Flavonoid myricetin, usually found in tea and medicinal plants, has antioxidant and anti-inflammatory effects. Our objectives in this study were to verify the effects of myricetin on periodontal ligament fibroblasts (PDLFs) under inflammatory conditions and to observe its effects on osteoclast generation and on cytokine expression in RAW264.7 cells. To determine the effects of myricetin on PDLFs, we examined the expression and activity of proteolytic enzymes, including MMP-1, MMP-2, and MMP-8, which all play an important role in chronic periodontitis. We observed the effects of myricetin on intracellular signal transduction to verify the molecular mechanism involved. By measuring the formation of TRAP–positive multinucleated cells and the expression and activity of MMP-8, we were able to assess the effects of myricetin on osteoclast generation. In addition, by measuring the secretion of IL-6 and NO, we could evaluate the effects of myricetin on inflammatory mediators. We found that Myricetin had no effect on the viability of the PDLFs in the presence of inflammation, but it did decrease both the expression of MMP-1 and MMP-8 and the enzyme activity of MMP-2 and MMP-8 in these fibroblasts. Myricetin also decreased the lipopolysaccharide-stimulated phosphorylation of JNK, p38 signaling, IKKB, AKT, and p65RelA in the PDLFs. In the RAW264.7 cells, myricetin inhibited both the expression and the activity of MMP-8. Furthermore, Myricetin not only suppressed the generation of LPS-stimulated osteoclasts, but it also slightly inhibited LPS-stimulated degradation of IkB and decreased the release of LPS-induced IL-6 and NO. These findings suggest that myricetin alleviates the tissue-destructive processes that occur during periodontal inflammation.
Chronic Periodontitis ; Fibroblasts ; Inflammation* ; Interleukin-6 ; Osteoclasts ; Peptide Hydrolases ; Periodontal Ligament ; Phosphorylation ; Plants, Medicinal ; Signal Transduction ; Tea

This study examined the anti-osteoclastogenic effects of baicalin on receptor activator of NF-kB ligand (RANKL)-induced RAW264.7 cells. Baicalin is a flavonoid that is produced by Scutellaria baicalensis and is known to have multiple biological properties, including antibacterial, anti-inflammatory and analgesic effects. The effects of baicalin on osteoclasts were examined by measuring 1) cell viability; 2) the formation of tartrate-resistant acid phosphatase (TRAP) (+) multinucleated cells; 3) RANK/RANKL signaling pathways and 4) mRNA levels of osteoclast-associated genes. Baicalin inhibited the formation of RANKL-stimulated TRAP (+) multinucleated cells and also suppressed the RANKL-stimulated activation of p-38, ERK, cSrc and AKT signaling. Baicalin also inhibited the RANKL-stimulated degradation of IkappaB in RAW264.7 cells. In addition, the RANKL-stimulated induction of NFATc1 transcription factors was found to be abrogated by this flavonoid. Baicalin was further found to decrease the mRNA expression of osteoclast-associated genes, including carbonic anhydrase II, TRAP and cathepsin K in the RAW264.7 cells. Our data thus demonstrate that baicalin inhibits osteoclastogenesis by inhibiting the RANKL-induced activation of signaling molecules and transcription factors in osteoclast precursors.
Acid Phosphatase ; Carbonic Anhydrase II ; Cathepsin K ; Flavonoids ; Isoenzymes ; NF-kappa B ; NFATC Transcription Factors ; Osteoclasts ; RNA, Messenger ; Scutellaria baicalensis ; Transcription Factors

Luteolin is a flavonoid that exists in a glycosylated form in celery and green pepper. Flavonoids possess antioxidant and anti-inflammatory properties and can reduce the expression of key inflammatory molecules in macrophages and monocytes. It has been reported also that some flavonoids have effects on bone metabolism. The effects of luteolin on the function of osteoblasts were investigated by measuring cell viability, alkaline phosphatase activity, type I collagen production, osteoprotegerin secretion, Wnt promoter activity, BMP-2 and Runx2 expression and calcified nodule formation. Luteolin has no effects upon osteoblast viability but induced an increase in alkaline phosphatase activity, type I collagen production and a decrease in osteoprotegerin secretion in these cells. Luteolin treatment also upregulated BMP-2 mRNA expression. These results suggest that luteolin may be a regulatory molecule that facilitates the differentiation of osteoblasts.
Alkaline Phosphatase ; Apium graveolens ; Capsicum ; Cell Survival ; Collagen Type I ; Flavonoids ; Luteolin ; Macrophages ; Monocytes ; Osteoblasts ; Osteoprotegerin ; RNA, Messenger

Recent study on the enamel matrix derivatives explained on the effects of new bone and new attachment formation in infrabony pocket of periodontal defects. The purpose of this study was to investigate on the biological effects of enamel matrix derivatives to attachment, proliferation and activation of periodontal ligament and osteoblast cells. After treatment of osteoblast and PDL cells with various Emdogain concentration level(0.03microgram/ml, 3microgram/ml, 300 microgram/ml), activation of osteogenetic factor, calcified nodule formation and measuring alkaline phosphatase activity(ALP) were performed. 1. Both osteoblast and PDL cell showed increasing initial cell attachment with 300microgram/ml Emdogain concentration. 2. At the level of 300microgram/ml, accelerated proliferation of oseoblast and PDL cell was appeared. 3. As Emdogain's concentration increased, increased ALP activation of osteoblast was shown. In case of PDL cell, Emdogain increased ALP activation prominently at the level of 300microgram/ml. 4. No statistically significant activating change were founded at all of the concentrations of Emdogain on the activating of transcript factor Runx2 for differentiating osteoblast. 5. At the level of 300microgram/ml, calcified nodule formation was increased prominently to compare with other concentration. These results indicated that Emdogain should activate initial attachment, proliferation and activation, but not on Runx2 activation and can be used for useful tool of the treatment of periodontal tissue regeneration.
Alkaline Phosphatase ; Dental Enamel* ; Fibroblasts* ; Osteoblasts* ; Periodontal Ligament* ; Regeneration

Rheumatoid arthritis, osteoarthritis, and periodontal disease are bone destructive diseases mainly caused by inflammation. Various studies are being conducted to develop treatments for inflammatory bone destructive diseases. Many of these studies involve plant-derived natural compounds. In these studies, cell differentiation, signal transduction pathways, and bone resorption were measured at the cellular level. In disease-induced animal models, the amount of inflammatory mediators or matrix destructive enzymes and serum metabolic markers were measured. This study examined the effects of plant-derived natural compounds, such as flavonoids, on inflammatory bone destructive diseases. In addition, we structurally classified various substances used to maintain bone health and summarized the biological effects and related mechanisms of the components.

Periodontal disease is an inflammatory disease that affects the destruction of the bone supporting the tooth and connective tissues surrounding it. Periodontal ligament fibroblasts (PDLFs) induce overexpression of matrix metalloproteinase (MMP) involved in periodontal diseaseʼs inflammatory destruction. Osteoclasts take part in physiological bone remodeling, but they are also involved in bone destruction in many kinds of bone diseases, including osteoporosis and periodontal disease. This study examined the effect of baicalin on proteolytic enzymesʼ production and secretion of inflammatory cytokines in PDLFs and RAW 264.7 cells under the lipopolysaccharide (LPS)-induced inflammatory conditions. Baicalin inhibited the expression of the protein, MMP-1 and MMP-2, without affecting PDLFs’ cell viability, suggesting its possibility because of the inhibition of phosphorylation activation of mitogen-activated protein kinase’s p38, and the signal transduction process of nuclear factor κB (NFκB)-related protein. Also, baicalin reduced the expression of MMP-8 and MMP-9 in RAW 264.7 cells. This reduction is thought to be due to the inhibition of the signal transduction process of NFκB-related proteins affected by inhibiting p65RelA phosphorylation. Also, baicalin inhibited the secretion of nitric oxide and interleukin-6 induced by LPS in RAW 264.7 cells. These results suggest that baicalin inhibits connective tissue destruction in periodontal disease. The inhibition of periodontal tissue destruction may be a therapeutic strategy for treating inflammatory periodontal-diseased patients.

Periodontal disease is an inflammatory disease that affects the destruction of the bone supporting the tooth and connective tissues surrounding it. Periodontal ligament fibroblasts (PDLFs) induce overexpression of matrix metalloproteinase (MMP) involved in periodontal diseaseʼs inflammatory destruction. Osteoclasts take part in physiological bone remodeling, but they are also involved in bone destruction in many kinds of bone diseases, including osteoporosis and periodontal disease. This study examined the effect of baicalin on proteolytic enzymesʼ production and secretion of inflammatory cytokines in PDLFs and RAW 264.7 cells under the lipopolysaccharide (LPS)-induced inflammatory conditions. Baicalin inhibited the expression of the protein, MMP-1 and MMP-2, without affecting PDLFs’ cell viability, suggesting its possibility because of the inhibition of phosphorylation activation of mitogen-activated protein kinase’s p38, and the signal transduction process of nuclear factor κB (NFκB)-related protein. Also, baicalin reduced the expression of MMP-8 and MMP-9 in RAW 264.7 cells. This reduction is thought to be due to the inhibition of the signal transduction process of NFκB-related proteins affected by inhibiting p65RelA phosphorylation. Also, baicalin inhibited the secretion of nitric oxide and interleukin-6 induced by LPS in RAW 264.7 cells. These results suggest that baicalin inhibits connective tissue destruction in periodontal disease. The inhibition of periodontal tissue destruction may be a therapeutic strategy for treating inflammatory periodontal-diseased patients.

PURPOSE: We performed the present study to investigate whether osteotropic hormomes play roles on the nitric oxide (NO) production in culture of ROS17/2.8 osteoblastic cells. MATERIALS AND METHODS: The osteoblastic cell line ROS17/2.8 cells were cultured in F12 medium supplemented with 5% fetal bovine serum (FBS) at 37 degrees C in a humidified atmosphere of 5% CO2 in air. ROS17/2.8 cells were plated in 96-well plates at a density of 2-3x10 (3) cells/well and grown to confluence. Then the cells were pretreated with osteotropic hormones (parathyroid hormone (PTH) 20-500 ng/mL, 1, 25-dihydroxycholecalciferol (1, 25[OH]2D3) 1-100 nM; prostaglandin E2 (PGE2) 20-500 ng/mL) in the medium supplemented with 0.4% FBS for 72 hours and the cells were treated with cytokines (TNFalpha and IFNgamma) in phenol red-free F12 medium for an additional 48 hours. NO synthesis was assessed by measuring the nitrite anion concentration, the reaction product of NO, in the cell culture medium using Griess reagent. RESULTS: PTH and 1, 25[OH]2D3 pretreatment induced a significant increase in NO production in the presence of TNFalpha and IFNgamma. PGE2 slightly induced NO production compared to the control group. But, PGE2 pretreatment did not affect in NO production in the presence of TNFalpha and IFNgamma. CONCLUSIONS: These results suggest that the actions of osteotropic hormones in bone metabolism may be partially mediated by NO in the presence of cytokines.
Atmosphere ; Cell Culture Techniques ; Cell Line ; Cytokines ; Dinoprostone ; Metabolism ; Nitric Oxide* ; Osteoblasts ; Phenol ; Tumor Necrosis Factor-alpha

Natriuretic peptides comprise a family of three structurally related peptides; atrial natriuretic peptide (ANP), brain natriuretic peptide (BNP), and C-type natriuretic peptide (CNP). The present study was performed to investigate the effect of ANP on the proliferation and activity of ROS17/2.8 and HOS cells which are well-characterized osteoblastic cell lines. ANP dose-dependently decreased the number of ROS17/2.8 and HOS cells after 48-hour treatment. ANP generally increased the alkaline phosphatase activity of ROS17/2.8 and HOS cells after 48 hr treatment, regardless of the fact that basal activity of alkaline phosphatase was much lower in HOS cells compared to that of ROS17/1.8 cells. ANP increased the NBT reduction by ROS17/2.8 and HOS cells. ANP showed the variable but no significant effect on the nitric oxide production by ROS17/2.8 and HOS cells. ROS17/2.8 and HOS cells produced and secreted gelatinase into culture medium, and this enzyme was thought to be the gelatinase A type with the molecular weight determination. The gelatinase activity produced by ROS17/2.8 cells was increased by the treatment of ANP. However, the enzyme activity was not affected by ANP treatment in the HOS cell culture. In summary, ANP decreased the proliferation and increased the alkaline phosphatase activity and NBT reduction of osteoblasts. These results indicate that ANP is one of the important regulators of bone metabolism.
Alkaline Phosphatase ; Atrial Natriuretic Factor ; Cell Culture Techniques ; Cell Line ; Gelatinases ; Humans ; Matrix Metalloproteinase 2 ; Metabolism ; Molecular Weight ; Natriuretic Peptide, Brain ; Natriuretic Peptide, C-Type ; Natriuretic Peptides ; Nitric Oxide ; Osteoblasts* ; Peptides