gut mucosal immunity

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GI Fellows Seminar serious 3 on GI Mucosal
immunity and microbiota

• By Wudassie melak

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Introducton

• GI tract is unique and Multifunctional
• largest organ exposed to external antigens
• GI tract requires regulation to balance immune
  reactions against foreign antigens while
  fostering the symbiotic, commensal microbiota

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Mucosal immunity

• Refers to immune responses that occur at mucosal
  sites.
• organisms and their products, along with ingested
  food, represent an enormous antigenic load that
  must be tolerated to maintain mucosal homeostasis
• Controlled or physiologic inflammation

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Functional anatomy of Mucosal immune system

• Inductive compartment
• Are organized lymphoid structures that bring
  together naive T cells, B cells, and APCs
• PPs? GALT
• Effector compartment
• Phagocytes that engulf and kill microbes
  cytotoxic T cells, B cells, helper T cells
• Lymphoid cells in the epithelium (IELs) ,LP
  (LPLs)
• Are innate and antigen-experienced memory cells

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Innate immunity in gastrointestinal tract

• Intestinal epithelial cells (IEC)
• Antimicrobial peptides
• Toll-like and NOD-like receptors
• Innate immune cells
• Intestinal microbiome

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Intestinal Epithelial Cells

• Largest of the body's mucosal surfaces
• A single layer of cells organized into crypts and
  villi
• Continually renewed by pluripotent epithelial
  stem cells
• The proliferation, differentiation and functional
  potential of epithelial cell progenitors is
  regulated - local stem cell niche

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• IECs are Secretory IECs, including EECs, goblet
  cells and Paneth cells-maintaining digestive or
  barrier function
• luminal secretion of mucins and antimicrobial
  proteins (AMPs)by goblet cells and Paneth cells
• Establishes a physical and biochemical barrier to
  microbial contact with the epithelial surface and
  underlying immune cells

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Paneth cells

• Derived from epithelial stem cells and reside in
  the base of the crypts of small intestine.
• Identified on stains -large eosinophilic
  cytoplasmic granules
• High concentrations of antimicrobial peptides,
  including a-defensins, lysozyme,
• Secrete these AMPs into the crypt, thus keeping
  it relatively sterile.
• Defensins are released upon stimulation by
  various bacterial ligands, including
  endotoxin--via TLRs and nuclear oligomerization
  domain 2 (NOD2).

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Goblet cells

• Specialized in the production of mucins and other
  molecules.
• In addition to their physical properties, mucins
  interact with AMPs to maintain their high
  concentration close to the epithelium
• Produce trefoil factor-3 (TFF-3), which can
  influence mucus viscosity and is important in
  epithelial repair after injury.
• Innate immune cells also interact with goblet
  cells increasing their number by stimulating
  enterocytes to differentiate into goblet cells.

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Tuft cells

• increased in number during parasite infection.
• Tuft cells are identified by a distinct
  morphology, characterized by microvilli
  projecting from the apical membrane.
• Discern helminths in the intestinal lumen
  chemosensors.
• Stimulates mucin production

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IEC Barrier function

• Intracellular components

• Extracellular components

• Tight junction and subjacent adherens junction
• Supported by a dense perijunictional ring of
  actin and myosin

• Mucin
• Prevents large particles, including most bacteria
• Unstirred layer
• Slows down nutrient absorption rate

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Antimicrobial peptides

• First protective barrier in mucous layer of
  esophagus mucin-2 and glycoprotein
• TFF (Trefoil factors) produced by goblet cells
  protease-resistant peptides promote cell
  survival and migration
• Mucous layer of small intestine sIgA and AMPs
• IL-1,IL-4,IL-6,IL-9,IL-13, TNF, type 1 IFN,
  neutrophilproducts
• microbial adhesive proteins increase mucingene
  expression
• Defensins

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Toll-like and NOD-like receptors

• Pattern recognition receptors (PRRs)
• PRRs recognize pathogen-associated molecular
  patterns (PAMPs) such as Lipopolysaccharide,
  flagellin, bacterial DNA and RNA
• PRRs fall into three families
• Toll-like receptors (TLRs)
• NOD-like receptors (NLRs)
• Retinoicacid-inducible gene I (RIG-I)-like
  receptors (RLRs)

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• Role of TLR in GI
• 1.Sensing bacteria in intestinal epithelium
• 2.Sensing intestinal injury
• 3.Regulate barrier function
• Trigger production of cytokines/chemokins
• TLR sensing epithelium injury-promote
  proliferation
• Inc strength of tight junction, IEC motility and
  proliferation

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• NOD like receptors (NLRs)- Intracellular innate
  immune proteins
• Enable detection of intracellular bacteria
• Promote clearance through initiation of a
  pro-inflammatory transcriptional programme and
  other host defence pathways, including autophagy
• Acivate both innate and adaptive immunity

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Innate immune cells

• Eosinophils, Mast cells
• ILC and multifunctional IgA plasma cells
• Macrophages highly phagocytic, generating
  inflammatory response without damage surrounding
  tissue
• Lamina propria contain a dense network of DC

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Adaptive immunity

• Also referred as the acquired immune system, is a
  subsystem of the immune system composed of
  specialized, systemic cells and processes that
  eliminates pathogens by preventing their growth.
• Functional anatomy of adaptive immunity in GI
  tract
• Humoral immunity and Cell-mediated immunity

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Functional anatomy of adaptive immunity

• Initiated in collections of lymphocytes and APC
  closely associated with the mucosal epithelial
  lining of bowel and in mesenteric LN
• GALT Peyers patches, found mainly in distal
  ileum or isolated follicles in appendix and colon
  
• GALT differs from LN
• Ratio of BT cells 5 X higher, non-capsulated,
  independent routes of Ag delivery

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Lymphoid site of immune response

• GALT
• Mesenteric LN serve some of the same function as
  GALT
• correct lymph-borne Ag
• 100-150 LN between membranous layer of mesentery
• Lingual and palatine tonsils-sites of immune
  response in oral cavity

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Antigen uptake
major pathway of antigen delivery from lumen to
GALT is through microfold(M) cells- overlying
Peyerspatchs and lamina propria lymphoid
follicles
M cells engage in transport of microbes or
molecules across barrier into GALT, where they
are handed off to dendriti ccells
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APCs

• APCs in the GI tract include
• Professional APCs such as DCs, B cells,
  macrophages
• Nonprofessional APCs such as epithelial cells
• During inflammatory states, epithelial cells of
  the esophagus and small intestine upregulate MHC
  II and can activate CD4 T cell

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Antigen sampling by dendritic cells
Some DC extend dendritic processes between IEC
into lumen to sample antigens
Other present in lamina propria, sample antigens
that derived from lumina contents through the
epithelial barrier
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Humoral immunity

• Immunoglobulins of the mucosal surface- Major
  function is to neutralize luminal microbes
• mediated mainly by IgA produced in the GALT
• IgA is produced in larger amounts than any other
  antibody isotype because of large number of
  IgA-producing plasma cells in GALT, (80 of all
  Ab-producing plasma cells in the body)
• selective induction of IgA isotype switching in B
  cells in GALT and MLN

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• SIgA is a dimeric form of IgA produced by plasma
  cells in the LP
• Two IgA molecules (homodimers) are bound together
  by J chain (produced by plasma cells).
• The pIgR is expressed on the BLM of IEC

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• Once bound to the pIgR on the IEC, SIgA is
  actively transported within vesicles to the
  apical membrane of the IEC.
• The vesicle fuses with the apical membrane, and
  the pIgR/ IgA complex is released into the
  intestinal lumen.
• pIgR serves to protect the SIgA dimer from
  degradation by luminal proteases and gastric
  acid.
• SIgA binds to mucus, enhancing its ability to
  bind and trap microbial products.

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Unique features of sIgA

• Has anti-inflammatory nature.
• Immune exclusion
• SIgA also may exert specific protective immunity
  against certain pathogens via more direct
  mechanisms
• Suppression of bacterial virulence,
• non-antigen specific binding to bacterial glycan
  residues

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• IgM is another antibody capable of binding the
  pIgR.
• IgM also uses J chain produced by plasma cells to
  form polymers a pentamer.
• Important in patients with IgA deficiency, where
  secretory IgM (SIgM) compensates
• IgE production may play an important role in the
  intestinal response to helminths and in food
  allergy

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Cell-mediated immunity

• T cells scattered throughout lamina propria ,
  submucosa and within Peyers patches
• Different subsets of effector T cells --Th17
  cell, Th2 cells

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Intestinal microbiota

• Microbiome
• Microbiota

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• The intestinal microbiome is a diverse ecosystem
  comprising microorganisms (bacteria, archaea,
  fungi, and viruses including bacteriophages),
  their genomes (i.e., genes), and the surrounding
  environmental conditions.
• The human genome consists of about 23 000 genes,
  whereas the microbiome encodes over 3 million
  genes producing thousands of metabolites, which
  replace many of the functions of the host,

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Where on a healthy human is the microbiome
located?

• Every human body surface which is exposed to the
  environment and every body part with an opening
  to the environment has a microbiome
• Microbiota diversitya measure of how many
  different species and, dependent on the diversity
  indices, how evenly distributed they are in the
  community.
• Dysbiosis
• Composition of the intestinal microbiota varies
  significantly among individuals

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CHARACTERSTICS and COMPOSITION OF GUT MICROBIOTA

• Weighs 1kg although is without distinct
  structure
• Is organized system of cells more akin to immune
  system than liver
• Is dominated by 4 large groups of bacteria or
  phyla Actinobacteria, Bacteroidetes, Firmicutes,
  Proteobacteria-
• The microbiota has a major impact on health
  through interactions with host cells (including
  components of the innate and adaptive immune
  systems)
• through extraction of nutrients and energy from
  the diet, and through complex biotransformations
  (e.g., detoxification) of a variety of ingested
  compounds, including potential carcinogens.

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Distribution of Gut Microbiota

• The intestine contains the largest collection of
  microbes among all of our body habitats
  (locations for microbial colonization).
• In the colon, for example, bacteria reach
  densities of 1011 cells per gram of luminal
  contents.
• Highly resilent

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Factors Affecting Gut Microbiota
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FACTORS AFFECTING INTESTINAL MICROBIOMEVARIABILIT
Y AND RESILIENCE

• Age --microbiota composition and function
  continues to change throughout life.
• Mode of delivery C/S or SVD
• Mode of feeding- Breat or Bottle formula
• Antibiotics in infancy
• Pre-adolescent intestinal microbiome is enriched
  in functions such as vitamin synthesis that
  support development

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Development of microbiota
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Early gut colonization has four phases

• Phase 1 Sterile gut
• Phase 2 Initial acquisition vagina, feces,
  hospital
• Phase 3 Breast feeding or bottle-feeding
• Breast fed more bifidobacteria (up to 90 of
  flora)
• Bottle fed more diverse more Bacteroides , and
  Clostridial species
• Phase 4 Start of solids move to adult flora
  (Fermicutes and bacteriotedes)
• Bifidobacteria remain key flora into adulthood

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• Gender- Women display higher levels of
  microbiota diversity and fn and a decreased
  abundance of Bacteroides and Prevotella species.
• Genetics, Geography and diet
• Medications -reduce microbial diversity
• Life style and habits.. Like stress level
  ,alcohol, smoking
• Microbe microbe signaling- microbial selfs
  election, bacteriocins,

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Gut Microbiota and Geographic Location

• African children have greater proportions of
  Bacteroidetes and Gram-positive organisms in
  their bowels, while a Western lifestyle appears
  to promote increases in Firmicutes and
  Gram-negative organisms.
• MACs found in fiber are one of the key sources of
  nutrients for intestinal microbes.
• In fermenting MACs, microbes produce SCFAs, which
  can help attenuate inflammation, serve as an
  energy source for IECs, and improve GI transit.
• The Western diet is low in MACs and has been
  associated with the risk of inflammatory and
  metabolic-related diseases

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Diet

• Low dietary fiber results in an increased
  reliance of intestinal microbes on the host
  epithelium and mucus, resulting in disruption of
  the epithelial barrier and an increased
  susceptibility to inflammation.
• A similar effect is also seen with a high-protein
  diet, which results in increased microbial
  density as well as an increased potential of the
  microbiome to cause colitis.
• Additives such as emulsifiers and substitutes
  such as artificial sweeteners can have
  deleterious effects on the intestinal microbiome
  and increase propensity for metabolic and
  inflammatory disorders

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• Dietary components serve as substrates for
  microbial metabolic pathways and hence can
  influence the generation of specific microbial
  metabolites, which influence host physiology.
• Some examples
• SCFAs from dietary carbohydrates, which can
  influence the intestinal serotonergic pathway,
  thereby altering GI motility and
• dietary fat-related free fatty acids and
  lipopolysaccharide, which are associated with
  enteric neurodegeneration, altered GI motility,
  and systemic effects contributing to obesity.

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• early antibiotic use is associated with delayed
  maturation of the microbiome and long-lasting
  changes in both microbiota composition and
  functionality
• association of early antibiotic use and increased
  risk for Crohndisease
• PPIs, laxatives, metformin, statins, hormones,
  benzodiazepines, anti depressants,NSAIDs, and
  antihistamines among others, are associated with
  changes in the composition of the intestinal
  microbiota

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Role of microbiota in Health

• Symbiotic relationship
• Shaping and maintaining immunity
• Innate immunity
• Adaptive immunity

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Host microbiome interactions With GUT immune
system

• Host

• Microbiome

• PAMPs are recognized by PRR- bearing cells of the
  innate immune system and many epithelial cells
• Microbiota stimulation leads to B cell switch to
  IgA, regulatory T cell induction, T cell
  differentiation to Th17
• IECs ? a defensins?limit contact with bacteria

• Produce PAMPs and metabolic byproducts and
  regulate intestinal immune responses
• Generate immune tolerance
• Commensal bacteria induce CD4T cell
  differentiation.
• Produce symbiosis factor ?decreases inflammation
  through SCF, TH17

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• Protective function (barrier effect) of microbes
• Compete and adhere to the attachment sites in the
  brush border of intestinal epithelial cells
• Compete for available nutrients.
• Produce antimicrobial (bacteriocins).
• All of this will prevent attachment and
  subsequent entry of pathogenic bacteria into the
  epithelial cells

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• Production of NT- Serotonin GI motility, inc
  transit time , Secretion and sensitivity (LPS,
  SCF, )
• Visceral hypersensitivity E coli
• Epithelial barrier maintainance - - change
  expression of TJ proteins, butyrate
• Bile acid decnjugation

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The microbiomegutbrain axis

• From bottom- up
• Alter ENS fn- through vagal pathways
• Metabolites
• SCF diffuse across BBB
• Association with human behavior, Autism

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Role of microbiota in Disease
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Diseases of the gut non mucosal diseases

• Malabsorption syndrome
• Malignancies Colorectal cancer
• Inflammatory Bowl disease (IBD)
• Irritable Bowl syndrome
• Diarrheal diseases
• Clostridium Difficile Infection (CDI)

• Obesity and metabolic syndrome
• Malignancies liver cancer, CRC
• Complications of liver cirrhosis
• Allergic conditions
• Autoimmune disorders (T1DM, arthritis )
• Autism and other neurological disorders
• Chronic fatigue syndrome
• Periodontal diseases

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IMT/FMT

• Administration of a solution of fecal matter from
  a donor into the intestinal tract of a recipient
  in order to directly change the recipients gut
  microbial composition and confer a health
  benefit.
• Re-establishes a balanced intestinal microbiota
• Potential Indications
• GI Disorders recurrent Clostridium difficile
  infection , IBD, IBS -C, and chronic constipation
• Non-GI disorders Obesity, Chronic Fatigue
  Syndrome, Autism

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PROBIOTICS /PREBIOTICS

• The microbiota can also be modified by adding
  live micro-organisms to food or by periods of
  fasting.
• Probiotics are live bacteria and yeasts that,
  when administrated in a viable form and in
  adequate amounts, are beneficial to human health.
  
• They are usually added to yoghurts or taken as
  food supplements.
• Prebiotics are defined as a substrate that is
  selectively used by host micro-organisms
  conferring a health benefit.
• Symbiotic contain a mixture of prebiotics and
  probiotics

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References

• sleisenger and fordtrans gastrointestinal and
  liver disease,11th edition
• Gut microbiome as a clinical tool in
  gastrointestinal disease management are we there
  yet? Nat. Rev. Gastroenterol. Hepatol.
  doi10.1038/nrgastro.2017
• Role of the gut microbiota in nutrition and
  health. Science and Politics of Nutrition. BMJ,
  13 June 2018.

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• Thank you

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