Cumulative developmental perturbations alter adult male behaviour associated with epigenetic, ribosomal and immune dysregulation in the blood and brain of C57bl/6 mice

Environmental exposures across critical developmental windows can significantly influence brain development and contribute to the risk of neurodevelopmental disorders (NDDs). Importantly, emerging clinical evidence suggests that multiple environmental factors during early development synergise and result in more pronounced disease phenotypes in offspring. To expand upon this existing notion, we developed a ‘triple-hit’ mouse model to examine the combined effects of maternal social stress, chronic high-fat diet consumption, and early life poly(I:C) exposure on long-term developmental outcomes in offspring. We observed that ‘triple-hit’ male offspring displayed autism-like social deficits and an overall increased susceptibility to NDD-like behavioural alterations in adulthood. Single-cell RNA (scRNA) transcriptomic and bulk proteomic analyses were performed in male triple hit offspring across both brain tissue and peripheral blood leukocytes. scRNA sequencing revealed consistent dysregulation in critical glial cell processes, ribosomal functions and chromatin remodelling across all brain glial cell types. Similar themes were observed across peripheral blood leukocytes - displaying immune and ribosomal dysregulation at the transcriptional level. However, innate and adaptive immune cells displayed variations in directional expression of change. Proteomics analyses revealed significant reductions in proteins associated with ribosomal biogenesis, translation and chromatin remodelling pathways between brain and peripheral blood cells. Furthermore, significantly upregulated proteins associated with synaptic structure and function were observed between both compartments. Overall, using our unique triple hit model and by harnessing a systems-level approach, we demonstrate that early life environmental perturbations synergistically drive NDD-associated behaviours and complimentary transcriptional reprogramming between the central nervous system (CNS) and periphery in male offspring