• Asian Pacific Journal of Cancer Prevention
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• 제15권12호
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• pp.5043-5047
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• 2014

Nowadays increasing effectiveness in cancer therapy and investigation of formation of new strategies that enhance antiproliferative activity against target organs has become a subject of interest. Although the molecular mechanisms of apoptosis can not be fully explained, it is known that cell suicide program existing in their memory genetically is activated by pathophysiological conditions and events such as oxidative stress. Low pressure (hypobaric) conditions that create hypoxia promote apoptosis by inhibiting cell cycling. In this study, determination of the effects of fractional hypobaric applications at different times on HeLa cells at cellular and molecular levels were targeted. Experiments were carried out under hypobaric conditions (35.2 kPa) in a specially designed hypobaric cabin including 2% $O_2$ and 98% N. Application of fractional hypobaric conditions was repeated two times for 3 hours with an interval of 24 hours. At the end of the implementation period cells were allowed to incubate for 24 hours for activation of repair mechanisms. Cell kinetic parameters such as growth rate (MTT) and apoptotic index were used in determination of the effect of hypobaric conditions on HeLa cells. Also in our study expression levels of the Bcl-2 gene family that have regulatory roles in apoptosis were determined by the RT-PCR technique to evaluate molecular mechanisms. The results showed that antiproliferative effect of hypobaric conditions on HeLa cells started three hours from the time of application and increased depending on the period of exposure. While there was a significant decrease in growth rate values, there was a significant increase in apoptotic index values (p<0.01). Also molecular studies showed that hypobaric conditions caused a significant increase in expression level of proapoptotic gene Bax and significant decrease in antiapoptotic Bfl-1. Consequently fractional application of hypobaric conditions on HeLa cell cultures increased both antiproliferative and apoptotic effects and these effects were triggered by the Bax gene.

• 대한한방내과학회지
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• 제27권2호
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• pp.429-443
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• 2006

Objectives : The Purpose of this study was to identify anti-tumor effects of Curcuma longa L. on some kinds of cancer cells through molecular biologic methods. Materials & Methods : We used 4 kinds of cancer cell lines such as glioma cells(A172), cervical cancer cells(HeLa), Prostate cancer cells(PC3), lung cancer cells(A549). We injected the boiled extract of Curcuma longa L. $5{\mu}g,\;10{\mu}g$ to culture media(ml) for 24 hours. We measured the cytotoxic effect on 4 kinds of cancer cells through trypan blue exclusion test and the suppressive effect on viability of 4 kinds of cancer cells via MTT assay. We measured the change of mitochondria membrane potential via flow cytometry. The quantitative RT-PCR was used to examine the effect on the revelation of Bcl-2 and Bax which genes are related to apoptosis. We examined the effect on the revelation of Bcl-2 protein and Bax protein by western blot analysis. Results: 1. Extract of Curcuma longa L. showed significant cytotoxic effect on A172, HeLa, PC3 compared to the control group with density dependent manner. 2. Extract of Curcuma longs L. showed significant suppressive effect on viability of A172, HeLa, PC3 compared to the control group with density dependent manner. 3. Curcuma longs L. induced apoptosis by decreasing the membrane potential of mitochondria in A172, HeLa, PC3. 4. In the test about the revelation of genes related to apoptosis, the revelation of Bcl-2 decreased and the revelation of Bax increased in A172. HeLa, PC3 treated with Curcuma longa L. with density dependent manner. 5. In the test about the revelation of protein related to apoptosis, the protein levels of Bcl-2 decreased and the protein levels of Bax increased in A172, HeLa, PC3 treated with Curcuma longa L. Conclusions: This experiment shewed that Curcuma longs L. has anti-tumor effect on glioma, cervical, Prostate cancer cells except on lung cancer. We hope that anti-tumor effects of Curcuma longa L. will be more Practically identified.

• 생명과학회지
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• 제27권2호
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• pp.164-171
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• 2017

본 연구에서는 인간 자궁경부암세포주에서 S-allylcysteine (SAC)이 세포자멸경로에 중요한 역할을 담당하는 initiator caspase의 하나인 caspase-9와 effector caspase에 속하는 caspase-3 및 caspase-7 그리고 DNA 복구에 관여하는 poly ADP-ribose polymerase (PARP)의 발현조절에 미치는 영향과, SAC에 의한 이러한 세포자멸 및 DNA 복구 관련 단백질의 발현변화가 세포증식억제를 통한 기능적 작용을 유발하는지를 조사하였다. 단백질 발현분석 결과, 특히 50 mM의 SAC로 48시간 동안 처리하였을 경우, procaspase-3, -7, -9 및 PARP의 발현은 각각 94%, 38%, 95% 및 64% 감소되었으며, 이와 반대로 caspase-3, -7, -9 및 cleaved-PARP의 발현은 현저히 증가되었다. 또한 cell proliferation assay 결과, 20 mM 이상의 SAC 처리는 6, 12, 24 및 48시간에서 농도 및 시간 의존적인 세포증식 억제효과를 나타내었다. 이러한 결과는 SAC 처리가 자궁경부암세포의 증식을 억제하며, 이에 대한 가능한 분자적 작용기전들 중의 하나로 세포자멸과정 중 initiator caspase의 하나인 caspase-9의 활성을 유도하고 이에 따른 effector caspase인 caspase-3과 caspase-7의 활성을 촉진시킬 뿐만 아니라 DNA 복구에 관여하는 PARP의 불활성화를 초래함으로써 세포자멸 유도에 관여하는 것으로 사료된다.

• 한국미생물·생명공학회지
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• 제31권4호
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• pp.362-367
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• 2003

운지버섯을 희석 감귤농축액과 일반 합성배지에서 각각 배양한 배양물을 열탕추출하고 여과하여 얻어진 버섯 배양추출물과 버섯을 접종하지 않은 희석 감귤농축액 자체에 대한 항산화 활성, 아질산 제거능, 그리고 항암활성을 조사 비교하였다. DPPH 전자공여를 이용한 항산화 활성에서 희석 감귤농축액에서 얻어진 배양추출물(Extract-I)은 89%, 일반 합성배지에서 얻어진 배양추출물(Extract-II)은 66%, 접종하지 않은 희석 감귤농축액 자체(Extract-III)는 22%의 항산화 활성을 나타내었다. 아질산 제거능은 배양추출물 모두 산성 조건일수록 증가하였으며, Extrac-I은 67%, Extract-II는 54%, 그리고 Extract-III는 34%의 아질산 제거능을 보였다. HeLa, PC-3, HepG2 및 A-549 세포에 대한 항암활성을 조사한 결과, Exract-I는 각각의 암세포들에 대해서 순차적으로 75%, 82%, 55%, 그리고 82%의 높은 생육 저해 활성을 보였다. 반면에 Extract-II는 HeLa cell에 대해서만 66%의 생육저해를 보였고, Exoact-III는 모든 암세포의 생육을 저해하지 않는 것으로 조사되었다. 따라서 운지버섯을 희석 감귤농축액에 배양함으로서 일반 합성배지에서 배양하거나 감귤농축액 자체 보다 항산화 활성과 항암활성을 현저히 증가시킬 수 있었다.

• Radiation Oncology Journal
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• 제7권2호
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• pp.149-156
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• 1989

HeLa 세포의 spheroid를 배 양시켜 cis-platinum과 carboplatin으로 처리한 후 그 반응을 세포의 생존분획으로 분석하였다. 체외실험 model인 spheroid를 사용하여 platinum유사체의 약효와 방사선 감수성을 평가하고 약제에 대한 단층 세포와 spheroid의 감수성 차이를 세포-생존곡선에서 규명하기 위하여 본 실험을 시행하였다. Cis-platinum 농도-곡선에서 spheroid의 $Cq=3.4{\mu}M$이고 $Co=1.2{\mu}M$이었다. 이 것은 단층세포에 비하여 Co는 큰 변화가 없으나 Cq가 증가되어 cis-platinum이 저산소층 세포보다 활동적으로 분화하는 표면세포에 주로 작용하였으며, 반대로 carboplatin의 효과는 spheroid에 대한 $(Co=15.0{\mu}M)$ 감수성이 단층세포$(Co=32.5{\mu}M)$에 비하여 크게 증가되어, spheroid의 심층 세포에 주로 작용하였다. 방사선과 carboplation의 병용효과를 세포 생존분획이 0.01 수준에서 isobologram으로 분석한 결과 상호작용으로 supra-additive 효과를 나타내었다. 따라서, carboplatin은 cis-platinum에 비하여 신장과 위장에 대한 독성작용이 적고, 방사선과 병용함으로써 향후 더욱 효과적 인 종양 치료에 중요한 역할을 할 것으로 기대한다.

• Radiation Oncology Journal
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• 제16권2호
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• pp.99-106
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• 1998

목적 : 임상에서 발생빈도가 높은 위선암, 폐선암, 망막세포종, 자궁경부 편평상피암의 4가지 인 체암세포주(MKN-45, PC-14, Y-79, HeL)를 이용하여 방사선조사후 세포생존분획 및 세포생존곡선의 모양을 결정하는 지표를 구하고 방사선조사후 손상 회복정도를 측정하여 이들 여러 지표간의 상관관계 여부를 구명하기 위하여 본실험을 시행하였다. 대상 및 방법 :각 세포주의 방사선감수성을 보기 위하여 1, 2, 3, 5, 7 및 10Gy의 방사선을 1 회 조사하였고 방사선조사후 준치사손상 회복정도를 보기 위하여 5Gy씩 2회의 방사선조사를 0, 1, 2, 3, 4, 6 및 24시간 간격으로 시행하였다. 세포의 생존분획은 $Sperman-K\"{a}rbor$ 방법으로 세포집락형성능력을 측정하여 산출하였으며 생존곡선의 수학적 분석은 linear-quadratic(LQ), multitarget-single hit(MS) 모형과 mean inactivation $dose(\v{D})$를 이용하였다. 결과 : 방사선조사후의 세포생존 실험결과 2Gy에서의 세포생존분획(SF2)은 0.174에서 0.85까지 다양하게 나타났으며 Y-79는 유의하게 낮은 SF2를, PC-14는 높은 SF2를 나타내었다(p<0.05, t-test). LQ model로 분석한 방사선 세포생존곡선의 분석결과 Y-79, MKN-45, HeLa, PC-14에서 ${\alpha}$가 각각 0.603, 0.355, 0.275, 0.102이었고 ${\beta}$는 각각 0.005, 0.016, 0.025, 0.027이었다. MS model로 분석한 결과는 Y-79, MKN-45, HeLa, PC-14에서 Do가 각각 1.59, 1.84, 1.88, 2.52였고 n은 0.97, 1.46, 1.52, 1.69를 보였다. 한편 Gauss-Laguerre방법으로 계산한 $\v{D}$는 Y-79, MKN-45, HeLa, PC-14에서 각각 1.62, 2.37, 2.61, 3.95였다. SF2가 감소함에 따라 ${\alpha}$값은 증가하였고 Do, $\v{D}$값은 감소하였으며 이들간의 Pearson 상관계수는 각각 0.953, 0.993, 0.999였다. (p<0.05). 분할조사에 의한 준치사손상 회복정도는 약 4시간 내외에 포화상태에 도달하였으며 포화상태의 recovery ratio(RR)는 2에서 3.79 사이였다. RR은 방사선감수성의 지표인 SF2, ${\alpha}$, ${\beta}$, Do, $\v{D}$과 통계학적으로 유의한 상관관계를 나타내지 않았다. 결론 : 본 연구의 결과 네가지 인체상피암세포주의 내재적 발사선감수성은 서로 상이하였으며 Y-79가 가장 민감하였고, MKN-45와 HeLa는 각각 중등도의 방사선감수성을 나타냈으며 PC-14는 방사선감수성이 가장 낮았다. 이와같은 감수성의 차이는 SF2, ${\alpha}$, Do와 $\v{D}$의 차이로 나타났으며 띠들간에는 밀접한 상관관계를 나타내었다. 방사선에 의한 준치사손상 회복력은 MKN-45와 HeLa에서 높게 나타났고 회복력과 방사선감수성과는 무관하였다. 각 암세포주에 따르는 이와같은 지표들은 향후 방사선치료 효과를 높이기 위한 방사선생물학 실험의 기초 자료로서 이용되어 질 수 있을 것이다.

• Asian Pacific Journal of Cancer Prevention
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• 제15권9호
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• pp.4101-4107
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• 2014

Protein phosphatase 1 (PP1) is a major serine/threonine phosphatase that controls gene expression and cell cycle progression. The active mutant IPP5 ($8-60hIPP5^m$), the latest member of the inhibitory molecules for PP1, has been shown to inhibit the growth of human cervix carcinoma cells (HeLa). In order to elucidate the underlying mechanisms, the present study assessed overexpression of $8-60hIPP5^m$ in HeLa cells. Flow cytometric and biochemical analyses showed that overexpression of $8-60hIPP5^m$ induced G2/M-phase arrest, which was accompanied by the upregulation of cyclin B1 and phosphorylation of G2/M-phase proteins ATM, p53, $p21^{cip1/waf1}$ and Cdc2, suggesting that $8-60hIPP5^m$ induces G2/M arrest through activation of the ATM/p53/$p21^{cip1/waf1}$/Cdc2/cyclin B1 pathways. We further showed that overexpression of $8-60hIPP5^m$ led to delayed nuclear translocation of cyclin B1. $8-60hIPP5^m$ also could translocate to the nucleus in G2/M phase and interact with $pp1{\alpha}$ and Cdc2 as demonstrated by co-precipitation assay. Taken together, our data demonstrate a novel role for $8-60hIPP5^m$ in regulation of cell cycle in HeLa cells, possibly contributing to the development of new therapeutic strategies for cervix carcinoma.

• Asian Pacific Journal of Cancer Prevention
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• 제15권11호
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• pp.4475-4482
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• 2014

Background: Cisplatin, a DNA damaging agent, induces apoptosis through increasing DNA fragmentation. However, identification of intrinsic resistance molecules against Cisplatin is vital to estimate the success of therapy. Bag-1 (Bcl-2-associated anthanogene) is one anti-apoptotic protein involved in drug resistance impacting on therapeutic efficiency. Elevated levels of this protein are related with increase cell proliferation rates, motility and also cancer development. For this reason, we aimed to understand the role of Bag-1 expression in Cisplatin-induced apoptosis in HeLa cervix cancer cells. Cisplatin decreased cell viability in time- and dose-dependent manner in wt and Bag-1L+HeLa cells. Although, $10{\mu}M$ Cisplatin treatment induced cell death within 24h by activating caspases in wt cells, Bag-1L stable transfection protected cells against Cisplatin treatment. To assess the potential protective role of Bag-1, we first checked the expression profile of interacting anti-apoptotic partners of Bag-1. We found that forced Bag-1L expression prevented Cisplatin-induced apoptosis through acting on Mcl-1 expression, which was reduced after Cisplatin treatment in wt HeLa cells. This mechanism was also supported by the regulation of heat shock protein (Hsp) family members, Hsp90 and Hsp40, which were involved in the regulation Bag-1 interactome including several anti-apoptotic Bcl-2 family members and c-Raf.

• BMB Reports
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• 제36권5호
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• pp.468-474
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• 2003

The molecular mechanism of hypoxia-induced apoptosis has not been clearly elucidated. In this study, we investigated the involvement of extracellular signal-regulated protein kinase (ERK 1/2) in hypoxia-induced apoptosis using cobalt chloride in HeLa human cervical cancer cells. The cobalt chloride was used for the induction of hypoxia, and its $IC_{50}$ was $471.4\;{\mu}M$. We demonstrated the DNA fragmentation after incubation with concentrations more than $50\;{\mu}M$ cobalt chloride for 24 h, and also evidenced the morphological changes of the cells undergoing apoptosis with electron microscopy. Next, we examined the signaling pathway of cobalt chloride-induced apoptosis in HeLa cells. ERK1/2 activation occurred 6 and 9 h after treatment with $600\;{\mu}M$ cobalt chloride. Meanwhile, the pretreatment of the MEK 1 inhibitor (PD98059) completely blocked the cobalt chloride-induced ERK 1/2 activation. At the same time, the activated ERK 1/2 translocated into the nucleus and phosphorylated its transcriptional factor, c-Jun. In addition, the pretreatment of PD98059 inhibited the cobalt chloride-induced DNA fragmentation and apoptotic cell death. These results suggest that cobalt chloride is able to induce apoptotic activity in HeLa cells, and its apoptotic mechanism may be associated with signal transduction via ERK 1/2.

• BMB Reports
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• 제41권3호
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• pp.230-235
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• 2008

LRP15 is a novel gene cloned from lymphocytic cells, and its function is still unknown. Bioinformatic data showed that LRP15 might be regulated by DNA methylation and had an important role in DNA repair. In this study, we investigate whether the expression of LRP15 is regulated by DNA methylation, and whether overexpression of LRP15 increases efficiency of DNA repair of UV-induced DNA damage in HeLa cells. The results showed (1) the promoter of LRP15 was hypermethylated in HeLa cells, resulting a silence of its expression. Gene expression was restored by a demethylating agent, 5-aza-2'-deoxycytidine, but not by a histone deacetylase inhibitor, trichostatin A; (2) overexpression of LRP15 inhibited HeLa cell proliferation, and the numbers of cells in the G2/M phase of the cell cycle in cells transfected with LRP15 increased about 10% compared with controls; (3) cyclin B1 level was much lower in cells overexpressing LRP15 than in control cells; and (4) after exposure to UV radiation, the LRP15-positive cells showed shorter comet tails compared with the LRP15-negative cells. From these results we conclude that the expression of LRP15 is controlled by methylation in its promoter in HeLa cells, and LRP15 confers resistance to UV damage and accelerates the DNA repair rate.