Mechanistic insights into retinoic-acid treatment for autism in the improvement of social behavior: Evidence from a multi omics study in rats.

Background: Autism spectrum disorder (ASD) is a lifelong condition. It is characterized by complex etiologies, including disruptions in exogenous retinoic acid (RA) signaling, which may serve as an environmental risk factor. Targeting the RA pathway presents a promising therapeutic avenue, though the precise mechanisms remain to be elucidated.

Methods: Female Sprague-Dawley rats were treated with valproic acid (VPA) during pregnancy to induce an ASD model in their offspring. Some offspring received RA treatment postnatally. Social behavior and brain-functional connectivity were assessed using behavioral tests and functional magnetic resonance imaging (fMRI), respectively. Transcriptomics analysis and proteomics analysis of the hypothalamus identified differentially expressed genes (DEGs) and differentially expressed proteins (DEPs). These were intersected with ASD pathogenic genes (APGs) and ASD pathogenic proteins (APPs) to identify differentially expressed APGs (DE-APGs) and differentially expressed APPs (DE-APPs), which were validated by real-time reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and western blotting. Analyses of enrichment of signaling pathways were done using the Kyoto Encyclopedia of Genes and Genomes database.

Results: RA treatment significantly improved social behaviors and revealed distinct patterns of hypo- and hyper-connectivity across various brain regions, with notable changes involving the hypothalamus and facial nerve. Differential analysis revealed 4165 DEGs (DEG 1) and 329 DEPs (DEP 1) between control and VPA groups, and 1610 DEGs (DEG 2) and 197 DEPs (DEP 2) between VPA and RA supplementation (RAS) groups. Twenty-two DE-APGs and five DE-APPs were identified, with key associations found between proteins such as Tbl1xr1 and Myo5a and >13 genes including Nrxn1, Cacna1e, and Gabrb2. Significant alterations in DE-APGs, including Grin2b, Nrxn1, Cacna1e, and Gabrb2, were confirmed via real-time RT-PCR and western blotting. In addition, 22 key signaling pathways were enriched in DEPs and DEGs.

Conclusion: RA supplementation in ASD rats induced by VPA may ameliorate social deficits and modulated functional connectivity, especially in the hypothalamus and facial nerve regions. This suggests potential therapeutic benefits for neural circuitry dysregulation in ASD. Additionally, RA altered critical gene and protein expressions in hypothalamus, implicating its role in modulating key signaling pathways to mitigate social deficits in ASD. This study provides new insights into the molecular mechanisms of ASD and supports the development of novel therapeutic strategies.