Intestinal Dysbiosis and BBB Integrity in Autism

Autism Spectrum Disorder (ASD) is a neurodevelopmental disorder characterized by impaired social interactions and communication, associated with repetitive behaviors and restricted interests. The pathophysiology of ASD is complex due to clinical heterogeneity and numerous comorbidities. Surprisingly, up to 70% of children with ASD have gastrointestinal (GI) disorders, correlated with the severity of ASD. These GI disorders are influenced by eating disorders linked to ASD. There is also an imbalance of the intestinal microbiota in ASD. In addition, the gut microbiota plays a key role in regulating the gut-brain axis. It is involved in brain development and neuroinflammation, one of the pathophysiological tracks in ASD. Neuroinflammation is also involved in regulating the integrity of the blood-brain barrier (BBB), which is affected in ASD. The question if intestinal dysbiosis influences the BBB integrity in ASD therefore arises.

The main objective is to determine if there is an association between the composition of the intestinal microbiota and the integrity of the BBB in ASD.

Secondary objectives are i/ to identify the pathophysiological pathways involved in this association, in particular via the neuroinflammation processes, ii/ to estimate the influence of this association on some ASD characteristics, iii/ to determine the influence of nutritional status on the measured biological and clinical parameters.

This is a monocentric, observational and cross-sectional study of 72 children with ASD, aged 6 to 16, living in the Languedoc-Roussillon region and from the ELENA cohort (this cohort includes 900 French children with an ASD, initially aged 2 to 16, and followed for 6 years).

Serum immunoassays will be assessed for the measurement of neuroinflammation, plasma assays for the intestinal inflammation and permeability. Clinical characteristics will be collected by a clinician and parental questionnaires (ASD severity test, level of social interaction, repetitive and stereotypical behavior, behavioral disorders, quality of life, intensity of GI disorders). Nutritional status will be assessed by serum metabolome analysis and anthropomorphic measures.

Participants will be stratified according to the intensity of their ASD characteristics and the presence or absence of GI disorders.

Descriptive analyses of microbiota composition, biological markers of intestinal permeability and inflammation, neuroinflammation and BBB integrity will be performed using an Heat Map Correlation. Principal component analysis will allow to identify patterns of microbiota compositions. Associations between the different groups of measured biomarkers (microbiota patterns, BBB integrity, intestinal permeability, intestinal inflammation and neuroinflammation) will be performed using multivariate regression models. The models will be systematically adjusted for age and gender.

The study will last 4 and a half years and will end in February 2025. The inclusions will last 42 months. No specific follow-up is planned outside the 6-month maximum period between the study proposal and the inclusion visit. The study will be presented to patients at the end of their visit at 3 or 6 years of follow-up as part of the ELENA cohort. For patients meeting the inclusion criteria and who agrees to participate in the study, two visits will occur at the Center of Autism Ressources of Languedoc-Roussillon (CRA-LR), for the collection of consent and the delivery of the material to be used at home. In the week preceding their second visit to CRA-LR, parents will collect a sample of their child's stool in a dedicated jar which will be packed in a plastic bag and kept in the home-deep-freeze until they reach CRA-LR. A Bristol scale - allowing the display of the child's stools -, a PedsQL GSS questionnaire - evaluating gastrointestinal disorders - and a RBS-R questionnaire - evaluating repetitive and stereotypical behaviors - will be completed by parents. During the inclusion visit, a clinical examination of children will be performed and a blood sample will be collected.

This study will concomitantly assess many clinical and biological parameters, some of whom never explored before in children with ASD. It will allow a better understanding of the implication of the gut-brain axis in the ASD pathophysiology and will open the way to new therapeutic targets.