• Asian Pacific Journal of Cancer Prevention
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• 제14권12호
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• pp.7339-7344
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• 2013

To analyze the effects of a new unknown peptide DEF on the growth of tumor cells, a fused polypeptide TAT-DV1-DEF was designed and synthesized. The lung adenocarcinoma cell line GLC-82 treated with TAT-DV1-DEF was analyzed with a cell counting kit 8, and the location of polypeptides in cells was observed under laser confocal microscopy. The efficiency of polypeptide transfection and changes in nuclear morphology were analyzed by flow cytometry and fluorescence microscopy, respectively. Finally, the mechanism of tumor cell growth inhibition was evaluated by Western blotting. We found that TAT-DV1-DEF could significantly inhibit the growth of the lung adenocarcinoma cell line GLC-82, but not the normal human embryonic kidney cell line HEK-293. Polypeptides were found to be mostly localized in the cytoplasm and some mitochondria. The efficiency of polypeptide transfection in the two cell types was approximately 99%. Apoptotic nuclei were observed under fluorescence microscopy upon treatment with polypeptides and DAPI staining. Western blot analyses indicated that the polypeptide inhibition of tumor cell growth was apoptosis dependent. In the present study, we demonstrated that fused polypeptides could induce apoptosis of the lung adenocarcinoma cell line GLC-82, indicating that the new unknown peptide DEF has antitumor effects.

• Journal of Microbiology and Biotechnology
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• 제12권6호
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• pp.929-935
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• 2002

The 5-stage biological nutrient removal (BNR) process with step feed system showed a very stable organic carbon and nutrient removal efficiency ($87\%\;COD\,;79\%\;nitrogen,\;and\;87\%$ phosphorus) for an operation period of 2 years. In each stage at the pilot plant, microbial communities, which are important in removing nitrogen and phosphorus, were investigated using fluorescence in-situ hybridization (FISH) and 165 rDNA characterization. All tanks of 5-stage sludge had a similar composition of bacterial communities. The totat cell numbers of each reactor were found to be around $2.36-2.83{\times}10^9$ cells/ml. About $56.5-62.0\%$ of total 4,6-diamidino-2-phenylindol (DAPI) cells were hybridized to the bacterial-specific probe EUB388. Members of ${\beta}$-proteobacteria were the most abundant proteobacterial group, accounting for up to $20.6-26.7\%$. The high G+C Gram-positive bacterial group and Cytophaga-Flexibacter cluster counts were also found to be relatively high. The beta subclass proteobacteria did not accumulate a large amount of polyphosphate. The proportion of phosphorus-accumulating organisms (PAOs) in the total population of the sludge was almost $50\%$ in anoxic-1 tank. The high G+C Gram-positive bacteria and Cytophaga-Flexibacter cluster indicate a key role of denitrifying phosphorus-accumulating organisms (dPAOs). Both groups might be correlated with some other subclass of proteobacteria for enhancing nitrogen and phosphorus removal in this process.

• 대한약침학회지
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• 제12권3호
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• pp.49-59
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• 2009

Background : Anticancer effects of herbal medicine have been reported in various types of cancer, but the systematic approaches to explain molecular mechanism(s) are not established yet. Objective : The purpose of this study is to investigate the apoptotic cell death by Egg White combined Chalcanthite in NCI-H460 human lung cancer cells. Methods : Inhibitory effects were estimated by the MTT-assay. Cancer cells were stained with DAPI and showed condensed and fragmented nuclei. The expression of cleaved caspase-3, bcl-2, and bax was detected by western blotting. To establish a basis of understanding for anti-cancer mechanism, whole proteins have been obtained from NCI-H460 harvested at 24 hrs after the treatment of Egg White combined Chalcanthite, protein expression has been profiled by 2DE-based proteomic approach. Results : NCI-H460 human lung cancer cells were treated by three samples of IS3, IS4 and IS5. IS4 inhibited most effectively the growth of NCI-H460 human lung cancer cells. The expression of cleaved caspase-3 increased in IS4 in a concentration-dependent manner. Various changes of the protein expression have been monitored, and most frequent dysregulation was found in Vimentin, Lamin-A/C. Conclusion : Egg White combined-Chacanthite inhibited the growth of NCI-H460 human lung cancer cells by inducing the apoptotic cell death via caspase-3 activation. Based upon the present findings, the further study will focus on monitoring various cancer survival factors after artificial regulation of the proteins identified, and it would be the basis for the understanding of the Chacabthite anticancer effect(s) at the molecular level.

• 대한한방부인과학회지
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• 제20권2호
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• pp.139-154
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• 2007

Purpose: This study examined the Cell differentiation and the anti-oxidative effect of Dioscoreae Rhizoma on HeLa cells. We are interested in whether Dioscoreae Rhizoma is capable of causing apoptosis processes on HeLa cell, and whether cotreatment of NCS with Dioscoreae Rhizoma reduces cell viability. Methods: We used aqueous extract to treat HeLa cell with different concentrations treated with a water or a MeOH extract of Dioscoreae Rhizoma (0, x10, x20, x40, x80). The MTT (3, (4, 5-dimethyl-thiazol) 2, 5-diphenyl-tetraxolium bromide) reduction assay was employed to quantify the differences in cell activity and viability. Cells were stained with DAPI and visualized by fluorescent Microscope. The caspase-3, Bcl-2, PARP, p53 expression level was monitored using western-blotting techniques. The patterns of the changes in expression were scanned and analyzed. Results: The survival rate of cells treated with Dioscoreae Rhizoma extracts increased by 20% at specific concentration. The other side Dioscoreae Rhizoma extracts induced apoptotic features including chromatin condensation and fragmentation. And Dioscoreae Rhizoma extracts increased the expression of caspase-3, p53 and the cleavage of PARP protein. However, co-treatment with Dioscoreae Rhizoma with NCS attenuated the activations of p53 and PARP protein that were mediated by NCS treatment alone. This is indicated Dioscoreae batatas extracts attenuated cytotoxicity induced by oxidative agents including NCS. Conclusion: Our results suggest Dioscoreae Rhizoma extracts induce cell differentiation or apoptosis connected with concentration. Further elucidation of concentration of Dioscoreae Rhizoma awaits many other biochemical investigative studies.

• 한방안이비인후피부과학회지
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• 제19권2호
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• pp.26-39
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• 2006

Objective : The effect of water extract of Chungjogupae-tang (CJGPT) was investigated on the growth of human lung carcinoma A549 cells. Methods : MTT assay and fluorescent microscope performed to compare and examine the efficacy of CJGPT treatment on the cytostaticity of lung cancer cells in proportion to time and doses, and DAPI staining and Western blot analysis were used to examine their effect on apoptosis. In addition the quantitative RT-PCR was used to examine to lung cancer cells growth and Progtaglandin E2 and Telomerase activity were measured Results : Exposure of A549 cells to CJGPT resulted in the growth inhibition and apoptosis in a dose-dependent manner as measured by MTT assay and fluorescent microscope. The antiuoliferative effect by CJGPT treatment in A549 cells was associated with morphological changes such as membrane shrinking and cell rounding up. CJGPT treatment resulted in an up-regulation of cyclin-dependent kinase inhibitor p21(WAF1/CIPl) in a p53-independent fashion. We found that CJGPT treatment decreased the levels of cyclooxygenase (COX)-2 and inducible nitric oxide synthease (iNOS) expression without significant changes in the expression of COX-1, which was correlated with a decrease in protaglandin E2 (PGE2) synthesis. CJGPT treatment also inhibited the levels of human telomerase reverse transcriptase (hTERT) and telomerase-associated protein (TEP)-1 mRNA expression, however the activity of telomerase was slightly increased by CJGPT treatment. Conclusion : These findings suggested that CJGPT-induced inhibition of human lung carcinoma A549 cell growth was connected with the induction of apoptotic cell death and the results provided important new insights into the possible molecular mechanisms of the anti-cancer activity of CJGPT.

• Journal of Acupuncture Research
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• 제22권6호
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• pp.37-50
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• 2005

Objectives : The purpose of this study was to investigate the anti-caner effect of Bee Venom and Melittin on the prostatic cancer cell(PC-3). The goal of study is to ascertain whether Bee Venom and Melittin inhibits the 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 Bee Venom and Melittin, we performed Fluorescence microscope, MTT assay, Western blotting, Flow cytometry, PAGE electrophoresis and Surface plasmon resonance analysis to identify the cell viability, apoptosis and gene related to apoptosis. Results : 1. Compared with Control cell, the inhibition of cell growth reduced in proportion with the dose of Bee Venom or Melittin($0{\sim}10{\mu}g/ml$) in PC-3. 2. In PC-3, Cell viabilities of Bee Venom or Melittin treatment was decreased significantly. 3. The nucli of Control cells were stained round and homogenous in DAPI staining, but those of PC-3 were stained condense and splitted. 4. In PC-3, apoptosis of Bee Venom or Melittin treatment was increased significantly. 5. Bax, Caspase-3 and P ARP of Bee Venom or Melittin treatment was increased significantly and Bcl-2 of Bee Venom or Melittin treatment was decreased significantly. Caspase-9 of Bee venom treatment was increased significantly. Conclusion : These results indicate that Bee Venom and Melittin inhibits the growth of prostate cancer cells, has anti-cancer effects by inducing apoptosis. We wish that the anti-cancer effects of Bee Venom and Melittin are used to clinical caner treatment.

• Journal of Acupuncture Research
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• 제29권1호
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• pp.37-45
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• 2012

목적 : 최근 NF-${\kappa}B$와 STAT3의 활성억제와 관련된 항암제 연구가 주목받고 있으며, 본 연구는 사독(蛇毒)이 세포자멸사 관련 단백질의 발현 조절을 통하여 세포자멸사를 유도하고, NF-${\kappa}B$와 STAT3의 활성억제를 유도하여 난소암 PA-1 세포의 성장을 억제하는지를 확인하고, 해당 기전을 살펴보고자 하였다. 방법 : 사독을 처리한 후 난소암 PA-1 세포의 세포자멸사의 관찰에는 DAPI, TUNEL staining assay를 시행하였고, 세포자멸사 조절단백질 및 NF-${\kappa}B$, STAT3의 활성 변동 관찰에는 western blot analysis를 시행하였다. 결과 : 1. 사독을 처리한 후 난소암 PA-1 세포에서 세포자멸사가 유도되어 암세포성장이 억제되었다. 2. 사독을 처리한 후 세포자멸사 관련 단백질 중 세포자멸사 촉진 단백질인 cleaved caspase-3, Bax의 발현은 증가되었고, 세포자멸사 억제 단백질인 Bcl-2의 발현은 감소되었다. 3. 사독을 처리한 후 난소암 PA-1 세포의 NF-${\kappa}B$와 STAT3 발현은 감소되었고, 각각의 길항제인 salicylic acid와 stattic 처리 후 NF-${\kappa}B$와 STAT3 발현은 더욱 감소되었다. 결론 : 사독은 난소암 세포의 세포자멸사 유발과, NF-${\kappa}B$와 STAT3의 활성억제를 통해 치료 효율이 높고, 내성이 적은 난소암 치료제의 개발에 도움이 될 것으로 기대된다.

• 한방재활의학과학회지
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• 제24권2호
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• pp.41-50
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• 2014

Objectives The purpose of this study was to evaluate the effects of Clematidis radix extract on $CoCl_2$-induced apoptosis in SH-SY5Y human neuroblastoma cells. Methods In order to investigate the protective effect of Clematidis radix on $CoCl_2$-induced cytotoxicity in neuronal cells, MTT(3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay, DAPI(4,6-diamidino-2-phenylindoleI) staining, TUNEL (terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling) assay, DNA fragmentation assay and western blotting were performed on SH-SY5Y human neuroblastoma cells. Results Cells treated with $CoCl_2$ exhibited several apoptotic features, while cells pre-treated with Clematidis radix prior to $CoCl_2$ exposure showed a decrease in the occurrence of apoptotic features. $CoCl_2$ increased HIF-$1{\alpha}$ expression, in contrast, Clematidis radix treatment decreased $CoCl_2$-induced HIF-$1{\alpha}$ expression. Pre-treatment with the extract of Clematidis radix suppressed Bax, cytochrome c, and caspase-3 expressions, and also increased Bcl-2 expression in SH-SY5Y human neuroblastoma cells. Conclusions These results suggest that Clematidis radix may exert a protective effect on $CoCl_2$-induced apoptosis in SH-SY5Y human neuroblastoma cells.

• Preventive Nutrition and Food Science
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• 제12권2호
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• pp.79-83
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• 2007

The effects of young radish (YR, yeolmu in Korean) on the induction of apoptosis were examined in AGS human gastric adenocarcinoma cells. The young radish kimchi (YRK) were made of YR cultivated in the soil without (Control YR kimchi: C-YRK) and with 1,818 g/m$^{3}$ sulfur (Sulfur YR kimchi: S-YRK), respectively. Methanol extracts from S-YRK exhibited higher inhibitory effect on the growth of AGS human gastric adenocarcinoma cells in a time dependent-manner compared to C-YRK at the same concentration. 4,6-diamidino-2-phenylindole (DAPI) staining showed that S-YRK induced apoptosis accompanied by the increased Bax but decreased Bcl-2 in mRNA expression. Moreover, S-YRK decreased the levels of cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) mRNA expressions. The results suggested that S-YRK cultivated in the presence of sulfur elicited stronger anticancer activity than C-YRK in vitro. Dietary intakes of S-YRK may be beneficial to decrease the risk of cancer.

• International Journal of Oral Biology
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• 제41권4호
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• pp.183-190
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• 2016

Anthricin (Deoxypodophyllotoxin), a naturally occurring flavolignan, has well known anti-cancer properties in several cancer cells, such as prostate cancer, cervical carcinoma and pancreatic cancer. However, the effects of Anthricin are currently unknown in oral cancer. We examined the anticancer effect and mechanism of action of Anthricin in human FaDu hypopharyngeal squamous carcinoma cells. Our data showed that Anthricin inhibits cell viability in a dose- and time-dependent manner ($IC_{50}$ 50 nM) in the MTT assay and Live & Dead assay. In addition, Anthricin treated FaDu cells showed marked apoptosis by DAPI stain and FACS. Furthermore, Anthricin activates anti-apoptotic factors such as caspase-3, -9 and poly (ADP-ribose) polymerase (PARP), suggesting that caspase-mediated pathways are involved in Anthricin- induced apoptosis. Anthricin treatment also leads to accumulation of the pro-apoptotic factor Bax, followed by inhibition of cell growth. Taken together, these results indicate that Anthricn-induced cell death of human FaDu hypopharyngeal squamous carcinoma cells is mediated by mitochondrial-dependent apoptotic pathway. In summary, our findings provide a framework for further exploration on Anthricin as a novel chemotherapeutic drug for human oral cancer.