Kim, B. M.;Lee, M.;Lee, J. H.;I. S. Doo;M. K. Son;S. H. Kang;Kim, W. B.;J. W. Kwon
대한화장품학회:학술대회논문집
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대한화장품학회 2003년도 IFSCC Conference Proceeding Book I
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pp.659-659
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2003
In this study, we have investigated the potent anti-aging effect of DA-3711, a cosmetic ingredient derived from artificial skin culture. The artificial skin was originally developed as a skin replacement for the treatment of chronic skin wounds. To produce DA-3711, neonatal human fibroblasts were seeded into biocompatible collagen/chitosan/glycosaminoglycan (GAG) scaffolds and cultured in Dulbecco's modified Eagle's medium (DMEM) supplemented with fetal bovine serum and nonessential amino acids. Analysis of the culture broth (DA-3711) showed that growth factors such as VEGF, TGF-$\beta$, KGF were present at significantly higher levels that in the culture broth of fibroblasts cultured in monolayer. The biological activity of DA-3711 was assessed by measuring in vitro cell proliferation and collagen synthesis of normal human fibroblasts. Fibroblasts treated with 10% DA-3711 showed a 2-fold higher proliferation and 2 to 4-fold higher collagen synthesis than untreated cells. DA-3711 also exhibited anti-oxidative effects, since cells under peroxide-induced oxidative stress showed a 30% higher viability in DA-3711-containing medium than in medium without DA-3711 addition. The results suggest that DA-3711 may have anti-aging effects by stimulating skin regeneration and protecting against oxidative stress.
Myocardial infarction is one of the leading causes of mortality globally. Currently, the pleiotropic inflammatory cytokine interleukin-6 (IL-6) is considered to be intimately related to the severity of myocardial injury during myocardial infarction. Interventions targeting IL-6 are a promising therapeutic option for myocardial infarction, but the underlying molecular mechanisms are not well understood. Here, we report the novel role of IL-6 in regulating adverse cardiac remodeling mediated by fibroblasts in a mouse model of myocardial infarction. It was found that the elevated expression of IL-6 in myocardium and cardiac fibroblasts was observed after myocardial infarction. Further, fibroblast-specific knockdown of Il6 significantly attenuated cardiac fibrosis and adverse cardiac remodeling and preserved cardiac function induced by myocardial infarction. Mechanistically, the role of Il6 contributing to cardiac fibrosis depends on signal transduction and activation of transcription (STAT)3 signaling activation. Additionally, Stat3 binds to the Il11 promoter region and contributes to the increased expression of Il11, which exacerbates cardiac fibrosis. In conclusion, these results suggest a novel role for IL-6 derived from fibroblasts in mediating Stat3 activation and substantially augmented Il11 expression in promoting cardiac fibrosis, highlighting its potential as a therapeutic target for cardiac fibrosis.
This study was carried out to investigate cytotoxicity of several herbicides (Bentazone, Butachlor. Paraquat and Ethalfluralin) in cultured mouse NIH 3T3 fibroblasts. Tetrazolium (MTT), neutral red (NR) and sulforhodamine protein B (SRB) of the colorimetric assays were performed to evaluate the cytotoxicity on cell organelles. 2 x 10$^4$cell/$m\ell$ of NIH 3T3 fibroblast in each well of 24 multidish were cultured. After 24 hours, the cells were treated with solution (1, 25, 50 or 100 $\mu$M) of each herbicide. After the NIH 3T3 fibroblasts of all groups were cultured in the same condition for 48 hours, MTT, NR and SRB assays were performed to evaluate the cytotoxicity. The light microscopic study was carried out to examine morphological changes of cultured NIH 3T3 fibroblasts. The MTT$_{50}$ of Bentazone, Butachlor, Paraquat and Ethalfluralin were 1560.97 $\mu$M, 56.15 $\mu$M, 3138.81 $\mu$M and 1301.82 $\mu$M, respectively. The NR$_{50}$ of Bentazone, Butachlor. Paraquat and Ethalfluralin were 1763.93 $\mu$M, 45.98 $\mu$M, 1030.85 $\mu$M and 1808.29 $\mu$M, respectively. The SRB$_{50}$ of Bentazone, Butachlor. Paraquat and Ethalfluralin were 1913.38 $\mu$M, 65.30 $\mu$M, 1860.73 $\mu$M and 1086.93 $\mu$M, respectively. The morphological changes of NIH 3T3 fibroblasts showed severe degeneration in Butachlor 50 $\mu$M and 100 $\mu$M concentrations. These results indicate that Butachlor has high cytotoxicity, Bentazone, Paraquat and Ethalfluralin very weak cytotoxicity against NIH 3T3 fibroblasts.lasts.
Cyclosporine A is an immunosuppressant commonly used for patients receiving organ transplants. Gingival overgrowth is an adverse side-effect seen in about 8-26% of patients taking cyclosporine A which have been shown to increase the DNA synthesis of gingival fibroblast at the concentration of $10^{-9}g/ml$ in vitro. Glycyrrhetinic acid is the active pharmacological ingredients of licorice which exerts steroid-like action and anti-viral activity. Oleanolic acid, which were isolated from Glechoma hederacea, has been shown to act as inhibitors of tumor promotion in vivo and to be less cytotoxic retinoic acid. This study has been performed to evaluate the effects of glycyrrhetinic acid and oleanolic acid on cyclosporine A induced cell activity in vitro. Human gingival fibroblasts were isolated from explant cultures of healthy gingiva of orthodontic patients. Gingival fibroblasts were trypsinized and transferred to the walls of microtest plates. Fibroblasts were cultured in growth medium added $10^{-9}g/ml$ cyclosporineA and $50{\mu}l/ml$ lipopolysaccharides. Cells between the 4th and 6th transfer in culture were used for this study. The morphology of gingival fibroblst were examined by inverted microscope. The effects of cyclosporine A on the time course of DNA sythesis by human gingival fibroblasts were assessed by $[^3H]-thymidine$ uptake assays. Cyclosporine A was found to stimulate DNA synthesis of human gingival fibroblast at a concentration of $10^{-9}g/ml$. In the presence of lipopolysaccharide derived from Fusobacterium nucleatum, addition of cyclosporine A results in reversal of inhibition at the concentration which normally inhibits gingival fibroblast proliferation. The cell acitivities in the presence of glycyrrhetinic acid and oleanolic acid were decreased, and increased cell acitivities by cyclosporine A were decreased by glycyrrhetinic acid and oleanolic acid at the concentration of $200{\mu}g/ml$. These results suggested that the increased cell activities by cyclosporine A modulated by glycyrrhetinic acid and oleanolic acid.
Based on the data from our previous studies, four new diabetic remedies were composed with the addition of Coicis Semen into Okchun-san (OCH), Commelinae Herba into Gangsim-tang (GST) , Scrophulariae Radix into Sunki-san (SKS), and Erythrinae Cortex into Yukmijihuang-hwan (YMG). The water extracts of these new remedies were treated in 3T3-L1 fibroblasts and adipocytes in order to investigate insulin-like substances and insulin sensitizers, respectively. With and without differentiation inducers, unmodified SKS (SKS-O) treatment induced 3T3-L1 fibroblasts into adipocytes more than the control. However, without inducers, YMG treatment, but not SKS, induced the differentiation more than the control among modified remedies. Without inducers, SKS, OCH as well as YMG increased the induction of differentiation from 3T3-L1 fibroblasts into adipocytes, compared to the control. The treatment of OCH and YMG with 1 ng/mL insulin increased glucose uptake much more than only insulin 1 ng/mL treatment. Thus, OCH and YMG contained increased insulin actions. In conclusions, the modified remedies, OCH and YMG, contained insulin-like substances and insulin sensitizers, and they can be improved the hypoglycemic effects.
An experimental study was performed for the preparation of living skin-equivalent by the using collagen gel contraction with human fibroblasts as neodermls and cultured human keratinocytes as neoderm is . The results were as follows ; 1) The rate of collagen gel contraction was dependent on the number of fibroblasts into the lattice and collagen contraction was progressed according to the increment of the number of the cells. 2) The rate of collagen gel contraction was progressed according to the decrement of the contraction of the collagen. 3) The rate of gel contraction was progressed according to the increment of serum concentration in the fixed concentration of the fibroblasts and collagen. 4) The lattice contraction was decreased according to the increment of the population doublings of the fibroblasts. 5) Macroscopically, the artificial dermis was gray white in color and tissue-like consistency and elas- ticity. 6) Microscopically, three dimensionally contracted artificial dermis showed more dense fibroblasts and its newly formed collagen fibrils in the matrix than one dimensionally contracted one. 7) Finally prepared skin-equivalent showed good attachment of living stratified keratinocytes to the dermal equivalent microscopically. It has been proposed that newly formed skin-equivalent is suitable for the graft of extensively and deeply burned patients. Shortening of the manufacturing period of skin-equivalent and development of conservation technique as a readily usable state are to be solved for our ongoing works.
The growth and synthetic activities of fibroblasts are regulated by cytokines and growth factors derived from activated inflammatory cells. Stimulatory effect of low level laser (LLL) radiation on wound healing seems to be in part due to direct stimulatory action on cell proliferation and synthetic activities of fibroblasts. Also indirect stimulatory effect on the fibroblast function through inflammatory or immune cells is another possible mechanism of biostimulatory action of LLL. This study was performed to determine the growth rate of human gingival fibroblasts obtained biopsy and culture, fibroblast cell line, and immune cell line by using $[^3H]-$ thymidine incorporation test. And gene expression pattern was also analyzed by using the DNA probe such as Hsp70, IL-1$\beta$, MIP-1$\alpha$ and actin cDNA. Proliferation rate of gingival fibroblast was increased by LLL irradiation, but no more effect was added by LPS or IL-1$\beta$ pretreatment Enhanced Hsp70 gene expression was found from gingival fibroblasts and fibroblast cell line COS by LLL irradiation., which was not more increased by LPS or IL-1$\beta$ pretreatment. LLL-irradiated promyelcytic cell line HL-60 and macrophage cell line RAW264.7 showed significant stimulatory effect of proliferation rate when compared with respective control. However there were no changes in growth rate of other immune cell tested in this study, such as B cell line WR19n.l and 230, helper T cell line Jurkat and Hut78, cytolytic T cell line CTLL-r8. By LLL-irradiation Hsp70 gene expression was increased in RAW246.7 and HL-60, not in CTLL-R8. And IL-1$\beta$ and MIP-1$\alpha$ gene expression were induced only from LLL-irradiated RAW264.7. These results led us to presume that LLL radiation may affect to the immune cells, especially to macrophage, through which it might promote wound healing process.
Kim, Kwang-Mi;Noh, Min-Soo;Kim, Soo-Hyun;Park, Mi-Kyung;Lee, Hye-Ja;Kim, Soo-Youl;Lee, Chang-Hoon
Biomolecules & Therapeutics
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제18권1호
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pp.71-76
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2010
Skin irritation caused by retinol and retinoic acid results in mild erythema called as retinoid dermatitis. To develop compounds modulating the retinoid dermatitis, we tried to establish the screening method for retinoid dermatitis. At first we examined the inflammatory cytokine profile in neonatal human dermal fibroblasts which are known to be one of main site of retinoid action. As a result, interleukin-8 (IL-8) and monocytes chemoattractant protein-1 (MCP-1) were significantly produced by all trans retinoic acid (ATRA) and all trans retinol (ATROL) in dermal fibroblasts. Especially the production of MCP-1 was more than that of IL-8. The production of MCP-1 by retinoid was dose-dependently increased, continuing up to 24 hrs. After then using ethacrynic acid (ECA) known to reduce mouse ear edema induced by ATRA, we checked whether ECA suppressed the production of MCP-1. As a result, ECA effectively suppressed the production of MCP-1 in the ATRA- or ATROL-treated-fibroblasts. These results suggested that screening method effectively reflects the in vivo anti-inflammatory activity of ECA. It was reported that citral inhibited the enzyme involved in the conversion of ATROL to ATRA. We showed that citral suppressed the production of MCP-1 in ATROL-treated fibroblasts. We expect these finding might be helpful to find useful compounds modulating the side effects of retinoid or retinoid dermatitis.
Some therapeutic agents and medicaments may lead to pathologic changes in the gingival tissue, especially on the cultured human gingival fibroblasts. The purpose of this study was to investigate on the effect of diphenylhydantoin, retinoic acid to the human gingival fibroblast. Human gingival fibroblasts were cultured from the healthy gingiva of patients with orthodontic patients. Gingival fibroblasts were trypsinized and transferred to the wells of 96 well microtest plates. Next day, the medium was removed, fibroblasts were washed with HBSS, and the washed cells were cultured in growth medium added 5 or $10{\mu}g/ml$ of diphenylhydantoin, $10^{-5}M$, $10^{-6}M$ and $10^{-7}M$ of retinoic acid and glycyrrhetinic acid. The passage number of cultured fibroblasts were fifth and eighth. The cell morphology was examined by inverted microscope, the cell number was counted by hemocytometer, and cell activity was measured by the growth and proliferatiton assay using MTT assay. The fifth experiments were performed and statistical significance was measured by ANOVA. The cell morphology in the presence of retinoic acid was round irrespective of the presence of diphenylhydantoin and glycyrrhetinic acid(Fig 2-6). The proliferation of cells was not changed by diphenylhydantoin(Table 1). The cell activity showed the tendency to increase at the concentration of $10{\mu}$'/, of diphenylhydantoin (Table 2). The cell activity in the presence of retinoic acid glycyrrhetinic acid was decreased, and the increased cell activity by diphenylhydantoin was decreased by retinoic acid and glycyrrhetinic acid at the concentration of $10^{-7}M$(Table 3-5). These results suggested that the increased cell activity by diphenylhydantoin might be modulated by retinoic acid and glycyrrhetinic acid.
Purpose: Oncostatin M(OSM) is a multifunctional cytokine that belongs to the interleukin(IL)-6 family. Although there have been a number of studies that focused on the role and mechanism of OSM in various organs and tissues, there are few reports on its effect on wound healing. The final purpose of this project is to evaluate the effect of OSM on wound healing. This pilot study was designed to investigate the effect of OSM on proliferation and matrix synthesis of human dermal fibroblasts, which are the major components of the wound healing. Methods: Excess skin that was obtained from patients who underwent skin grafts, was used for this study. From this material, fibroblasts were isolated and cultured. The cultured fibroblasts were treated with one of four concentrations of OSM. The OSM concentrations used were 0, 50, 100, and 200 ng/ml, respectively. After the OSM treatment, cell proliferation was determined by the MTT assay, collagen synthesis by the C1CP method, GAG levels by the Blyscan Dye method. The parameter levels of each group were compared. Results: OSM treatment increased all the components tested in the study. In particular, cell proliferation, GAG synthesis demonstrated statistically significant increases(p<0.05 in the Mann-Whitney U-test). The highest increase in all the components was obtained at a 100 ng/ml concentration of OSM.Conclusion: The results of the present study indicate that OSM stimulates proliferation and matrix synthesis of human dermal fibroblast and the optimal concentration for wound healing is 100 ng/mL.
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