The wet surfaces, immunity and autism


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Autsim File Conference Presentation 2010

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  • Epigenetic mechanisms are affected by several factors and processes including development in utero and in childhood, environmental chemicals, drugs and pharmaceuticals, aging, and diet. DNA methylation is what occurs when methyl groups, an epigenetic factor found in some dietary sources, can tag DNA and activate or repress genes.  Histones are proteins around which DNA can wind for compaction and gene regulation. Histone modification occurs when the binding of epigenetic factors to histone “tails” alters the extent to which DNA is wrapped around histones and the availability of genes in the DNA to be activated. All of these factors and processes can have an effect on people’s health and influence their health possibly resulting in cancer, autoimmune disease, mental disorders, or diabetes among other illnesses.
  • The dualistic and the continuum models of the microbial and immunological worlds. The ancestral view of immunology states that the immune system is educated not to react to self and to react to non-self. In that view, self is good and induces tolerance during lymphocyte maturation and selection, or anergy of self-reacting mature lymphocytes. Non-self, including microbes, as well as allotypic and xenotypic tissues, are evil and elicit an immune response aimed at destroying non-self. All can be boiled down to self/non-self discrimination.
  • The modern view of immunology states that the immune system reacts primarily to danger signals.The immune system is still educated to be tolerant to self, but the notion of self is modified. Good includes normal self and mutualistic microbes, whereas evil includes altered self such as dead cells releasing danger signals and pathogenic microbes that alter the antigenic landscape of normal self. In that context, the normal self induces a physiological level of inflammation that contributes to homeostasis through for example containment of the intestinal microbiota, whereas injury and pathogens induce pathological inflammation that leads to “full-blown” inflammation. The continuum model states that the perceived duality of mutualistic and pathogenic microbes, normal and altered self, and regulatory or inflammatory immunity, represents extremes of a continuous reality. Microbes can express different levels of mutualistic or pathogenic properties and these levels can vary during interaction with the host. Similarly, the state of self and of immune responses can navigate between well-described extremes, and the most likely states are combination of these extremes
  • The continuum of microbial states and immune responses: a dynamic equilibirium. Microbes, including members of the symbiotic microbiota, are not inherently mutualistic or pathogen, but navigate between shades of mutualism and parasitism. Facing the microbes, the immune system is not designed to discriminate between mutualist or pathogens, but merely to react to signals, including MAMPs and antigens. The nature of the immune response is not purely regulatory or inflammatory, but more generally adjusts to the nature of the trigger it faces, like a spring that is pulled by the intensity of the microbial challenge. Furthermore, the immune system has the capacity to evolve when challenged, through generation of different types of lymphocyte subsets such as Th1, Th2, Th17, Treg, Th22 cells, and follicular T helper cells, and generation of memory lymphocytes and lymphoid tissues. This adds another level of adaptability to the immune system and provides it with the necessary flexibility to maintain homeostasis of the superorganism. MAMP, microbe-associated molecular pattern.
  • a | A healthy microbiota contains a balanced composition of many classes of bacteria. Symbionts are organisms with known health-promoting functions. Commensals are permanent residents of this complex ecosystem and provide no benefit or detriment to the host (at least to our knowledge). Pathobionts are also permanent residents of the microbiota and have the potential to induce pathology. b | In conditions of dysbiosis there is an unnatural shift in the composition of the microbiota, which results in either a reduction in the numbers of symbionts and/or an increase in the numbers of pathobionts. The causes for this are not entirely clear, but are likely to include recent societal advances in developed countries. The result is non-specific inflammation, which may predispose certain genetically susceptible people to inflammatory disease and may be caused by pathogens, which are opportunistic organismsNature Reviews Immunology 9, 313-323 (May 2009) | doi:10.1038/nri2515The gut microbiota shapes intestinal immune responses during health and diseaseJune L. Round1 & Sarkis K. Mazmanian
  • The role of the serotonergic system in the neuroplastic events that create, repair, and degenerate the brain has been explored. Synaptic plasticity occurs throughout life and is critical during brain development. Evidence from biochemical, pharmacological, and clinical studies demonstrates the huge importance of an intact serotonergic system for normal central nervous system (CNS)function. Serotonin acts as a growth factor during embryogenesis, and serotonin receptor activity forms a crucial part of the cascade of events leading to changes in brain structure. The serotonergic system interacts with brain-derived neurotrophic factor (BDNF), S100beta, and other chemical messengers, in addition to ts cross talk with the GABAergic, glutamatergic, and dopaminergic neurotransmitter systems. Disruption of these processes may contribute to CNS disorders that have been associated with impaired development. Furthermore, many psychiatric drugs alter serotonergic activity and have been shown to create changes in brain structure with long-term treatment. However, the mechanisms for their therapeutic efficacy are still unclear. Treatments for psychiatric illness are usually chronic and alleviate psychiatric symptoms, rather than cure these diseases. Therefore, greater exploration of the serotonin system during brain development and growth could lead to real progress in the discovery of treatments for mental disorders.
  • Department of Pediatrics, Duke University Medical Center, Durham, NC 27710, USA. Delon006@mc.duke.eduOne hundred and twenty-nine children, 2 to 8 years old, with idiopathic autistic spectrum disorder diagnosed by standard instruments (Childhood Austim Ratings Scale and Autism Diagnostic Observation Schedule) were treated with fluoxetine (0.15 to 0.5mg/kg) for 5 to 76 months (mean 32 to 36 months), with discontinuation trials. Response criteria are described. Family histories were obtained using the family history method in repeated interviews. Fluoxetine response, family history of major affective disorder, and unusual intellectual achievement, pretreatment language, and hyperlexia were used to define a coherent subgroup of autistic spectrum disorder. Statistical analyses were post hoc. Of the children, 22 (17%) had an excellent response, 67 (52%) good, and 40 (31%) fair/poor. Treatment age did not correlate with response. Fluoxetine response correlated robustly with familial major affective disorder and unusual intellectual achievement, and with hyperlexia in the child. Family history of bipolar disorder and of unusual intellectual achievement correlated strongly. Five children developed bipolar disorder during follow-up. Fluoxetine response, family history of major affective disorder (especially bipolar), unusual achievement, and hyperlexia in the children appear to define a homogeneous autistic subgroup. Bipolar disorder, unusual intellectual achievement, and autistic spectrum disorders cluster strongly in families and may share genetic determinants.
  • Immune activation is accompanied by induction of indoleamine (2,3)-dioxygenase (IDO), an enzyme which degrades tryptophan, a phenomenon which plays a role in the pathophysiology of major depression and post-natal depression and anxiety states. TRYCATs - tryptophan catabolites along the IDO pathway - such as kynurenine, kynurenic acid, xanthurenic acid, andquinolinic acid, have multiple effects, e.g. apoptotic, anti- versus pro-oxidant, neurotoxic versus neuroprotective, andanxiolytic versus anxiogenic effects. The aim of the present study was to study the immune effects of the above TRYCATS. Toward this end we examined the effects of the above TRYCATs on the LPS + PHA-induced production of interferon-gamma (IFNgamma), interleukin-10 (IL-10), andtumor necrosis factor-alpha (TNFalpha) in 18 normal volunteers. We found that the production of IFNgamma was significantly decreased by all 4 catabolites. Xanthurenic acid andquinolinic acid decreased the production of IL-10. Kynurenine, kynurenic acid, andxanthurenic acid, decreased the IFNgamma/IL-10 production ratio, whereas quinolinic acid increased this ratio. Kynurenic acid significantly reduced the stimulated production of TNFalpha. It is concluded that kynurenine, kynurenic acid, andxanthurenic acid have anti-inflammatory effects trough a reduction of IFNgamma, whereas quinolinic acid has pro-inflammatory effects in particular via significant decreases in IL-10. Following inflammation-induced IDO activation, some TRYCATs, i.e. kynurenine, kynurenic acid, andxanthurenic acid, exert a negative feedback control over IFNgamma production thus downregulating the initial inflammation, whereas an excess of quinolinic acid further aggravates the initial inflammation.
  • In a little more than 10 years, the kynurenine metabolites of tryptophan have emerged from their former position as biochemical curiosities, to occupy a prominent position in research on the causes and treatment of several major CNS disorders. The pathway includes two compounds, quinolinic acid and kynurenic acid, which are remarkably specific in their pharmacological profiles: one is a selective agonist at receptors sensitive to NMDA, whereas the other is a selective antagonist at low concentrations at the strychnine-resistant glycinemodulatory site associated with the NMDA receptor. It has been argued that these agents cannot be of physiological or pathological relevance because their normal extracellular concentrations, in the nanomolar range, are at least 3 orders of magnitude lower than those required to act at NMDA receptors. This is a facile argument, however, that ignores at least two possibilities. One is that both quinolinate and kynurenate may be present in very high concentrations locally at some sites in the brain that cannot be reflected in mean extracellular levels. Similar considerations apply to many neuroactive agents in the CNS. The fact that both compounds appear to be synthesised in, and thus emerge from, glial cells that are well recognised as enjoying a close physical and chemical relationship with some neurones in which the intercellular space may be severely restricted may support such a view. Certainly the realisation that NMDA receptors may not be fully saturated functionally with glycine would be consistent with the possibility that even quite low concentrations of kynurenate could maintain a partial antagonism at the glycine receptor. A second possibility is that there may be a subpopulation of NMDA receptors (or, indeed, for a quite different amino acid) that possesses a glycinemodulatory site with a much lower sensitivity to glycine or higher sensitivity to kynurenate, making it more susceptible to fluctuations of endogenous kynurenine levels. Whatever the specific nature of their physiological roles, the presence of an endogenous selective agonist and antagonist acting at NMDA receptors must continue to present exciting possibilities for understanding the pathological basis of several CNS disorders as well as developing new therapeutic approaches. An imbalance in the production or removal of either of these substances would be expected to have profound implications for brain function, especially if that imbalance were present chronically.(ABSTRACT TRUNCATED AT 400 WORDS)
  • AIM: Indoleamine 2,3-dioxygenase (IDO) catalyzation of tryptophan is the first rate-limiting step of the kynurenine pathway in the majority of tissues. The kynurenine pathway produces neurotoxic metabolites such as 3-hydroxykinurenine andquinolinic acid. IDO is inducible by the cytokine interferon-gamma (IFN-gamma) and has been proposed to mediate the sickness behavior of patients with infectious or other inflammatory diseases.To better understand the neuroendocrine component of cytokine induced sickness behavior we determined the effects of the pro-inflammatory cytokine IFN-gamma and the anti-inflammatory cytokine IL-10 on IDO expression in cells derived from the hypothalamic-pituitary-adrenal axis (HPA): GT1-7 hypothalamic, AtT-20 pituitary, and Y-1 adrenal cells. 2. METHODS: Reverse transcriptase polymerase chain reaction (RT-PCR) was performed to check the IDO expression from IFN-gamma and IL-10 treated cells such as GT1-7, AtT-20 and Y-1 cells. 3. RESULTS: We found that IFN-gamma induces IDO expression after 4 h treatment in GT1-7 and AtT-20 cells. IL-10 was also able to suppress IFN-gamma induced IDO expression in these cells. In Y-1 adrenal cells, IFN-gamma treatment had no effect on IDO expression. 4. CONCLUSIONS: Our results indicate that cytokines such as IFN-gamma and IL-10 are able to regulate IDO expression in cells of hypothalamic and pituitary origin. The ability of IL-10 to suppress IFN-gamma induced IDO expression implies that IL-10 has a putative neuroprotective role in the HPA axis. It can act at two levels, systemically by inhibiting sickness behavior-related Th1 cytokine synthesis and more centrally by inhibiting the kynurenine pathway.
  • Figure 2. In populations adequately exposed to old friends, the levels of priming of regulatory cells (Treg) are sufficient to suppress inappropriate inflammation (right side of large balance). When the priming of Treg is too low, the population is at risk of a variety of syndromes attributable to inadequate termination of inappropriate inflammatory responses. Some individuals have a genetic background and immunological history that make their Th1 mechanisms more likely to become pathologically uncontrolled, and these people are at risk of Th1-mediated conditions (type 1 diabetes, multiple sclerosis, Crohn’s disease). In other individuals it is the Th2 response that is most liable tolack of control, resulting in allergic disorders. A further category of individuals suffering from failure to terminate inflammation does not develop any gross pathology, but is susceptible to CNS effects of chronic cytokine exposure. The symptoms tend toward anxiety when these are Th2, and toward depression when these are Th1, IL-1 and TNF-α.It is relatively common, however, for the patients to have both the inflammatory disorder and the psychiatric disturbance.
  • Figure 1. Environmental organisms, that are part of human evolutionary history (‘old friends’), are recognized as harmless by pattern recognition receptors on DC. The DCmature into regulatory DC (DCreg) that drive regulatory T cell responses to the antigens of these organisms (Treg). The continuing presence of these antigens (left), eitherbecause they are inhaled into the lungs, present in the gut flora, ingested with food or water or resident as parasites that must be tolerated (such as microfilariae), leads tohigh continuous background release of regulatory cytokines, exerting bystander suppression of inflammatory responses and counteracting activation of the innate immunesystem. Meanwhile, the increased numbers of DCreg lead to increased processing by such DC of self antigens, gut content antigens and allergens (right). Therefore, thenumbers of Treg specific for these antigens are also increased, downregulating autoimmunity, IBD and allergies, respectively.
  • Retinoic acid, the oxidized form of vitamin A, has long beenviewed as important to both the innate and the acquired immunesystems. The transcription factor RORt, which if overexpressed,induces T-cell differentiation to Th17, belongs to a retinoidnuclear receptor superfamily (17). Kim (29) provides an excellentreview of published research data referencing the regulationof regulatory T cells by retinoids. Recently, researchers haverecognized retinoic acid's regulation in immune responses (17,30).It was shown to suppress memory cell IFN- production and increaseIL-4 secretion (17). Mucida and colleagues (30) completed astudy that measured the effect of retinoic acid on Th17 celldevelopment both in vitro and in vivo. In each case, retinoicacid greatly reduced RORt expression, which resulted in a measurablereduction of Th17 mucosal T cells. Further, all-trans retinoicacid (ATRA) has been reported to inhibit Th17 production andto promote FoxP3 expression, thereby affecting the Th17/Tregpolarization (17). Retinoic acid's importance to the immunesystem cannot overshadow its promotion of glial and neuronaldifferentiation. Within the CNS (central nervous system), retinoicacid has been shown to enhance axonal regeneration of differentiatedretinal ganglion cells and peripheral sensory neurons (31).Therefore, supplementation with low doses of vitamin A may beuseful for treatment of neuroimmune disorders.
  • The wet surfaces, immunity and autism

    1. 1. The Wet Surfaces, The Immune System & The Brain. Why And How they May Affect Behaviour And Function In People With Autism Michael Ash BSc (Hons) DO ND F.DipION
    2. 2. • The aetiology of ASD is largely unknown, but genetic, environmental, immunological, and neurological factors are thought to play a role in the development of ASD. • Recently, focused on the connections between the immune system and the nervous system. • These neuroimmune interactions begin early during embryogenesis and persist throughout an individual’s lifetime, with successful neurodevelopment contingent upon a normal balanced immune response Ashwood P, Wills S, Van de Water JThe immune response in autism: a new frontier for autism research. . J Leukoc Biol. 2006 Jul;80(1):1-15. Epub 2006 May 12. Review. View Paper
    3. 3. Genetic Determinism • Your genome, or any part of it, is not you • The concept that our DNA sequence our genome- does not equal or predict our destiny has been extremely difficult for some geneticists to accept. • We don’t even have a good idea how many genes there are, let alone how these genes work with each other and the environment to orchestrate human development. • LL McCabe & ERB McCabe. DNA Promise and Peril. University of California Press; 1 edition (1 Mar 2008)
    4. 4. Oct 2010
    5. 5. • Autism may in fact be a systemic disorder with connections to abnormal immune responses. • M Careaga, J Van de Water, and P AshwoodImmune dysfunction in autism: a pathway to treatment. Neurotherapeutics, July 1, 2010; 7(3): 283-92. View Abstract • Immunological factors have provided more support for a probable immunological process or for processes that may play a role in the acquisition of an autistic condition. • MG Chez and N Guido-Estrada. Immune therapy in autism: historical experience and future directions with immunomodulatory therapy. Neurotherapeutics, July 1, 2010; 7(3): 293-301. View Abstract • ....significantly shifted cytokine profiles in ASD. These findings suggest that ongoing inflammatory responses may be linked to disturbances in behaviour. • P Ashwood, P Krakowiak, I Hertz-Picciotto, R Hansen, I Pessah, and J Van de WaterElevated plasma cytokines in autism spectrum disorders provide evidence of immune dysfunction and are associated with impaired behavioral outcome. Brain Behav Immun, August 9, 2010; View Abstract
    6. 6. About to be Published data --- • altered activation profile for T cells in ASD. Overall these data indicate significantly altered adaptive cellular immune function in children with ASD that may reflect dysfunctional immune activation, along with evidence that these perturbations may be linked to disturbances in behaviour and developmental functioning
    7. 7. Traditional View of Immune System • Seen as a system of organs, molecules and tissues that defend us from disease by eliminating bad guys. • This promotes the dichotomy model good vs bad • There have been changes to the model of self/nonself and they are relevant to autism.
    8. 8. Changing Models • Immunology is no longer good Vs bad – • it is the homeostasis of the superorganism! • Complexity is required to apply therapy well
    9. 9. Immune Concepts Evil- Microbiota Non-Self Evil- Altered Self Pathogens Good – Self Tolerance/ ignorance Good- Normal Self Mutualists Continuum Model Ancestral Dualistic Model Modern Dualistic Model
    10. 10. • The continuum model states that the perceived duality of mutualistic and pathogenic microbes, normal and altered self, and regulatory or inflammatory immunity, represents extremes of a continuous reality. • Microbes can express different levels of mutualistic or pathogenic properties and these levels can vary during interaction with the host. • Similarly, the state of self and of immune responses can navigate between well-described extremes, and the most likely states are combination of these extremes.
    11. 11. Danger Good – Regulation/Tolerance Mutualists/Normal self Evil- Inflammation Pathogens/A ltered Self Strangers Pathological Inflammation Friends Physiological Inflammation Health of Host Health of Local tissue Timing Genes Nutri- genomics Nutrition Modern Dualism Continuum Model
    12. 12. Mutualist Pathogen Regulation/Tolerance Inflammation Development of Adaptive Immunity (AgS) Control by Innate Immunity (MAMPs) Lymphoid Tissues Lymphocyte Sub Sets Lymphocyte Memory Microbes Immune Responses IL-2 IL-10 TGFB RA IL-23 IL-12 IL-1 TNFα
    13. 13. • Cytokines • Inflammation • Old Friends • Mucosal Tolerance Hygiene Hypothesis • Kyneurenines • Enzymes • Cytokines • Stressors • IDO Gastro- Neuro- degeneration hypothesis ASD
    14. 14. Wet Surfaces -Massive Area of Contact – GI. Mucosal surface area 300m2 • Respiratory mucosa 100m2 • Skin 1.5 m2 • Thickness 0.03mm (1/2 width of 60gsm) ‘Tightly regulated mucosal Immunity is essential to maintain health’
    15. 15. “the mucosa is directly exposed to the external environment and taxed with antigenic loads consisting of commensal bacteria, dietary antigens, and viruses at far greater quantities on a daily basis than the systemic immune system sees in a lifetime”. Mayer L. Mucosal immunity. Pediatrics. 2003 Jun;111(6 Pt 3):1595-600.
    16. 16. Confusion • “People tend to get the immune system the wrong way around; • We’re so focussed on the immune system responding to things, that we forget that 99.99% of the time, its job is NOT to respond to things. • There’s you, your breakfast and your gut, for a start. That’s a lot of stuff not to respond to”. Handley C. Should auld acquaintance be forgot… EMBO Reports Vol 5, No 12, 2004
    17. 17. Mucosal Immune System For Host Protection The mucosal immune system consists of an integrated network of: Macrophages and dendritic cells, T lymphocytes and their cytokines play a key role in orchestrating a specific mucosal immune response. An uncontrolled mucosal immune system may lead to immunologic diseases such as allergy and hypersensitivity.
    18. 18. Humans Are Superorganisms Cells 1 Trillion human cells 10 Trillion bacterial in and on 10 x ratio DNA Humans have approx 25,000 genes and bacteria have an estimated 100 times more. Who has more genes – grape, chicken or human?
    19. 19. Our relationship extends Beyond the shared Environmental benefits – and extends to: •Nutrient harvesting •Gene expression •Mood regulation •Illness prevention •Illness resolution •Weight management •Immune control This raises the possibility that the mammalian immune system, which seems to be designed to control microorganisms, is in fact controlled by microorganisms. Nature Reviews Immunology 9, 313-323 (May 2009) | doi:10.1038/nri2515 The gut microbiota shapes intestinal immune responses during health and disease “We respond to our microbiota from birth to death” Swidinki,A
    20. 20. Evidence Is Accumulating • to suggest that gut microbes (microbiota) may be involved in neural development and function, both peripherally in the enteric nervous system and centrally in the brain.
    22. 22. NKc
    24. 24. Immune- Cytokines - Autism • Immunological findings in autism. HH Cohly and A Panja Int Rev Neurobiol, January 1, 2005; 71: 317- Link • Activation of the inflammatory response system in autism. J Croonenberghs, M Maes Neuropsychobiology, January 1, 2002; 45(1): 1-6. Link • Elevated cytokine levels in children with autism spectrum disorder. CA Molloy, et al. J Neuroimmunol, March 1, 2006; 172(1-2): 198-205. Link • It is hypothesised that increased production of proinflammatory cytokines could play a role in the pathophysiology of autism.
    25. 25. Cytokines - Autism 2010 • In conclusion, using larger number of participants than previous studies, we report significantly shifted cytokine profiles in ASD. These findings suggest that ongoing inflammatory responses may be linked to disturbances in behaviour • Ashwood P, Krakowiak P, Hertz-Picciotto I, Hansen R, Pessah I, Van de Water J. Elevated plasma cytokines in autism spectrum disorders provide evidence of immune dysfunction and are associated with impaired behavioral outcome. Brain Behav Immun. 2010 Aug 10 View Abstract
    26. 26. The Innate Immune Response In Autism
    27. 27. • Dysregulated innate immune responses in young children with autism spectrum disorders: their relationship to gastrointestinal symptoms and dietary intervention.H Jyonouchi, et al. Neuropsychobiology, January 1, 2005; 51(2): 77-85. • Impact of innate immunity in a subset of children with autism spectrum disorders: a case control study. H Jyonouchi, et al. J Neuroinflammation, January 1, 2008; 5: 52. • Proinflammatory and regulatory cytokine production associated with innate and adaptive immune responses in children with autism spectrum disorders and developmental regression. H Jyonouchi, et al J Neuroimmunol, November 1, 2001; 120(1-2): 170-9. • Intrinsic defects of innate immune responses in GI(+) ASD children but not in NFH* or GI(-) ASD children, suggesting a possible link between GI and behavioural symptoms mediated by innate immune abnormalities. • TNFα, IL-1, IL-6, • IL-10, TGF-β • *Non Allergic Food Hypersensitivity
    28. 28. Regulatory Cytokine Transforming Growth Factor Beta-1 • Decreased serum levels of transforming growth factor-beta1 in patients with autism. K Okada, et al. Prog Neuropsychopharmacol Biol Psychiatry, Jan 2007; 31(1): 187-90. • Decreased transforming growth factor beta1 in autism: a potential link between immune dysregulation and impairment in clinical behavioural outcomes. P Ashwood, et al. J Neuroimmunol, Nov 2008; 204(1-2): 149-53. • These findings suggest that decreased levels of TGF-beta1 may be implicated in the pathophysiology of autism. • Such that lower TGF beta 1 levels were associated with lower adaptive behaviours and worse behavioural symptoms.
    29. 29. Autism & The Gut
    30. 30. Comparisons of the prevalence of gastrointestinal symptoms between autistic children and their healthy siblings
    31. 31. • Reasons to consider that microorganisms may be involved in late‐onset autism include • onset of the disease often follows antimicrobial therapy, • gastrointestinal symptoms are common at onset and often persist, • antimicrobials (e.g., oral vancomycin) may lead to a clear‐cut response and relapse may occur when the vancomycin is discontinued, and • some patients have responded to several courses of vancomycin and relapsed each time it was discontinued. Clinical Infectious Diseases 2002; 35(Suppl 1):S6–16 Link
    32. 32. Regulation Inflammation Symbionts Commensals Pathobionts Immunological Equilibrium Regulation Inflammation Immunological Disequilibrium Dysbiosis/Pathogens The gut microbiota shapes intestinal immune responses during health and disease June L. Round & Sarkis K. Mazmanian. Nature Reviews Immunology 9, 313-323 (May 2009)
    33. 33. N C Occludin Cgn PILT bCtnn ZONAB Epithelial Tight Junctions F-Actin F-Actin Cell Adhesion Cell Adhesion Cell Migration Cytoskeletal Organization Vesicular Trafficking Cell Cycle Progression Intracellular Membrane Transport and Exocytosis Vesicular Trafficking Cytoskeletal Organization Exocyst Complex Assembly of v-SNARE and t-SNARE Complexes Cell Proliferation Cell Adhesion G1/S Phase Transition mRNA Cleavage and Processing Cell Adhesion and Apoptosis Epithelial Cell Proliferation and Differentiation Non-CiliatedEpithelial Cell CiliatedEpithelial Cell RhoA PathwayActin Based Motility Actin Nucleation and Branching ZONAB CDK4 AP-1 CEBP Paracellular Signaling RhoA SAFB ZO2 Ras Stx4 mLGL Myosin Crb1,3 Afadin SMURF1 PTEN HSF1 CSTF aCtnn Rab GTPases PIP3 PIP2 Cell Adhesion Ga12 Ga12 ARP 2/3 PAR-3 Rac1 TGFbR C C JAMs JAMs TIAM1 PAR-3 CDC42 MAGI 2,3 GSK3ILK JNK Sympk TNFaR TNFa p130 CAS Integrins MAGI1 N C Claudins N C Occludin N C Claudins PKA VAP33 Akt Plasmamembrane 2009 C
    34. 34. Leaky Gut and Autism • CONCLUSIONS: The results obtained support the leaky gut hypothesis and indicate that measuring IP could help to identify a subgroup of patients with autism who could benefit from a gluten-free diet. The IPT alterations found in first-degree relatives suggest the presence of an intestinal (tight-junction linked) hereditary factor in the families of subjects with autism. • de Magistris L, Familiari V, Pascotto A, Sapone A, Frolli A, Iardino P, Carteni M, De Rosa M, Francavilla R, Riegler G, Militerni R, Bravaccio C.Alterations of the Intestinal Barrier in Patients With Autism Spectrum Disorders and in Their First-degree Relatives. J Pediatr Gastroenterol Nutr. 2010 Jul 28 View Abstract
    35. 35. SIgA Vs Autism
    36. 36. • The production of immunoglobulin A (IgA) in mammals exceeds all other isotypes, and it is mostly exported across mucous membranes. 60mg/kg daily • It is now clear that SIgA can function in high- affinity modes for neutralisation of toxins and pathogenic microbes, and as a low-affinity system to contain the dense commensal microbiota within the intestinal lumen. • A J Macpherson, K D McCoy, F-E Johansen and P Brandtzaeg. The immune geography of IgA induction and function. Mucosal Immunology (2008) 1, 11– 22. doi:10.1038/mi.2007.6 Link
    37. 37. SIgA • When depleted the mucosal barrier management of antigen is compromised and microbial translocation (LPS) can occur, a consequence of which: • Increased pro inflammatory cytokines • Increased IL-17
    38. 38. Conditions That Can Change The Level Of Secretory IgA In Oral Fluid • Increased SIgA Level • Acute stress • Some medications • Orophanrygeal carcinoma • Chronic oral infection • Chronic GI infection • Heavy smoking • Alcoholism • Periodontitis • Dental plaque accumulation • Intestinal barrier dysfunction • Multiple Myeloma • Decreased SIgA Level • Chronic stress (frustration) • Some medications • Adrenal insufficiencies • Bacterial colonisation on molar surfaces • Recurrent tonsillitis • Adenoid hyperplasia • Cutaneous candidiasis • Asthmatic with recurrent respiratory tract infection • Intestinal barrier dysfunction • Nutritional deficiencies • Recurrent herpes infection • Coeliac, Crohn’s, Ulcerative colitis
    39. 39. SIgA Levels In Clinic Tests M.Ash 2000 SIgA 10 100 200 300 400 500 600 700 1 2 4 6 8 10 12 14 16 18 20 22 24 26 28 SIgA 1 28 Children with Autism 26♂2♀ ave age 7 Ug/ml
    40. 40. Serotonin & Autism
    41. 41. Serotonin Vs Autism • Studies demonstrating impaired serotonin synthesis in the brains of autistic individuals • Chugani DC, Muzik O, Behen M, Rothermel R, Janisse JJ, Lee J, Chugani HT. Developmental changes in brain serotonin synthesis capacity in autistic and nonautistic children. Ann Neurol. 1999;45:287-295. FULL TEXT • Chugani DC, Muzik O, Rothermel R, Behen M, Chakraborty P, Mangner T, da Silva EA, Chugani HT. Altered serotonin synthesis in the dentatothalamocortical pathway in autistic boys. Ann Neurol. 1997;42:666-669. FULL TEXT • -a worsening of repetitive behaviours after tryptophan depletion… • McDougle CJ, Naylor ST, Goodman WK, Volkmar FR, Cohen DJ, Price LH. Acute tryptophan depletion in autistic disorder: a controlled case study. Biol Psychiatry. 1993;33:547-550. FULL TEXT
    42. 42. Serotonergic Neurons • are generated early in brain development and establish extensive cortical and subcortical connections. • Serotonin regulates growth cone motility, synaptogenesis, synaptic plasticity, and the development and activity of multiple neuronal subtypes. • Sodhi MS, Sanders-Bush E. Serotonin and brain development. Int Rev Neurobiol. 2004;59:111-174. Link
    43. 43. SSRI Study Response • 129 children – 2-8 yrs of age • 17% excellent, 52% good, 31% fair to poor • DeLong GR, Ritch CR, Burch SA: Fluoxetine response in children with autistic spectrum disorders: correlation with familial major affective disorder and intellectual achievement. Devel Med Child Neurol 2002; 44(10):652–659 Link 1/2
    44. 44. Serotonin Vs Indolamine 2,3 • It is concluded that the (TRYCATS) kynurenine, kynurenic acid, and xanthurenic acid have anti-inflammatory effects trough a reduction of IFNgamma, • Whereas quinolinic acid has pro-inflammatory effects in particular via significant decreases in IL-10. • M Maes, I Mihaylova, MD Ruyter, M Kubera, and E Bosmans The immune effects of TRYCATs (tryptophan catabolites along the IDO pathway): relevance for depression - and other conditions characterized by tryptophan depletion induced by inflammation. Neuro Endocrinol Lett, December 1, 2007; 28(6): 826-31 LINK
    45. 45. Indolamine (IDO) TRYCATS & Autism
    46. 46. TDO IDO Tryptophan IFN γ TNFα Kyneurenine 3-Hydroxykynurenine Quinolinic acid Nicotinamide NMDA Receptor Kyneurenic acid 5 - Hydroxytryptophan Serotonin P38MAPk + + Stressor Immune Induction Of Neurotoxins BBB Liver 95% Brain 5%> IDO – Indolamine 2,3 dioxygenase TDO – Tryptophan 2,3 dioxygenase N-methyl-D-aspartate
    47. 47. Quinolinate/Kyneurenate • An imbalance in the production or removal of either of these substances would be expected to have profound implications for brain function, especially if that imbalance were present chronically. • Identified in Organic Acid test • Stone TW. Neuropharmacology of quinolinic and kynurenic acids. Pharmacol Rev. 1993 Sep;45(3):309-79. Link
    48. 48. Quin leads acutely to neuronal death or chronically to neuronal function by at least 6 mechanisms. 1. Activation of the NMDA receptor in pathophysiological concentrations 2. Increase glutamate release by neurons, inhibition of glutamate uptake by astrocytes, decrease in glutamine synthetase activity. 3. Lipid peroxidation of the membrane 4. Quin can potentiate its own toxicity and that of other excitotoxins (NMDA and glutamate) in the context of energy depletion (mitochondrial dysfunction)
    49. 49. 5. Quin induces iNOS (astrocyte) and nNOS (neuron) leading to over production of NO (= Oxidative stress) 6. Destabilisation of the cellular cytoskeleton Nady Braidy, Ross Grant, Seray Adams, Bruce J. Brew and Gilles J. Guillemin Mechanism for Quinolinic Acid Cytotoxicity in Human Astrocytes and Neurons Neurotoxicity Research Volume 16, Number 1, 77-86, DOI: 10.1007/s12640-009- 9051-z View Abstract
    50. 50. • Proinflammatory cytokines induce IDO under stress, promote the KYN pathway, deprive the 5-HT pathway of TRP, and reduce 5-HT synthesis. The resultant decrease in 5-HT production may relate to the monoamine hypothesis of major depression. • The hippocampal atrophy that appears in chronic depression may be associated with imbalances in KP neurotoxic/neuroprotective metabolites.
    51. 51. Quinolinate acts as a most potent endogenous excitotoxin to neurons.
    52. 52. Cytokine regulation of tryptophan metabolism in the hypothalamic- pituitary-adrenal (HPA) axis: implications for protective and toxic consequences in neuroendocrine regulation. • Our results indicate that cytokines such as IFN- gamma and IL-10 are able to regulate IDO expression in cells of hypothalamic and pituitary origin. The ability of IL-10 to suppress IFN-gamma induced IDO expression implies that - • IL-10 has a putative neuroprotective role in the HPA axis. It can act at two levels, systemically by inhibiting sickness behaviour-related Th1 cytokine synthesis and more centrally by inhibiting the kynurenine pathway. • Tu H, Rady PL, Juelich T, Smith EM, Tyring SK, Hughes TK.. Cytokine regulation of tryptophan metabolism in the hypothalamic-pituitary-adrenal (HPA) axis: implications for protective and toxic consequences in neuroendocrine regulation. Cell Mol Neurobiol. 2005 Jun;25(3-4):673-80 Link Cytokine regulation of tryptophan metabolism in the hypothalamic- pituitary-adrenal (HPA) axis: implications for protective and toxic consequences in neuroendocrine regulation.
    53. 53. Bacteria – Friend and Foe & Autism
    54. 54. Bacterial Species In The Genomic Era • RECENT FINDINGS: • Upwards of 40,000 bacterial species are estimated to comprise the collective gastrointestinal microbiome, most of which have not been characterised by culture. • Frank DN, Pace NR. Gastrointestinal microbiology enters the metagenomics era. Curr Opin Gastroenterol. 2008 Jan;24(1):4- 10.
    55. 55. Yet … • About 200 different bacterial species are known to cause human disease. • Mascie-Taylor, C. G. & Karim, E. The burden of chronic disease. Science 302, 1921−1922 (2003). Article
    56. 56. Treg - Immunommodulation Low ratio of Treg to effector Tcells Normal ratio of Treg to effector T cells Effective levels of sIgA, IL-10 & TGF-β to control adverse inflammation ASD Anxiety and Allergies IL-10, TGF-β, sIgA Th1 Th2 (IFN-γ, IL-1, TNF-α) (IL-4, IL-5, IL-13) Th17 Retinoic Acid 59
    57. 57. Probiotics Old friends (innocuous environmental microorganisms, helminths) Treg T Bystander suppression Old friends IL-10 TGF-β Retinoic Acid TregT IL-10 TGF-β Retinoic Acid Specific suppression Self/gut contents allergens T cells Immature DC Regulatory DC (DCreg) sIgA sIgA Reduction of peripheral/CNS inflammation 60
    58. 58. Probiotics May Ease Anxiety: Pilot study • “Two months of supplementation with the bacterial strain from a sachet was associated with a decrease in anxiety symptoms.”
    59. 59. 62 For instance a specific part of the microbiota has been shown to cooperate With the development of regulatory instead of the inflammatory IL-17 producing T helper cells in the small intestine. Ivanov II, Frutos Rde L, Manel N, Yoshinaga K, Rifkin DB, Sartor RB, Finlay BB, Littman DR Specific microbiota direct the differentiation of IL-17-producing T-helper cells in the mucosa of the small intestine.Cell Host Microbe. 2008 Oct 16;4(4):337-49. View Full Paper
    60. 60. 63 Tolerance Vit A TH-17 Treg TGF-β & Autism
    61. 61. Retinoic Acid • Is autism a G-alpha protein defect reversible with natural vitamin A? MN Megson Med Hypotheses, June 1, 2000; 54(6): 979-83. • Symptomatic Vitamin A and D Deficiencies in an Eight-Year-Old With Autism Joseph H. Clark, Donna K. Rhoden, and Denece S. Turner JPEN J Parenter Enteral Nutr, May 1993; 17: 284 – 286 • Serum values of cytokines in children with vitamin A deficiency disorders JY Leal, HV Castejon, T Romero, P Ortega, G Gomez, D Amaya, and J Estevez Invest Clin, September 1, 2004; 45(3): 243-56 • Lack of Vitamin A - Reduced IL-10 and reduced Treg and increased TH-17 IL-17 production
    62. 62. TGFβ • TH17 • IL-17 • IL-6 • IL-21 • Retinoic Acid • IL-10 The Role of Th17 in Neuroimmune Disorders: A Target for CAM Therapy. Part III Aristo Vojdani, Jama Lambertand,Gottfried Kellermann eCAM Advance Access published online on July 21, 2009 eCAM, doi:10.1093/ecam/nep064 •In each case, retinoicacid greatly reduced RORt expression •which resulted in a measurable reduction of Th17 mucosal T cells. •IL-17-producing CD4(+) T-helper cells (Th17) contribute to chronic autoimmune inflammation in the brain
    65. 65. NEURODEGENERATION ALZHEIMERS SENILITY AUTISM ROS/NS IL-1β, TNFα CORTISOL INDUCED ATROPHY DEPRESSION Imbalanced Cytokine Induction and Binding Is ‘Regressive Autism’ an Immune Mediated Disorder
    66. 66. What Can You Do?
    67. 67. Antioxidants Inhibit IDO • Thomas SR J Immunol. 2001 May 15;166(10):6332-40. • Antioxidants inhibit indoleamine 2,3- dioxygenase in IFN-gamma-activated human macrophages: post translational regulation by pyrrolidine dithiocarbamate. • Glutathione (NAC)
    68. 68. Clinical Strategies • Assess patient as potential GIT candidate • Improve daily nutrition • Include probiotics (Human Strain) • Build Beneficial Bio Film • Remove non beneficial Bio Films • Include PUFA’s for brain function and improving Bacterial/Immunological cross talk &Treg • Pro-biotics do not appear to have any contraindications with medications involved in mood disorders.
    69. 69. Supermarket dairy shelves are filled with yogurt products containing live cultures of 'probiotic' bacteria — species that live in the human gut and are proposed to deliver health benefits when eaten at high levels. Three probiotic species seem to alter gene expression in the gut lining of volunteers consuming the cultures. The effect was similar to that of drugs for conditions including inflammation and high blood pressure. Michiel Kleerebezem at NIZO Food Research in Ede, the Netherlands, and his co-workers analysed the gene-expression profiles of tissue taken from the small intestinal inner lining of seven healthy volunteers who had eaten a placebo and three probiotic cultures — Lactobacillus acidophilus, L. casei and L. rhamnosus — in a random order. The altered gene-expression profiles resembled those associated with the regulation of immune responses, cell growth, metabolism and even wound repair
    70. 70. Protocol • Saccharomyces Boulardii 150-600mg • Lactobacillus GG 30-609 CFU • Lactobacillus Caseii 20-609 CFU • Lactobacillus Paracaseii 20-609 CFU • Lactobacillus, plantarum, rhamnosus, salivarius 20-609 CFU • Bifido-Bifidus 20-609 CFU • DHA > EPA (CLO) concentrate 2-4gms (GPR120) • Vit A 5000 -12000 iu & Vit D 6-12000iu
    71. 71. Current Understood Uses of PB’s 21 3 4
    72. 72. • The gut maintains an extensive and highly active immune system, environmental factors can induce dysregulation of the mucosal immune system and potentially damage tissue locally and systemically. • In the healthy gut, Th1,Th2,Th17 responses are carefully managed by regulatory T cells (CD4+CD25+) expressing IL- 10 and TGF-β. • Depletion of IL-10- and TGF-β-producing regulatory T cells, or homing of CD4+CD25RBHIGH T cells in the GI tissue of children with autism, may be responsible for GI pathology reported by different investigators in autism. Conclusions
    73. 73. Conclusions • Regulatory T cells and TGF-β production measured in the blood of children with autism are inconsistent. T cell subtypes are different TH1> Adversity. TH2 < ASD symptoms • Immune function abnormalities, in particular, low natural killer cell activity, low glutathione and abnormal cytokine production, is part of the illness in autism. • Abnormal levels of neurotransmitters such as serotonin, Indolamine (Quin/Kyn), dopamine, epinephrine, and norepinephrine are detected in children with autism.
    74. 74. • A gluten- and casein-free diet or a low antigen diet, with clean omega-3 and omega 6 oils, strain specific probiotics, and moderate to high doses of vitamin- A - can be extremely helpful towards mucosal immune recovery for a ‘subgroup’ of children with autism. • A low risk to reward strategy, and can be observed to have effect in weeks vol
    75. 75. The End Thank You