Chronic and selective inhibition of basolateral membrane Na-K-ATPase uniquely regulates brush border membrane na absorption in intestinal epithelial cells

Palanikumar Manoharan, Swapna Gayam, Subha Arthur, Balasubramanian Palaniappan, Soudamani Singh, Gregory M. Dick, Uma Sundaram

Research output: Contribution to journalArticle

4 Scopus citations

Abstract

Na-K-ATPase, an integral membrane protein in mammalian cells, is responsible for maintaining the favorable intracellular Na gradient necessary to promote Nacoupled solute cotransport processes [e.g., Na-glucose cotransport (SGLT1)]. Inhibition of brush border membrane (BBM) SGLT1 is, at least in part, due to the diminished Na-K-ATPase in villus cells from chronically inflamed rabbit intestine. The aim of the present study was to determine the effect of Na-K-ATPase inhibition on the two major BBM Na absorptive pathways, specifically Na-glucose cotransport and Na/H exchange (NHE), in intestinal epithelial (IEC-18) cells. Na-K-ATPase was inhibited using 1 mM ouabain or siRNA for Na-K-ATPase-α1in IEC-18 cells. SGLT1 activity was determined as 3-O-methyl-D-[3H]glucose uptake. Na-K-ATPase activity was measured as the amount of inorganic phosphate released. Treatment with ouabain resulted in SGLT1 inhibition at 1 h but stimulation at 24 h. To further characterize this unexpected stimulation of SGLT1, siRNA silencing was utilized to inhibit Na-K-ATPase-α1. SGLT1 activity was significantly upregulated by Na-K-ATPase silencing, while NHE3 activity remained unaltered. Kinetics showed that the mechanism of stimulation of SGLT1 activity was secondary to an increase in affinity of the cotransporter for glucose without a change in the number of cotransporters. Molecular studies demonstrated that the mechanism of stimulation was not secondary to altered BBM SGLT1 protein levels. Chronic and direct silencing of basolateral Na-KATPase uniquely regulates BBM Na absorptive pathways in intestinal epithelial cells. Specifically, while BBM NHE3 is unaffected, SGLT1 is stimulated secondary to enhanced affinity of the cotransporter.

Original languageEnglish
Pages (from-to)C650-C656
JournalAmerican Journal of Physiology - Cell Physiology
Volume308
Issue number8
DOIs
StatePublished - 2015

Keywords

  • Intestinal Inflammation
  • Na-glucose cotransporter
  • Na-k-atpase
  • Ouabain
  • Sglt1

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