Figure 5. Effect of T. denticola ggt gene deletion on death of hGF cells.

Wild type T. denticola (wt Td) or Δggt mutant bacteria and glutathione (gsh) were added in various combinations to different tissue culture flasks of human gingival fibroblast cells (hGFs) at near80% confluency. After 24 hours incubation, cell death (apoptosis plus necrosis) of hGFs was measured by flow cytometry. The experiment was done four times (biological replicates). The levels of H₂S in the media were also measured in one experiment. (A) The hGF cells from the samples indicated were stained with annexin V and propidium iodide followed by flow cytometry analysis (30). Annexin V levels were measured with a FITC-A filter set and propidium iodide staining was measured with a PerCP-A filter set. The results of one representative experiment (of four) are shown. (B) Quantification of the four flow cytometry plots, where % death of hGF cells in the indicated samples equals the percentage of cells that stained with Annexin V, propidium iodide, or both. (C) H₂S levels in the culture supernatants of the indicated samples. *, p = 0.001 per Tukey’s test analysis.