Effect of heat stress on the expression levels of receptor activator of NF-κB ligand and osteoprotegerin in human periodontal ligament cells.

  To investigate the expression levels of the receptor activator of NF-κB ligand (RANKL) and osteoprotegerin (OPG) in human periodontal ligament fibroblasts (PDLs) when stimulated with heat.   Periodontal ligament fibroblasts were subjected to various temperature increases for 5 min and then maintained at 37 °C. After that, cell viability was determined using an MTT assay. The expression levels of RANKL and OPG were investigated using real-time RT-PCR and ELISA. As a control, the cells were cultured at 37 °C. Data were analysed using one-way anova at a significant level of P = 0.05. Results  Cell viability was reduced significantly in the heated groups (P < 0.05). Heat stress downregulated the mRNA expression levels of RANKL and OPG (P < 0.05). When the cells were heated at 39 °C, the protein release of OPG was increased (P < 0.05). There were no significant differences between the heated groups and the control in the release of soluble RANKL (P > 0.05). The relative RANKL/OPG expression ratios were decreased at 39, 43 and 50 °C (P < 0.05), but increased at 47 °C (P < 0.05).   Heat influenced the balance between RANKL and OPG in PDLs. Low heat downregulated their relative ratio, whilst high heat upregulated it.

Zhang L ，Zhou X ，Wang Q ，Wang Y ，Tang L ，Huang D ... -  《-》

Effects of Enterococcus faecalis lipoteichoic acid on receptor activator of nuclear factor-κB ligand and osteoprotegerin expression in periodontal ligament fibroblasts.

To investigate the influence of Enterococcus faecalis lipoteichoic acid (LTA) on the key bone resorption-regulating proteins, receptor activator of nuclear factor-κB ligand (RANKL) and osteoprotegerin (OPG) in human periodontal ligament fibroblasts (PDL cells). Periodontal ligament cells were subjected to various concentrations of LTA. Cell viability was then determined by methyl thiazolyl tetrazolium (MTT) assay, whilst the expression levels of RANKL and OPG were investigated by enzyme-linked immunosorbent assay (ELISA) and Western blotting. The effect of the inhibitors [IL-1 receptor-associated kinase (IRAK)-1/4, p38 mitogen-activated protein kinase (MAPK) (SB203580)] on LTA-stimulated RANKL/OPG activation was examined. Cell viability and RANKL/OPG ratio in PDL cells were also analysed by MTT assay and Western blotting. Data were analysed using one-way anova or t-test at a significance level of P = 0.05. Cell viability was reduced significantly in the LTA group in a dose-dependent fashion (P < 0.05). In addition, LTA was found to upregulate the protein expression of RANKL, OPG and their relative ratio in PDL cells (P < 0.05). The optimal concentration of LTA used in PDL cells was determined to be 10 μg mL(-1) . Following IRAK1/4 and p38MAPK inhibition, LTA-stimulated increases of RANKL/OPG ratio were significantly reduced (P < 0.05). Enterococcus faecalis LTA could upregulate the expression of RANKL and OPG at different rates, suggesting a potential role for LTA in the bone resorption process of refractory apical periodontitis through the regulation of RANKL and OPG. In addition, IRAK1/4 and p38MAPK signalling involving RANKL/OPG may contribute to inflammatory responses from PDL cells.

Zhao L ，Chen J ，Cheng L ，Wang X ，Du J ，Wang F ，Peng Z ... -  《-》

Osteogenic induction and 1,25-dihydroxyvitamin D3 oppositely regulate the proliferation and expression of RANKL and the vitamin D receptor of human periodontal ligament cells.

Tang X ，Meng H 《-》

Effect of hydrogen sulphide on the expression of osteoprotegerin and receptor activator of NF-κB ligand in human periodontal ligament cells induced by tension-force stimulation.

Hydrogen sulphide (H2S) is an endogenous gaseous signalling molecule, the generation rate of which is affected by mechanical force in cells. Recently, it was reported that mechanical force plays an important role in some pathways such as osteoprotegerin (OPG) and the receptor activator of nuclear factor kappa B (NF-κB) ligand (RANKL) in human periodontal ligament cells (hPDLCs). Here, we investigated whether H2S played a regulatory role within the periodontal remodelling process by addressing the expression level of OPG/RANKL in hPDLCs. hPDLCs were first applied with tension force for 0-120min to select the optimal time for force application. These cells were treated with H2S for 24h followed by stimulation with tension-force application. Cell proliferation and apoptosis were assessed by the methyl thiazolyl tetrazolium (MTT) assay and flow cytometry analysis. For OPG, RANKL and cystathionine-γ-lyase (CSE), real-time polymerase chain reaction (PCR) was used to analyse the messenger RNA (mRNA) expression; enzyme-linked immunosorbent assay (ELISA) was used to detect the secretion of OPG and soluble RANKL (sRANKL). Tension force promoted the mRNA expression of CSE and the optimal application time was 60min. The expression of OPG was increased in a concentration-dependent manner by H2S treatment. Importantly, the relative OPG/RANKL expression ratio was significantly increased upon induction by H2S, an effect that was enhanced by tension-force application. H2S could promote osteogenic differentiation by regulating the relative OPG/RANKL expression ratio of hPDLCs, which is enhanced by tension force. These findings may be valuable for understanding the mechanism of H2S in the periodontal remodelling, especially in the process of tooth movement.

Liao C ，Hua Y 《-》

Effects of IL-10 and glucose on expression of OPG and RANKL in human periodontal ligament fibroblasts.

Zhang L ，Ding Y ，Rao GZ ，Miao D ... -  《-》