Inducible Nitric Oxide Regulates Brush Border Membrane Na-Glucose Co-transport, but Not Na:H Exchange via p38 MAP Kinase in Intestinal Epithelial Cells

Palanikumar Manoharan,Shanmuga Sundaram,Soudamani Singh,Uma Sundaram

doi:10.3390/cells7080111

Palanikumar Manoharan, Shanmuga Sundaram + Show 2 more

Open Access

https://doi.org/10.3390/cells7080111

Copy DOI

Journal: Cells	Publication Date: Aug 19, 2018
Citations: 8	License type: CC BY 4.0

Affiliation: University of Cincinnati, Marshall University

Abstract

During chronic intestinal inflammation in rabbit intestinal villus cells brush border membrane (BBM) Na-glucose co-transport (SGLT1), but not Na/H exchange (NHE3) is inhibited. The mechanism of inhibition is secondary to a decrease in the number of BBM co-transporters. In the chronic enteritis mucosa, inducible nitric oxide (iNO) and superoxide production are known to be increased and together they produce abundant peroxynitrite (OONO), a potent oxidant. However, whether OONO mediates the SGLT1 and NHE3 changes in intestinal epithelial cells during chronic intestinal inflammation is unknown. Thus, we determined the effect of OONO on SGLT1 and NHE3 in small intestinal epithelial cell (IEC-18) monolayers grown on trans well plates. In cells treated with 100 μM SIN-1 (OONO donor) for 24 h, SGLT1 was inhibited while NHE3 activity was unaltered. SIN-1 treated cells produced 40 times more OONO fluorescence compared to control cells. Uric acid (1mM) a natural scavenger of OONO prevented the OONO mediated SGLT1 inhibition. Na+/K+-ATPase which maintains the favorable trans-cellular Na gradient for Na-dependent absorptive processes was decreased by OONO. Kinetics studies demonstrated that the mechanism of inhibition of SGLT1 by OONO was secondary to reduction in the number of co-transporters (Vmax) without an alteration in the affinity. Western blot analysis showed a significant decrease in SGLT1 protein expression. Further, p38 mitogen-activated protein (MAP) kinase pathway appeared to mediate the OONO inhibition of SGLT1. Finally, at the level of the co-transporter, 3-Nitrotyrosine formation appears to be the mechanism of inhibition of SGLT1. In conclusion, peroxynitrite inhibited BBM SGLT1, but not NHE3 in intestinal epithelial cells. These changes and the mechanism of SGLT1 inhibition by OONO in IEC-18 cells is identical to that seen in villus cells during chronic enteritis. Thus, these data indicate that peroxynitrite, known to be elevated in the mucosa, may mediate the inhibition of villus cell BBM SGLT1 in vivo in the chronically inflamed intestine.

Highlights

In the mammalian small intestine, glucose is primarily absorbed by Na-glucose co-transport (SGLT1; Slc5a1)
The uptake of 3-O-methyl glucose (OMG) was done in the presence and absence of extracellular Na in IEC-18 cells and Na stimulated 3-OMG uptake was measured as SGLT1 activity
SIN-1 treatment was used to demonstrate the effect of OONO on activity Na-glucose co-transport

Summary

Introduction

In the mammalian small intestine, glucose is primarily absorbed by Na-glucose co-transport (SGLT1; Slc5a1). The glucose transporter SGLT1 uses a Na+ gradient to transport Na+ and glucose at a. Cells 2018, 7, 111 with Na+ across the cell, which is accompanied by 260 water molecules [2]. This mechanism was calculated to account for five L of water absorbed per day in the human intestine and formed the molecular basis of oral rehydration therapy aimed to control mortality associated with cholera and other infectious diarrheal diseases [3]. All of these essential transport processes are found on the brush border membrane (BBM)

Methods

Results

Discussion

Conclusion