Glucose and Palmitate Differentially Regulate PFKFB3/iPFK2 and Inflammatory Responses in Mouse Intestinal Epithelial Cells

Rachel Botchlett,Jiajia Zhao,Honggui Li,Juan Zheng,Ting Qi,Guang Chen,Qifu Li,Xin Guo,Xiaoqiu Xiao,Chaodong Wu,Yuqing Huo,Xiang Hu,Ya Pei,Sijun Yang,Ting Guo,Mengyang Liu,Shih-Lung Woo

doi:10.1038/srep28963

Abstract

The gene PFKFB3 encodes for inducible 6-phosphofructo-2-kinase, a glycolysis-regulatory enzyme that protects against diet-induced intestine inflammation. However, it is unclear how nutrient overload regulates PFKFB3 expression and inflammatory responses in intestinal epithelial cells (IECs). In the present study, primary IECs were isolated from small intestine of C57BL/6J mice fed a low-fat diet (LFD) or high-fat diet (HFD) for 12 weeks. Additionally, CMT-93 cells, a cell line for IECs, were cultured in low glucose (LG, 5.5 mmol/L) or high glucose (HG, 27.5 mmol/L) medium and treated with palmitate (50 μmol/L) or bovine serum albumin (BSA) for 24 hr. These cells were analyzed for PFKFB3 and inflammatory markers. Compared with LFD, HFD feeding decreased IEC PFKFB3 expression and increased IEC proinflammatory responses. In CMT-93 cells, HG significantly increased PFKFB3 expression and proinflammatory responses compared with LG. Interestingly, palmitate decreased PFKFB3 expression and increased proinflammatory responses compared with BSA, regardless of glucose concentrations. Furthermore, HG significantly increased PFKFB3 promoter transcription activity compared with LG. Upon PFKFB3 overexpression, proinflammatory responses in CMT-93 cells were decreased. Taken together, these results indicate that in IECs glucose stimulates PFKFB3 expression and palmitate contributes to increased proinflammatory responses. Therefore, PFKFB3 regulates IEC inflammatory status in response to macronutrients.

Highlights

It is well established that inactivity and overnutrition are major determinants in the development of obesity and contribute to obesity-related metabolic diseases such as type 2 diabetes, fatty liver disease, and inflammatory atherosclerosis[1,2,3,4,5]
Considering that the intestine is responsible for digestion, absorption, and assimilation of nutrients and that the nutrients absorbed by intestine have undergone metabolism whose dysregulation accounts for increased proinflammatory responses, intestinal cells, in particular intestinal epithelial cells (IECs), may respond to nutrient overload to regulate its own inflammatory status prior to regulating inflammatory responses in distal organs
PFKFB3/iPFK2 is involved in the effect of rosiglitazone, one of the only two currently prescribed medicines as insulin-sensitizers for the treatment of type 2 diabetes, on suppressing intestinal inflammation

Summary

Introduction

It is well established that inactivity and overnutrition are major determinants in the development of obesity and contribute to obesity-related metabolic diseases such as type 2 diabetes, fatty liver disease, and inflammatory atherosclerosis[1,2,3,4,5]. The intestine has recently been implicated as another key organ that critically contributes to the development of obesity-associated chronic inflammation and systemic insulin resistance, and metabolic dysregulation[15,16,17,18]. Recent studies by Huo et al.[13,14,23] and Guo et al.[13,14,23] have further demonstrated that PFKFB3/iPFK2 critically determines the balance of metabolic fluxes through glycolysis and fatty acid oxidation, thereby suppressing the generation of reactive oxygen species and proinflammatory responses in adipocytes[13,14,23]. A previous study by Guo et al.[22] showed increased PFKFB3 expressions in the small intestine in response to HFD feeding, as well as increased inflammation. This study sought to first determine how macronutrients influence PFKFB3/iPFK2 expression in IECs and secondly, how this relates to the IEC inflammatory status

Methods

Results

Conclusion