en Behavioral Neuroscience Behavioral Neuroscience Original Original <div style="text-align: justify;"><strong>Introduction</strong>: This study aimed to elucidate the molecular mechanism underlying the anti-anxiety effects of flaxseed (Linum usitatissimum) by investigating its role in modulating the oxytocin (OXT) signaling pathway through protein-protein interaction (PPI) network analysis.<br> <strong>Methods</strong>: Gene expression profiles from the Gene Expression Omnibus (GEO) dataset (GSE36422) were analyzed to identify flaxseed-induced differentially expressed genes (DEGs). Bioinformatics tools, including functional enrichment (KEGG, GO) and PPI network construction (STRING, Cytoscape), were used to map key genes and pathways. Hub and bottleneck proteins were identified using Cyto-Hubba&rsquo;s Maximal Clique Centrality (MCC) and Betweenness algorithms, focusing on genes shared between flaxseed and OXT signaling.<br> <strong>Results</strong>: Of 98 initially DEGs, 56 were significantly regulated by flaxseed. Three critical genes CALM3&nbsp;(upregulated),&nbsp;NFATC4, and&nbsp;RAF1&nbsp;(both downregulated) emerged as shared mediators between flaxseed and OXT pathways. CALM3, a calcium-sensing hub protein, exhibited extensive network connectivity, interacting with 58% of OXT pathway proteins and functioning as a bottleneck regulator. Pathway analysis revealed flaxseed&rsquo;s influence on OXT signaling, cellular senescence, and the cyclic guanosine monophosphate-activated protein kinase G&nbsp;(cGMP-PKG) pathway. NFATC4 downregulation suggested disinhibition of OXT signaling, while RAF1&rsquo;s role in MAPK cascades further supported anxiolytic effects.<br> <strong>Conclusion</strong>: Flaxseed exerts its anti-anxiety effects primarily through&nbsp;CALM3-mediated OXT signaling, leveraging calcium-dependent pathways. These findings provide mechanistic evidence supporting flaxseed as a natural anxiolytic and highlight its potential as a dietary intervention for anxiety disorders.</div> Linum usitatissimum, Oxytocin (OXT) signaling, Anxiety, CALM3, Protein-protein interaction (PPI) network 1169 1178 http://bcn.iums.ac.ir/browse.php?a_code=A-10-8066-1&slc_lang=en&sid=1 Hajar Sadat Sadeghi Darya.sadeghi0073@gmail.com 13700319475328460056661 13700319475328460056661 No Department of Traditional Pharmacy, Traditional Medicine and Materia Medica Research Center, School of Traditional Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran. Mostafa Rezaei Tavirani Tavirany@yahoo.com 13700319475328460056662 13700319475328460056662 Yes Proteomics Research Center, Shahid Beheshti University of Medical Sciences, Tehran, Iran. Melika Abrishami abrishamimelika28@gmail.com 13700319475328460056663 0009-0001-9487-7799 No Proteomics Research Center, Shahid Beheshti University of Medical Sciences, Tehran, Iran. Maryam Hamzeloo-Moghadam mhmoghadam@sbmu.ac.ir 13700319475328460056664 13700319475328460056664 No Department of Traditional Pharmacy, Traditional Medicine and Materia Medica Research Center, School of Traditional Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran.