IADR Abstract Archives
Endotoxin Tolerance in Human Periodontal Ligament Cells
Objectives: The endotoxin tolerance (ET) is a preventive mechanism developed by immune cells and other cytokine producing cells, characterized by a state of hyporesponsiveness due to treatment with endotoxins such as lipopolysaccharide (LPS) in low concentrations.
The aim of this study was to investigate, whether the fibroblast-like human periodontal ligament cells (hPDLC) are able to develop ET like state upon stimulation with Porphyromonas (P.) gingivalis LPS in low concentrations. Further, possible alteration in toll-like receptor (TLR) 2- and TLR4- induced response upon pre-stimulation with P. gingivalis LPS was assessed.
Methods: Primary hPDLC isolated from five healthy donors were pretreated with P. gingivalis LPS in a concentration of 0.1µg/ml that leads to a submaximal inflammatory response as determined in preliminary tests. After 24 hours, pretreated and non-pretreated cells were stimulated with one of the following stimuli: P. gingivalis LPS (1µg/ml); TLR4 agonist E. coli LPS (1µg/ml); TLR2 agonist Pam3CSK4 (1µg/ml).
The protein expression of interleukin (IL)-6, IL-8, and monocyte chemotactic protein (MCP)-1 was analyzed with qPCR and ELISA. Gene expression levels of TLR2 and TLR4 were determined by qPCR.
Results: Stimulation of hPDLC with P. gingivalis LPS, E. coli LPS, and Pam3CSK4 resulted in significantly increased production of IL-6, IL-8, and MCP-1. Pretreatment of cells with low concentration of P. gingivalis LPS had no significant effect on the production of these proteins. The gene expression of TLR2 showed no significant difference between the pretreated and non-pretreated group, whereas gene expression of TLR4 was significantly decreased in pretreated cells compared to non-pretreated cells.
Conclusions: The results of this study show that hPDLC do not develop ET, suggesting an important role in the progression of periodontal disease by excessive cytokine expression. Furthermore, this study demonstrates that the amplitude of the inflammatory response of hPDLC to bacterial LPS shows no significant dependency on TLR4 expression levels.
The aim of this study was to investigate, whether the fibroblast-like human periodontal ligament cells (hPDLC) are able to develop ET like state upon stimulation with Porphyromonas (P.) gingivalis LPS in low concentrations. Further, possible alteration in toll-like receptor (TLR) 2- and TLR4- induced response upon pre-stimulation with P. gingivalis LPS was assessed.
Methods: Primary hPDLC isolated from five healthy donors were pretreated with P. gingivalis LPS in a concentration of 0.1µg/ml that leads to a submaximal inflammatory response as determined in preliminary tests. After 24 hours, pretreated and non-pretreated cells were stimulated with one of the following stimuli: P. gingivalis LPS (1µg/ml); TLR4 agonist E. coli LPS (1µg/ml); TLR2 agonist Pam3CSK4 (1µg/ml).
The protein expression of interleukin (IL)-6, IL-8, and monocyte chemotactic protein (MCP)-1 was analyzed with qPCR and ELISA. Gene expression levels of TLR2 and TLR4 were determined by qPCR.
Results: Stimulation of hPDLC with P. gingivalis LPS, E. coli LPS, and Pam3CSK4 resulted in significantly increased production of IL-6, IL-8, and MCP-1. Pretreatment of cells with low concentration of P. gingivalis LPS had no significant effect on the production of these proteins. The gene expression of TLR2 showed no significant difference between the pretreated and non-pretreated group, whereas gene expression of TLR4 was significantly decreased in pretreated cells compared to non-pretreated cells.
Conclusions: The results of this study show that hPDLC do not develop ET, suggesting an important role in the progression of periodontal disease by excessive cytokine expression. Furthermore, this study demonstrates that the amplitude of the inflammatory response of hPDLC to bacterial LPS shows no significant dependency on TLR4 expression levels.