• 폴리머
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• 제27권3호
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• pp.226-234
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• 2003

이프리플라본은 이소플라본의 파생물로서 골의 재흡수를 방지하여 골의 재형성을 방해함으로써 골 형성에 도움을 준다. 이프리플라본은 칼슘의 양을 안정적으로 증가시킴과 함께 골수 줄기 세포의 작용으로 세포층에 칼슘을 침착시킨다. 조직공학적 골을 형성시키기 위해 락타이드-글리를라이드 공중합체 (PLGA)에 이프리플라본을 함유시킨 지치체를 용매 캐스팅/염 추출법으로 제조하였다. 수은 다공도계, 주사 전자 현미경, 시차 주사 열량계, X선 회절기를 이용하여 특성결정을 수행하였고, 이프리플라본이 함유된 지지체와 이프리플라본이 함유되지 않은 지지체를 면역이 결핍된 쥐의 피하에 삽입하여 이들의 골 형성을 비교하였다. 조직을 hematoxylin & eosin, 본쿠사 염색과 면역화학적 염색법인 콜라겐 I 형과 오스테오칼신 염색을 하였다. 이프리플라본이 함유된 담체의 다공도는 91.7% 이상이었고, 평균 다공 크기도 101 $\mu\textrm{m}$였다. PLGA로만 제조된 지지체와 이프리플라본을 50% 함유시킨 지지체를 동물 실험을 수행한 결과 이프리플라본은 피하 층과 다른 연조직에서 미분화 줄기 세포가 칼슘 침착, 골아 세포, 골상으로의 유도에 더 많은 영향을 주는 것을 관찰하였다. 결론적으로 이프리플라본을 함유한 지지체에서 이프리플라본이 골형성에 중요한 요인으로 작용한다고 사료된다.

• Journal of Periodontal and Implant Science
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• 제34권2호
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• pp.269-279
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• 2004

백악질 세포의 분리 및 배양방법을 확립하고, 이를 이용하여 백악질 세포의 형질특성을 알아보고자 하였다. 교정목적으로 발거된 소구치를 이용하여, 치은섬유아세포, 치주인대 세포 및 백악질 유래세포를 분리, 배양하였다. 백악질 유래 세포 배양시에는 백악질을 절제한 후 Collagenase P를 이용하여 백악질 유래 세포 외의 다른 세포의 개제를 배제하였고, 기질을 분해하여 세포의 분리 및 배양이 용이하도록 하였다. 분리 및 배양시기의 세포의 형태를 광화현미정을 이용하여 관찰하였다. 조골세포의 특성을 가지는 SaOs-2 세포를 대조군으로 이용하여 분리 및 배양된 세포군들을 동일한 조건으로 배양하였다. 3일 및 7일째에 세포증식도를 측정하였고 7일째에 ALPase 효소 활성도를 측정하였다. 각 세포의 형질 특성을 알아보기 위해 RT-PCR을 실시하여 조골세포 분화 표식자와 연관된 osteopontin(OPN), Alkaline phosphatase(ALP), type I collagen(COL-I), Bone sialoprotein(BSP), BMP-2 및 osteocalcin(OC)의 발현을 비교 관찰하였다. 백악질 유래 세포의 분리 및 배양을 시도한 5명의 치아 중에서 3명의 치아에서 세포군을 배양해 낼 수 있었다. 배양한 백악질 유래 세포는 섬유아세포와 유사한 형태와 증식을 보였다. ALPase 효소 활성도 검사 결과 백악질 유래 세포는 SaOs-2 세포보다 낮은 활성도를 나타내었으며, 배양된 세포의 RT-PCR 결과 백악질 유래 세포군에서는 ALPase의 발현이 나타나지 않았고, 다른 조골세포 표식자의 발현도 낮게 나타났다. 이는 백악질 유래 세포가 조골세포 및 다른 대조군의 세포와는 다른 형질 특성을 가지고 있다는 것을 시사한다. 이상의 관찰결과로 사람의 백악질 유래 세포롤 백악질의 절제 및 효소처리 방법으로 효과적인 분리 및 배양이 가능하며, 이는 향후 백악질 세포의 형질 특성 및 백악질 형성의 분자적 기전을 파악하는 중요한 연구자료로 활용 될 수 있을 것으로 사료된다.

• Maxillofacial Plastic and Reconstructive Surgery
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• 제27권1호
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• pp.1-8
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• 2005

This study was aimed to characterize osteogenic potential of rat bone marrow stromal cells (BMSC) isolated with standard flushing method and investigate the plasticity of transdifferentiation between osteoblastic and adipocytic lineage of cultured BMSC. Unlike aspiration method in human, rat bone marrow was extracted by means of irrigation with culture media that elevates the possibility of co-extraction of committed osteoprogenitor, or preosteoblast or other progenitor cells of several types present inside bone marrow. The cultured stromal cells showed high ALP activity which is representative marker of osteoblast without any treatment. Osteogenic inducers such as Dex and BMP-2 were examined for the evaluation of their effect on osteogenic and adipocytic differentiation of stromal cells, because they function as osteoinductive agent in stromal cells, but simultaneously induce adipogenic differentiation. Osteogenic differentiation was evaluated by measuring alkaline phosphatase activity or mRNA expression of osteoblast markers such as osteopontin, bone sialoprotein, collagen type I and CbfaI, and in vitro matrix mineralization by von Kossa staining. Oil red staining method was used to detect adipocyte and adipocytic marker, aP2 and $PPAR{\gamma}2$ expression was examined using RT-PCR. It can be supposed that irrigation procedure resulted in high portion of already differentiation-committed osteoprogenitor cell showing elevated ALP activity and strong mineralization only under the supplement of $100{\mu}M$ ascorbic 2-phosphate and 10mM ${\beta}$-glycerophosphate without any treatment of osteogenic inducers such as Dex and BMP-2. Dex and BMP-2 seemed to transdifferentiate osteoprogenitor cells having high ALP activity into adipocytes temporarily, but continuous treatment redifferentiated into osteoblast and developed in vitro matrix mineralization. This property must be considered either in tissue engineering for bone regeneration, or in research of characterization of osteogenic differentiation, with rat BMSC isolated by the standard irrigation method.

• Journal of Periodontal and Implant Science
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• 제36권2호
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• pp.385-396
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• 2006

For the regeneration of periodontal tissues, the microenvironment for new attachment of connective tissue fibers should be provided, At this point of view, cementum formation in root surface plays a key role for this new attachment. This study was performed to figure out which factor promotes differentiation of cementoblast Considering anatomical structure of tooth, we selected the cells which may affect the differentiation of cementoblast - Ameloblast, OD11&MDPC23 for odontoblasts, NIH3T3 for fibroblsts and MG63 for osteoblasts. And OCCM30 was selected for cementoblast cell line. Then, the cell lines were cultured respectively and transferred the conditioned media to OCCM30. To evaluate the result, Alizarin red S stain was proceeded for evaluation of mineralization. The subjected mRNA genes are bone sialoprotein(BSP), alkaline phosphate(ALP) , osteocalcin(OC), type I collagen(Col I), osteonectin(SPARC ; secreted protein acidic and rich in cysteine). Expression of the gene were analysed by RT-PCR, The results were as follows: 1. For alizarin red S staining, control OCCM30 didn't show any mineralized red nodules until 14 days. But red nodules started to appear from about 4 days in MDPC-OCCM30 & OD11-OCCM30. 2. For results of RT-PCR, ESP mRNAs of control-OCCM30 and others were expressed from 14 days, but in MDPC23-OCCM30 & OD11-OCCM30 from 4 days. Like this, the gene expression of MDPC23-OCCM30 & OD11-OCCM30 were detected much earlier than others. 3. For confirmation of odontoblast effect on cementoblast, conditioned media of osteoblasts(MG63) which is mineralized by producing matrix vesicles didn't affect on the mineralized nodule formation of cementoblasts(OCCM30). This suggest the possibility that cementoblast mineralization is regulated by specific factor in dentin matrix protein rather than matrix vesicles. Therefore, we proved that the dentin/odontoblast promotes differentiation/mineralization of cementoblasts. This new approach might hole promise as diverse possibilities for the regeneration of tissues after periodontal disease.

• Molecules and Cells
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• 제42권11호
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• pp.763-772
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• 2019

Periodontitis is characterized by the loss of periodontal tissues, especially alveolar bone. Common therapies cannot satisfactorily recover lost alveolar bone. Periodontal ligament stem cells (PDLSCs) possess the capacity of self-renewal and multilineage differentiation and are likely to recover lost alveolar bone. In addition, periodontitis is accompanied by hypoxia, and hypoxia-inducible $factor-1{\alpha}$ ($HIF-1{\alpha}$) is a master transcription factor in the response to hypoxia. Thus, we aimed to ascertain how hypoxia affects runt-related transcription factor 2 (RUNX2), a key osteogenic marker, in the osteogenesis of PDLSCs. In this study, we found that hypoxia enhanced the protein expression of $HIF-1{\alpha}$, vascular endothelial growth factor (VEGF), and RUNX2 ex vivo and in situ. VEGF is a target gene of $HIF-1{\alpha}$, and the increased expression of VEGF and RUNX2 proteins was enhanced by cobalt chloride ($CoCl_2$, $100{\mu}mol/L$), an agonist of $HIF-1{\alpha}$, and suppressed by 3-(5'-hydroxymethyl-2'-furyl)-1-benzyl indazole (YC-1, $10{\mu}mol/L$), an antagonist of $HIF-1{\alpha}$. In addition, VEGF could regulate the expression of RUNX2, as RUNX2 expression was enhanced by human VEGF ($hVEGF_{165}$) and suppressed by VEGF siRNA. In addition, knocking down VEGF could decrease the expression of osteogenesis-related genes, i.e., RUNX2, alkaline phosphatase (ALP), and type I collagen (COL1), and hypoxia could enhance the expression of ALP, COL1, and osteocalcin (OCN) in the early stage of osteogenesis of PDLSCs. Taken together, our results showed that hypoxia could mediate the expression of RUNX2 in PDLSCs via $HIF-1{\alpha}$-induced VEGF and play a positive role in the early stage of osteogenesis of PDLSCs.

• 동의생리병리학회지
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• 제33권2호
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• pp.102-108
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• 2019

This study aims to evaluate the anti-inflammatory effect of Coptidis Rhizoma (CR) extract for atopic dermatitis through maintaining skin barrier and regulating Th2 cell differentiation. We divided NC/Nga mice into 3 groups as follows; atopy-like dermatitis induced group with CR treatment (CT, n=10), no treatment group(Ctrl), atopy-like dermatitis elicited group(AE). Atopy-like dermatitis was induced to NC/Nga mice by sensitizing with dermatophagoides farinae(DfE) on 7, 8, 9, 11, 12, and 13th week. After inducing atopic dermatitis, CR extract was administered 20 mg/kg daily for the experimental duration to the CT group. We measured the integrity of lipid layers in the epidermis and Th2 differentiation through immunohistochemical staining against filaggrin, loricrin, IL-4, and IL-13. We also measured the distribution of subcutaneous collagen fibers by the Masson's trichrome staining. Administration of CR significantly inhibited the reduction of lipid layers in the skin that caused atopy. The expression of IL-4, IL-13, each of which is a cytokine secreted by T helper type 2 (Th2) cells, was markedly suppressed in the CT group as compared with AE group (p<0.05). CR treatment also decreased the expression of iNOS, $p-I{\kappa}B$. Atopic dermatitis induced dermatological damage to skin, such as hyperplasia of epithelium, and capillary proliferation was significantly reduced by CR administration. CR effectively inhibited the thinning of the skin barrier and inflammatory responses in atopic dermatitis-induced mice. In particular, it showed anti-inflammatory effects by reducing the expression of IL-4 and IL-13, Th2 cell cytokines, which play a crucial role in development of atopic dermatitis. Therefore, CR can be a good candidate to ameliorate and treat atopic dermatitis.

• 한방재활의학과학회지
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• 제31권1호
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• pp.33-46
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• 2021

Objectives The aim of this study is to evaluate anti-inflammatory and anti-arthritic effects of Sogyunghwalhyel-tang-gamibang (SGHHTGB) in cell and animal models and also to suggest one of putative mechanisms underlying its anti-arthritic effects. Methods Enzyme-linked immunosorbent assay was applied to measure the concentrations of tumor necrosis factor-α (TNF-α), interleukin (IL)-1β, IL-6 and prostaglandin E2 (PGE2) in culture medium and blood serum and nitric oxide (NO) was assayed by Griess reagent. The expressions of inducible NO synthase (iNOS) and cyclooxygenase-2 (COX-2) were analyzed by Western blot method. Results In a cell model using RAW264.7 macrophages stimulated with the endotoxin lipopolysaccharide (LPS), the drug, at its non-cytotoxic concentrations, inhibited the production of the pro-inflammatory cytokine TNF-α, IL-1β and IL-6. In addition, it suppressed the expression of the inflammatory enzyme iNOS and COX-2, and reduced the synthesis of the enzyme product NO (as stable nitrite) and PGE2 in activated macrophages. Meanwhile, in an animal model using rheumatic arthritis (RA) mice induced with injection of type II collagen antibody (CAb) and LPS, the drug improved clinical symptom of arthritis and reduced paw thickness and inflammatory cell infiltration. In blood of RA mice, the drug reduced serum levels of TNF-α, IL-1β, IL-6, nitrite, and PGE2, all inflammatory mediators produced by activated macrophages. Conclusions SGHHTGB may ameliorate CAb and LPS-induced RA in mice, presumably by inactivating macrophages that are capable of initiating joint inflammation by producing pro-inflammatory cytokines and expressing inflammatory enzymes.

• Journal of Korean Neurosurgical Society
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• 제65권2호
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• pp.204-214
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• 2022

Objective : Osteoporosis result from age-related decline in the number of osteoblast progenitors in the bone marrow. Probiotics have beneficial effects on the host, when administered in appropriate amounts. This study investigated the effects of probiotics expressing specific genes, especially the effects of genetically modified bone morphogenetic protein (BMP)-2-expressing Lactobacillus plantarum CJNU 3003 (LP) on ovariectomized rats. Methods : Twenty-eight female Wistar rats (250-300 g, 12 weeks old) were divided into four groups : the sham (control), the ovariectomy (OVX)-induced osteoporosis group (OVX), the OVX and LP (OVX/LP), OVX and genetically modified BMP-2-expressing LP (OVX/LP with BMP) groups. The three groups underwent bilateral OVX and two of these groups were administered two different types of LP via oral gavage daily. At 16 weeks post-OVX, blood was collected from the heart and the bilateral tibiae were extracted and were scanned by ex-vivo micro-computed tomography and stained with hematoxylin-and-eosin (H&E) and Masson's trichrome stain for pathological assessment. The serum levels of osteocalcin (OC), rat C-telopeptide of type I collagen (CTX-I), BMP-2, and receptor activator of nuclear factor-ĸB ligand (RANKL) were measured. Results : The 3D-micro-computed tomography images showed that the trabecular structure in the OVX/LP with BMP group was maintained compared with OVX and OVX/LP groups. No significant differences were detected in trabecular thickness (Tb.Th) between control and OVX/LP with BMP groups (p>0.05). Furthermore, a tendency toward increased BMD, trabecular bone volume, Tb.Th, and trabecular number and decreased trabecular separation was found in rats in the OVX/LP with BMP groups when compared with the OVX and OVX/LP groups (p>0.05). The H&E and Masson's trichrome stained sections showed a thicker trabecular bone in the OVX/LP with BMP group compared with the OVX and OVX/LP groups. There was no difference in serum levels of OC, CTX and RANKL control and OVX/LP with BMP groups (p>0.05). In contrast, significant differences were found in OC and CTX-1 levels between the OVX and OVX/LP with BMP groups (p<0.05). Conclusion : Our results showed that the expression of genetically modified BMP-2 showed inhibition effect for bone loss in a rat model of osteoporosis.

• 환경위생공학
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• 제7권2호
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• pp.95-110
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• 1992

Much progress has been made in understanding the subcellular events of the human lung injuries after acute exposure to environmental air pollutants. Host of those events represent oxidative damages mediated by reactive oxygen species such as superoxide, hydrogen peroxide, and the hydroxy, free radical. Recently, nitric oxide (NO) was found to be endogenously produced by endothelial cells and cells of the reticulo-endothelial system as endothelialderived relaxation factor (EDRF) which is a vasoactive and neurotransmitter substance. Together with superoxide, NO can form another strong oxidant, peroxonitrite. The relative importance of exogenous sources of $N0/N0_2$ and endogenous production of NO by the EDRF producing enzymes in the oxidative stresses to the heman lung has to be elucidated. The exact events leading to chronic irreversible damage are still yet to be known. From chronic exposure to oxidant gases, progressive epithelial and interstitial damages develop. Type I epithelial cells become thicker and cover a smaller average alveolar surface area while thee II cells proliferate instead. Under acute damages, the extent of loss of the alveolar epithelial cell lining, especially type II cells appears to be a good predictor of the ensuing irreversible damage to alveolar compartment. Interstitial matrix undergo remodeling during chronic exposure with increased collagen fibers and interstitial fibroblasts. However, Inany of these changes can be reversed after cessation of exposure. Among chronic lung injuries, genetic damages and repair responses received particular attention in view of the known increased lung cancer risks from exposure to several air pollutants. Heavy metals from foundry emission, automobile traffics, and total suspended particulate, especially polycystic aromatic hydrocarbons have been positively linked with the development of lung cancer. Asbestos in another air pollutant with known risk of lung cancer and mesothelioma, but asbestos fibers are nonauthentic in most bioassays. Studies using the electron spin resonance spin trapping method show that the presence of iron in asbestos accelerates the production of the hydroxy, radical in vitro. Interactions of these reactive oxygen species with particular cellular components and disruption of cell defense mechanisms still await further studies to elucidate the carcinogenic potential of asbestos fibers of different size and chemical composition. The distribution of inhaled pollutants and the magnitude of their eventual effects on the respiratory tract are determined by pollutant-independent physical factors such as anatomy of the respiratory tract and level and pattern of breathing, as well as by pollutant-specific phyco-chemical factors such as the reactivity, solubility, and diffusivity of the foreign gas in mucus, blood and tissue. Many of these individual factors determining dose can be quantified in vitro. However, mathematical models based on these factors should be validated for its integrity by using data from intact human lungs.

• Childhood Kidney Diseases
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• 제8권1호
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• pp.18-25
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• 2004

목 적 : 알포트 증후군은 감각신경성 난청을 동반하는 만성 진행성 유전성 신질환으로, 질환의 경과의 다양성으로 인하여 진행성의 예후인자에 특히 유전학적인 역할에 대한 관심이 커지고 있다. 본 연구에서는 만성 진행성 신질환에서 신부전으로의 진행에 관여하는 것으로 알려진 안지오텐신전환효소 유전자 다형성의 발현빈도 및 병의 진행경과와의 관계를 조사하고, 신부전으로의 진행을 억제하는 치료제로서 안지오텐신전환효소 억제제의 가능성을 알아보고자 하였다. 방 법 : 임상양상 및 신조직검사상 알포트 증후군으로 진단된 12명의 환아와 발병하지 않은 12명의 가족들에서 안지오텐신전환효소 다형성의 빈도를 비교하였다. 대상 환아를 신염의 발생 후 10세 이후까지 정상적인 신기능을 유지하고 있는 신기능 유지군과, 발병 후 10세 이전에 만성신부전으로 진행을 시작한(신기능 유지기간 5년 이하) 조기 신부전 진행군으로 구분하여 신기능 감소의 진행시기와 다형성과의 관계를 비교하였고, 발병이 없는 가족에서 다형성의 빈도 및 양상을 관찰하였다. 정상 대조군으로 고혈압, 신장 혹은 심장질환이 없고, 신질환의 가족력, 안지오텐신전환효소 억제제 혹은 다른 항고혈압제의 사용경력이 없는 소아환자 35명을 정상대조군으로 선정하였다. 결과 : 1) 발병연령은 신기능 유지군이 평균 $3.45{\pm}2.4$세였고, 조기 신부전 진행군이 평균 $4.4{\pm}1.2$세로 의미있는 차이가 없었으며, 남녀비는 각각 5:3, 2:1이었다. 2) 12례의 환아 중 4례(33%)는 10세 이전에 평균 8.9세 이전에 만성신부전으로 진행하여 발병에서부터 평균 4.5년 안에 신부전으로 빠를 진행을 보여주는 조기 신부전 진행군이었고, 8례(67%)는 10세 이후까지 이상 신기능이 유지되는 신기능 유지군으로 발병 이후 평균 10.6년(최소 8년, 최대 15년 이상) 이상 신기능을 유지하고 있었으며, 두 군간의 발병 당시 신기능 및 임상양상의 의미있는 차이는 없었다. 3) 알포트 환자의 안지오텐신전환효소 유전자형태는 II type이 3례(25.0%), ID type이 5례(41.7%). DD type이 4례(33.3%)로 정상대조군의 IItype 44.3%, ID type 40.9%, DD type 14.8%와 비교하여 DD type이 많은 경향을 보였으나 두 군사이의 의미 있는 차이는 없었다. 4) 조기 신부전 진행군 4례 중 3례(75%)에서 DD type을 보였고, 신기능 유지군 8례 중 1례(12.5%)에서 DD type을 보여 두 군간의 의미있는 차이를 보였다(p<0.05). 5) 신질환을 나타내지 않은 가족을 대상으로한 조사에서는 총 12례 중 II type 5례(41.7%), ID type 5례(41.7%), DD type 2례(16.6%)로 역시 정상대조군과 유의한 차이를 보이지 않았으며, 신기능 유지여부와 가족내의 DD발현 빈도는 의미있는 관계가 없었다. 결 론 : 소아 알포트 증후군에서 안지오텐신전환효소 유전자의 다형성의 빈도는 정상대조군에 비하여 의미있는 차이가 없었으나 조기에 신부전으로 진행한 환아에서의 DD genotype의 발현율이 의미있게 높게 나타나, 예후인자로서 가능성을 시사하였으며, 따라서 안지오텐신전환효소 억제제 치료가 신부전으로의 진행을 억제하는데 도움을 줄 수 있을 것으로 생각되었다.