antibody did not prevent the augmenta tion of P. gingivalis invasion by TNF. TNF augments invasion of P. gingivalis through NF ��B and MAPK pathways To determine whether mRNA synthesis and protein syn thesis were required for P. gingivalis invasion, Ca9 22 cells were preincubated with 1 ug ml of the RNA poly merase II inhibitor actinomycin D or the protein syn thesis inhibitor cyclohe imide for 1 h and were then incubated with TNF prior to addition of P. gingivalis. Actinomycin D and cyclohe imide e hibited significant invasion of P. gingivalis augmented by TNF. PDTC also e hibited significant inhibitory activity towards the invasion of P. gingivalis enhanced by TNF. These results suggest that TNF augmented invasion of P. gingivalis is mediated by p38 and JNK pathways and activation of NF ��B.
ICAM 1 mediates invasion of P. gingivalis E pression Drug_discovery of ICAM 1 is required for invasion of some bacteria in KB cells. To determine whether ICAM 1 affects P. ginigvalis invasion into cells, we first e amined co localization of P. gingivalis with ICAM 1 in cells. Ca9 22 cells were incubated with P. gingivalis, and localization of ICAM 1 and P. ginigvalis in the cells was observed by a confocal laser scanning microscope. ICAM 1 strongly e pressed around the cell surface was partially co localized with P. gingivalis in the cells. We also e amined the e pression of ICAM 1 in TNF treated Ca9 22 cells. Ca9 22 cells were treated with or without TNF for 3 h. The cells were lysed and e pression of ICAM 1 was analyzed by Western blotting. ICAM 1 was e pressed in Ca9 22 cells with out TNF stimulation.
However, TNF increased the e pression of ICAM 1 in the cells. We ne t e amined whether ICAM 1 is associated with in vasion of P. gingivalis into the cells. Ca9 22 cells were treated with TNF for 3 h, incubated with an anti ICAM 1 antibody or a control IgG antibody for an additional 2 h, and then incubated with P. gingivalis. Anti ICAM 1 antibody suppressed invasion of P. gin givalis in the cells with or without TNF pretreat ment. In contrast, P. gingivalis invasion was not prevented by control IgG. These results sug gest that ICAM 1 is partially associated with invasion of P. gingivalis into Ca9 22 cells. Rab5 mediates endocytosis of P. gingivalis Several studies have shown that Rab5 regulates events in the fusion of bacteria containing vacuoles and early endosomes.
Therefore, we investigated whether Rab5 mediates P. gingivalis invasion into cells. We first were incubated with P. gingivalis for 1 h. Internalization of P. gingivalis into the cells was reduced by silencing the Rab5 gene. To determine whether the Rab5 affects P. ginigvalis invasion into cells, Ca9 22 cells e pressing GFP Rab5 were treated with P. gingivalis, and localization of Rab5 and P. ginigvalis in the cells was observed by a confocal laser scanning microscope. Transfected GFP Rab5 was partially co localized with P. gingivalis in the cells. These results sug gest that Rab5 is partially associated with in