FIG. 7.

H. somnus culture filtrates induce endothelial cell apoptosis. Volumes containing 5 × 10⁴ adherent endothelial cells were treated for 6 h with culture filtrates prepared from H. somnus organisms cultured in BHI-YT (■) (33% final volume; prepared with approximately 10⁸ CFU) at 37°C with 5% CO₂, using the same isolates described in Fig. 5. Heat-inactivated (□) (60°C for 60 min) culture filtrates were tested in the same experiments. Controls consisted of untreated cells (UT), cells treated with BHI-YT (33% final volume), and staurosporine (200 nM; Stauro)-treated endothelial cells. The data illustrate the mean ± SEM of three separate experiments. Statistical analysis was performed by ANOVA followed by the Tukey-Kramer significant difference test. Significant levels of endothelial cell apoptosis (P < 0.05) were detected when endothelial cells were incubated with H. somnus culture filtrates or heat-inactivated filtrates and were compared to control (untreated) endothelial cells. Heat inactivation of culture filtrates significantly reduced the level of endothelial cell apoptosis (P < 0.05) compared to that of culture filtrates.