Atopic diseases and inflammation of the brain in the pathogenesis of autism spectrum disorders

Abstract

Autism spectrum disorders (ASDs) affect as many as 1 in 45 children and are characterized by deficits in sociability and communication, as well as stereotypic movements. Many children also show severe anxiety. The lack of distinct pathogenesis and reliable biomarkers hampers the development of effective treatments. As a result, most children with ASD are prescribed psychopharmacologic agents that do not address the core symptoms of ASD. Autoantibodies against brain epitopes in mothers of children with ASD and many such children strongly correlate with allergic symptoms and indicate an aberrant immune response, as well as disruption of the blood-brain barrier (BBB). Recent epidemiological studies have shown a strong statistical correlation between risk for ASD and either maternal or infantile atopic diseases, such as asthma, eczema, food allergies and food intolerance, all of which involve activation of mast cells (MCs). These unique tissue immune cells are located perivascularly in all tissues, including the thalamus and hypothalamus, which regulate emotions. MC-derived inflammatory and vasoactive mediators increase BBB permeability. Expression of the inflammatory molecules interleukin (IL-1β), IL-6, 1 L-17 and tumor necrosis factor (TNF) is increased in the brain, cerebrospinal fluid and serum of some patients with ASD, while NF-kB is activated in brain samples and stimulated peripheral blood immune cells of other patients; however, these molecules are not specific. Instead the peptide neurotensin is uniquely elevated in the serum of children with ASD, as is corticotropin-releasing hormone, secreted from the hypothalamus under stress. Both peptides trigger MC to release IL-6 and TNF, which in turn, stimulate microglia proliferation and activation, leading to disruption of neuronal connectivity. MC-derived IL-6 and TGFβ induce maturation of Th17 cells and MCs also secrete IL-17, which is increased in ASD. Serum IL-6 and TNF may define an ASD subgroup that benefits most from treatment with the natural flavonoid luteolin. Atopic diseases may create a phenotype susceptible to ASD and formulations targeting focal inflammation of the brain could have great promise in the treatment of ASD.

OPEN

Citation: Transl Psychiatry (2016) 6, e844; doi:http://dx.doi.org/10.1038/tp.2016.77

Web End =10.1038/tp.2016.77

http://www.nature.com/tp

Web End =www.nature.com/tp

REVIEW

Atopic diseases and inammation of the brain in the pathogenesis of autism spectrum disorders

TC Theoharides1,2,3,4, I Tsilioni1, AB Patel1,2 and R Doyle5

Autism spectrum disorders (ASDs) affect as many as 1 in 45 children and are characterized by decits in sociability and communication, as well as stereotypic movements. Many children also show severe anxiety. The lack of distinct pathogenesis and reliable biomarkers hampers the development of effective treatments. As a result, most children with ASD are prescribed psychopharmacologic agents that do not address the core symptoms of ASD. Autoantibodies against brain epitopes in mothers of children with ASD and many such children strongly correlate with allergic symptoms and indicate an aberrant immune response, as well as disruption of the bloodbrain barrier (BBB). Recent epidemiological studies have shown a strong statistical correlation between risk for ASD and either maternal or infantile atopic diseases, such as asthma, eczema, food allergies and food intolerance, all of which involve activation of mast cells (MCs). These unique tissue immune cells are located perivascularly in all tissues, including the thalamus and hypothalamus, which regulate emotions. MC-derived inammatory and vasoactive mediators increase BBB permeability. Expression of the inammatory molecules interleukin (IL-1), IL-6, 1 L-17 and tumor necrosis factor (TNF) is increased in the brain, cerebrospinal uid and serum of some patients with ASD, while NF-kB is activated in brain samples and stimulated peripheral blood immune cells of other patients; however, these molecules are not specic. Instead the peptide neurotensin is uniquely elevated in the serum of children with ASD, as is corticotropin-releasing hormone, secreted from the hypothalamus under stress. Both peptides trigger MC to release IL-6 and TNF, which in turn, stimulate microglia proliferation and activation, leading to disruption of neuronal connectivity. MC-derived IL-6 and TGF induce maturation of Th17 cells and MCs also secrete IL-17, which is increased in ASD. Serum IL-6 and TNF may dene an ASD subgroup that benets most from treatment with the natural avonoid luteolin. Atopic diseases may create a phenotype susceptible to ASD and formulations targeting focal

INTRODUCTION

Autism spectrum disorders (ASDs) are pervasive neurodevelop-mental disorders characterized by decits in communication and social interactions, as well as the presence of stereotypic behaviors.13 Numerous gene mutations have been identied in patients with ASD, but no direct link has so far been uncovered except for a small percentage of cases associated with Tuberous Sclerosis, Fragile X syndrome, Rett syndrome and PTEN deciency.4,5 As a result, even though there are a number of genetically-engineered mice with phenotypes resembling autism,6 they do not adequately reect ASD and there is an urgent need for appropriate animal models of ASD.7 In fact, mouse models are increasingly considered unreliable with respect to inamma-tory human diseases.8 We recently reported that a small number of bull terriers develop symptoms consistent with autism and have increased serum neurotensin (NT) and corticotropin-releasing hormone (CRH), also found to be elevated in children with ASD.9

ASD may affect as many as 1 in 45 children in the USA,10 but the

global prevalence is still under-recognized.11 The lack of reliable biomarkers12 and specic pathogenesis,13 as well as the existence

inammation of the brain could have great promise in the treatment of ASD.

Translational Psychiatry (2016) 6, e844; doi:http://dx.doi.org/10.1038/tp.2016.77

Web End =10.1038/tp.2016.77 ; published online 28 June 2016

of subgroups or comorbidities14 (Table 1), makes the development of specic treatments and conducting clinical studies difcult.13 As

a result, child and adolescent outpatient mental health services in the USA have increased considerably.15 Moreover, the annual economic burden for ASD was recently estimated at $268 billion for 2015 and is projected to reach $416 billion in 2025.16

A number of perinatal allergic, genetic, environmental, immune and infectious factors may increase the risk of or contribute to the pathogenesis of ASD1719 (Table 2). These could act through activation of a unique tissue immune cell, the mast cell (MC).20,21

MCs derive from bone marrow progenitors and mature in tissues depending on microenvironmental conditions.22 In addition to histamine, stimulated MCs secrete other vasoactive and proinammatory mediators such as the preformed kinins and proteases, as well as the de novo synthesized leukotrienes, prostaglandins, chemokines (CCXL8, CCL2), cytokines (interleukin(IL)-4, IL-6, IL-1, tumor necrosis factor (TNF)) and vascular endothelial growth factor (VEGF).20

MCs are not only considered critical for the development of allergic reactions,20 but also for immunity22 and inammation.23

1Molecular Immunopharmacology and Drug Discovery Laboratory, Department of Integrative Physiology and Pathobiology, Tufts University School of Medicine, Boston, MA, USA;

2Sackler School of Graduate Biomedical Sciences, Program in Cell, Molecular and Developmental Biology, Tufts University, Boston, MA, USA; 3Department of Internal Medicine, Tufts University School of Medicine and Tufts Medical Center, Boston, MA, USA; 4Department of Psychiatry, Tufts University School of Medicine and Tufts Medical Center, Boston, MA, USA and 5Department of Child Psychiatry, Harvard Medical School, Massachusetts General Hospital and McLean Hospital, Boston, MA, USA. Correspondence: Professor TC Theoharides, Molecular Immunopharmacology and Drug Discovery Laboratory, Department of Integrative Physiology and Pathobiology, Tufts University School of Medicine, 136 Harrison Avenue, Suite J304, Boston, MA 02111, USA.

E-mail: mailto:[email protected]

Web End [email protected] Received 13 January 2016; revised 23 February 2016; accepted 17 March 2016

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Table 1. ASD comorbidities or subgroups

ADDADHDAtopic diseasesFood intolerance Gastrointestinal symptoms Mitochondrial dysfunction PANDASPTEN mutationsSeizures

Abbreviations: ADD, attention decit disorder; ADHD, attention decit hyperactivity disorder; ASD, autism spectrum disorder; PANDAS, pediatric autoimmune neuropsychiatric disorders associated with streptococcal infections; PTEN, phosphatase and tensin homolog.

Table 2. Perinatal conditions increasing the risk of ASD

Strong evidenceAllergiesAsthmaBrain autoantibodies Brain hemorrhage InfectionLow birth weight Obesity Preeclampsia Prematurity PsoriasisStress

Moderate evidenceCesarean section with general anesthesia Environmental toxin exposureOxytocin, prolonged use for labor induction Psychotropic medication useSexual abuse

Abbreviation: ASD, autism spectrum disorder.

In fact, many studies have reported that allergic diseases in preschoolers are strongly associated with psychological and behavioral problems.24 We had proposed that MC-derived mediators could disrupt the bloodbrain barrier (BBB) and cause 'allergy of the brain25 or focal encephalitis,26 thus contributing to the pathogenesis of ASD.26,27 A number of recent reviews have

now conrmed and expanded on these ndings.28,29

MATERNAL HEALTH, PREMATURITY AND LOW BIRTH WEIGHT ARE LINKED TO INCREASED RISK OF ASD

Obesity during gestation has been strongly associated with prematurity and low birth weight.30,31 Obesity is considered as an inammatory state32 and has been associated with activation of MCs.33,34 Moreover, MCs secrete leptin35 and its deciency switches MC to an anti-inammatory phenotype.36 Leptin is increased both in obesity37 and ASD.38 Premature births account for about 15% of all births in the USA and premature infants (32 36 weeks) make up most of the increased rate of prematurity.39

Such infants are at risk for neurologic injury40,41 associated with decreased attention, increased anxiety, as well as social interaction and learning difculties.42

A retrospective study reported that children o33 weeks gestation were associated with a twofold higher risk of ASD.43

One prospective study found that 26% very low birth weight (o1500 g) infants ((n = 91), mean age of 22 months) developed

ASD.44 There was a higher risk of infantile autism among children

with low birth weight especially in mothers 435 years, foreign born and those who had psychoactive medicines during pregnancy.45 Another case-control population-based cohort study among Swedish children (n = 408, born 19741993), reported that the risk of ASD was associated with being small for gestational age, daily maternal smoking in early pregnancy, maternal birth outside Europe and North America, a 5-min APGAR score o7 and congenital malformations.46

Perinatal stress has been linked to increased risk of ASD.18,47

Such stress may be linked to sexual abuse that has been associated with higher risk of ASD.48,49 ASD patients are prone

to stress50 and a meta-analysis showed a strong correlation between the presence of anxiety disorders and ASD.51 In fact,

anxiety was signicantly correlated with repetitive behaviors in children with ASD.52 We reported that the peptides NT53 and CRH9 secreted under stress were increased in the serum of young children with ASD, as compared with normal controls.53 The

highest expression of NT receptors in the human brain is in the amygdala,54 hypothalamus and area of Broca,55 which regulate emotions and language, respectively. Stress can activate MCs through CRH leading to increased BBB permeability.56 Moreover, CRH has synergistic actions with NT, stimulating secretion of VEGF and increasing vascular permeability.53 Human MCs express CRHR-1,57 activation of which by CRH leads to VEGF secretion and BBB disruption58 and NT stimulates secretion of VEGF.57

A recent review concluded that stress during gestation increases the risk for developing atopic diseases in infants.59

Moreover, stress has been associated with precipitating or worsening asthma60 and multiple sclerosis.61

ATOPIC DISEASES ARE STRONGLY CORRELATED WITH INCREASED RISK OF ASD

Recent studies have shown strong associations between allergies, asthma, autoimmune diseases and psoriasis in the mother with increased risk for ASD in their children.6264 Moreover, mothers with mastocytosis or MC activation syndrome were much more likely to have children who developed ASD.65

Allergies66 and auto-immune diseases67,68 have been increasing signicantly. Early reports indicated more frequent allergies in ASD children,69,70 with food allergies being the most prevalent complaint, often in the absence of elevated serum IgE or positive skin tests.7173

A large epidemiological study of noninstitutionalized children (n = 92 642; 017 years old) showed that eczema was strongly associated with ASD and attention decit hyperactivity disorder.74

Another study of atopic subjects (n = 14 812; 3 years old) and nonatopic subjects (n = 6944) also showed a strong association between atopy and risk of both ASD and attention decit hyperactivity disorder.75 A case control study of children and young patients with ASD (n = 5565) and controls (n = 27 825) matched to birth year (19802003) and sex reported that allergies, asthma and auto-immune disorders were diagnosed more frequently, with psoriasis occurring more than twice as often, in ASD patients than controls.76

An experimental study actually reported neurochemical changes and autistic-like behavior in a mouse model of food allergy.77

MCs can be activated by fungi,78 such as Aspergillus fumigatus which triggers IgE-independent MC degranulation79 and fungal

zymosan induces leukotriene production from human MCs.80

Moreover, MCs can be stimulated by aluminum and mercury.81,82

PERINATAL EPIGENETIC ENVIRONMENTAL TRIGGERS CONTRIBUTE TO INFLAMMATION OF THE BRAIN AND INCREASE RISK OF ASD

Environmental triggers have been increasingly invoked in ASD.17,19,8386 Chemical intolerant mothers were three times more

likely to have a child who developed ASD and these children were

more prone to allergies and sensitivities, including odors.87

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Exposure to mold has been linked to decreased cognitive function in children88 and volatile mycotoxins have been reported to induce neuropsychiatric symptoms.89

Both mercury90 and aluminum91,92 have been associated with

symptom severity in children with ASD and both can stimulate MCs.81 Aluminum has replaced mercury as an adjuvant in vaccines, but aluminum can cause DNA damage93 and induce

microglia TNF release.94 The adjuvant activity of aluminum was shown to be mediated through DNA released from dying cells, possibly through production of IgE and IgG1, known MC triggers.95

Such damage-associated molecular patterns can act as alarmins96 and cause inammatory responses through toll-like receptors, which participate in immunity against bacterial infections97,98 and are also expressed on MCs.99

Stimulated human MCs can secrete mitochondrial DNA (mtDNA) and ATP extracellularly without cell death.100 These mitochondrial components augmented allergic responses101 and could act as

innate pathogens triggering inammation and potentially contributing to ASD.102 mtDNA is also directly neurotoxic in rat brain slices.103 We reported that serum mtDNA is elevated in young autistic children as compared with controls.104 The pathological importance of extracellular mtDNA could be even more relevant in the subgroup of ASD patients with mitochondrial dysfunction.105

MCs are therefore considered important for inammation.23,106

EVIDENCE FOR INFLAMMATION OF THE BRAIN IN ASD PATIENTS

Increasing evidence indicates that perinatal brain inammation,18,107 may contribute to the pathogenesis of neuropsychiatric disorders,108,109 including ASD.26,110 It was previously reported that ASD pathogenesis involves some immune17,111113 and autoimmune102,114 components. Circulating

auto-antibodies directed against fetal brain proteins have been reported in mothers of children with ASD115,116 and in about 37%

of ASD patients,117 implying BBB disruption which is regulated through MCs.56,118 The presence of auto-brain antibodies

signicantly correlated with allergic symptoms.119

A number of inammatory molecules have been shown to be increased in the brain and cerebrospinal uid of many ASD patients including IL-1, IL-6, TNF, MCP-1 and CCL8 (IL-8) 120122

(Table 3). Plasma levels of IL-1, IL-6 and IL-8 were increased in children with ASD and correlated with regression, as well as impaired communication and aberrant behavior.123

Analysis of cytokines in neonatal blood showed that IL-1 and IL-4 linked to severe ASD.124 In a previous study by some of the same authors, these cytokines were not detected apparently due to the sensitivity of the assay used.125 Increased maternal serum concentrations of IFN-, IL-4 and IL-5 during midgestation were signicantly associated with a 50% increased risk of ASD.126

MC-derived TNF can promote Th17-dependent neutrophil recruitment.127 Moreover, MC-derived IL-6 and TGF promote the devlopment of Th17 cells.128 In fact, MCs can also secrete IL-17129 and IL-17 was reported to be increased in the serum of children with ASD.130 There was an increased IL-17 production from peripheral blood immune cells following mitogen stimulation, and IL-17 was further increased in ASD children with comorbid asthma.131 A recent paper reported that selective elimination of Th17 cells in the maternal immune activation (MIA) mouse model prevented the development of autism-like behaivor in the offspring.132

The MIA model was also associated with increased serum IL-6,133 and the autism-like behavior was absent in IL-6/ mice.134 We had reported that acute stress signicantly increases serum IL-6 in mice that was entirely dependent on MCs, as it was absent in MC-decient W/Wv mice.135 In fact, human MC can undergo selective release of IL-6 without degranulation.136 Mastocytosis patients have increased serum IL-6 that correlates with disease activity137139 and children with mastocytosis had a 10-fold higher risk of developing ASD than the general population,65 implying activation of MCs.27

MCP-1 in amniotic uid was strongly correlated with increased risk for infantile autism140 and MCP-1 was also elevated in archived neonatal blood specimens.125 MCP-1 is chemotactic for MCs,23 which can secrete both pre-formed and newly synthesized TNF.141 TGF-beta has been reported to be low in the brains of children with ASD,142 a nding that may contribute to the inammatory state since TGF-beta inhibits MCs.143,144

Peripheral blood mononuclear cells from patients with ASD (n = 23) produced twice as much TNF as those from controls (n = 13) when stimulated even by gliadin, cows milk protein or soy.145 NF- DNA-binding activity, involved in TNF production, was twice as much in peripheral blood from patients with ASD (n = 67) than controls (n = 29).146 Neurons, astrocytes and micro-glia from patients with ASD had higher expression of NF- p65 as compared with matched controls.147 Moreover, signaling through NF- was prominent in interacting gene networks constructed from brains of ASD patients.148

MCs have recently been considered important in neuroinammation.149

MCMICROGLIA INTERACTIONS IN THE PATHOGENESIS OF ASD

Microglia, the innate brain immune cells,150 are important during healthy brain development because they may prune neural circuits.151,152 However, abnormal microglia activation and pro

liferation could lead to focal inammation and choking of normal synaptic trafc as has been reported in brains of patients with ASD.39,153155 A recent study of the transcriptomes from 104

human brain cortical tissue samples from patients with ASD identied gene clusters associated with increased microglia activation (M2) and decreased neuronal activity.156 As a result,

microglia are now considered an important component of the pathogenesis of ASD.157,158

Human microglia express functional CRHR1159 and NTR3

(sortilin), activation of which leads to microglia proliferation.160

NTR3 has been implicated in neuronal viability and function161

and increased soluble sortilin has been associated with depression, corresponding to elevated levels of BDNF and VEGF.162 NT

can be neurotoxic by facilitating N-Methyl-D-aspartate-induced excitation of cortical neurons.163 We recently reported that NT

Table 3. Evidence for inammation of the brain

BrainMicroglia activation Microglia proliferation IL-1

IL-6

IL-17 TNF

BloodAuto-brain antibodies

IL-1

IL-6

IL-17 TNF NF-B

Neonatal bloodMCP-1

Midgestational bloodAuto-brain antibodies

IL-4, IL-5, IFN-

Abbreviations: IL, interleukin; TNF, tumor necrosis factor.

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Figure 1. Schematic representation of the interactions among mast cellsmicroglianeurons and the bloodbrain barrier. Curved arrows, along with mediators associated with them, indicate action from one type of cell to another. The inhibitory action of luteolin (in box) is indicated by the inhibitory symbols (T). CRH, corticotropin-releasing hormone; IL, interleukin; NT, neurotensin; TNF, tumor necrosis factor; VEGF, vascular endothelial growth factor.

stimulates activation and proliferation of human microglia.164

We believe this is the rst time that a neuropeptide elevated in ASD is shown to stimulate human microglia that are now believed to play a major role in the pathogenesis of ASD.39,153,154 NT can

therefore stimulate both microglia and MCs (Figure 1).53

Signaling through the mammalian target of rapamycin (mTOR) has been implicated in ASD5,165 and mutations of the mTOR upstream regulatory molecule phosphatase and tensin homolog (PTEN)166 and tuberous sclerosis complex 1 and 2 (TSC 1/2)167 have been associated with higher risk of ASD.167 We recently

showed that activation of NTR3 induced activation of human cultured microglia, which was regulated by mTOR.164 PTEN and

mTOR are also involved in MC activation and proliferation.168

MC-derived histamine169 and tryptase170 can stimulate micro-glia, ndings that have led to the proposal that MC-microglia interactions are important in neuroinammation.171,172 Stimula

tion of brain MC in mice was recently shown to induce microglia activation and brain inammation, inhibited by a MC stabilizer.172

It is, therefore, important to address neuroinammation as a possible treatment option for ASD.

TREATMENT APPROACHESMost children with ASD are often prescribed psychotropic medications,173 primarily risperidone and aripiprazole to reduce disruptive and aggressive behaviors, but these drugs have no effect on the core symptoms of ASD.174,175 In fact, recent studies

have questioned the benet of psychotropic agents and have highlighted frequent adverse effects such as weight gain, sedation, tremor, movement disorders and drooling.176 As a

result, there is increased polypharmacy174,177 and risk of unwanted

drug interactions.178

There should be concerted efforts toward developing effective treatments for ASD, such as the European Autism Interventions-A

MultiCentre Study for Developing New Medications (EU-AIMS) Initiative.179

Immunomodulatory treatments have been considered for ASD,180 but few studies have been published. Some reports have hypothesized that the increase in ASD is linked to the increased use of the antipyretic acetaminophen.181 On the contrary, some families report that high fever reduces symptoms temporarily.182

Immune IgIntravenous (i.v.) immunoglobulin treatment (commonly known as immune Ig) has been used in ASD.183,184 In one study, i.v. Ig (200

to 400 mg kg1, every 6 weeks 2) was administered to children with ASD (n = 10) with one child showing signicant and four children showing mild improvement.185 Three pilot open clinical trials showed some benet.186188

The usefulness of this approach may be even more apparent in children with ASD whose plasma levels of IgG and IgM were reported to be low in spite of apparently normal numbers of B cells.189

Macrophage activating factor (GcMAF)

This molecule, an endogenous glycosylated vitamin D-binding protein, which promotes macrophage cell activation, down-regulated the over-activation of blood monocyte-derived macrophages observed in autistic children (n = 22, 311 years old) compared with age-matched healthy developing controls (n = 20).190

Antioxidant compoundsA recent double-blind, placebo-controlled, study using the broccoli-derived anti-oxidant sulforaphane in adult patients with ASD (n = 40, 1327 years old, selected for their history of reduced symptoms during febrile episodes) for 18 weeks showed

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signicant improvement (34%) in social interaction and communication using the Aberrant Behavior Checklist (ABC) scale;191

however, the apparent signicance was due to the uncharacteristically low placebo effect (o3.3%). Placebo effects have been reported as high as 4060% in studies of neuropsychiatric diseases.192

Another antioxidant, N-acetylcysteine (NAC), has also been tested. In one randomized, placebo-controlled, trial (n = 13) increasing doses of NAC (900 mg per day 4 weeks, then 900 mg twice daily 4 weeks and nally 900 mg three times daily 4 weeks) found no difference on the total ABC, but signicant improvement on the irritability subscale.193 In another

also randomized, double-blind, placebo-controlled, study (n = 40), NAC added to a stable dose of risperidone, again had no effect on total ABC, but decreased the irritability subscale.194,195 NAC

treatment appears to be safe and well-tolerated.195 Similar results were obtained in a more recent randomized, double-blind, placebo-controlled clinical trial of children with ASD (n = 40) who were given NAC (600900 mg per day) and risperidone titrated (between 1 and 2.0 mg per day); by week 10, the NAC group showed signicantly less irritability using the ABC-C irritability subscale (P = 0.02).196

Anti-inammatory compoundsAn open-label study investigated a formulation containing the natural avonoids luteolin and quercetin ((100 mg each per softgel capsule in olive kernel oil to increase oral absorption) 1 capsule per 10 kg weight per day for 6 months) in children with ASD (410 years old, n = 50) and reported signicant (Po0.005)

improvement in attention and behavior (34% in total ABC and8.43 months in age-equivalent scores in the VABS communications domain).197 A subgroup of children in that study improved even more (65%) and were the ones with highest serum TNF and IL-6 at the beginning of the study, the levels of which dropped below basal levels at the end of treatment.122 These results indicate that objective inammation markers may identify a subgroup of children with ASD, who are most amenable to treatment with luteolin or quercetin. A case series using the same formulation in children with ASD and atopic diseases (n = 17, 412 years old) reported 65% improvement in attention and communication.198 Luteolin also improved brain fog, characterized by reduced attention span, memory and learning199 in adults.

Luteolin (5, 7, 3, 4-tetrahydroxyavone) is naturally found in green plants, herbs and seeds 200 and is structurally related to 7, 8-dihydroxyavone, which was shown to have brain-derived neurotrophic factor (BDNF) activity201 (Table 3). Low BDNF was associated with autistic-like-behavior in mice202 and 7, 8-

dihydroxyavone reduced symptoms in a mouse model of Rett syndrome,203 which is strongly associated with ASD.204

Luteolin and its structurally related avonol quercetin (5, 7, 11, 3, 4-pentahydroxyavonol) inhibit histamine, IL-6, IL-8, TNF and tryptase release from human MCs.205,206 We recently showed that

tetramethoxyluteolin is a more potent inhibitor of human MCs than luteolin.207 Luteolin also inhibits microglial activation and proliferation,208 especially IL-6 release,209 and is neuroprotective.210 Luteolin also prevented autism-like behavior in a mouse model of autism.211 Flavonoids are generally considered safe212,213 and now being increasingly discussed for the treatment

of neurodegenerative disorders.214

CONCLUSIONSSubstantial evidence indicates that the presence of atopic diseases increases the risk of ASD and that inammation of the brain may be involved in the pathogenesis of ASD. Addressing allergic symptoms, as well as reducing BBB permeability and inammation of the brain, could provide signicant benet in ASD. Luteolin

analogs with better bioavailability and BDNF activity should be investigated further. Intranasal administration to penetrate the brain through the cribriform plexus could deliver anti-inammatory molecules directly to the brain. Such formulations could further be prepared in liposomes to contain molecules that target them to microglia.

CONFLICT OF INTEREST

TCT has been awarded US Patents No 8268365; 9050275 and 9176146 covering treatment of brain inammation and of ASD. He has developed the luteolin-containing dietary supplements BrainGain, NeuroProtek and NeuroProtek-low phenol. The remaining authors declare no conicts of interest.

ACKNOWLEDGMENTS

Aspects of our work discussed were funded in part by the Autism Research Institute, the National Autism Association, Safe Minds, the Nancy Lurie Marks Foundation and The Jane Botsford Johnson Foundation.

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