CHD8 regulates gut epithelial cell function and affects autism-related behaviours through the gut-brain axis

Background

Autism spectrum disorder (ASD) is a neuropsychiatric and neurodevelopmental condition Hundreds of gene mutations have been identified as potential causes of ASD, both in mice and humans (Mohrle et al. 2020). In particular, de novo mutations in the chromodomain helicase DNA binding protein 8 (CHD8) gene, also called DUPLIN, are strongly associated with ASD (O’Roak et al. 2011; O’Roak et al. 2012). The main symptoms of CHD8-related ASD are macrocephaly, intellectual disability, attentional deficit, gastrointestinal (GI) problems and sleeping disorder (Bernier et al. 2014). Knocking out of Chd8 in mice causes embryonic lethality (Nishiyama et al. 2004), suggesting the essential role of Chd8 in early neurodevelopment. Thus, only Chd8^+/- mice could serve as a rodent model of Chd8-related ASD. Chd8^+/- mice have anxiety-like, repetitive behavior and defects in sociability compared with wild type (WT) mice (Katayama et al. 2016). Moreover, Chd8^+/- mice also have slower GI movement, similar to human ASD patients (Bernier et al. 2014). Unfortunately, the connection between classic ASD symptoms and GI disorders have not been clarified so far.

The goal of this preprint by Elliott et al. was to reveal to possible relationship between ASD-related anxiety-like behavior and GI disorders in Chd8^+/- mice.

 

Key findings

Global mutant Chd8^+/-, gut epithelial cell specific mutant Chd8^ΔIEC and WT mice were subjected to various behavioral tests. These included elevated plus maze (EPM) and light-dark box (assessment of anxiety-like behavior); open-field and marble burying (repetitive behavior) and three chamber tests (social interaction). Less time spent in the open arms and center is associated with anxiety in the EPM and open-field test, respectively. The three-chamber test consists of a center chamber, a chamber of a stranger mouse and an empty chamber.  Mice with social deficits tend to visit the empty chamber over the chamber of a stranger mouse. All of these tests examine behaviors associated with ASD in mice models.

Small intestine and colon samples were collected for transcriptomics and histological analysis, while fecal samples were used for 16S rRNA gene sequencing, for investigating the role of microbiome in autism. Antibiotic treatment was an effective for certain types of ASD in the past (Kuhn et al. 2012). To test the effects of antibiotics on CHD8-related ASD, ciprofloxacin, metronidazole and vancomycin were added to the drinking water of Chd8^+/- and WT mice.

Gut permeability assay: serum samples of fasted mice were obtained after FITC Dextran administration.

Anxiety-like behavior of Chd8^ΔIEC mice.

In line with the previously reported data of Chd8^+/- mice, Chd8^ΔIEC mice exhibited anxiety-like behavior, compared to the WT mice.

Chd8^ΔIEC mice did not exhibit repetitive behavior and deficits in social behavior

The effect of antibiotic treatment (ciprofloxacin, metronidazole and vancomycin) on the social and anxiety-like behavior of Chd8^+/- mice.

Anxiety-like phenotype of Chd8^+/- mice was not changed after antibiotic treatment in any tests. However, social deficit behavior was rescued by antibiotic treatment; Chd8^+/- mice spent significantly more time with the stranger mouse in the three-chamber test compared to WT group.

Chd8^+/- mice showed an elevation in bacterial load in the colon samples and increased gut permeability, compared with WT mice.

Colon transcriptomics

581 genes were downregulated and 339 genes were upregulated in the gut epithelial cells of Chd8^+/- mice. Downregulated genes are involved in mitochondrial function while cell cycle-related genes were upregulated. In addition, genes of antimicrobial peptides were also upregulated, suggesting the response of the immune system against bacterial infection.

 

Why I liked this preprint

The authors used gut epithelial cell specific Chd8^ΔIEC mutant mice to test the symptoms of ASD, bringing a new insight into the discussion about CHD8 haploinsufficiency-related ASD.

 

Questions for the authors

1. Three different type of antibiotics were added to the drinking water of WT and Chd8^+/- mice. Did you check the serum levels of the antibiotics? What was the reason of the selection of those antibiotics? Did the antibiotic treatment reverse the anxiety-like behavior of Chd8^ΔIEC mutant mice as well?
2. Katayama and his research group tested the oligodendrocyte-specific Chd8^+/- KO mice, and found similar anxiety-like behavior (Kawamura et al. 2020), as in their original publication, using Chd8^+/- global mutant mice (Katayama et al. 2016). In your preprint, I did not find the Chd8^+/- vs. Chd8^ΔIEC comparisons. Are you planning to compare the anxiety-like behavior of Chd8^+/- and Chd8^ΔIEC mice?
3. In another recent article, cerebellar granule neuron progenitor (GNP)-specific Chd8 KO mice were characterized and cerebellar GNP cells was proposed to be a key regulator of CHD8-related ASD pathogenesis (Kawamura et al. 2021). How do these novel data fit into your hypothesis about the prominent role of colon microbiome changes in CHD8 haploinsufficiency-related ASD?

 

References

 

Bernier R., Golzio C., Xiong B., Stessman H.A., Coe B.P., Penn O., Witherspoon K., Gerdts J., Baker C., Vulto-van Silfhout A.T., Schuurs-Hoeijmakers J.H., Fichera M., Bosco P., Buono S., Alberti A., Failla P., Peeters H., Steyaert J., Vissers L., Francescatto L., Mefford H.C., Rosenfeld J.A., Bakken T., O’Roak B.J., Pawlus M., Moon R., Shendure J., Amaral D.G., Lein E., Rankin J., Romano C., de Vries B.B.A., Katsanis N. & Eichler E.E. (2014) Disruptive CHD8 mutations define a subtype of autism early in development. Cell 158, 263-76.

Katayama Y., Nishiyama M., Shoji H., Ohkawa Y., Kawamura A., Sato T., Suyama M., Takumi T., Miyakawa T. & Nakayama K.I. (2016) CHD8 haploinsufficiency results in autistic-like phenotypes in mice. Nature 537, 675-9.

Kawamura A., Katayama Y., Kakegawa W., Ino D., Nishiyama M., Yuzaki M. & Nakayama K.I. (2021) The autism-associated protein CHD8 is required for cerebellar development and motor function. Cell Rep 35, 108932.

Kawamura A., Katayama Y., Nishiyama M., Shoji H., Tokuoka K., Ueta Y., Miyata M., Isa T., Miyakawa T., Hayashi-Takagi A. & Nakayama K.I. (2020) Oligodendrocyte dysfunction due to Chd8 mutation gives rise to behavioral deficits in mice. Hum Mol Genet 29, 1274-91.

Kuhn M., Grave S., Bransfield R. & Harris S. (2012) Long term antibiotic therapy may be an effective treatment for children co-morbid with Lyme disease and autism spectrum disorder. Med Hypotheses 78, 606-15.

Mohrle D., Fernandez M., Penagarikano O., Frick A., Allman B. & Schmid S. (2020) What we can learn from a genetic rodent model about autism. Neurosci Biobehav Rev 109, 29-53.

Nishiyama M., Nakayama K., Tsunematsu R., Tsukiyama T., Kikuchi A. & Nakayama K.I. (2004) Early embryonic death in mice lacking the beta-catenin-binding protein Duplin. Mol Cell Biol 24, 8386-94.

O’Roak B.J., Deriziotis P., Lee C., Vives L., Schwartz J.J., Girirajan S., Karakoc E., Mackenzie A.P., Ng S.B., Baker C., Rieder M.J., Nickerson D.A., Bernier R., Fisher S.E., Shendure J. & Eichler E.E. (2011) Exome sequencing in sporadic autism spectrum disorders identifies severe de novo mutations. Nat Genet 43, 585-9.

O’Roak B.J., Vives L., Fu W., Egertson J.D., Stanaway I.B., Phelps I.G., Carvill G., Kumar A., Lee C., Ankenman K., Munson J., Hiatt J.B., Turner E.H., Levy R., O’Day D.R., Krumm N., Coe B.P., Martin B.K., Borenstein E., Nickerson D.A., Mefford H.C., Doherty D., Akey J.M., Bernier R., Eichler E.E. & Shendure J. (2012) Multiplex targeted sequencing identifies recurrently mutated genes in autism spectrum disorders. Science 338, 1619-22.

 

 

doi: https://doi.org/10.1242/prelights.30965

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