INTRODUCTION: Enterococcus faecalis is commonly detected in persistent apical periodontitis characterized by unimproved periradicular bone resorption. The aim of the present study was to examine the effect of lipoteichoic acid (LTA), a major virulence factor of E. faecalis, on apoptosis of osteoblasts. METHODS: Human osteoblast-like MG63 cells were treated with LTA from E. faecalis at a series of concentrations for 48 hours. The proliferation of the MG63 cells was assessed by 3-(4,5-dimethyl-thiazol-2-yl)-2,5- diphenyl-tetrazolium bromide assay. To examine the apoptosis, the LTA-treated cells were analyzed by flow cytometry by using annexin V-fluorescein isothiocyanate and propidium iodide double staining. Hoechst 33258 staining was performed to observe the morphologic changes of the cells. To investigate the apoptosis at the molecular level, the protein levels of Bcl-2 and Bax were determined by Western blot. Meanwhile, the caspase-3 activity was detected with caspase colorimetric protease assay. RESULTS: The proliferation of MG63 cells was inhibited by E. faecalis LTA in a dose-dependent manner. Flow cytometry assay indicated that LTA had a stimulating effect on MG63 cell apoptosis. Typical morphologies of apoptotic cells were observed under fluorescence microscope. Furthermore, the cell apoptosis was confirmed by (1) the down-regulation of the antiapoptotic protein (Bcl-2), (2) the up-regulation of the proapoptotic protein (Bax), and (3) the elevated caspase-3 activity. CONCLUSIONS: LTA of E. faecalis could inhibit the proliferation and induce apoptosis of human osteoblast-like MG63 cells.
INTRODUCTION:Enterococcus faecalis is commonly detected in persistent apical periodontitis characterized by unimproved periradicular bone resorption. The aim of the present study was to examine the effect of lipoteichoic acid (LTA), a major virulence factor of E. faecalis, on apoptosis of osteoblasts. METHODS:Human osteoblast-like MG63 cells were treated with LTA from E. faecalis at a series of concentrations for 48 hours. The proliferation of the MG63 cells was assessed by 3-(4,5-dimethyl-thiazol-2-yl)-2,5- diphenyl-tetrazolium bromide assay. To examine the apoptosis, the LTA-treated cells were analyzed by flow cytometry by using annexin V-fluorescein isothiocyanate and propidium iodide double staining. Hoechst 33258 staining was performed to observe the morphologic changes of the cells. To investigate the apoptosis at the molecular level, the protein levels of Bcl-2 and Bax were determined by Western blot. Meanwhile, the caspase-3 activity was detected with caspase colorimetric protease assay. RESULTS: The proliferation of MG63 cells was inhibited by E. faecalisLTA in a dose-dependent manner. Flow cytometry assay indicated that LTA had a stimulating effect on MG63 cell apoptosis. Typical morphologies of apoptotic cells were observed under fluorescence microscope. Furthermore, the cell apoptosis was confirmed by (1) the down-regulation of the antiapoptotic protein (Bcl-2), (2) the up-regulation of the proapoptotic protein (Bax), and (3) the elevated caspase-3 activity. CONCLUSIONS:LTA of E. faecalis could inhibit the proliferation and induce apoptosis of human osteoblast-like MG63 cells.
Authors: Katja Hattar; Christian P Reinert; Ulf Sibelius; Mira Y Gökyildirim; Florentine S B Subtil; Jochen Wilhelm; Bastian Eul; Gabriele Dahlem; Friedrich Grimminger; Werner Seeger; Ulrich Grandel Journal: Cancer Immunol Immunother Date: 2017-03-17 Impact factor: 6.968