인체 구강상피에서 Langerhans cell 분포에 관한 면역조직화학적 연구

김흥중,박주철,조재오 朝鮮大學校 口腔生物學硏究所 1994 口腔生物學硏究 Vol.18 No.2

28. Human Gingiva에서의 Langerhans Cell 분포와 형태에 관한 면역조직화학적 연구

김흥중,조재오,장인엽 대한체질인류학회 1990 대한체질인류학회 학술대회 연제 초록 Vol.33 No.-

RNA interference를 이용한 OD314 유전자의 발현억제가 상아질모세포 전구세포에 미치는 영향

김흥중,정문지,손호현,박주철 대한체질인류학회 2004 해부·생물인류학 (Anat Biol Anthropol) Vol.17 No.2

Tooth development depends on reciprocal interactions between oral epithelium and ectomesenchyme. The ectomesenchyme-derived odontoblasts secrete several collagenous and non-collagenous proteins to form a unique extracellular matrix. OD314 was obtained by subtractive hybridization between odontoblasts and osteoblast/pulp cells, and differentially or predominantly expressed in odontoblasts of rat incisors compared to osteoblasts and pulp cells. However, little is known about the function of OD314 in odontoblast differentiation. In this study, to better understand the function of OD314, we inactivated the OD314 gene in mouse MDPC23 cells using U6 promoter-driven siRNA method. After the characterization of mineralized nodule formation and molecular expression of MDPC23 cell, Inactivation effects of the OD314 were evaluated by RT-PCR and western blot. 1. Mineralized nodule formation was observed after 14 days of culture in MDPC23 cells. 2. The OD314, OC and DSPP mRNA were highly expressed throughout the cultures, while the expression of ON decreased as the MDPC23 cells differentiated. 3. The OD314 protein was weakly expressed at 7 days of culture, but increased gradually as MDPC23 cells reached mineralization stage. 4. The inactivation of OD314 by RNA interference downregulated the expressions of OD314, DSPP, OC, and ON mRNA and OD314 protein in MDPC23 cells. These results suggest that OD314 may play a important role in mineralization process of odontoblast and also regulate odontoblast-related genes such as OC, ON, and DSPP. 상아질모세포는 신경능선세포에서 기원하며 상아질의 유기질을 형성하는데 상아질모세포의 분화과정이 나 상아질의 형성과정과 관련한 분자생물학적 기전은 아직 명확히 밝혀져 있지 않다. OD314는 흰쥐의 치아발생 과정과 세포배양실험에서 상아질 형성과정 특히 석회화결절이 형성되는 시기에 그 발현이 현저하며 상아질의 석 회화과정에 중요한 기능을 암시한다. 본 연구에서는 OD314 유전자의 발현을 RNA interference를 이용하여 억제하 여 상아질모세포내 다양한 유전자들의 단백질들을 평가함으로서 상아질의 형성과정에서 OD314의 기능을 알아보 고자 하였다. 1. MDPC23 세포에 ascorbic acid와 β-glycerophosphate 첨가하여 석회화 결절 형성을 유도한 실험에서 배양 후 14 일에 석회화 결절이 관찰되었다. 2. MDPC23 세포의 석회화 결절 형성과정에서 OD314, OC 및 DSPP mRNA는 배양초기부터 석회화 결절이 형성 된 14일에도 동일한 발현을 보인 반면, ON mRNA는 석회화 결절이 형성되기 시작하는 7일부터 발현이 감소되 었다. 3. MDPC23 세포의 배양과정에서 OD314 단백질은 17 kDa 정도의 크기로 배양 시작부터 7일까지 동일한 발현을 보였으며 석회화 결절이 형성된 14일에는 더욱 강한 발현을 보였다. 4. MDPC23 세포에 OD314 siRNA를 transfection한 실험에서 OD314, OC, ON 및 DSPP의 mRNA의 발현이 감소 하였고 OD314의 단백질 발현 또한 OD314 siRNA를 transfection한 세포에서 감소하였다. 이상의 결과를 종합하면 OD314가 상아질모세포의 주요 유전자들의 조절에 관여할 것으로 사료되나 이를 명확히 하기 위하여 앞으로 OD314에 대한 기능과 상아질모세포 관련인자들과의 상호연관성에 대한 향후 보완 연구가 필요할 것으로 사료된다.

공초점레이저 주사현미경의 원리와 생물학적 응용

김흥중,김현섭 朝鮮大學校 口腔生物學硏究所 2000 Oral Biology Research (Oral Biol Res) Vol.24 No.1

Conventional light microscope have been used to study cellular structure, function and regulation. However, the conventional microscope has a limitation that out-of-focus information interferes in-focus image. A solution for this problem is confocal laser scanning microscope(CLSM). Compared to conventional microscope, confocal laser scanning microscope increased contrast and resolution of specimen. The name of confocal laser scanning microscope was derived from the principles of the microscope; simultaneous focusing(confocal), using laser(laser) and scanning a sample(scanning). Confocal laser scanning microscope has several advantages over conventional microscope. The first advantage of CLSM is 3-D image construction. 3-D formation can be constructed form serial sectioned images. And individual images saved in computer can be processed into various results such as 3-D structure, stereo image, color depth coding and fluorescence intensity. Second, another advantage of CLSM is optical sectioning. Optical sectioning allows us to analyzing real-time changes of living cells such as Ca^+, cAMP and H_2O_2.

흰쥐에서 치아손상 후 삼차신경절에서 vasoactive intestinal polypeptide 면역반응세포에 관한 연구

김흥중,박주철,김두현,문주훈 朝鮮大學校 口腔生物學硏究所 1999 Oral Biology Research (Oral Biol Res) Vol.23 No.2

The effect of tooth injury on the expression of vasoactive intestinal polypeptide(VIP) in trigeminal ganglion was examined immunohistochemically in rat. The animals were divided into normal and 2 weeks experimental groups. The trigerminal granglion was removed and immersed in the 4% paraformaldehyde in 0.1M phosphate buffer. Serial forzon sections were gained with a cryostat. The rabbit anti-VIP was used as primary antibody and fluorescene isothiocynate(FITC)- conjugated anti-rabbit IgG as secondary antibody in immunofluorescence staining. The stained slides were observed under confocal laser scanning microscope. The results were as follows; 1. The positive ratio of VIP immunoreactive neurons; The positive ratio of VIP immunoreactive neurons in the mandibular part of trigeminal ganglion was 6.52±2.31% in normal group and 24.73±7.27 in 2 weeks after tooth injury group. 2. The relative fluorescence intensity of VIP immunoreactive neurons; The relative fluorescence intensity of VIP immunoreactive neurons in the mandibular part of trigeminal ganglion was 96.92±14.95 in normal group and 165.37±25.73 in 2 weeks after tooth injury group. The relative fluorescence intensity of w weeks experimental group was increased 68.5,% than intensity of normal group. 3. These results indicate that immunoreactivity and fluorescence intensity of VIP immunoreactive neurons were increased in trigeminal ganglion following tooth injury.

부도를 성공의 거름으로

김흥중 샘터사 1997 월간 샘터 Vol.28 No.6

치주조직의 발생과 분화과정에서 치주인대-특이 유전자 PDLs22의 발현

김흥중,정문진,김병옥,국중기,김종관,박주철 대한구강악안면병리학회 2005 대한구강악안면병리학회지 Vol.29 No.1

Periodontalligament (PDL) fibroblasts have an ectomesenchymal origin and are known to participate not only in formation of PDL but also in the repair and regeneration of the a이acent alveolar bone and cementum. However, little is known about the molecular mechanism which is related to the development and differentiation of PDL cells. Recendy, we reported the PDLs (a periodontalligament-specific) 22 as a PDL fibroblast-specific mRNA which is not expressed in gingival fibroblasts. In this study, to examine the expression and functional characterization of PDμ22 mRNA and prαein in development and differentiation of periodontal 따sue , we carried out northem analysis, insitu hybridization, immunofluorescence and immunohistochemistry. The expression of PDLs22 mRNA was increased with PDL cell differentiation from the confluent to multilayer stage but decreased slighdy with mineralized nodule formation in vitro. πle PDLs22 protein was localized on the nuclear membrane and expressed throughout the differentiation of PDL fibroblasts in vitro. The PDLs22 mRNA and protein were expressed in the differentiating cementoblasts, PDL fibroblasts and osteoblasts along the r∞t surface and alveolar bone of the developing rat teeth. These results indicate that the PDLs22 plays an irnportant role in the differentiation of cementoblasts and osteoblasts and thus homeostasis of cementum, PDL and alveolar bone.

Cortical Bone Thickness for Mini-implant Placement in Korean

김흥중 KOREAN ACADAMY OF ORAL BIOLOGY 2011 International Journal of Oral Biology Vol.36 No.2

Recently, mini-implant is popular in the orthodontic treatment due to its simplicity and convenient surgical procedure. The objective of this study is to provide the anatomical guideline for mini-implant placement by analysing the cortical bone thickness in Korean. Hemi-sections of sixteen maxillae and twenty-two mandibles with normal teeth were used. Interdental areas between the 1st premolar and the 2nd premolar (Group 1), the 2nd premolar and the 1st molar (Gruop 2), and the 1st molar and the 2nd molar (Group 3) were sectioned and then scanned. After setting the axis of teeth, the cortical bone thickness was measured at the distance of 2 mm, 4mm, 6 mm, and 8 mm from alveolar crest. The mean thickness of cortical bone in the maxilla according to distance from alveolar crest was 1.30 ± 0.63 mm (2 mm), 1.49 ± 0.62 mm (4mm), 1.72 ± 0.64 mm (6mm), and 1.90 ± 0.90 mm (8 mm) at the buccal side and 1.33 ± 0.47 mm, 1.31 ± 0.45 mm, 1.37 ± 0.55 mm, and 1.39 ± 0.58 mm at the palatal side. In the mandible, that was 3.14 ± 1.71 mm, 4.31 ± 2.22 mm, 4.23 ± 1.94 mm, and 4.30 ± 1.57 mm at the buccal side and 1.98 ± 0.88 mm, 2.79 ± 1.01 mm, 3.35 ± 1.27 mm, and 3.93 ± 1.38 mm at the lingual side. The buccal cortical bone thickness in the maxilla was decreased from Group 1 to Group 3, while the thickness of palatal side was no change. In the mandible, it did not show a tendency at the buccal side and it was decreased from Group 1 to Group 3 without significant difference at the lingual side. Therefore, the buccal side of the Group 1 and Group 2 in both the maxilla and mandible seems to be the most appropriate site for a mini-implant placement with taking the stability and retention.

치수제거후 흰쥐 삼차신경절에서 VIP 면역반응세포의 변화: 공초점레이저주사현미경적 연구

김흥중,김승재,박주철,박주철,이상호 대한소아치과학회 2001 大韓小兒齒科學會誌 Vol.28 No.1

말초신경 손상에 의한 VIP의 변화를 연구하기 위해 흰쥐 하악대구치 치수제거 후 삼차신경 절에서 VIP의 분포 및 반응강도를 공초점레이저주사현미경을 이용하여 관찰하였다. 체중 200g내외의 Sprague-Dawley계 흰쥐를 대조군과 하악대구치 치수제거 후 14일군으로 분리하여 희생시켰다. 1차 항체로 rabbit anti-VIP, 2차 항체로 fluorescein isothiocyanate(FITC) conjugated anti-rabbit IgG를 사용하여 면역형광염색을 시행한 후 공초점레이저주사현미경으로 관찰하여 다음과 같은 결론을 얻었다. 1.삼차신경절 하악부위에서 VIP 양성반응세포의 비율은 대조군에서 7.40%를, 실험군에서는 28.42%를 보였다. 대조군에 비해 실험군에서 양성반응세포의 증가를 보였다. 2.삼차신경절 하악부위에서 VIP 면역반응세포체에 대한 상대성 형광강도는 대조군에서 87.78을, 실험군에서는 138.65를 보였다. 대조군과 비교하였을 때 실험군에서 상대성 형광강도의 증가를 보였다. 3.실험군의 광연속절편(1㎛) 관찰에서 VIP 면역반응세포는 9개의 절편 대부분에서 강하게 나타났다. 축삭의 면역반응을 살펴보면, 대조군의 축삭에서는 약한 반응을 보였으며, 실험군의 축삭에서는 강한 면역반응을 보였다. 또한 양성반응 세포체의 크기는 20∼25㎛의 중간 크기의 세포체에서 강한 면역반응을 보였다. 위의 결과로 보아 치수제거 후에 삼차신경절 하악부위에서 VIP면역반응세포의 증가와 함께 상대성 형광강도가 높아졌음을 알 수 있었다. The purpose of this study was to investigate the distribution and fluorescence intensity of vasoactive intestinal polypeptide immunoreactive(VIP-IR) cells in rat trigeminal ganglion following pulp extirpation of rat mandibular molar. The animals were divided into control group(n=6) and experimental group(n=6). The experimental animals were sacrificed at 14 days after pulp extirpation. The trigeminal ganglion was removed and immersed in the 4% paraformaldehyde in 0.1M phosphate buffer. Serial frozen sections about 20㎛ in thickness were cut with a cryostat The immunofluorescence staining was performed. The rabbit anti-VIP(1:8,000) was used as primary antibody and fluorescene isothiocynate(FITC) cojugated anti-rabbit IgG(1:80) as secondary antibody. The slides were observed under confocal laser scanning microscope(CLSM). Unprocessed optical sections were obtained and stored on a optical disk. Color pictures were printed by a video copy processor. The results were as follows ; 1. The positive ratio of VIP-IR cells in mandibular part of trigeminal ganglion were 7.40% in control group and 28.42% in experimental group(14 days affter pulp extirpation). 2. The relative fluorescence intensity of VIP-IR cells in mandibular part of trigeminal ganglion were 87.78 in control group and 138.65 in experimental group. The relative fluorescence intensity of experimental group was 58% higher than that of control group. 3. In optical serial section analysis of VIP-IR cells of experimental group, most of the 9 section showed high fluorescence intensity. At high magnification, axons of the experimental group displayed greater VIP-IR than in the control group, and the positive cells were mainly of medium size. The result indicate that number and fluorescence intensity of VTP-IR cells were increased in the mandibular part of trigeminal ganglion following pulp extirpation of mandibular molar, and it suggests that VIP could play a role in processing of nociception.

흰쥐에서 치수노출 후 삼차신경절의 신경절아교세포에서 GFAP-IR의 변화

김흥중,문주훈 大韓齒科保存學會 1997 Restorative Dentistry & Endodontics Vol.22 No.2

Glial fibrillary acidic protein(GFAP) are a group of intermediate filaments that are distributed in the cytoplasm of many type of glial cells. The purpose of this study was to determine change of GFAP immunoreactivity (GFAP-IR) in rat trigeminal ganglion satellite cells in response to pulp exposure. The immunohistochemistry was carried out using the avidinbiotin-peroxidase complex(ABC) method and subsequently stained with AEC(3-aminoethyl-9-carbasol). 1. Contol group; Central root astrocytes had strong GFAP-IR, but ganglion satellite cells occasionlly had GFAP-IR. This reaction patterns of ganglion satellite cells was not concenturated in any specific region of trigeminal ganglion. 2. Three day pulp exposure group' There was a highly GFAP-IR in satellite cells of trigeminal ganglion in maxillary region. GFAP-IR in neighboring mandibular and ophthalmic regions was less intense compared to maxillary region. 3. Seven day pulp exposure group; In this group, GFA-IR that was increased compared to control group was seen in the maxillary region. But GFAP-IR was less intense compared to three day pulp exposure group. These results suggest that GFAP in satellite cell increase in specific region of trigeminal ganglion after pulp exposure and offer useful tool in trigeminal pain research.