• Journal of Periodontal and Implant Science
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• v.30 no.2
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• pp.347-360
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• 2000

Bone remodeling results from the combined process of bone resorption and new bone formation which is regulated in part by some of the polypeptide growth factors such as platelet derived growth factor(PDGF), which has been known to be an important local regulator of bone cell activity and participate in normal bone remodeling. This process includes strictly regulated gene expression of several bone matrix proteins such as type I collagen and osteopontin, a 44 kDa phosphorylated glycoprotein, which has important roles in bone formation. The purpose of this study is to evaluate the effecs of PDGF-BB on the mRNA expression of bone matrix protein, type I collagen and osteopontin, in MC3T3- E1 cell culture. Cells were seeded at $5{\times}10^5$ cells in 10 ml of minimum essential medium alpha(${\alpha}-MEM$) containig 10% fetal bovine serum, 10 mM beta glycerophosphate. 0.1, 1, 10 ng/ml PDGF-BB were added to the cells for the day 3, 7, 14, 21, 28 and cultured for 24 hours. Type I collagen cDNA, Hf677, and osteopontin cDNA were used as probes for northern blot analysis. Total cellular RNA was purified at indicated day and northern blot analysis was performed. The results were as follows : Type I collagen mRNA expressions were higher at the day 3 and 7, and lower in the day 14, 21 in the control groups. In the experimental groups, mRNA expressions were increased when 0.1 ng/ml PDGF-BB were added on the day 3, 7, 21, and decreased in dose-dependent manner on the day 14, decreased at all added dose on the day 28. Osteopontin mRNA expressions were highest in the day 21 groups and lowest in the day 14 groups in the control groups. Interesting results were shown in the day 14 and 21 groups. We found that osteopontin mRNA level was increased in dose dependent manner in the day 14 groups, and decreased dose dependent manner in the day 21 groups. In conclusion, PDGF-BB may have various control effects on type I mRNA expression in the growth and differentiation process of MC3T3-E1 cells and may have contrary regulatory effects on osteopontin mRNA expression. For examples, when the baseline level of osteopontin mRNA was low, as in the day 14, PDGF-BB up-regulated osteopontin mRNA expression in dose dependent manner, and when the baseline level was high as in the day 21, PDGF-BB down-regulated dose dependent manner. Thus, it may be useful for clinical application in periodontal regeneration procedure if further study were performed.

• The Journal of Internal Korean Medicine
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• v.36 no.4
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• pp.518-526
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• 2015

Objectives This study investigated the effect of purified safflower (Carthamus tinctorius Linne) and safflower seed (Carthamus tinctorius L. seed; CS) extract, using hot water and ethanol extract methods , on the osteogenic differentiation of MC3T3E1 cells.Methods The safflower and safflower seed were extracted with hot water and ethanol. The samples were concentrated by a rotary evaporator and then freeze-dried using a freeze-dryer. The MC3T3E1 cells were propagated and maintained in DMEM (Gibco) containing 10% FBS and a 1% antibiotic antimycotic solution. To induce osteogenic differentiation, the cells were treated for 14 days with DMEM with 10 mM β-glycerophosphate and 50 μM ascorbic acid. Extract doses were confirmed by the results of an MTT assay, and treatment of the extracts was performed in a differentiation medium every two days. The ALP staining and activity were tested after osteogenic differentiation for five days, and after 14 days, osteogenic differentiation was determined by alizarin red S staining. The mRNA expressions of osteogenic-related genes were quantified using quantitative real-time PCR.Results In the results of the MTT assay, all concentrations of safflower extracts had no toxicity in the MC3T3El cells. But in the groups of 100 ng/ml and 200 ng/ml concentrations of safflower seed extracts, the cell viability was significantly reduced by up to 40-50%. So we fixed the treatment concentration of the extract at 50 ng/ml. In the ALP and alizarin red S staining, all extract groups increased osteogenic differentiation compared with the control group. The water-safflower extract group showed the highest mRNA level of Alp, Runx2, and Dlx5 genes. The mRNA level of Ocn, an osteogenic gene related to late-stage differentiation, in the ethanol-safflower extract group increased the mineralization more significantly than in other groups.Conclusions These data suggest that the extract of safflower increases the osteoblastic differentiation activates of MC3T3E1 cells like the extract of safflower seed. The water-extract and ethanol-extract of safflower have effects on different stages of osteogenesis in MC3T3El. Not only safflower seed but also safflower will be useful therapeutic reagents for age-associated chronic diseases such as osteoporosis.

• Journal of the Korean Association of Oral and Maxillofacial Surgeons
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• v.35 no.4
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• pp.205-212
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• 2009

Purpose : The purpose of this study was to examine the expression of various angiogenic factors during osteoblastic differentiation of periostealderived cells and the effects of osteogenic inductive medium of periosteal-derived cells on the proliferation of endothelial progenitor cells. Materials and methods : Periosteal-derived cells were obtained from mandibular periosteums and introduced into the cell culture. After passage 3, the cells were divided into two groups and cultured for 21 days. In one group, the cells were cultured in the DMEM supplemented with osteogenic inductive agent, including 50g/ml L-ascorbic acid 2-phosphate, 10 nM dexamethasone and 10 mM -glycerophosphate. In the other group, they were cultured in DMEM supplemented without osteogenic inductive agent. VEGF isoforms, VEGFR-1, VEGFR-2, and neuropilin-1 mRNA expression was observed. Human umbilical cord blood-derived endothelial progenitor cell proliferation was also observed. Results : The expression of VEGF isoforms was higher in osteogenic inductive medium than in non-osteogenic inductive medium. The expression of VEGFR-2 was also higher in osteogenic inductive medium than in non-osteogenic inductive medium. However, the expression of VEGFR-1 and neuropilin-1 was similar in both osteogenic inductive medium and non-osteogenic inductive medium. In addition, conditioned medium from differentiated periosteal-derived cells stimulated human umbilical cord blood-derived endothelial progenitor cell numbers compared to conditioned medium from non-differentiated periosteal-derived cells. Conclusion : These results suggest that in vitro osteoblastic differentiation of periosteal-derived cells has angiogenic capacity to support endothelial progenitor cell numbers.

• Maxillofacial Plastic and Reconstructive Surgery
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• v.29 no.6
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• pp.474-484
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• 2007

Purpose: The effect of platelet-rich plasma(PRP) on osteogenesis of marrow-derived osteoblasts on histomorphometric analysis in the mandible of rabbit was assessed. Materials and Method: Bone marrow cells were obtained from iliac bone of rabbits and were cultured in a Dulbecco's Modified Eagle's Medium(DMEM) with Dexamethasone, L-Ascortic acid, ${\beta}$-Glycerophosphate to proliferate and differentiate into osteoblasts for $4{\sim}5$ weeks. The expression of osteogenic mar-kers was detected by reverse transcription-polymerase chain reaction(RT-PCR) and silver nitrate stain. Then we prepared bony defects in the mandible of rabbit, 10.0mm in diameter and 4.0mm deep, by trephine bur. In the control group, the defects were filled with autogenous bone and cultured osteoblasts. In the experimental group, the defects were filled with autogenous bone, cultured osteoblasts and PRP. 2 weeks, 4 weeks, 8 weeks later, each group was evaluated with histological and histomorphometric analyses. Results: In vitro, osteoblasts were identified on RT-PCR and silver nitrate stain. According to histological observation, at 2 weeks well-developed anasto-mosing newly-formed woven bone was observed, at 4 weeks anastomosing newly-formed woven bone having osteoblastic activation was observed, and at 8 weeks thick newly-formed woven bone was observed in both control and experimental groups. According to histomorphometric analysis, there were 1.5% more newly-formed bone volume in experimental group than control group at 2 weeks, 28.4% more at 4 weeks, 4.3% more at 8 weeks. Particularly there were significant differences in bone volume at 4 weeks and 8 weeks new bone. Conclusion: Our results demonstrated PRP may enhance osteogenesis of marrow-derived osteoblasts at 4 weeks, 8 weeks.

• Journal of Periodontal and Implant Science
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• v.26 no.2
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• pp.491-510
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• 1996

This study was performed to evaluate the effects of platelet-derived growth factor (PDGF) and epidermal growth factor (EGF) on the characteristics of beagle dog's periodontal ligament (BPD) cells and bone marrow (BBM) cells which have the important role on the early stage of periodontal tissue regeneration in vitro. In control group, the cells ($1.5{\times}10^5$cells/ml) were cultured alone with Dulbecco's Modified Eagle's Medium contained with 10% fetal bovine serum, $50{\mu]g/ml$ ascorbic acid, and 10mM/ml ${\beta}-glycerophosphate$. In experimental groups, growth factors, PDGF or EGF(10ng/ml), were added into the above culture condition. And then each group was characterized by examining the cell proliferation rate, amount of total protein synthesis, alkaline phosphatase activity at 1, 5, 9, 13, 17th day after seeding of cells into the culture wells. The results were as follows: 1. Both BPD and BBM cells in PDGF-treated group proliferated more rapidly than non-treated cells. This finding also was observed in EGF-treated group but it was not as prominent as that of PDGF-treated group. The proliferation rates of both cells showed the time-dependent pattern during experimental periods in all three groups. 2. Amount of total protein synthesis was more increased in PDGF-treated group than in control group. But no significant difference between EGF-treated group and control group was observed throughout experimental periods even though the tendency of amount of protein synthesis was time-dependent pattern. 3. Alkaline phosphatase activity also more increased in PDGF-treated group than control group. But slight decrease tendency was seen in both cells of EGF-treated group. From the above results, PDGF appeared to enhance the proliferation and cellular activities including amount of total protein synthesis and alkaline phosphatase activity of BPD and BBM cell, but EGF did not show notable effects. The optimal application of these growth factors was thought to be useful as the adjunctive means in periodontal regeneration procedures.

• Maxillofacial Plastic and Reconstructive Surgery
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• v.29 no.5
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• pp.383-393
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• 2007

The aim of the present study is to investigate the effect of anodized surface of osseointegration implants by using of resonance frequency analysis (RFA) and histomorphometric analysis. A total of 96 screw-shaped implants were devided into 4 groups. Seventy-two implants were prepared by electrochemical oxidation with 3 different ways; Group 1 (n=24) were prepared at galvanostatic mode in 0.25M sulfuric acid and phosphoric acid, Group 2 (n=24) were prepared at galvanostatic mode in calcium glycerophosphate and calcium acetate, and Group 3 (n=24) were prepared at galvanostatic mode in 0.25M sulfuric acid and phosphoric acid followed by Calcium metaphosphate(CMP) coating. Control group (n=24) were the RBM surfaced implants. The implants were placed in the mandibles of 12 mini pigs. Bone tissue responses were evaluated by resonance frequency analysis(RFA) and histomorphometric analysis that were undertaken at 2, 4 and 6weeks after implant placement. The following result were obtained. 1. Twenty-two of 96 implants (4 in control group, 5 in group 1, 7 in group 2, and 6 in group 3) were failed due to faliure of osseoitegration. The failure rate of osseointegration was 22.9%. 2. The mean values of RFA in control, group 2 and groups 3 showed the similar values, but there was no significant difference among groups. 3. Histomorphometric evaluation demonstrated significantly higher bone-to-implant contact ratio in group 2 at 3 and 4 weeks after implant placement than other groups (p<0.05), but there was no significant difference among groups at 6weeks after implant placement.

• Journal of the Korean Association of Oral and Maxillofacial Surgeons
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• v.34 no.4
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• pp.419-427
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• 2008

The present study aimed to investigate the osteogenic potentials of differentiated osteoblast-like cells (DOCs) induced from bone marrow-derived mesenchymal stem cells (MSCs) on ${\beta}-tricalcium$ phosphate (${\beta}-TCP$) with recombinant human bone morphogenetic protein (rhBMP-2) in vitro. Osteoblast differentiation was induced in confluent cultures by adding 100 nM dexamethasone, 10 mM ${\beta}$-glycerophosphate, 50 mM L-ascorbic acid. The Alizarin red S staining and reverse transcriptase-polymerase chain reaction (RT-PCR) were perfomed to examine the mRNA expression of alkaline phosphatase (ALP), bone sialoprotein (BSP), osteocalcin (OCN), receptor activator for nuclear factor ${\kappa}B$ ligand (RANKL), runt-related transcription factor 2 (RUNX2), collagen-Ⅰ (COL-Ⅰ). There were no significant differences in the osteogenic potentials of DOCs induced from MSCs on ${\beta}-TCP(+/-)$. According to the incubation period, there were significant increasing of Alizadin red S staining in the induction 3 weeks. The mRNA expression of ALP, RUNX2, and RANKL were higher in DOCs/${\beta}-TCP(-)$ than DOCs/${\beta}-TCP(+)$. According to rhBMP-2 concentrations, the mRNA expression of BSP was significantly increased in DOCs/${\beta}-TCP(+)$ compared to that of DOCs/${\beta}-TCP(-)$ on rhBMP 10 ng/ml. Our study presented the ${\beta}-TCP$ will have the possibility that calcium phosphate directly affect the osteoblastic differentiation of the bone marrowderived MSCs.

• The Sea:JOURNAL OF THE KOREAN SOCIETY OF OCEANOGRAPHY
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• v.19 no.4
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• pp.232-242
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• 2014

We investigated the interspecific competition between the harmful dinoflagellate Cochlodinium polykrikoides and diatom Skeletonema sp. based on the utilization and uptake of dissolved organic nutrients. C. polykrikoides and S. costatum were able to grow using dissolved organic nitrogen (DON) and dissolved organic phosphorus (DOP) as well as dissolved inorganic nitrogen (DIN) and dissolved inorganic phosphorus (DIP). This result indicates that the utilization of dissolved organic nutrients may play a role in surviving strategy in the DIN or DIP-limited environments. The half-saturation constants (Ks) of urea and glycerophosphate (glycero-P) calculated from uptake kinetics experiment of C. polykrikoides was lower than those of Skeletonema sp. This result indicates that Skeletonema sp. have higher affinity for dissolved organic nutrients, such as urea and glycero-P, than C. polykrikoides. Although Skeletonema sp. have higher affinity of dissolved organic nutrients, C. polykrikoides could effectively uptake for urea and glycero-P at sub-saturating nutrient concentrations (${\alpha}$ (${\rho}_{max}/Ks$) of C. polykrikoides was higher than Skeletonema sp.. Therefore, C. polykrikoides by utilization and effectively uptake of dissolved organic nutrients under monoculture may have an advantageous position in the interspecific competition with Skeletonema sp. in the low nutrient environments.

• The Korean Journal of Oral and Maxillofacial Pathology
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• v.42 no.5
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• pp.111-118
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• 2018

Nicotine of tobacco component has a controversial impact in the clinical outcome of dental implants. Although numerous nicotine effects on bone healing around implants have been presented, it is rarely reported in vitro study about normal human osteoblast(NHost) from oral and maxillofacial area at the surface of implants. The purpose of the present study was to evaluate the effect of nicotine on the proliferation and differentiation response of NHost to plasmatic and salivary levels of nicotine reported in smokers at the surface of screw-type plasma-sprayed titanium implants. NHosts were seeded on the surface of titanium implants and cultured for 21 days in ${\alpha}-MEM$ supplemented with 10% FBS, 50mg/ml ascorbic acid, 5mM ${\beta}$-glycerophosphate and 100nM dexamethasone. Seeded implants were exposed to various nicotine concentration(0.05-0.5mg/ml) from 1 to 21 days, and characterized for cell morphology, proliferation, differentiation, alkaline phosphatase(ALP) activity and ionized calcium concentration(Cai) of medium. Continuous exposure to higher nicotine concentration(above 0.3mg/ml) induced a dose- and time-dependent vacuolation of the cytoplasm, and a tendency to detach from the implant surface. 0.05mg/ml(lower nicotine concentration) did not cause significant effects in the cell proliferation and ALP activity. 0.1-0.2mg/ml caused evident dose-dependent effects in increased cell proliferation, ALP activity and earlier onset of matrix mineralization at levels up to 0.2mg/ml, while a dose-dependent inhibitory effect at 0.3-0.5mg/ml. Cai concentration of control group was decreased at 14 days. Increased Cai concentration at 0.1-0.2mg/ml, decreased Cai concentration at 0.3mg/ml and no change at 0.5mg/ml during the culture period were seen. It suggested that nicotine concentration could paly an role in modulating NHost activity as a contributing factor associated with proliferation and differentiation of NHost at the surface of implants.

• Journal of Food Hygiene and Safety
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• v.20 no.3
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• pp.134-140
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• 2005

Boiling point and distillation range, melting range, and identification methods in general test method of Korea, Japan, Joint FAO/WHO Expert Committee of Food Additives (JECFA), and USA on chemical food additives were compared. Boiling point of propylene glycol was indicated as boiling point in Korea, distillate in Japan, distillation range in JECFA and USA, and its value was up to the standard. Distillation range of propionic acid was indicated as distillate in Korea and Japan, distillation range in JECFA and USA, and its value was up to the standard. There is no standard on distillation range of isopropyl alcohol in Japanese method. Test method of melting range on synthetic food additives was identical in all organizations, and there are 28 items to which this test method applies in Korean Food Additives Code. The standards on molting range of D-mannitol were different in various organizations, and in USA method there are no standards to which L-ascorbic acid, calciferol, and fumaric acid apply. Synthetic food additives performing the identification test were 251 items in Korean Food Additives Code, but there are no items to which manganese, glycerophosphate, bromate, thiosulfate, and bromide apply. Calcium benzoate was dissolved by heating in benzoate test and we could not identify the citrate in ferric citrate by method (2) of Korea and Japan. Identification test methods for ammonium, lactate, magnesium, copper, sulfate, phosphate, and zinc were identical in all organizations, and these could be identifed by current identification methods.