Ok-Jin Park1, Jiseon Kim1, Jihyun Yang1, Cheol-Heui Yun2, Seung Hyun Han3. 1. Department of Oral Microbiology and Immunology, DRI and BK21 Plus Program, School of Dentistry, Seoul National University, Seoul, Republic of Korea. 2. Department of Agricultural Biotechnology and Research Institute for Agriculture and Life Sciences, Seoul National University, Seoul, Republic of Korea. 3. Department of Oral Microbiology and Immunology, DRI and BK21 Plus Program, School of Dentistry, Seoul National University, Seoul, Republic of Korea. Electronic address: shhan-mi@snu.ac.kr.
Abstract
INTRODUCTION: Enterococcus faecalis is commonly found in root canals of patients with refractory apical periodontitis, often accompanying inflammation and malfunctioning bone regeneration. In this study, we investigated the effect of E. faecalis on osteoblast differentiation and the ability to induce chemokine expression to recruit inflammatory cells. METHODS: Osteoblast precursors from mouse calvaria were differentiated into osteoblasts with ascorbic acid and β-glycerophosphate in the absence or presence of heat-killed E. faecalis (HKEF). Alizarin red S staining was performed to determine the degree of mineralization. Reporter gene and reverse-transcription polymerase chain reaction assays were performed to examine the activity of the Runx2 transcription factor and the expression of osteogenic marker genes, respectively. Secretion of the chemokines keratinocyte-derived chemokine and monocyte chemotactic protein-1 was measured by the enzyme-linked immunosorbent assay, and their functions were analyzed by measuring the migration of peripheral blood mononuclear cells using a transwell system. RESULTS: HKEF inhibited osteoblast mineralization and Runx2 transcriptional activity, which are typical features of osteoblast differentiation. HKEF also decreased the expression of Runx2, osterix, β-catenin, osteocalcin, and type I collagen. Interestingly, however, the expression of keratinocyte-derived chemokine and monocyte chemotactic protein-1 was increased by HKEF, and the culture supernatant of HKEF-stimulated osteoblasts increased the transmigration of peripheral blood mononuclear cells. CONCLUSIONS: HKEF inhibits osteoblast differentiation and induces chemokine expression, which might be involved in refractory apical periodontitis and bone loss.
INTRODUCTION:Enterococcus faecalis is commonly found in root canals of patients with refractory apical periodontitis, often accompanying inflammation and malfunctioning bone regeneration. In this study, we investigated the effect of E. faecalis on osteoblast differentiation and the ability to induce chemokine expression to recruit inflammatory cells. METHODS: Osteoblast precursors from mouse calvaria were differentiated into osteoblasts with ascorbic acid and β-glycerophosphate in the absence or presence of heat-killed E. faecalis (HKEF). Alizarin red S staining was performed to determine the degree of mineralization. Reporter gene and reverse-transcription polymerase chain reaction assays were performed to examine the activity of the Runx2 transcription factor and the expression of osteogenic marker genes, respectively. Secretion of the chemokines keratinocyte-derived chemokine and monocyte chemotactic protein-1 was measured by the enzyme-linked immunosorbent assay, and their functions were analyzed by measuring the migration of peripheral blood mononuclear cells using a transwell system. RESULTS: HKEF inhibited osteoblast mineralization and Runx2 transcriptional activity, which are typical features of osteoblast differentiation. HKEF also decreased the expression of Runx2, osterix, β-catenin, osteocalcin, and type I collagen. Interestingly, however, the expression of keratinocyte-derived chemokine and monocyte chemotactic protein-1 was increased by HKEF, and the culture supernatant of HKEF-stimulated osteoblasts increased the transmigration of peripheral blood mononuclear cells. CONCLUSIONS: HKEF inhibits osteoblast differentiation and induces chemokine expression, which might be involved in refractory apical periodontitis and bone loss.