• International Journal of Oral Biology
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• 제38권3호
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• pp.127-134
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• 2013

Xylitol is a sugar alcohol with a variety of functions including bactericidal and anticariogenic effects. However, the cellular mechanisms underlying the role of xylitol in bone metabolism are not yet clarified. In our present study, we exploited the physiological role of xylitol on osteoclast differentiation in a co-culture system of osteoblastic and RAW 264.7 cells. Xylitol treatment of these co-cultures reduced the number of tartrate-resistant acid phosphatase (TRAP)-positive multinucleated cells induced by 10 nM $1{\alpha},25(OH)_2D_3$ in a dose-dependent manner. A cell viability test revealed no marked cellular damage by up to 100 mM of xylitol. Exposure of osteoblastic cells to xylitol decreased RANKL, but not OPG, mRNA expression in the presence of $10^{-8}M$ $1{\alpha},25(OH)_2D_3$ in a dose-dependent manner. Furthermore, bone resorption activity, assessed on bone slices in the coculture system, was found to be dramatically decreased with increasing xylitol concentrations. RANKL and OPG proteins were assayed by ELISA and the soluble RANKL (sRANKL) concentration was decreased with an increased xylitol concentration. In contrast, OPG was unaltered by any xylitol concentration in this assay. These results indicate that xylitol inhibits $1{\alpha},25(OH)_2D_3$-induced osteoclastogenesis by reducing the sRANKL/OPG expression ratio in osteoblastic cells.

• Animal cells and systems
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• 제14권4호
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• pp.283-289
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• 2010

Ssanghwa-tang (SHT) is a traditional Korean herbal medicine widely prescribed to decrease fatigue following an illness. The purpose of this study was to investigate the effects of SHT on osteoclast differentiation in vitro, and on bone loss in ovariectomized (OVX) rats in vivo. SHT significantly reduced the receptor activator for the nuclear factor ${\kappa}B$ (NF-${\kappa}B$) ligand (RANKL)-induced tartrate-resistant acid phosphatase (TRAP) activity, and multinucleated osteoclast formation in RAW264.7 cells without affecting cell viability. In addition, SHT significantly attenuated RANKL-induced mRNA expression levels of c-Src and cathepsin K. To examine the in vivo effect of SHT on OVX-induced bone loss in OVX rats, we administered SHT (0.6 g/kg BID) orally to OVX rats for 12 weeks. SHT administration significantly blocked OVX-induced decrease of femoral bone mineral density (BMD) and femoral trabeculae in OVX rats. In conclusion, these results suggest that SHT treatment effectively prevents OVX-induced bone loss, and this effect may result from its inhibitory effect on osteoclast differentiation.

• 대한본초학회지
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• 제37권5호
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• pp.1-8
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• 2022

Objectives : This study aimed to investigate the synergistic effect of combining Cynanchi Wilfordii Radix extract with Humuli Lupuli Flos extract on estrogenic and anti-osteoclastogenic activity. Methods : Estrogenic effect of a mixture of Cynanchi Wilfordii Radix extract and Humuli Lupuli Flos extract (CWHL), Cynanchi Wilfordii Radix extract, Humuli Lupuli Flos extract, caudatin (an active ingredient of Cynanchi wilfordii Radix extract) and 8-prenylnaringenin (an active ingredient of Humuli Lupuli Flos extract) were examined by proliferation E-screen assay and expression of estrogen inducible gene, pS2 via Real Time-PCR (RT-PCR) in MCF-7 estrogen responsive cells. And their estrogenic activities were investigated how to modulate Estrogen receptor 𝛽 by binding affinity assay. Inhibitory effect of CWHL, Cynanchi Wilfordii Radix extract, Humuli Lupuli Flos extract, caudatin and 8-prenylnaringenin on RANKL-induced osteoclast differentiation were tested by TRAP (Tartrate-resistant acid phosphatase) staining in osteoclastogenic RAW 264.7 cells. Results : CWHL, Humuli Lupuli Flos extract and 8-prenylnaringenin accelerated the proliferation of MCF-7 and the expression of pS2 in MCF-7. CWHL, Cynanchi Wilfordii Radix extract, Humuli Lupuli Flos extract, caudatin and 8-prenylnaringenin bind to estrogen receptor 𝛽. CWHL, Cynanchi Wilfordii Radix extract, Humuli Lupuli Flos extract, caudatin and 8-prenylnaringenin inhibited RANKL-induced osteoclastogenesis in osteoclastogenic RAW 264.7. CWHL is more effective for all markers than Cynanchi Wilfordii Radix extract or Humuli Lupuli Flos extract alone. Conclusions : CWHL may a potential therapeutic agent for menopause and osteoporosis as a natural food resource. CWHL as a natural food source has therapeutic potential in cases of menopause and osteoporosis.

• Journal of the Korean Association of Oral and Maxillofacial Surgeons
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• 제33권6호
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• pp.684-689
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• 2007

The aim of this study is to examine the change of the osteoclastic activity in the surrounding bone with or without tooth movement after corticotomy by histologic study. Eighteen male Sprague Dawley rats with an average body weight of 300 g(range 250-350 g) were used. The rats were divided into three groups of six animals. They were operated corticotomy-assisted tooth movement and killed after 1 week, 2 weeks, and 3 weeks after tooth movement. Corticotomy was done in the surrounding of the both upper first molar. A split mouth design was used by referring to the contralateral side as control. After flap suturing, the upper left first molar was moved anteriorly by closed coil spring. The force applied was 1 N. The average of tooth movement of the 1 week group was $0.24{\pm}0.09mm,\;0.20{\pm}0.26mm$ in 2 weeks group and $0.41{\pm}0.39mm$ in 3 weeks group, respectively. The difference between the 1 week and the 2 weeks groups was very small to compare with the 3 weeks group. In the treatment group, the average numbers of cells that positively reacted to TRAP were 14.5 in the 1 week group, 12.0 in the 2 weeks group, and 6.0 in the 3 weeks group. In the control group, the numbers were 8.3 in the 1 week group, 12.8 in the 2 week group, and 1.5 in the 3 week group, respectively. The amount of tooth movement of the 3 week group was about twice as large as those of the 1 week and 2 week groups. From the standpoint of histology, the average number of cells that positively reacted to TRAP was initially larger in the treatment group than in the control group, similar in both group in 2 weeks, and became less in the treatment group in 3 weeks. Additionally, in the control group, their activity of osteoclast was higher in 2 weeks than in 1 week, and decreased rapidly in 3 weeks.

• 대한치과교정학회지
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• 제25권6호
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• pp.715-722
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• 1995

Orthodontic tooth movement in response to orthodontic force results from actions of osteoclasts and osteeoblasts in the cell level. Convincing evidence has now been provided to support the view that osteoclasts are derived from mononuclear cells that originate in the bone marrow or other hematopoietic organs and they migrate to the bones via vascular routes. Nitric oxide(NO), which accounts for the biological properties of endothelium-derived relaxing factor(EDRF), is the endogenous stimulator of soluble guanylate cylase. The discovery of the formation of nitric oxide(NO) from L-arginine in mammalian tissues and its biological roles has, in the last 7 years, thrown new light onto many areas of research. Data from experiments in vitro showed that N-metyl-L-arginine(L-NMA) and L-nitro-L- arginine(L-NAME) are competitive inhibitors of nitric oxide synthase. This study suggest that the multinucleated cells in our culture have characteristics of osteoclasts and that the potential bone cell activity of nitric oxide in vitro may be mediated in part by stimulation of marrow mononuclear cells to form osteoclast-like cells. Bone marrow cells were obtaineed from tibia of 19-days old chick embryo. After sacrifice, tibia was quickly dissected and the bone were then split to expose the medullary bone. The cells were attached for 4 hours and the nonadherent cells were collected. Marrow cells weere cultured in 96-well plate in medium 199. To examine the number of TRAP-positive multinucleated cells(MNCs), $10^{-8}\;M\;Vit=D_3$ and various concentration of L-NMA and L-NAME weere added at the beginning of cultures and with each medium change. After 7 days of culture. tartrate-resistant acid phosphatase(TRAP) staining was performed for microscopic evaluation. Cells haying more than three nuclei per cell were counted as MNCs. The obsrved results were as follows;1. 1,25-dihydroxyvitamine $D_3$ stimulated the osteoclast-like multinucleated cells in cultures of chick embryo bone marrow. 2. Nitric oxide synthase inhibitors(NOSI ; N-NMA, N-NAME) stimulated the osteoclast-like cells in cultures of chick embry bone marrow. 3. 1,25-dihydroxyvitamine$D_3$ and nitric oxide synthase inhibitors did not appear to have additive effect on the generation of TRAP-positive MNCs. These results suggest that nitric oxide synthase inhibitors may stimulate the osteoclast-like multinucleated cell formation and fusion in cultures of chick bone marrow.

• IMMUNE NETWORK
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• 제21권6호
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• pp.43.1-43.14
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• 2021

Group 3 innate lymphoid cells (ILC3), which express IL-22 and IL-17A, has been introduced as one of pathologic cells in axial spondyloarthritis (axSpA). Dyslipidaemia should be managed in axSpA patients to reduce cardiovascular disease, and dyslipidaemia promotes inflammation. This study aimed to reveal the role of circulating ILC3 in axSpA and the impact of dyslipidaemia on axSpA pathogenesis. AxSpA patients with or without dyslipidaemia and healthy control were recruited. Peripheral blood samples were collected, and flow cytometry analysis of circulating ILC3 and CD4⁺ T cells was performed. The correlation between Ankylosing Spondylitis Disease Activity Score (ASDAS)-C-reactive protein (CRP) and circulating immune cells was evaluated. The effect of oxidized low-density lipoprotein cholesterol (oxLDL-C) on immune cell differentiation was confirmed. AxSpA human monocytes were cultured with with oxLDL-C, IL-22, or oxLDL-C plus IL-22 to evaluate osteoclastogenesis using tartrate-resistant acid phosphatase (TRAP) staining and real-time quantitative PCR of osteoclast-related gene expression. Total of 34 axSpA patients (13 with dyslipidaemia and 21 without) were included in the analysis. Circulating IL-22⁺ ILC3 and Th17 were significantly elevated in axSpA patients with dyslipidaemia (p=0.001 and p=0.034, respectively), and circulating IL-22⁺ ILC3 significantly correlated with ASDAS-CRP (Rho=0.4198 and p=0.0367). Stimulation with oxLDL-C significantly increased IL-22⁺ ILC3, NKp44^- ILC3, and Th17 cells, and these were reversed by CD36 blocking agent. IL-22 and oxLDL-C increased TRAP⁺ cells and osteoclast-related gene expression. This study suggested potential role of circulating IL-22⁺ ILC3 as biomarker in axSpA. Furthermore, dyslipidaemia augmented IL-22⁺ ILC3 differentiation, and oxLDL-C and IL-22 markedly increased osteoclastogenesis of axSpA.

• 한국식품영양과학회지
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• 제40권4호
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• pp.525-530
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• 2011

아시아 지역 널리 자생하고 있는 소나무는 항암, 조골세포의 콜라겐 합성 등 다양한 연구결과가 보고된 바 있으나, 항산화 활성에 따른 파골세포의 증식 및 분화에 대한 연구는 미비한 실정이다. 따라서 본 연구에서는 적송잎 추출물별 항산화 활성과 RAW 264.7 세포를 이용하여 적송잎 추출물이 파골세포의 증식과 TRAP 활성에 미치는 영향에 대해 검토하였다. 적송잎 추출물의 총 폴리페놀 함량을 측정한 결과, 열수에탄올 추출물이 140.54 mg/g으로 가장 높은 함량을 나타내었으며, 그 다음은 에탄올 추출물, 열수 추출물, 열수헥산 추출물, 헥산 추출물 순으로 나타났다. SOD 유사 활성을 검색한 결과, 열수, 에탄올 및 열수에탄올 추출물이 proanthocyanidin의 47.31%보다 높은 SOD 유사활성을 보였다. MTT assay에 의한 파골세포의 생존율을 측정한 결과, 에탄올 추출물 경우 $1{\mu}g$/mL의 농도에서 54.04%로 가장 낮은 생존율을 나타내었고 적송잎 헥산 추출물 또한 70% 이하의 생존율을 나타내어, 각 추출물 간의 생존 비율에 차이는 있으나 모든 적송잎 추출물에 있어 파골세포의 성장을 억제하는 결과가 나타났다. 적송잎 추출물의 파골세포 분화억제 효과를 알아보기 위해 TRAP staining 한 결과, 모든 추출물에서 대조군보다 낮은 TRAP 활성이 나타났다. 특히 열수 및 에탄올 추출물의 경우, $1{\mu}g$/mL의 낮은 농도에서 각각 67.8 및 66.3%로 파골세포 분화를 감소시켰으나, 헥산 추출물은 약 80%의 분화 감소율을 나타내었다. $100{\mu}g$/mL의 고농도 첨가군에서는 오히려 에탄올 추출물의 파골세포 분화 감소율이 낮게 나타나, 저농도에서의 효과가 우수한 것으로 나타났다. 따라서 항산화 활성을 가지는 적송잎 추출물이 파골세포의 증식과 분화를 억제하여 골흡수 억제에 효과를 준다는 것이 확인되었으며, 구체적인 기작 연구와 in vivo 연구가 병행된다면 노화 및 골다공증 예방과 관련된 기능성 천연소재로 개발이 가능하리라 사료된다.

• Journal of Periodontal and Implant Science
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• 제26권2호
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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.

• 동의생리병리학회지
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• 제26권2호
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• pp.160-165
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• 2012

Osteoporotic fracture became a serious social problem, which related with mortality and morbidity in old age population. Osteoclast which is responsible for bone resorption is originated from hematopoietic cell line and plays a key role osteoporotic bone loss. Cynanchum wilfordii (Asclepiadaceae) roots have been used in Korean folk medicine for the treatment of diabetes mellitus and aging progression. Also, recent studies have shown that the extract and fractions of Cynanchi Wilfordii Radix have various pharmacological actions including scavenging free radicals, enhancing immunity, reducing high serum cholesterol, and anti-tumor activity. However, the effect of extract of Cynanchi Wilfordii Radix in osteoclast differentiation had not been reported. Thus, we evaluated the effect of Cynanchi Wilfordii Radix on receptor activator of nuclear factor-${\kappa}B$ ligand (RANKL)-induced osteoclast differentiation. Through our study, we found that Cynanchi Wilfordii Radix significantly inhibited osteoclast differentiation induced by RANKL. Cynanchi Wilfordii Radix suppressed the activation of p38 pathway and $NF{\kappa}B$ in bone marrow macrophages (BMMs) treated with RANKL. Also, Cynanchi Wilfordii Radix significantly inhibited the mRNA expression of c-Fos, tartrate-resistant acid phosphatase (TRAP), osteoclast-associated receptor (OSCAR), nuclear factor of activated T cells (NFAT)c1 and cathepsin K in BMMs treated with RANKL. Particularly, Cynanchi Wilfordii Radix inhibited the protein expression of c-fos and NFATc1. Taken together, our results demonstrated that Cynanchi Wilfordii Radix may be useful treatment option of bone-related disease such as osteoporosis leads to fracture of bone and rheumatoid arthritis.

• Molecules and Cells
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• 제39권12호
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• pp.855-861
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• 2016

Ginsenosides, which are the active materials of ginseng, have biological functions that include anti-osteoporotic effects. Aqueous ginseng extract inhibits osteoclast differentiation induced by receptor activator of NF-${\kappa}B$ ligand (RANKL). Aqueous ginseng extract produces chromatography peaks characteristic of ginsenosides. Among these peaks, ginsenoside Re is a major component. However, the preventive effects of ginsenoside Re against osteoclast differentiation are not known. We studied the effect of ginsenoside Re on osteoclast differentiation, RANKL-induced tartrate-resistant acid phosphatase (TRAP) activity, and formation of multinucleated osteoclasts in vitro. Ginsenoside Re hampered osteoclast differentiation in a dose-dependent manner. In an in vivo zebrafish model, aqueous ginseng extract and ginsenoside Re had anti-osteoclastogenesis effects. These findings suggest that both aqueous ginseng extract and ginsenoside Re prevent bone resorption by inhibiting osteoclast differentiation. Ginsenoside Re could be important for promoting bone health.