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Figure 7 | BMC Microbiology

Figure 7

From: TNF-α augmented Porphyromonas gingivalis invasion in human gingival epithelial cells through Rab5 and ICAM-1

Figure 7

Rab5 mediates endocytosis of P. gingivalis. (A) Ca9-22 cells were transfected with 100 pmol siRNA specific for Rab5 or control siRNA using Lipofectamine 2000 reagent, as described by the manufacturer. Then expression of Rab5 in the cells was examined by Western blotting. (B) Rab5 siRNA-transfected Ca9-22 cells were incubated with P. gingivalis for 1 h. Viable P. gingivalis in the cells was determined as described in Methods. (Means ± standard deviations [SD] [n = 3]. **, P < 0.01 versus control siRNA. (C) Ca9-22 cells were transfected with expression vectors with inserted genes of GFP alone and GFP-Rab5. The cells were incubated with P. gingivalis for 1 h. The cells were then stained using anti-P. gingivalis antiserum. Each molecule was visualized as follows: GFP and GFP-Rab5 (green) and P. gingivalis (red). Bars in each panel are 10 μm. (D) Active form of Rab5 colocalizes with P. gingivalis in Ca9-22 cells. Ca9-22 cells were transfected with vectors with inserted genes of GFP alone (control), GFP-Rab5 (S34N) (inactive form of Rab5), and GFP-Rab5 (Q79L) (active form of Rab5). The cells were incubated with P. gingivalis for 1 h. Then localization of P. gingivalis and Rab5 in the cells was observed by a confocal laser scanning microscope. Each molecule was visualized as follows: GFP and GFP-Rab5 (green) and P. gingivalis (red). Bars in each panel are 10 μm. (E) Overexpression of the active form of Rab5 increased invasion of P. gingivalis in Ca9-22 cells. Ca9-22 cells were transfected with expression vectors with inserted genes of GFP alone (Control), GFP-Rab5 (S34N) and GFP-Rab5 (Q79L). Viable P. gingivalis in the cells was determined as described in Methods. (Means ± standard deviations [SD] [n = 3]). *, P < 0.05 versus control; **, P < 0.01 versus GFP alone.

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