• Journal of Korean Neurosurgical Society
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• 제29권6호
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• pp.725-730
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• 2000

Purpose : The subcellular localization of E1B-19k has been known cytosol or nuclear membrane by immunohistochemical staining and could dimerize with Bax to regulate cell death also known by the in-vitro immunoprecipitation. We planed to confirm this dimerization of E1B-19k with Bax in vivo in Cos-7 cells by using green fluorescent protein. Material and Method : We cloned E1B-19k and Bax into C3-EGFP. C3-EGFP-E1B-19k, C3-EGFP-Bax, and C3-EGFP-E1B-19k and pcDNA3-Bax were transfected into Cos-7 cells. We explored location of E1B-19k and Bax, and confirmed its dimerization with Bax in transfected living healthy Cos-7 cells by following green fluorescent protein of E1B-19k on the confocal microscope. Results : E1B-19k was located diffusely in cytoplasm and in nucleus but not in mitochondria. It prevented cell death from the apoptosis by staurosporine but its location was not changed. GFP-E1B-19k is not changed its intracellular location with Bax even with staurosporine. Conclusion : These results support that E1B-19k does not localize in mitochondria nor dimerize with Bax even with staurosporine. We could anticipate E1B-19k prevent cell death via the other dimerizing partner or pathways.

• 생명과학회지
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• 제20권4호
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• pp.572-577
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• 2010

본 연구에서 60 주령된 쥐를 대상으로 8 주간 트레드밀 운동강도와 근섬유 유형에 따른 apoptosis 관련 인자를 연구하였다. 결론적으로 노후된 쥐의 apoptosis 관련 인자는 운동강도와 근육유형에 따라 차이가 났다. EDL이 Soleus보다 apoptosis인자의 변화가 운동강도에 의해 더 민감하게 나타났다. 또한 casepase-3의 발현은 고강도 운동에서 나타났으나 DNA 절편화는 나타나지 않았기에 실질적인 apoptosis는 발생되지 않았다. 이는 노후된 개체의 운동 수행방법으로 고강도 운동은 적합하지 않으며, 고강도 운동상태를 지속할 경우 근섬유의 apoptosis를 유도할 가능성이 있다는 것을 시사한다. 그러므로 추후 실험에서는 운동시간과 빈도에 의한 apoptosis의 time-course impact 관찰과 외재적 경로(caspase independent apoptosis)에서의 apoptosis 관련 단백질 변화에 대한 연구를 통하여 노화에 의한 근손실의 원인 규명이 요구된다.

• 국제물리치료학회지
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• 제6권2호
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• pp.846-851
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• 2015

This neurological damage accelerates the infection reaction of cells and apoptosis at the time of reperfusion after ischemia occurs. BCL-2/BCL-2 allogeneic begeminum has a function of suppressing the apoptosis of cells, and thus it is inferred that the susceptibility of cells to apoptosis is determined by the amount of allogeneic begeminum present which is determined based on the amount of BAX. Ischemia was induced in SD mice by occluding the common carotid artery for 5 minutes, after which blood was re-perfused. NEES was applied to acupuncture points, at 12, 24, and 48 hours post-ischemia on the joksamri, Hapgok. Protein expression was investigated through BAX antibody immuno-reactive cells in the cerebral nerve cells and Western blotting. The results were as follows: In the present study as well, as a result of observation of the change in the number of the BAX reaction cells after the inducement of GI, there was the aspect of most of the BAX reaction cells being observed in the corpus striatum area of the GI group 24 hours after the inducement of ischemia. This revealed the same results as those of previous studies in which the change in the number of BAX reaction cells occurred in all areas while ischemia was in progress. The change in the expression of BAX protein after 24 hours showed that there was a very significant reduction in the NEES group compared to the GI group (p<.01). As a result, a greatest amount of change in the number of BAX immunoreactive cells related to apoptosis 24 hours after ischemia appeared in the NEES group. This study that ischemia increases the expression of BAX that induces apoptosis. Thus, it is determined that ischemia is the main cause of the apoptosis of neurons, and this study reveals that low frequency needle electrode electrical stimulation has the effect of blocking the apoptosis of neurons by reducing protein related to the apoptosis of cells that has increased after ischemia has occurred.

• 한국동물번식학회:학술대회논문집
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• 한국동물번식학회 2001년도 춘계학술발표대회
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• pp.16-16
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• 2001

Apoptosis로 일컬어지는 예정된 세포사멸(programmed cell death)은 개별 세포의 입장에서는 곧바로 사멸을 의미하지만, 정상적인 고등 생물의 입장에서는 개체의 발생과 분화하는데 프로그램된 과정이다. 자발적 세포사멸은 다른 조직에 비해 생식 조직인 난소나 정소에서 복잡한 apoptosis 기작들을 가지리라 사료된다. 본 연구는 Bcl-2 family중 apoptotic protein인 Bax에 대해 suppression하는 유전자를 yeast system을 활용하여 돼지 정소와 난소로부터 각각 cDNA library를 구축한 후 탐색하였다. 탐색에 활용된 cDNA library는 돼지의 정소와 난소로부터 mRNA를 분리하여 yeast vector인 pAD-GAL4-2.1에 구축하였고, 마우스 bax 유전자는 gal 1 promoter의 조절 하에 glucose 배지에서는 유도되지 않고, galactose 배지에서만 선택적으로 Bax를 발현할 수 있는 효모 vector(pL19-bax)를 구축하였다. Bax에 의한 apoptosis suppressor를 탐색하기 위해 우선 효모 W303에 pL19-bax를 transform하여 glucose 배지에서 Bax의 발현을 억제하였다. pL19-bax를 가진 효모에 정소와 난소로부터 구축된 cDNA library를 transform 시키고, transform된 효모는 각각 Bax에 의한 toxicity를 저해하는 유전자를 찾기 위해 스크린되었다. 이러한 방법으로 정소 cDNA library 탐색에서는 5 $\times$ $10^{6}$ transformant중 39개, 난소cDNA library 탐색에서는 2 $\times$ $10^{6}$ transformant중 26개의 콜로니가 생존하였다. 이들 콜로니로부터 유전자를 분리하여 분석해 본 결과 여러 그룹으로 분류할 수 있었다. 각 그룹의 관련 유전자는 protein synthesis/degradation 12종, oxidation/reductation 5종, detoxin/ cell cycle promoter 3종, signal transduction/growth factor 5종, 그리고 알려지지 않은 유전자 9종이었다. 그 중, bax-toxicity inhibition에 강력한 survival phenotype을 가지는 유전자(pSEDL)를 동정하였다. 이것은 T3-4-1 콜로니로부터 분리하였는데 140개 아미노산으로 이루어진 인간 SEDL(GenBank, XM_013096) 유전자와 매우 유사한 homology를 가지며, bax와 관련된 기능은 밝혀져 있지 않다. 이외에도 분리된 유전자에는 NADH, thioreduction, 그리고 cytochrome oxidase와 같은 positive 유전자 군이 크로닝되어, Bax를 이용한 효모에서 apoptosis suppressor에 관련된 유전자를 손쉽게 스크린하는 것이 가능하고, 분리된 유전자의 기능을 예측할 수 있어 지금까지 보고된 유전자 크로닝법 보다는 강력한 수단으로 활용될 수 있다는 사실을 시사하였다. 그러나, ORF에 관계없이 Bax 발현에 저항하는 유전자군이 선발된다든지 하는 문제점은 금후 검토가 필요하리라 사료된다.

• 한국식품영양과학회지
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• 제35권6호
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• pp.671-676
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• 2006

본 연구는 생강의 대표적인 비휘발성 매운맛 성분인 [6]-gingerol이 인체 유방암세포 MDA-MB-231에서 세포사멸에 미치는 영향을 살펴보았으며 그 결과는 다음과 같다. 세포사멸을 억제하는 단백질인 Bcl-2의 발현은 [6]-gingerol의 농도가 증가할수록 발현이 감소되었으며, mRNA 수준에서도 같은 양상을 보였다. 세포사멸을 유도하는 Bax의 단백질 발현은 [6]-gingerol의 농도가 증가되어도 유의적인 차이는 나타나지 않았으며 mRNA 수준에도 별 영향을 미치지 않았다. 그러나 세포사멸의 지표로 사용되는 Bcl-2/Bax의 비율은 [6]-gingerol의 농도가 증가할수록 감소를 보였다. 그리고 [6]-gingerol의 농도가 증가할수록 caspase-3의 활성이 증가하였다. 이상의 결과들로 볼 때, 인체 유방암 세포인 MDA-MB-231에서 [6]-gingerol은 암세포의 증식을 억제하고, 세포사멸을 유도하는 효과가 있는 것으로 사료된다.

• 한국식품영양과학회지
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• 제37권9호
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• pp.1114-1119
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• 2008

ECCG는 녹차 카테킨의 주요 성분으로 항산화작용으로 인한 항암작용이 보고되고 있다. 본 연구는 EGCG가 전이성이 강한 인체 유방암 세포인 MDA-MB-231의 세포사멸에도 영향을 주는지를 알아보고자 하였다. 인체 유방암 세포 배양액에 EGCG를 0, 5, 10, $20\;{\mu}M$로 첨가시켜, 세포사멸과 관련된 단백질들의 단백질과 mRNA 발현, caspase-3 활성을 관찰하였다. EGCG 첨가 농도가 $5\;{\mu}M$ 이상부터 세포사멸을 억제하는 단백질인 bcl-2의 단백질과 mRNA 발현이 감소하였으며, 세포사멸을 유도하는 단백질인 bax의 단백질과 mRNA 발현은 유의적으로 증가하여 결과적으로 EGCG 첨가에 따라 bcl-2/bax의 비율이 유의적으로 감소하였다. 또한 세포사멸의 마지막 단계인 caspase-3의 활성은 EGCG 농도가 증가할수록 유의적으로 증가하였다. 본 연구 결과를 종합해 보면 전이성이 강한 인체 유방암 세포 MDA-MB-231에서 EGCG는 암세포에서 bcl-2의 발현은 억제시키고 bax의 발현은 증가시키며, caspase-3의 활성을 증가시켜 세포사멸을 유도하는 것으로 확인하였다.

• BMB Reports
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• 제36권3호
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• pp.269-274
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• 2003

In the fight against cancer, novel chemotherapeutic agents are constantly being sought to complement existing drugs. Various studies have presented evidence that the apoptosis that is induced by these anticancer agents is implicated in tumor regression, and Bcl-2 family genes play a part in apoptosis following treatment with various stimuli. Here, we present data that a styrylpyrone derivative (SPD) that is extracted from the plant Goniothalamus sp. showed cytotoxic effects on the human breast cancer cell line MCF-7. SPD significantly increased apoptosis in MCF-7 cells, as visualized by phase contrast microscopy and evaluated by the Tdt-mediated dUTP nick end-labeling assay and nuclear morphology. Western blotting and immunostaining revealed up-regulation of the proapoptotic Bax protein expression. SPD, however, did not affect the expression of the anti-apoptotic protein, Bcl-2. These results, therefore, suggest SPD as a potent cytotoxic agent on MCF-7 cells by inducing apoptosis through the modulation of Bax levels.

• BMB Reports
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• 제34권5호
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• pp.391-394
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• 2001

The balance between life and death of a cell regulates essential developmental processes in multicellular organisms. Apoptotic cell death is a complex, stepwise program involving multiple protein components that trigger and execute the demise of the cell. Of the many triggers of apoptosis, most are not well understood, but some key components have been identified, such as those of the Bcl-2 family, which function as anti-apoptotic or proapoptotic factors. Bax, a pro-apoptotic member of this family, has been shown to serve as a component of many apoptotic triggering cascades and its mechanism of action is the focus of intense study. Herein we discuss current, differing ideas on the function of Bax and its structure, and suggest novel mechanisms for how this death protein targets mitochondria, triggering apoptosis.

• Asian Pacific Journal of Cancer Prevention
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• 제15권17호
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• pp.7065-7068
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• 2014

To investigate the anti-proliferative mechanism of mangiferin in a human nasopharyngeal carcinoma cell line, CNE2 cells were incubated with different concentrations of mangiferin (12.5, 25, 50, 100, 150 and $200{\mu}M$) or with PBS as a control for 72 hours. Analyses were made of the cell cycle and apoptosis with measurement of mRNA and protein levels of two apoptosis-related genes, Bcl-2 and Bax. Flow cytometry assays showed mangiferin could inhibit CNE2 cell proliferation via G2/M arrest and induction of early apoptosis. Real time PCR and Western blotting showed the mRNA and protein level of Bcl-2 to be down-regulated, while those of Bax were upregulated, when CNE2 cells were treated with mangiferin. This investigation indicated anti-proliferation effects of mangiferin through induction of cell apoptosis regulated by Bcl-2 and Bax expression.

• Toxicological Research
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• 제18권2호
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• pp.183-190
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• 2002

The mechanism by which catechin-mediated cytotoxicity against tumor cells remains to be elusive. To elucidate the mechanical mights of anti-tumor effects, (-)epigallocatechin-gallate (EGCG) of catechin was applied to human prostate cancer DU 145 cells. Cell viability was measured by crystal violet staining. Cell lysates were wed to measure the catalytic activity of caspases by using fluorogenic peptide: Ac-DEVD-AMC for caspase-3 protease, Z-IETD-AFC for caspase-8 protease, Ac-LEHD-AFC for caspase-9 protease as substrates. The equal amounts of protein from cell lysate was separated on SDS-PAGE and analyzed by western blotting with anti-Fas antibody, anti-FasL antibody, anti-BCL2 antibody and anti-Bax antibody. (-)EGCG induced the death of DUl45 cells, which was revealed as apoptosis shown by DNA fragmentation. (-)EGCG induced the activation of caspase family cysteine proteases including caspase-3, -8 and -9 proteases in DU145 cells. Also, (-)EGCG increased the expression of Fas and Fas ligand (FasL) protein in DU145 colls. The expression level of BCL2 was decreased in (-)EGCG treated DU145 cells, whereas Bax protein was increased in a time-dependent manner. We suggest that (-)EGCG-induced apoptosis of DU145 cells is mediated by signaling pathway involving caspase family cysteine protease, mitochondrial BCL2-family protein and Fas/FasL.