Milk fat globule membrane supplementation modulates the gut microbiota and attenuates metabolic endotoxemia in high-fat diet-fed mice
Graphical abstract
MFGM improved the gut microbiota dysbiosis induced by high-fat diet in C57BL/6J mice. MFGM also alleviated high-fat diet-induced intestinal inflammation, metabolic endotoxemia and systemic inflammation by decreasing the levels of toll-like receptor 4 (TLR4), pro-inflammatory cytokine release, lipopolysaccharides (LPS) and the expression of tight junction proteins including zonulin-1 (ZO-1) and occludin. These findings demonstrated that MFGM supplementation ameliorated obesity-related inflammation and endotoxemia partly via modulating the composition of gut microbiota in mice challenged with a high-fat diet. Red solid line arrows represented changes in response to HFD; Green solid line arrows represented changes in HFD-fed C57BL/6J mice receiving MFGM.
Introduction
Obesity is a serious and alarming global health problem closely linked to diet and lifestyle factors, as it contributes to a higher risk of developing other metabolic disorders including non-alcoholic fatty liver disease (NAFLD), type 2 diabetes, hypertension and certain cancers (Cani et al., 2008). Gut microbiota dysbiosis has been shown to be associated with the development of obesity and its related metabolic disorders (Kim, Gu, Lee, Joh & Kim, 2012). About 90% of the normal gut microbiota consists of Bacteroidetes and Firmicutes phylum in human subjects and rodents. Obesity leads to the decrease in the ratio of Bacteroidetes to Firmicutes phylum, which can lead to dysbiosis (Gil-Cardoso et al., 2016). Strikingly, an obese phenotype has been successfully induced by transferring gut microbes from obese mice to lean gnotobiotic ones, which strengthens the crucial role of gut microbiota in energy homeostasis and metabolic state (Turnbaugh et al., 2006). Therefore, the modulation of gut microbiota dysbiosis induced by high-fat diet should be targeted as an effective intervention approach to prevent obesity and its related chronic metabolic disorder (Wang et al., 2015).
Low-grade chronic inflammation is a common characteristic for obesity and associated metabolic disorders, despite the fact that the molecular mechanism of the inflammation is still unclear (Esser, Legrand-Poels, Piette, Scheen & Paquot, 2014). Lipopolysaccharides (LPS), major components of the external cell wall of Gram-negative bacteria, elicit strong immune responses as endotoxins (Zhou et al., 2014). The presence of LPS at a normal range in the intestinal lumen does not cause deleterious effects under normal conditions (Geurts, Neyrinck, Delzenne, Knauf & Cani, 2014). However, increased LPS are transferred into circulatory system under high-fat diet because of gut microbiota dysbiosis and related gut barrier impairment, which results in metabolic endotoxemia (Moreira, Texeira, Ferreira, Do Carmo Gouveia Peluzio & de Cássia Gonçalves Alfenas, 2012). The elevated bacterial LPS levels cause inflammation mainly by interacting with toll-like receptor 4 (TLR4) (de La Serre et al., 2010). Recognition of LPS by TLR4 of host cells increases the expression of downstream pro-inflammatory factors including IL-6 and TNF-α, thus triggering inflammatory events that contributes to the development of obesity and metabolic disorders (Cani et al., 2007). In addition, gastrointestinal tract, acting as the first line to defend the ingested toxic substance, prevents bacteria and pathogens from entering circulatory circulation (Groschwitz & Hogan, 2009). Intercellular tight junctions, which has as a vital role for maintaining gut barrier integrity, consist of transmembrane proteins (occludin, claudins, and junctional adhesion molecule (JAM)), junctional complex proteins (such as ZO-1, symplekin, and cingulin), and actin cytoskeleton (Suzuki, 2013). Gut barrier integrity can be damaged by obesity, being most obvious in obese rodent models (Cani et al., 2009). The activation of TLR4 also alters tight junctions and increases intestinal permeability, leading to intestinal inflammation (Guo et al., 2015). Therefore, prevention of gut microbiota dysbiosis and maintaining gut epithelial barrier function are critical for treating metabolic disorders and endotoxemia associated with obesity.
In recent years, increasing attention is attached to bioactive compounds derived from milk as components of functional foods in pharmaceutical and food industries. Milk fat globule membrane (MFGM), derived from the membrane of the mammary epithelial cells, is a tri-layer rich in polar lipids, membrane-specific proteins, glycoproteins, enzymes and vitamins, resulting in health-promoting functions (Baars et al., 2016). Lactadherin, one of the major protein components associated with MFGM, has been demonstrated to be capable of binding and neutralising pathogenic bacteria, which attenuated intestinal inflammation and maintained the homeostasis of intestinal epithelium (Bu et al., 2007). The polar lipids from MFGM displayed bactericidal activity against enterotoxigenic pathogens and reduced intestinal inflammation (Sánchez-Juanes, Alonso, Zancada & Hueso, 2009). MFGM addition reduced the postprandial insulinaemic and inflammatory response in overweight and obese individuals induced by high-saturated fat diet (Demmer et al., 2016). MFGM supplementation in vivo reduced pathogen colonization and translocation by inhibiting mucosal pathogen adherence (Sprong, Hulstein, Lambers & van der Meer, 2012). MFGM-rich milk fat diet significantly reduced systemic inflammation and gut barrier disruption in LPS-induced mice (Snow, Ward, Olsen, Jimenez-Flores & Hintze, 2011). Recently, MFGM supplementation in infant formula was reported to promote intestinal epithelial cell proliferation and tight junction protein patterns by modulating the neonatal gut microbiome (Bhinder et al., 2017). However, it is still unclear whether MFGM addition would regulate gut microbiota and alleviate inflammatory status against obesity induced by high-fat diet.
In this study, the effects of MFGM on the distribution profile of gut microbiota of high-fat diet-fed mice were investigated. Moreover, the impact of MFGM against endotoxin was also assessed to elucidate the possible mechanisms by which MFGM may exert its protective effects against obesity associated metabolic disorders.
Section snippets
Materials
Lacprodan® MFGM-10 was provided by Arla Co. (Sønderhøj, Denmark). Antibodies against TLR4, ZO-1, occludin and β-actin were purchased from Bioss Inc. (Bios, Beijing, China)
Animals and treatments
Male C57BL/6J mice were purchased from Beijing Vital River Laboratory Animal Technology Co., Ltd (Beijing, China) at the age of 5 weeks. The animals were housed in a controlled environment (12 h light-dark cycle). After 7 days of acclimation, mice were randomly divided into two groups. The control group (ND group) contained 10
MFGM modulated the composition of the gut microbiota in high-fat diet-fed mice
The overall composition of the bacterial community was influenced by diet (Fig. 1), with significant alterations among ND, HFD and MFGM group. As shown in Fig. 1A and B, at the phylum level, the gut microbiota of HFD group showed decreases in the relative abundance of Bacteroidetes and the Bacteroidetes/Firmicutes ratio compared with the ND group (p < 0.05). However, MFGM increased the relative abundance of Bacteroidetes and up-regulated the Bacteroidetes/Firmicutes ratio compared with the HFD
Discussion
Gut microbiota plays a role in obesity and related metabolic disorders, while many bioactive compounds from diet have a significant impact on its composition and could be useful tools against obesity (Dong et al., 2016, Jiang et al., 2016). In this study, MFGM supplementation of high-fat diet-mice favors an interplay between microbiota and host metabolism by modulating the composition of gut microbiota, increasing the expression of tight junction proteins and lowering the levels of serum
Conclusions
In conclusion, this study demonstrated that MFGM intake could modulate the gut microbiota composition of high-fat diet fed mice. Concomitantly, MFGM alleviated high-fat diet-induced colon inflammation, systematic inflammation and metabolic endotoxemia. In addition, the expression of tight junction proteins was increased with MFGM supplementation. These findings indicated that MFGM might play an important role against obesity-related metabolic and inflammatory disorders. The current results may
Conflicts of interest
The authors declare no conflict of interest.
Ethics statement
The animal protocols (Approval No. KY160018) were approved by the Animal Experimentation Committee of China Agricultural University (Beijing, China). All mouse experimental procedures were performed in accordance with the Regulations for the Administration of Affairs Concerning Experimental Animals approved by the State Council of People’s Republic of China.
Acknowledgments
This work was supported by the National Natural Science Foundation of China (Grant No. 31371753), and the National Dairy Industry Technology System-Beijing Innovation Team (NDITS-BIT).
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