Without a mucosal immune system an animal would probably die, so the simple answer as to why it is important the study of mucosal immunology is important is that it is relevant not only to immunologists but also to clinicians. Without some basic knowledge of the normal GI mucosal immune system it is impossible to understand why diseases such as food allergy and inflammatory bowel disease develop, or how pathogens cause GI disease and can invade the body, or the rationale for conventional and novel therapies for immune-mediated GI diseases.

The mammalian gastrointestinal (GI) tract is a highly complex organ that has to digest and absorb food, and excrete faeces. These functions are achieved in a controlled and coordinated manner through direct cell-cell interactions and through neuro-humoral and chemical messaging. There is a large range of chemicals and enteric hormones involved, and it is estimated that there are as many neurones in the canine GI tract as there are in the brain. This complex organ is also, remarkably, the largest immunological organ in the mammalian body: the number of immune cells in the GI tract exceeds the total number in the rest of the body.

As well as being the largest immune system in the body, the gut associated lymphoid tissue (GALT) is also probably the most complex in the body. It comprises one arm of the mucosal associated lymphoid tissue (MALT) with constant trafficking of cells between mucosal surfaces (i.e., GI, respiratory, urogenital and mammary surfaces), mesenteric lymph nodes, the lamina propria, and the intestinal epithelium, whilst also interacting with the systemic immune system. And at the mucosal surface these immune cells interact with other cells such as enterocytes and enteric neurones, through various chemical messengers (cytokines), thereby modulating their function.

When one considers the role of the intestine, the need for such a complex immune system is perhaps not surprising. The GI tract is required to digest and absorb nutrients whilst excluding toxins and mounting an immune response against potential pathogens at the same time as recognising but maintaining a state of tolerance towards non-harmful dietary antigens and commensal bacteria. It is continually bombarded by dietary and bacterial antigens: it has been calculated that the average 20 kg dog eats over one tonne of dry dog food in a 10 year lifespan and estimated that the average diet contains over 2000 different antigens. Furthermore, the commensal gut flora comprises more cells than the whole mammalian body; half of faecal output is actually bacteria. Finally the gut is repeatedly challenged with potential pathogens frequently ingested with food; like the skin the GI mucosa is in direct contact with the environment, but is also bathed continuously in food antigens and bacteria! So what is perhaps more surprising is that dysfunction is so relatively uncommon.

Introductory Immunology

The GI immune system has to be highly controlled: it needs the ability to mount a vigorous immune response to invading pathogens, whilst selectively ignoring antigens of benefit to the host (i.e., commensal flora, food antigens) by a process (which is not fully understood) known as mucosal (oral) tolerance (1).

Innate immunity provides a primitive 'first line' defense, mediating antigen exclusion. It is comprised of:

 Commensal microflora, which may exclude pathogens by competing for colonisation sites, sometimes by the release of antibacterial substance

 Peristalsis and digestion

 Epithelial barrier

 Secretions containing antimicrobial substances and polyreactive immunoglobulins (IgA and IgM)

 Leukocytes, including neutrophils, mast cells, macrophages, dendritic cells, natural killer (NK) cells and some T lymphocyte subtypes (e.g., intraepithelial lymphocytes)

The adaptive (or specific) immune system comprises:

 Antigen presenting cells (APCs), such as macrophages and dendritic cells

 Enterocytes, which can also act as APCs

 Various lymphocyte populations (B and T lymphocytes, including T-helper, T-cytotoxic and T-regulatory or suppressor cells)

 Antibody-secreting plasma cells

 high-specificity antibodies of the IgA, IgG, IgM or IgE classes

 Interleukins mediating communication between immune cells

 Chemotactic cytokines directing the movement of immune cells

Initially, foreign antigen is taken up by APCs, broken into small peptides, and presented on the cell surface in association with class I or II molecules of the major histocompatibility complex (MHC). The MHC-peptide complex is recognised by T helper lymphocytes which, in the presence of other co-stimulatory surface molecules and cytokine signaling, become activated and thereby activate B cells binding the same antigen.

After activation, antigen-relevant T and B cells proliferate within the lymphoid tissue, but are then exported to the bloodstream where they 're-circulate' until they reach the tissue where the antigen was originally encountered. Here, they selectively exit the circulation by virtue of the interaction with specific molecules (addressins) expressed by the local vascular endothelium. The subsequent effector phase of the immune response results in antigen elimination by antibody-mediated phagocytosis and complement activation, and by cell-mediated immunity. The large numbers of antigen-specific cells are normally controlled before they damage normal tissue by populations of T regulatory (suppressor) cells. Memory T and B cells are retained to mount the more potent secondary immune response on re-exposure to antigen.

Anatomy of the GI Immune System

There are three specific immunological compartments within the GI tract:

 Organised, unencapsulated, lymphoid tissue in Peyer's patches

 The enterocyte lining of the intestine

 Diffuse lymphoid tissue scattered throughout the lamina propria

Normal GI Immune Responses

A wide range of pathogens can potentially infect the intestine, and will tend to provoke a protective immune response. Pathogen-derived antigen is first taken up, usually via the microfold (M) cells overlying Peyer's patches, and presented to T cells, initiating an immune response. Antigen-activated lymphocytes 'home' to the lamina propria via mesenteric lymph and blood, through interaction with addressins (particularly MAdCAM-1) on the vascular endothelium.

The effector immune response is determined by the nature and amount of antigen and its route of presentation, but may primarily involve:

 IgA production and secretion across the epithelium

 Classical type 1 response controlled by T helper 1 (Th1) lymphocytes: cell-mediated T cell cytotoxicity, IgG and cytokines (e.g., IFN-gamma)

 Classical type 2 response controlled by T helper 2 (Th2) lymphocytes: IgE-mast cell and eosinophil dominated

Tolerance

Whilst an active immune response to pathogens is not surprising, the more striking feature of the GALT is the ability to selectively recognise and ignore non-pathogenic antigens. Such antigens may preferentially enter the lamina propria compartment through the enterocytes or lamina propria dendritic cells rather than the Peyer's patches, and activate regulatory and suppressor T cells. Such T-reg cells are characterised by the dominant production of the down-regulatory cytokine IL-10, whilst T helper 3 (Th3) cells predominantly express transforming growth factor (TGF-beta). Both mediate local and systemic unresponsiveness, i.e., oral tolerance.

Appropriate Inflammatory Responses

The normal intestinal mucosa has an intact mucosal barrier and an environment dominated by down-regulatory cytokines e.g., IL-10 and TGF-beta. Therefore, most immune responses that develop are tolerance responses, with IgA expression and immune exclusion. Yet the GALT must decide when to generate specific immune responses (i.e., towards a pathogen), and when to remain tolerant (i.e., to commensal bacteria or food). The best hypothesis for how such decisions are made is currently the 'Danger theory', based on the supposition that the type of response depends upon the amount of antigen and the context in which it is presented.

When the mucosa is invaded by a pathogen or toxin, cell damage leads to release of 'danger signals' i.e., inflammatory mediators (e.g., prostaglandins, leukotrienes), pro-inflammatory cytokines (e.g., IL-1, IL-6 and TNF-alpha) and chemokines (e.g., IL-8). In this altered micro-environment, the immune response generated changes from tolerance to an active immune response. This can either be 'Th1-dominated' (e.g., cytotoxicity and IgG responses) or 'Th2-dominated' (e.g., IgE responses).

Inappropriate Intestinal Inflammation

If the antigenic challenge to the GALT is contained, the 'danger' signals diminish and the normal 'tolerogenic' environment returns. However, if mucosal barrier remains breached, or the pathogenic insult continues unabated, or there is an inherent immune abnormality, chronic inflammation ensues. This may also lead to a breakdown in tolerance to harmless environmental antigens (food components and commensal bacteria), and food allergy or inflammatory bowel disease may develop. Whilst the presence of either a disrupted mucosal barrier, or immune system dysregulation, or both, are required for development of uncontrolled inflammation, the presence of the specific food antigen or the enteric flora respectively are essential for the expression of these diseases. Ultimately any histopathological changes are similar as there is a final common for the development of intestinal inflammation whatever the cause.

The role of the GALT in the aetiology of spontaneous idiopathic IBD and food allergy in will be discussed in subsequent lectures, but may be associated with one or a combination of:

 Altered mucosal permeability

 Abnormal antigen presentation

 Dysregulation of the mucosal immune system

Any of these mechanisms may depend on genetic susceptibility, and perhaps be damaged by infective agents. Development of dietary sensitization or intestinal inflammation may follow a primary insult to the GI tract, and then become self-perpetuating. Treatment is aimed at trying to restore the normal mucosal barrier and switch off the inappropriate immune response.

The help of Prof. MJ Day and Dr AJ German in the production of this abstract is acknowledged.

References

1.  German AJ, Hall, EJ,Day MJ. Chronic intestinal inflammation and intestinal disease in dogs. J Vet Intern Med 2003; 17:8-20.

2.  Hall EJ, German AJ. Diseases of the Small Intestine. In: Textbook of Veterinary Internal Medicine. SJ Ettinger and EC Feldman (eds). Elsevier, Philadelphia 2005, pp 1332-1340