laxatives and water transport. In the present study, we investigated the role of AQP3 in the colon on the laxative effect of MgSO 4 , a widely used osmotic laxative. First, rats were administered MgSO 4 , and faecal water content was measured over time as an indicator of diarrhoea. Then, by analyzing the expression level of genes that sharply increase as a result of increased osmotic pressure, we investigated the relationship between diarrhoea and osmotic pressure. Next, protein expression levels of AQP3, AQP4, and AQP8 in the rat colon were analyzed by immunostaining in order to investigate the distribution and intensity of AQP expression. By analyzing the expression of AQP3 in the colons of rats administered MgSO 4 , the relationship between the expression level and the laxative effects of MgSO 4 was then investigated. MATERIALS AND METHODS AnimalsMale Wistar rats (10 weeks old) were purchased from Sankyo Labo Service Corp., Inc. (Tokyo, Japan). Each rat was caged separately and kept at room temperature (24Ϯ1°C) and 55Ϯ5% humidity with 12 h of light (artificial illumination; 08:00-20:00). The present study was conducted in accordance with the Guiding Principles for the Care and Use of Laboratory Animals, as adopted by the Committee on Animal Research at Hoshi University.Treatment Rats were fasted for 18 h before MgSO 4 administration (water provided ad libitum). An aqueous solut...