Spleen-tonifying formula alleviates social deficits, gut dysbiosis, and hypomyelination in a perinatal injury model.

Perinatal injury has been shown to induce autism spectrum disorder (ASD)-like phenotypes in offspring, including social behavioral deficits, hypomyelination, and gut microbial imbalances. Spleen-tonifying formulas (STFs), derived from traditional Chinese medicine, have demonstrated efficacy in modulating the brain-gut axis, particularly by alleviating hypomyelination and microbial dysbiosis in offspring with maternal immune activation-induced perinatal injury. However, whether STF can improve behavioral deficits in such offspring with perinatal injury remains unclear. To evaluate the therapeutic efficacy of STF, a battery of behavioral tests-including the three-chamber social interaction, grooming, rotarod, and open-field tests-was conducted in a two-hit perinatal injury rat model induced by prenatal lipopolysaccharide (LPS) exposure and postnatal hypoxic stress. Starting at 6 weeks of age, rats received daily oral administration of STF extract or distilled water for 14 days before behavioral assessments. Myelination and oligodendrocyte density in the prefrontal cortex were examined via immunohistochemistry. Fecal microbiota composition was profiled using 16S rRNA gene sequencing, and in vitro oxidative stress assays were performed in human oligodendroglioma (HOG) cells. STF treatment significantly improved social behavior and restored myelination in offspring with perinatal injury, as evidenced by increased MBP expression and elevated numbers of Olig2+ and NG2+ cells in the prefrontal cortex. In vitro, STF protected differentiated HOG cells from H2O2-induced oxidative stress and cell death by sustaining AKT activation and inhibiting ERK signaling. Gut microbiota analysis revealed that STF effectively ameliorated dysbiosis, notably enriching bacterial families such as Oscillospiraceae and Colidextribacter, which were positively correlated with improved social behavior, suggesting potential as biomarkers of therapeutic response. Our results indicate that oral STF treatment is associated with improvements in social behavior, myelination, and gut microbial composition in offspring with perinatal injury, underscoring the need for future studies to elucidate the potential causal relationships among these outcomes.