When specific gut microbes reveal a possible link between hepatic steatosis and adipose tissue

Abstract

Gut-adipose tissue axis in hepatic fat accumulation in humansJournal of HepatologyVol. 61Issue 1PreviewRecent evidence suggests that in animals gut microbiota composition (GMC) affects the onset and progression of hepatic fat accumulation. The aim of this study was to investigate in humans whether subjects with high hepatic fat content (HHFC) differ in their GMC from those with low hepatic fat content (LHFC), and whether these differences are associated with body composition, biomarkers and abdominal adipose tissue inflammation. Full-Text PDF The gut microbiota is considered as a factor involved in the regulation of numerous metabolic pathways by impacting different functions of the host. Among these regulations, the influence of gut microbes on energy homeostasis is of particular interest because it has been suggested to be a driving force in the pathogenesis of metabolic diseases associated with obesity (such as insulin resistance, diabetes, cardiovascular disease, and non-alcoholic fatty liver disease (NAFLD)) [1Cani P.D. Metabolism in 2013: the gut microbiota manages host metabolism.Nat Rev Endocrinol. 2014; 10: 74-76Crossref PubMed Scopus (95) Google Scholar, 2Moschen A.R. Kaser S. Tilg H. Non-alcoholic steatohepatitis: a microbiota-driven disease.Trends Endocrinol Metab. 2013; 24: 537-545Abstract Full Text Full Text PDF PubMed Scopus (126) Google Scholar]. Intestinal microbes have developed a mutualistic relationship with their host and can influence physiological systems by modulating gut motility, intestinal barrier homeostasis, nutrient absorption, fat distribution, and liver fat accumulation [3Everard A. Cani P.D. Diabetes, obesity and gut microbiota.Best Pract Res Clin Anaesthesiol. 2013; 27: 73-83Scopus (377) Google Scholar, 4Zhao L. The gut microbiota and obesity: from correlation to causality.Nat Rev Microbiol. 2013; 11: 639-647Crossref PubMed Scopus (509) Google Scholar].The relationship between changes in gut microbiota and the development and progression of liver disease has been known for over fifty years. Endotoxemia and gut-derived toxins are suggested to have causative roles in the onset and progression of liver inflammation and damage in chronic liver diseases [[5]Nolan J.P. Leibowitz A.I. Endotoxins in liver disease.Gastroenterology. 1978; 75: 765-766PubMed Google Scholar]. Similar to the mechanisms underlying metabolic endotoxemia (i.e., increased blood lipopolysaccharides (LPS) levels) and inflammation described in our previous work [[6]Cani P.D. Possemiers S. Van de W.T. Guiot Y. Everard A. Rottier O. et al.Changes in gut microbiota control inflammation in obese mice through a mechanism involving GLP-2-driven improvement of gut permeability.Gut. 2009; 58: 1091-1103Crossref PubMed Scopus (1767) Google Scholar], intestinal bacterial overgrowth, gut leakiness and increased endotoxin absorption have all been associated with hepatic fat accumulation and inflammation (NAFLD and non-alcoholic steatohepatitis (NASH)) in both rodents and human patients [7Farhadi A. Gundlapalli S. Shaikh M. Frantzides C. Harrell L. Kwasny M.M. et al.Susceptibility to gut leakiness: a possible mechanism for endotoxaemia in non-alcoholic steatohepatitis.Liver Int. 2008; 28: 1026-1033Crossref PubMed Scopus (169) Google Scholar, 8Wigg A.J. Roberts-Thomson I.C. Dymock R.B. McCarthy P.J. Grose R.H. Cummins A.G. The role of small intestinal bacterial overgrowth, intestinal permeability, endotoxaemia, and tumour necrosis factor alpha in the pathogenesis of non-alcoholic steatohepatitis.Gut. 2001; 48: 206-211Crossref PubMed Scopus (682) Google Scholar, 9Zhu L. Baker S.S. Gill C. Liu W. Alkhouri R. Baker R.D. et al.Characterization of gut microbiomes in nonalcoholic steatohepatitis (NASH) patients: a connection between endogenous alcohol and NASH.Hepatology. 2013; 57: 601-609Crossref PubMed Scopus (955) Google Scholar]. However, the role of specific bacteria, metabolites coming from the gut microbiota or both remained to be demonstrated.In NAFLD patients changes in tight junction protein expression and distribution are suggested as critical factors in the impairment of gut barrier function and subsequent alterations in gut permeability [[10]Miele L. Valenza V. La T.G. Montalto M. Cammarota G. Ricci R. et al.Increased intestinal permeability and tight junction alterations in nonalcoholic fatty liver disease.Hepatology. 2009; 49: 1877-1887Crossref PubMed Scopus (951) Google Scholar]. As a consequence, metabolic endotoxemia exposes the liver to gut-derived toxins resulting in the release of numerous pro-inflammatory cytokines that ultimately lead to hepatic injury and fibrosis [[11]Seki E. Brenner D.A. Toll-like receptors and adaptor molecules in liver disease: update.Hepatology. 2008; 48: 322-335Crossref PubMed Scopus (550) Google Scholar]. However, although previous studies have shown that the treatment with antibiotics or loss of endotoxins receptors (Toll-like receptor-(TLR) 4) significantly attenuates the development of hepatic steatosis in mice [12Thuy S. Ladurner R. Volynets V. Wagner S. Strahl S. Konigsrainer A. et al.Nonalcoholic fatty liver disease in humans is associated with increased plasma endotoxin and plasminogen activator inhibitor 1 concentrations and with fructose intake.J Nutr. 2008; 138: 1452-1455PubMed Google Scholar, 13Cani P.D. Bibiloni R. Knauf C. Waget A. Neyrinck A.M. Delzenne N.M. et al.Changes in gut microbiota control metabolic endotoxemia-induced inflammation in high-fat diet-induced obesity and diabetes in mice.Diabetes. 2008; 57: 1470-1481Crossref PubMed Scopus (3174) Google Scholar], it has also been suggested that other TLRs than TLR4 contributed to the onset of NAFLD [[14]Wagnerberger S. Spruss A. Kanuri G. Volynets V. Stahl C. Bischoff S.C. et al.Toll-like receptors 1–9 are elevated in livers with fructose-induced hepatic steatosis.Br J Nutr. 2012; 107: 1727-1738Crossref PubMed Scopus (91) Google Scholar]. In light of these recent findings, understanding the role of specific microbes or microbial communities as well as identifying the cross-talks existing between organs and the gut microbiota related to metabolism regulation is of the utmost importance.The present study by Munukka et al. in this issue of the Journal of Hepatology provides a putative link between the abundance of one specific bug, namely Faecalibacterium prausnitzii, hepatic fat accumulation and adipose tissue inflammation. By using proton magnetic resonance spectroscopy (1H MRS) they classified their population comprising of 31 subjects into two distinct groups, that is high hepatic fat content (>5%, n = 10) and low hepatic fat content (<5%, n = 21). Munukka et al. also investigated whether hepatic fat content was associated with body composition, adipose tissue inflammation (by microarray) and specific gut microbes (by 16S rRNA hybridization and flow cytometry). The major limitations of this study rely on the fact that the number of patients was relatively small and more importantly because of the use of a targeted approach to investigate the gut microbiota. But, besides these points, the authors found that a lower abundance of F. prausnitzii is associated with hepatic fat accumulation. Moreover, this remained significant after adjustment for gender, age and weight, which reinforce a putative link between this bacterium and host metabolism.Conversely with previous reports [[15]Mouzaki M. Comelli E.M. Arendt B.M. Bonengel J. Fung S.K. Fischer S.E. et al.Intestinal microbiota in patients with nonalcoholic fatty liver disease.Hepatology. 2013; 58: 120-127Crossref PubMed Scopus (485) Google Scholar], they found that the abundance of the Bacteroides group was positively correlated with insulin resistance index (HOMA-IR). When they combined the data obtained for F. prausnitzii and Bacteroides as a ratio (F. prausnitzii/Bacteroides), the authors found that the level of several genes expressed in the subcutaneous adipose depots and related with inflammation were negatively associated with this ratio. Interestingly, no significant differences were found at the level of several genes involved in lipid metabolism. It is worth noting that in the present study the authors had access to subcutaneous adipose tissues which is likely less prone to directly induce an overflow of the liver with fatty acids, and this in comparison with the visceral adipose-derived fatty acids. Thus, given that in this study the patients with a higher liver fat accumulation exhibited more visceral fat accumulation (2-fold more) it is likely that both type of fat depots contribute to the development of hepatic fat accumulation via complementary mechanisms, that are directly or indirectly associated with the gut microbiota. Nevertheless, in this study Munukka et al. unequivocally found that the increased inflammatory tone observed in the subcutaneous adipose tissue was correlated with the presence of F. prausnitzii and Bacteroides and eventually suggest a link between the gut, subcutaneous adipose tissue, and liver fat accumulation.Although this direct link was not shown in the present study, it has been previously demonstrated that higher abundance of F. prausnitzii was associated with an improved inflammatory status in obese patients [[16]Furet J.P. Kong L.C. Tap J. Poitou C. Basdevant A. Bouillot J.L. et al.Differential adaptation of human gut microbiota to bariatric surgery-induced weight loss: links with metabolic and low-grade inflammation markers.Diabetes. 2010; 59: 3049-3057Crossref PubMed Scopus (857) Google Scholar]. Moreover, a reduction in a cluster of genes belonging to F. prausnitzii was identified as a discriminant marker for the prediction of diabetic status in European women [[17]Karlsson F.H. Tremaroli V. Nookaew I. Bergstrom G. Behre C.J. Fagerberg B. et al.Gut metagenome in European women with normal, impaired and diabetic glucose control.Nature. 2013; 498: 99-103Crossref PubMed Scopus (1756) Google Scholar], which may support the link with F. prausnitzii observed in the present study and HOMA-IR. Moreover, rodents studies have clearly associated the role of F. prausnitzii with intestinal barrier function [[18]Martin R. Chain F. Miquel S. Lu J. Gratadoux J.J. Sokol H. et al.The commensal bacterium Faecalibacterium prausnitzii is protective in DNBS-induced chronic moderate and severe colitis models.Inflamm Bowel Dis. 2014; 20: 417-430Crossref PubMed Scopus (148) Google Scholar].Strikingly, the decreased abundance of several genera including Bacteroides (which was decreased in the present study) and F. prausnitzii has also been found in type 2 diabetic Chinese subjects [[19]Zhang X. Shen D. Fang Z. Jie Z. Qiu X. Zhang C. et al.Human gut microbiota changes reveal the progression of glucose intolerance.PLoS One. 2013; 8: e71108Crossref PubMed Scopus (508) Google Scholar]. Moreover, Balamurugan et al. found that F. prausnitzii was increased in obese Indian children compared to the lean controls [[20]Balamurugan R. George G. Kabeerdoss J. Hepsiba J. Chandragunasekaran A.M. Ramakrishna B.S. Quantitative differences in intestinal Faecalibacterium prausnitzii in obese Indian children.Br J Nutr. 2010; 103: 335-338Crossref PubMed Scopus (140) Google Scholar]. Thus these discrepancies strongly highlight once again that the specificity of the population, age, and diets in phenotype and taxonomy associations have to be taken into account before drawing any clear conclusions on the role of one specific bacterium on the onset or the protection against metabolic disorders associated with fat accumulation.Growing evidence suggests that cross-talks between gut microbes and the host are achieved through specific metabolites such as for example short chain fatty acids or specific molecular patterns of microbial membranes (e.g., LPS) that may contribute to the activation of TLR’s.In this study, Munukka et al. found a positive association between Enterobacteriaceae family (i.e., Gram negative bacteria) and triglycerides, but they did not find any differences in metabolic endotoxemia measured in the peripheral blood of both groups. Besides the fact that measuring blood LPS is highly tricky, and may be an important confounding factor here, previous studies have clearly associated plasma LPS levels with triglycerides [[21]Lassenius M.I. Pietilainen K.H. Kaartinen K. Pussinen P.J. Syrjanen J. Forsblom C. et al.Bacterial endotoxin activity in human serum is associated with dyslipidemia, insulin resistance, obesity, and chronic inflammation.Diabetes Care. 2011; 34: 1809-1815Crossref PubMed Scopus (275) Google Scholar].Thus, in this study, we may not exclude that numerous parameters may have contributed to the regulation of hepatic fat accumulation. Is the phenotype starting from the gut barrier dysfunction and eventually linked with the presence of LPS or any other molecular patterns of microbial membranes? Is the higher inflammatory tone observed in the adipose tissue directly or indirectly associated with any modification of the gut barrier function, or changes in the gut microbiome or both? What is the abundance of other gut bacteria measured by using high-throughput methods (e.g., sequencing)? All these questions remain still unanswered, and merit further investigations.Conflict of interestThe author declare that he not have anything to disclose regarding funding or conflict of interest with respect to this manuscript. The gut microbiota is considered as a factor involved in the regulation of numerous metabolic pathways by impacting different functions of the host. Among these regulations, the influence of gut microbes on energy homeostasis is of particular interest because it has been suggested to be a driving force in the pathogenesis of metabolic diseases associated with obesity (such as insulin resistance, diabetes, cardiovascular disease, and non-alcoholic fatty liver disease (NAFLD)) [1Cani P.D. Metabolism in 2013: the gut microbiota manages host metabolism.Nat Rev Endocrinol. 2014; 10: 74-76Crossref PubMed Scopus (95) Google Scholar, 2Moschen A.R. Kaser S. Tilg H. Non-alcoholic steatohepatitis: a microbiota-driven disease.Trends Endocrinol Metab. 2013; 24: 537-545Abstract Full Text Full Text PDF PubMed Scopus (126) Google Scholar]. Intestinal microbes have developed a mutualistic relationship with their host and can influence physiological systems by modulating gut motility, intestinal barrier homeostasis, nutrient absorption, fat distribution, and liver fat accumulation [3Everard A. Cani P.D. Diabetes, obesity and gut microbiota.Best Pract Res Clin Anaesthesiol. 2013; 27: 73-83Scopus (377) Google Scholar, 4Zhao L. The gut microbiota and obesity: from correlation to causality.Nat Rev Microbiol. 2013; 11: 639-647Crossref PubMed Scopus (509) Google Scholar]. The relationship between changes in gut microbiota and the development and progression of liver disease has been known for over fifty years. Endotoxemia and gut-derived toxins are suggested to have causative roles in the onset and progression of liver inflammation and damage in chronic liver diseases [[5]Nolan J.P. Leibowitz A.I. Endotoxins in liver disease.Gastroenterology. 1978; 75: 765-766PubMed Google Scholar]. Similar to the mechanisms underlying metabolic endotoxemia (i.e., increased blood lipopolysaccharides (LPS) levels) and inflammation described in our previous work [[6]Cani P.D. Possemiers S. Van de W.T. Guiot Y. Everard A. Rottier O. et al.Changes in gut microbiota control inflammation in obese mice through a mechanism involving GLP-2-driven improvement of gut permeability.Gut. 2009; 58: 1091-1103Crossref PubMed Scopus (1767) Google Scholar], intestinal bacterial overgrowth, gut leakiness and increased endotoxin absorption have all been associated with hepatic fat accumulation and inflammation (NAFLD and non-alcoholic steatohepatitis (NASH)) in both rodents and human patients [7Farhadi A. Gundlapalli S. Shaikh M. Frantzides C. Harrell L. Kwasny M.M. et al.Susceptibility to gut leakiness: a possible mechanism for endotoxaemia in non-alcoholic steatohepatitis.Liver Int. 2008; 28: 1026-1033Crossref PubMed Scopus (169) Google Scholar, 8Wigg A.J. Roberts-Thomson I.C. Dymock R.B. McCarthy P.J. Grose R.H. Cummins A.G. The role of small intestinal bacterial overgrowth, intestinal permeability, endotoxaemia, and tumour necrosis factor alpha in the pathogenesis of non-alcoholic steatohepatitis.Gut. 2001; 48: 206-211Crossref PubMed Scopus (682) Google Scholar, 9Zhu L. Baker S.S. Gill C. Liu W. Alkhouri R. Baker R.D. et al.Characterization of gut microbiomes in nonalcoholic steatohepatitis (NASH) patients: a connection between endogenous alcohol and NASH.Hepatology. 2013; 57: 601-609Crossref PubMed Scopus (955) Google Scholar]. However, the role of specific bacteria, metabolites coming from the gut microbiota or both remained to be demonstrated. In NAFLD patients changes in tight junction protein expression and distribution are suggested as critical factors in the impairment of gut barrier function and subsequent alterations in gut permeability [[10]Miele L. Valenza V. La T.G. Montalto M. Cammarota G. Ricci R. et al.Increased intestinal permeability and tight junction alterations in nonalcoholic fatty liver disease.Hepatology. 2009; 49: 1877-1887Crossref PubMed Scopus (951) Google Scholar]. As a consequence, metabolic endotoxemia exposes the liver to gut-derived toxins resulting in the release of numerous pro-inflammatory cytokines that ultimately lead to hepatic injury and fibrosis [[11]Seki E. Brenner D.A. Toll-like receptors and adaptor molecules in liver disease: update.Hepatology. 2008; 48: 322-335Crossref PubMed Scopus (550) Google Scholar]. However, although previous studies have shown that the treatment with antibiotics or loss of endotoxins receptors (Toll-like receptor-(TLR) 4) significantly attenuates the development of hepatic steatosis in mice [12Thuy S. Ladurner R. Volynets V. Wagner S. Strahl S. Konigsrainer A. et al.Nonalcoholic fatty liver disease in humans is associated with increased plasma endotoxin and plasminogen activator inhibitor 1 concentrations and with fructose intake.J Nutr. 2008; 138: 1452-1455PubMed Google Scholar, 13Cani P.D. Bibiloni R. Knauf C. Waget A. Neyrinck A.M. Delzenne N.M. et al.Changes in gut microbiota control metabolic endotoxemia-induced inflammation in high-fat diet-induced obesity and diabetes in mice.Diabetes. 2008; 57: 1470-1481Crossref PubMed Scopus (3174) Google Scholar], it has also been suggested that other TLRs than TLR4 contributed to the onset of NAFLD [[14]Wagnerberger S. Spruss A. Kanuri G. Volynets V. Stahl C. Bischoff S.C. et al.Toll-like receptors 1–9 are elevated in livers with fructose-induced hepatic steatosis.Br J Nutr. 2012; 107: 1727-1738Crossref PubMed Scopus (91) Google Scholar]. In light of these recent findings, understanding the role of specific microbes or microbial communities as well as identifying the cross-talks existing between organs and the gut microbiota related to metabolism regulation is of the utmost importance. The present study by Munukka et al. in this issue of the Journal of Hepatology provides a putative link between the abundance of one specific bug, namely Faecalibacterium prausnitzii, hepatic fat accumulation and adipose tissue inflammation. By using proton magnetic resonance spectroscopy (1H MRS) they classified their population comprising of 31 subjects into two distinct groups, that is high hepatic fat content (>5%, n = 10) and low hepatic fat content (<5%, n = 21). Munukka et al. also investigated whether hepatic fat content was associated with body composition, adipose tissue inflammation (by microarray) and specific gut microbes (by 16S rRNA hybridization and flow cytometry). The major limitations of this study rely on the fact that the number of patients was relatively small and more importantly because of the use of a targeted approach to investigate the gut microbiota. But, besides these points, the authors found that a lower abundance of F. prausnitzii is associated with hepatic fat accumulation. Moreover, this remained significant after adjustment for gender, age and weight, which reinforce a putative link between this bacterium and host metabolism. Conversely with previous reports [[15]Mouzaki M. Comelli E.M. Arendt B.M. Bonengel J. Fung S.K. Fischer S.E. et al.Intestinal microbiota in patients with nonalcoholic fatty liver disease.Hepatology. 2013; 58: 120-127Crossref PubMed Scopus (485) Google Scholar], they found that the abundance of the Bacteroides group was positively correlated with insulin resistance index (HOMA-IR). When they combined the data obtained for F. prausnitzii and Bacteroides as a ratio (F. prausnitzii/Bacteroides), the authors found that the level of several genes expressed in the subcutaneous adipose depots and related with inflammation were negatively associated with this ratio. Interestingly, no significant differences were found at the level of several genes involved in lipid metabolism. It is worth noting that in the present study the authors had access to subcutaneous adipose tissues which is likely less prone to directly induce an overflow of the liver with fatty acids, and this in comparison with the visceral adipose-derived fatty acids. Thus, given that in this study the patients with a higher liver fat accumulation exhibited more visceral fat accumulation (2-fold more) it is likely that both type of fat depots contribute to the development of hepatic fat accumulation via complementary mechanisms, that are directly or indirectly associated with the gut microbiota. Nevertheless, in this study Munukka et al. unequivocally found that the increased inflammatory tone observed in the subcutaneous adipose tissue was correlated with the presence of F. prausnitzii and Bacteroides and eventually suggest a link between the gut, subcutaneous adipose tissue, and liver fat accumulation. Although this direct link was not shown in the present study, it has been previously demonstrated that higher abundance of F. prausnitzii was associated with an improved inflammatory status in obese patients [[16]Furet J.P. Kong L.C. Tap J. Poitou C. Basdevant A. Bouillot J.L. et al.Differential adaptation of human gut microbiota to bariatric surgery-induced weight loss: links with metabolic and low-grade inflammation markers.Diabetes. 2010; 59: 3049-3057Crossref PubMed Scopus (857) Google Scholar]. Moreover, a reduction in a cluster of genes belonging to F. prausnitzii was identified as a discriminant marker for the prediction of diabetic status in European women [[17]Karlsson F.H. Tremaroli V. Nookaew I. Bergstrom G. Behre C.J. Fagerberg B. et al.Gut metagenome in European women with normal, impaired and diabetic glucose control.Nature. 2013; 498: 99-103Crossref PubMed Scopus (1756) Google Scholar], which may support the link with F. prausnitzii observed in the present study and HOMA-IR. Moreover, rodents studies have clearly associated the role of F. prausnitzii with intestinal barrier function [[18]Martin R. Chain F. Miquel S. Lu J. Gratadoux J.J. Sokol H. et al.The commensal bacterium Faecalibacterium prausnitzii is protective in DNBS-induced chronic moderate and severe colitis models.Inflamm Bowel Dis. 2014; 20: 417-430Crossref PubMed Scopus (148) Google Scholar]. Strikingly, the decreased abundance of several genera including Bacteroides (which was decreased in the present study) and F. prausnitzii has also been found in type 2 diabetic Chinese subjects [[19]Zhang X. Shen D. Fang Z. Jie Z. Qiu X. Zhang C. et al.Human gut microbiota changes reveal the progression of glucose intolerance.PLoS One. 2013; 8: e71108Crossref PubMed Scopus (508) Google Scholar]. Moreover, Balamurugan et al. found that F. prausnitzii was increased in obese Indian children compared to the lean controls [[20]Balamurugan R. George G. Kabeerdoss J. Hepsiba J. Chandragunasekaran A.M. Ramakrishna B.S. Quantitative differences in intestinal Faecalibacterium prausnitzii in obese Indian children.Br J Nutr. 2010; 103: 335-338Crossref PubMed Scopus (140) Google Scholar]. Thus these discrepancies strongly highlight once again that the specificity of the population, age, and diets in phenotype and taxonomy associations have to be taken into account before drawing any clear conclusions on the role of one specific bacterium on the onset or the protection against metabolic disorders associated with fat accumulation. Growing evidence suggests that cross-talks between gut microbes and the host are achieved through specific metabolites such as for example short chain fatty acids or specific molecular patterns of microbial membranes (e.g., LPS) that may contribute to the activation of TLR’s. In this study, Munukka et al. found a positive association between Enterobacteriaceae family (i.e., Gram negative bacteria) and triglycerides, but they did not find any differences in metabolic endotoxemia measured in the peripheral blood of both groups. Besides the fact that measuring blood LPS is highly tricky, and may be an important confounding factor here, previous studies have clearly associated plasma LPS levels with triglycerides [[21]Lassenius M.I. Pietilainen K.H. Kaartinen K. Pussinen P.J. Syrjanen J. Forsblom C. et al.Bacterial endotoxin activity in human serum is associated with dyslipidemia, insulin resistance, obesity, and chronic inflammation.Diabetes Care. 2011; 34: 1809-1815Crossref PubMed Scopus (275) Google Scholar]. Thus, in this study, we may not exclude that numerous parameters may have contributed to the regulation of hepatic fat accumulation. Is the phenotype starting from the gut barrier dysfunction and eventually linked with the presence of LPS or any other molecular patterns of microbial membranes? Is the higher inflammatory tone observed in the adipose tissue directly or indirectly associated with any modification of the gut barrier function, or changes in the gut microbiome or both? What is the abundance of other gut bacteria measured by using high-throughput methods (e.g., sequencing)? All these questions remain still unanswered, and merit further investigations. Conflict of interestThe author declare that he not have anything to disclose regarding funding or conflict of interest with respect to this manuscript. The author declare that he not have anything to disclose regarding funding or conflict of interest with respect to this manuscript. P. D. Cani is a research associate from the FRS-FNRS (Fonds de la Recherche Scientifique) and a recipient of grants from the FNRS and PDR (Projet de recherche, Belgium), from ARC (Actions de Recherche Concertées – Communauté française de Belgique convention :12/17-047). Moreover, P. D. Cani is a recipient of the ERC Starting Grant 2013 (European Research Council, Starting grant 336452-ENIGMO).