The Impact of High-Fat Diet on Metabolism and Immune Defense in Small Intestine Mucosa

Improved
procedures for sample preparation and proteomic data analysis
allowed us to identify 7700 different proteins in mouse small intestinal
mucosa and calculate the concentrations of >5000 proteins. We compared
protein concentrations of small intestinal mucosa from mice that were
fed for two months with normal diet (ND) containing 34.4% carbohydrates,
19.6% protein, and 3.3% fat or high-fat diet (HFD) containing 25.3%
carbohydrates, 24.1% protein, and 34.6% fat. Eleven percent of the
quantified proteins were significantly different between ND and HFD.
After HFD, we observed an elevation of proteins involved in protein
synthesis, protein N-glycosylation, and vesicle trafficking.
Proteins engaged in fatty acid absorption, fatty acid β-oxidation,
and steroid metabolism were also increased. Enzymes of glycolysis
and pentose phosphate cycle were decreased, whereas proteins of the
respiratory chain and of ATP synthase were increased. The protein
concentrations of various nutrient transporters located in the enterocyte
plasma membrane including the Na+-d-glucose cotransporter
SGLT1, the passive glucose transporter GLUT2, and the H+-peptide cotransporter PEPT1 were decreased. The concentration of
the Na+,K+-ATPase, which turned out to be the
most strongly expressed enterocyte transporter, was also decreased.
HFD also induced concentration changes of drug transporters and of
enzymes involved in drug metabolism, which suggests effects of HFD
on pharmacokinetics and toxicities. Finally, we observed down-regulation
of antibody subunits and of components of the major histocompatibility
complex II that may reflect impaired immune defense and immune tolerance
in HFD. Our work shows dramatic changes in functional proteins of
small intestine mucosa upon excessive fat consumption.

优化后的样品制备与蛋白质组学数据分析（proteomic data analysis）流程，使我们得以在小鼠小肠黏膜中鉴定出7700种不同蛋白质，并定量了超过5000种蛋白质的浓度。我们对饲喂时长为2个月的小鼠小肠黏膜蛋白质浓度开展了比较：两组小鼠分别接受正常膳食（Normal Diet, ND）与高脂膳食（High-Fat Diet, HFD）饲喂，其中正常膳食含34.4%碳水化合物、19.6%蛋白质与3.3%脂肪，高脂膳食含25.3%碳水化合物、24.1%蛋白质与34.6%脂肪。在完成定量的蛋白质中，有11%在ND组与HFD组间存在显著表达差异。高脂膳食饲喂后，我们观察到参与蛋白质合成、蛋白质N-糖基化及囊泡运输的蛋白质浓度升高；参与脂肪酸吸收、脂肪酸β-氧化与类固醇代谢的蛋白质丰度同样出现上调。糖酵解与磷酸戊糖循环相关酶的浓度则呈下调趋势，而呼吸链及ATP合酶（ATP synthase）相关蛋白质的浓度有所升高。位于肠上皮细胞质膜上的多种营养物质转运蛋白，包括Na+依赖性葡萄糖协同转运蛋白SGLT1、被动性葡萄糖转运蛋白GLUT2以及H+肽协同转运蛋白PEPT1，其浓度均出现下调。作为肠上皮细胞中表达量最高的转运蛋白，钠钾ATP酶（Na+,K+-ATPase）的浓度同样有所下降。高脂膳食还可诱导药物转运蛋白及药物代谢相关酶的浓度发生改变，这提示高脂膳食会对药物代谢动力学与毒性产生影响。最终，我们观察到抗体亚基与主要组织相容性复合体II（major histocompatibility complex II, MHC II）的组成成分出现表达下调，这或可反映高脂膳食状态下机体免疫防御与免疫耐受功能受损。本研究揭示了过量脂肪摄入后小肠黏膜功能蛋白质组发生的显著变化。