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The Effects of Proinflammatory Cytokines and TGF-beta, on The Fibroblast Proliferation (Proinflammatory Cytokines과 TGF-beta가 섬유모세포의 증식에 미치는 영향)

  • Kim, Chul;Park, Choon-Sik;Kim, Mi-Ho;Chang, Hun-Soo;Chung, Il-Yup;Ki, Shin-Young;Uh, Soo-Taek;Moon, Seung-Hyuk;Kim, Yong-Hoon;Lee, Hi-Bal
    • Tuberculosis and Respiratory Diseases
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    • v.45 no.4
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    • pp.861-869
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    • 1998
  • Backgrounds: The injury of a tissue results in the infalmmation, and the imflammed tissue is replaced by the normal parenchymal cells during the process of repair. But, constitutional or repetitive damage of a tissue causes the deposition of collagen resulting in the loss of its function. These lesions are found in the lung of patients with idiopathic pulmonary fibrosis, complicated fibrosis after diffuse alveolar damage (DAD) and inorganic dust-induced lung fibrosis. The tissue from lungs of patients undergoing episodes of active and/or end-stage pulmonary fibrosis shows the accumulation of inflammatory cells, such as mononuclear cells, neutrophils, mast cells and eosinophils, and fibroblast hyperplasia. In this regard, it appears that the inflammation triggers fibroblast activation and proliferation with enhanced matrix synthesis, stimulated by inflammatory mediators such as interleukin-1 (IL-1) and/or tumor necrosis factor (TNF). It has been well known that TGF-$\beta$ enhance the proliferation of fibroblasts and the production of collagen and fibronectin, and inhibit the degradation of collagen. In this regard, It is likely that TGF-$\beta$ undergoes important roles in the pathogenesis of pulmonary fibrosis. Nevertheless, this single cytokine is not the sole regulator of the pulmonary fibrotic response. It is likely that the balance of many cytokines including TGF-$\beta$, IL-1, IL-6 and TNF-$\alpha$ regulates the pathogenesis of pulmonary fibrosis. In this study, we investigate the interaction of TGF-$\beta$, IL-1$\beta$, IL-6 and TNF-$\alpha$ and their effect on the proliferation of fibroblasts. Methods: We used a human fibroblast cell line, MRC-5 (ATCC). The culture of MRC-5 was confirmed by immunofluorecent staining. First, we determined the concentration of serum in cuture medium, in which the proliferation of MRC-5 is supressed but the survival of MRC-5 is retained. Second, we measured optical density after staining the cytokine-stimulated cells with 0.5% naphthol blue black in order to detect the effect of cytokines on the proliferation of MRC-5. Result: In the medium containing 0.5% fetal calf serum, the proliferation of MRC-5 increased by 50%, and it was maintained for 6 days. IL-1$\beta$, TNF-$\alpha$ and IL-6 induced the proliferation of MRC-5 by 45%, 160% and 120%, respectively. IL-1$\beta$ and TNF-$\alpha$ enhanced TGF-$\beta$-induced proliferation of MRC-5 by 64% and 159%, but IL-6 did not affect the TGF-$\beta$-induced proliferation. And lNF-$\alpha$-induced proliferation of MRC-5 was reduced by IL-1$\beta$ in 50%. TGF-$\beta$, TNF-$\alpha$ and both induced the proliferation of MRC-5 to 89%, 135% and 222%, respectively. Conclusions: TNF-$\alpha$, TGF-$\beta$ and IL-1$\beta$, in the order of the effectiveness, showed the induction of MRC-5 proliferation of MRC-5. TNF-$\alpha$ and IL-1$\beta$ enhance the TGF-$\beta$-induced proliferation of MRC-5, but IL-6 did not have any effect TNF-$\alpha$-induced proliferation of MRC-5 is diminished by IL-1, and TNF-$\alpha$ and TGF-$\beta$ showed a additive effect.

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Effects of Activation Treatments and Culture Condition on In Vitro Development of Caprine In Vivo and In Vitro Oocytes (재래산양의 체내 및 체외유래 난자의 활성화 처리방법 및 배양조건이 단위발생란의 체외발달에 미치는 영향)

  • Park H. S.;Kim T. S.;Lee Y. H.;Jung S. Y.;Lee M. Y.;Jin J. I.;Park J. K.;Lee J. S.;Kim C. H.
    • Reproductive and Developmental Biology
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    • v.28 no.3
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    • pp.181-185
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    • 2004
  • This study was conducted to examine whether activation treatments, source of oocytes and culture conditions affect in vitro developmental ability of caprine oocytes. Mature Korean native goats were pretreated with intravaginal CIDR for 10 days. The goats were then treated with a single intramuscular injection of 1,000 IU PMSG on Day 8 or twice daily injection of a total of 70 mg FSH for 3 days from Day 8 of CIDR insertion for superovulation. All the goats were injected with 10 mg PGF/sub 2a/ on Day 8 and 400 IU hCG on Day 10 of CIDR. Oocytes were surgically collected by oviduct flushing(in vivo maturation) or direct follicle aspiration(in vitro maturation) through mid-ventral incision at 35 h after hCG injection. Fifteen to twenty oocytes were placed in TCM-199 medium containing 25 mM Hepes and hormones under mineral oil at 39℃ in a humudified atmosphere of 5% CO₂ in air for 22 to 24 h. After maturation, the oocytes were activated by electric stimulation or ionomycin + 6-DMAP. The activated oocytes were then cultured in M16, TCM-199 and mSOF media supplemented with proteins at 39℃ for 6 to 7 days. Activation treatments did not affect cleavage of the oocytes. The cleavage rates were 64.1% (41/64) in oocytes activated by electric stimulation and 76.5% (218/285) in oocytes activated by ionomycin + 6-DMAP. The proportion of development to blastocyst was 15.6% (34/218) in oocytes activated by ionomycin + 6-DMAP, but activation by electric stimulation did not support embryos developed beyond morula stage. There were no differences in the cleavage rates of activated oocytes experiencing in vivo (86.8%, 66/76) and in vitro maturation (69.0%, 127/184). However, the development rate to blastocyst stage was significantly (P<0.05) higher for oocytes matured in vivo (50.0%, 33/66) compared to in vitro (0.8%, 1/127). Culture conditions did not affect the cleavage of -activated oocytes. The cleavage rates were 51.6% (49/95) in M16, 64.3% (18/28) in TCM-199 and 81.0% (145/179) in mSOF, respectively. By contrast, the development rate of activated oocytes to stage was greater (P<0.05) for oocytes cultured in mSOF medium (23.4%, 34/145) than in M16 or TCM-199 (0.0%). Our results suggest that source of oocytes and culture conditions are major factors affecting in vitro development of caprine parthenogenetic oocytes.

THE EFFECTS OF VARIOUS TETRACYCLINE HCL CONCENTRATION TREATED ROOW SURFACES ON PROLIFERATION AND SPREADING OF PERIODONTAL LIGAMENT CELLS (다양한 농도의 테트라사이클린로 처리된 치근면이 치주인대세포의 증식과 전개에 미치는 영향)

  • Jung, Oh-Chul;Sun, Jo-Young
    • Journal of Periodontal and Implant Science
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    • v.24 no.3
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    • pp.581-596
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    • 1994
  • This in vitro study was undertaken to obtain optimal tetracycline concentration that aids proliferation and spreading of human periodontal ligament cells, for clinical application in root surfaces of periodontally diseased teeth. Periodontal ligament cells used in this study were obtained from explants of periodontal ligament of 1st premolar teeth which were extracted for the purpose of orthodontic treatment. The cells were cultured in Dulbecco's Modified Eagle Medium(DMEM) supplemented with 100 U/ml penicillin, $100\;{\mu}g/ml$ streptomycin and 10% FBS at $37^{\circ}C$, 100% humidity, 5% $CO_2-95%$ air. Cells were used between the third to 4th passage. After root planing of periodontally extracted teeth, the root slabs were cut with carborundum disk. In the cell proliferation experiment, experimental groups were root planing only group, immersed groups in 25, 50, 75, 100, 150mg/ml aqueous solution of Tetracycline HCl followed by a vigorous rinse in PBS. Human PDL cells at concentration of $1{\times}10^5\;cells/ml$ were seeded in each culture well which contained root slabs and incubated for 6 hours. Then, all of the root slabs were moved into new 24 culture well and incubated 24, 48 and 72 hours. The cell counting was done by inverted phase contrast microscope after trypsinization. The following results were obtained. The cell number was increased in order root planing only group, 25, 150, 50, 75, 100mg/ml of Tetracycline HCl treated group in 24, 48 and 72 hours. The maximal cell number was obtained when the root slabs were immersed in solution with 100mg/ml of Tetracycline HCl. There were statistically significant between the root planing only group and 75, 100 mg/ml of Tetracycline HCl treated group in 24 hours, between the root planing only group and 100mg/ml of Tetracycline HCl treated group in 48 hours, between the root planing only group and 50, 75, 100mg/ml of Tetracycline HCl treated group, between 25 and 100mg/ml of Tetracycline HCl treated group in 72 hours(p<0.05). In the cell spreading experiment, after 30 minutes of incubated, in the root planing only group, the cells were generally round in shape. The cell surface was mostly covered with blebs. The cells started to attach to root surface by cytoplasmic extension in 50, 100mg/ml of Tetracycline HCl treated groups, more numerous cells attached to root surface than root planing only group. Many orifices of dentinal tubule were exposed, cells showed radially spreaded cytoplasm and unspreaded central region of the cell was covered with blebs. After 6 hours of incubation, in the root planing only group, cells showed radially spreaded cytoplasm and were attached flat appearance. In 50, 100mg/ml of Tetracycline HCl treated groups, cellular margin was concaved and cytoplasm showed elongated appearance with polarity. After 24 hours of incubation, in the root planing group, cells showed characteristic polarity. In 50, 100mg/ml of Tetracycline HCl treated groups, cells showed more elongated and spindle - like appearance.

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The Effect of TGF-{\beta}_1 on Cellular Activity of Periodontal Ligament Cells activated by PDGF-BB (PDGF-BB에 의한 치주인대세포활성에 대한 TGF-{\beta}의 효과)

  • Baek, Sang-Churl;Park, Jin-Woo;Suh, Jo-Young
    • Journal of Periodontal and Implant Science
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    • v.32 no.3
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    • pp.457-473
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    • 2002
  • The purposes of this study is to evaluate the combination effects of TGF-${\beta}_1$ and PDGF-BB on the periodontal ligament cells to use 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/100% FBS at the $37^{\circ}C$, 5% $CO_2$ incubator. Authors measured the DNA synthesis, total protein, collagen and noncollagenous protein synthesis according to the concentration of TGF-${\beta}_1$,(1,5ng/ml) and PDGF-BB (1,10 ng/ml) in combination. To explore further this delayed effect of TGF-${\beta}_1$, we preincubated human periodontal ligament cells with TGF-${\beta}_1$ for 4 or 24 hours before PDGF-BB stimulation. The results were as follows: The DNA synthetic activity was increased dose dependently by TGF-${\beta}_1$, PDGF-BB. The combination of TGF-${\beta}_1$ and PDGF-BB consistently enhanced the DNA synthetic activity to PDGF-BB alone. The ability of TGF-${\beta}_1$ to enhance DNA synthetic activity in PDGF-BB treated periodontal ligament cells was dose dependent. The maximum mitogenic effect was at the 5ng/ml of TGF-${\beta}_1$ and l0ng/ml of PDGF-BB. Preincubation of cell with TGF-${\beta}_1$ resulted in significantly greater response to PDGF-BB at all TGF-${\beta}_1$ concentration studied, and may be useful for clinical application in periodontal regenerative procedures. The total protein, collagen and noncollagen synthesis was increased dose pendently by TGF-${\beta}_1$, PDGF-BB. The % of collagen was slightly decreased according to the concentration of TGF-${\beta}_1$, PDGF-BB. The effect of TGF-${\beta}_1$, PDGF-BB were not specific for collagen synthesis since it also increased noncollagenous protein synthesis. This study demonstrates that PDGF-BB is major mitogens for human periodontal ligament cells in vitro, and supports a role for TGF-${\beta}_1$ as a regulation of the mitogenic and total protein formation to PDGF-BB in these cells.

Documentation of Intangible Cultural Heritage Using Motion Capture Technology Focusing on the documentation of Seungmu, Salpuri and Taepyeongmu (부록 3. 모션캡쳐를 이용한 무형문화재의 기록작성 - 국가지정 중요무형문화재 승무·살풀이·태평무를 중심으로 -)

  • Park, Weonmo;Go, Jungil;Kim, Yongsuk
    • Korean Journal of Heritage: History & Science
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    • v.39
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    • pp.351-378
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    • 2006
  • With the development of media, the methods for the documentation of intangible cultural heritage have been also developed and diversified. As well as the previous analogue ways of documentation, the have been recently applying new multi-media technologies focusing on digital pictures, sound sources, movies, etc. Among the new technologies, the documentation of intangible cultural heritage using the method of 'Motion Capture' has proved itself prominent especially in the fields that require three-dimensional documentation such as dances and performances. Motion Capture refers to the documentation technology which records the signals of the time varing positions derived from the sensors equipped on the surface of an object. It converts the signals from the sensors into digital data which can be plotted as points on the virtual coordinates of the computer and records the movement of the points during a certain period of time, as the object moves. It produces scientific data for the preservation of intangible cultural heritage, by displaying digital data which represents the virtual motion of a holder of an intangible cultural heritage. National Research Institute of Cultural Properties (NRICP) has been working on for the development of new documentation method for the Important Intangible Cultural Heritage designated by Korean government. This is to be done using 'motion capture' equipments which are also widely used for the computer graphics in movie or game industries. This project is designed to apply the motion capture technology for 3 years- from 2005 to 2007 - for 11 performances from 7 traditional dances of which body gestures have considerable values among the Important Intangible Cultural Heritage performances. This is to be supported by lottery funds. In 2005, the first year of the project, accumulated were data of single dances, such as Seungmu (monk's dance), Salpuri(a solo dance for spiritual cleansing dance), Taepyeongmu (dance of peace), which are relatively easy in terms of performing skills. In 2006, group dances, such as Jinju Geommu (Jinju sword dance), Seungjeonmu (dance for victory), Cheoyongmu (dance of Lord Cheoyong), etc., will be documented. In the last year of the project, 2007, education programme for comparative studies, analysis and transmission of intangible cultural heritage and three-dimensional contents for public service will be devised, based on the accumulated data, as well as the documentation of Hakyeonhwadae Habseolmu (crane dance combined with the lotus blossom dance). By describing the processes and results of motion capture documentation of Salpuri dance (Lee Mae-bang), Taepyeongmu (Kang seon-young) and Seungmu (Lee Mae-bang, Lee Ae-ju and Jung Jae-man) conducted in 2005, this report introduces a new approach for the documentation of intangible cultural heritage. During the first year of the project, two questions have been raised. First, how can we capture motions of a holder (dancer) without cutoffs during quite a long performance? After many times of tests, the motion capture system proved itself stable with continuous results. Second, how can we reproduce the accurate motion without the re-targeting process? The project re-created the most accurate motion of the dancer's gestures, applying the new technology to drew out the shape of the dancers's body digital data before the motion capture process for the first time in Korea. The accurate three-dimensional body models for four holders obtained by the body scanning enhanced the accuracy of the motion capture of the dance.

STUDIES ON THE DIMORPHISM AND FERTILITY OF PERSICARIA JAPONICA (MEISSNER) GROSS ET NAKAI (Persicaria Japonica (MISSNER) Gross et Nakai의 이형화와 수정력에 관한 연구)

  • HARN, Chang Yawl
    • Journal of Plant Biology
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    • v.3 no.1
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    • pp.1-15
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    • 1960
  • HARN, Chang Yawl : Studies on the dimorphism and Fertility of Persicaria japonica (MEISSNER) Gross et Nakai. Kor Jour. Bot. 3(I) 1-15 1960 Numerous investigations, since the works of DARWIN, have been made regarding the heterostylous plants by JOST (1907), CORRENS (1924), LAIBACK (1924), LEWIS (1943), and many others. Studies on the heterostylous Polygomum, however, were not reported except for the buckwhent, Fagopyrum esculentum, which was investigated by SCHOCH-BODMER (1930), EAST (1934), FROLOVA & Co-Workers (1946), MORRIS (1947, 1951) TATEBE (1949, 1951, 1953), present author (1957), and others. It is because no heterostylous species, besides buckwheat, have been known to exist in the Polygonum family. The author, during his studies on both heterostylism and fertility of Polygonaceae, has found that the species, persicaria japonica (Meissner) Gross et Nakai, is not diecious as has been known in taxonomy, but in reality beterostylous both morphologically and physiologically. It was found that this plant, regarded by taxonomist, as a male plant setting no seed, actually set seed (botanical fruit) when legitimate combination was made. Since his brief report on the dimorphic phenomens of this plant in 1956, the author's further research on the manner of fertilization has revealed that this species is a peculiar type whose dimorphism has undergone extreme specialization structurally and physiologically, the short-styled individual behaving in nature as a male plant and the long-styled individual, as female, whereas in controllled pollination the plant shows highly differentiated typical dimorphism. When compared with the other dimorphous species of this family, F. esculentum and P. sentiosa. it has been clarified that these three species differ in the degree of differentiation of their dimorphism morphologically and physiologically. That is, P. japonica has developed such a high specialization as to mislead the taxonomists, while P. senticosa shows almost no noticeable difference between long- and shortstyled individuals retaining most of the inherent physiological character cmmon to the genus except for the fact that it has two forms of flowers. F. esculentum appears to have taken the intermediate position in every respect. The result obtained in the present experiment are summarized as follows: 1) P. japonica has two kinds of individuals, one long style-short stamened; the other, short style-long stamened. The floral structure of this plants shows typical characteristics of dimorphic heterostylism. The differentiation between the two forms of flower has proceeded so highly both in primary and secondary difference of flower structure that this may be regarded as the most specialized form of dimorphism. 2) The differences of floral structure between the long and short styled individuals are remarkable compared with the other dimorphic species of the family. 3) The stamens of long styled plants show the sign of deteriolation whereas those of the short styled flower are well-developed. 4) When legitimate combinations are made, both L- and S-styled individuals are fertilized well and set seed (fruit), while in the illegitimate combination no fertilization and seed setting occur. Physiologically this species exhibits the typical behavior of dimorphic plants. 5) The self-fertile character, so common in other species of the other non-heterostyle Polygonum family, has disappeared completely. 6) Under natural conditions, no or few seed setting is observed in short styled individuals that behave as if they were male plants. 7) In hand pollination, the combination of both $L{\times}S$ and $S{\times}L$ alike yield relatively good fertility and seed-formation, the behavior of short styled individuals in artificial pollination differing remarkably from that in nature. 8) Under controlled pollination, $L{\times}S$ combination sets far more seed than in the combination of $S{\times}L$. In the S-styled individuals, the fertilized flower has the tendency of its seed more readily falling off in every stage of seed development than in the L-styled individuals. 9) The behaviors of pollen tubes just parallels the results of fertility test. That is, in the illegitimate combination, L-selfed, $L{\times}L$, S-selfed, and $S{\times}S$, the growth of pollen tubes is checked in the style, while in legitimately combined $L{\times}S$ and $S{\times}L$, the pollen tubes grow well reaching the ovaries within 40-50 minutes after pollination. The response of short styled individuals, known as male plant among taxonomists, is identical, as far as behavior fo pollen tube growth and fertilization are concerned, to that of long styled individuals, the so-called female plant. 10) The pollen grains from the short-styled plants are complete and fertile, whereas 70% of those of L-styled are found to be abortive, i.e., empty contents. 11) The remaining 30% of pollen of L-plant shows varied degree of stainability when stained with iron-aceto-carmine......mostly light red, while the pollen grains of S-style individuals are dark brown indicating complete fertility and viability. 12) The abundance of sterile pollen in L-styled and the nature of seed-dropping which occurs in S-styled individuals appear to be the main causes why the short styled individuals bear no seed in nature. Under controlled legitimate union, $S{\times}L$, the careful and elaborate pollination would give the S-styoled flowers the opportunities to receive the fertile pollens, though few in number, from L-styled plant, thus enabling S-plant to bear seed. 13) This species is not dioecious as is regarded by taxonomists, but typical dimorphic plant which has so highly specialized in floral structures and funcitons that the long-styled plant behaves just like a female individual; and the short-styled, like a male.

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