Effect of interleukin-1ß on ICAM-1 expression of dental pulp cells: role of PI3K/Akt, MEK/ERK, and cyclooxygenase.

OBJECTIVE: Interleukin-1ß (IL-1ß) is an important inflammatory mediator of the dental pulp. IL-1ß stimulates cyclooxygenase-2 (COX-2) expression and prostanoid production of pulp cells and affects the inflammatory and healing processes of the dental pulp. There are two interleukin-1 (IL-1) receptors, IL-1RI and IL-1RII, with opposing effect after activation. However, the expression of IL-1Rs, the effects of IL-1ß on intercellular adhesion molecule-1 (ICAM-1) of dental pulp cells, and its relation to protein kinase B (Akt) signaling and COX activation are not clear. METHOD: Human dental pulp cells were treated with IL-1ß with/without pretreatment and co-incubation by LY294002 (a PI3K/Akt inhibitor), U0126 [a mitogen-activated protein kinase kinase/extracellular signal-regulated kinase (MEK/ERK) inhibitor], aspirin (a COX inhibitor), or eugenol (a COX inhibitor) for different time periods. The expression of ICAM-1, IL-1RI, and IL-1RII messenger RNA (mRNA) was evaluated by reverse transcriptase-polymerase chain reaction (RT-PCR). ICAM-1 protein expression was examined by western blotting. Soluble ICAM-1 (sICAM-1) level in the culture medium was determined by enzyme-linked immunosorbent assay (ELISA). Activation of Akt and ERK by IL-1ß was measured by Pathscan p-Akt ELISA or western blot. Viable cell number was evaluated by 3-(4,5-dimethyldiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay. RESULTS: Dental pulp cells expressed IL-1RI, but little IL-1RII. IL-1ß stimulated COX-2 and ICAM-1 mRNA and protein expression as well as sICAM-1 production of pulp cells. Aspirin and eugenol enhanced the IL-1ß-induced sICAM-1 production and ICAM-1 expression. IL-1ß rapidly activated Akt and ERK. LY294002 and U0126 attenuated IL-1ß-induced ICAM-1 expression and sICAM-1 production. CONCLUSIONS: These results reveal that IL-1ß may be involved in the pulpal inflammatory processes by stimulating ICAM-1 expression and secretion. These events are associated with IL-1RI expression and differential activation of PI3K/Akt and MEK/ERK and COX. CLINICAL RELEVANCE: Pharmacological inhibition of IL-1ß, IL-1RI, COX-2, ICAM-1, and related signaling pathways (MEK/ERK and PI3K/Akt) may be useful for the control of pulpal inflammation.

In vitro effect of laser irradiation on cementum-bound endotoxin isolated from periodontally diseased roots.

BACKGROUND: In a previous study, we evaluated the in vivo effects of an Nd:YAG laser on periodontal disease by measuring crevicular interleukin (IL)-1beta levels before and after laser application. It was found that laser therapy was less effective than traditional scaling and root planing. These results might be due to incomplete removal of microbial residues and cementum-bound endotoxin on root surfaces by the laser. In this study, we explored the in vitro effectiveness of an Nd:YAG laser for the elimination of cementum-bound endotoxin by measuring IL-1beta changes in stimulated monocytes. METHODS: Fresh human monocytes were harvested from adults without periodontitis and grown in RPMI 1640 medium. Diseased cementum particles were collected and prepared from teeth with untreated periodontitis and were irradiated with 5 levels of laser energy. Cementum particles were subjected to endotoxin testing by a limulus amebocyte lysate (LAL) assay and then were incubated with cultured monocytes. Production of IL-1beta in stimulated monocytes was measured by enzyme-linked immunosorbent assay and quantified by spectrophotometry. RESULTS: The endotoxin unit (EU) of diseased cementum was 18.4 EU/mg, which seemed to be remarkably lower than that of common periodontal pathogens including Porphyromonas gingivalis (381) at 15,300 EU/mg/ml, Prevotella intermedia (ATCC 25611) at 227 EU/mg/ml, and Fusobacterium nucleatum (ATCC 25586) at 1,987 EU/mg/ml. Monocytes subjected to stimulation by diseased cementum particles without laser irradiation produced 124 to 145 pg/ml IL-1beta, 9- to 18-fold higher than that of unstimulated monocytes (7.07 to 15.95 pg/ml). Diseased cementum particles after irradiation with various energy levels of the Nd:YAG laser could still stimulate monocytes to secrete 89 to 129 pg/ml IL-1beta. No statistically significant difference was found in the production of IL-1beta induced by diseased-bound cementum with or without laser irradiation. CONCLUSIONS: The Nd:YAG laser varying from 50 mJ, 10 pps to 150 mJ, 20 pps, for 2 minutes, did not seem to be effective in destroying diseased cementum endotoxin.