• 생명과학회지
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• 제27권5호
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• pp.501-508
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• 2017

Bone morphogenetic proteins (BMPs)는 다양한 세포기능을 조절하는 중요한 사이토카인 중 하나이다. 최근 BMP와 일주기 유전자들이 연관되어 있다는 연구결과들이 보고되고 있지만 조골세포에서 일주기 유전자인 Per1의 역할은 아직 명확하지 않다. 본 연구에서는 조골세포 분화에서 Per1의 역할을 조사하였다. MC3T3-E1 세포에서 BMP2 처리에 의해 Per1 mRNA 발현과 luciferase 활성이 증가하는 것을 확인하였다. 또한 Per1 과발현 실험을 통해서 Per1 유전자가 Runx2, ALP, OC의 발현을 증가시켰으며 ascorbic acid와 ${\beta}$-glycerophosphate에 의한 ALP 염색과 석회화가 Per1 과발현에 의해 더욱 증가하는 것을 확인하였다. 이상의 결과는 일주기 리듬을 조절하는 Per1 유전자가 조골세포의 분화를 촉진하는 인자로 작용함을 시사한다.

• 치위생과학회지
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• 제20권4호
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• pp.221-229
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• 2020

Background: The purpose of this study was to investigate the anti-oral microbial activity and anti-inflammatory effects of rosmarinic acid (RA) in lipopolysaccharide (LPS)-stimulated MC3T3-E1 osteoblastic cells on a titanium (Ti) surface during osseointegration, and to confirm the possibility of using RA as a safe natural substance for the control of peri-implantitis (PI) in Ti-based dental implants. Methods: A disk diffusion test was conducted to confirm the antimicrobial activity of RA against oral microorganisms. In order to confirm the anti-inflammatory effects of RA, inflammatory conditions were induced with 100 ng/ml of LPS in MC3T3-E1 osteoblastic cells on the Ti surface treated with or without 14 ㎍/ml of RA. The production of nitric oxide (NO) and prostaglandin E2 (PGE2) in LPS-stimulated MC3T3-E1 osteoblastic cells on the Ti surface was confirmed using an NO assay kit and PGE2 enzyme-linked immunosorbent assay kit. Reverse transcription polymerase chain reaction and western blot analysis were performed to confirm the expression of interleukin (IL)-1β and tumor necrosis factor (TNF)-α in total RNA and protein. Results: RA showed weak antimicrobial effects against Streptococcus mutans and Escherichia coli, but no antimicrobial activity against the bacteria Aggregatibacter actinomycetemcomitans and the fungus Candida albicans. RA reduced the production of pro-inflammatory mediators, NO and PGE2, and proinflammatory cytokines, TNF-α and IL-1β, in LPS-stimulated MC3T3-E1 osteoblastic cells on the Ti surface at the protein and mRNA levels. Conclusion: RA not only has anti-oral microbial activity, but also anti-inflammatory effects in LPS-stimulated MC3T3-E1 osteoblasts on the Ti surface, therefore, it can be used as a safe functional substance derived from plants for the prevention and control of PI for successful Ti-based implants.

• Food Science and Biotechnology
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• 제16권5호
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• pp.807-811
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• 2007

Diabetes is marked by high glucose levels and is associated with decreased bone mass and increased fracture rates. To determine if [6]-gingerol could influence osteoblast dysfunction induced by 2-deoxy-D-ribose (dRib), osteoblastic MC3T3-E1 cells was treated with dRib and [6]-gingerol and markers of osteoblast function and oxidized protein were examined. [6]-Gingerol ($10^{-7}\;M$) significantly increased the growth of MC3T3-E1 cells in the presence of 30 mM dRib (p<0.05). [6]-Gingerol ($10^{-7}\;M$) caused a significant elevation of alkaline phosphatase (ALP) activity, collagen content, and osteocalcin secretion in the cells. We then examined the effect of [6]-gingerol on the production of osteoprotegerin and protein carbonyl in osteoblasts. Treatment with [6]-gingerol ($10^{-9}$ and $10^{-7}\;M$) increased osteoprotegerin secretion in osteoblastic cells. Moreover, [6]-gingerol ($10^{-9}$ and $10^{-7}\;M$) decreased protein carbonyl contents of osteoblastic MC3T3-E1 cells in the presence of 30 mM dRib. Taken together, these results demonstrate that [6]-gingerol inhibits dRib-induced damage and may be useful in the treatment of diabetes related bone diseases.

• 한국환경농학회지
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• 제16권2호
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• pp.149-155
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• 1997

Carbofuran에 의하여 손상된 NIH3T3 섬유모세포(mouse skin fibroblast)를 보상시킬 수 있는 물질을 개발하기 위한 기초자료를 제시하고자 carbofuran이 NIH3T3 섬유모세포에 미치는 세포독성을 검정하고, carbofuran의 독성에 대한 phenobarbital sodium(PB) 및 3-methylcholantherene(3-MC)의 보상효과를 비교${\cdot}$분석하였다. Carbofuran의 $IC_{50}$ 결정은 배양중인 NIH3T3 섬유모세포의 각 well당 1, 25, 50, $100{\mu}M$의 carbofuran을 첨가하여 48시간 배양한 후 MTT(Tetrazolium MTT), NR(Neutral red) 및 SBR(Sulforhodamine B protein)정량을 실시하여 이들에 대한 각각의 $IC_{50}$을 구하였으며, 여기에서 구한 carbofuran의 $IC_{50}$농도와 여러 농도의 PB 또는 3-MC를 배양액에 첨가하여 48시간 배양한 후 MTT, NR 및 SRB 정량을 실시하여 보상효과를 측정하고 광학현미경적 관찰을 실시하였다. Carbofuran의 세포독성 실험결과를 보면 MTT흡광도는 carbofuran의 농도증가에 따라 감소하였으며 $MTT_{50}$은 $60.7{\mu}M$이었고, NR흡광도는 $100{\mu}M$농도에서 급격히 감소하였으며, $NR_{50}$은 $82.5{\mu}M$이었고, SRB흡광도는 $50{\mu}M$농도에서 급격히 감소하였으며 $SRB_{50}$은 $87.0{\mu}M$로써 50%의 세포독성을 나타냈다. 보상효과 실험에서는 carbofuran IC50과 PB의 조합처리의 경우 MTT 정랑과 NR정량에서는 유사하게 PB $100{\mu}M$처리군에서부터 유의성있는 보상효과가 나타났으나, SRB정량에서는 보상효과가 인정되지 않았다. Carbofura $IC_{50}$과 3-MC의 조합처리의 경우 MTT정량은 3-MC $50{\mu}M$처리군에서부터, NR정량과 SRB정량의 경우는 동일하게 3-MC $100{\mu}M$처리군에서부터 유의성있는 보상효과가 나타났다. 세포의 광학현미경적 관찰소견의 경우에서도 carbofuran과 PB 또는 3-MC 조합처리 실험군 모두에서 세포가 회복되는 것을 관찰할 수 있었다. 이상의 결과에서 PB와 3-MC 모두 carbofuran의 세포독성을 감소시킬 수 있는 물질임을 알 수 있었다.

• Natural Product Sciences
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• 제11권3호
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• pp.139-144
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• 2005

Bioflavone quercetin is believed to play an important role preventing bone loss by affecting osteoclastogenesis and regulating many systemic and local factors including hormones and cytokines. This study examined how quercetin acts on tumor necrosis factor-alpha ($TNF-{\alpha}$)-mediated apoptosis in MC3T3-E1 osteoblastic cells. Apoptosis assays revealed the dose-dependent acceleration of quercetin on $TNF-{\alpha}-induced$ apoptosis in MC3T3-E1 cells, which was demonstrated by the increased number of positively stained cells in the trypan blue staining and TUNEL assay, and the migration of many cells to the $sub-G_0/G_1$ phase in flow cytometric analysis. In particular, quercetin treatment alone increased the expression of p53 and p21 proteins in the cells. Consequently, this study showed that quercetin accelerates the $TNF-{\alpha}-induced$ apoptosis in MC3T3-E1 osteoblastic cells.

• BMB Reports
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• 제44권10호
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• pp.659-664
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• 2011

As part of the search for biologically active anti-osteoporotic agents that enhance differentiation and mineralization of osteoblastic MC3T3-E1 cells, we identified the ginsenoside Rh2(S), which is an active component in ginseng. Rh2(S) stimulates osteoblastic differentiation and mineralization, as manifested by the up-regulation of differentiation markers (alkaline phosphatase and osteogenic genes) and Alizarin Red staining, respectively. Rh2(S) activates p38 mitogen-activated protein kinase (MAPK) in time- and concentration-dependent manners, and Rh2(S)-induced differentiation and mineralization of osteoblastic cells were totally inhibited in the presence of the p38 MAPK inhibitor, SB203580. In addition, pretreatment with Go6976, a protein kinase D (PKD) inhibitor, significantly reversed the Rh2(S)-induced p38 MAPK activation, indicating that PKD might be an upstream kinase for p38 MAPK in MC3T3-E1 cells. Taken together, these results suggest that Rh2(S) induces the differentiation and mineralization of MC3T3-E1 cells through activation of PKD/p38 MAPK signaling pathways, and these findings provide a molecular basis for the osteogenic effect of Rh2(S).

• Nutritional Sciences
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• 제8권4호
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• pp.242-249
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• 2005

It is well established that zinc plays an important role in bone metabolism and mineralization. The role of zinc in bone formation is well documented in animal models, but not much reported in cell models. In the present study, we evaluated zinc deficiency effects on osteoblastic cell proliferation, alkaline phosphatase activity and expression, and extracellular matrix bone nodule formation and bone-related gene expression in osteoblastic MC3T3-E1 cells. To deplete cellular zinc, chelexed-FBS and interpermeable zinc chelator TPEN were used. MC3T3-E1 cells were cultured in zinc concentration-dependent (0-15 ${\mu}M\;ZnCl_2$) and time-dependent (0-20 days) manners. MC3T3-E1 cell proliferation by MTT assay was increased as medium zinc level increased (p<0.05). Cellular Ca level and alkaline phosphatase activity were increased as medium zinc level increased (p<0.05). Alkaline phosphatase expression, a marker of commitment to the osteoblast lineage, measured by alkaline phosphatase staining was increased as medium zinc level increased. Extracellular calcium deposits measured by von Kossa staining for nodule formation also appeared higher in Zn+(15 ${\mu}M\;ZnCl_2$) than in Zn-(0 ${\mu}M\;ZnCl_2$). Bone formation marker genes, alkaline phosphatase and osteocalcin, were also expressed higher in Zn+ than in Zn-. The current work supports the beneficial effect of zinc on bone mineralization and bone-related gene expression. The results also promote further study as to the molecular mechanism of zinc deficiency for bone formation and thus facilitate to design preventive strategies for zinc-deficient bone diseases.

• International Journal of Oral Biology
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• 제34권1호
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• pp.37-42
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• 2009

The attachment and adhesion of RAW 264.7 and MC3T3-E1 cells to titanium (Ti) discs with various degrees of roughness was investigated. The attachment, adhesion, and proliferation of these cells were evaluated after 4 hr, 24 hr and 7 day incubations. Both RAW 264.7 and MC3T3-E1 cells showed a time-dependant correlation between attachment and adhesion on the surface of the titanium discs. Both types of cells tended to have higher survival rate on these discs as the surface roughness increased. The percentage of adherent inflammatory RAW 264.7 cells was greater than MC3T3-E1 cells at 24 hr, but this was reversed at 7 days in culture. The morphology of osteoblastic MC3T3-E1 cells at 24 hr, determined using a surface emission microscope (SEM), appeared flattened and spread out while inflammatory RAW 264.7 cells were predominantly spherical in shape. The adhesion of both cell types on the titanium discs was dependant on the levels of fibronectin adsorbed on the disc surface, indicating that serum constituents modulate the efficient adhesion of these cells. Our data indicate that the cellular response to the titanium surface is dependent on the types of cells, surface roughness and serum constituents.

• International Journal of Oral Biology
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• 제39권2호
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• pp.121-128
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• 2014

Sambucus sieboldiana (SS) is a member of the family Caprifoliaceae and has been recommended as a functional material because of its several bioactivities. Although numerous literatures are available on the pharmacological and biological activities, the biological activity of SS in bone regeneration process has not yet been well-defined. Therefore, in this study, the effect of SS was investigated in the proliferation and differentiation of MC3T3-E1 osteoblastic cell line. The treatment of SS did not significantly affect the cell proliferation in MC3T3-E1 cells. SS significantly accelerated the mineralization and significantly increased the expression of alkaline phosphatase (ALP) and osteocalcin (OC) mRNAs, compared to the control, in the differentiation of MC3T3-E1 cells. SS significantly accelerated the decrease of osteonectin (ON) mRNA expression as compared with the control in a time-dependent manner in the differentiation of MC3T3-E1 cells. These results suggest that the SS facilitate the osteoblast differentiation and mineralization in MC3T3-E1 osteoblastic cells. Therefore, there may be potential properties for development and clinical application of bone regeneration materials.

• 대한기계학회:학술대회논문집
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• 대한기계학회 2008년도 추계학술대회A
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• pp.1411-1414
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• 2008

It is reported that mechanical stimulation takes a role in improving cell growth in skeletal system. And various research groups have showed that developed bioreactor to stimulate cell-seeded and threedimensional scaffold. In this study, we designed a custom-made bioreactor capable of applying controlled compression to cell-seeded agarose gel. This device consisted of a circulation system and compression system. In circular system, culture chamber was sealed for prohibiting contamination and media solution was circulated by pump. In compression system, mechanical stimuli were controlled by LabVIEW software and mechanical transfer system. Cell-encapsulated agarose gels were cultured for up to 7 days. There were significant differences between the number of cells grown in dynamic cell culture and in static cell culture from 3 days to 7 days.