Transcriptome profile of human macrophages co-cultured with human supracrestal gingival connective tissue-derived MSCs

Objectives: The supracrestal gingival connective tissue (SG) plays a pivotal role in maintaining periodontal health due to its proximity to the gingival sulcus, where it is exposed to microbial biofilms. Disruption of this tissue can lead to inflammation, apical migration of the junctional epithelium, and loss of periodontal support. Mesenchymal stem cells derived from the supracrestal gingival connective tissue (SG-MSCs) may contribute to local immune regulation and tissue homeostasis; however, their role in modulating macrophage function during periodontal inflammation remains unclear. This study aimed to investigate the molecular interactions between SG-MSCs and human macrophages through transcriptomic profiling, focusing on immunomodulatory pathways. Methods: Peripheral blood monocytes from three healthy donors were differentiated into human monocyte-derived macrophages (HMDMs), while SG-MSCs were isolated from a healthy donor and characterized for MSC markers, colony-forming ability, and trilineage differentiation potential. HMDMs were cultured alone or co-cultured with SG-MSCs under transwell conditions to allow paracrine interaction. Total RNA was extracted from HMDMs and subjected to RNA sequencing. Differentially expressed genes (DEGs) were identified and analyzed using bioinformatics tools for functional enrichment and pathway analysis (GO and KEGG). Results: Transcriptomic analysis revealed substantial transcriptional alterations in HMDMs co-cultured with SG-MSCs compared to monocultured controls. Enriched biological processes included cell chemotaxis, extracellular matrix organization, and cytokine-mediated signaling. Molecular function analysis showed significant upregulation of oxidoreductase activity, while cellular component enrichment indicated enhanced extracellular matrix-related gene expression. KEGG pathway analysis identified upregulation of the cytokine–cytokine receptor interaction pathway. Notably, key DEGs such as CCL1, CCL24, CXCL5, CXCR5, IL1B , and TGFB2 were upregulated, suggesting enhanced macrophage chemotactic and immunomodulatory activity in response to SG-MSC interaction. Conclusions: SG-MSCs modulate macrophage gene expression toward a pro-regenerative and chemotactic phenotype, highlighting their role in regulating inflammatory responses and promoting tissue remodeling. These findings provide molecular evidence supporting the therapeutic potential of SG-MSCs in periodontal inflammation control and regeneration strategies.