• 대한치과보철학회지
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• 제44권5호
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• pp.574-583
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• 2006

Statement of problem : The use of permanent magnetics is increasing in implant dentistry. Purpose : This study is to know the effect of permanent magnetics on bone matrix formation of osteoblasts. Materials and methods : The konus abutment-shaped permanent magnetics were connected to the implant fixture, and placed on the culture plate. The osteoblast-like cell Mc3T3E1 were used for cell culture. As the control group, the implants were connected to titanium healing caps, and cultured in the same conditions of experimental group. After 3. 7, 14 days, cells were cultured, and we measured and compared the amount of collagen type I, osteocalcin, which is bone matrix protein by Western immunoblotting analysis. Results: As a result of Western immunoblotting analysis for estimating the amount of bone extracellular matrix, there was no difference between osteoblast of the experimental group and the control group during 3 and 7day-osteoblast culturing. However when cells were cultured for 14days, the amount of bone extracellular matrix was increased, on the experimental group. Conclusion: From these results, magnetic field of permanent magnetics might have effect on bone formation of osteoblast, especially at initial stage of implant placement. Therefore, their clinical application for implant or bone graft could be possible.

• International Journal of Oral Biology
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• 제31권2호
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• pp.53-65
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• 2006

Calcium concentration in the bone resorption lacunae is high and is in the mM concentration range. Both osteoblast and osteoclast have calcium sensing receptor in the cell surface, suggesting the regulatory role of high extracellular calcium in bone metabolism. In vitro, high extracellular calcium stimulated osteoclastogenesis in coculture of mouse osteoblasts and bone marrow cells. Therefore we examined the genes that were commonly regulated by both high extracellular calcium and $1,25(OH)_2vitaminD_3(VD3)$ by using mouse oligo 11 K gene chip. In the presence of 10 mM $[Ca^{2+}]e$ or 10 nM VD3, mouse calvarial osteoblasts and bone marrow cells were co-cultured for 4 days when tartrate resistant acid phosphatase-positive multinucleated cells start to appear. Of 11,000 genes examined, the genes commonly regulated both by high extracellular calcium and by VD3 were as follows; 1) the expression of genes which were osteoclast differentiation markers or were associated with osteoclastogenesis were up-regulated both by high extracellular calcium and by VD3; trap, mmp9, car2, ctsk, ckb, atp6b2, tm7sf4, rab7, 2) several chemokine and chemokine receptor genes such as sdf1, scya2, scyb5, scya6, scya8, scya9, and ccr1 were up-regulated both by high extracellular calcium and by VD3, 3) the genes such as mmp1b, mmp3 and c3 which possibly stimulate bone resorption by osteoclast, were commonly up-regulated, 4) the gene such as c1q and msr2 which were related with macrophage function, were commonly down-regulated, 5) the genes which possibly stimulate osteoblast differentiation and/or mineralization of extracellular matrix, were commonly down-regulated; slc8a1, admr, plod2, lox, fosb, 6) the genes which possibly suppress osteoblast differentiation and/or mineralization of extracellular matrix, were commonly up-regulated; s100a4, npr3, mme, 7) the genes such as calponin 1 and tgfbi which possibly suppress osteoblast differentiation and/or mineralization of extracellular matrix, were up-regulated by high extracellular calcium but were down-regulated by VD3. These results suggest that in coculture condition, both high extracellular calcium and VD3 commonly induce osteoclastogenesis but suppress osteoblast differentiation/mineralization by regulating the expression of related genes.

• 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.

• Preventive Nutrition and Food Science
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• 제16권4호
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• pp.291-298
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• 2011

Yam extracts (Dioscorea batatas) have been reported to possess a variety of functions. However, studies on its osteogenic properties are limited. In this study, we investigated the effect of ethanol and water extracts on osteoblast proliferation and bone matrix protein synthesis, type I collagen and alkaline phosphatase (ALP), using osteoblastic MC3T3-E1 cell model. MC3T3-E1 cells were cultured with yam ethanol and water extracts (0~30 mg/L) within 39 days of osteoblast differentiation period. Cell proliferation was measured by MTT assay. Bone matrix proteins were assessed by the accumulation of type I collagen and ALP activity by staining the cell layers for matrix staining. Also, the secreted (media) matrix protein concentration (type I collagen) and enzyme activity (ALP) were measured colorimetrically. Yam ethanol and water extracts stimulated cell proliferation within the range of 15~30 mg/L at 15 day treatment. The accumulation of type I collagen in the extracellular matrix, as well as secreted collagen in the media, increased with increasing doses of yam ethanol (3~15 mg/L) and water (3~30 mg/L) extracts. ALP activity was not affected by yam ethanol extracts. Our results demonstrated that yam extracts stimulated osteoblast proliferation and enhanced the accumulation of the collagenous bone matrix protein type I collagen in the extracellular matrix. These results suggest that yam extracts may be a potential activator for bone formation by increasing osteoblast proliferation and increasing bone matrix protein type I collagen. Before confirming the osteogenic action of yam, further studies for clarifying how and whereby yam extracts can stimulate this ostegenesis action are required.

• Journal of Periodontal and Implant Science
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• 제47권6호
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• pp.363-371
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• 2017

Purpose: The purpose of this study was to investigate the feasibility of regenerative therapy with a collagenated bone graft and resorbable membrane in intrabony defects, and to evaluate the effects of the novel extracellular matrix (ECM)-based membrane clinically and radiologically. Methods: Periodontal tissue regeneration procedure was performed using an ECM-based resorbable membrane in combination with a collagenated bovine bone graft in intrabony defects around the teeth and implants. A novel extracellular matrix membrane (NEM) and a widely-used membrane (WEM) were randomly applied to the test group and the control group, respectively. Cone-beam computed tomography images were obtained on the day of surgery and 6 months after the procedure. Alginate impressions were taken and plaster models were made 1 week and 6 months postoperatively. Results: The quantity of bone tissue, the dimensional changes of the surgically treated intrabony defects, and the changes in width and height below the grafted bone substitutes showed no significant difference between the test and control groups at the 6-month examination. Conclusions: The use of NEM for periodontal regeneration with a collagenated bovine bone graft showed similar clinical and radiologic results to those obtained using WEM.

• The Journal of Advanced Prosthodontics
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• 제6권5호
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• pp.351-360
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• 2014

PURPOSE. The aim of present study was to identify characteristic and response of mouse bone marrow (BM) derived low-adherent bone marrow mesenchymal stem cells (BMMSCs) obtained by quantification of extracellular matrix (ECM). MATERIALS AND METHODS. Non-adherent cells acquired by ECM coated dishes were termed low-adherent BMMSCs and these cells were analyzed by in vitro and in vivo methods, including colony forming unit fibroblast (CFU-f), bromodeoxyuridine (BrdU), multi-potential differentiation, flow cytometry and transplantation into nude mouse to measure the bone formation ability of these low-adherent BMMSCs. Titanium (Ti) discs with machined and anodized surfaces were prepared. Adherent and low-adherent BMMSCs were cultured on the Ti discs for testing their proliferation. RESULTS. The amount of CFU-f cells was significantly higher when non-adherent cells were cultured on ECM coated dishes, which was made by 7 days culturing of adherent BMMSCs. Low-adherent BMMSCs had proliferation and differentiation potential as adherent BMMSCs in vitro. The mean amount bone formation of adherent and low-adherent BMMSCs was also investigated in vivo. There was higher cell proliferation appearance in adherent and low-adherent BMMSCs seeded on anodized Ti discs than machined Ti discs by time. CONCLUSION. Low-adherent BMMSCs acquired by ECM from non-adherent cell populations maintained potential characteristic similar to those of the adherent BMMSCs and therefore could be used effectively as adherent BMMSCs in clinic.

• Journal of Nutrition and Health
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• 제51권5호
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• pp.379-385
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• 2018

Purpose: Zinc (Zn) is an essential trace element for bone mineralization and osteoblast function. We examined the effects of Zn deficiency on osteoblast differentiation and mineralization in MC3T3-E1 cells. Methods: Osteoblastic MC3T3-E1 cells were cultured at concentration of 1 to $15{\mu}M$ $ZnCl_2$ (Zn- or Zn+) for 5, 15 and 25 days up to the calcification period. Extracellular matrix mineralization was detected by staining Ca and P deposits using Alizarin Red and von Kossa stain respectively, and alkaline phosphatase (ALP) activity was detected by ALP staining and colorimetric method. Results: Extracellular matrix mineralization was decreased in Zn deficiency over 5, 15, and 25 days. Similarly, staining of ALP activity as the sign of an osteoblast differentiation, was also decreased by Zn deficiency over the same period. Interestingly, the gene expression of bone-related markers (ALP, PTHR; parathyroid hormone receptor, OPN; osteopontin, OC; osteocalcin and COLI; collagen type I), and bone-specific transcription factor Runx2 were downregulated by Zn deficiency for 5 or 15 days, however, this was restored at 25 days. Conclusion: Our data suggests that Zn deficiency inhibits osteoblast differentiation by retarding bone marker gene expression and also inhibits bone mineralization by decreasing Ca/P deposition as well as ALP activity.

• 대한치과의사협회지
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• 제55권1호
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• pp.30-41
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• 2017

Inadequate keratinized mucosa around dental implants can lead to more plaque accumulation, tissue inflammation, marginal recession and attachment loss. We evaluated the effects of free gingival and extracellular matrix membrane grafts performed to increase the insufficient width of keratinized tissue around dental implants in the posterior mandible. A 47-year-old female patient presented with discomfort due to swelling of the lower right second premolar area. Due to severe destruction of alveolar bone, the tooth was extracted. After 3 months, a guided bone regeneration (GBR) procedure was performed and then a dental implant was placed 6 months later. During the second-stage implant surgery, free gingival grafting was performed to increase the width of the keratinized tissue. After 12 months, a clinical evaluation was performed. A 64-year-old female patient had a missing tooth area of bilateral lower molar region with narrow zone of keratinized gingiva and horizontal alveolar bone loss. Simultaneous implant placement and GBR were performed. Five months after the first-stage implant surgery, a gingival augmentation procedure was performed with an extracellular matrix membrane graft to improve the width of the keratinized tissue in the second-stage implant surgery. After 12 months, a clinical evaluation was performed. In these two clinical cases, 12 months of follow-up, revealed that the increased width of the keratinized tissue and the deepened oral vestibule was well maintained. A patient showed a good oral hygiene status. In conclusion, increased width of keratinized tissue around dental implants could improve oral hygiene and could have positive effects on the long-term stability and survival rate of dental implants. When planning a keratinized tissue augmentation procedure, clinicians should consider patient-reported outcomes.

• Experimental and Molecular Medicine
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• 제50권11호
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• pp.2.1-2.16
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• 2018

Supplementation of mesenchymal stem cells (MSCs) at sites of bone resorption is required for bone homeostasis because of the non-proliferation and short lifespan properties of the osteoblasts. Calcium ions ($Ca^{2+}$) are released from the bone surfaces during osteoclast-mediated bone resorption. However, how elevated extracellular $Ca^{2+}$ concentrations would alter MSCs behavior in the proximal sites of bone resorption is largely unknown. In this study, we investigated the effect of extracellular $Ca^{2+}$ on MSCs phenotype depending on $Ca^{2+}$ concentrations. We found that the elevated extracellular $Ca^{2+}$ promoted cell proliferation and matrix mineralization of MSCs. In addition, MSCs induced the expression and secretion of osteopontin (OPN), which enhanced MSCs migration under the elevated extracellular $Ca^{2+}$ conditions. We developed in vitro osteoclast-mediated bone resorption conditions using mouse calvaria bone slices and demonstrated $Ca^{2+}$ is released from bone resorption surfaces. We also showed that the MSCs phenotype, including cell proliferation and migration, changed when the cells were treated with a bone resorption-conditioned medium. These findings suggest that the dynamic changes in $Ca^{2+}$ concentrations in the microenvironments of bone remodeling surfaces modulate MSCs phenotype and thereby contribute to bone regeneration.

• Nutritional Sciences
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• 제9권4호
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• pp.233-239
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• 2006

Porous matrices of bioactive polymers such as polyglycolic acid (PGA) or polylactic acid (PLA) can be used as scaffolds in bone tissue growth during bone repair process. These polymers are highly porous and serve as a template for the growth and organization of new bone tissues. We evaluated the effect of PGA and PLA polymers on osteoblastic MC3T3-E1 cell extracellular mineralization. MC3T3-E1 cells were cultured in a time-dependent manner -1, 15, 25d as appropriate - for the period of bone formation stages in one of the five culture circumstances, such as normal osteogenic differentiation medium, PGA-plated, fetal bovine serum (FBS)-plated, PGA/FBS-coplated, and PLA-plated For the evaluation of bone formation, minerals (Ca, Mg, Mn) and alkaline phosphatase activity, a marker for osteoblast differentiation, were measured Alizarin Red staining was used for the measurement of extracellular matrix Ca deposit During the culture period, PGA-plated one was reabsorbed into the medium more easily and faster than the PLA-plated one. At day 15, at the middle stage of bone formation, cellular Ca and Mg levels showed higher tendency in PGA- or PLA-plated treatments compared to non-plated control and at day 25, at the early late stage of bone formation, all three cellular Ca, Mg or Mn levels showed higher tendency as in order of PGA-related treatments and PLA-plated treatments, compared to control even without significance. Medium Ca, Mg or Mn levels didn't show any consistent tendency. Cellular ALP activity was higher in the PGA- or PLA-plated treatments compare to normal osteogenic medium treatment PGA-plated and PGA/FBS-plated treatments showed better Ca deposits than other treatments by measurement of Alizarin Red staining, although PLA-plated treatment also showed reasonable Ca deposit. The results of the present study suggest that biodegradable material, PGA and also with less extent for PLA, can be used as a biomaterial for better extracellular matrix mineralization in osteoblastic MC3T3-E1 cells.