• Title/Summary/Keyword: [$^3$H]thymidine incorporation

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THE EFFECT OF THE GENISTEIN ON THE PROLIFERATION OF HT1080 AND EXPRESSION OF MEMBRANE TYPE 1-MATRIX METALLOPROTEINASE (MT1-MMP) mRNA (Genistein이 사람 섬유육종 세포주 증식 및 Membrane Type 1-Matrix Metalloproteinase (MT1-MMP) mRNA 발현에 미치는 영향)

  • Kang, Jin-Han;Myoung, Hoon;Kim, Myung-Jin
    • Journal of the Korean Association of Oral and Maxillofacial Surgeons
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    • v.27 no.4
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    • pp.314-320
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    • 2001
  • Matrix metalloproteinases have long been viewed as ideal candidates for proteinases that enables tumor cells to permeated basement membrane defenses and invade surrounding tissue. There is growing evidence that the MMPs have an expanded role, as they are important for the creation and maintenance of a microenvironment that facilitates growth and angiogenesis of tumors at primary and metastatic sites. MT-MMPs are not secreted but instead remaining attached to cell surfaces. Although not all of the MT-MMPs are fully characterized, MT-MMPs have important role in localizing and activating secreted MMPs. The MMP genes are transcriptionally responsive to a wide variety of oncogene, growth factors, cytokine, and hormones. Currently, a number of MMP inhibitors are being developed and some have reached clinical trials as anti-metastatic or anti-cancer therapies. MT1-MMP is involved in the activation of proMMP-2. MT1-MMP is significant not only as a tumor marker but as a new target for chemotherapy against cancer. The purpose of this study was to evaluate the effects of protein kinase C inhibitor(genistein) on the proliferation of HT1080 and expression of MT1-MMP mRNA. Human fibrosarcoma cell line HT1080 was cultured and divided 2 groups. The experimental group was treated with $100{\mu}M$ genistein and incubated 12h, 24h for $[3^H]-thymidine$ uptake assay and northern hybridization individually. And the control group was treated with same amount of PBS for the above procedures. $[3^H]-thymidine$ incorporation was measured with ${\beta}$ ray detector. And RT-PCR and northern blotting for MT1-MMP mRNA was performed. The results were as follows 1. $[3^H]-thymidine$ uptake was reduced in experimental group with statistical significance. 2. MT1-MMP mRNA expression was significantly reduced in experimental group. These results showed that protein kinase C inhibitor (genistein) inhibited proliferation of HT1080 and almost completely blocked transcription of MT1-MMP mRNA. So, it is possible to use the protein kinase inhibitor (genistein) as anti-metastatic and anti-proliferative agent.

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THE EFFECT OF STATIC MAGNETIC FIELDS ON MOLECULAR AND CELLULAR ACTIVITIES (정자기장이 효소와 세포 활성에 미치는 영향)

  • Park, Jae-Gu;Hwang, Hyeon-Shik
    • The korean journal of orthodontics
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    • v.27 no.6 s.65
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    • pp.929-941
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    • 1997
  • Optimal force for orthodontic treatment is the force that produces a rapid rate of tooth movement without discomfort to the Patient or ensuing tissue damage. Recently considerable interest has been generated in the application of magnets as a way to obtain an optimal force. The purpose of the present study was to investigate the effect of static magnetic fields of Sm-Co magnets on molecular and cellular activities. The distance of erythrocyte sedimentation was measured directly, and the activities and the syntheses of $Fe^{2+}$-related enzymes (catalase and NO synthase) and non $Fe^{2+}$-related enzyme (lactic dehydrogenase) were assayed by the spectrophotometer. The growth and the proliferation of osteoblast-like cells $MC_3T_3-E_1$ were determined by the crystal violet staining and the ${^3}H$-thymidine incorporation. The erythrocytes were exposed to the pole face flux density of 1,400 G (gauss), and the enzymes and osteoblast-like cells $MC_{3}T_3-E_1$ were exposed to the flux density of 7,000 G. The results obtained were as follows: 1. The distance of sedimentation of erythrocyte was not affected by the static magnetic fields. 2. The activities of catalase and lactic dehydrogenase were not affected by the static magnetic fields. 3. The intracellular syntheses of NO synthase and lactic dehydrogenase were not affected by the static magnetic fields. 4. The growth and the proliferation of cultured osteoblast-like cells $MC_{3}T_3-E_1$ were not affected by the static magnetic fields. These results suggested that the molecular and cellular activities were not significantly influenced by the static magnetic fields.

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THE EFFECT OF SODIUM FLUORIDE AND SODIUM ORTHOVANADATE ON OSTEOBLASTIC CELL LINE MC3T3-E1 CELLS (Sodium fluoride와 Sodium orthovanadate가 조골세포주 MC3T3-E1에 미치는 영향에 관한 연구)

  • Kim, Won-Jin;Chung, Kyu-Rhim
    • The korean journal of orthodontics
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    • v.21 no.1 s.33
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    • pp.97-111
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    • 1991
  • It is the aim of this study to investigate the effects of sodium fluoride and sodium orthovanadate upon the proliferation and activity of the osteoblast (MC3T3-E1 cells). MC3T3-E1 cells were cultured in $\alpha-MEM$ containing $10\%$ FBS and various concentration of sodium fluoride and sodium orthovanadate was appended to serum free media. DNA synthesis was examined through the $[^3H]$ thymidine incorporation into DNA. Collagen synthesis was examined through the $[^3H]$ proline incorporation into collagenase digestible protein and noncollagen protein. The following results were drawn; 1. Sodium fluoride stimulated the DNA synthesis of osteoblast significantly in dose-dependent manner within the concentration from $2{\mu}M$ to $10{\mu}M$ (P < 0.005). 2. Sodium orthovanadate stimulated the DNA synthesis of osteoblast significantly in dose-dependent manner within the concentration from $2{\mu}M\;to\;8{\mu}M$, however showed diminution at $10{\mu}M$ (P < 0.001). 3. Sodium fluoride and sodium orthovanadate stimulated the percent collagen synthesis of osteoblast significantly in dose-dependent manner within the concentration from $5{\mu}M$ to $10{\mu}M$ (P < 0.001). 4. Sodium fluoride and sodium orthovanadate stimulated the noncollagen synthesis of osteoblast significantly in dose-dependent manner within the concentration from $5{\mu}M\;to\;10{\mu}M$ (P < 0.001). In conclusion, sodium fluoride and sodium orthovanadate stimulate the proliferation and activity of osteoblast by stimulation of DNA synthesis and collagen and noncollagen synthesis in osteoblast.

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Hepatoprotective Activity of Salviae miltiorrhizae Radix Extract (단삼(丹蔘) 엑기스의 간보호작용)

  • Eun, Jae-Soon;Lim, Jong-Pil;Park, Yi-Kyu;Yeom, Jeong-Ryeol;Choi, Dong-Seong;Ahn, Moon-Seng
    • Korean Journal of Pharmacognosy
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    • v.22 no.2
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    • pp.95-100
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    • 1991
  • Salviae miltiorrhizae(SM) Radix extract increased $[^3H]-thymidine$ incorporation into rat hepatocytes at the concentration ranging from $5{\times}10^{-5}\;to\;5{\times}10^{-1}mg/ml$. It decreased the activities of s-GOT and s-GPT in cirrhotic rats induced by $CCl_4$, TAA and D-GalN, respectively and reduced the sleeping time induced by hexobarbital in $CCl_4$,TAA and D-GalN intoxicated mice, respectively. SM extract shortened the half-life of sulfobromophthalein in $CCl_4$ intoxicated rats.

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A Microtechnique for Mixed Lymphocyte Culture in the Sheep (면양(緬羊)의 혼합임파구배양(混合淋巴球培養)의 Micro-technique 정립(定立)에 관(關)한 연구(硏究))

  • Jun, Moo-Hyung
    • The Journal of the Korean Society for Microbiology
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    • v.16 no.1
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    • pp.65-70
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    • 1981
  • A microculture technique for the mixed lymphocyte culture test(MLC) in the sheep is described. The optimal incubation periods for the MLC are varied from 4 to 6 days depending on the sources of lymphocytes. It is found that a 2.5:1 stimulator-responder cell ratio and the responder cell concentration of $1{\times}10^5$ cells/well produce the highest($^3H$)-thymidine incorporation. Moreover cryopreservation of bovine lymphocytes results in satisfactory and reproducible MLC reaction. It is also evident that the MLC reactions of the sheep are under the control of histocompatibility matching between the stimulator cells and the responder cells. Significance of the technique as a too! for study on cell mediated immunity in sheep system, either normal or squamous cell carcinoma-bearing, are discussed.

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Effects of Polyamines on DNA Synthesis in Nicotiana tabacum L. Suspension Cultured Cells (담배(Nicotiana tabacum L.) 현탁배양 세포에서 DNA 합성에 미치는 Polyamine의 효과)

  • 남경희
    • Journal of Plant Biology
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    • v.36 no.1
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    • pp.19-27
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    • 1993
  • Effects of polyamines on DNA synthesis were studied in synchronized culture of Nicotiana tabacum L. When DFMO and DFMA, inhibitors of ornithine decarboxylase and arginine decarboxylase, respectively were initially applied to the cells, the polyamine contents were rapidly dropped and [methyl-3H] thymidine incorporation into DNA was markedly reduced during the early stage of culture period. Inhibition of DNA synthesis, however, was partially reversed when these inhibitors were applied simultaneously with putrescine. In addition, exogenous administration of putrescine also increased the DNA synthesis during the all over the culture period. In vitro activity of DNA polymerase from Nicotiana tabacum L. was promoted by increasing concentrations of polyamines in the reaction mixture. Maximal activity was shown at 5 mM putrscine, 0.5 mM spermidine and spermine, respectively. Lack of Mg2+ ion in the reaction buffer resulted in an inhibition of the enzyme activity by about 30%. The inhibition could not be completely reversed by application of polyamines at optimal concentrations. These results suggest that polyamines promote the DNA synthesis in vivo and in vitro by stabilizing the DNA-helix upon binding to negatively charged groups on DNA or increasing the activity of DNA polymerase in Nicotiana tabacum L.

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Toxicological Evaluation of Medicinal Plants Used for Herbal Drugs (II) -Acute Toxicity and Effects on DNA Biosynthesis in Bone Marrow Cells and Hemoglobin Content in Blood- (한국산 생약의 약리작용 및 독성연구 (제2보) -급성 독성 및 골수세포의 DNA생합성에 미치는 영향-)

  • Chang, Il-Moo;Kim, Young-Soo;Han, Byung-Hoon
    • Korean Journal of Pharmacognosy
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    • v.13 no.1
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    • pp.14-19
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    • 1982
  • Potential toxicity of 15 medicinal plants used for herbal drugs, which were also described as being tonic for hematopoietic system or being toxic for the system in a oriental book 'Dong Ee Bo Gam', were evaluated in mice. Six plants among 15 plants tested appeared to exhibit acute toxicity along with bone marrow depression or with abnormally enhancing the $^3H-thymidine$ incorporation into DNA biosynthesis in bone marrow cells. Six plants were Paeonia albiflora, Pharbitis nil, Cemphalia lapidescens, Scutellaria baicalensis, Akebia quinata and Glycyrriza uralensis.

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Kaempferol inhibits the platelet-derived growth factor $\beta$-receptor tyrosine-phosphorylation and its downstream intracellular signal transduction pathway in rat aortic vascular smooth muscle cells

  • Kim, Soo-Yeon;Kim, Jin-Ho;Lim, Yong;Yoo, Hwan-Soo;Yun, Yeo-Pyo
    • Proceedings of the PSK Conference
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    • 2003.10b
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    • pp.108.2-108.2
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    • 2003
  • Kaempferol, a flavonol compound, has been reported as the anti-oxidant and anti-angiogenic agent and it has been found to inhibit cell growth in vitro. Abnormal proliferation of vascular smooth muscle cells (VSMCs) plays an important role in development of atherosclerosis. In this study, we examined the anti-proliferative effect and its mechanism on rat aortic VSMCs treated by kaempferol. kaempferol significantly inhibited the platelet-derived growth factor (PDGF)-BB-induced proliferation of rat aortic VSMCs in concentration-dependent manner by cell count and [$^3$H]-thymidine incorporation assay. (omitted)

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Role of Integrin, FAK (Focal Adhesion Kinase) and ERK (Extracellular Signal Regulated Kinase) on the Suppressed Cell Proliferation of Endometrial Cancer Cells by GnRH (Gonadotropin-Releasing Hormone) (GnRH (Gonadotropin-Releasing Hormone)에 의한 자궁내막암 유래 세포주의 세포 증식 억제 기전에 있어서 Integrin, FAK (Focal Adhesion Kinase) 및 ERK (Extracellular Signal Regulated Kinase)의 역할)

  • Choi, Jong Rak;Park, Dong Wook;Choi, Dong Soon;Min, Churl K.
    • Clinical and Experimental Reproductive Medicine
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    • v.33 no.2
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    • pp.115-123
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    • 2006
  • Objective: To investigate new signal transduction cascade through integrin, FAK and ERK in the suppressed cell proliferation by GnRH-I and -II. Method: Human endometrial cancer cells (HEC1A) were cultured under the following condition: DMEM/F12 (10% FBS). GnRH-I and -II were treated time (0, 5, 10, 15, 20, 30 min; 100 nM) and dose (10 nM or 100 nM; 20 min) dependent manner according to experimental purposes. Cell proliferation was measured using [$^3H$] thymidine incorporation assay. Immunoblotting was utilized to detect proteins. Results: GnRH-I and -II inhibited proliferation of HEC1A cells and induced expression of integrin ${\beta}3$. Phosphorylation of FAK and ERK were induced by GnRH-I and -II. Conclusion: GnRH inhibited cell proliferation via the expression of integrin and FAK, ERK phosphorylation.

ppGalNAc T1 as a Potential Novel Marker for Human Bladder Cancer

  • Ding, Ming-Xia;Wang, Hai-Feng;Wang, Jian-Song;Zhan, Hui;Zuo, Yi-Gang;Yang, De-Lin;Liu, Jing-Yu;Wang, Wei;Ke, Chang-Xing;Yan, Ru-Ping
    • Asian Pacific Journal of Cancer Prevention
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    • v.13 no.11
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    • pp.5653-5657
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    • 2012
  • Objectives: To investigate the effect of glycopeptide-preferring polypeptide GalNAc transferase 1 (ppGalNAc T1 ) targeted RNA interference (RNAi) on the growth and migration of human bladder carcinoma EJ cells in vitro and in vivo. Methods: DNA microarray assays were performed to determine ppGalNAc Ts(ppGalNAc T1-9) expression in human bladder cancer and normal bladder tissues. We transfected the EJ bladder cancer cell line with well-designed ppGalNAc T1 siRNA. Boyden chamber and Wound healing assays were used to investigate changes of shppGalNAc T1-EJ cell migration. Proliferation of shppGalNAc T1-EJ cells in vitro was assessed using [3H]-thymidine incorporation assay and soft agar colony formation assays. Subcutaneous bladder tumors in BALB/c nude mice were induced by inoculation of shppGalNAc T1-EJ cells and after inoculation diameters of tumors were measured every 5 days to determine gross tumor volumes. Results: ppGalNAc T1 mRNA in bladder cancer tissues was 11.2-fold higher than in normal bladder tissues. When ppGalNAc T1 expression in EJ cells was knocked down through transfection by pSUPER-shppGalNAc T1 vector, markedly reduced incorporation of [3H]-thymidine into DNA of EJ cells was observed at all time points compared with the empty vector transfected control cells. However, ppGalNAc T1 knockdown did not significantly inhibited cell migration (only 12.3%). Silenced ppGalNAc T1 expression significantly inhibited subcutaneous tumor growth compared with the control groups injected with empty vector transfected control cells. At the end of observation course (40 days), the inhibitory rate of cancerous growth for ppGalNAc T1 knockdown was 52.5%. Conclusion: ppGalNAc T1 might be a potential novel marker for human bladder cancer. Although ppGalNAc T1 knockdown caused no remarkable change in cell migration, silenced expression significantly inhibited proliferation and tumor growth of the bladder cancer EJ cell line.