• 치과방사선
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• 제7권1호
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• pp.9-15
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• 1977

The author observed the effect of X-ray irradiation on the secondary palate formation of the rat fetuses. The mothers were exposed to X-radiation on the 10½th, 11½th and 12½th day of gestation with respectively 150, 200, 250, 300 and 350 rads. The fetuses were removed from mothers on 15½h, 16½th and 18½th day of gestation. Morphological changes in palate formation were examined and histochemical preparations were made. 1. In control fetuses, the secondary palates were fully developed on the 15½th to 18½th day of gestation. But in experimental fetuses, many cleft palates were observed in accordance with increase of X-radiation dose. 2. Frequency of incidence of horizontal position of both palated shelves in cleft palate was highest. 3. Accordig to the dislocation of palatal processes, the stain ability of palatal crest was varied. 4. The thickened area of palatal epithelium of palatal crest showed intense methyl green-pyronin and PAS reaction 5. Mesenchymal cell condensation was appeared under the thickened epithelium of palatal process and this mesenchymal tissue showed strong colloidal iron reaction. 6. The stain ability of alizarin red S and alkaline phosphatase reaction of tectal ridge were decreased, in accordance with increase of irradiation doses.

• Archives of Plastic Surgery
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• 제34권5호
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• pp.537-542
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• 2007

Purpose: Adipose tissue contains a population of pluripotent stem cells capable of differentiating along multiple mesenchymal cell lineages. It is well known that fat depots from different part of our body shows different nature not only in morphological aspect but also physiologic aspect. The authors compared the adipogenic potentials and osteogenic potentials of adipose stem cells from different anatomical sites of human. Methods: After laparotomy by surgery team, the authors isolated these adipose stem cells successfully from 7 men with an average age of 58, and induced differentiation along adipogenic and osteogenic lineages in vitro. On the 14th day, cells cultured in adipogenic media differentiated into adipocytes in vitro, as evidenced by positive Oil Red O staining of lipid vacuoles. On the 21st day, cells cultured in osteogenic media differentiated into osteoblasts in vitro as demonstrated by Alizarin red staining of a calcified extracellular matrix. Results: After exposure to adipogenic and osteogenic differentiation medium, subcutaneous adipose stem cells were found to possess greater adipogenic and osteogenic potentials than cells isolated from visceral adipose tissues. Conclusion: This study indicates that adipogenic and osteogenic potentials of adipose stem cells vary by their anatomical sites, with subcutaneous adipose stem cells exhibiting higher adipogenic and osteogenic potential than those isolated from visceral fat.

• Journal of Periodontal and Implant Science
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• 제34권1호
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• pp.93-100
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• 2004

많은 연구들에서 hMSC를 얻기 위해 centrifugation, fluoroscence activated cell sorter(FACS), magnetic activated cell sorter(MACS)가 이용되어져 왔다. 그러나 centrifugation만을 이용한 경우 순도가 떨어지며 FACS나 MACS의 경우에는 비용, 시간이 많이 드는 단점이 있다. 따라서 이 연구에서는 antibody cocktail을 이용하여 hMSC를 좀더 쉽게 얻어내는 방법에 대해 알아보았다. 사람의 골반에서 12G의 바늘을 이용하여 골수를 흡입한 후 heparin이 들어있는 시험관에 넣고 처리과정을 시행하기 전에 냉장고에 보관하며 가능한 한 빨리 처리 과정을 실시한다. 얻은 골수에 적당량의 RosetteSep( Stemcell Technologies)을 첨가한 후 실온에서 20분간 반응시킨다. 그 후 적당량의 Ficoll-paque위에 골수와 RosetteSep의 혼합물을 섞이지 않게 올리고 원심분리를 이용하여 원하는 세포층을 얻어낸다. 이 세포층을 따로 분리한 뒤 배양한다. 배양 시 세포가 80%이상 차기 전에 계속 passage를 시행하며 배양한다. 이는 세포가 밀도가 높아져 원치 않는 세포로 분화되는 것을 막기 위함이다. 배양된 세포가 다양한 분화능력을 가지고 있는지 알아보기 위해 세 가지로 분화를 유도하였다. 적절한 배지와 적절한 환경에서 배양함으로써 얻어진 세포를 osteoblast, chondroblast, adipocyte로 분화를 유도하였다. 분화된 세포가 원하는 형질의 세포로 분화되었는지를 확인하기 위하여 osteoblast의 경우 alizarin red staining, alkaline phosphatase activity, chondroblast의 경우 toluidine blue staining, adipocyte의 경우 Oil-Red-O staining으로 염색하여 분화를 확인하였다. 분리해낸 세포는 각각 세 가지 세포로 분화가 되었으며 이는 RosetteSep이 hMSC를 성공적으로 분리해냈다는 것을 보여준다. 그러나 모든 세포가 분화를 보이지는 않았으며 따라서 hMSC의 순도를 높이기 위한 연구가 더 필요하다. RosetteSep을 이용하면 다른 방법들 보다 쉽게 hMSC를 얻을 수 있으나 기존의 방법과 순도의 측면에서 더 비교할 필요가 있다.

• The Korean Journal of Physiology and Pharmacology
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• 제24권6호
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• pp.463-472
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• 2020

Direct reprogramming, also known as a trans-differentiation, is a technique to allow mature cells to be converted into other types of cells without inducing a pluripotent stage. It has been suggested as a major strategy to acquire the desired type of cells in cell-based therapies to repair damaged tissues. Studies related to switching the fate of cells through epigenetic modification have been progressing and they can bypass safety issues raised by the virus-based transfection methods. In this study, a protocol was established to directly convert fully differentiated fibroblasts into diverse mesenchymal-lineage cells, such as osteoblasts, adipocytes, chondrocytes, and ectodermal cells, including neurons, by means of DNA demethylation, immediately followed by culturing in various differentiating media. First, 24 h exposure of 5-azacytidine (5-aza-CN), a well-characterized DNA methyl transferase inhibitor, to NIH-3T3 murine fibroblast cells induced the expression of stem-cell markers, that is, increasing cell plasticity. Next, 5-aza-CN treated fibroblasts were cultured in osteogenic, adipogenic, chondrogenic, and neurogenic media with or without bone morphogenetic protein 2 for a designated period. Differentiation of each desired type of cell was verified by quantitative reverse transcriptase-polymerase chain reaction/western blot assays for appropriate marker expression and by various staining methods, such as alkaline phosphatase/alizarin red S/oil red O/alcian blue. These proposed procedures allowed easier acquisition of the desired cells without any transgenic modification, using direct reprogramming technology, and thus may help make it more available in the clinical fields of regenerative medicine.

• Nutrition Research and Practice
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• 제15권5호
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• pp.579-590
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• 2021

BACKGROUND/OBJECTIVES: Petasites japonicus Maxim (P. japonicus) has been used as an edible and medicinal plant and contains many bioactive compounds. The purpose of this study is to investigate the effect of P. japonicus on osteogenesis. MATERIALS/METHODS: The leaves and stems of P. japonicus were separated and extracted with hot water or ethanol, respectively. The total phenolic compound and total polyphenol contents of each extract were measured, and alkaline phosphatase (ALP) activity of each extract was evaluated to determine their effect on bone metabolism. To investigate the effect on osteoblast differentiation of the aqueous extract of P. japonicus leaves (AL), which produced the highest ALP activity among the tested extracts, collagen content was measured using the Sirius Red staining method, mineralization using the Alizarin Red S staining method, and osteocalcin production through enzyme-linked immunosorbent assay analysis. Also, real-time reverse transcription polymerase chain reaction was performed to investigate the mRNA expression levels of Runt-related transcriptional factor 2 (Runx2) and Osterix. RESULTS: Among the 4 P. japonicus extracts, AL had the highest values in all of the following measures: total phenolic compounds, total polyphenols, and ALP activity, which is a major biomarker of osteoblast differentiation. The AL-treated MC3T3-E1 cells showed significant increases in induced osteoblast differentiation, collagen synthesis, mineralization, and osteocalcin production. In addition, mRNA expressions of Runx2 and Osterix, transcription factors that regulate osteoblast differentiation, were significantly increased. CONCLUSIONS: These results suggest that AL can regulate osteoblasts differentiation, at least in part through Runx2 and Osterix. Therefore, it is highly likely that P. japonicus will be useful as an alternate therapeutic for the prevention and treatment of osteoporosis.

• 한국응용과학기술학회지
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• 제38권1호
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• pp.168-177
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• 2021

본 연구는 방사선 육종 차조기와 백출 복합물의 조골세포 분화 활성 및 파골세포 형성 억제를 조사하였다. 차조기와 백출 복합물은 MG-63 세포에서 ALP 활성 및 arlizarin red 염색을 확인하였고 조골세포 형성의 영향은 RAW 264.7 세포에서 TRAP 활성과 TRAP 염색을 진행하였다. 세포 독성시험에서 차조기와 백출 복합물은 50 ㎍/㎖ 농도 이하에서 안전한 것으로 확인되었다. ALP 활성 및 골석회화 형성 능력은 대조군보다 활성이 낮았으나, 파골세포에서 TRAP 활성을 유의적으로 감소시켰으며, 효과적으로 TRAP(+) 다핵세포를 억제하였다. 따라서 차조기와 백출 복합물은 골 흡수 억제 활성을 향상시켜 뼈 관련 질환의 예방 및 치료에 효과적인 것으로 보여진다.

• Nutrition Research and Practice
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• 제18권4호
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• pp.479-497
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• 2024

BACKGROUND/OBJECTIVES: Activating brown adipose tissue (BAT) and browning of white adipose tissue (WAT) can protect against obesity and obesity-related metabolic conditions. Cryptotanshinone (CT) regulates lipid metabolism and significantly ameliorates insulin resistance. Adenosine-5'-monophosphate (AMP)-activated protein kinase (AMPK), a receptor for cellular energy metabolism, is believed to regulate brown fat activity in humans. MATERIALS/METHODS: The in vivo study included high-fat-fed obese mice administered orally 200/400 mg/kg/d CT. They were evaluated through weight measurement, the intraperitoneal glucose tolerance test (IPGTT), intraperitoneal insulin tolerance test (IPITT), cold stimulation test, serum lipid (total cholesterol, triglycerides, and low-density lipoprotein) measurement, hematoxylin and eosin staining, and immunohistochemistry. Furthermore, the in vitro study investigated primary adipose mesenchymal stem cells (MSCs) with incubation of CT and AMPK agonists (acadesine)/inhibitor (Compound C). Cells were evaluated using Oil Red O staining, Alizarin red staining, flow cytometry, and immunofluorescence staining to identify and observe the osteogenic versus adipogenic differentiation. Quantitative real-time polymerase chain reaction and the Western blot were used to observe related gene expression. RESULTS: In the diet-induced obesity mouse model mice CT suppressed body weight, food intake, glucose levels in the IPGTT and IPTT, serum lipids, the volume of adipose tissue, and increased thermogenesis, uncoupling protein 1, and the AMPK pathway expression. In the in vitro study, CT prevented the formation of lipid droplets from MSCs while activating brown genes and the AMPK pathway. AMPK activator enhanced CT's effects, while the AMPK inhibitor reversed the effects of CT. CONCLUSION: CT promotes adipose tissue browning to increase body thermogenesis and reduce obesity by activating the AMPK pathway. This study provides an experimental foundation for the use of CT in obesity treatment.

• 한국균학회소식:학술대회논문집
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• 한국균학회 2014년도 춘계학술대회 및 임시총회
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• pp.43-43
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• 2014

White rot fungi have been useful source of enzymes for the degradation of environmental pollutants including polycyclic aromatic hydrocarbons (PAHs) and synthetic dyes. PAHs are widespread organic compounds present in fossil fuels and are routinely generated by incomplete fuel combustion. PAHs are some of the major toxic pollutants of water and soil environments. Synthetic dyes are major water-pollutants, which are toxic to organisms in water environments and interfere photosynthesis of water plants. Removal of PAHs and synthetic dyes has been of interests in the environmental science especially in the environmental microbiology. Mushrooms are fungal groups that function as primary degraders of wood polyphenolic lignin. The ligninolytic enzymes produced by mushroom, including manganese peroxidase, lignin peroxidase, and laccase, mediate the oxidative degradation of lignin. The catalytic power of these enzymes in the degradation of aromatic ring compounds has been sought for the degradation of various organic compounds. In this project, we have screened 60 wild mushroom strains for their degradation activity against two representative PAHs, naphthalene and anthracene, and five aromatic dyes, including alizarin red S, crystal violet, malachite green, methylene blue, rose bengal. The degradation of PAHs was measured by GC while the decolorization of dyes was measured by both UV spectrophotometer and HPLC. As results, 9 wild mushroom strains showed high activity in degradation of PAHs and textile dyes. We also describe the secretive enzyme activities, the transcription levels, and cloning of target genes. In conjunction with this, activities of degradative enzymes, including laccase, lignin peroxidase, and Mn peroxidase, were measured in the liquid medium in the presence of PAHs and dyes. Our results showed that the laccase activity was directed correlated with the degradation, indicating that the main enzyme acts on PAHs and dyes is the laccase. The laccase activity was further simulated by the addition of $Cu^{2+}$ ion. Detailed studies of the enzyme system should be sought for future applications.

• 대한치과보철학회지
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• 제29권3호
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• pp.55-74
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• 1991

The purpose of this study was to investigate the influence of early functional load around osseointegrated titanium implants. 24 titanium plasma spray coated implants (ITI HS-type) were placed into the previously extracted site in the mandible of six adult dogs. The implants were divided into three groups : the control group was the implants without abutment during the experimental period; the experimental group I was loaded by connecting the contoured abutment after 6 weeks of healing; the experimental group II was loaded after 12 weeks of healing: and the mandibular second premolar and surrounding tissues were selected for natural tooth group to compare the implanted group. All dogs were injected intravenously tetracycline, alizarin red S, and calcein for bone labeling. After the experimental period of 18 weeks, the dogs were sacrificed and longitudinal sections of the bone-implant interface were cut and observed using light microscope, scanning electron microscope, and fluorescence microscope. The results of the study were as follows: 1. Light and scanning electron microscopically, all implant surfaces were well contact with bone tissue at the cortical layer, but some areas of cancellous bone were not contact directly. 2. Fluorescence microscopically, number and size of the new secondary osteons around the implant were increased than those of the natural tooth. 3. Fluorescence microscopically, linear and concentrical fluorescence was observed at or near the surface of all implants, and the bone formation and remodeling of the implants loaded after 6 week of healing were great, and unloaded implants were worst. 4. Fluorescence microscopically, endosteal bone formation was greater than periosteal bone formation at or near the implants. 5. Fluorescence microscopically, number and size of linear and concentric fluorescence was increased at the lingual side than the buccal side of the loaded implants. The result of the study indicate the possibility of the early load to the implant via a prosthesis.

• Journal of Periodontal and Implant Science
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• 제47권5호
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• pp.273-291
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• 2017

Purpose: Although static magnetic fields (SMFs) have been used in dental prostheses and osseointegrated implants, their biological effects on osteoblastic and cementoblastic differentiation in cells involved in periodontal regeneration remain unknown. This study was undertaken to investigate the effects of SMFs (15 mT) on the osteoblastic and cementoblastic differentiation of human osteoblasts, periodontal ligament cells (PDLCs), and cementoblasts, and to explore the possible mechanisms underlying these effects. Methods: Differentiation was evaluated by measuring alkaline phosphatase (ALP) activity, mineralized nodule formation based on Alizarin red staining, calcium content, and the expression of marker mRNAs assessed by reverse transcription polymerase chain reaction (RT-PCR). Signaling pathways were analyzed by western blotting and immunocytochemistry. Results: The activities of the early marker ALP and the late markers matrix mineralization and calcium content, as well as osteoblast- and cementoblast-specific gene expression in osteoblasts, PDLCs, and cementoblasts were enhanced. SMFs upregulated the expression of Wnt proteins, and increased the phosphorylation of glycogen synthase $kinase-3{\beta}$ ($GSK-3{\beta}$) and total ${\beta}-catenin$ protein expression. Furthermore, p38 and c-Jun N-terminal kinase (JNK) mitogen-activated protein kinase (MAPK), and nuclear $factor-{\kappa}B$ ($NF-{\kappa}B$) pathways were activated. Conclusions: SMF treatment enhanced osteoblastic and/or cementoblastic differentiation in osteoblasts, cementoblasts, and PDLCs. These findings provide a molecular basis for the beneficial osteogenic and/or cementogenic effect of SMFs, which could have potential in stimulating bone or cementum formation during bone regeneration and in patients with periodontal disease.