Figure 4. Inhibition of ICAM-1-dependent PMN adhesion to oral epithelial cells by Rgp. (A) HSC-2 cells were pre-treated with or without 10³ U/mL IFN- for 3 days at 37°C. Cells were then treated with or without 0.3 µmol/L RgpB for 30 min at 37°C. After cells were washed with PBS, the expression of ICAM-1 was analyzed by flow cytometry. Representative findings of three independent experiments are expressed as the means of the MFI. **P < 0.01 vs. respective control. HSC-2 cells (B) and primary oral (gingival) epithelial cells (C) were pre-treated with or without 10³ U/mL IFN- for 3 days at 37°C. Cells were then treated with 0.3 µmol/L RgpB in the presence of 3 µmol/L FPR-cmk, anti-ICAM-1 mAb (10 µg/mL), or isotype-matched control IgG1 (10 µg/mL) for 30 min at 37°C. After the treatment, they were washed with warmed medium three times, and co-cultured with calcein-labeled PMNs (5 x 10⁵ cells/well) for 30 min at 37°C. At the end of the incubation, non-adherent PMNs were washed away with warmed PBS three times, and adherent PMNs were evaluated by means of a spectrophotofluorometer. We used serial dilutions of calcein-labeled PMNs as a standard to calculate the number of adherent PMNs. Results are expressed as the number of adherent cells. Representative findings of three independent experiments are expressed as the mean ± SD of the binding of PMNs. **P < 0.01 vs. respective control.

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