• Restorative Dentistry and Endodontics
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• v.26 no.3
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• pp.191-199
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• 2001

목적 - 기질금속단백분해효소(matrix metalloproteinase)는 조직의 염증 및 치유과정에서 숙주세포에서 생성, 분비되어 세포외기질(extracellular matrix)의 분해에 작용한다. 다양한 염증반응에 기질금속단백분해효소가 중요한 역할을 하는것으로 보고되고 있으나 치수 및 치근단 질환에서의 그 역할은 거의 알려져 있지 않은 상태이다. 본 연구에서는 염증이 있는 사람의 치수 및 치근단 조직을 채취하여 Enzymeimmunoassay 및 면역조직화학적 검색을 통해 제1형, 2형, 3형 기질금속단백분해효소의 수준 및 그 분포를 측정하여 치수 및 치근단 병소에서 이 효소의 작용을 알아보는 것을 목적으로 한다. 방법 - 연구재료는 근관치료를 위해 서울대학교 병원 치과 진료부 보존과에 내원한 환자를 대상으로 34개의 치아에서 통상의 근관치료 중 발수한 치수조직과 치근단 수술중 얻은 치근단 병소(n=10)를 이용하였다. 치수는 발수 전에 임상진단을 통해 급성 치수염(n=12), 만성 치수염(n=12), 정상 치수(n=10)로 구분하고 정상치수로 진단된 것을 대조군으로 설정하였다. 채취된 표본은 둘로 나누어 절반은 30분 이내에 5$\mu\textrm{m}$ 두께로 동결절단을 시행하여 조직표본을 제작하였고 deep freezer에 보관하였다가 헤마톡실린-에오신 염색 및 면역조직화학적 검색을 시행하였다. 나머지 조직은 ELISA를 위해 액체 질소에 보관하였다. ELISA를 시행하기전 표본의 단백질 정량을 시행하여 모든 표본의 단백질 양을 50mg/$\mu\textrm{l}$로 일치시키고 Amersham사의 ELISA kit를 사용하여 제1형, 2형, 3형의 기질금속단백분해효소의 양을 측정하였으며 그 결과를 Mann-Whitney U test를 사용하여 각 군간의 통계학적 유의성을 검증하였다. 결과 1. ELISA의 결과 제1형 기질금속단백분해효소의 농도는 모든 실험군에서 대조군보다 유의성있게 높게 나타났다.(p<.05). 또한 급성치수염군의 제1형 기질금속단백분해효소의 농도가 다른 실험군보다 유의성있게 높았다(p<.05). 2. 제2형 기질금속단백분해효소의 경우 급성치수염군과 대조군에서만 유의성있는 차이를 보였다(p<.05). 3. 제3형 기질금속단백분해효소의 경우 급성치수염군에서 대조군이나 만성치수염군보다 유의성 있는 높은 수치를 보였다(p<.05). 4. 면역조직화학검색 결과 염증성 치수에 존재하는 급성 및 만성염증세포 주위로 기질금속단백분해효소에 대한 면역 반응이 존재하였으며 주로 제1형과 제3형 기질금속단백분해효소의 경우 대식세포 및 림파구 주위로 강한 발색제의 침윤양상이 관찰되었다. 5. 치근단병소의 면역조직화학적 검색 결과 만성염증 세포 주변으로 미약한 발색제의 침윤양상이 관찰되었다.

• Journal of the korean academy of Pediatric Dentistry
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• v.45 no.2
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• pp.242-249
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• 2018

The purpose of this study is to compare the properties of dental pulp and periodontal ligament stem cells from extracted supernumerary teeth by quantitative real-time PCR. Impacted supernumerary teeth in the maxillary anterior region were extracted. Dental pulp and periodontal ligament cells were collected from extracted supernumerary teeth on the same day. After isolation and culture of cells, compare characterization of them by using qRT-PCR. Primer sequences for odontoblasts are ONT, ALP, OCN, DMP-1 and DSPP. On dental pulp group, ONT has the largest quantity of gene expression, followed by OCN, ALP, DMP-1 and DSPP. On periodontal ligament group, ONT has the largest quantity of gene expression, followed by OCN, ALP, DSPP and DMP-1. Analysis of quantitative gene expression data using relative quantification showed that the expression of all genes decreased in periodontal ligament cells. Dental pulp and periodontal ligament stem cells from supernumerary teeth have the properties of odontoblasts. Considering that properties, supernumerary teeth were considered a useful donor site of dental pulp and periodontal ligament stem cells.

• Restorative Dentistry and Endodontics
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• v.35 no.3
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• pp.152-163
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• 2010

This study investigated the changes in gene expression when mineral trioxide aggregate (MTA) was applied in vitro to human dental pulp cells (HDPCs). MTA in a teflon tube (diameter 10 mm, height 2 mm) was applied to HDPCs. Empty tube-applied HDPCs were used as negative control. For microarray analysis, total RNA was extracted at 6, 24, and 72 hrs after MTA application. The results were confirmed selectively by performing reverse transcriptase polymerase chain reaction for genes that showed changes of more than two-fold or less than half. Of the 24,546 genes, 109 genes were up-regulated greater than twofold (e.g., FOSB, THBS1, BHLHB2, EDN1, IL11, FN1, COL10A1, and TUFT1) and 69 genes were down-regulated below 50% (e.g., SMAD6 and DCN). These results suggest that MTA, rather than being a bio-inert material, may have potential to affect the proliferation and differentiation of pulp cells in various ways.

• Restorative Dentistry and Endodontics
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• v.36 no.1
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• pp.26-36
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• 2011

Objectives: In the present study, three kinds of tissues cells (pulp, gingiva, and periodontal ligament) were investigated if those cells express MMP and TIMP when they were stimulated with neuropeptides (substance P, CGRP) or proinflammatory cytokine, TNF-$\alpha$. Materials and Methods: The cells cultured from human dental pulp (PF), gingiva (GF) and periodontal ligament were (PDLF) stimulated with Mock, SP, TNF-$\alpha$, and CGRP for 24 hrs and 48 hrs. for an RNase protection assay and Enzyme Linked Immunosorbent Assay. Cells (PF, GF and PDLF) seeded in 100 mm culture dish were stimulated with SP ($10^{-5}$, $10^{-8}\;M$) or only with medium (Mock stimulation) for 4hrs and for 24 hrs for RNase Protection Assay, and they were stimulated with CGRP ($10^{-5}\;M$) and TNF-$\alpha$(2 ng/mL) for 24 hrs and with various concentraion of TNF-$\alpha$(2, 10, and 100 ng/mL) for Rnase Protection Assay with a human MMP-1 probe set including MMP 1, 2, 8, 7, 8, 9, 12, and TIMP 2, 3. In addition, cells (PF, GF and PDLF) were stimulated with Mock and various concentraion of TNF-$\alpha$(2, 10, and 100 ng/mL) for 24 hrs and with TNF-$\alpha$(10 ng/mL) for 48 hrs, and the supernatents from the cells were collected for Enzyme Linked Immunosorbent Assay (ELISA) for MMP-1 and MMP-13. Results: The expression of MMPs in PF, GF, PDLF after stimulation with SP and CGRP were not changed compared with Mock stimulation for 4 hrs and 24 hrs. The expression of MMP-1, -12, -13 24 hrs after stimulation with TNF-$\alpha$ were upregulated, however the expression of TIMP-3 in PF, GF, PDLF after stimulation with TNF-$\alpha$ were downregulated. TNF-$\alpha$(2 ng/mL, 10 ng/mL, 100 ng/mL) increased MMP-1 and MMP-12 expression in PF dose dependently for 24 hrs. Conclusions: TNF-$\alpha$ in the area of inflammation may play an important role in regulating the remodeling of dentin, cementum, and alveolar bone.

• Journal of the korean academy of Pediatric Dentistry
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• v.27 no.2
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• pp.318-332
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• 2000

The purpose of this study was to investigate the effects of demineralized freeze-dried bone (DFDB) on mechanically exposed pulp of dog by evaluating the pulpal inflammation and healing process, formation of dental hard tissue, and structural changes of fibroblasts of the remaining pulp tissue. Teeth of 4 dogs, weighing 10kg, were used in this study. Class V cavities were prepared followed by exposed the pulp tissue mechanically by sterilized round bur. In control group, exposed pulps were capped with calcium hydroxide paste followed by sealed with IRM. In experimental groups, the exposed pulps of one group were capped with the collagen and those of the other group were capped with DFDB. All cavities were sealed with same manor as control group. The animals were sacrificed at the intervals of 3, 7, 14, and 28 days for histopathlogic evaluation. The specimens were observed by the light microscope and trans-electron microscope. The results were as follows: 1. Pulp necrosis was not observed in all groups. Inflammatory response was disappeared from 1 week in control group and group 2. But it was not disappeared until 2 weeks and also irregular arrangement of odontoblasts was showed at the lateral walls of root canal just beneath the amputated site of the pulp in group 1. 2. Dentinal bridge was formed incompletely at 2 weeks but it was formed completely at 4 weeks in control group. Odontoid tissue was also found in control group at 4 weeks from treatment. Amputated site of pulp was encapsulated with fibrous tissue and odontoblast and dentinal bridge was not found in group 1. Preodontoid tissue and reparative dentin which were formed by odontoblast differentiated around DFDB were found, but dentinal bridge was not found in group 2. 3. Cell with large basophillic-stained nuclei infiltrated to amputated site and DFDB at 1 week from treatment in control group and group 2. They were found more in group 2 than in control group. Odontoblasts arranged more regularly and reparative dentin was found more as time elapsed. 4. Dentin-formative odontoblasts which showed ultramicrostructure of cytoplasm with polarized nucleus, rEM, Golgi complex, secretory granules, secretion of organic matrix in control of group and group 2. In regards to above results, the demineralized freeze-dried bone(DFDB) induce odontoblastic differentiation and further come up to the dentin formation in amputated pulp.

• Restorative Dentistry and Endodontics
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• v.35 no.4
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• pp.238-245
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• 2010

Numerous cases about additional growth of roots or pulp tissue regeneration by using various intracanal medicaments in immature permanent teeth with periapical or pulpal disease have been reported. The underlying mechanism has not been clearly delineated, but it has been widely accepted that undifferentiated mesenchymal cells and stem cells are involved. Moreover, the growth and deposition of osteoid or cementoid tissues have been observed in regenerated pulp and roots. This new and non-invasive treatment has brightened the future of endodontics, and enlarged the vision of regenerative root canal treatment with multi-potent stem cells and various tissue engineering techniques.

• Journal of dental hygiene science
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• v.13 no.3
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• pp.296-303
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• 2013

Lysyl oxidase (LOX), extracellular matrix enzyme, is catalyzing lysine-derived crosslinks in collagen and elastin. Recently, several LOX-like proteins (LOXL, LOXL2, LOXL3 and LOXL4) have been identified in human but their specific functions are still largely unknown. The purpose of this study was to evaluate the function of the LOX family genes during odontoblastic differentiation of human dental pulp (HDP) cells induced with odontogenic supplement (OS). The messenger RNA (mRNA) expression of LOX family genes and differentiation markers was assessed by reverse transcriptase polymerase chain reaction analysis (RT-PCR). The formation of mineralization nodules was evaluated by alrizarin red S staining. Amine oxidase activity of HDP cells was measured by peroxidase-coupled fluormetric assay. The expressions of differentiation markers, such as alkaline phosphatase (ALP), osteopontin (OPN), osteocalcin (OCN), dentin matrix protein1 (DMP1), dentin sialophosphoprotein (DSPP) in HDP cells were increased after treatment with OS media. The LOX and LOXL mRNA expression were gradually increased in OS media, whereas LOX enzyme activities were markedly detected on day 7. The mRNA expression and LOX enzyme activity of collagen type I was very similar to the pattern of LOX gene. In this study, the expression of LOX and its isoforms, and activity of LOX were highly regulated during odontoblastic differentiation. Thus, these results suggest that LOX plays a key role in odontoblastic differentiation of HDP cells.

• Journal of the korean academy of Pediatric Dentistry
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• v.41 no.1
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• pp.1-7
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• 2014

If it is possible to preserve and culture cells from exfoliated deciduous teeth in a readily available storage medium within each family, more stem cells would be obtained. This research is about the effect of storage media and time on pulpal cell viability of exfoliated deciduous teeth. 330 exfoliated deciduous teeth were randomly divided into 11 groups; fresh group, dry group, groups stored in cell culture medium (2, 4, 7 days each), in milk (2, 4, 7 days each), and in saline (2, 4, 7 days each). Primary culture of pulpal cells was conducted in each group and the success rates were compared by calculating the number of teeth with viable cells. The result of primary culture shows that the success rate decreases as the time of storage gets longer. There was no statistical difference between groups stored in the cell culture medium, milk, and saline for 2 and 4 days. However, the groups stored in milk and saline for 7 days showed dramatic decrease in success rate compared to the group stored in the cell culture medium. In conclusion, exfoliated or extracted deciduous teeth can be used to culture pulpal cells when they are stored in milk and saline for a certain period of time; however obtaining viable pulpal cells becomes harder as the storage time gets longer.

• Restorative Dentistry and Endodontics
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• v.36 no.5
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• pp.397-408
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• 2011

Objectives: This study investigated changes in gene expressions concerning of differentiation, proliferation, mineralization and inflammation using Human-8 expression bead arrays when white Mineral Trioxide Aggregate and calcium hydroxide-containing cement were applied in vitro to human dental pulp cells (HDPCs). Materials and Methods: wMTA (white ProRoot MTA, Dentsply) and Dycal (Dentsply Caulk) in a Teflon tube (inner diameter 10 mm, height 1 mm) were applied to HDPCs. Empty tube-applied HDPCs were used as negative control. Total RNA was extracted at 3, 6, 9 and 24 hr after wMTA and Dycal application. The results of microarray were confirmed by reverse transcriptase polymerase chain reaction. Results: Out of the 24,546 genes, 43 genes (e.g., BMP2, FOSB, THBS1, EDN1, IL11, COL10A1, TUFT1, HMOX1) were up-regulated greater than two-fold and 25 genes (e.g., SMAD6, TIMP2, DCN, SOCS2, CEBPD, KIAA1199) were down-regulated below 50% by wMTA. Two hundred thirty nine genes (e.g., BMP2, BMP6, SMAD6, IL11, FOS, VEGFA, PlGF, HMOX1, SOCS2, CEBPD, KIAA1199) were up-regulated greater than two-fold and 358 genes (e.g., EDN1, FGF) were down-regulated below 50% by Dycal. Conclusions: Both wMTA and Dycal induced changes in gene expressions related with differentiation and proliferation of pulp cells. wMTA induced changes in gene expressions related with mineralization, and Dycal induced those related with angiogenesis. The genes related with inflammation were more expressed by Dycal than by wMTA. It was confirmed that both wMTA and Dycal were able to induce gene expression changes concerned with the pulp repair in different ways.

• Journal of the korean academy of Pediatric Dentistry
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• v.43 no.4
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• pp.419-426
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• 2016

For this research 20 supernumerary teeth impacted in the maxillary anterior have been extracted and pulp cells have been collected from them. From the collected pulp cells, total of 17 (10 males, 7 females) have been selected as subjects. From this research, the run-time of successive culture of the cell from tooth number pulp tissue was $2.91{\pm}0.29$ days. From the gathering of cells from the initial pulp tissue until gaining 80% confluency took $4.53{\pm}0.94$ which was the longest. The following successive cultures took $2.73{\pm}0.32$ days. Average runtime for female was $2.81{\pm}0.27$ days whereas male had average runtime of $2.98{\pm}0.29$ days. Average run-time for inversion was $2.94{\pm}0.30$ days and for normal location, $2.80{\pm}0.20$ days. Average runtime was $2.92{\pm}0.31$ days and other forms took $2.88{\pm}0.22$ days. In the future, follow up research would be needed to evaluate the efficiency of the cells collected from the initial passage and the latter passage as stem-cells and taking into consideration the less than 3 days'time for the subculture, it could be concluded that the research efficiency and fast cultivation would be sufficiently effective.