뼈흡수유도호르몬이 ROS17/2.8세포로부터 Nitric Oxide 형성에 미치는 영향

고선일,김민성,한원정,김세원,김정근 대한구강악안면방사선학회 2005 Imaging Science in Dentistry Vol.35 No.3

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?C in a humidified atmosphere of 5% CO₂ in air. ROS17/2.8 cells were plated in 96-well plates at a density of 2-3×10³ 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 (TNFα and IFNγ) 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 TNFα and IFNγ. PGE₂ slightly induced NO production compared to the control group. But, PGE2 pretreatment did not affect in NO production in the presence of TNFα and IFNγ. Conclusions : These results suggest that the actions of osteotropic hormones in bone metabolism may be partiallymediated by NO in the presence of cytokines.

Interleukin-1β와 tumor necrosis factor-α가 마우스 조골세포의 nitric oxide 생성에 미치는 영향

채종성,고선일,김정근 단국대학교 치의학연구소 1996 논문집 Vol.8 No.1

Bone remodeling is characterized by the coupling of osteoclast-mediated bone resorption and osteoblast-mediated bone formation. The process is tightly regulated at the local level by an incompletely known complex network of humoral factors produced in the bone microenvironment, including peptide as well as non-peptide molecules. Nitric oxide(NO) is recently identified messenger molecule regulating a wide range of functions throughout the body but little is known about its possible role in skeletal metabolism. Many other cells, including macrophages, hepatocytes, chondrocytes and bone marrow cells, also produce NO. Recently, NO was reported to inhibit osteoclastic bone resorption in vitro, and there are indications of an arginine-dependent NO pathway in osteoblasts and osteoclasts. In this study we present evidence that mouse osteoblasts can be induced to produce NO by cytokines (interleukin-1β (IL-1β), tumor necrosis factor-α (TNT-α) known to modulate bone cell activity. The observed results were as follows. 1. In case of 24 hr incubation with IL-1β, NO production of MC3T3/E1 cells was not stimulated. But significant amounts of NO were measured after 48 hr incubation. In case of TNT-α, significant amounts of NO were measured after 24 hr, and increased further during the next 48 hr followed by a much slower increase up to 96 hrs. 2. IL-1β stimulated NO production into cultured medium in cultures of MC3T3/E1 cells dose dependently (0.4, 0.2, 1 ng/ml). 3. TNT-α stimulated NO production into cultured medium in cultures of MC3T3/E1 cells dose dependently (0.2, 1, 5 ng/ml). 4. Cytokine combination increased NO production dose dependently. And synergistic effects of the two cytokines were observed at all dose combinations.

수종의 cytokine이 배양중인 조골세포에 미치는 영향

李鐘烈,高鮮一,金正根 단국대학교 치의학연구소 1995 단국대치의학연구소논문집 Vol.7 No.1

수종의 cytokine이 배양중인 조골세포에 미치는 영향

李鍾烈,高鮮一,金正根 단국대학교 치의학연구소 1995 논문집 Vol.7 No.1

Evidence indicates that cytokines are involved in the initiation and progression of chronic inflammatory diseases. Recent evidence demonstrates major roles of growth factors and cytokines as mediators for bone growth and remodeling. Bone remodeling is a local phenomenon which occurs in discrete packets throughout the skeleton. The cellular events which comprise the remodeling sequence are controlled by cytokines which are generated in the microenvironment of the bone resorbing area. These cytokines are derived from marrow mononuclear cells or from bone cells themselves, or they are incorporated into the bone matrix and released in biologically active form as bone resorb. But evidence is accumulating that some of these cytokines play an important role not just in physiological bone remodeling, but also in common diseases of bone remodeling such as osteoporosis, osteopetrosis, Paget's diseases and malignant diseases which involve bone and chronic inflammatory diseases such as rheumatoid arthritis and periodontal diseases. Also, these cytokines are derived from bone cells themselves, these receptors are present in bone cells. Therefore, this experiment developed a culture of osteoblastic cells from embryonic chicken calvaria to examine the possible role of some cytokine in osteoblast function. This experiment was performed 1) to examine the effect of cytokines on the acid phosphatase activity of chicken osteoblast and 2) to examine the effect of cytokines on the alkaline phosphatase activity of chicken osteoblast and 3) to study the effect of cytokines on the bone nodule formation of chicken osteoblast in long-term cultures. The observed results were as follows. 1. Cytokines and cytokine combination stimulate the release of acid phosphatase into cultured medium in cultures of chicken osteoblast. 2. Cytokines and cytokine combination increase the activity of acid phosphatase of cell extract in cultures of chicken osteoblast. 3. Cytokines and cytokine combination decrease the activity of alkaline phosphatase of cell extract in cultures of chicken osteoblast. 4. Less bone nodules were formed by cytokines than in control group.

The effect of combined Rehmannia glutinosa Libosch and Eleutherococcus senticosus Max(OPB) extracts on bone mineral density in ovariectomized rats.

Kim,Jung-Keun 대한구강생물학회 2007 International Journal of Oral Biology Vol.32 No.4

This study was conducted to investigate the preventingeffects of OPB (Rehmannia glutinosa Libosch and Eleuth-erococcus senticosus Max extracts) and combined OPB/Calcium therapy on bone loss in ovariectomized rats. SixtySprague Dawley rats of 12-week-old were divided into eightgroups: OVX (ovariectomized), OPBL (OPB 50mg/kg),OPBM (OPB100mg/kg), OPBH (OPB 200mg/kg), OPBL/CAL(OPBL+CAL), OPBM/CAL (OPBM+CAL), OPBH/CAL (OPBH+CAL) and CAL (Calcium citrate 88.33mg/kg+1α, 25-dihydroxy-vitamin D3 33.33IU/kg). Bone mineraldensity (BMD), bone mineral content (BMC), bone strengthindices and cortical thickness were analyzed by peripheralquantitative computerized tomography (pQCT). pQCTscanning showed that OVX induced a significant decreasein trabecular bone mineral density and bone mineralcontent in the proximal tibia (-36.4±2.4%, -21.8±12.7%).These decreases were significantly prevented by theadministration of OPBM and OPBM/CAL. Cortical BMDand BMC of tibia were slightly enhanced by OPB and OPB/CAL. However there was no significant difference betweenOVX and OPB, OPB/CAL treated group. Bone strengthindices and cortical thickness were not significantly different.Our results suggest that OPB and combined OPB/Calciumtherapy are effective in preventing the development of boneloss induced by ovariectomy in rats.

The Molecular Mechanism of Baicalin on RANKL-induced Osteoclastogenesis in RAW264.7 Cells

Seon-Yle Ko The Korean Academy of Oral Biology 2013 International Journal of Oral Biology Vol.38 No.2

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, antiinflammatory 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 IĸB 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.

Inhibitory Effect of Myricetin on Matrix Metalloproteinase Expression and Activity in Periodontal Inflammation

Seon-Yle Ko 대한구강생물학회 2016 International Journal of Oral Biology Vol.41 No.4

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.

Effects of plant-derived natural products on inflammatory bone destructive disease

Seon-Yle Ko 대한구강생물학회 2019 International Journal of Oral Biology Vol.44 No.4

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.

Effects of Leptin on Osteoclast Generation and Activity

Ko, Seon-Yle,Cho, Sang-Rae,Kim, Se-Won,Kim, Jung-Keun Korean Academy of Oral Biology and the UCLA Dental 2005 International Journal of Oral Biology Vol.30 No.2

Leptin, the product of the obese gene, is a circulating hormone secreted primarily from adipocytes. Several results suggest that leptin is important mediators of bone metabolism. The present study was undertaken to determine the effects of leptin on anti-osteoclastogenesis using murine precursors cultured on Ca-P coated plates and on the production of osteoprotegerin (OPG) in osteoblastic cells. Additionally, this study examined the possible involvement of prostaglandin E₂ (PGE₂)/protein kinase C (PKC)-mediated signals on the effect of leptin on anti-osteoclastogenesis to various culture systems of osteoclast precursors. Osteoclast generation was determined by counting tartrate-resistant acid phosphatase positive [TRAP (+)] multinucleated cells (MNCs). Osteoclastic activity was determined by measuring area of resorption pits formed by osteoclasts on Ca-P coated plate. The number of 1,25-dihydroxycholecalciferol (1,25[OH]₂D₃)- or PGE₂- induced TRAP (+) MNCs in the mouse bone marrow cell culture decreased significantly after treatment with leptin. The number of receptor activator of NF-kB ligand (RANKL)-induced TRAP (+) MNCs in MCSF dependent bone marrow macrophage (MDBM) cell or RAW264.7 cell culture decreased significantly with leptin treatment. Indomethacin inhibited osteoclast generation induced by 1,25[OH]₂D₃ and dexamethasone, however, no significant differences were found in the leptin treated group when compared to the corresponding indomethacin group. Phorbol 12-myristate 13-acetate (PMA), a PKC activator, inhibited osteoclast generation induced by 1,25[OH]₂D₃. The number of TRAP (+) MNCs decreased significantly with treatment by PMA at concentrations of 0.01 and 0.1 μM in culture. Leptin inhibited PMA-mediated osteoclast generation. Isoquinoline-5-sulfonic 2-methyl-l piperazide dihydrochloride (117) had no effect on osteoclast generation induced by 1,25[OH]₂D₃. Cell culture treatment with leptin resulted in no significant differences in osteoclast generation compared to the corresponding 117 group. Indomethacin showed no significant effect on TRAP (+) MNCs formation from the RAW264.7 cell line. PMA inhibited TRAP (+) MNCs formation induced by RANKL in the RAW264.7 cell culture. H7 had no effect on osteoclast generation from the RAW264.7 cell line. There was no difference compared with the corresponding control group after treatment with leptin. 1,25[OH]₂D₃- or PGE₂-induced osteoclastic activity decreased significantly with leptin treatment at a concentration of 100 ng/ml in mouse bone marrow cell culture. Indomethacin, PMA, and H7 significantly inhibited osteoclastic activity induced by 1,25[OH]₂D₃ in mouse bone marrow cell culture. No significant differences were found between the leptin treated group and the corresponding control group. 'The secretion of OPG, a substance known to inhibit osteoclast formation, was detected from the osteoblasts. Treatment by leptin resulted in significant increases in OPG secretion by osteoblastic cells. Taken these results, leptin may be an important regulatory cytokines within the bone marrow microenvironment.

Baicalin suppresses lipopolysaccharide-induced matrix metalloproteinase expression: action via the mitogen-activated protein kinase and nuclear factor κB-related protein signaling pathway

Ko, Seon-Yle The Korean Academy of Oral Biology 2021 International Journal of Oral Biology Vol.46 No.1

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 disease's 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 enzymes' 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.