Fisetin Inhibits Periodontal Pathogen-Induced EMT in Oral Squamous Cell Carcinoma via the Wnt/β-Catenin Pathway

<b>Objective</b>: Previous reports showed that periodontopathic bacteria induce epithelial-mesenchymal transition (EMT) in oral squamous cell carcinoma (OSCC). Fisetin, a foodborne flavonoid, is reportedly associated with anticancer potential in various carcinogenic processes. This study aimed to elucidate the effects of fisetin on <i>Fusobacterium nucleatum</i>- and <i>Porphyromonas gingivalis</i>-induced EMT in OSCC cells. <b>Methods</b>: OSCC cells were co-cultured with live and heat-killed forms of <i>F. nucleatum</i> and <i>P. gingivalis</i>. The concentration of fisetin was set at 10 μM. Morphological changes in the OSCC cells were observed under a light microscope. Cell viability was measured using the Cell Counting Kit-8 assay, whereas migration was examined via wound healing. The mRNA expression of EMT-related markers was quantified using quantitative real-time polymerase chain reaction (PCR), and the expression of EMT-related markers and Wnt pathway-associated proteins was examined via Western blotting. <b>Results</b>: At a multiplicity of infection (MOI) of 300:1 for <i>F. nucleatum</i> and 100:1 for <i>P. gingivalis</i>, OSCC cell viability remained unchanged; however, wound closure rates increased significantly relative to the control. Likewise, treatment with fisetin (10 µM) did not materially alter viability; nevertheless, it attenuated promigratory effects induced by heat-killed periodontal pathogens at 3 h and 6 h. The OSCC cells exhibited EMT-like morphological changes after 6 h of co-culture with heat-killed pathogens. Consistently, reverse-transcriptase quantitative PCR and Western blot analyses showed increased expression of TWIST, ZEB1, and N-cadherin, accompanied by decreased E-cadherin expression, which was more pronounced in <i>F. nucleatum</i> than in <i>P. gingivalis</i>. However, fisetin reversed these trends. Moreover, co-culture with heat-killed pathogens markedly elevated β-catenin protein levels. In line with modulation of canonical Wnt/β-catenin signaling, fisetin and a Wnt inhibitor reduced β-catenin expression, whereas co-treatment with a Wnt agonist restored β-catenin levels in the presence of fisetin. <b>Conclusions</b>: Heat-killed <i>F. nucleatum</i> and <i>P. gingivalis</i> induced EMT in OSCC cells, with <i>F. nucleatum</i> exerting the strongest effect. Fisetin suppressed pathogen-driven EMT, at least partly via canonical Wnt/β-catenin signaling, highlighting its potential therapeutic value and warranting further investigation.