The Physiological Relevance of the Intestinal Microbiota ...

Review

The Physiological Relevance of the Intestinal Microbiota - Contributions to Human Health Kelly A. Tappenden, Ph.D, RD, and Andrew S. Deutsch, BS Division of Nutritional Sciences and Department of Food Science and Human Nutrition, University of Illinois at Urbana-Champaign, Urbana, Illinois The intestinal commensal microbiota is a dynamic mixture of essential microbes that develops under key influences of genetics, environment, diet and disease. Population profiles differ along the gastrointestinal tract, from the lumen to the mucosa, and among individuals. The total microbiota population outnumbers the cells in the human body and accounts for 35–50% of the volume of the colonic content. Key physiological functions of the commensal microbiota include protective effects exerted directly by specific bacterial species, control of epithelial cell proliferation and differentiation, production of essential mucosal nutrients, such as short-chain fatty acids and amino acids, prevention of overgrowth of pathogenic organisms, and stimulation of intestinal immunity. Oral probiotics are living microorganisms that upon ingestion in specific numbers exert health benefits beyond those of inherent basic nutrition. Emerging evidence indicates prophylactic and therapeutic utility for probiotic consumption in gastrointestinal health and disease.

INTRODUCTION

bad breath/halitosis, rosacea, acne, fatigue, irritability, anorexia and/or bulimia, stuffy nose, increased mucus production or symptoms of PMS, perimenopause, or menopause, worsening sensitivity to sugar and fermented products and worsening symptoms of inflammatory conditions, like asthma”. These marketing materials imply that such claims are confirmed by legitimate research, doctors, and patient testimonials; however transparent scientific support for these broad indications is not apparent. The title of this supplement is ‘The Host-Microbiota Connection - Is It Important Beyond the Intestinal Lumen?’ and the series of papers contained herein strive to provide: 1.) the physiological basis for the intestinal microbiota including important implications for intestinal health and disease; 2.) evidence supporting ingestion of probiotics preparations during various gastrointestinal ailments; 3.) insight into new experimental models highlighting the therapeutic utility and potential mechanisms underlying probiotic consumption, and; 4.) information on how probiotic therapy can impact immune function and a growing prominence in preventative medical nutrition therapy. The purpose of this initial paper is to review the physiological significance of the commensal microbiota that paves the way for contemplating research investigating the

Probiotics have been ingested by humans for hundreds of years, however clear scientific evidence supporting the benefit for their consumption has only begun to emerge recently. Common food sources of probiotic bacteria include yogurt and cheese, however various over the counter preparations are widely available. Unfortunately, some facets of the supplement industry have marketed certain preparations as “miracle drugs” and elixirs to cure all ills. The internet perpetuates these myths by indiscriminately crediting probiotics with such traits as “managing lactose intolerance, preventing colon cancer and infection, lowering cholesterol and blood pressure, improving immune function and mineral absorption, reducing inflammation, and preventing harmful bacteria growth under stress”. One company selling the “Friendly Colonizer”, a capsule containing six probiotic strains of unknown quantity, tells customers to expect these widespread positive results. Other marketing materials promote use if “you’ve been sick or taken antibiotics recently or have symptoms such as GI sensitivity (cramps, diarrhea/constipation), bloating or foul-smelling gas, including IBS or partially-digested stools, yeast infections, thrush, cold sores, diaper rash, headaches, migraines, joint aches, chronic

Address reprint requests to: Kelly A. Tappenden, Ph.D., R.D., 443 Bevier Hall, 905 South Goodwin Avenue, Department of Food Science and Human Nutrition, University of Illinois at Urbana-Champaign, Urbana, IL 61801. E-mail: [email protected] Disclosures: Presented at American College of Nutrition 47th Annual Meeting, Reno, Nevada, October 2006.

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Intestinal Microbiota and Human Health impact of altering the abundance and composition of this resident microbial ecosystem on human health.

THE GASTROINTESTINAL MICROBIOTA - FACTS FAR MORE THAN TRIVIA The sheer magnitude of the intestinal microbiota demands consideration for its role in human health and disease. The human gastrointestinal tract contains approximately ten times more prokaryotic cells than the total number of eukaryotic cells within the human body, equating to 1010–1011 CFU per gram of luminal contents at its peak within the colon, and up to half the volume of feces [1]. However, it is the dynamic role of this complex microbial community, composed primarily of bacteria but also fungi and protozoa, that warrants our attention regarding human health and disease. The commensal microbiota in the adult human is composed of greater than 500 species. The precise number varies among individuals and is somewhat unclear due to methodological limitations imposed by sampling limitations and standard culturing techniques that predated the development of molecular techniques in the field of microbiology [2]. The colonization process of the gastrointestinal tracts begins at birth and is highly impacted by the route of that process (vaginal versus cesarian), the method of feeding (human milk versus formula), genetics, environment, diet and disease. The microbial ecosystem varies among individuals, just as it does along the length of the gastrointestinal tract (Table 1), and even between the lumen and epithelium at any distinct gastrointestinal location.

PHYSIOLOGICAL FUNCTIONS OF THE COMMENSAL MICROBIOTA Production of Essential Mucosal Nutrients The significance of the commensal microbiota in salvaging energy and producing vitamins has been clearly demonstrated in animals reared in a sterile environment that prevents colonization of the gastrointestinal tract. Compared to conventional rodents, germ-free rodents consuming standard chow require 30% more energy in their diet, and supplementation with vitamin K and various B vitamins are mandatory to support growth and development. In the human, the liberation of energy by the commensal microbiota is estimated to be approximately 10% of that absorbed, but is highly dependent upon the diet consumed [3]. The ability of the commensal microbiota to salvage nutrients entering the colon is particularly relevant in both healthy and malabsorptive states. Patients with intestinal failure require

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Table 1. Abundance of Numerically Dominant Microbial Genera Along the Length of the Human Gastrointestinal Tract Esophagus Stomach

No own microbiota 104 CFU/g contents

Duodenum

103–104 CFU/g contents

Jejunum


Ileum


Colon


Candida albincans Helicobacter pylori Lactobacillus Streptococcus Bacteroides Candida albicans Lactobacillus Streptococcus Bacteroides Candida albincans Lactobacillus Streptococcus Bacteroides Clostridium Enterobacteriae Enterococcus Lactobacillus Veillonella Bacteroides Bacillus Bifidobacterium Clostridium Enterococcus Eubacterium Fusobacterium Peptostreptococcus Ruminococcus Streptococcus

parenteral nutrition due to malabsorption of consumed nutrients. Nordgaard and colleagues [4] demonstrated that when a functional colon is present, energy absorption can be maximized by consumption of a diet high in carbohydrate rather than fat. The microbiota contained within the colon of these patients served a critical function of fermenting the malabsorbed carbohydrate to short-chain fatty acids, which are subsequently absorbed and supply energy. Even in individuals with an intact and normally functioning gastrointestinal tract, the commensal microbiota provide an important function by fermenting dietary fiber into usable short-chain fatty acids, thereby salvaging important nutrients that would be lost in the feces due to the inability of the human intestine to digest dietary fiber. The relevance of short-chain fatty acid production extends beyond the energy that is salvaged by the microbial fermentation of malabsorbed carbohydrate. There is a growing body of evidence revealing a trophic role for SCFAs in the gastrointestinal tract. Intestinal adaptation following resection is increased with the consumption of fermentable dietary fiber [5]. Furthermore, total parenteral nutrition supplemented with short-chain fatty acids significantly reduces the ileal mucosa atrophy associated with TPN and enhances adaptive markers following small bowel resection [6,7]. Work from our laboratory has

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Intestinal Microbiota and Human Health determined that the supplementation of parenteral nutrition with SCFA enhanced structural and functional adaptation in both adult rats [6,8] and neonatal piglets [9] following massive small bowel resection. It appears that butyrate is the SCFA responsible for augmenting structural aspects of intestinal adaptations by increasing proliferation and decreasing apoptosis as early as 4 h post-resection [9]. Further, it is not clear if butyrate mediates these responses directly or a potential mechanism(s) relating to induced expression of the intestinotrophic peptide, glucagon-like peptide-2 (GLP-2). Despite the hypothesis yet to be tested regarding these novel nutrients, the fermentation of dietary carbohydrate to short-chain fatty acids represents an important physiological function served by the commensal microbiota.

Regulation of Intestinal Structure The structure of the gastrointestinal tract varies along its length and is a critical attribute that supports the digestive, absorptive and immune function within each segment. The surface area for these important functions are increased by 3-, 30- and 600-fold by the Folds of Kerckring, cryptvillus axis, and brush border membrane, respectively, over that provided by length of this cylindrical organ alone [10]. Importantly, the commensal microbiota appears to be very important for maintaining the cell proliferation, differentiation and function necessary for maintaining these critical structural attributes [11]. Beyond the effect of trophic nutrients that is discussed in the previous section, the intestinal microbiota has a direct impact on the structure of the intestine. Compared to conventional animals, germ-free animals have altered patterns of stem cell commitment to the various epithelial cell types, which alters both the timing of differentiation and relative abundance of cell types expressed within the intestinal epithelium [11]. Members of the commensal microbiota also degrade the mucous glycoproteins produced by and cloaking the epithelium. Due to the absence of this function, germ free animals develop an enlarged cecum because of the accumulation of intact mucous [12]. Convincingly, this effect can be rapidly reversed by monoassociation with Peptostreptococcus micros [13]. Intestinal motility is stimulated by the commensal microbiota such that germ-free animals display restricted and slower migrating motor complexes than conventional animals [14]. These effects, which can be reversed by colonization with the microbiota from a conventional animal [15], provoke hypotheses that the commensal microbiota may initiate these effects through interactions with enteroendocrine cells within the intestinal epithelium and/or the enteric nervous system. Certainly these basic physiological observations provide potential insight wherein the regulation of the intestinal microbiota impacts states such as irritable bowel syndrome.

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Protection against Incoming Microbes The intestinal microbiota is an important constituent in the mucosal defense barrier. Indeed, studies have demonstrated that animals bred in a germ-free environment are highly susceptible to pathogenic infections. Distinct from its stimulatory effects on the immune system (discussed below), the phenomenon in question is termed “colonization resistance” [16,17], wherein the stable ecosystem provided by the commensal bacteria compete for the same nutrients and attachment sites as pathogenic bacteria. Further, through a coordinated effort, the intestinal epithelium produces several compounds that inhibit the growth of pathogens and other transient incoming bacteria that are not members of the residing intestinal microbiota, the opportunity for these pathogens to invade is limited. A clinically relevant example of the colonization resistance provided by the microbiota is the common and difficult to treat antibiotic-associated diarrhea caused by Clostridium difficile that occurs following ablation of the intestinal microbiota with broad spectrum antibiotic therapy. Ingestion of live bacteria aimed at reestablishing the commensal microbiota following antibiotic therapy has been shown to ameliorate antibioticassociated diarrhea [18].

Maturation and Function of the Mucosal Immune System Despite the constant presence of a myriad of antigens from the food and microorganisms that we eat, the commensal microbiota stimulates and then coordinates the gastrointestinal associated immune systems to achieve a disease-free state in any one individual. At birth, the immune system is immature and develops upon exposure to the commensal microbiota and other organisms within the gastrointestinal tract. The regulatory mechanisms governing the development [17] and optimization of these effects have been reviewed elsewhere [19,20]. However, the commensal microbiota are critical for stimulating the number of Peyer’s patches, immunoglobulin or IgA producing cells, dendritic cell recognition of the commensal microbiota [21], and the coordination of pro- and anti-inflammatory signals [22] all serving to regulate the abundance and promoting the efficacy of the immunological barrier provided by the intestinal mucosa. This powerful regulatory influence provided by the intestinal microbiota to the mucosal immune system provides potential therapeutic strategies ranging from heightened host defense and containment of inflammation [23].

Probiotics - ‘Beneficial Bugs’ Given the benefits of a healthy and robust commensal microbiota outlined above, it is not surprising that efforts to optimize this microbiome date back to over 2000 years ago when the Nomads consumer soured milks. However, this concept officially entered the scientific literature around the turn of

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Intestinal Microbiota and Human Health the 20th century. Ilya Metchnikoff, a Russian-born bacteriologist who won the Nobel Prize in Physiology or Medicine in 1908 for his work describing phagocytosis, observed the complex microbial population of the colon [24]. As history tells it, he became very concerned with this ‘autointoxication’ and for a period of time colectomy was recommended to relieve patients of this source of infection. However, Dr. Metchnikoff went on to study the lifestyle and diet in Bulgarians (87y versus 48y in U.S.) and linked their longevity (87 years versus 48 y in the United States at that time) to their consumption of a fermented milk product called Kefir that contained lactobacillus. He convinced many that “friendly-bacteria” colonizing the intestine normalized bowel habits and fought disease-carrying bacteria and wrote that the secret to longevity was in the Russian mountains. As a result, colectomy was abandoned for this purpose and the era of probiotic use began. Dr. Metchnikoff succeeded in isolating the two types of bacilli that are responsible for changing milk into yogurt, and named the primary yogurt-culturing bacteria Lactobacillus bulgaricus, in honor of the long-living Bulgarians who inspired his discovery. The word ‘probiotic’ is a compound of a Latin and Greek word meaning ‘favorable to life’. The Food and Agricultural Organization of the United Nations (FAO) defines a probiotic as a ‘live microorganisms administered in adequate amounts which confer a beneficial health effect on the host’ [25] (Table 2). The microorganisms referred to in this definition are nonpathogenic bacteria (small, single celled organisms which do not promote or cause disease), and one yeast, Saccharomyces. Probiotic supplements are produced for both animals and humans and are available in many forms and preparations ranging from probiotic foods to dietary supplements. The most common probiotics produced are in the form of dairy products containing two types of microbes—lactobacilli and bifidobacteria.

CONCLUSION Given the critical effect of the gastrointestinal microbiota on human health and disease, probiotic consumption has been implicated in a laundry list of human ailments that far exceed the scientific evidence necessary to recommend therapeutic consumption. Exciting data has emerged in areas such as diarrhea, immune function, irritable bowel syndrome and necrotizing enterocolitis. However, the purpose of this supplement is Table 2. Characteristics of a Probiotic Nonpathogenic Resistant to technological processing, storage and delivery Resistant to gastric acidity and lysis by bile Viable in the gastrointestinal environment May adhere to the epithelium Produces antimicrobial substances Modulates the host’s immune response Modulates the host’s metabolism

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not to review the evidence supporting each purported indication but to identify emerging areas of probiotic consumption for distinct purposes that warrant our strong consideration and research efforts.

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Intestinal Microbiota and Human Health 16. van der Waaij D: The ecology of the human intestine and its consequences for overgrowth by pathogens such as Clostridium difficile. Annu Rev Microbiol 43:69–87, 1989. 17. Bauer E, Williams BA, Smidt H, Verstegen MW, Mosenthin R: Influence of the gastrointestinal microbiota on development of the immune system in young animals. Curr Issues Intest Microbiol 7:35–51, 2006. 18. Szajewska H, Ruszczynski M, Radzikowski A: Probiotics in the prevention of antibiotic-associated diarrhea in children: a metaanalysis of randomized controlled trials. J Pediatr 149:367–372, 2006. 19. Rakoff-Nahoum S, Medzhitov R: Role of the innate immune system and host-commensal mutualism. Curr Top Microbiol Immunol 308:1–18, 2006. 20. O’Hara AM, Shanahan F: Mechanisms of action of probiotics in intestinal diseases. Scientific World Journal 7:31–46, 2007.

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21. Niess JH, Reinecker HC: Dendritic cells in the recognition of intestinal microbiota. Cell Microbiol 8:558–564, 2006. 22. Clavel T, Haller D: Molecular interactions between bacteria, the epithelium, and the mucosal immune system in the intestinal tract: implications for chronic inflammation. Curr Issues Intest Microbiol 8:25–43, 2007. 23. O’Hara AM, Shanahan F: Gut microbiota: mining for therapeutic potential. Clin Gastroenterol Hepatol 5:274–284, 2007. 24. Nobelprize.org. 25. World Health Organization: “Health and Nutritional Properties of Probiotics in Food including Powder Milk with Live Lactic Acid Bacteria.” Co´rdoba, Argentina: Food and Agriculture Organization of the United Nations and World Health Organization, 2001.

Received September 14, 2007

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