The Impacts of TNF-α-Induced Inflammation on Amnion Epithelial Cells: Exploring Stem Cell Gene Expression, Senescence, Inflammatory Responses, and Cellular Transition.

BACKGROUND: It is crucial to understand the relationship between inflammation and adverse pregnancy outcomes such as preterm birth and premature membrane rupture. Inflammation alters stem cell factors and cell transitions, disrupting immune balance and leading to adverse outcomes. OBJECTIVE: This study explored the effects of the pro-inflammatory cytokine TNF-α on Amnion epithelial cells (AECs), focusing on stem cell transcription factors (TFs), senescence, and the epithelial-mesenchymal transition (EMT). METHODS: Transcriptomic data were generated to compare the stem cell TFs expression (KLF4, c-MYC, OCT-4, SOX2, NANOG) between the placenta and fetal membrane (FM); AEC were treated with 50 ng/mL TNF-α for 48 h. Gene expression of stem cell TFs was assessed using qPCR; p38 MAPK activation and differential expression of KLF4, c-MYC were subsequently verified by Western blotting. Cellular senescence was evaluated using SA-β-Gal staining, and the pro-inflammatory cytokines IL-6 and IL-8 were measured using ELISA. The EMT was determined by measuring cell shape index and vimentin/CK-18 immunofluorescence. RESULTS: The placenta and the FM expressed higher mRNA levels of KLF4 than c-MYC, SOX2, OCT-4, and NANOG. AEC exhibited significantly higher KLF4 and c-MYC (p < 0.05). KLF4 downregulation (p < 0.05) and an increase in c-MYC (p < 0.01) in mRNA and protein levels were observed. The p38 MAPK activation (p < 0.01) increased cellular senescence (p < 0.05) and increased IL-6, IL-8 production (p < 0.01) with no change in cellular transition. CONCLUSION: This study shows how TNF-α affects stem cell TFs and impacts cellular integrity, senescence, and inflammatory responses, thus influencing adverse pregnancy outcomes.