Endocannabinoids and Inflammatory Response in Periodontal Ligament Cells

Abstract

Endocannabinoids are associated with multiple regulatory functions in several tissues. The main endocannabinoids, anandamide (AEA) and 2-arachidonylglycerol (2-AG), have been detected in the gingival crevicular fluid of periodontitis patients, but the association between periodontal disease or human periodontal ligament cells (hPdLCs) and endocannabinoids still remain unclear. The aim of the present study was to examine the effects of AEA and 2-AG on the proliferation/viability and cytokine/chemokine production of hPdLCs in the presence/absence of Porphyromonas gingivalis lipopolysaccharide (P. gingivalis LPS). The proliferation/viability of hPdLCs was measured using 3,4,5-dimethylthiazol-2-yl-2,5-diphenyl tetrazolium bromide (MTT)-assay. Interleukin-6 (IL-6), interleukin-8 (IL-8), and monocyte chemotactic protein-1 (MCP-1) levels were examined at gene expression and protein level by real-time PCR and ELISA, respectively. AEA and 2-AG did not reveal any significant effects on proliferation/viability of hPdLCs in the absence of P. gingivalis LPS. However, hPdLCs viability was significantly increased by 10–20 µM AEA in the presence of P. gingivalis LPS (1 µg/ml). In the absence of P. gingivalis LPS, AEA and 2-AG did not exhibit any significant effect on the expression of IL-8 and MCP-1 expression in hPdLCs, whereas IL-6 expression was slightly enhanced by 10 µM 2-AG and not affected by AEA. In P.gingivalis LPS stimulated hPdLCs, 10 µM AEA down-regulated gene-expression and protein production of IL-6, IL-8, and MCP-1. In contrast, 10 µM 2-AG had an opposite effect and induced a significant up-regulation of gene and protein expression of IL-6 and IL-8 (P<0.05) as well as gene-expression of MCP-1 in P. gingivalis LPS stimulated hPdLCs. Our data suggest that AEA appears to have an anti-inflammatory and immune suppressive effect on hPdLCs’ host response to P.gingivalis LPS, whereas 2-AG appears to promote detrimental inflammatory processes. In conclusion, AEA and 2-AG might play an important role in the modulation of periodontal inflammation.

Human periodontal ligament cells exhibit no endotoxin tolerance upon stimulation with Porphyromonas gingivalis lipopolysaccharide

“…Isolation of mRNA and transcription into cDNA was performed using the TaqMan Gene Expression Cells‐to‐CT kit (Ambion/Applied Biosystems, Foster City, CA, USA) as described in our previous studies . The mRNA expression levels of interleukin (IL)‐6, IL‐8, monocyte chemotactic protein (MCP)‐1, TLR2 and TLR4 were determined by qPCR with the GAPDH encoding gene as internal reference.…”

Methanandamide diminish the Porphyromonas gingivalis lipopolysaccharide induced response in human periodontal ligament cells

“…Cell proliferation/viability was measured by the MTT method, as described in our previous study [18]. hPdLCs were seeded in 24 well plates at a density of 2 × 10 4 cells in 500 μL of DMEM supplemented with 10% FBS.…”

“…In vitro studies show that anandamide stimulates the proliferation of human gingival fibroblasts [15] and diminishes cytokine production by these cells in response to stimulation with P. gingivalis LPS [14]. Our recent study shows that AEA and 2-AG have a different effect on P. gingivalis LPS induced production of interleukin (IL)-6, IL-8, and monocyte chemoattractant protein (MCP)-1 [18]. Mainly, P. gingivalis LPS induced response was inhibited by AEA and enhanced by 2-AG [18].…”

“…Our recent study shows that AEA and 2-AG have a different effect on P. gingivalis LPS induced production of interleukin (IL)-6, IL-8, and monocyte chemoattractant protein (MCP)-1 [18]. Mainly, P. gingivalis LPS induced response was inhibited by AEA and enhanced by 2-AG [18]. An in vivo study using the ligature periodontitis model in rats shows that the local application of AEA decreases the content of tumor necrosis factor-alpha and IL-1β in gingival tissue [19].…”

A new target to ameliorate the damage of periodontal disease: The role of transient receptor potential vanilloid type‐1 in contrast to that of specific cannabinoid receptors in rats

“…In recent years, many studies have focused on the research of the ECS and its implication on immune and inflammatory diseases, including periodontal disease. [31][32][33] Most of these studies have approached this issue through cells cultures such as human gingival fibroblasts and ligament periodontal cells. 34 However, our research group reported anti-inflammatory effects on gingival tissue and osteoprotective effects on alveolar bone induced by both a CB2r and a CB1r agonist (HU308 and meth-AEA, respectively), in a model of LPS-induced periodontal disease in vivo.…”