- The Effect of Sodium Butyrate (SB), Halofuginone Hydrobromide (HH) and High Glucose Concentration on Cell Growth and Gene Expression in Human Aortic Smooth Muscle Cell.
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June Ho Park, Sang Jun Lee, Yeo Jun Ki, Young Jung Jeon, Ki Young Kwon, Hong Suk Song, Seung Beom Han, In Kyu Lee, Jung Chul Kim
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J Korean Endocr Soc. 2000;15(2):272-285. Published online January 1, 2001
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- BACKGROUND
Vascular smooth muscle cell (VSMC) proliferation associated with arterial injury causes restenosis, which remains to be resolved in cardiovascular disease, especially after balloon angioplasty. Although numerous factors including hyperglycemia, hyperinsulinemia, angiotensin, basic fibroblast growth factor (BFGF), etc are suggested as potent mitogens for VSMCs, other mechanisms are still needed to take into new consideration. Advances in molecular biology have led to the development of powerful methods for the analysis of differential gene expression. There, we clarified the effect of glucose, sodium butyrate and halofuginon hydrobromide on gene expression which play a role in VSMC growth. METHODS: Therefore, we evaluate the changes of gene expression in response to high glucose concentration, sodium butyrate which is an inhibitor of platelet-derived growth factor (PDGF), and halofuginon hydrobromide which is an inhibitor of specific type 1 collagen, using differntial expressed sequence tag (EST) sequencing and cDNA microarray hybridization. Human mammary artery VSMC isolated from patients undergoing coronary bypass surgery. Cells from passage 3 to 5 were used in experiment with serum-free media with varying conditions. RESULTS: After 6 days of culture, the cells (VSMC) were resuspended with PBS and counted in a hemocytometer, and viable cells were counted using the trypan blue test. VSMC number reached 36?04 cell under high glucose concentration (H/G: 22mM) and 29?04 cell under low glucose concentration(L/G: 5.5 mM) at 6 day of culture (p<0.01). Sodium butyrate(SB) inhibited VSMC growth at varying butyrate concentrations (0.625, 1.25, 2.5, 5.0, 10.0mM) by 84%, 87%, 94%, 96%, 98%, respectively. Halofuginon hydrobromide(HH) also inhibited VSMC growth at varying halofuginon concentrations (10-11, 10-9, 10-7, 10-5mM) by 15%, 30%, 85%, 100%, respectively. Using a differential EST screening technique to assay the relative level of expression of each of large numbers of cloned cDNA sequences after treatment with high glucose concentration (22mM), sodium butyrate (5 mM), and halofuginon (1microM). Among the total 1,730 cDNA clones, 6 cDNA clones were down-regulated after treatment with sodium butyrate (5mM) and halofuginon (1microM). Those were revealed homology to genes encoding connective tissue growth factor (cTGF), Betaig-H3, nm23-H1 nm23-H2, enigma and copine 1. On the contrary, four clones were up-regulated after treatment with high glucose concentration (22mM). Those clones (BO94-5, K1316-5, K1764-5, B1835-5) didn't match any sequence in the public data base. CONCLUSION: These results indicate that this EST analysis is useful technique in targeting genes which are associated with atherosclerosis in VSMC. These identified clones may be used to assist in the positional cloning of genes which are related with atherosclerosis.
- Analysis of Cytokine Gene Expression in Thyroid Aspirates and Peripheral Blood Mononuclear Cell and in vitro Production of Interferon - Gamma by Peripheral Blood Mononuclear Cell Culture.
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Sung Rae Cho, Keun Yong Park, Young June Jeon, Hong Suk Song, Ki Young Kwon, Yun Nyun Kim, Seung Beom Han, In Kyu Lee
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J Korean Endocr Soc. 1995;10(1):13-25. Published online November 6, 2019
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- Cytokine production was studied in thyroid fine needle aspirates and peripheral blood and the production of interferon-gamma by peripheral blood mononuclear cell(PBMC) culture in response to interleukin-2(IL-2) stimulation was also studied from patients with hyperthyroidism, non toxic goiter, thyroid nodule. The expression of glycer aldehyde 3-phosphate dehydrogenase(GAPDH), interleukin-1beta(IL-1beta), IL-2, interleukin-8(IL-8), platelet- derived growth factor-A(PDGF-A) and interferon-gamma(IFN-gamma) chain was assessed by RT-PCR(reverse transcriptase polymerase chain reaction) in fine needle aspirates of thyroid and peripheral blood mononuclear cell : the samples were obtained from 7 patients with hyperthyroidism, 6 patients with non toxic goiter, 7 patients with thyroid nodule. A dose of IL-2(25 U/ml) was utilized to induce IFN-gamma production by PBMC from all patients.The results were as follows:1) In case of cytokine expression of fine-needle aspirates, GAPDH and IL-1beta, IL-8 were expressed highly but IFN-gamma, IL-2 were not expressed in hyperthyroidism and non-toxic goiter, thyroid nodule. PDGF-A was expressed in hyperthyroidism and thyroid nodule but not in non toxic goiter. 2) In case of cytokine expression of PBMC, GAPDH, IL-1beta were expressed in hyperthyroidism and non toxic goiter, thyroid nodule and highly expressed after IL-2 stimulation than before. But PDGF-A was more expressed in non toxic goiter and thyroid nodule than hyperthyroidism. Also, IFN-gamma was less expressed in thyroid nodule than hyperthyroidism and non toxic goiter. 3) The incremental increase in IFN-gamma value in supernatants of PBMC culture was significantly higher in patients with non toxic goiter than that in PBMC from hyperthyroidism and thyroid nodule(p<0.05).Therefore it seems that the cytokine production was found in hyperthyroidism and non toxic goiter and thyroid nodule. There were variability in their distribution each other, in general, higher expressed in hyperthyroidism than non toxic goiter. And RT-PCR Method that employed should be sufficiently sensitive to permit the analysis of cytokine gene expression in fine needle aspiration biopsies from patients with thyroid disease.
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