• Journal of Periodontal and Implant Science
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• v.24 no.2
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• pp.321-339
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• 1994

The cytokines released by osteoblasts induce bone resorption via the differentiation of osteoclast precursors. In this process, $interleukin-1{\beta}$($IL-1{\beta}$)-induced bone resorption is mediated by granulocyte macrophage-colony stimulation factor(GM-CSF), interleukin-6 (IL-6), and tumor necrosis factor ${\alpha}$($TNF-{\alpha}$) released from osteoblasts. Since these cytokines (GM-CSF, IL-6, $TNF-{\alpha}$) are produced by not only osteoblasts but also monocytes, and interleukin-10(I1-10) inhibits the secretion of these cytokines from monocytes, it may be speculated that IL 10 could modulate the production of GM-CSF, IL-6, and $TNF-{\alpha}$ by osteoblasts, then control $IL-1{\beta}-induced$ bone resorption. Therefore, the aims of the present study were to examine the effects of IL-10 on bone resorption. The sixten or seventeen-day pregnant ICR mice were injected with $^{45}Ca$ and sacrificed one day after injection. Then fetal mouse calvaria prelabeled with $^{45}Ca$ were dissected out. In order to confirm the degree of bone resorption, mouse calvaria were treated with Lipopolysaccharide(LPS), $TNF-{\alpha}$, $IL-1{\alpha}$, IL-8, $IL-1{\beta}$, and $IL-1{\alpha}$, Then, IL-10 and $interferon-{\gamma}$ ($IFN-{\gamma}$) were added to calvarial medium, in an attempt to evaluate the effect of $IL-1{\beta}-induced$ bone resorption. In addition, osteoclasts formation in bone marrow cell cultures, and the concentration of IL-6, $TNF-{\alpha}$, and GM-CSF produced from mouse calvarial cells were investigated in response to $IL-1{\beta}$ alone and simultaneously adding f $IL-1{\beta}$ and IL-10. The degree of bone resorption was expressed as the ratio of $^{45}Ca$ release(the treated/the control). The osteoclasts in bone marrow cultures were indentified by tartrate resistant acid phosphatase(TRAP) stain and the concentration of the cytokines was quantified using enzyme linked immunosorbent method. As results of these studies, bone resorption was induced by LPS(1 ng/ml ; the ratio of $^{45}Ca$ release, $1.14{\pm}0.07$). Also $IL-1{\beta}$(1 ng/ml), $IL-1{\alpha}$(1 ng/ml), and $TNF-{\alpha}$(1 ng/ml) resulted in bone resorption(the rations of $^{45}Ca$ release, $1.61{\pm}0.26$, $1.77{\pm}0.03$, $1.20{\pm}0.15$ respectively), but IL-8 did not(the ratio of $^{45}Ca$ release, $0.93{\pm}0.21$). The ratios of $^{45}Ca$ release in response to IL-10(400 ng/ml) and $IFN-{\gamma}$(100 ng/ml) were $1.24{\pm}0.12$ and $1.08{\pm}0.04$ respectively, hence these cytokines inhibited $IL-1{\beta}$(1 ng/ml)-induced bone resorption(the ratio of $^{45}Ca$ release $1.65{\pm}0.24$). While $IL-1{\beta}$(1 ng/ml) increased the number of TRAP positive multinulcleated cells in bone marrow cultures($20{\pm}11$), simultaneously adding $IL-1{\beta}$(1 ng/ml) and IL-10(400 ng/ml) decreased the number of these cells($2{\pm}2$). Nevertheless, IL-10(400 ng/ml) did not affect the IL-6, GM-CSF, and $TNF-{\alpha}$ secretion from $IL-1{\beta}$(1 ng/ml)-activated mouse calvarial cells. From the above results, it may be suggested that IL-10 inhibites $IL-1{\beta}-induced$ osteoclast differntiation and bone resorption. However, the inhibitory effect of IL-10 on the osteoclast formation seems to be mediated not by the reduction of IL-6, GM-CSF, and $TNF-{\alpha}$ production, but by other mechanisms.

• Journal of Nutrition and Health
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• v.48 no.2
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• pp.157-166
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• 2015

Purpose: The effects of water extract and distillate from the mixture of black goat meat and medicinal herb on MG-63 osteoblast proliferation and mouse bone marrow derived osteoclast formation were investigated. Methods: Proximate composition, volatile basic nitrogen (VBN), mineral content, free amino acid composition and free fatty acid composition in black goat meat were determined. Water extract and distillate were prepared with three groups; goat meat only (BG-E, BG-D), six herbs added group (BG-E6, BG-D6), and eight herbs added group (BG-E8, BG-D8). Osteoblast proliferation, mineralization and calcium uptake activity of MG-63 cells were measured and tartrate resistant acid phosphatase activity of osteoclasts was analyzed. Results: Black goat meat had remarkably low fat and high level of calcium. Glutamic acid was the most abundant amino acid. Herbs added extract groups (BG-E6 and BG-E8) showed increased MG-63 cell proliferation in a concentration dependent manner, while all the distillates did not show the effect. All extracts and distillates showed significantly increased osteoblast mineralization depending on the concentration. In particular, herb added extract, BG-E6, increased 170.3% of control and the distillate of BG-D and BG-D6 increased up to 168.5% and 159.8%, respectively. Calcium uptake activities of all water extracts showed remarkable increase of BG-E6 and BG-E8 up to 615.5% and 628.1% of control, respectively. Ditillates had no effect except BG-D6. All water extracts significantly reduced the activity of tartrate-resistant acid phosphatase (TRAP) in osteoclasts derived from mouse bone marrow. Conclusion: Combination of black goat meat and medicinal herb increased the MG-63 cell proliferation and effectively inhibited osteoclast differentiation in both water extracts and distillate of them, which implies that they could be used as potent functional food materials for bone health.

• Journal of the Korean Society of Food Science and Nutrition
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• v.37 no.8
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• pp.992-997
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• 2008

In this study, we investigated the indirect effect of silk-fibroin on osteoclastic differentiation of RAW264.7 cells. The conditioned medium were collected from MC3T3-E1 osbeoblasts treated with $0.001\;mg/mL{\sim}0.1\;mg/mL$ silk fibroin for 6 days, mixed in 1:1 ratio with osteoclast medium, and then added into RAW264.7 cells with receptor activator of nuclear factor kappa B ligand (RANKL), a differentiation inducer for 3 days. Of osteoclastic cytokines in the conditioned medium, the protein expression of osteoprotegerin (OPG) with silk-fibroin was not significantly different. However, the protein expression of interleukin (IL)-$1{\beta}$ was specifically lower in a dose dependent manner. In RAW264.7 cells, the conditioned medium with silk-fibroin inhibited RANKL induced osteoclastic differentiation as total number of multinucleated tartrate-resistant alkaline phosphatase (TRAP)-positive osteoclasts in a dose dependent manner. Taken together, we demonstrated that the conditioned medium of silk-fibroin treated osteoblasts inhibits RANKL induced differentiation of osteoclasts with inhibiting selective expression of IL-$1{\beta}$.

• The korean journal of orthodontics
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• v.28 no.2 s.67
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• pp.297-310
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• 1998

The purpose of this study was to investigate the initial tissue change, to repair on the teeth & surrounding tissue under the intrusive orthodontic forces by use of elastic chain, through the microscopic findings. For this study, three young adult mongrel dogs were used, and were divied into three group : the control group was deliveried only casting crown and the experimental group 1 was equipped with energy chain during 1 week and experimental 2 group was deliveried using energy chain during 1 week and 3 weeks observation. All experimental groups and control groups were sacrificed to make the samples for microscopic findings on premolar teeth. All samples were examed and compared the histologic changes through the microscopic with H-E stain. The obtained results were as follows. 1. In hematoxylin-eosin stain of the control group, the periodontal ligament was constant width from apical third to cervical third of the root, and the periodontal fiber arrangement was horizontal or oblique in cervical third, oblique in middle and apical third of the root. 2. In Masson Trichrome stain of the control group, osteoblast and osteoclast appeared in cervical third of root, and bone resorption and new bone formation was observed in middle and apical third of the root. 3. In experimental 1, osteoclasts were increased highly, and hyperemia of blood vessels and new bone formation and bone resorption by reversal line in apical third of the root were seen. PDL width was increased apprarently from crest to apex of the root and more in apical third. 4. In experimental 2, osteoclasts and hyperemia of blood vessels were more increased than control material in apical third of the root. PDL width was increased more than control group in root apex, and was seen less than experimental 1. PDL arrangement was similar to experimental 1 and was mixed only in root apex. Therefore, in premolar intrusion of the young adult dog, there were increased osteoclast, hyperemia and dilation of blood vessel, resorption of alveolar bone and cementum and different arrangement of PDL in initial tissue change. There was not observed complete repair after remove intrusive force.

• Journal of Life Science
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• v.30 no.11
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• pp.983-989
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• 2020

The balance between bone-resorbing osteoclasts and bone-forming osteoblasts is key to bone health. An imbalance between osteoclasts and osteoblasts leads to various bone-related disorders, such as osteoporosis, osteomalacia, and osteopetrosis. However, the bone-resorption inhibitor drugs that are currently used may cause side effects. Natural substances have recently received much attention as therapeutic drugs for the treatment of bone health. This study was designed to determine the effect of Tenebrio molitor larvae ethanol extract (TME) on receptor activator of nuclear factor-κB ligand (RANKL)-induced osteoclast differentiation. To measure the effect of TME on osteoclast differentiation, RAW264.7 cells were treated with RANKL with or without TME for 5 days. The tartrate-resistant acid phosphatase (TRAP) activity was significantly inhibited by treatment of TME without cytotoxicity up to 2 mg/ml. In addition, TME effectively suppressed expression of osteoclast differentiation-related marker genes and proteins such as TRAP, NFATc1, and c-Src. TME also significantly inhibited the p38 mitogen-activated protein kinase (MAPK) signaling pathway without affecting ERK and JNK signaling in RANKL-induced RAW264.7 cells. Consequently, we conclude that TME suppresses osteoclast differentiation by inhibiting RANKL-induced osteoclastogenic genes expression through the p38 MAPK signaling pathways. These results suggest that TME and its bioactive components are potential therapeutics for bone-related diseases such as osteoporosis.

• The korean journal of orthodontics
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• v.27 no.4 s.63
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• pp.585-597
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• 1997

The purpose of this study was to evaluate the effect of estrogen on the periodontium and alveolar bone tissue response during experimental tooth movement in ovariectomized rats. Eighty female rats, 250gm in body weight, were classified into four groups ; sham operated group(NN), ovariectomized group(ON), ovariectomized & estrogen injected group(OE), sham operated & estrogen injected group(NE). flats were ovariectomized before 3 weeks to begin the experiment, which resulted in estrogen-deficient osteoporosis. In OE group & NE group, estrogen was injected $50{\mu}g/kg\;B.W.$ every other days. The left maxillary 1st molar was moved mesially with 60g force. Each foot rats were sacrificed after 1, 3, 7, 15 days from application of orthodontic appliance and alter additional 7 days from removal of orthodontic appliance. Histological findings on mesial roots of upper 1st molar in pressure and tension side are observed. The results were summarized as follows ; 1. In pressure side of alveolar bone, the number of osteoclasts and Howship's lacuna of ON group was significantly more than that of NN group from 1 day to 15 days(P<0.05). Especially the number of Howship's lacuna of ON group was significantly more than that of OE group during all experimental period(P<0.05). 2. In tension side of alveolar tune, the number of osteoclasts of ON group was significantly increased from 1 day to 3 days and decreased after 7 days. But the number of osteoclast of ON group was significantly mote than that of NN group during all experimental period(P<0.05). Also the number of Howship's lacuna of all groups was abruptly increased at 1 day, but slowly decreased till experimental 15 days. And the number of Howship's lacuna of of group was significantly more than that of NN group from 0 hr to 7 days(P<0.05). 3. The speed of tooth movement of OE group & NE group was similar to that of NN group(P>0.05). The amount of tooth movement of ON group between 7 days and 15 days was significantly greater than those of other groups(P<0.05). 4. The degree of relapse of ON group after 7 days from removal of orthodontic appliance was similar to those of other groups(P>0.05).

• Journal of Periodontal and Implant Science
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• v.26 no.2
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• pp.448-468
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• 1996

To maintain its functional integrity, bone is continuously remodelled by a process involving resorption by osteoeclasts and formation by osteoblasts, In order to respond to changes in the physical environment or to trauma with the relevant action, this process is strictly regulated by locally synthesized or systemic fators, Prostaglandin $E_2(PGE_2$) is perhaps one of the best studied factors, having been known to affect bone cell function for several decades.$PGE_2$ has both anabolic and catabolic activities. Excess of $PGE_2$ has been implicated in a number of pathological states associated with bone loss in a number of chronic inflammatory conditions such as periodontal disease and rheumatoid arthritis. $PGE_2$ and other arachidonic acid metabolites have been shown to be potent stimulators of osteoclastic bone resorption in organ culture. The anabolic effects of $PGE_2$ were first noticed when an increase in periosteal woven bone formation was seen after the infusion of $PGE_2$ into infants in order to prevent closure of the ductus arteriosus. The cellular basis for the catabolic actions of $PGE_2$ has been well characterized. $PGE_2$increases osteoclast recruitment in bone marrow cell cultures. Also $PGE_2$ has a direct action on osteoclast serving to inhibit activity and can also indirectly activate osteoclast via other cells in the vicinity, presumably osteoblast. The cellular mechanisms for the anabolic actions of $PGE_2$ are not nearly so well understood. The purpose of this paper was to study the effects of $PGE_2$ and dibutyl(DB)cAMP on osteoblastic clone MC3T3El cells and on the generation of osteoclasts from their precursor cells. The effect of $PGE_2$ and DBcAMP on the induction of alkaline phoaphatase(AlP) was investigated in osteoblastic clone MC3T3El cells cultured in medium containing 0.4% fetal bovine serum. $PGE_2$ and DBcAMP stimulated ALP activity and MTT assay in the cells in a dose-dependent manner at concentrations of lO-SOOng/ml. Cycloheximide, protein synthesis inhibitor, inhibited the stimulative effect of $PGE_2$ and DBcAMP on ALP activity in the cells. $PGE_2$also increased the intracellular cAMP content in a dose-dependent fashion with a maximal effect at 500ng/ml. The effect of $PGE_2$ on the generation of osteoclasts was investigated in a coculture system of mouse bone marrow cells with primary osteoblastic cells cultured in media containing 10% fetal bovine serum.After cultures, staining for tartrate-resistant acid phosphatase(TRAP)-marker enzyme of osteoclast was performed. The TRAP(+) multinucleated cells(MNCs), which have 3 or more nuclei, were counted. More TRAP(+) MNCs were formed in coculture system than in control group. $PGE_2(10^{-5}10^{-6}M)$ stimulated the formation of osteoclast cells from mouse bone marrow cells in culture. $PGE_2(10^{-6}M)$ stimulated the formation of osteoclast cells from mouse bone marrow cells in coculture of osteoblastic clone MC3T3E1 cells This results suggest that $PGE_2$ stimulates the differentiation of osteoblasts and generation of osteoclast, and are involved in bone formation, as well as in bone resorption.

• Journal of the korean academy of Pediatric Dentistry
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• v.33 no.2
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• pp.290-303
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• 2006

Tooth eruption is a complex and tightly regulated process that involves cells of the tooth organ and the surrounding alveolus. Osteoclast precursors must be recruited into the dental follicle prior to the onset of eruption. This function of dental follicle may be regarded as the ability of bone remodeling characterized by the interaction of osteoclasts and osteoblasts. This is because tooth eruption is a localized event in which many of the genes required for eruption are expressed in the dental follicle. RANKL is a membrane-bound protein that is a member of the TNF ligand family. which is present on bone marrow stromal cells and osteoblasts, and induces osteoclast formation and activation from precursor cell. The biologic effect of RANKL is inhibited by OPG and, in bone, the relative ratio of RANKL and OPG modulates osteoclastogenesis. To evaluate the roles of RANKL and OPG in tooth eruption and the relations with the expression pattern of Runx2, in situ hybridization was performed with mandibles of mice at postnatal stage 1, 3, 5, 7, 9 and 11. mRNA of RANKL, OPG, and Runx2 are expressed in dental follicle and surrounding tissue from P1 to 11. To determine the sites of osteoclastic activity during tooth eruption, mandibles were dissected. Peak osteoclastic activity in alveolar bone along the occlusal and basal regions was observed from P5 to 9, with osteoclasts in these regions being large and strongly TRAP-positive The specific spatio-temporal expression patterns of RANKL, OPG, and Runx2 in our study suggest that tooth eruption could be progressed through the interactions of molecular signaling among dental follicle, dental organ and alveolar bone, furthermore it means that dental follicle is quite important in tooth eruption In addition, it indicates that these genes (RANKL, OPG, and Runx2) play critical roles in tooth eruption.

• Journal of Korean Academy of Oral and Maxillofacial Radiology
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• v.22 no.2
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• pp.241-252
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• 1992

The purpose of this study was to investigate the changes of morphology and structure of bone tissue in the irradiated mandibular bone in rats which were fed a low calcium diet. In order to carry out this experimant, 64 seven-week old Sprague-Dawley strain rats weighing about 150gms were selected and equally divided into one experimental group of 32 rats and one control group with the remainder. The experimental group and the control group were then subdivided into two groups when the rats reached ten-week old, 16 were assigned rats for each subdivided group, exposed to irradiation. The two irradiation groups received a single dose of 20 Gy in the jaws area only and irradiated with a cobalt-60 teletherapy unit. The rats in the control and experimental groups were serially terminated by fours on the 3rd, the 7th, the 14th, and the 21st day after irradiation. Following termination, both sides of the dead rats mandibular bodies were removed and fixed with 10% neutral formalin. One side of the mandibular body was radiographed with a soft X-ray apparatus. Thereafter, the obtained microradiographs were observed by a light microscope. The remaining side of the mandibular bone was further decalcified and embedded in paraffin as using the general method. The specimen sectioned and stained with hematoxylin and eosin, and Rabit Anti-Human Tumor Necrosis Factor-a, observed by a light microscope. The obtained results were as follows: 1. Microradiogram revealed that thinning of the cortex and a decrease in the trabecula of the interradicular bone and mandibular body were observed and noted from the start to finish throughout the experiment in the non-irradiated rats on the low calcium diet rather than in the non-irradiated rats on the normal diet. In microscopic observations, there were marked osteolytic changes in the center of the bone marrow. 2. Microradiogram revealed that thinning of the cortex and a decrease in the trabecula of interradicular bone and mandibular body were more marked after 7 days in the irradiated rats on the low calcium diet rather than in the non-irradiated rats on the low clacium diet. In microscopic observations, osteoblasts were decreased markedly after 7 days, and a few osteoclasts were observed. 3. Microradiogram revealed that thinning of the cortex and a decrease in trabecula of interradicular bone and the mandibular body were more marked from the strart to finish troughout the experiment in the irradiated rats on the low calcium diet rather than in the irradiated rats on the normal diet. In microscopic observations, there were no osteolytic changes noted in the irradiated rats on the normal diet. 4. In immunocytochemical findings, a few bone marrow cells including Tumor Necrosis Factor were observed in the irradiated rats on the normal diet, but was not observed in the rats on the low calcium diet.

• International Journal of Oral Biology
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• v.33 no.1
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• pp.33-43
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• 2008

In the process of bone remodeling, mineral phase of bone is dissolved by osteoclasts, resulting in elevation of calcium concentration in micro-environment. This study was performed to explore the effect of high extracellular calcium ($Ca{^{2+}}_e$) on mineralized nodule formation and on the expression of progressive ankylosis (Ank), plasma cell membrane glycoprotein-1 (PC-1) and osteopontin by primary cultured mouse calvarial cells. Osteoblastic differentiation and mineralized nodule formation was induced by culture of mouse calvarial cells in osteoblast differentiation medium containing ascorbic acid and ${\beta}$-glycerophosphate. Although Ank, PC-1 and osteopontin are well known inhibitors of mineralization, expression of these genes were induced at the later stage of osteoblast differentiation during when expression of osteocalcin, a late marker gene of osteoblast differentiation, was induced and mineralization was actively progressing. High $Ca{^{2+}}_e$(10 mM) treatment highly enhanced mRNA expression of Ank, PC-1 and osteopontin in the late stage of osteoblast differentiation but not in the early stage. Inhibition of p44/42 MAPK activation but not that of protein kinase C suppressed high $Ca{^{2+}}_{e^-}$induced expression of Ank, PC-1 and osteopontin. When high $Ca{^{2+}}_e$(5 mM or 10 mM) was present in culture medium during when mineral deposition was actively progressing, matrix calcifiation was significantly increased by high $Ca{^{2+}}_e$. This stimulatory effect was abolished by pyrophosphate (5 mM) or levamisole (0.1-0.5 mM), an alkaline phosphatase inhibitor. In addition, probenecid (2mM), an inhibitor of Ank, suppressed matrix calcification in both control and high $Ca{^{2+}}_{e^-}$treated group, suggesting the possible role of Ank in matrix calcification by osteoblasts. Taken together, these results showed that high $Ca{^{2+}}_e$ stimulates expression of Ank, PC-1 and osteopontin as well as matrix calcification in late differentiation stage of osteoblasts and that p44/42 MAPK activation is involved in high $Ca{^{2+}}_{e^-}$induced expression of Ank, PC-1 and osteopontin.