• 치위생과학회지
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• 제23권4호
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• pp.282-295
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• 2023

Background: Secretory leukocyte protease inhibitor (SLPI) protects tissues from proteases and promotes cell proliferation and healing. SLPI also reduces periodontal inflammation and alveolar bone resorption by inhibiting proinflammatory cytokine expression in rat periodontal tissues and osteoblasts. However, little is known of the role of SLPI in the expression of osteoclast regulatory factors from osteoblasts, which are crucial for the interaction between osteoblasts and osteoclasts. Therefore, we aimed to determine the effects of SLPI on the regulation of osteoclasts and osteoblasts in LPS-treated alveolar bone and osteoblasts. Methods: Periodontitis was induced in rats using LPS. After each LPS injection, SLPI was injected into the same area. Immunohistochemical analysis was performed with antibodies against SLPI, RANKL, OPG, and Runx2 in the periodontal tissue. RT-PCR and western blotting were performed to determine the expression levels of SLPI, RANKL, OPG, and Runx2 in LPS- and SLPI/LPS-treated MC3T3-E1 cells. SLPI/LPS-treated MC3T3-E1 cells were also stained with Alizarin Red S. Results: Immunohistochemical analysis showed that the expression levels of SLPI, OPG, and Runx2 were higher while that of RANKL was lower in the LPS/SLPI group relative to those in the LPS group. The mRNA and protein expression of SLPI, OPG, and Runx2 was higher in SLPI/LPS/MC3T3-E1 cells than in LPS/MC3T3-E1 cells, and RANKL expression was lower. During differentiation, OPG and Runx2 protein levels were higher whereas RANKL levels were lower in SLPI/LPS/MC3T3-E1 than in LPS/MC3T3-E1 cells on days 0, 4, 7, and 10. In addition, mineralization and matrix deposition were higher in SLPI/LPS/MC3T3-E1 than in LPS/MC3T3-E1 on days 7 and 10. SLPI decreased RANKL expression in LPS-treated alveolar bone and osteoblasts but increased the expression of OPG and Runx2. Conclusion: SLPI can be considered as a regulatory molecule that indirectly regulates osteoclast activation via osteoblasts and promotes osteoblast differentiation.

• International Journal of Oral Biology
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• 제33권3호
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• pp.105-111
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• 2008

Thrombospondins (TSP-1, TSP-2) are secretory extracellular glycoproteins that are involved in a variety of physiological processes such as tumor cell adhesion, invasion, and metastasis. The present study was undertaken to elucidate the involvement of thrombospondins in the adhesion of osteoblast-like cells using the TSP-1 or TSP-2 antisense MG63 and MC3T3-E1 cell lines. For downregulation of TSPs expression, we prepared antisense constructs for TSP-1 and TSP-2 using the pREP4 an episomal mammalian expression vector, which be able to produce the specific antisense oligonucleotides around chromosome. MG63 and MC3T3-E1 osteoblast-like cells were transfected with the antisense constructs and nonliposomal Fugene 6, and then selected under hygromycin B (50 ${\mu}g/ml$) treatment for 2 weeks. Western blot analysis revealed that expression of the TSP proteins was downregulated in the antisense cell lines. The cell adhesion assay showed that adhesive properties of TSP-1 and TSP-2 antisense MG63 cells on the polystyrene culture plate were reduced to 17% and 21% of the control cells, respectively, and those of the TSP-1 and TSP-2 antisense MC3T3-E1 cells also decreased to 19% and 27% of control, respectively. Adhesion of TSP-1 and TSP-2 antisense MC3T3-E1 cells on Type I collagen-coated culture plate decreased to 27% and 76%, respectively. These results indicate that TSP-1 and TSP-2 proteins may have an important role in adhesion of osteoblast-like cells to extracellular matrix.

• Parasites, Hosts and Diseases
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• 제35권4호
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• pp.259-264
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• 1997

폐흡충 감염시 일어나는 비만세포 (mastcell)의 반응이 감염경로에 따라 어떠한 영향을 미치는지 알아보고자 비호적숙주인 마우스에 폐흡충의 피낭유충 10개를 피하 또는 경구 감염시킨 후 감염 시기별로 진피 내에 동원되는 비만세포의 숫적 변동 및 탈과립률 (%)을 경시적으로 관찰하였다. 피하로 감염시킨 마우스는 감염 1주 ($38.3/\textrm{mm}^2$)부터 진피 내 비만세포의 수가 PBS주입군 (대조군) (범위: $19.4-25.1/\textrm{mm}^2$)에 비해 유의한 수준 (P<0.05)으로 증가하기 시작하여 전 실험기간 동안 증가하였다. 이때 비만세포는 충체의 낭 (cyst)주위의 피하층에도 집중적으로 침윤되어 있었다. 경구로 감염시킨 마우스 는 감염 2주 ($33.5/\textrm{mm}^2$)부터 진피 내 비만세포의 수가 비감염군 (대조군)(범위: $17.4-22.3/\textrm{mm}^2$)에 비 해 유의 한 수준 (p<0.05)으로 증가하기 시작하여 전 실험 기간인 감염 6주 ($38.4/\textrm{mm}^2$)까지 높게 운지 되었다. 진피 내에 동원된 비만세포의 평균 탈과림률 (%)은 피하 감염 및 경구 감염군 모두 감염 2주부 터 6주까지 60%이상을 보인 반면 PBS주입군 (대조군)및 비감염군 (대조군)에 있어서는 전 실험기간 동안 10%정도의 탈과립률이 관찰되었다. 이상의 결과로 보아 폐흡충을 마우스에 감염시켰을 때 일어 나는 진피 내 비만세포 반응은 감염경로와는 상관없이 충체에서 분비되는 배설-분비 항원의 자극과 깊은 관계가 있음을 알 수 있었다.

• 한국어류학회지
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• 제28권1호
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• pp.19-27
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• 2016

은대구의 소화관 상대길이 비는 1.52 (n=12)이며, 소화관은 위의 후방부에 5~6개의 유문수를 가진다. 점막주름의 형태는 식도와 위에서는 미분지형이지만 장에서는 분지형이다. 횡단면에서 소화관은 조직학적으로 점막층, 점막하층, 근육층 및 장막으로 구분할 수 있다. 식도의 점막상피층은 단층이며, 원주섬모상피세포들과 점액세포들로 구성된다. 위 점막층의 위선은 관상선으로 주세포, 벽세포 및 뮤신분비세포들로 구성된다. 뮤신분비세포는 원주형으로 AB-PAS (pH 2.5) 반응에서 분홍색과 푸른색을 나타내는 분비과립을 가진다. 장의 점막상피층은 단층이며, 원주섬모상피세포들과 배상세포들로 구성된다. 점막하층은 소성결합조직층으로 주로 교원섬유들로 구성되며, 식도에서 잘 발달되어 있다. 소화관의 근육층은 종주근층과 환상근층으로 구분되며, 위에서 잘 발달되어 있다. 간은 다수의 소엽구조와 담관들로 이루어져 있으며, H-E 염색에서 간세포의 세포질은 호산성이며, 핵과 인은 호염기성을 보였다. 췌장조직은 소화관 주변의 지방조직에 산재하며, 다수의 외분비세포들로 구성된 포상선이었다. H-E 염색에서 외분비선세포의 세포질은 호염기성을 나타내며, 다수의 호산성 전효소 과립들을 함유한다.

• 한국임상수의학회지
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• 제33권1호
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• pp.74-79
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• 2016

A 7-year-old female Pointer dog with multiple masses in the axilla, mammary gland, and bladder was submitted to the Pathology Department of the College of Veterinary Medicine in the Jeju National University. Grossly, mass between right axilla and 1st mammary gland, $15{\times}10cm$ in size, was well delineated and firm, slightly soft center, oval shape. And masses in right 1st, 3rd and 5th mammary gland were well delineated and sulphur yellow in color on the cut-surface. Numerous round to oval shaped masses, 0.3 to 2 cm in diameter were existed in the lung. Urinary bladder mucosa had rough and thick and round to oval papillary masses, 0.1 to 2 cm in diameter, on surface. Microscopically, masses in right axilla, 1st mammary gland, lung and axillary lymph node were composed of poorly differentiated tubules originated from apocrine gland. Lining neoplastic epithelium showed high mitotic figures, typical apical secretory blebs, and PAS-positive diastase-resistant cytoplasmic granules. Masses in 3rd and 5th mammary gland were confirmed as mammary complex adenoma and simple adenoma respectively. The masses in the urinary bladder were covered with stratified transitional epithelium with marked cellular atypia and high mitotic figures. Some neoplastic cells showed focal invasion into substantia propria of bladder. Immunohistochemaically, neoplastic transitional epithelium demonstrated positive reactions for cytokeratin 7, AE1/AE3, and MNF116. Based on the gross, histopathologic and immunohistochemical characteristics, this dog was diagnosed as apocrine carcinoma, mammary gland tumor including simple adenoma and complex adenoma and bladder transitional cell carcinoma. And distant metastases of apocrine carcinoma in right axilla were observed in axillary lymph node and lungs. This is the first report for concurrent occurrence of apocrine carcinoma, mammary gland tumor, and transitional cell carcinoma in a same dog.

• Journal of Periodontal and Implant Science
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• 제36권3호
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• pp.613-625
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• 2006

Periodontal ligament(PDL) cells and human gingival fibroblasts(HGFs) play important roles in development, regeneration, normal function, and pathologic alteration. PDL cells and HGFs have the similarity related with general characteristics of fibroblast such as spindle shaped morphology, the presence of vimentin intermediate filament and the synthesis of interstitial collagens and fibronectin. There were many studies about the differences between PDL cells and HGFs, but they were not about whole gene level. In this study, we tried to explain the differences of gene expression profiles between PDL cells and HGFs, and the differences among three individuals by screening gene expression patterns of PDL cells and HGFs, using cDNA microarray. Although there were some variants among three experiments, a set of genes were consistentely and differentially expressed in one cell type. Among 3,063 genes, 49 genes were more highly expressed in PDL cells and 12 genes were more highly expressed in HGFs. The genes related with cell structure and motility were expressed more highly in PDL cells. These are cofilin 1, proteoglycan 1 secretory granule, collagen type I(${\alpha}$ 1), adducin gamma subunit, collagen type III(${\alpha}$ 1), fibronectin, lumican(keratan sulfate proteoglycan), and ${\alpha}$ -smooth muscle actin. Tissue inhibitor of metalloproteinase known as the enzyme controlling extracellular matrix with matrix metalloproteinase is more highly expressed in PDL cells, osteoprotegerin known as osteoclastogenesis inhibitory factor is more highly expressed in HGFs. We performed northern blot to verify cDNA microarray results on selected genes such as tissue inhibitor of metalloproteinase, fibronectin, osteoprogeterin. The result of northern blot analysis showed that each cell expressed the genes in similar pattern with cDNA microarray result. This result indicates that cDNA microarray is a reliable method in screening of gene expression profiles.

• Animal cells and systems
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• 제4권2호
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• pp.157-163
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• 2000

Yeast pheromone a-factor is a 13-amino acid peptide hormone that is synthesized as a part of a larger precursor, prepro-$\alpha$-factor, consisting of a signal peptide and a proregion of 64 amino acids. The carboxy-terminal half of the precursor contains four tandem copies of mature $\alpha$-factor. To investigate the molecular basis of intracellular sorting, proteolytic processing, and storage of the peptide hormone, yeast prepro-$\alpha$-factor precursors were heterologously expressed in rat pituitary $GH_3 cells. When cells harboring the precursor were metabolically labeled, a species of approximately 27 kD appeared inside the cells. Digestion with peptide: N-glycosidase F (PNG-F) shifted the molecular mass to a 19 kD, suggesting that the 27 kD protein was the glycosylated form as in yeast cells. The nascent polypeptide is efficiently targeted to the ER in the $GH_3 cells, where it undergoes cleavage of its signal peptide and core glycosylation to generate glycosylated pro-a-factor. To look at the post ER intracellular processing, the pulse-labelled cells were chased up to 2 hrs. The nascent propeptides disappeared from the cells at a half life of 30 min and only 10-25% of the newly synthesized, unprocessed precursors were stored intracellularly after the 2 h chase. However, about 20% of the pulse-labeled pro-$\alpha$-factor precursors were secreted into the medium in the pro-hormone form. With increasing chase time, the intracellular level of propeptide decreased, but the amount of secreted propeptide could not account for the disappearance of intracellular propeptide completely. This disappearance was insensitive to lysosomotropic agents, but was inhibited at $16^{circ}C or 20^{\circ}C$, suggesting that the turnover of the precursors was not occurring in the secretory pathway to trans Golgi network (TGN) or dependent on acidic compartments. From these results, it is concluded that a pan of these heterologous precursors may be processed at its paired dibasic sites by prohormone processing enzymes located in TGN/secretpry vesicles producing small peptides, and that the residual unprocessed precursors may be secreted into the medium rather than degraded intracellularly.

• 한방재활의학과학회지
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• 제34권2호
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• pp.15-28
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• 2024

Objectives We used the D-galactose (D-gal) induced C2C12 myoblast senescence model to investigate whether ethanol extract of Perilla. fructescens leaves (EEPF) could delay cellular senescence and regulate related mechanisms. Methods C2C12 myogenic cells were cultured in an incubator under 37 ℃ and 5% CO₂ conditions. EEPF, dried perilla leaves were pulverized and extracted at 1:10 (v/v) at 50 ℃ for 4 hours. Cell counting kit-8 and western blot analysis was performed. Annexin V-FITC apoptosis detection kit and DAPI staining was applied. Catalase (CAT), glutathione peroxidase (GSH-Px), total antioxidant capacity (T-AOC), superoxide dismutase (SOD), and malondialdehyde analysis kits were used. To measure the level of reactive oxygen species generation, staining and flow cytometry was used. To analyze the mitochondrial activity, membrane potential changes were measured using JC-1. 𝛽-gal activity was analyzed using SA-𝛽-gal staining solution, and DNA damage was analyzed by using 𝛾-H2AX. Quantikine ELISA kit was used to analyze inflammatory cytokine production. Results According to the results of this study, EEPF significantly alleviated the decrease in cell viability in C2C12 cells treated with D-gal and suppressed the decrease in the expression of proliferating cell nuclear antigen. EEPF also markedly blocked D-gal-induced C2C12 cell apoptosis and restored reduced activity of CAT, GSH-Px, T-AOC, SOD. In addition, EEPF suppressed the decrease in 𝛽-galactosidase activity, the induction of DNA damage and the increase in expression of senescence-associated secretory phenotype proteins such as p16, p53 and p21 in D-gal-treated C2C12 cells. Furthermore, EEPF significantly attenuated D-gal-induced production and expression of inflammatory cytokines such as interleukin (IL)-6 and IL-18. Conclusions The results of this study indicate that EEPF can be used as a potential candidate for the prevention and treatment of muscle aging.

• Tuberculosis and Respiratory Diseases
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• 제80권1호
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• pp.77-82
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• 2017

Background: Delayed hypersensitivity plays a large role in the pathogenesis of tuberculous pleural effusion (TPE). Macrophages infected with live Mycobacterium tuberculosis (MTB) increase the levels of adenosine deaminase2 (ADA2) in the pleural fluid of TPE patients. However, it is as yet unclear whether ADA2 can be produced by macrophages when challenged with MTB antigens alone. This study therefore evaluated the levels of ADA2 mRNA expression, using monocyte-derived macrophages (MDMs) stimulated with MTB antigens. Methods: Purified monocytes from the peripheral blood mononuclear cells of healthy volunteers were differentiated into macrophages using granulocyte-macrophage colony-stimulating factor (GM-CSF) or macrophage colony-stimulating factor (M-CSF). The MDMs were stimulated with early secretory antigenic target protein 6 (ESAT6) and culture filtrate protein 10 (CFP10). The mRNA expression levels for the cat eye syndrome chromosome region, candidate 1 (CECR1) gene encoding ADA2 were then measured. Results: CECR1 mRNA expression levels were significantly higher in MDMs stimulated with ESAT6 and CFP10, than in the unstimulated MDMs. When stimulated with ESAT6, M-CSF-treated MDMs showed more pronounced CECR1 mRNA expression than GM-CSF-treated MDMs. Interferon-${\gamma}$ decreased the ESAT6- and CFP10-induced CECR1 mRNA expression in MDMs. CECR1 mRNA expression levels were positively correlated with mRNA expression of tumor necrosis factor ${\alpha}$ and interleukin 10, respectively. Conclusion: ADA2 mRNA expression increased when MDMs were stimulated with MTB antigens alone. This partly indicates that pleural fluid ADA levels could increase in patients with culture-negative TPE. Our results may be helpful in improving the understanding of TPE pathogenesis.

• 한국가금학회지
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• 제28권3호
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• pp.267-274
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• 2001

부화후 성장하는 꿩 송과샘의 미세구조적 발달을 구명하고자 1일령, 1, 2 및 6개월령을 희생시켜 광학 및 전자현미경을 이용하여 다음과 같은 결과를 얻었다. 광학현미경적 관찰에서 꿩 송과샘의 실질은 피막에 의해 싸여 있고 여기에서 기시하는 결합조직에 의해 불완전한 소엽 (lobule)으로 구분되었다. 송과샘실질은 소엽상과 불완전한 여포살이었으며, 둥근 핵과 염색성이 옅은 송과샘세포와 약간 짙고 길쭉한 핵을 갖는 지주세포로 구성되어 있었다. 전자현미경적 관찰에서 송과샘실질에는 길쭉한 송과샘세포와 지주세포가 소엽의 중심부를 향해 배열되어 있었고 광학현미경에서 소엽의 중심부에는 매우 불규칙한 막성층판복합체이나 포상구조물이 출현하였다. 실질은 비교적 밝은 솔과 샘세포와 짙은 핵을 갖는 지주세포로 구성되어 있었다. 송과샘세포는 잔유성 광수용세포의 형태를 취하였는데, 그 첨단부위의 밝고 팽대된 세포질에는 세포소기관이 빈약하나 섬모, 약간의 유리리보솜과 사립체가 출현하였다. 첨단부위세포질과 핵위부위세포질 사이의 좁아진 경부는 지주세포와 연접복합체를 이루고 목부위세포질내에는 미세소관이 풍부하였다. 핵주위세포질은 풍부하고 다수의 사립체, 잘 발달된 골지장치, 풍부한 과립형질내세망 및 유리 리보솜 등을 함유하고 있었다. 또한 핵아래부위세포질에서 여포의 기저부위로 기저돌기를 내고 있었다. 송과샘세포의 핵아래부위 세포질 및 기저돌기에서 60∼90nm 크기의 치밀소포가 관찰된 것이 특이하였고 짙은 핵을 갖고 긴 독기를 갖는 것이 특징이었다. 이상의 결과는 꿩 송과샘세포가 광수용능은 것의 없고 분비기능을 주로 수행하는 소견이었으며, 성숙 꿩의 송과샘의 미세구조적 연구를 확인하여 번식기와 비번식기에 따른 비교연구가 수행되어야 할 것으로 사료된다.