골재생유도술에 의한 골재생과정에서의 골기질 유전자 발현 양상

이창곤,류현모,신홍인,Lee, Chang-Kon,Ryoo, Hyun-Mo,Shin, Hong-In 대한악안면성형재건외과학회 1999 Maxillofacial Plastic Reconstructive Surgery Vol.21 No.3

골재생유도술에 의한 골재생 과정에서의 생물학적 현상을 보다 구체적으로 이해하고자 인위적으로 골결손부를 형성하고, 비흡수성 비공유성 차폐막을 이용하여 골성회복 시 주위 연조직의 유입을 차단한 다음 골수강 및 골면으로부터 유래되는 세포들에 의한 골성회복 양상과 이때 이들 세포의 조골세포로의 분화정도를 판정하기 위하여 비교원성 골기질 단백질인 OSN, OPN 그리고 OSC mRNA의 발현 양상을 비교 검토하여 다음과 같은 결과를 얻었다. 실험 전기간에 걸쳐 골재생유도술을 시행한 군에서 보다 신속하고 양호한 골성회복을 보였다. 차폐막을 처리한 실험군에서는 인접골의 주변부로부터 신생골이 형성되어 조골세포의 분화가 조기에 골결손부에 국한되어 유도된 반면, 대조군에서는 주변연조직의 개입으로 인하여 실험군 보다는 약 1주정도 신생골의 형성이 지연되었으며, 따라서 골수강 내의 기질세포의 조골세포로의 분화 역시 지연되었다. 이상의 사실에서 골창상부에서 차폐막에 의해 형성된 차폐공간은 기질 세포들의 보다 신속한 조골세포로의 분화 증식과 이들에 의해 신생된 골주들의 빠른 골 개조를 조장하였음을 시사한다. To investigate the expression pattern of noncollagenous bone matrix proteins such as osteonectin(OSN), osteopontin(OPN) and osteocalcin(OSC) mRNA during bony healing procedure induced by guided bone regeneration method, we made artificial defects on bilateral femur of rats. Then induced bony healing by application of a nonabsorbable PTFE membrane in experimental sites and without its application in control sites for 3 weeks. The mRNA expression pattern at specimens obtained at 1, 2 and 3 weeks after operation was detected by in situ hybridization method using its antisense mRNA probes. The experimental sites revealed more rapid and favorable bony healing than control sites and new bone formation was limited within defected area by inhibitory activity of bone marrow cells. In experimental sites, the OSN and OSC mRNA were expressed strongly on osteoblasts of regenerating cortical bone at 1st week and on osteoblasts lining the trabecular bone in marrow space at 3rd week, whereas, in control sites, their expression were noted on osteoblasts lining the reactively formed sponge bones at 2nd and 3rd week. In addition, the OPN mRNA was expressed on osteoblasts and osteoclasts at sites of remodeling and osteocytes of remained trabecular bone of defected area in experimental sites and on macrophages at 1st week and osteoclasts at sites of remolding at 2nd and 3rd week in control sites. The above findings suggest that the more rapid and favorable bony healing might be induced by blocking of invading fibrous connective tissue into bony defects. And the earlier expression of OSN and OSC mRNA on osteoblasts of experimental sites suggest that the formation and resorption of regenerating bone was more rapidly progressed in confined spaces made by applicate membranes.

PDGF와 IGF-I 병용 사용시 치주인대세포의 증식과 세포활성에 미치는 영향에 관한 연구

서조영,신홍인,경희문,Suh, Jo-Young,Shin, Hong-In,Kyung, Hee-Moon 대한치주과학회 1996 Journal of Periodontal & Implant Science Vol.26 No.2

Current acceptable methods for promoting periodontal regeneration are based on removal of diseased soft tissue. root treatment, guided tissue regeneration, introduction of new graft materials and biological mediators. Insulin-like growth factor-I(IGF-I) and Platelet-derived growth factor-BB(PDGF-BB), the members of the polypeptuyde growth factor family have been reported as the biological mediators which regulate a variety cellular matrix biologic activities of wound healing process including the cell proliferation, migration and extracellular matrix synthesis.The purposes of this study is to evaluate the combination effects of IGF-I and PDGF-BB on the cellular activity of the periodontal ligament cells to act as a regeneration promoting agent of periodontal tissue. Human periodontal ligament cells were prepared from the first premolar tooth extracted for the orthodontic treatment and were cultured in DMEM containing 10% FBS at the $37^{\circ}C$, 5% CO2 incubator. Author measured the DNA synthetic activity, and total protein, collagen and noncollagenous protein synthetic activities according to the concentration of 10,100ng/ml IGF-I and1,10 ng/ml PDGF-BB in combination. The results were as follows: Significantly increased in the 1 ng/ml PDGF-BB alone compared to the 10 ng/ml PDGF-BB alone(P<0.01) and in the 1 ng/ml PDGF-BB and 10, 100ng/ml IGF-I in combination compared to the 1 ng/ml PDGF-BB alone(P<0.05, P<0.0l). The synthetic activity of the total protein and collagen is significantly increased like to the synthetic activity of the DNA(P<0.05). The synthetic activity of the noncollagenous protein is increased according to the concentration of IGF_I, but not statistically statistically significant(P>0.05). The percent of the collagen is significantly in the 1ng/ml PDGF-BB and 10ng/ml IGF-I in combination compared to the 1ng/ml PDGF-BB alone(P<0.05) and in the 10ng/ml IGF-I in combination compared to the 10ng/ml PDGF-BB alone(P<0.05). The synthetic activity of the DNA is In conclusions, the percent study shows that PDGF-BB and IGF-I in combination have a potentiality to enhance the DNA synthesis and the total protein and collagen synthesis of The periodontal ligament cells, especially it is more significant in the low concentration of PDGF-BB compared to the high one. Thus, the PDGF-BB and IGF-I in combination may have important roles in promotion of periodontal litgment healing, and consequently, may useful for clinical application in periodontal regenerative procedures.

동종이식골의 탈회정도가 이소성 골형성유도에 미치는 영향

윤홍식,진병로,신홍인,Yun, Hong-Sik,Chin, Byung-Rho,Shin, Hong-In 대한악안면성형재건외과학회 1998 Maxillofacial Plastic Reconstructive Surgery Vol.20 No.2

동종 이식골 내의 탈회 정도에 따른 잔존 칼슘치가 이소성 골형성능에 미치는 영향을 파악하고자 Sprague-dowley계 백서에서 채취한 경골 및 대퇴골의 골간부를 0.5cm 크기로 절단하여 부착 연조직은 제거한 다음, 초음파 세척장치를 이용하여 $60^{\circ}C$ 0,6N HCl용액으로 5분, 10분, 15분, 20분, 25분, 30분, 35분, 40분간 각각 처리하여 탈회 동종 이식골을 준비하였다. 이때 탈회시간에 따른 시편의 무게를 측정하였고, 각 탈회용액으로부터 1cc를 취한 다음 Sigma사의 진단용 칼슘치 측정kit를 이용하여 spectrophotometer로 600nm 파장하에서 칼슘치를 측정하였다. 그리고 탈회 정도에 따른 이식골편의 골형성 유도능을 확인하기 위하여 24마리의 Sprague-dowley계 백서를 탈회시간별로 8군으로 나누어 배부에 0.5cm 크기로 4군데의 피하낭을 형성한후 각각의 처리된 탈회 이식골편을 이식하였다. 매식된 동종 이식골편들을 술후 1, 2, 3주째 채취하여 통법에 따라 H&E염색 표본을 제작하여 광학현미경으로 관찰하였다. 이상의 실험에서 다음과 같은 결과를 얻었다. 1. 탈회 30분까지의 용출되는 칼슘 농도 변화는 평균 15.91mg/ml로, 탈회 20분 이후에 평균 99.65%의 탈회정도를 보였다. 2. 동종골 무게 변화량은 탈회시작 25분까지 뚜렷한 무게변화를 보였으나, 그 이후에는 변화의 정도가 미약하였다. 3. 탈회된 이식 통종골에의한 이소성 골형성 유도능 비교에서 20분에서 30분간 탈회된 군에서 가장 양호하였고 그 외의 군에서는 그 정도가 열등하였다. 이상의 결과는 탈회된 동종골에 있어 골형성 유도능을 극대화하기 위해서는 골기질내 무기성분의 완전한 탈회가 선행되어야 하며, 탈회골 기질의 변성이 최소화되어야 함을 시사한다. This study has been performed to evaluate the relationship between the remained mineral components in a decalcified bone matrix and an ectopic bone formation efficiency. The freezed rat diaphyseal cortical bones measuring 0.5cm in length were demineralized in heated 0.6N HCl at $60^{\circ}C$ for 5, 10, 15, 20, 25, 30, 35, 40 minutes, respectively, using a controlled heat ultrasonic cleaner. Each 1cc of decalcifying solution taken during decalcification procedure was used to calculate calcium content using calcium dignostics kit under 600nm of spectrophotomer. After decalcification, each specimen was also weighed. Then each prepared specimen was implanted into the dorsal pouch of 24 Sprague-Dawley rats divided into 8 groups by time course. The implants were harvested at 1, 2, and 3 weeks and prepared for routine H-E stain specimens to evaluate osteogenic activity. The results are as follows: 1. There was statistical significant difference in change of calcium concentration up to demineralization of 30 minutes and each allogenic bones decalcifed up to 20 minutes revealed 99.65% of decalcification in average. 2. There was statistical significant difference in change of weight in demineralized allogenic bone up to 20 minutes treatment but, no significant change was noted after that time. 3. The histologic analysis revealed active ectopic bone formation in the implanted allografts demineralized for 20, 25, 30 minutes, respectively. However, the other groups of allografts showed relatively poor osteoinductive activity. These findings suggest that complete decalcification with a minimized degeneration of collagen matrix is necessary to induce maximal osteogenesis by decalcified bone allograft.

골재생유도술에 의한 골재생시 미세혈관 구축 양상

최두희,류현모,신홍인,Choi, Du-Hee,Ryoo, Hyun-Mo,Shin, Hong-In 대한악안면성형재건외과학회 1999 Maxillofacial Plastic Reconstructive Surgery Vol.21 No.3

골재생유도술에 의한 골재생과정의 생물학적 현상을 보다 구체적으로 이해하고자, 백서의 대퇴골에 인위적인 골결손부를 형성하고 비흡수성 차폐막을 설치한 다음 골재생유도 과정에서의 미세혈관의 구축 양상을 통상적인 광학 현미경적 소견 및 미세혈관주형 표본 관찰법을 중심으로 관찰한바 다음과 같은 결과를 얻었다. 광학 현미경적 소견의 초기 즉 술후 1주 및 2주 소견상 차폐막에 의해 피개된 실험부에서 보다 정연한 골성회복이 이루어졌으며, 이는 차폐막으로 인한 혈관망 형성의 양상에 의해 영향을 받는 것으로 나타났다. 즉, 차폐막에 의한 연조직 침입이 차단됨으로 인해 인접 골조직으로부터 수평적으로 들어온 혈관에 의해 규칙적인 혈관 분포를 나타내나, 대조군의 경우 연조직에서 유입된 혈관망에 의해 불규칙한 혈관망을 나타내었다. 시간이 경과되면서 재생된 결손부의 골은 재구성되면서 대조군과 실험군 사이에 골성회복의 양상은 유사하였으나, 인접 실험군의 골조직으로부터 유입된 혈관에 의해 형성된 규칙적인 혈관망이 연조직으로부터 침입한 혈관에 의해 그 규칙적인 배열이 흐트러진 대조군에 비하여 골성회복의 속도가 빨라진다는 사실을 확인하였다. 이상의 결과는 차폐막에 의한 골재생 유도과정에서 혈관의 유래와 혈관망의 정렬상태가 골성회복의 속도와 밀접한 관련이 있음을 보여주는 것이다. To investigate the sequential changes in microvascular architecture and osseous regeneration during the bony healing after an application of the guided tissue regeneration method, we made artificial defects measuring $0.7cm{\times}0.3cm$ in size on femoral bones of rats measuring about 200gm and applied non-absorbable TEFE membrane at experimental sites but not at control sites. Then we observed the sequential changes and correlations between new vacuolation and bony regeneration using microvascular corrosion cast method and routine light microscopic observation at 1, 2 and 3 weeks after operation, respectively. The results showed that there were close relationships between regeneration of microvasculature and bone. In early phase, the invasion of granulation tissue at control sites delayed bony regeneration, however, in later phase, there was no remarkable differences in bony regeneration between control and experimental sites. The placement of barrier also affected in revascularization of regenerating bony defects. This is, the experimental sites showed parallel arranged nutritional vessels along long axis with well developed retiform plexus whereas the control revealed vertical invasion of microvasculature from outside of marrow space through bony defects which was also rearrange with time into parallel pattern with a vertical plexus but lesser organized than that of experimental sites. These findings suggest that the reconstruction of regenerating vasculature within the marrow cavity only may be sufficient and/or more be efficient in regeneration of bony defects.

이중구조 합성 폴리머 차폐막에 위한 골성 회복능 평가

손정오 ( Jung Oh Sohn ),박의균 ( Eui Kyun Park ),이진호 ( Jin Ho Lee ),신홍인 ( Hong In Shin ) 한국조직공학·재생의학회 2012 조직공학과 재생의학 Vol.9 No.1s

Bone defects occur in a variety of clinical situation, and their reconstruction to provide mechanical integrity to the skeleton is a necessary step in the patient rehabilitation. Bones can regenerate themselves to repair defects up to a certain size but sometimes the suitable implant materials have to be applied to facilitate the bone repair. As a part of the effort to improve the efficiency of bone repair, we evaluated the hydrophilized asymmetric pore sized polydioxanone membrane for guided bone regeneration. The polydioxanone membrane was fabricated to have different pore size at inner (around 50 nm in diameter) and outer surface (around 50 μm in diameter) with an average 0.4 mm thickness. The cytotoxic effect of it was analyzed in vitro and the histocompatibility and guided bone regeneration effects were evaluated in vivo, respectively. The rat tibia bone defects measuring 7 mm×3 mm in size were treated with polydioxanone membrane with or without application of 10 mM LiCl or 100 μg/ml T-CAM. The defects were evaluated at 3weeks after treatment by radiography and histology. The polydioxanone membrane was relatively resilient with some cushion. It was no cytotoxic and evoked neither an immune nor an inflammatory response. It was gradually absorbed by numerous multinucleated giant cells with time and completely disappeared within 8 weeks at rat subcutaneous pouches. The application of polydioxanone membrane at rat tibia bone defects induced more effective bone repair with matured cortical plate regeneration compared to none membrane applied group. In addition, the treatment of 10 mM LiCl and 100 μg/ml T-CAM within polydioxanone membrane facilitated bony healing. These results suggest that the combined application of bioactive molecules such as 10 mM LiCl or 100 μg/ml T-CAM with hydrophilized sized polydioxanone membrane can facilitate the guided bone regeneration.

여러 종류의 성장인자와 덱사메타손이 골수간엽줄기세포의 증식에 미치는 효과

손민정 ( Min Jung Shon ),이선영 ( Sun Young Lee ),김태호 ( Tae Ho Kim ),김석영 ( Suk Young Kim ),강길선 ( Gil Son Khang ),손영숙 ( Young Sook Son ),신홍인 ( Hong In Shin ),김신윤 ( Shin Yoon Kim ),박의균 ( Eui Kyun Park ) 한국조직공학과 재생의학회 2006 조직공학과 재생의학 Vol.3 No.4

Bone marrow stromal cells(BMSCs) are pluripotent cells capable of differentiating into several types of cells and are thus an attractive cell source for connective tissue engineering. For use of BMSCs in there applications, ex vivo expansion is necessary to obtain sufficient numbers of cells. Various growth factors regulate the recruitment of progenitor cells and their proliferation and differentiation into matured cells. In this study we have investigated the effects of different growth factors and in combination with dexamethasone(Dex) on the proliferation of BMSCs. We found that at a low concentration of growth factors(1 ng/mL) including fibroblast growth factor(FGF)-2, epidermal growth factor(EGF), hepatocyte growth factor(HGF), transforming growth factor-ß(TGF-ß), vescular endothelial growth factor(VEGF) and platelet derived growth factor-BB(PDGF-BB), FGF-2 was most effective in stimulation of proliferation in BMSCs in vitro. In addition, when FGF-2, PDGF and EGF were combined with Dex, proliferation of BMSCs was further stimulated. Among the growth factors, it was shown that FGF-2 in combination with Dex showed most prominent effect on stimulation of BMSCs. These results demonstrate that a low concentration of FGF-2 incombination with Dex may be useful stimulants for ex vivo expansion of BMSCs.

BMP-교원질 섬유막 복합체에 의한 이소성 골형성

신홍인,서조영,Shin, Hong-In,Suh, Jo-Young 대한치주과학회 1996 Journal of Periodontal & Implant Science Vol.26 No.1

To investigate the efficiency of a fibrous collagen membrane(FCM) composed of bovine skin type I atelocollagen as a carrier for BMP, partially purified bovine BMP/FCM($0.3mg/10{\times}5{\times}1mm$) composites were implanted into the dorsal subcutaneous tissue of rats. FCM alone was also implanted as a control. The implants were harvested at 1, 2, 3, and 10 weeks after implantation, then prepared for routine light microscopic observation. The FCM alone did not induce osteogenesis and revealed no specific foreign body reaction nor was there any definite resorptive evidence for 10 weeks after implantation, while the BMP/FCM composites induced favorable bone formation in a process that resembled an endochondral and direct ossification mode. At 10 weeks, the well formed bone confined to embedded collagen fibers revealed hematopoietic marrow between bony trabeculae without evidence of resorptive or degenerative changes . These findings support the suggestion that BMP may induce undifferentiated mesenchymal cells into either chondroblasts or osteoblasts or both independantly according to the chemico- physical characteristics of the carrier, which develops the endochondral and/or direct bone formation process, and suggest that the FCM may be a favorable carrier for BMP.

mRNA Expression of Cytokines and Release of Metalloproteinases around Loose Cemented Total Hip Arthroplasty

Shin-Yoon Kim(김신윤),Hee-Soo Kyung(경희수),Hong-In Shin(신홍인),Jae-Yong Choi(최제용),Moon-Kyu Kim(김문규),Jung-Chul Kim(김정철) 대한정형외과학회 1998 대한정형외과학회지 Vol.33 No.6

Interrelationship of Matrix Metalloproteinase and $TNF-{\alpha}$ in Human Gingiva with Chronic Periodontitis associated to Type 2 Diabetes Mellitus

김도훈,박의균,신홍인,조제열,서조영,이재목,Kim, Doe-Heun,Park, Eei-Kyun,Shin, Hong-In,Cho, Je-Yeol,Suh, Jo-Young,Lee, Jae-Mok The Korean Academy of Periodontoloy 2006 Journal of Periodontal & Implant Science Vol.36 No.2

치주질환의 병원균은 세포벽의 항원에 의하여 조직내 존재하는 mononuclear phagocytes가 활성화되어 cytokine들이 생성됨 으로써 치주 결체조직의 파괴를 진행시킨다. 이런 관련된 cytokine들은 순차적으로 상주하는 치은세포 및 대식세포가 Matrix metalloproteinase 합성을 하도록 유도하여 조직파괴를 시작한다. 이들 Matrix metalloproteinase중 MMP-2, MMP-9 (Gelatinase A,B)는 type IV collagen 및 변성된 interstitial collagen을 파괴하며 치주환자의 치은 열구액, 치은조직, 타액 네에서 높게 보고 되어왔다. 당뇨병은 치주질환의 위험요소중 하나로 달뇨 환자에서는 치주질환의 유병율이 일반인에 비해 높고 치주질환의 중증도도 더 심하여 진행도 빠르다고 알려져 있다. 그 병리 기전 중 하나로는 당뇨 환자에서는 치은 열구액 내 중성구 유래의 Matrix metalloproteinase의 활성 증가 및$TNF-{\alpha}$ 의 활성 증가가 추정되고 있다. 본 실험에서는 제2형 당뇨병 환자와 비당뇨 환자들에서 만성 치주염 부위의 치은 및 건강한 치은에서 염증매개체 중 하나인 MMP-2, MMP-9 및 $TNF-{\alpha}$ 의 발현에 대해 상호 비교 분석함으로서 염증, 혈당이 미치는 영향을 밝히고 제 2형 당뇨병 환자에서 심한 치주조직 파괴의 기전을 연구하고자 하였다. 경북대학교병원 치주과 내원환자 중 제2형 당뇨병 환자와 비당뇨 환자들 및 치주질환이 없는 건강인 대조군을 대상으로 여러 가지 환자요소, 임상 치주상태를 기록하고, 전신적으로 건강한 환자의 건강한 부위(n=8,Group 1), 전신적으로 건강한 환자으 만성 치주염 부위(n=8, Group 2), 제2형 당뇨병 환자의 만성 치주염 부위 (n=8,Group 3)에서 각각 변연치은을 채득하고 액화질소에 급속 동결하였다. Western blotting을 이용하여 각 조직 내 MMP-2, MMP-9 및 $TNF-{\alpha}$ 의 발현을 관찰, densitometer를 이용하여 상대적 발현을 정량, 각 조직의${\beta}-actin$을 이용하여 표준화하여 실험군과 대조군들의 평균치를 비교하였다. 비당뇨 환자들의 만성 치주염 부위 및 제2형 당뇨병 환자의 만성 치주염 부위에서 모두 건강 대조군에 비해 MMP-2와 MMP-9 의 발현이 증가되었다. 또한 MMP-2와 MMP-9는 2형 당뇨 환자의 만성 치주염 부위가 비당뇨 환자의 만성 치주염 부위보다 증가된 발현양상을 보였으며, $TNF-{\alpha}$ 발현 비교시 각 군간 유의성 있는 변화는 없었으나 2형 당뇨환자군에서 MMP-2 및 MMP-9의 증가와 함께 다소 증가 양상을 보였다. 결론적으로 본 실험에서 MMP-2 및 MMP-9의 증가가 만성 치주염 및 2형 당뇨 환자에서의 만성치주염에서 비당뇨환자 보다 MMP-2, MMP-9의 증가양상을 보여 주었으며 $TNF-{\alpha}$ 가 2형 당뇨환자의 만성치주염 진행과정에 기여인자로써 역할을 하는 것으로 생각된다.

Desflurane 마취에서 근이완제를 투여하지 않은 기관내삽관을 위한 Remifentanil의 효과처농도와 저혈압 유발

인준용 ( Jun Yong In ),신홍일 ( Hong Il Shin ),정승현 ( Seung Hyun Chung ),김경옥 ( Kyoung Ok Kim ),최준권 ( Jun Gwon Choi ),이윤석 ( Youn Suk Lee ),조헌 ( Hun Cho ) 대한마취과학회 2008 Korean Journal of Anesthesiology Vol.55 No.1