Dataset Information

Rapamycin Inhibition of mTOR Reduces Levels of the Na+/H+ Exchanger 3 in Intestines of Mice and Humans, Leading to Diarrhea.

ABSTRACT: BACKGROUND & AIMS:The immunosuppressant rapamycin frequently causes noninfectious diarrhea in organ transplant recipients. We investigated the mechanisms of this process. METHODS:We performed a retrospective analysis of renal transplant recipients treated with rapamycin from 2003 through 2010 at Albany Medical College, collecting data on serum levels of rapamycin. Levels of the Na+/H+ exchanger 3 (NHE3) were measured in human ileal biopsy specimens from patients who did and did not receive rapamycin (controls), in ileum tissues from rats or mice given rapamycin, and in mice with intestine-specific disruption of mammalian target of rapamycin (Mtor) (mTOR(f/f):Villin-cre mice) or Atg7 (Atg7(flox/flox); Villin-Cre). Exchange activity and intestinal water absorption were measured using a pH-sensitive dye and small intestine perfusion, respectively. RESULTS:Episodes of noninfectious diarrhea occurred in organ recipients after increases in serum levels of rapamycin. The expression of NHE3 was reduced in the ileal brush border of patients with diarrhea. In rats and mice, continuous administration of low doses of rapamycin reduced levels of NHE3 in intestinal tissues; this effect was not observed in mice with intestinal deletion of ATG7, indicating that autophagy is required for the reduction. Administration of single high doses of rapamycin to mice, to model the spikes in rapamycin levels that occur in patients with severe diarrheal episodes, resulted in reduced phosphorylation of S6 and AKT in ileal tissues, indicating inhibition of the mTOR complex (mTORC1 and mTORC2). The intestines of mice with intestine-specific deletion of mTOR were dilated and contained large amounts of liquid stools; they also had reduced levels of total NHE3 and NHERF1 compared with control mice. We observed a significant reduction in Na(+)/H(+) exchange activity in ileum tissues from these mice. CONCLUSIONS:Rapamycin inhibition of mTOR reduces levels of NHE3 and Na(+)/H(+) exchange activity in intestinal tissues of patients and rodents. This process appears to require the autophagic activity mediated by ATG7. Loss of mTOR regulation of NHE3 could mediate the development of diarrhea in patients undergoing rapamycin therapy.

SUBMITTER: Yang J

PROVIDER: S-EPMC4849539 | biostudies-literature | 2015 Jul

REPOSITORIES: biostudies-literature

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Rapamycin Inhibition of mTOR Reduces Levels of the Na+/H+ Exchanger 3 in Intestines of Mice and Humans, Leading to Diarrhea.

Yang Jun J Zhao Xiaofeng X Patel Archana A Potru Rachana R Azizi-Ghannad Sadra S Dolinger Michael M Cao James J Bartholomew Catherine C Mazurkiewicz Joseph J Conti David D Jones David D Huang Yunfei Y Zhu Xinjun Cindy XC

Gastroenterology 20150330 1

<h4>Background & aims</h4>The immunosuppressant rapamycin frequently causes noninfectious diarrhea in organ transplant recipients. We investigated the mechanisms of this process.<h4>Methods</h4>We performed a retrospective analysis of renal transplant recipients treated with rapamycin from 2003 through 2010 at Albany Medical College, collecting data on serum levels of rapamycin. Levels of the Na+/H+ exchanger 3 (NHE3) were measured in human ileal biopsy specimens from patients who did and did no ...[more]

PMID: 25836987

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Project description:Transmissible gastroenteritis virus (TGEV) is member of the family Coronaviridae and mainly causes acute diarrhea. TGEV infection is characterized by vomiting, watery diarrhea, and severe dehydration, resulting in high mortality rates in neonatal piglets. TGEV infection symptoms are related to an imbalance of sodium absorption in small intestinal epithelial cells; however, the etiology of sodium imbalance diarrhea caused by TGEV remains unclear. In this study, we performed transcriptomic analysis of intestinal tissues from infected and healthy piglets and observed that the expression of NHE3, encoding Na+/H+ exchanger 3 (NHE3), the main exchanger of electroneutral sodium in intestinal epithelial cells, was significantly reduced upon TGEV infection. We also showed that specific inhibition of intestinal NHE3 activity could lead to the development of diarrhea in piglets. Furthermore, we revealed an interaction between TGEV N protein and NHE3 near the nucleus. The binding of TGEV N to NHE3 directly affected the expression and activity of NHE3 on the cell surface and affected cellular electrolyte absorption, leading to diarrhea. Molecular docking and computer-aided screening techniques were used to screen for the blocker of the interaction between TGEV N and NHE3, which identified irinotecan. We then demonstrated that irinotecan was effective in relieving TGEV-induced diarrhea in piglets. These findings provide new insights into the mechanism of TGEV-induced sodium imbalance diarrhea and could lead to the design of novel antiviral strategies against TGEV. IMPORTANCE A variety of coronaviruses have been found to cause severe diarrhea in hosts, including TGEV; however, the pathogenic mechanism is not clear. Therefore, prompt determination of the mechanism and identification of efficient therapeutic agents are required, both for public health reasons and for economic development. In this study, we demonstrated that NHE3 is the major expressed protein of NHEs in the intestine, and its expression decreased by nearly 70% after TGEV infection. Also, specific inhibition of intestinal NHE3 resulted in severe diarrhea in piglets. This demonstrated that NHE3 plays an important role in TGEV-induced diarrhea. In addition, we found that TGEV N directly regulates NHE3 expression and activity through protein-protein interaction, which is essential to promote diarrhea. Molecular docking and other techniques demonstrated that irinotecan could block the interaction and diarrhea caused by TGEV. Thus, our results provide a basis for the development of novel therapeutic agents against TGEV and guidance for the development of drugs for other diarrhea-causing coronaviruses.

Dataset Information

Rapamycin Inhibition of mTOR Reduces Levels of the Na+/H+ Exchanger 3 in Intestines of Mice and Humans, Leading to Diarrhea.

Publications

Rapamycin Inhibition of mTOR Reduces Levels of the Na+/H+ Exchanger 3 in Intestines of Mice and Humans, Leading to Diarrhea.

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