Bioinformatics and machine learning identify ITIH4 and ZC3H13 as novel mRNA biomarkers for major depressive disorder that promote microglial M1 polarization.

Major depressive disorder (MDD), a prevalent mental illness, currently lacks reliable biomarkers and depends predominantly on subjective diagnostic criteria. Neuroinflammation, particularly microglial M1 polarization, plays a pivotal role in MDD pathogenesis. Inter-α-trypsin inhibitor heavy chain 4 (ITIH4) and zinc finger CCCH-type containing 13 (ZC3H13) are inflammation-related genes, but their roles in MDD remain unclear. The GSE217811 dataset was analyzed via limma to screen differentially expressed mRNAs; 21 inflammation-related ones underwent heatmap and LASSO (5-fold cross-validation), with candidate gene differential expression confirmed by unpaired t-tests. Candidate genes were initially screened and subsequently validated in an expanded cohort using qRT-PCR, with diagnostic efficacy evaluated by receiver operating characteristic (ROC) curves. siRNA-mediated silencing and plasmid-based overexpression of the two genes (ITIH4 and ZC3H13) were performed in BV2 microglial; the expression levels of target proteins, inflammatory cytokines, and the M1 marker CD86 were assessed using Western blot, ELISA, and flow cytometry. LASSO identified four candidate genes, with ITIH4 and ZC3H13 significantly upregulated in MDD. qRT-PCR confirmed their elevated serum expression, and ROC analysis showed high diagnostic efficacy. Silencing ITIH4 and ZC3H13 inhibited LPS-induced inflammation and M1 polarization, while overexpression exacerbated these effects. ITIH4 and ZC3H13 are potential MDD biomarkers that regulate microglial inflammation and M1 polarization, providing insights for MDD diagnosis and targeted therapy. GRAPHICAL ABSTRACT: The novel mRNA biomarkers ITIH4 and ZC3H13, upregulated in MDD serum with high diagnostic value, promote neuroinflammation via microglial M1 polarization, bridging MDD diagnosis and neuroinflammatory pathogenesis.