colonic bacterial metabolism.docx

1、colonic bacterial metabolismAm J Physiol Gastrointest Liver Physiol.2012 January;302(1): G1G9.Functional analysis of colonic bacterial metabolism: relevant to health?Henrike M. Hamer,Vicky De Preter,Karen Windey, andKristin VerbekeTranslational Research Center for Gastrointestinal Disorders and Leuv

2、en Food Science and Nutrition Research Center, University Hospital Gasthuisberg, Katholieke Universiteit Leuven, Leuven, BelgiumCorresponding author.Address for reprint requests and other correspondence: K. Verbeke, TARGID, Univ. Hospital Leuven, Herestraat 49, 3000 Leuven, Belgium (e-mail:kristin.v

3、erbekemed.kuleuven.be).Received February 7, 2011; Accepted October 16, 2011.Copyright 2012 the American Physiological SocietyAbstractWith the use of molecular techniques, numerous studies have evaluated the composition of the intestinal microbiota in health and disease. However, it is of major inter

4、est to supplement this with a functional analysis of the microbiota. In this review, the different approaches that have been used to characterize microbial metabolites, yielding information on the functional end products of microbial metabolism, have been summarized. To analyze colonic microbial met

5、abolites, the most conventional way is by application of a hypothesis-driven targeted approach, through quantification of selected metabolites from carbohydrate (e.g., short-chain fatty acids) and protein fermentation (e.g.,p-cresol, phenol, ammonia, or H2S), secondary bile acids, or colonic enzymes

6、. The application of stable isotope-labeled substrates can provide an elegant solution to study these metabolic pathways in vivo. On the other hand, a top-down approach can be followed by applying metabolite fingerprinting techniques based on1H-NMR or mass spectrometric analysis. Quantification of k

7、nown metabolites and characterization of metabolite patterns in urine, breath, plasma, and fecal samples can reveal new pathways and give insight into physiological regulatory processes of the colonic microbiota. In addition, specific metabolic profiles can function as a diagnostic tool for the iden

8、tification of several gastrointestinal diseases, such as ulcerative colitis and Crohns disease. Nevertheless, future research will have to evaluate the relevance of associations between metabolites and different disease states.Keywords:fermentation, metabolites, metabolomics, microbiota, short-chain

9、 fatty acidsthe human intestinal microbiotacomplements our physiology with functions that we have not had to develop on our own. In fact, the intestinal microbiota have a metabolic capacity that is comparable to that of the liver (83). The human colon contains an extremely complex and dynamic microb

10、ial ecosystem with high densities of living bacteria in concentrations of 1011-1012cells/g of luminal contents belonging to more than 1,000 different species. In healthy adults, 80% of the identified fecal microbiota can be classified into three dominant phyla: Firmicutes, Bacteroidetes, and Actinob

11、acteria, but there is substantial variation in the species composition between individuals (30,101).The intestinal microbiota plays an important role in human physiology. For example, the intestinal microbiota is responsible for the further metabolism and energy harvest from nondigested nutrients, i

12、s involved in the synthesis of vitamins such as B and K and metabolism of polyphenols, provides colonization resistance toward potential pathogens, is involved in the metabolism of bile acids, and stimulates the immune function of the host (74,83).Molecular approaches, mainly based on the 16S riboso

13、mal RNA gene, have revolutionized the field of gut microbial ecology. Nowadays, the uncultured and complex microbial communities can be characterized with greater sensitivity by using high-throughput technologies, such as pyrosequencing (5) and phylogenetic microarrays (79), compared with former mol

14、ecular fingerprinting methods, such as PCR-denaturing gradient gel electrophoresis (DGGE). Complementary quantitative technologies, such as fluorescence in situ hybridization (FISH) and real-time quantitative PCR, can be used to confirm shifts in particular groups or species (113).In recent years an

15、 increasing amount of literature has demonstrated that several diseases are related to alterations in the intestinal microbiota (known as dysbiosis), such as irritable bowel syndrome (54), inflammatory bowel disease (93), diabetes (55), atopic diseases (76), cancer (85), and obesity (7). For example

16、, a reduction in the abundance and diversity of Firmicutes is frequently associated with inflammatory bowel disease and irritable bowel syndrome (105,113). These studies have mainly shown that differences in the composition of the intestinal microbiota are associated with disease.Functional Capacity

17、 of the MicrobiotaThe human microbiota is characterized by a significant degree of functional redundancy, meaning that different bacteria can perform similar functions and metabolize the same substrate (60,64). Therefore, not only the composition but also the functional capacity of the intestinal mi

18、crobiota is highly important regarding the clinical end points.Next-generation pyrosequencing can be applied to further evaluate the functional capacity of the colonic microbiota by creating a catalog of the genetic potential of the microbiota. However, it has to be realized that the detection of ge

19、nes in a metagenomic library does not necessarily mean that these are functionally important (113). To gain better insight into the activity and functionality of the intestinal microbiota, other meta-“omics” approaches can be applied that use RNA, proteins, and metabolites as targets. In this review

20、, we summarize the different approaches that have been used to quantify and characterize the metabolites produced by the microbiota, which yield information on the actual end products of metabolism. Colonic microbial fermentation results in the production of large amounts of different end products.

21、The type and amount of these fermentation-derived metabolites largely depend on the composition of the microbiota, transit time, and the substrates available for fermentation (95). Some of these end products have been shown to be protective to the colonic epithelium, and others have proved to be pro

22、inflammatory or procarcinogenic metabolites (3,48). By using knowledge of these specific metabolites, a hypothesis-driven targeted approach can be applied to evaluate changes in colonic metabolism following dietary interventions or during different disease states, for example, through quantification

23、 of selected metabolites from carbohydrate and protein fermentation, secondary bile acids, or colonic enzymes. On the other hand, a top-down approach can be followed by applying metabolite fingerprinting techniques based on1H-nuclear magnetic resonance (NMR) or mass spectrometric analysis. By follow

24、ing this approach, novel metabolites and mechanisms can be identified that are involved in health and disease. This is, however, not an easy task, since the signals first have to be identified and their metabolic roles elucidated.For analyses of metabolites as end products of intestinal metabolism i

25、n humans, we mainly rely on fecal samples or on breath (for example, hydrogen, methane, and carbon dioxide), urine, and plasma samples due to the relative inaccessibility of the colon to sample at different locations (Fig. 1) (50). In these human studies, an elegant solution to study metabolic pathw

26、ays in vivo is the application of stable isotope tracers.Types of FermentationThe colonic microbiota ferment endogenous host-derived substrates such as mucus, pancreatic enzymes, and exfoliated epithelial cells, as well as dietary components that escape digestion in the upper gastrointestinal tract.

27、 Two main types of colonic microbial fermentation can be distinguished, including saccharolytic fermentation of carbohydrate and proteolytic fermentation of protein (Fig. 2). In the proximal part of the colon, mainly saccharolytic fermentation takes place, since most microorganisms preferentially fe

28、rment carbohydrates and switch to protein fermentation when carbohydrate sources are depleted (75). The main products of carbohydrate metabolism are short-chain fatty acids (SCFA), mainly acetate, propionate, and butyrate, which have been shown to contribute to colonic health. SCFA provide energy to

29、 the colonic epithelial cells, decrease luminal pH, and improve mineral absorption. Furthermore, butyrate has been shown to possess an anti-inflammatory and anticarcinogenic potential (46,63). Besides their contribution to gut health and maintenance, SCFA may provide further benefits for the systemi

30、c metabolism. For example, it has been shown that acetate and propionate affect hepatic lipid metabolism. Acetate is the primary substrate for cholesterol synthesis, whereas propionate can inhibit cholesterol synthesis (27,110). Since the different SCFA may show distinct effects, not only the amount

31、 of SCFA produced but also their ratio is of importance with regard to these health effects. Furthermore, SCFA stimulate increased plasma levels of satiety hormones such as peptide YY (PYY), leptin, and glucagon-like peptide-1 (GLP-1) and may attenuate insulin resistance (1,34). These effects may oc

32、cur due to the fact that SCFA are ligands for G protein-coupled receptors (GPR) 41 and 43, expressed on adipocytes, enteroendocrine L-cells, and immune cells (88). In addition, recent studies have linked SCFA activation of GPR 43 to the suppression of colon cancer (94,96). Proteolytic fermentation a

33、lso leads not only to the production of SCFA (lower amounts than produced from carbohydrates) and branched-chain fatty acids but also to potentially toxic metabolites such as phenolic compounds, sulfur-containing compounds, amines, and ammonia (Fig. 2) (48). The toxicity of these protein fermentation metabolites has mainly been es