Porphyromonas gingivalis LPS inhibits osteoblastic differentiation and promotes pro-inflammatory cytokine production in human periodontal ligament stem cells
Introduction
One of the most common causes of tooth loss is periodontal disease, which is defined as the destruction of the alveolar bone and periodontal ligament by bacterial infection.1 Many of the bacteria that cause periodontal disease are Gram-negative bacilli, such as Porphyromonas gingivalis (P. gingivalis), Fusobacterium nucleatum, and Aggregatibacter actinomycetemcomitans.2, 3 P. gingivalis has been implicated as the causal pathogen in most cases of periodontal disease.4, 5, 6 Lipopolysaccharide (LPS) is an outer membrane component of Gram-negative bacterial cell walls and an endotoxin with a wide range of biological activities. LPS induces the production of inflammatory cytokines, such as interleukin-1 β (IL-1β), IL-6, and IL-8.7, 8 These cytokines and LPS induce alveolar bone resorption and destroy periodontal tissue.9
Periodontal tissue is composed of the periodontal ligament (PDL), cementum, alveolar bone, and gingiva. The PDL is composed of fibrous connective tissue that connects the tooth root to the alveolar bone.10 PDL cells have roles in maintaining tissue regeneration11. PDL cells produce bone-related proteins; they also express alkaline phosphatase (ALP) activity, collagen type 1 alpha 1, osteocalcin, and possess mineralisation capacity.12, 13, 14 Although PDL cells are a component of connective tissues, they also exhibit osteoblastic differentiation capacity.
Periodontal ligament stem cells (PDLSCs) exist in periodontal tissues and participate in periodontal tissue regeneration.15 Similar to other mesenchymal stem cells (MSCs), PDLSCs have the potential to differentiate into osteoblasts.16, 17 PDLSCs have tissue regeneration capacity.18, 19, 20 PDLSCs exhibit greater potential to regenerate periodontal tissues than other MSCs, such as bone marrow stromal cells.21 Therefore, PDLSCs play a particularly important role in periodontal tissue regeneration. Moreover, PDLSCs are predicted to be an important source of cells in periodontal tissue regeneration therapy. However, no studies have reported the effects of LPS on PDLSCs; therefore, this area of investigation is critical.
The aim of this study was to characterize the effects of P. gingivalis LPS on osteoblastic differentiation and mineralisation in PDLSCs. Additionally, we examined the production of inflammatory cytokines, such as IL-1β, IL-6, and IL-8 in the PDLSCs.
Section snippets
Culture and isolation of human PDLSCs
The human tissue experiments were performed in accordance with the guidelines of the Osaka Dental University for Medical Ethics, and all experiments were approved by the Osaka Dental University Medical Ethics Committee (approval no. 110712). All participants provided written informed consent to participate in this study, and the study design was approved by the appropriate ethics review boards. PDLSCs were isolated and cultured as described in our previous study.22 After extraction, the teeth
Isolation and characterisation of PDLSCs
The PDL-derived cells formed clonogenic clusters of fibroblast-like cells (Fig. 1A). No staining was detected in control (Fig. 1B). The PDLSCs derived from the PDL cells were positive for vimentin (Fig. 1C). The PDLSCs were also positive for STRO-1 (Fig. 1D) and SSEA-4 (Fig. 1E), which are MSC markers. The isolated PDLSCs were capable of differentiating into osteogenic cells (Fig. 1F).
Cell proliferation
The proliferation of the PDLSCs in the normal culture medium containing P. gingivalis LPS was significantly
Discussion
The results of this study indicated that P. gingivalis LPS enhanced the proliferation of PDLSC cells. The osteoblastic differentiation assay revealed that the ALP activity, COL1A1 production, osteocalcin production, and mineralisation were significantly suppressed by P. gingivalis LPS. P. gingivalis LPS also enhanced the production of IL-1β, IL-6, and IL-8.
PDLSCs express STRO-1, a marker of bone marrow mesenchymal stem cells (BMSCs).16 SSEA-4 is also a marker of MSCs23 and PDLSCs.24 In this
Funding
No additional external funding was received for this study. The funders played no role in the study design, data collection and analysis, decision to publish, or manuscript preparation.
Competing interests
The authors report no conflicts of interest related to this study.
Ethical approval
Not required.
Acknowledgments
This study was partially supported by Grants-in-Aid for Scientific Research from the Japan Society for the Promotion of Science (C) (no. 24593138 to A.T. and no. 24792345 to Y.T.).
References (50)
- et al.
Periodontal Dis
Lancet
(2005) - et al.
Protein expression and functional difference of membrane-bound and soluble receptor activator of NF-kappaB ligand: modulation of the expression by osteotropic factors and cytokines
Biochem Biophys Res Commun
(2000) - et al.
Investigation of multipotent postnatal stem cells from human periodontal ligament
Lancet
(2004) - et al.
Periodontal ligament stem cell-mediated treatment for periodontitis in miniature swine
Stem Cells
(2008) - et al.
Comparison of different tissue-derived stem cell sheets for periodontal regeneration in a canine 1-wall defect model
Biomaterials
(2011) - et al.
SSEA-4 identifies mesenchymal stem cells from bone marrow
Blood
(2007) - et al.
Lipopolysaccharide from Escherichia coli but not from Porphyromonas gingivalis induce pro-inflammatory cytokines and alkaline phosphatase in dental follicle cells
Arch Oral Biol
(2012) - et al.
Lipopolysaccharides can protect mesenchymal stem cells (MSCs) from oxidative stress-induced apoptosis and enhance proliferation of MSCs via Toll-like receptor(TLR)-4 and PI3K/Akt
Cell Biol Int
(2009) - et al.
Effects of lipopolysaccharide on newly established rat dental pulp-derived cell line with odontoblastic properties
J Endod
(2007) - et al.
Molecular mechanisms of the inhibitory effect of lipopolysaccharide (LPS) on osteoblast differentiation
Biochem Biophys Res Commun
(2010)
The bone-resorbing activities in tissue culture of lipopolysaccharides from the bacteria Actinobacillus actinomycetemcomitans, Bacteroides gingivalis and Capnocytophaga ochracea isolated from human mouths
Arch Oral Biol
Structure of periodontal tissues in health and disease
Periodontology 2000
The breadth of bacterial diversity in the human periodontal pocket and other oral sites
Periodontology 2000
Immune responses and vaccination against periodontal infections
J Clin Periodontol
Distribution of 3-hydroxy iC17:0 in subgingival plaque and gingival tissue samples: relationship to adult periodontitis
Infect Immun
Relationship between hydroxy fatty acids and prostaglandin E2 in gingival tissue
Infect Immun
Bacteria induce osteoclastogenesis via an osteoblast-independent pathway
Infect Immun
Review of osteoimmunology and the host response in endodontic and periodontal lesions
J Oral Microbiol
Periodontal regeneration: a challenge for the tissue engineer?
Proc Inst Mech Eng H
Collagen resorption by fibroblasts. A theory of fibroblastic maintenance of the periodontal ligament
J Periodontol
Expression of extracellular matrix proteins in human periodontal ligament cells during mineralization in vitro
J Periodontol
Biological effects of cementum and bone extracts on human periodontal fibroblasts
J Periodontol
Storage conditions of avulsed teeth affect the phenotype of cultured human periodontal ligament cells
J Periodontal Res
Stem cell properties of human periodontal ligament cells
J Periodontal Res
Isolation and characterization of human periodontal ligament (PDL) stem cells (PDLSCs) from the inflamed PDL tissue: in vitro and in vivo evaluations
J Clin Periodontol
Cited by (167)
Controlled drug delivery from metronidazole-containing bioactive endodontic cements
2023, Dental MaterialsInhibition of Wnt11 impairs the osteogenesis and aggravates the inflammatory response of human mesenchymal stem cells under LPS-induced inflammatory condition
2023, Biochemical and Biophysical Research CommunicationsEffects of a Low Glucose Condition on Proliferation, Differentiation and Autophagy in Mouse Osteoblast-Like Cells
2024, Journal of Hard Tissue BiologyProanthocyanidin enhances the endogenous regeneration of alveolar bone by elevating the autophagy of PDLSCs
2023, Journal of Periodontal Research