Basic Study
Copyright ©The Author(s) 2016. Published by Baishideng Publishing Group Inc. All rights reserved.
World J Gastrointest Pathophysiol. Feb 15, 2016; 7(1): 138-149
Published online Feb 15, 2016. doi: 10.4291/wjgp.v7.i1.138
Differential expression of pancreatic protein and chemosensing receptor mRNAs in NKCC1-null intestine
Emily M Bradford, Kanimozhi Vairamani, Gary E Shull
Emily M Bradford, Department of Internal Medicine, Division of Gastroenterology, University of Kentucky, Lexington, KY 40536-0298, United States
Kanimozhi Vairamani, Gary E Shull, Department of Molecular Genetics, Biochemistry and Microbiology, University of Cincinnati College of Medicine, Cincinnati, OH 45267-0524, United States
Author contributions: Bradford EM and Shull GE conceived the study, analyzed the data, and wrote the manuscript; Vairamani K analyzed and compiled microarray data; Bradford EM performed all of the experiments.
Supported by National Institutes of Health to Gary E Shull, No. DK050594.
Institutional review board statement: Because human subjects or tissues were not used in this study, approval from the institutional review board was not required. Ethical issues relating to the animal protocol were reviewed and approved by the Institutional Animal Care and Use Committee of the University of Cincinnati.
Institutional animal care and use committee statement: All procedures involving animals were reviewed and approved by the Institutional Animal Care and Use Committee (IACUC) of the University of Cincinnati (protocol number: 06-06-22-02).
Conflict-of-interest statement: The authors have no conflict of interest related to this manuscript.
Data sharing statement: The microarray data have been deposited in the National Center for Biotechnology Gene Expression Omnibus and can be freely accessed as described in Methods. The Slc12a2 (NKCC1) knockout mouse model has been deposited in a publically available repository and can be accessed as described in Methods.
Open-Access: This article is an open-access article which was selected by an in-house editor and fully peer-reviewed by external reviewers. It is distributed in accordance with the Creative Commons Attribution Non Commercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited and the use is non-commercial. See: http://creativecommons.org/licenses/by-nc/4.0/
Correspondence to: Gary E Shull, PhD, Professor, Department of Molecular Genetics, Biochemistry and Microbiology, University of Cincinnati College of Medicine, 231 Albert Sabin Way, Cincinnati, OH 45267-0524, United States. shullge@ucmail.uc.edu
Telephone: +1-513-5580056 Fax: +1-513-5591885
Received: June 29, 2015
Peer-review started: July 2, 2015
First decision: September 22, 2015
Revised: October 10, 2015
Accepted: December 17, 2015
Article in press: December 18, 2015
Published online: February 15, 2016
Processing time: 216 Days and 7.8 Hours
Abstract

AIM: To investigate the intestinal functions of the NKCC1 Na+-K+-2Cl cotransporter (SLC12a2 gene), differential mRNA expression changes in NKCC1-null intestine were analyzed.

METHODS: Microarray analysis of mRNA from intestines of adult wild-type mice and gene-targeted NKCC1-null mice (n = 6 of each genotype) was performed to identify patterns of differential gene expression changes. Differential expression patterns were further examined by Gene Ontology analysis using the online Gorilla program, and expression changes of selected genes were verified using northern blot analysis and quantitative real time-polymerase chain reaction. Histological staining and immunofluorescence were performed to identify cell types in which upregulated pancreatic digestive enzymes were expressed.

RESULTS: Genes typically associated with pancreatic function were upregulated. These included lipase, amylase, elastase, and serine proteases indicative of pancreatic exocrine function, as well as insulin and regenerating islet genes, representative of endocrine function. Northern blot analysis and immunohistochemistry showed that differential expression of exocrine pancreas mRNAs was specific to the duodenum and localized to a subset of goblet cells. In addition, a major pattern of changes involving differential expression of olfactory receptors that function in chemical sensing, as well as other chemosensing G-protein coupled receptors, was observed. These changes in chemosensory receptor expression may be related to the failure of intestinal function and dependency on parenteral nutrition observed in humans with SLC12a2 mutations.

CONCLUSION: The results suggest that loss of NKCC1 affects not only secretion, but also goblet cell function and chemosensing of intestinal contents via G-protein coupled chemosensory receptors.

Keywords: SLC12a2; Chemosensory; Chemosensitivity; Gastrointestinal; Dyspepsia

Core tip: The NKCC1 Na+-K+-2Cl- cotransporter is a major mechanism of Cl- uptake in support of secretion. To investigate its intestinal functions we analyzed mRNA expression changes in NKCC1-null intestines. Differentially expressed genes included digestive enzymes and a large number of olfactory and other G-protein coupled receptors that function in chemical sensing. This suggests that loss of NKCC1 affects not only secretion, but also digestion and chemosensing of components of the intestinal contents. The results likely have relevance to recent evidence showing that mutations in the human NKCC1 gene cause unexplained food intolerance and failure of intestinal function requiring parenteral nutrition.