Our gastrointestinal tract is home to no less than 1012 to 1014 microorganisms, i.e., 2 to 10 times more than the number of cells which make up our bodies. This collection of non-pathogenic bacteria, viruses, parasites and fungi makes up our intestinal microbiota (or intestinal flora). Its role is becoming increasingly well known, and researchers are now attempting to clarify the relationship between imbalances in the microbiota and certain disorders, particularly autoimmune and inflammatory disorders.
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Understanding the role of the intestinal microbiota
The microbiota is a collection of microorganisms - non-pathogenic bacteria, viruses, parasites and fungi, known as commensal microorganisms - which live in a specific environment. Different microbiotas exist in the body, on the skin, in the mouth, in the vagina, etc. The intestinal microbiota is the most extensive, with 1012 to 1014 microorganisms: 2 to 10 times more than the number of cells which make up our bodies, weighing 2 kg!
The intestinal microbiota is mainly found in the small intestine and colon – the stomach acids make the stomach wall practically sterile. It is distributed between the lumen of the gastrointestinal tract and the protective biofilm formed by the intestinal mucus on its internal wall (intestinal epithelium).
The presence of microorganisms in the intestine has been known for over a century and it was soon assumed that there was genuine symbiosis between our body and this flora. However, until recently, limited technical resources were available to study this interaction in detail: only a minority of the bacterial species constituting the microbiota could be cultured in vitro. The development of high-throughput sequencing techniques for genetic material has given new impetus to this research, and there is a real trend today in research aiming to describe the nature of host-microbiota interactions, interactions between the microorganisms, and their impact in terms of health.
Hence, the role of the intestinal microbiota is becoming increasingly clear. It is now known to play a role in digestive, metabolic, immune and neurological functions. Consequently, dysbiosis, i.e., qualitative and functional impairment of the intestinal flora, is a serious avenue for understanding the cause of certain disorders, particularly those with underlying autoimmune or inflammatory mechanisms. This has become a central theme in biological and medical research.
MetaHIT: Flora with unparalleled wealth
The MetaHIT study, initiated in 2008, and coordinated by Inra, aimed to identify all intestinal microbial genomes (metagenome) by high-throughput sequencing. It also made it possible to outline the interactions between the metagenome and health. This study, the first of its kind, is based on the analysis of stool samples collected from 124 participants. It thus identified a total of 3.3 million different genes, belonging to more than 1,000 different species, the vast majority of bacterial origin. From an individual perspective, it also showed that each person had on average 540,000 microbial genes, i.e., approximately 160 species, divided into seven different phyla (family groups). Lastly, MetaHIT was the first study to demonstrate the extreme wealth of the intestinal flora, by identifying certain as yet unknown bacterial species.
Like fingerprints, the intestinal microbiota is specific to each individual: it is unique in both qualitative and quantitative terms. Out of the 160 species of bacteria on average found in the microbiota of a healthy individual, half of these species are commonly found in one individual to another. Furthermore, a common foundation of 15 to 20 species is said to exist, responsible for essential microbiota functions. Although this is somewhat controversial, it appears that homogeneous population groups can be distinguished according to the predominant species found in their microbiota: three main groups – or enterotypes – are distinguished: Bacteroides, Prevotella and Clostridiales.
Bacterial viruses (which infect bacteria) are also found in very large numbers in the microbiota. These can modify the genetic inheritance of intestinal bacteria or its expression. Hence, the virome is undoubtedly another part of the puzzle regarding the physiopathology specific to the intestinal flora, like the fungal microbiota which includes yeasts and molds. Other study subjects to be explored.
A unique ecosystem formed at birth
A person's microbiota is created at birth, upon contact with the vaginal flora after natural childbirth, or upon contact with environmental microorganisms for infants born by C-section. Bacterial colonization occurs gradually, in a clearly defined order: the first intestinal bacteria need oxygen to multiply (aerobic bacteria: Enterococci, Staphylococci, etc.). By consuming the oxygen present in the intestine, they then promote the introduction of bacteria which only proliferate in the absence of this gas (anaerobic bacteria: Bacteroides, Clostridium, Bifidobacterium, etc.).
Due to the influence of a varied diet, genetics, hygiene level, medical treatments administered and the environment, the composition of the intestinal microbiota then develops qualitatively and quantitatively during the first few years of life. The qualitative and quantitative composition of the microbiota then remains fairly stable thereafter. Fluctuations in sex hormones – testosterone and estrogens – may nonetheless have an impact on its composition. Medical treatments, changes in lifestyle and various events can also modify the microbiota, in a more or less durable manner. For instance, antibiotic treatment reduces the quality and quantity of the microbiota over several days to several weeks. The initial species are largely capable of reestablishing themselves, although differences may persist. Repeated antibiotic therapy during a person's life could thus lead to gradual, permanent, potentially harmful changes in the microbiota. However, we do not all appear to be equal with regard to this risk: some individuals apparently have a more stable microbiota than others, when faced with the same disruptive events.
When the microbiota benefits the body
The intestinal microbiota regulates its own metabolism by drawing on our food (particularly fiber-rich food). At the same time, its microorganisms play a direct role in digestion:
- they ferment indigestible food residues and substrates;
- they facilitate the assimilation of nutrients thanks to a collection of enzymes otherwise not found in the body;
- they hydrolyze starch, cellulose, and polysaccharides, etc.
- they help synthesize certain vitamins (vitamin K, B12, B8).
- they regulate several metabolic pathways: absorption of fatty acids, calcium, magnesium, etc.
Animals reared without a microbiota (known as axenic animals) thus have energy requirements 20 to 30% higher than those of a normal animal.
The microbiota also acts on intestinal epithelial function: gastrointestinal motility is slower in axenic animals. The cells constituting this epithelium are incompletely differentiated and its irrigating vascular network is less dense than in normal animals. Now, this vascular system plays a decisive role in nutritional and hormonal metabolism, and for the storage of immune cells within the intestinal wall.
The intestinal microbiota fully contributes to intestinal immune system function: the latter is essential to the barrier role of the intestinal wall, subjected to the flow of antigens of dietary or microbial origin, from birth. Hence, bacteria such as Escherichia coli directly fight against the colonization of the gastrointestinal tract by pathogenic species, due to a competing phenomenon and by producing bactericidal substances (bacteriocins). At the same time, from the first few years of life, the microbiota is essential to helping the intestinal immune system to learn how to distinguish between friendly (commensal) and pathogenic species. Studies show that the immune system in axenic mice is immature and incomplete compared with mice reared under normal conditions: in the intestinal epithelium of these mice, Peyer's patches, inducers of intestinal immunity, are immature, and lymphocytes, effectors of immune responses, are limited in number. The spleen and lymph nodes, which are important immune system organs for the body's immunity in general, thus display structural and functional abnormalities.
Microbiota and inflammation
Inflammation is an important element, closely correlated with immunity: there is both a vital physiological aspect of inflammation, notably controlling the microbiota, and important inflammatory responses triggered in the presence of pathogenic species. The latter mechanism is notably based on the presence of inflammatory bacterial components, such as lipopolysaccharides (LPS) found on the surface of certain (Gram-negative) bacteria. These antigens induce an immune response by intestinal macrophages which then produce proinflammatory mediators (cytokines). These trigger local inflammation and increase the permeability of the intestinal wall. LPS may then cross the latter, enter the blood circulation, and induce an inflammatory phenomenon in other target tissues.
Challenges facing research
Exploration of the intestinal microbiota has recently become central to health research.
Inflammatory bowel disease: an obvious link
Chronic inflammatory bowel disease (IBD), such as Crohn's disease and ulcerative colitis, is related to inappropriate activation of the intestinal immune system. Onset thereof is triggered by underlying genetic and environmental factors (diet, age, etc.). At the same time, the improvement in symptoms of patients receiving antibiotic therapy and the resolution of intestinal inflammatory lesions in individuals whose intestinal wall is no longer in contact with feces (fecal diversion) have also pointed to the role of the microbiota.
A microbiota which is imbalanced in terms of proinflammatory and anti-inflammatory bacterial species, and the predominant nature of certain classes of bacteria (Enterobacteria, Fusobacteria), or the rare nature of other species (Clostridia, Faecalibacterium) have been described in individuals suffering from IBD. At present, it is not possible to ascertain whether this is a cause or consequence of these diseases, or to determine whether dysbiosis behind the disease is innate or the result of another environmental factor (diet, medication, etc.). An attractive hypothesis has been put forward: dysbiosis is said to develop due to the influence of genetic and environmental factors, but apparently plays a role in triggering, maintaining and the degree of severity of inflammation, resulting in a vicious circle.
Furthermore, among the dozens of IBD predisposition genes currently identified, some play a decisive role with regard to the microbiota. Mutation of the NOD2 gene is most frequently observed in patients suffering from Crohn's disease: this gene codes for an innate immunity receptor responsible for detecting a component of the bacterial wall. Once it has mutated, it can no longer fulfill this function or help preserve the intestinal barrier. Other mutations have been reported, for example, concerning the ATG16L1 gene, involved in the autophagy of immune cells in the presence of bacteria, or the MUC2 gene, which plays a role in synthesis of the intestinal mucus.
The microbiota is one of the preferred therapeutic target in these inflammatory diseases. Until now, the first clinical trials conducted with probiotics or prebiotics have been inconclusive. However, new studies are expected, based on a more rational selection of microorganisms or compounds to be utilized. At the same time, certain teams are attempting to create genetically modified probiotics which would make it possible to implant the microorganism of interest while giving it additional properties, such as the secretion of immunomodulatory mediators.
Dysbiosis and metabolism
Diabetes and obesity have multifaceted causes, genetic, nutritional and environmental. The respective role of each of these factors varies from one individual to another, and the molecular mechanisms implied for each of them have yet to be accurately described.
Hence, increasing fats in a regular diet increases the proportion of Gram-negative bacteria. As a result, this increases the presence of inflammatory LPS locally, then, once the LPS enter the blood circulation, in the liver, adipose tissue, muscle tissue, etc. This low-grade chronic inflammation which becomes established in these tissues promotes insulin-resistance prior to diabetes and obesity. Furthermore, in axenic mice, the introduction of microbiota from obese mice rapidly causes considerable weight gain.
Other mechanisms involving the microbiota may also be implicated: in addition to LPS, the increase in epithelial permeability could allow whole bacteria to pass through. Lasting implantation in adipose, muscle and hepatic tissue supposedly promotes in situ persistence of inflammation. At the same time, certain circulating bacterial metabolites are said to play a decisive role in the mechanism for renal regulation of blood pressure, or in the development of atheromatous plaque.
The idea, today, is to develop tailored strategies, in which the prebiotics, probiotics or symbiotics provided are suited to each individual patient's specific requirements. In the longer term, preventive treatment could be developed with a view to avoiding the occurrence of these diseases.
From carcinogenesis to cancer treatment
In the field of cancer, the microbiota comes into play on two levels: firstly in carcinogenesis itself. Several data effectively confirm that certain tumors are related to the presence of specific microorganisms, or even intestinal dysbiosis. For example, an imbalanced microbiota favorable to certain species(fusobacterium) would increase the risk of colorectal cancer, while the presence of Helicobacter pylori promotes the onset of stomach cancer. Data collected in animal studies also indicate an increase in the incidence and severity of mammary gland tumors in mice subjected to frequent antibiotic regimens. These data are correlated with an epidemiological study in which young women having received on average more than two courses of antibiotics per year have a higher risk of breast cancer compared to other women. However, in this field, the difficulty lies in making a distinction between the role of the microbiota and other carcinogenic risk factors – tobacco, alcohol, etc.- which also promote dysbiosis.
In addition to carcinogenesis, the efficacy of cancer treatments is also said to be influenced by the microbiota. Certain cancer drugs are thought to act in synergy with the intestinal flora: the efficacy of cyclophosphamide - widely used in oncology - is known to be influenced by the microbiota which promotes intestinal permeability and the migration of immunogenic bacteria to the tumor-immune system. These bacteria are said to induce an immune response in synergy with the cancer drug.
Immunotherapy, recently used the treatment of melanoma, together with lung cancer and cancer of the kidney, is also thought to benefit from a boost by Bacteroides type bacteria. These apparently influence the ability of the immune system to naturally resist the melanoma. Furthermore, the efficacy of anti-melanoma treatment with ipilimumab is correlated with the presence of one of the two species of bacteria in the Bacteroides class.
Other therapies (platinum salts and nivolimab) and other cancer targets could respond to the same mechanisms. The therapeutic prospects are vast: analysis of the microbiota could become a routine test prior to initiating treatment, predictive of the therapeutic response. If necessary, treatments specific to the microbiota would be added: probiotics known to be capable of boosting intratumoral lymphocytes could be combined with conventional cancer treatments.
Neuropsychiatry influenced by the gut-brain axis
The nervous system which regulates the intestine contains 200 million neurons. Its primary function is to ensure intestinal motility; however, 80% of these nerve cells are afferent, i.e., they convey information from the intestine to the brain. This is why the enteric nervous system is described as a second brain. Researchers soon put forward the hypothesis that a change in the microbiota could modify the information transmitted to the central nervous system. Several clinical findings have been reported, such as a significant improvement in autistic symptoms following antibiotic treatment. Although this correlation seemed unlikely a few years ago, it has since been taken seriously.
The role of the microbiota is suggested in numerous neuropsychiatric disorders: autism, schizophrenia, anxiety and depression or bipolar disorders. There are still insufficient scientific arguments in the majority of cases, although preliminary evidence has recently been published. This would add to the numerous genetic, epigenetic, environmental, and psychological factors, etc. which also play a decisive role in triggering these disorders.
In individuals suffering from schizophrenia or bipolar disorders, the balance between the different proinflammatory and anti-inflammatory cytokines in the blood is disrupted, mediated by factors including LPS and other bacterial translocation markers.
In autism, it has also been shown that mice could develop anxious and self-harming behavior if the composition of their microbiota was significantly modified during a specific stage of their development. Researchers suggest that a similar phenomenon could occur in children and promote the development of autism.
Recently, a number of studies have suggested that the microbiota could play a decisive role in neurodegenerative diseases: it is said to be involved in brain inflammation found in Alzheimer's disease. The severity of Parkinson's symptoms is also correlated with the concentration of a specific species (Enterobactericeae). All of these different phenomena could be mediated by bacterial neuroactive substances. Furthermore, the development of transcriptomic data (on gene expression) and metabolomic data (relating to metabolites) should facilitate their identification.
The therapeutic prospects are vast: preliminary studies have shown that administration of certain probiotics was able to improve symptoms of anxiety or depression in unwell and healthy individuals; others have shown that adapting the diet was able to improve cognitive decline. These avenues of exploration are still in the very early stages, and need to be confirmed.
Treatment: Six therapeutic avenues for modifying the composition of the microbiota?
Disorders triggered or maintained by dysbiosis could be treated by means of six different therapeutic measures:
- a diet promoting the development of bacteria beneficial to the gastrointestinal system.
- antibiotic treatment targeting harmful species involved in the physiopathology of the disease. This option cannot, however, be envisaged as a long-term treatment owing to its potential for selection pressure; it could thus give rise to new disorders.
- oral administration of probiotics, non-pathogenic live microorganisms, shown to be beneficial to the intestinal flora.
- administration of prebiotics, indigestible dietary components, which are helpful for the growth or activity of certain intestinal bacterial populations.
- symbiotics, which combine pre- and probiotics.
- fecal transplantation, which involves administering a bacterial suspension prepared from the stools of a healthy individual through a nasogastric tube or via an enema. This enables a normal microbiota to be introduced into an unwell patient. This therapeutic option has already proved effective and is used for severe intestinal infections caused by Clostridium difficile.