• Maxillofacial Plastic and Reconstructive Surgery
• /
• v.29 no.4
• /
• pp.279-288
• /
• 2007

In the present study, we focused on stem cells in the dental papilla of the tooth germ. The tooth germ, sometimes called the tooth bud, is the primordial structure from which a tooth is formed. The tooth germ consists of the enamel organ, the dental papilla, and the dental follicle. The dental papilla lies below a cellular aggregation of the enamel organ. Mesenchymal cells within the dental papilla are responsible for formation of dentin and pulp of a tooth. Tooth germ disappears as a tooth is formed, but that of a third molar stays in the jawbone of a human until the age of 10 to 16, because third molars grow slowly. Impacted third molar tooth germs from young adults are sometimes extracted for orthodontic treatment. In the present study, we evaluated the osteogenic activity and mineralization of cultured human dental papilla-derived cells. Dental papillas were harvested from mandible during surgical extraction of lower impacted third molar from 3 patients aged 13-15 years. After passage 3, the dental papilla-derived cells were trypsinized and subsequently suspended in the osteogenic induction DMEM medium supplemented with 10% fetal bovine serum, 50 g/ml L-ascorbic acid 2-phosphate, 10 nM dexamethasone and 10 mM -glycerophosphate at a density of $1\;{\times}10^6\;cells/dish$ in a 100-mm culture dish. The dental papilla-derived cells were then cultured for 6 weeks and the medium was changes every 3 days during the incubation period. Dental papilla-derived cells showed positive alkaline phosphatase (ALP) staining during 42 days of culture period. The formation of ALP stain showed its maximal manifestation at day 7 of culture period, then decreased in intensity during the culture period. ALP mRNA level was largely elevated at 1 weeks and gradually decreased with culture time. Osteocalcin mRNA expression appeared at day 14 in culture, after that its expression continuously increased in a time-dependent manner up to day 28. The expression remained constant thereafter. Runx2 expression appeared at day 7 with no detection thereafter. Von Kossa-positive mineralization nodules were first present at day 14 in culture followed by an increased number of positive nodules during the entire duration of the culture period. Osteocalcin secretion was detectable in the culture medium from 1 week. The secretion of osteocalcin from dental papilla-derived cells into the medium greatly increased after 3 weeks although it showed a shallow increase by then. In conclusion, our study showed that cultured human dental papilla-derived cells differentiated into active osteoblastic cells that were involved in synthesis of bone matrix and the subsequent mineralization of the matrix.

• Maxillofacial Plastic and Reconstructive Surgery
• /
• v.32 no.3
• /
• pp.199-206
• /
• 2010

This study investigated the effects of strontium on osteoblastic phenotypes of cultured human periostealderived cells. Periosteal tissues were harvested from mandible during surgical extraction of lower impacted third molar. Periosteal-derived cells were introduced into cell culture. After passage 3, the periostealderived cells were further cultured for 28 days in an osteogenic induction DMEM medium supplemented with fetal bovine serum, ascorbic acid 2-phosphate, dexamethasone and at a density of $3{\times}10^4$ cells/well in a 6-well plate. In this culture medium, strontium at different concentrations (1, 5, 10, and 100 ${\mu}g$/mL) was added. The medium was changed every 3 days during the incubation period. We examined the cellular proliferation, histochemical detection and biochemical measurements of alkaline phosphatase (ALP), the RT-PCR analysis for ALP and osteocalcin, and von Kossa staining and calcium contents in the periostealderived cells. Cell proliferation was not associated with the addition of strontium in periosteal-derived cells. The ALP activity in the periosteal-derived cells was higher in 5, 10, and 100 ${\mu}g$/ml strontium-treated cells than in untreated cells at day 14 of culture. Among the strontium-treated cells, the ALP activity was appreciably higher in 100 ${\mu}g$/ml strontium-treated cells than in 5 and 10 ${\mu}g$/ml strontium-treated cells. The levels of ALP and osteocalcin mRNA in the periosteal-derived cells was also higher in strontium-treated cells than in untreated cells at day 14 of culture. Their levels were increased in a dose-dependent manner. Von Kossa-positive mineralization nodules were strongly observed in the 1 ${\mu}g$/ml strontium-treated cells at day 21 and 28 of culture. The calcium content in the periosteal-derived cells was also higher in 1 ${\mu}g$/ml strontium-treated cells at day 28 of culture. These results suggest that low concentration of strontium stimulates the osteoblastic phenotypes of more differentiated periosteal-derived cells, whereas high concentration of strontium stimulates the osteoblastic phenotypes of less differentiated periosteal-derived cells. The effects of strontium on osteoblastic phenotypes of periosteal-derived cells appear to be associated with differentiation-extent.

• Maxillofacial Plastic and Reconstructive Surgery
• /
• v.29 no.3
• /
• pp.197-205
• /
• 2007

Angiogenesis is a essential part for bone formation and bone fracture healing. Vascular endothelial growth factor (VEGF), one of the most important molecules among many angiogenic factors, is a specific mitogen for vascular endothelial cells. VEGF-mediated angiogenesis is required for bone formation and repair. However, the effect of VEGF on osteoblastic cells during osteogenesis is still controversial. In recent days, substantial progress have been made toward developing tissue-engineered alternatives to autologous bone grafting for maxillofacial bony defects. Periosteum has received considerable interest as a better source of adult stem cells. Periosteum has the advantage of easy harvest and contains various cell types and progenitor cells that are able to differentiate into a several mesenchymal lineages, including bone. Several studies have reported the bone formation potential of periosteal cells, however, the correlation between VEGF signaling and cultured human periosteal cell-derived osteogenesis has not been fully investigated yet. The purpose of this study was to examine the correlation between VEGF signaling and cultured human periosteal-derived cells osteogenesis. Periosteal tissues of $5\;{\times}\;20\;mm$ were obtained from mandible during surgical extraction of lower impacted third molar from 3 patients. Periosteal-derived cells were introduced into the cell culture and were subcultured once they reached confluence. After passage 3, the periosteal-derived cells were further cultured for 42 days in an osteogenic inductive culture medium containing dexamethasone, ascorbic acid, and ${\beta}-glycerophosphate$. We evaluated the alkaline phosphatase (ALP) activity, the expression of Runx2 and VEGF, alizarin red S staining, and the quantification of osteocalcin and VEGF secretion in the periosteal-derived cells. The ALP activity increased rapidly up to day 14, followed by decrease in activity to day 35. Runx2 was expressed strongly at day 7, followed by decreased expression at day 14, and its expression was not observed thereafter. Both VEGF 165 and VEGF 121 were expressed strongly at day 35 and 42 of culture, particularly during the later stages of differentiation. Alizarin red S-positive nodules were first observed on day 14 and then increased in number during the entire culture period. Osteocalcin and VEGF were first detected in the culture medium on day 14, and their levels increased thereafter in a time-dependent manner. These results suggest that VEGF secretion from cultured human periosteal-derived cells increases along with mineralization process of the extracellular matrix. The level of VEGF secretion from periosteal-derived cells might depend on the extent of osteoblastic differentiation.

• Korean Journal of Food Science and Technology
• /
• v.30 no.5
• /
• pp.1097-1106
• /
• 1998

Biodegradable edible films were prepared from rice protein concentrates (RPC) made from rice wine meal by alkaline extraction and isoelectric precipitation. The effect of film forming solution pH and plasticizers were studied, and cross-linkers were added to improve mechanical properties and water vapor permeabilities (WVP) of films. Films could be formed within pH $8{\sim}11$ with tensile strength (TS) of 4.3{\sim}5.7\;MPa$. Films produced under pH 11 had the highest TS (5.7 MPa) and the lowest WVP $(0.44\;ng{\cdot}m/m^2{\cdot}s{\cdot}Pa)$. Added glycerol, polyethylene glycol 200 (PEG) and its mixture (GLY:PEG=50:50) as plasticizers also affected the mechanical properties and WVP of films. TS and elongation at break (E) of films at various plasticizer levels were $5.5{\sim}1.0\;MPa$ and $3.6{\sim}24.3%$, respectively. At the same plasticizer concentration, the highest TS was observed when glycerol was used whereas the highest E was measured when mixture was used as plasticizer. WVPs of films with thickness of $60\;{\mu}m$ were $0.39{\sim}0.54\;ng{\cdot}m/m^2{\cdot}s{\cdot}Pa$. WVP of films decreased as the ratio of glycerol/PEG 200 was decreased, and WVP increased as the total amount of plasticizer added to the films increased. Film strength was improved by the addition of small amount of sodium hydrogen sulfate, succinic anhydride, ascorbic acid and citric acid, whereas TS of films containing $0.5{\sim}2.0%$ of NaCl and $CaCl_2$ were lower than those without the salts. The highest TS (6.3 MPa) was achieved with films containing 0.1% of succinic anhydride.

• Journal of Periodontal and Implant Science
• /
• v.31 no.4
• /
• pp.833-846
• /
• 2001

Recently, use of natural medicine is getting more attention, and some of them are believed to be effective in　the treatment of periodontitis. Among them, the seeds of safflower(Carthamus tinctrorius L.) have been proven to be effective through its use in bone diseases such as fracture and osteoporosis. During the last few years, studies using the seeds of safflower gown in Korea have been active, and it has been reported that safflower seed extract increase the proliferation and the alkaline phosphatase(ALP) activity of human periodontal ligament fibroblast(hPDLF), osteoblast, and that they promote the mineralization process. In animal studies, when safflower seed extract were administered orally new bone formation was promoted. Recently, in an effort to find out the most effective osteogenic components, among many components of the safflower seed, various safflower seed fraction extracts were obtained by multistep extraction of the safflower components using various solvents. Among these, saf-M-W fraction extracted by methanol and water was most effective in increasing osteogenic potential of osteoblasts. In this study, the effect of safflower seed fraction extract, saf-M-W, on the growth and differentiation of hPDLF and MC3T3-E1 cell was investigated. The toxicity of saf-M-W on both cells was measured using M'IT(3-(4,5dimethylthiazol-2-y1)-2,5-diphenyl tetrazolium bromide) test, and ALP activity was measured using the colorimetric assay of hPDLF. In addition, in MC3T3-El cells, the expression of ALP, bone sialoprotein(BSP) mRNA was observed using Northern blot, and the mineralized nodule formation Was observed using von Kossa stain and phase-contrast microscope. 1. In concentrations below $10{\mu}g/ml$, saf-M-W didn't show any toxicity on hPDLF and MC3T3-El cell. 2. The change in saf-M-W concentration had no effect on the ALP activity of hPDLF. 3. In MC3T-E1 cells, mRNA expressions of ALP and BSP were greater in the experimental group treated with $10{\mu}g/ml$ concentration of saf-M-W compared with the control group. 4. In MC3T3-El cells, abundance of mineralized nodules were formed in the experimental group treated with $10{\mu}g/ml$ Concentration of saf-M-W, while no mineralized nodule was formed in the control group. These results suggest that safflower seed fraction extract, saf-M-W. didn't show any toxicity on hPDLF and MC3T3-E1 cell at concentrations below $10{\mu}g/ml$ and effectively enhanced the differentiation and osteogenic potential of MC3T3-El cell.

• Journal of the Korean Wood Science and Technology
• /
• v.7 no.2
• /
• pp.3-30
• /
• 1979

Larch ($\underline{Larix}$ $\underline{leptolepis}$ GORDON), one of the major afforestation species in Korea in view of its growing stock and rate of growth, is not favored as a raw material for pulp due to its low yield of pulp and difficulties with bleaching arising from the high content of extractives in wood, and the high heartwood ratio and the active phenolics, respectively. The purpose of this study is to investigate the characteristics of firstly pulping with various additives of cellulose protector for the yield of pulp, and secondly bleaching with oxygen for chlotination-alkali extraction of five stage-sequence to reduce chlorine compounds in bleaching effluents. The kraft cooking liquor for five age groups of larchwood was 18 percent active alkali with 25 percent sulfidity and 5 : 1 liquor-to-wood ratio, and each soda liquor for sap-and heart-wood of the 15-year-old larchwood was 18 percent alkali having one of the following cellulose protectors as the additive; magnesium sulfate ($MgSO_4$, 2.5%), zinc sulfate ($ZnSO_4$, 2.5%), aluminium sulfate ($Al_2(SO_4)_3$, 2.5%), potasium iodide (KI, 2.5%), hydroquinone (HQ, 2.5%), anthraquinone (AQ, 0.1%) and ethylene diamine (EDA, 2.5%). Then each anthraquinone-soda liquor for the determination of suitable cooking condition was the active alkali level of 15, 17 and 19 percent with 1.0, 0.5 and 0.1 percent anthraquinone, respectively. The cooking procedure for the pulps was scheduled to heat to 170$^{\circ}C$ in 90 minutes and to cook 90 minutes at the maximum temperature. The anthraquinone-soda pulps from both heartwood and sapwood of 15-year-old larchwood prepared with 0.5 percent anthraquinone and 18 percent active alkali were bleached in a four-stage sequency of OCED. (O: oxygen bleaching, D: chlorine dioxide bleaching and E: alkali extraction). In the first stage oxygen in atmospheric pressure was applied to a 30 percent consistency of pulp with 0.1 percent magnesium oxide (MgO) and 3, 6, and 9 percent sodium hydroxide on oven dry base, and the bleached results were compared pulps bleached under the conventional CEDED (C: chlorination). The results in the study were summarized as follows: 1. The screened yield of larch kraft pulp did not differ from particular ages to age group, but heartwood ratio, basic density, fiber length and water-extractives contents of wood and the tear factor of the pulp increased with increasing the tree age. The total yield of the pulp decreased. 2. The yield of soda pulp with various chemicals for cellulose protection of the 15-year-old larchwood increased slightly more than that of pure soda pulp and was slightly lower than that of kraft pulp. The influence of cellulose protectors was similar to the yield of pulps from both sapwood and heartwood. The effective protectors among seven additives were KI, $MgSO_4$ and AQ, for which the yields of screened pulp was as high as that of kraft pulp. Considering the additive level of protector, the AQ was the most effective in improving the yield and the quality of pulp. 3. When the amount of AQ increased in soda cooking, the yield and the quality of the pulp increased but rejects in total yield increased with decreasing the amount of active alkali from 19 to 15 percent. The best proportion of the AQ seemed to be 0.5 percent at 17 percent active alkali in anthraquinone-soda pulping. 4. On the bleaching of the AQ-soda pulp at 30 percent consistency with oxygen of atomospheric pressure in the first stage of the ODED sequence, the more caustic soda added, the brighter bleached pulp was obtained, but more lignin-selective bleaching reagent in proportion to the oxygen was necessary to maintain the increased yield with the addition of anthraquinone. 5. In conclusion, the suitable pulping condition for larchwood to improve the yield and quality of the chemical pulp to the level for kraft pulp from conventional process seemed to be. A) the selection of young larchwood to prevent decreasing in yield and quality due to the accumulation extractives in old wood, B) the application of 0.5 percent anthraquinone to the conventional soda cooking of 18 percent active alkali, and followed, C) the bleaching of oxygen in atmospheric pressure on high consistency (30%) with 0.1 percent magnesium oxide in the first stage of the ODED sequence to reduce the content of chlorine compounds in effluent.