Gut-Brain Inflammation Due to Toxin-Activated Mast Cells and Microglia in Autism Spectrum Disorder

1. Introduction

Autism Spectrum Disorder (ASD) is increasing at an alarming rate and now impacts 1 in 31 children in the United States [2], with a projected total cost of $461 billion by 2025 [3]. While genetic vulnerabilities are known to be a significant contributing factor to ASD, environmental factors, particularly environmental toxicants, may be another reason for the increased prevalence of ASD [4,5]. The microbiome and total gut physiology can also influence the onset of ASD and can be highly predictive of ASD [6].

Environmental toxins, including glyphosates, heavy metals, synthetic compounds and plastics, mold toxins, air pollution, COVID, and excessive use of antibiotics are just some examples of factors that can compromise gut permeability, induce gut-mediated inflammation, and disrupt the microbiomes [3,7,8,9,10,11,12,13]. Neonatal events such as C-section delivery and early use of antibiotics are also known to disrupt the microbiome [14] and may contribute to an increased risk of ASD [15,16,17]. What is not yet well understood is how transient, yet significant, exposures to environmental toxins and stressors contribute to ASD.

This paper discusses how such exposures disrupt the gut-brain axis leading to a chronic state of neuroinflammation primarily via activation of mast cells and microglial that can explain the physical, cognitive, psychological, and social findings in at least a subset of individuals with ASD. It further offers a simple, safe, and inexpensive set of recommendations to mitigate the inflammatory response and improve subjects with ASD.

2. Relevant Clinical Findings

Individuals with ASD have abnormal neuronal apoptosis [18], myelination [19] and neuroplasticity [20]. They also have dysfunction in multiple areas of cognition including: attention, executive functioning, working memory, praxis and motor planning [21]. Moreover, individuals with ASD experience significantly higher rates of anxiety [22], obsessive compulsive disorder (OCD) [23], social anxiety [24], abnormal perception of fear [25], anger and aggression [26]. This chronic state of psychological stress can trigger inflammation within the nervous system [27] and gastrointestinal tract [28]. These individuals also struggle with abnormalities in the processing of complex sensory information [29], with sensory processing disorders affecting up to 80-95% of individuals with ASD. The severity of these sensory processing disorders is a significant predictor of ASD severity and everyday functioning [30,31,32].

Children with ASD are also at a 500% higher risk of developing feeding problems [33], such as food selectivity, food refusal, and poor oral intake, as compared to neurodevelopmentally normal children [34,35]. Children with ASD have texture aversion and strong preferences for foods like carbohydrates and processed foods [36,37], and a higher risk of healthy food avoidance: vegetables (56% refusal), eggs (43%), fruits (42%), chicken (35%), and meat (24%) [35].

Individuals with ASD have a significantly higher prevalence of intestinal inflammation and inflammatory bowel disease [38] and are four times more likely to have gastrointestinal symptoms vs controls [36,39], including constipation (odds ratio 3.86), diarrhea (OR 3.63), abdominal pain (OR 2.45) [39], and these symptoms can begin as early as 6-18 months of age [40]. Individuals with ASD also have abnormal intestinal permeability [41], abnormal microbiomes [42] with a possible [43] higher prevalence of candida versus controls [44]. These individuals also have complex patterns of systemic immune dysregulation [45,46], with brain inflammation including microglia activation [47] and elevations of various cytokines in the cerebrospinal fluid [46,48], including the neuropilin disruptor matrix metalloproteinase-9 (MMP-9) [49]. The immune dysregulation also presents as higher rates of food allergies [50] and atopic diseases including asthma, allergies, and eczema, which are also strongly correlated with the risk of ASD50.

3. How Environmental Exposures Create Chronic Gastrointestinal Inflammation and Dysfunction

ASD is likely due to genetic vulnerabilities activated by to environmental toxins and stressors [51], with a significant contribution from the environmental factors themselves [52,53]. Some of these vulnerabilities occur at the gut level [54], including how host genetics interact with the gut microbes to shape the immune and metabolic state of ASD [55]. One such example is the role of the human leukocyte antigen (HLA) haplotypes, of which HLA haplotypes A2, DR4, and DR11 have been found to create major susceptibility for ASD45. These haplotypes can determine the specificity of T lymphocyte and natural killer (NK) cell responses, which then influence the makeup of the commensal bacteria [56].

As such, each individual has a genetically determined threshold of gut resilience and tolerance to various stressors or toxins. Thus, if an environmental factor or combination of factors supersede this threshold, the gastrointestinal tract and microbiome can become compromised setting off a complex cascade of gut-immune-brain dysregulation. At the center of this dysregulated axis are the gastrointestinal mast cells (Figure 1).

4. The Role of Mast Cells in Chronic Gastrointestinal Inflammation

Mast cells are the sentinels of our immune system [57] and can be activated by many of the same triggers that disrupt the gut, including: heavy metals, herbicides (including glyphosates), polychlorinated biphenyl (PCB), lipopolysaccharides (LPS), mycotoxins and molds, as well as pathogens [58] including Borrelia [59,60], and Helicobacter pylori [61] (Figure 2). Once activated, they orchestrate complex arrays of immune activation including mediating the allergy response [62], a finding common in individuals with ASD.

The largest population of mast cells is in the gastrointestinal tract, where they play a fundamental role in maintaining the intestinal barrier by regulating epithelial function and integrity, managing the defensive and immuno-regulatory function, modulating both innate and adaptive mucosal immunity, and maintaining neuro-immune interactions. These functions all play a fundamental role in the health or disease of the gut [63,64].

The gastrointestinal tract harbors trillions of bacteria, fungi (the mycobiome), viruses, LPS, and dietary antigens. The microbiota are well known to play an important role in neuropsychiatric disorders [65]. Separating this complex and potentially deadly “internal” gut content from our circulation and organ systems, including the brain, is a tightly regulated interface consisting of a mucus layer, immunoglobulin A, defensins, a single epithelial layer held together by tight junction proteins, and a carefully orchestrated immune surveillance in the lamina propria of the gut [66]. As long as this gut barrier is healthy and intact, and the bacterial and fungal communities are healthy, the likelihood of systemic and chronic inflammation is low. If there is a disruption in the microbial communities or gut barriers, the likelihood of a chronic systemic and nervous system inflammatory responses rise dramatically [67,68,69,70].

Zonulin and occludin are two important tight junction proteins that ensure the integrity of the intestinal barrier. Aberrant mast cell activation can cause profound disruption to the gut through damage to these tight junction proteins via mediators such as histamine, MMP-9, TNF-α and tryptase, resulting in increased intestinal permeability and inflammation, a critical step that allows the translocation of the commensal bacteria [63,66,71]. Through the release of numerous mediators mast cells can further diminish the integrity of the gut barrier, sensitize dendritic cells to microbial signals, including LPS, and influence the behavior of the innate and adaptive immune response [72,73,74]. Small intestinal mucosal damage may decrease the activity of diamine oxidase (DAO) [75], a key enzyme that degrades histamine, and thus exacerbate the detrimental effects of histamine that is released by activated mast cells. Mast cells can directly and indirectly suppress the function of T regulatory cells, while simultaneously promoting the activation, recruitment, proliferation and cytokine secretion in multiple T-cell subsets [62].

These combined changes to the gastrointestinal tract barrier and microbiome are sufficient for the pathogenesis of food allergies [76]. Especially in young children, even minor changes to the barrier function early in life can lead to exposure to luminal antigens which can result in allergies in later stages of life [77], as found in individuals with ASD. These gut abnormalities can also explain the gastrointestinal symptoms found in individuals with ASD, including abdominal pain, constipation, and diarrhea [78,79,80].

Perhaps, the most significant consequence of this toxin-mediated gut dysfunction is how it influences the makeup of the microbiome. Abnormalities in T-cell activity directly impact microbial diversity [81]. Inflammation-mediated dysbiosis, an emerging concept where gut inflammation alters the intestine’s oxidative and metabolic environment, can also profoundly influence the survival and growth of the gut bacteria and lead to a deviation of the commensal population from a healthy, diverse symbiotic profile to microbial communities with reduced complexity and over-representation of particular taxa of microbes [66,82].

Furthermore, certain toxins, like mycotoxins can alter the production of intestinal specific immunoglobulins [83], which then significantly influence the composition and behavior of the bacterial and fungal makeup of the microbiome [84,85,86,87,88]. Mast cell activation and the concurrent gut and microbiome disruptions can also induce candida species to switch from a harmless commensal to a virulent pathobiont, which is then able to invade tissues and disseminate in the body [71,89,90]. In turn, the invasive form of candida, and the toxin its hyphae form secretes, candidalysin [91], can trigger significant mast cell activation [92,93], thus creating a vicious cycle of chronic gut and immune dysfunction. Further complicating this picture, candida can alter the makeup of the microbiome through multiple mechanisms [94] and prevent the regrowth of lactobacilli after antibiotic treatment, while promoting the colonization of enterococcus [95]. All of these factors in turn perpetuate candida’s virulent pathobiont state. These mycobiome abnormalities also influence the maturation and priming of the immune system [96] and create an additional factor to induce or exacerbate the allergic state [97] that, as noted, is more prevalent in ASD [98].

These findings may explain the significant microbiome and mycobiome abnormalities found in individuals with ASD [99,100]. Because this dysbiosis is largely immune-mediated, it often responds poorly to conventional gut treatments, such as probiotics.

5. Gut Disruption and Bacterial Translocation

The innate immune system within the gut has pattern recognition receptors (PRRs) whose primary purpose is to detect pathogens, including the commensal bacteria, by recognizing molecules and structures on the pathogens called pathogen-associated molecular patterns (PAMPs) [101]. The disruption of the gut barrier allows pathogenic commensal bacteria to breach the gut, at which point their PAMPs are recognized by the PRRs, which then activate the intestinal epithelial cells (IECs) to secrete cytokines and chemokines. This inflammatory reaction triggers a larger immune response to clean up the foreign and damaging antigens and threats, thus resulting in further inflammation and cytokine release [66]. These inflammatory cytokines further disrupt the epithelial barrier by downregulating tight junctions (claudin-1, occludin, zonula occludens protein-1) and adherens junctions (E-cadherin) in IECs [66]. One consequence of this pathogenic breach of the gut barrier and the ensuing inflammatory response becomes the key step in the induction and persistence of gut inflammation inflammatory bowel diseases [102], which is more prevalent in children with ASD versus healthy controls [103].

Another serious consequence of the compromised gut barrier is the entry of dangerous bacterial components into the systemic circulation. The gram-negative bacterial surface glycolipid lipopolysaccharide ( LPS), is among the most potent pro-inflammatory neurotoxins [104,105]. Gut dysbiosis and a disrupted gut barrier allow for the translocation of LPS through the gut lining into the systemic circulation to allow the development of a low-grade, chronic generalized toxin-associated effects [106], as is found in individuals with ASD [107]. The toxicity of the LPS increases with the degree of dysbiosis [105]. Numerous animal models suggest a strong link between generalized toxin effects and the features of ASD [108,109,110,111]. One study in rats reported autistic features after a single prenatal exposure to LPS [112]. Propionic acid is also known to cause similar effects [113,114].

Once absorbed, LPS can trigger systemic inflammation, with a reduction in T regulatory (Tregs) lymphocytes, increase in Th17 and Th1 lymphocytes, along with increased TNF-α, NFkB, IL-6, IL-8, IL- 10, and IL-12 [105,115]. Children with ASD also have activation of inflammasome complexes, including the NLRP3 inflammasome [116], which are complex systems that play a critical role in the regulation and activation of the body’s inflammatory response. LPS can rapidly prime and activate this inflammasome [117], which could be further primed by candidalysin [117,118]. Furthermore, within the intestinal lining, LPS binds to PRRs (TLR4) on intestinal cells [106] and mast cells [119,120], thus triggering additional inflammation within the gut and further disrupting the tight junction proteins and gut barrier.

The most concerning effect of LPS occurs within the nervous system. The microglia are the innate immune cells that modulate the inflammatory response within the central nervous system [46]. Peripheral gut inflammation and damage can activate the microglia [121,122,123]. In healthy adult volunteers, 1 ng/kg of LPS (similar levels found in some healthy adults) can trigger robust microglial activation in most areas of the brain as measured by PET within three hours of injection [115]. In rodents, a single intraperitoneal injection of 5 mg LPS/kg causes microglial activation that persists for at least 12 months [115]. Several other animal studies have also shown systemic LPS can activate the microglia [124,125,126,127]. If animals are given multiple doses of 1 mg LPS/kg (over several days), a model for chronic generalized toxin effects, they experience neuroinflammation, BBB permeability, and rapid neurodegeneration [115], findings that are common in individuals with ASD [128,129].

Even low levels of LPS can induce sickness behavior through elevation of inflammatory cytokines [115,130,131,132]. Sickness behavior, an adaptive change in behavior as a result of inflammation, has been described in individuals with ASD and can present as anxiety, appetite loss, depression, headache, impaired alertness and focus, lethargy, muscle pain, and social withdrawal [115,133].

The impact of LPS on the CNS is likely through multiple mechanisms, including the vagus nerve (Figure 2) [123]. LPS cannot pass through or directly disrupt the blood-brain barrier (BBB) [115]. On the other hand, central histamine, various toxins, a high fat, high sugar diet are just a few of the factors that can compromise the BBB [134,135,136,137,138], thus allowing LPS to directly enter with CNS. Stress via the release of corticotropin-releasing hormone CRH) can also disrupt the BBB through activation of mast cells [139,140,141].

Microglia shape neuronal plasticity and connectivity, and synaptic function and wiring through neuronal pruning [142,143,144]. They regulate myelin growth and integrity [145] and when activated may cause severe demyelination [146]. They are also involved in abnormal neuronal apoptosis [147]. By disrupting neuronal circuitry, microglial activation impairs the processing and integration of various sensory and emotional responses as part of the presentation of ASD [46,148,149]. Individuals with ASD have been found to have microglial activation [150], also confirmed through postmortem findings [151] and on functional positron emission tomography [152].

Microglial activation also alter the neuronal pathways of the amygdala, which has been shown to disrupt the fear threshold within human beings and may present as ASD [153]. Disruptions in the amygdala can also play a significant role in the pathogenesis of aggressive behavior [154], addictive behaviors [155,156], anxiety disorders [157], impulse control disorder [158], attention deficit disorder [159], depression, and a host of other neuropsychiatric findings [160]. These behavioral and psychiatric findings are all noted in individuals with ASD [161,162,163,164,165].

Furthermore, activated microglia may induce post-synaptic calcium elevation causing increased neuronal reactivity and disrupt glutamate signaling [144]. Microglia can also alter the levels of quinolinic acid [166], a potent neurotoxin implicated in ASD [167]. Microglial activation can also disrupt the behavior of the astrocytes [168], which in turn can further disrupt glutamate homeostasis, upset GABA regulation, and neuronal pruning [144].

Beyond the effects of the microglia, human and animal studies have shown that endotoxin-induced inflammation can also increase the neural responses in the anterior cingulate cortex and prefrontal regions further impacting the processing of social and emotional information [169]. Beyond the microglia are the mast cells, which serve as an “immune gate to the brain” and communicate with microglia in a two-way activation [170,171,172,173].

6. Mast Cells and ASD

Peripheral gut inflammation activates mast cells within the nervous system [174]. This is further impacted by psychological stress [175,176]. Mast cell activity is intimately tied to microglial activity, and the activation of one cell line can lead to the activation of other immune cells through multiple pathways (Figure 3) [168].

The role of mast cell and microglial activity has been well described by Theoharides, et al. in individuals with ASD [46,177,178,179]. Mast cell activity can disrupt the blood-brain barrier, and release multiple chemokines, cytokines, tissue disruptors and neurotoxic molecules, like IL-6 that create localized inflammation in the area of the basic ganglia that disrupts neuronal connectivity and contributes to ASD-related behaviors [180]. Mast cells also trigger a neuroinflammatory response through multiple mechanisms [46,47,181]. They can induce mitochondrial translocation to the cell surface with the secretion of extracellular mitochondrial adenosine triphosphate (ATP) and DNA that is then detected by the immune system as ‘innate pathogens’ triggering a significant inflammatory response, potentially contributing to ASD [182]. This mitochondrial DNA may also induce a neuro-inflammatory response, which has been found to alter behavior in mouse models [183]. Theoharides has suggested that this profound sequence of neuro-inflammatory response is sufficient to cause ASD in some [180].

Gut-mediated mast cell activation (and gut histamine) can increase histamine levels within the CNS, likely through multiple pathways, although the exact mechanisms are not yet clear [184]. It is clear that activated microglia and mast cells within the brain produce 50% of all brain histamine [185], and in animal models, intraperitoneal LPS injection can activate brain mast cells and cause a rapid elevation of central histamine within 6 hours [186].

Histamine plays a critical role in modulating the nervous system [187]. It regulates alertness and is also a key wake-promoting neurotransmitter that influences the circadian rhythm and sleep-wake behavior [185]. Sleep disruption is a common feature of ASD [188]. Elevated levels of central histamine can also disrupt the vestibular system, which is critical for balance, motor planning, and sensory perception [187]. Histamine has also been shown in animal models to directly or indirectly influence various sensory pathways: sound processing [189], tactile sensation [190], and olfactory perception [187,191]. The disruption of these sensory pathways may explain the host of sensory findings found in individuals with autism, including eating problems and food texture avoidance [192]. The intensity of sensory issues has been associated with more significant social difficulties, lower adaptive functioning, and lower or divergent visual exploration of social environments in children with ASD [193]. These sensory abnormalities are also strongly associated with social motivation and difficulties with initiating or maintaining social interactions, or feeling tense in social situations [193].

Furthermore, because of these sensory abnormalities and eating problems, individuals with ASD avoid fiber-rich vegetables and foods that are often rich in simple carbohydrates, fats, and sometimes highly processed foods [35,194]. These dietary preferences can exacerbate any microbiome disruption [195] and increase the abundance of candida [196]. The high amounts of carbohydrates and fats, and few dietary fibers can also dramatically enhance the absorption of LPS and induce an inflammatory response [197]. However, certain fruits and vegetable contain histamine (e.g. avocado, pineapple, spinach, tomatoes) that should best be avoided, especially in those with polymorphisms in the histamine degrading enzymes diamine oxidase (DAO) and histamine N-methyl transferase (HNMT). [198]

7. How to Address Gut-Brain Inflammation Effectively

While it is beyond the scope of this paper, numerous options are available to address many parts of this proposed cycle of physiological dysfunction and endogenous toxicity (Table 1).

Theoharides, et al. have demonstrated significant clinical improvements of symptoms in children with ASD from a dietary supplement which contains liposomal (formulated in olive pomace oil) luteolin (100 mg/softgel) quercetin (70 mg/softgel), and the quercetin glycoside rutin (30 mg/softgel)(NeuroProtek) [199,200,201]. The noted studies all suggest that these supplements are safe and well tolerated.

The terahydroxyflavone luteolin is a well-studied bioflavonoid with a host of anti-inflammatory properties [202,203,204]. Luteolin has been found to inhibit mast cell and T cell activation [205], and decrease levels of histamine and TNF [206,207]. In fact, luteolin was recently shown to be a more potent inhibitor that the “mast cell stabilizer” drug cromolyn [208]. Luteolin can also enter the brain and reduce microglial activation [209], particularly as a result of LPS [210], as well as have antibacterial properties [211]. Pentaxydroxyflavonol quercetin can inhibit mast cells and is more effective than cromolyn at blocking mast cell cytokine release [212]. It can also balance the Th1/Th2 immune response [213], reduces gut permeability while improving microbial diversity [214], and protects against LPS-induced gut damage through multiple mechanisms [215]. Rutin is another important compound because of its ability to liberate quercetin in the gut. The delivery base is just as important, since olive pomace oil that not only increases absorption from the gut, but it offers the well-known cytoprotective properties of olive oil [216]. Unfortunately, many cheaper preparations of luteolin and quercetin in powder form have flooded the market but are either of low purity or the daily dose requires multiple capsules [217]. The common notion that if you take higher amounts of luteolin or quercetin in powder form will eventually allow some of the flavonoids to be absorbed is not only wrong but dangerous as the unabsorbed flavonoids accumulate in the gut and disrupt the microflora [218].

The structural luteolin analogue, tetramethoxyluteolin, is even more potent than luteolin in inhibiting both mast cells and microglia [219,220,221,222,223], and has been incorporated in the novel anti-allergic skin lotion (GentleDerm) [224], which is particularly useful in those individuals with both eczema and ASD.

Two additional compounds that may play an useful role in the treatment of ASD are palmitoylethanolamide (PEA) and diamine oxidase (DAO) enzymes. PEA is a naturally occurring fatty acid amine found in soybean lecithin, egg yolk, and peanut meal. PEA has noticeable anti-inflammatory properties and can regulate mast cell activation [225,226,227] by reducing release of TNF-a and histamine [228]. Furthermore, PEA displays neuroprotective properties and can inhibit microglial activation [229], particularly as a result of LPS exposure [230]. In isolated case reports, PEA has been shown to be beneficial in ASD [230].

DAO enzymes are naturally occurring enzymes within the gastrointestinal tract that are responsible for the degradation of histamine within the gut. Since intestinal mucosal damage may decrease the DAO activity [75], additional supplementation may play an important role in normalizing histamine levels within the gut and reducing the inflammatory and possibly neurological findings. Histamine intolerance has also been associated with anxiety disorders [231]. Exogenous supplementation with DAO enzymes can significantly reduce histamine levels within the gastrointestinal tract and the signs and symptoms of histamine intolerance [232], including extra-intestinal symptoms such as headaches [233]. To our knowledge, DAO enzymes are safe and well tolerated. However, DAO preparations vary considerably in their stated activity and most the enzyme will be degraded by the stomach acidity unless they are in acid resistant formulations.

Addition of berberine, lactoferrin and Bifidobacteria infantis/longum could provide additional benefits as they all have both antibacterial and anti-inflammatory properties [234,235,236,237,238,239]. The combination of these naturally occurring compounds may prove useful in the treatment of ASD.

Addition of folinic acid (Calcium folinate, Leucovorin) has been shown to significantly improve brain health, cognition and language by bypassing surface folate receptors and the enzyme MTHFR, especially in those with anti-folate receptor antibodies and MTHFR polymorphisms (Table 1) [240].

There are obvious limitations to the suggested treatment approached discussed above. The clinical efficacy of these compounds may be limited in the face of significant gastrointestinal disease or serious environmental exposures, such as living in a home with severe mold contamination.

8. Beyond the Clinical Benefits

Unfortunately, at the time of this publication, there are few clinically available and reliable diagnostic tools that can help clinicians accurately assess these assessing gut-mediated pathophysiological events, including:

(1)

Assessing candida and the mycobiome – There are many commercially available stool kits to assess the bacterial component of the microbiome. Unfortunately, these kits lack the sensitivity to accurately detect disturbances of candida or other fungal components of the microbiome (mycobiome).

(2)

Assessing microglial activation – No commercially available diagnostic modalities are available to accurately assess microglial activation.

(3)

Assessing mast cell activity and histamine – Serum histamine has a half-life of less than two minutes and thus cannot be used to accurately detect histamine imbalances. Serum tryptase can be used to assess significant mast cell burden (e.g. systemic mastocytosis), which may limit its ability to detect more subtle forms of mast cell activation, including within the central nervous system. Urinary N-methylhistamine, Prostaglandin F2 alpha and leukotrience E4 must be collected cold in 24-hour urine and most clinical labs do not perform them.

(4)

Assessing endotoxemia – At this time there is no commercially available diagnostic tool available to directly assess endotoxemia.

(5)

Assessing total toxin load – Currently, only specialty tests are available to assess select categories of toxins. These tests are not FDA-approved and their results are at times called into question.

These points demonstrate the profound limitations in our current diagnostic tools to detect these critical physiological processes in individuals with ASD. They also highlight how important future well-designed research studies are in helping us further our understanding of ASD, and to allow this science to be effectively utilized in the clinical setting.

9. Conclusion

There is no singular trigger, event, genetic or physiological process that is solely responsible for the onset of ASD in the majority of cases. It is the total load of pathogenic and environmental toxins, which varies from individual to individual, compounded by other infectious [241], microbiome/gut, physiological and psychological stressors that may be responsible for the onset of ASD in some individuals [242]. We believe that in these individuals, there is a moment during gestation/delivery, in infancy or early childhood where the gut barriers, microbiome and gut-mediated immune responses surpass a threshold of homeostasis, and enter into a perpetual cycle of neuroinflammation.

While there is sufficient data to identify the individual factors contributing to ASD risk, additional research is needed to bring this vast array of findings into one cohesive model that has the power to assess the unique exposome for each individual and appreciate the total physiological impact it can have, including on the mitochondria and other organ system dysfunctions that have been associated with ASD. Additional research is also needed to assess the role that bacterial endotoxins may play in the manifestation of ASD.

In the meantime, the proposed interventions can inhibit some of the pathogenetic pathways and allow the gut-brain axis to recover as has been the case in numerous individuals with ASD.