• Journal of Periodontal and Implant Science
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• v.41 no.5
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• pp.218-226
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• 2011

Purpose: Micro-computed tomography (micro-CT) has been widely used in the evaluation of regenerated bone tissue but the reliability of micro-CT has not yet been established. This study evaluated the correlation between histomorphometric analysis and micro-CT analysis in performing new bone formation measurement. Methods: Critical-size calvarial defects were created using a 8 mm trephine bur in a total of 24 Sprague-Dawley rats, and collagen gel mixed with autogenous rat bone marrow stromal cells (BMSCs) or autogenous rat BMSCs transduced by adenovirus containing bone morphogenic protein-2 (BMP-2) genes was loaded into the defect site. In the control group, collagen gel alone was loaded into the defect. After 2 and 4 weeks, the animals were euthanized and calvaria containing defects were harvested. Micro-CT analysis and histomorphometric analysis of each sample were accomplished and the statistical evaluation about the correlation between both analyses was performed. Results: New bone formation of the BMP-2 group was greater than that of the other groups at 2 and 4 weeks in both histomorphometric analysis and micro-CT analysis (P=0.026, P=0.034). Histomorphometric analysis of representative sections showed similar results to histomorphometric analysis with a mean value of 3 sections. Measurement of new bone formation was highly correlated between histomorphometric analysis and micro-CT analysis, especially at the low lower threshold level at 2 weeks (adjusted $r^2=0.907$, P<0.001). New bone formation of the BMP-2 group analyzed by micro-CT tended to decline sharply with an increasing lower threshold level, and it was statistically significant (P<0.001). Conclusions: Both histomorphometric analysis and micro-CT analysis were valid methods for measurement of the new bone in rat calvarial defects and the ability to detect the new bone in micro-CT analysis was highly influenced by the threshold level in the BMP-2 group at early stage.

• Korean Journal of Veterinary Research
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• v.60 no.3
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• pp.145-153
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• 2020

This study aimed to evaluate the effects of a bioabsorbable bone hemostatic agent comprising poly (ethylene glycol-propylene glycol) copolymers (PEG-PPG) on hemostasis and osteogenesis. Bilateral 3 mm diameter calvarial defects were created in 99 male Sprague-Dawley rats. The defects were filled with PEG-PPG or bone wax. The defects of control group were left unfilled. Virtual autopsy was performed to evaluate bioabsorption. The calvaria were subjected to x-ray microtomography (microCT) and histological examination. Bone volume fraction (BV/TV) and bone mineral density (BMD) were measured using microCT; furthermore, white blood cell count and histological examination were performed. After application of PEG-PPG and bone wax, immediate hemostasis was achieved. Autopsy revealed that PEG-PPG disappeared within 48 h at the application site; in contrast, bone wax remained until 12 weeks. The PEG-PPG and control groups showed significantly more osteogenesis than the bone wax group with respect to BV/TV and BMD at 3, 6, and 12 weeks (p < 0.05). Histology revealed that the bone wax group exhibited little bone formation with inflammation. In contrast, PEG-PPG and control groups showed significantly more qualitative osteogenesis than the bone wax group (p < 0.01). In conclusion, PEG-PPG showed immediate hemostasis and was absorbed to allow progressive osteogenesis.

• Maxillofacial Plastic and Reconstructive Surgery
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• v.32 no.4
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• pp.313-323
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• 2010

Purpose: This study was conducted to evaluate the effect of beta-tricalcium phosphate (Cerasorb$^{(R)}$, Germany) and deproteinized bovine bone (Bio-Oss$^{(R)}$, Switzerland) grafted to the defect of rat calvaria artificially created and the effect of use of absorbable membrane (BioMesh$^{(R)}$, Korea) on new bone formation. Materials and Methods: Transosseous circular calvarial defects with diameters of 5 mm were prepared in the both parietal bone of 30 rats. In the control group I, no specific treatment was done on the defects. In the control group II, the defects were covered with absorbable membrane. In the experimental group I, deproteinized bovine bone was grafted without absorbable membrane; in the experimental group II, deproteinized bovine bone was grafted with absorbable membrane; in the experimental group III, beta-tricalcium phosphate was grafted without absorbable membrane; in the experimental group IV, beta-tricalcium phosphate was grafted with absorbable membrane. The animals were sacrificed after 3 weeks and 6 weeks respectively, and histologic and histomorphometric evaluations were performed. Results: Compare to the control groups, the experimental groups showed more newly formed bone. Between the experimental groups, beta-tricalcium phosphate showed more resorption than deproteinized bovine bone. Stabilization of grafted material and interception of the soft tissue invasion was observed in the specimen treated with membrane. There was no statistical difference between the experimental group I, III and experimental group II, IV classified by graft material, but statistically significant increase in the amount of newly formed bone was observed in the experimental group I, II and II, IV classified by the use of membrane (P<0.05). Conclusion: Both beta-tricalcium phosphate and deproteinized bovine bone showed similar osteoconductibility, but beta-tricalcium phosphate is thought to be closer to ideal synthetic graft material because it showed higher resorption rate in vivo. Increased new bone formation can be expected in bone graft with use of membrane.

• Maxillofacial Plastic and Reconstructive Surgery
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• v.21 no.1
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• pp.13-22
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• 1999

The purpose of this investigation was to evaluate the acceptability of the collagen-based xenograft ($Laddec^{(R)}$). Full thickness bone defects were prepared in the calvaria of the rats. In the experimental groups the bone defects were filled with a kind of collagen based xenograft. And bone defects, which left without filling, were used as control groups. Sequential sacrifice was performed at the 1st, 2nd, 4th, 8th, and 16th weeks of experiment. 1. At the 1st week of experiment, infiltration of chronic inflammatory cell was observed in all groups. In the experimental group, resorption of the xenograft was initiated. 2. At the 2nd week of experiment, infiltration of chronic inflammatory cells was decreased in all groups. In the experimental group, active resorption of xenograft and new bone formation from the periphery of the xenograft was observed. 3. At the 4th and 8th weeks of experiment, more resorption of the xenograft and new bone formation with calcification was observed in the experimental group. 4. At the 16th week of experiment, small bone trabecula was formed partially in the control group but that couldn't fill the whole bone defect. In the experimental group, more advanced resorption of xenograft and more new bone formation was observed. However mid portion of the xenograft was still remained without resorption. 5. From this experiment, we concluded that the collagen-based xenograft had some osteoconductive but no osteoinductive property. So the xenograft would be used for the bone defect filling material where rapid bone remodeling is not required.

• Journal of the Korean Association of Oral and Maxillofacial Surgeons
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• v.28 no.3
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• pp.188-195
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• 2002

The purpose of this investigation was to evaluate the relationship between the hydration time of demineralized freeze-dried bone (DFDB) and early new bone formation in rat calvarial defects filled with DFDB. Rats (n = 43) were divided into 4 experimental groups. Standard, transosseous circular defects of the calvaria were made midparietally. In experimental group 1, the defect was grafted immediately after soaking the DFDB. In experimental group 2, the defects were grafted with DFDB after soaking the DFDB for 10 minutes. In experimental groups 3 and 4, the defects were filled after soaking the DFDB for 30 and 60 minutes, respectively. Graft sites were analyzed histologically after healing periods of 1, 2, or 4 weeks. Each group showed similar bone regeneration at each time point by histological analysis. The results of this study were as follows: 1. After 1 week, a significant amount of inflammation, granulation tissue, and edema were found. A small amount of bone was seen, but the amount of bone did not differ between groups. 2. After 2 weeks, a small amount of new bone formation and DFDB resorption were observed. 3. After 4 weeks, a greater amount of new bone formation was observed. The greatest amount of bone formation occurred in experimental group 4 after 4 weeks. We conclude that the hydration time of DFDB does not affect new bone formation and that it is very important to control inflammation in bone grafting.

• Journal of Periodontal and Implant Science
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• v.27 no.4
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• pp.867-882
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• 1997

Safflower(Carthamus tinctorius $Linn\acute{e}$ has been traditionally used for the treatment of blood stasis, and Dipsasi Radix has been used as a drug for fracture in Chinese medicine. The purpose of present study was to examine the biologic effects of safflower extract and Disasi radix extracts on the periodontal. ligament cells and osteoblastic cells and on the wound healing of rat calvarial defect. The ethanolic extract of safflower blossom, safflower seed and Dipsasi Radix(125, 250, and 500 ${\mu}g/ml$) were prepared as test group, and PDGF-BB(lOng/ml) and unsafonifiable fraction of Zea Mays L.(125, 250, and 500 ${\mu}g/ml$) were employed as positive control. The effects of each agents on the growth and survival, ALPase activity, expression of PDGF-BB receptor, chemotactic response of PDL cell and ATCC human osteosarcoma MG63 cells in vitro were examined. The tissue regenerative effect of each extracts was evaluated by histomorphometric measuring of newly formed bone on the 8mm defect in rat calvaria after oral administration of 3 different dosages groups : 0.02, 0.1 and 0.35g/kg, per day. It was also employed the same dosages of unsaponifiable fraction of Zea Mays L. as positive controls. Safflower blossom extract, safflower seed extract, and Dipsasi Radix extract stimulate the cellular activity of MG63 cells in concentration range of $125-500{\mu}g/ml$, and safflower bolssom extract and safflower seed extract stimulate also the cellular activity of periodontal ligament cells in concentration range of $250-500{\mu}g/ml$. In activity of ALPase, $250-500{\mu}g/ml$ of safflower blossom extracts showed significant stimulating effects on MG63 cells, and the same concentration range of safflower seed extracts showed significant effect on periodontal ligament cells. In the recovery on PDGF-BB receptor expression which was depressed by $IL-1{\beta}$, $125-250{\mu}g/ml$ of safflower blossom extracts and $250-500{\mu}g/ml$ of safflower seed extracts showed significant increasing effect on MG63 cells, and $500{\mu}g/ml$ of safflower blossom extract and $250-500{\mu}g/ml$ of safflower seed extracts showed significant effect on periodontal ligament cells. In chemotactic response, among all tested group, safflower seed extracts only were chemotactic to MG63 cells and periodontal ligament cells in concentration range of $125-500{\mu}g/ml$. Also in the view of bone regeneration in rat calvarial defect model, the only group that was orally administrated 0.35g/kg, day of safflower seed extract showed significant new bone formation. These results suggested that safflower extracts might have a potential possibilities as an useful drug for adjunct to treatment for regeneration of periodontal defect.

• Journal of Periodontal and Implant Science
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• v.28 no.4
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• pp.545-559
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• 1998

Magnoliae cortex has been used as a drug for treatment of fractures in Chinese medicine and safflower(Carthamus tinctorius $Linn{\acute{e}}$) has been traditionally used for treatment of blood stasis. The purpose of present study was to examine the biologic effects of magnoliae cortex extract and safflower extract mixture(MSM) on human periodontal ligament cells and fetal rat calvarial osteoblasts and on healing of rat calvarial defects. The ethanolic extracts of magnoliae cortex(MCE), safflower seed(SSE), Zea May L(ZML) were prepared as positive control group. MSM mixed to the ratios of 1 : 1, 1 : 2, 1 : 5 and 1 : 10 were used as test group. The effects of each agents on the growth and survival, ALPase activity, cell proliferation and tissue regenerative effect of each extracts was evaluated by histomorphometric measuring of newly formed bone on the 8 mm defect in rat calvaria after oral administration of 2 ratio groups(1 : 5 and 1 : 10) at 3 different doses (0.1, 0.25 and 0.5g/kg per day). MSM stimulated the growth and survival rate of osteoblasts and PDL cells more than any other agents. The growth and survival rate were increased as the proportion of safflower seed extract was increased. MCE, SSE, ZML stimulated the ALPase activity of osteoblast and PDL cell in comparison to the negative control group. But all groups of MSM regardless of ratio of safflower seed extract stimulated the ALPase activity than any other agent. The ALPase activity was also increased as the proportion of safflower seed extract was increased. Although MCE, SSE, ZML stimulated the proliferation of osteoblasts. 1 : 5 and 1 : 10 ratio MSM showed significant increase in stimulation of proliferation of osteoblasts. No agent significantly increased proliferation of PDL cells. Significant new bone formation were seen where 1 : 5 ratio, 0.5g/kg group and 1 : 10 ratio, 0.25, 0.5g/kg groups were used. These results show that magnoliae cortex extract and safflower seed extract mixture can potentially increase bone regeneration ability.

• Journal of Periodontal and Implant Science
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• v.49 no.1
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• pp.2-13
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• 2019

Purpose: The aim of this study was to conduct a histologic evaluation of irradiated calvarial defects in rats 4 weeks after applying fibroblast growth factor-2 (FGF-2) with hyaluronan or biphasic calcium phosphate (BCP) block in the presence or absence of adjunctive hyperbaric oxygen (HBO) therapy. Methods: Twenty rats were divided into HBO and non-HBO (NHBO) groups, each of which was divided into FGF-2 and BCP-block subgroups according to the grafted material. Localized radiation with a single 12-Gy dose was applied to the calvaria of rats to simulate radiotherapy. Four weeks after applying this radiation, 2 symmetrical circular defects with a diameter of 6 mm were created in the parietal bones of each animal. The right-side defect was filled with the materials mentioned above and the left-side defect was not filled (as a control). All defects were covered with a resorbable barrier membrane. During 4 weeks of healing, 1 hour of HBO therapy was applied to the rats in the HBO groups 5 times a week. The rats were then killed, and the calvarial specimens were harvested for radiographic and histologic analyses. Results: New bone formation was greatest in the FGF-2 subgroup, and improvement was not found in the BCP subgroup. HBO seemed to have a minimal effect on new bone formation. There was tendency for more angiogenesis in the HBO groups than the NHBO groups, but the group with HBO and FGF-2 did not show significantly better outcomes than the HBO-only group or the NHBO group with FGF-2. Conclusions: HBO exerted beneficial effects on angiogenesis in calvarial defects of irradiated rats over a 4-week healing period, but it appeared to have minimal effects on bone regeneration. FGF-2 seemed to enhance new bone formation and angiogenesis, but its efficacy appeared to be reduced when HBO was applied.

• Maxillofacial Plastic and Reconstructive Surgery
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• v.34 no.1
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• pp.1-11
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• 2012

Purpose: The purpose of this study was to evaluate the bone regeneration capacity of silk fibroin (SF) when combined with beta tricalcium phosphate (${\beta}$-tricalcium phosphate [TCP]) and rh-bone morphogenetic protein (BMP) in vivo by micro-computed tomography (CT), soft x-ray, and histological analysis. Methods: A total of 56 critical size defects formed by a trephine bur made on 28 adult female Spague-Dawley rats were used for this study and the defect size was 5.0 mm in diameter. The defects were transplanted with (1) no graft material (raw defect), (2) autogenous bone, (3) SF ($10{\mu}g$), (4) SF-BMP ($10{\mu}g$, $0.8{\mu}g$ each), and (5) SF+${\beta}$-TCP ($10{\mu}g$). At 4 and 8 weeks after operation, the experimental animals were sacrificed. Samples were evaluated with soft x-ray, histological examinations and 3-dimensional micro-CT analysis. Results: In the 3-dimensional micro-CT evaluation, bone volume and bone surface data were higher in the SF-BMP ($12.8{\pm}1.5$, $138.6{\pm}45.0$ each) (P<0.05) and SF-TCP ($12.3{\pm}1.5$, $144.9{\pm}30.9$ each) group than in the SF group ($6.1{\pm}3.3$, $77.2{\pm}37.3$ each) (P<0.05), except for the autogenous group ($15.0{\pm}3.0$, $190.7{\pm}41.4$ each) at 4 weeks. At 8 weeks, SF-BMP ($16.8{\pm}3.5$, $173.9{\pm}34.2$ each) still revealed higher (P<0.05) bone volum and surface, but SF-TCP ($11.3{\pm}1.5$, $1132.9{\pm}52.1$ each) (P=0.5, P=0.2) revealed the same or lower amount compared with the SF group ($13.8{\pm}2.7$, $127.5{\pm}44.8$ each). The % of bone area determined by radiodensity was higher in the SF-TCP ($31.4{\pm}9.1%$) and SF-BMP ($36.2{\pm}16.2%$) groups than in the SF ($19.0{\pm}10.4$) group at the period of 4 weeks. Also, in the histological evaluation, the SF-BMP group revealed lower inflammation reaction, lower foreign body reaction and higher bone healing than the SF group at postoperative 4 weeks and 8 weeks. The SF-TCP group revealed lower inflammation at 4 weeks, but accordingly, as the TCP membrane was absorbed, inflammatory and foreign body reaction are increased at 8 weeks. Conclusion: The current study provides evidence that the silk fibrin can be used as an effective grafted material for tissue engineering bone generation through a combination of growth factor or surface treatment.

• The Journal of Korean Academy of Prosthodontics
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• v.48 no.1
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• pp.16-27
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• 2010

Purpose: The aim of this experimental study is to observe the effect of platelet-rich plasma (PRP) on early bone regeneration of rats both in normal condition and in osteoporosis induced by ovariectomy. Material and methods: Total 40 Sprague-Dawley female rats were divided into 4 groups. A 8-mm-diameter calvarial critical-sized defect (CSD) was made by drilling with trephine at the center of calvaria in cranium of every rat. Every CSD was augmented with an osteoconductive synthetic alloplastic substitute ($MBCP^{TM}$) and PRP as follows. Group A; 10 non-ovariectomized rats grafted with only $MBCP^{TM}$. Group B; 10 non-ovariectomized rats grafted with $MBCP^{TM}$ and PRP. Group C; 10 ovariectomized rats grafted with only $MBCP^{TM}$. Group D; 10 ovariectomized rats grafted with $MBCP^{TM}$ and PRP. At 4 weeks after graft, every rat was sacrificed. And histomorphometric analysis was performed by measuring calcified area of new bone formation within the CSD. Results: Comparing four groups, results were obtained as follows. 1. In non-ovariectomized groups, PRP showed a positive effect somewhat but not significant (P > .05). 2. In ovariectomized groups, PRP showed a positive effect significantly (P < .05). 3. In PRP untreated groups, ovariectomy diminished bone regeneration significantly (P < .05). 4. In PRP treated groups, ovariectomy diminished bone regeneration somewhat but not significant (P > .05). Conclusion: Within the limitation of this study, it can be concluded that PRP in combination with an osteoconductive synthetic alloplastic substitute has an effect on bone regeneration more significantly in ovariectomized osteoporotic rats than in normal rats.