Fig. 4: Butyrate provokes cellular senescence in HDFs.

a The concentrations of SCFAs present in indicated bacterial conditioned media or plain bacterial culture media of GAM (Mock) or modified GAM (Mock-mGAM) were shown. b–e Early passage TIG-3 cells were cultured for 9 days with tissue culture media containing indicated bacterial conditioned media at the ratio of 1/30 or indicated each SCFA or the mixture of these SCFAs in the same concentrations present in the bacterial conditioned media for 9 days. Representative photographs of the cells in the indicated culture conditions are shown. Scale bars represent 100 μm (b). Cells were then subjected to RT-qPCR analysis for indicated genes (one-way ANOVA followed by two-tailed Dunnett’s test for comparing with SCFAs mixture of E. coli. P values < 0.05 were considered significant) (c), or subjected to RNA sequencing analysis (d, e). Multi-dimensional scaling (MDS) plots based on the differential gene expression analysis were shown (d). Heatmap represents the ratio of gene expression between indicated cells and Mock-treated cells. The colours in the heatmap represent log fold-change (LogFC) relative to mock, with blue indicating three-fold lower expression and red three-fold higher expression (e). Three biological replicates of Mock and two biological replicates of the other conditions were analysed (d, e). The concentrations of doxorubicin (DXR) and Sodium butyrate used were 200 ng/ml or 3 mM, respectively. f Measurements of butyrate concentrations in the indicated bacterial conditioned media (left). Wild-type strain represents P. gingivalis (ATCC 33277) and Mutant strain represents its butyrate synthesis defective mutant (PGAGU 118)⁵³. Error bars indicate mean ± s.d. with three biologically independent replicates. Statistical significance was determined with a two-tailed Student’s t-test. P value < 0.05 was considered significant. Early passage TIG-3 cells were cultured with tissue culture media containing indicated bacterial conditioned media at the ratio of 1/30 for 9 days and representative photographs of the cells cultured with indicated bacterial conditioned media are shown (right). Source data are provided as a Source Data file.