• Journal of Acupuncture Research
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• v.22 no.3
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• pp.169-183
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• 2005

Objective : The purpose of this study was to investigate the anti-caner effect of cobrotoxin on the prostatic cancer cell line (PC-3).The goal of study is to ascertain whether cobrotoxin inhibits tile cell growth and cell cycle of PC-3, or the expression of relative genes and whether the regression of PC-3 cell growth is due to cell death or the expression of gene related to apoptosis. Methods : After the treatment of Pc-3 cells with cobrotoxin, we performed 형광현미경, MTT assay, Western blotting, Flow cytometry, PAGE electrophoresis and Surface plasmon resonance analysis to identify the cell viability, cell death, apoptosis, the changes of cell cycle and the related protein, Adk, MAP kinase. Results : 1. Compared with normal cell, the inhibition of cell growth reduced in proportion with the dose of cobrotoxin(0-16nM) in PC-3. 2. Cell viabilities of 0.1, 1, 4nM cobrotoxin treatment were decreased and those of 8, 16nM were decreased significantly. 3. S phase of cell cycle was decreased at the group of 1, 2, 4, 8, 16nM cobrotoxin, but M phase was increased at 0.1, 1, 2, 4, 8, 16nM cobrotoxin. 4. Cox-2 expression after cobrotoxin was peaked at 12hours and was decreased significantly after 6, 12, 24 hours. 5. The expression of Cdk4 was decreased dose-dependently at 1, 2, 4, 8nM cobrotoxin and was decreased siginificantly at 4, 8nM Cyclin D1 was decreased at 1, 2, 4, 8nM and Cycline E was not changed. Cycline B was decreased at 1, 2, 4, 8nM dose-dependently and was decreased siginificanlty at 2, 4, 8nM. 6. The expression of Akt was decreased at 1, 2, 4, 8nM dose-dependently and was decreased significantly at 2, 4, 8nM. 7. ERK was increased at 1, 2nM and decreased at 4, 8nM, p-ERK was increased at 1, 2, 4 nM, but decreased at 8nM. JNK and p-JNK were increased at 1, 4, 8 nM. p38 was increased at 2nM p-p38 was increased at lnM but decreased significantly at 2, 4, 8nM. 8. The nucli of normal cells were stained round and homogenous in DAPI staining, but those of PC-3 were stained condense and splitted. Apoptosis was increased dose-dependently at 2, 4, 8, 16nM and increased significantly at 2, 4, 8, 16nM. 9. Bax wasn`t changed at 1, 2, 4, 8nM and Bcl-2 was decreased significantly at 1, 2, 4, 8nM. Caspase 3 and 9 weren`t changed at 1, 2, 4nM but were decreased significantly at 8nM. Conclusions : These results indicate that cobrotoxin inhibits the growth of prostate Cancer cells, has anti-cancer effects by inducing apoptosis.

• Korean Journal of Environmental Biology
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• v.24 no.1 s.61
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• pp.81-88
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• 2006

The cell abundance and marker pigment distribution patterns of picophytoplankton in the Chuuk Lagoon, tropical South Pacific, were analyzed flow cytometry and HPLC. Also, respective contribution of Synechococcus, Prochlorococcus and picoeukaryotes on estimated carbon biomass was evaluated. Synechococcus and Prochlorococcus showed contrasting distributional patterns in the waters of Chuuk Lagoon. Relatively high concentration of Synechococcus was observed near Weno Island but the concentration decreased toward the Northeast Passage. However, Prochlorococcus showed an opposite distributional pattern. Picoeukaryotes did not show any significant variable difference. The range of divinyl chlorophyll a (Chl. $\alpha$) concentration, marker pigment of Prochlorococcus, was $1.2\sim180.3\;ng\;L^{-1}$ and higher concentrations were observed at the stations near the Northeast Passage than stations near Weno Island. This pigment pattern was similar to cell abundance pattern indicating that chi. a2 may be a useful biomass indicator. On the other hand, the range of zeaxanthin concentrations was $61.4\sim135.8\;ng\;L^{-1}$ showing comparatively less significant variation indicating zeaxanthin influence derived from Prochlorococcus. Estimated carbon biomass of Synechococcus contributed 68% of total picophytoplankton biomass. Prochlorococcus and picoeukaryotes respectively contributed 17.1% and 14.9% of total picophytoplankton biomass.

• Korean Journal of Environmental Biology
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• v.24 no.1 s.61
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• pp.67-80
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• 2006

The effect of environmental forcing on picophytoplankton distribution pattern was investigated in the tropical and subtropical western Pacific (TSWP) and the East Sea in September, 2002, and the continental shelf of the East China Sea (C-ECS) in August, 2003. The abundance of picophytoplankton populations, Synechococcus, Prochlorococcus and picoeukaryotes were determined by flow cytometry analyses. Picophytoplankton vertical profiles and integrated abundance $(0\sim100\;m)$ were compared with these three physiochemically different regions. Variation patterns of integrated cell abundance of Synechococcus and Prochlorococcus in these three regions showed contrasting results. Synechococcus showed average abundance of $84.5X10^{10}\;cells\;m^{-2}$, in the TSWP, $305.6X10^{10}\;cells\;m^{-2}$ in the C-ECS, and $125.4X10^{10}\;cells\; m^{-2}$ in the East Sea where increasing cell concentrations were observed in the region with abundant nutrient. On the other hand, Prochlorococcus showed average abundance of $504.5X10^{10}\;cells\;m^{-2}$ in the TSWP, $33.2x10^{10}\;cells\;m^{-2}$ in the C-ECS, and $130.2X10^{10}\;cells\;m^{-2}$ in the East Sea exhibiting a distinctive pattern of increasing cell abundance in oligotrophic warm water. Although picoeukaryotes showed a similar pattern to Synechococcus, the abundance was 1/10 of Synechococcus. Synechococcus and picoeukaryotes showed ubiquitous distribution whereas Prochlorococcus generally did not appear in the C-ECS and the East Sea with low salinity environment. The average depth profiles for Synechococcus and Prochlorococcus displayed uniform abundance in the surface mixed layer with a rapid decrease below the surface mixed layer. for Prochlorococcus, a similar rapid decreasing trend was not observed below the surface mixed layer of the TSWP, but Prochlorococcus continued to show high cell abundance even down to 100 m depth. Picoeukaryotes showed uniform abundance along $0\sim100\;m$ depth in the C-ECS, and abundance maximum layer appeared in the East Sea at $20\sim30\;m$ depth.

• Journal of Acupuncture Research
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• v.22 no.3
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• pp.155-167
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• 2005

Objectives : The purpose of this study was to investigate the anti-inflammatory effect of Cobrotoxin on binding affinity of cobrotoxin with P50, $IKK{\alpa}$ and $IKK{\beta}$, activities of NF-${\kappa}B$, Cell viability of astrocyte, expressions of protein molecules of NF-${\kappa}B$ such as P50, P-$1{kappa}B$, $1{\kappa}B$ and iflammation related genes such as Cox-2, iNOS, cPLA2 in the SNP or LPS induced Inflammatory pathway of Rats' astrocytes. Methods : In this study, The expression of cytosolic phospholipase A2, Nitric oxcide, Cyclooxygenase-2 and inducible nitrogen oxide synthase was determined by western blotting with corresponding antibodies, and the generation of NF-${\kappa}B$ was assayed by EMSA method in astrocytes of rats. The Cell viability of astrocytes was determined by MTT assay, and Binding affinity of Cobrotoxin with P50, $IKK{\alpha}$ and $IKK{\beta}$ was assayed by Surface plasmon resonance analysis, and NF-${\kappa}B$ dependent luciferase activity was determined by luciferase analysis, and Uptake of cobrotoxin in astrocytes was identified by Confocal laser scanning microscope Results : 1. Compared with control, LPS-induced NF-${\kappa}B$ DNA binding activity was decreased significantly by 0.1, $0.5{\mu}g/m{\ell}$ of Cobrotoxin in Astrocyte. 2. Compared with control, LPS-induced NF-kB dependent luciferase expression was decreased significantly by 0.1, 0.5 and $1{\mu}g/m{\ell}$ of Cobrotoxin in Astrocyte. 3. Compared with control, SNP induced P50, $I{\kappa}B$ expressions in astrocyte were decreased significantly by 0.1, 0.5 and $1{\mu}g/m{\ell}$ of Cobrotoxin and P-$1{\kappa}B$ expression was decreased significantly by 0.5 and $1{\mu}g/m{\ell}$ of Cobrotoxin. 4. Compared with control, LPS induced P50, $1{\kappa}B$ expressions in astrocyte were decreased significantly by 0.5 and $1{\mu}g/m{\ell}$ of Cobrotoxin. 5. Compared with control, SNP induced Cox-2, iNOS, CPLA2 expressions in astrocyte were decreased significantly by $1{\mu}g/m{\ell}$ of Cobrotoxin. 6. Compared with control, LPS induced Cox-2, cPLA2 expressions in astrocyte were decreased significantly by 0.1, 0.5, $1{\mu}g/m{\ell}$ of Cobrotoxin and iNOS expression was decreased significantly by 0.5, $1{\mu}g/m{\ell}$ of Cobrotoxin. 7. Compared with $0.5{\mu}g/m{\ell}$ of Cobrotoxin, SNP-induced NF-${\kappa}B$ DNA bindins activity in astrocyte was increased significantly by Cobrotoxin $0.5{\mu}g/m{\ell}$ with DTT 1mM and Cobrotoxin $0.5{\mu}g/m{\ell}$ with DTT 5mM. 8. Compared with $0.5{\mu}g/m{\ell}$ of Cobrotoxin, LPS-induced NF-${\kappa}B$ DNA binding activity in astrocyte was increased significantly by Cobrotoxin $0.5{\mu}g/m{\ell}$ with DTT 1mM, Cobrotoxin $0.5{\mu}g/m{\ell}$ with DTT 5mM, Cobrotoxin $0.5{\mu}g/m{\ell}$with GSH 1mM and Cobrotoxin $0.5{\mu}g/m{\ell}$ with GSH 5mM 9. Compared with $0.1{\mu}g/m{\ell}$ of cobrotoxin, SNP induced P50 expressions in astrocyte were increased significantly by Cobrotoxin $0.5{\mu}g/m{\ell}$ with DTT 1mM, Cobrotoxin $0.5{\mu}g/m{\ell}$ with DTT 5mM Cobrotoxin $0.5{\mu}g/m{\ell}$ with GSH 1mM and Cobrotoxin $0.5{\mu}g/m{\ell}$ with GSH 5mM. 10. The uptake of the labeled cobrotoxin into the cells was shown under a confocal laser scanning microscope. cobrotoxin was uptaken into the membrane and nucleus of astrocytes. Conclusions : In summary, the present results demonstrate that cobrotoxin directly binds to sulfhydryl group of p50 and IKKS resulting In the reduction of translocation of p50 and IkB release, thereby inhibits activation of NF-${\kappa}B$, and suggest that pico to nanomolar range of cobrotoxin could inhibit the expression of genes in the NF-${\kappa}B$ signal pathway.