Lipopolysaccharide Activates the Unfolded Protein Response in Human Periodontal Ligament Fibroblasts.

Abstract

The activation of the unfolded protein response (UPR) has been demonstrated in periodontal diseases. However, the cellular and molecular mechanisms by which the UPR is induced in periodontitis remain unclear. In this study, the effects of lipopolysaccharide (LPS) on the induction of the UPR in human periodontal ligament fibroblasts (HPDLFs) in vitro are investigated. HPDLFs isolated from human PDLs were stimulated with various concentrations of Escherichia coli LPS (0.1, 1, and 10 μg/mL) for the indicated time points (0, 3, 6, 9, 12, and 18 hours). The messenger RNA (mRNA) and protein levels of molecular markers associated with UPR activation, such as glucose-regulated protein 78 (GRP78), X-box binding protein 1 (XBP1), and C/EBP homologous protein (CHOP), were measured at different time points of LPS treatment. Apoptosis of HPDLFs was assessed by Annexin V-FITC and propidium iodide staining, followed by flow cytometry. LPS treatment of HPDLFs increased GRP78 mRNA and protein levels in a concentration-dependent manner. Additionally, LPS also induced the expression, splicing, and activation of XBP1 mRNA. Moreover, LPS-induced CHOP expression was concentration dependent: a lower concentration of LPS (0.1 μg/mL) had no effect on CHOP mRNA levels, but higher concentrations of LPS (1 and 10 μg/mL) markedly increased CHOP mRNA and protein expression without inducing apoptosis. The findings demonstrate that activating the Toll-like receptor-4 signaling pathway in HPDLFs using LPS triggers the UPR in vitro, warranting additional investigation into the precise mechanisms by which pathways promote this response under inflammatory conditions.