Essay: Autism Spectrum Disorder (ASD)

Introduction
Autism Spectrum Disorder (ASD) refers to a group of neurodevelopmental disorders characterized by core symptoms that include persistent deficits in social communication and interaction as well as restricted repetitive patterns of behaviors and interests (American Psychiatric Association, 2013). In order to reach diagnostic criteria, core symptoms must be present early in development and cause significant impairment in social and occupational functioning (American Psychiatric Association, 2013). Other symptoms often experienced include decreased verbal skills and social withdrawal, insistence to routines and unusual response to sensory stimuli (Critchfield, 2011). In recent years, ASD diagnoses have been increasing dramatically, now with roughly every 1 in 68 children diagnosed in the United States (Christensen, 2012). ASD is known to be much more common among boys (1 in 42) than among girls (1 in 189) (Christensen, 2012) and occurs in all racial, ethnic, and socioeconomic groups (Durkin, 2010).
Along with the strain put on individuals with ASD as well as their families, ASD is accompanied by staggering economic costs. In fact, it has been found that the total cost per year for children with ASD in the United States was estimated to be between $11.5 billion and $60.9 billion in 2011. This cost accounts for a variety of direct and indirect costs, from medical care to special education to lost parental productivity (Buescher, 2014. Lavelle, 2014). In addition to medical costs, intensive behavioral interventions for children with ASD cost $40,000 to $60,000 per child per year (Amendah, 2011).
Despite the economic, societal and personal difficulties associated with ASD, the exact cause is far from understood and there remains to be no US Food and Drug Administration (FDA)-approved pharmaceutical treatment to alleviate core symptoms (Neul, 2015). In the search for a cause, much emphasis has been placed on genetic vulnerability, heredity, functional and structural brain abnormalities as well as environmental factors. Recent advances in the field have shown that the most important environmental factor may have been inside of us all along: the microbiome. The ASD microbiome has become the new frontier in ASD research and has the potential to be the basis for innovative new therapies through which patients can be relieved of core symptoms and have an increased quality of life for themselves and those around them. Crucial to this understanding of the microbiome’s role in ASD is knowledge of the healthy intestinal microbiota, the gut-brain axis, the relationship between the microbiome and the immune system, and the ultimate connection between the microbiome, the immune system, and the brain in ASD. It is this critical connection that will ultimately lead to a more thorough understanding of the ASD brain and body as well as push for the use of microbiota transfer therapy to provide ASD patients with a safe and effective, long-lasting treatment.
The Intestinal Microbiota
The formation and maturation of the human intestinal microbiome is integral to a healthy immune system and standard neurodevelopment. Many varieties of microorganisms including bacteria, viruses, protozoa, archaea and fungi make up the microbiota of the gastro-intestinal (GI) tract (Sekirov, 2010). These organisms colonize the external body surfaces at the time of birth. This initial colonization is affected by the mode of delivery (vaginal or C-section), external environment (i.e. hospital), type of feeding (breast milk or formula), mother’s diet and use of antibiotics (Matamoros, 2013; Sekirov, 2013). The first three years of life are seen as critical in humans for the creation and maturation of the intestinal microbiome (Matamoros, 2013). It is at this same time that a considerable amount of synaptogenesis is occurring in the brain (Tau, 2010). This co-occurring evolution between the microbiota and the nervous system continues throughout life, suggesting their possible influence on one another’s development.
The intestinal microbiota of healthy individuals is largely comprised of anaerobic bacteria that are mostly non-pathogenic. These microorganisms are crucial to the health of the host as they aid in various functions such as the absorption of nutrients, production of short-chain fatty acids and vitamins, amino acid synthesis from ammonia or urea and maintenance of the intestinal barrier (Hooper, 2012; Sekirov, 2010). Dysbiosis is the term for microbial imbalance. Since these functions carried out by the microbiota are so crucial for a healthy host, dysbiosis can be detrimental.
The Gut-Brain Axis
The gut-brain axis (GBA) comprises the central nervous system (CNS), the autonomic nervous system (ANS), the enteric nervous system (ENS) and the hypothalamic pituitary adrenal (HPA) axis (Rhee, 2009). The role of the GBA is to monitor and integrate intestinal functions and to link the emotional/cognitive centers of the brain with intestinal functions and mechanisms. These mechanisms include immune activation and intestinal permeability (Rhee, 2009). The enteric microbiota, which is distributed throughout the gastrointestinal tract, is believed to have important metabolic and physiological functions and is crucial to the maintenance of homeostasis and significantly impact the GBA (Mayer, 2014). The vagus nerve has been indicated as the main communication pathway between microbiota and the brain (Bravo, 2011). The microbiota profile of healthy individuals is relatively similar and major changes could lead to a wide array of gastrointestinal or even neurological disorders (Mayer, 2014).
Sudo et al. (2004) conducted a ground-breaking study that began autism research’s new frontier into the intestinal microbiota’s impact on the brain. This study was done using germ free (GF) mice. GF mice are mice that are completely sterile and have never had contact with any microorganisms (Sudo, 2004). It is this lack of a stable intestinal microbiome that allows these GF mice to be a model for the microbial dysbiosis that is seen in many individuals with ASD. It was found that these GF mice show an exaggerated stress response when compared to mice with normal microbiota (SPF mice), most likely due to dysfunction of the HPA axis. It was also found that administration of the bacterium Bifidobacterium infantis rescued the HPA stress response (Sudo, 2004) in GF mice, supporting evidence for the relationship between the intestinal microbiota and the HPA axis. Additionally, fecal transplant of microbiota from SPF mice to GF mice at an early stage successfully restored the stress response to levels similar to that of SPF mice (Sudo, 2004). This is evidence for the idea of microbiota transfer therapy as a treatment for individuals with ASD, which will be discussed later. However, this same procedure was not effective at a later age (Sudo, 2004), suggesting that early on in development there exists a sort of critical period for the action of the microbiota on brain function and plasticity.
The Microbiome and the Immune System
The microbiome and the immune system interact in a variety of ways. First of all, the gut microbiota produce microbial-associated molecular patterns (MAMPS) and metabolites that can stimulate immune cells and eventually influence brain chemistry and behavior (Mu, 2016). A major way in which the microbiota interacts with the immune system is through microglia. Microglia are a type of glial cell that acts as the first line of immune defense in the central nervous system. It has been found that depletion of intestinal microbiota dramatically affects microglia development and function in adult mice, showing that the intestinal microbiota are important for the maturation of microglial cells (Erny, 2015).
A large part of the immune system is centered around the intestinal mucosa, as the intestinal microbiota are involved in the maturation of the immune system (Hooper, 2001) and play a role in the regulation of various immune functions (Delcenserie, 2008). However, the intestinal mucosal barrier may be compromised in those with ASD (White, 2003). In particular, intestinal permeability is increased in autistic children, and is referred to as “leaky gut”. It is hypothesized that this increased intestinal permeability allows various byproducts of bacteria and food derived peptides to escape into the blood, leading to immune responses that may affect neuronal signaling. In addition to leaky gut, many studies support the existence of abnormal immune activation, with such findings that include an abnormal ratio of CD4+ and CD8+ T-cells, abnormal T helper cells, elevated blood monocyte counts, decreased lymphocyte numbers, self-reactive antibodies, and neuroinflammation (Careaga, 2010).
Microbiota Abnormalities in those with ASD
Gastrointestinal (GI) disorders are among the most common comorbidities that are associated with autism. The Center for Disease Control recently found that children with ASD are more than 3.5 times more likely to suffer GI symptoms than normally developing children of the same age (Schieve, 2012). Common symptoms often include diarrhea, constipation, vomiting and reflux, abdominal pain and discomfort, gaseousness and unusually foul-smelling stools (Horvath, 2002. Molloy, 2003. Nikolov, 2009). In addition, individuals with ASD who have GI symptoms often display significantly higher measures of irritability, anxiety, and social withdrawal when compared to those without GI symptoms (Nikolov, 2009). It has also been found that there is a strong positive association between autism severity and GI dysfunction (Adams, 2011). It is this correlation that emphasizes the fact that therapies and treatments targeting the enteric microbiota and GI symptoms could be very effective in reducing the ASD-symptoms of autism as well.
In studying the intestinal microbiota of ASD patients, researchers have found many differences from the microbial norm. Specifically, it has been found that children with ASD have lower abundances of fermentative bacteria, low overall bacterial diversity, and higher levels of the toxin-producing Clostridium species (Finegold, 2002. Kang, 2013). It is this microbial imbalance that researchers hypothesize may contribute to ASD behavioral symptoms. The evidence supporting Clostridium’s role in ASD is presented in the next section.
Proposed Mechanisms
Clostridium & Sutterella
One well documented prediction regarding the mechanism underlying the link between the microbiota and ASD is the Clostridium Hypothesis. The Clostridium Hypothesis states that GI symptoms in ASD and the corresponding behavioral/social deficits are ultimately caused by toxin-producing clostridia species. Many parents of children with regressive-onset ASD have reported noticing the onset of social and behavioral changes characteristic of the disorder following repeated courses of antibiotics, accompanied by diarrhea (Bolte, 1998). It is from this observation that it was proposed that the disruption of intestinal microbiota caused by the antibiotics triggers the release of toxins by the clostridia. This hypothesis has been supported by a number of studies.
One such study found that children with ASD had a 10 fold higher level of Clostridium than healthy controls (Finegold, 2002). Another prominent study was conducted by Sandler (2000), in which children with ASD were given 6 weeks of oral vancomycin, an antibiotic that is known to work against Clostridium. At the end of treatment, participants were found to have significant improvement in both neurobehavioral as well as GI symptoms, supporting the idea that Clostridium is a main underlying cause of ASD (Sandler, 2000). However, after treatment ended symptoms returned, proving vancomycin treatment to not be a viable opportunity for therapy development (Sandler, 2000).
The bacteria Sutterella is also hypothesized to play a role in ASD. A significantly higher prevalence of Sutterella species was found in the GI tract of ASD children with GI disturbances compared to controls (no ASD) with GI disturbances (Williams, 2012). These findings suggest that Sutterella is a major component of the intestinal microbiota of children with autism and GI dysfunction and is absent in children with only GI dysfunction. Wang et al. (2013) also demonstrated elevated numbers of Sutterella in the feces of children with ASD as compared to healthy controls. In addition to this increased prevalence of Sutterella, (Kang, 2013) found reduced abundance of the species Prevotella in the intestinal microbiota of children with ASD. Prevotella is a bacteroidete, which are thought to be associated with good GI health (Kang, 2013). Therefore, this decrease in Prevotella combined with the increase in Sutterella suggests that ASD may result from a decrease in good bacteria and an increase in harmful bacteria. Unlike Clostridia, no mechanism for ASD has been suggested.
Intestinal Permeability & Immune Activation/Dysfunction
Another proposed mechanism underlying the relationship between the intestinal microbiota, the immune system and the brain in ASD is intestinal permeability, immune activation and dysfunction. It is known that the flow of substances between the intestines and the bloodstream is maintained by tight junctions (Hollander, 1999). The intestinal microbiota are essential to the maintenance of these junctions and therefore to the integrity of the intestinal barrier (Hsiao, 2013). When dysbiosis occurs the barrier may become compromised, referred to as “leaky gut,” introduced previously (Fasano, 2012). Leaky gut has been linked to a wide range of intestinal disorders (Fasano, 2012). This increased intestinal permeability likely allows for the passage of bacteria and their toxins into the bloodstream from the intestines that lead to immune activation.
One such compound that leaks from the intestines to the bloodstream is bacteria-derived lipopolysaccharide (LPS). LPS is a part of the cell membrane of gram-negative bacteria (Alexander, 2001). LPS in the bloodstream will elicit both an immunologic and inflammatory response (Qin, 2007). This response is characterized by increased systemic cytokines (Qin, 2007). Cytokines are cell-signaling molecules that facilitate cell-to-cell communication during immune responses. While cytokines are necessary for normal neurodevelopment, changes in their activities may negatively affect this development, as may be the case in ASD. Children with autism have been found to have increased levels of cytokines (Ashwood, 2011). Specifically, they show increased plasma levels of pro-inflammatory cytokines including IL-1B, IL-6, IL-8, and IL-12p40 (Onore, 2012). These elevated cytokine levels have been associated with characteristic behaviors, including poor communication and impaired social communication (Onore, 2012).
In postmortem studies of individuals with ASD, neuroinflammation in the brain has been a consistent finding (Li, 2009; Morgan, 2010). One study (Vargas, 2005) analyzed the autopsies and CSF of individuals with ASD and found evidence of excess microglial activation and increased cytokines compared to healthy controls. As discussed previously, microglia are the immune defense of the CNS, but they are also crucial in synaptogenesis and pruning (Bessis, 2007; Paolicelli, 2011). It is from this evidence that it is now hypothesized that this microglial deficit present in ASD results in weak synaptic transmission, decreased functional brain connectivity, deficits in social interactions, and increased repetitive behavior characteristic of ASD (Zhan, 2014).
Microbiota Transfer Therapy
Microbiota Transfer Therapy (MTT) is a modified fecal microbiota transplant (FMT) protocol and is a way to transplant healthy microbiota into the intestines of those with ASD. Specifically, a patient first receives 14 days of oral vancomycin treatment followed by 12-24 hours fasting with bowel cleansing. Repopulation of the gut microbiota occurs by administering a high initial dose of Standardized Human Gut Microbiota (SHGM) (Hamilton, 2012) either orally or rectally followed by daily, lower maintenance oral doses with a stomach acid suppressant for 7-8 weeks. In the study published by Kang et al in 2017, participants were followed for an additional 8 weeks after treatment ended, to determine if treatment effects were temporary or long-lasting (Kang, 2017).
Kang et al. found multiple significant results in those ASD patients who underwent MMT. First, GI symptoms were significantly improved for abdominal pain, indigestion, diarrhea and constipation. In addition, ASD-related behavior was also improved. Most importantly, it was found that the GI symptom improvement as well as ASD-related improvement were both present 8 weeks after treatment ended. In terms of intestinal microbiota, it was found that bacterial diversity increased in children with ASD and remained higher 8 weeks post treatment.
Overall, this study supports the validity of MMT while also showing that it is safe and well tolerated in children with ASD. By utilizing MMT as a therapeutic approach for children with ASD, patients will be able to see improvements in both GI- and ASD related symptoms that are long lasting. This finding that a change in intestinal microbiota of children with ASD towards that of a neurotypical child serves to support the hypotheses that intestinal microbiota are at least partially responsible for GI and ASD symptoms.
Summary/Discussion
While the exact cause of ASD remains unclear, it is certain that the imbalance in enteric microbiota found in those with ASD plays a role in not only GI symptoms common with the disorder, but in behavioral symptoms as well. It is through an understanding of the gut-brain axis, the relationship between the microbiome and the immune system, and the connection between the microbiome, the immune system, and the brain in ASD that researchers can begin to push for new therapies and treatments that bring about long-term, significant results in system improvement and long term change in the ASD community.