• Animal cells and systems
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• 제6권2호
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• pp.171-180
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• 2002

Nitric oxide has high affinity for iron, and thus it can cause intracellular iron loss. We tested the idea that intracellular iron can be the primary target of NO toxicity by comparing the signaling mechanisms involved in cell death caused by iron depletion and that caused by NO. Treatment of HL-60 cells with a NO donor, S-nitroso-N-acetyl-DL-penicillamine (SNAP), decreased the intracellular iron level rapidly as that observed with the iron chelator deferoxamine (DFO). Iron chelators such as DFO and mimosine could induce death of human leukemic HL-60 cells by a mechanism requiring activation of p38 kinase, c-Jun N-terminal kinase, caspase-3 and caspase-8. DFO and SNAP also caused release of cytochrome c from mitochondria. Inhibition of p38 kinase by a selective inhibitor, SB203580, abolished the NO and DFO-induced cell death, release of cytochrome c, and activation of caspase-3 and caspase-8, thus indicating that p38 kinase lies upstream in the cell death processes. In a parallel situation, the cells that are sensitive to NO showed similar sensitivity to DFO. Moreover, simultaneous addition of ferric citrate, an iron-containing compound, inhibited the SNAP and DFO-induced activation of caspases and also blocked the NO-mediated cell cycle arrest at $G_1$ phase. Collectively, our data implicate that the NO-induced cell death of tumor cells including HL-60 cells is mediated by depletion of iron and further suggest that activation of p38 kinase lies upstream of cytochrome c release and caspase activation involved in this apoptotic process.

• 생명과학회지
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• 제25권2호
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• pp.249-255
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• 2015

오미자 종자에서 추출된 정유(Schisandrae Semen essential oil, SSeo)의 항암활성 및 작용 기전 해석을 위하여 U937 백혈병 세포를 대상으로 apoptosis 유도 여부를 조사하였다. SSeo 처리에 의한 U937 세포의 증식 억제는 apoptosis 유도와 연관성이 있음을 DAPI 염색을 통한 apoptotic body 출현의 증가, agarose gel 전기영도에 의한 DNA의 단편화 유도 및 flow cytometry 분석에 의한 Sub-G1기 세포 빈도의 증가로 확인하였다. SSeo 처리에 의한 apoptosis 유도에서 IAP family 단백질에 속하는 XIAP, cIAP-1 및 survivin의 발현 감소와 anti-apoptotic Bcl-2 단백질의 발현 저하, DR4 및 DR5의 발현 증가와 연관성이 있었다. SSeo 처리는 또한 Bid truncation, 미토콘드리아 기능 손상, caspases (-3, -8 and -9)의 활성화와 활성형 caspase-3의 기질 단백질인 PARP의 단편화를 동반하였다. 본 연구의 결과는 오미자 정유의 생화학적 항암기전 해석을 이해하고 향후 지속적인 연구를 위한 기초자료로서 활용될 수 있을 것으로 생각된다.

• 한국축산식품학회지
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• 제26권3호
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• pp.402-409
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• 2006

Although microorganisms are, in fact, the most diverse and abundant type of organism on Earth, the ecological functions of microbial populations remains poorly understood. A variety of bacteria including marine Vibrios encounter numerous ecological challenges, such as UV light, predation, competition, and seasonal variations in seawater including pH, salinity, nutrient levels, temperature and so forth. In order to survive and proliferate under variable conditions, they have to develop elaborate means of communication to meet the challenges to which they are exposed. In bacteria, a range of biological functions have recently been found to be regulated by a population density-dependent cell-cell signaling mechanism known as quorum-sensing (QS). In other words, bacterial cells sense population density by monitoring the presence of self-produced extracellular autoinducers (AI). N-acylhomoserine lactone (AHL)-dependent quorum-sensing was first discovered in two luminescent marine bacteria, Vibrio fischeri and Vibrio harveyi. The LuxI/R system of V. fischeriis the paradigm of Gram-negative quorum-sensing systems. At high population density, the accumulated signalstrigger the expression of target genes and thereby initiate a new set of biological activities. Several QS systems have been identified so far. Among them, an AHL-dependent QS system has been found to control biofilm formation in several bacterial species, including Pseudomonas aeruginosa, Aeromonas hydrophila, Burkholderia cepacia, and Serratia liquefaciens. Bacterial biofilm is a structured community of bacterial cells enclosed in a self-produced polymeric matrix that adheres to an inert or living surface. Extracellular signal molecules have been implicated in biofilm formation. Agrobacterium tumefaciens strain NT1(traR, tra::lacZ749) and Chromobacterium violaceum strain CV026 are used as biosensors to detect AHL signals. Quorum sensing in lactic acid bacteria involves peptides that are directly sensed by membrane-located histidine kinases, after which the signal is transmitted to an intracellular regulator. In the nisin autoregulation process in Lactococcus lactis, the NisK protein acts as the sensor for nisin, and NisR protein as the response regulator activatingthe transcription of target genes. For control over growth and survival in bacterial communities, various strategies need to be developed by which receptors of the signal molecules are interfered with or the synthesis and release of the molecules is controlled. However, much is still unknown about the metabolic processes involved in such signal transduction and whether or not various foods and food ingredients may affect communication between spoilage or pathogenic bacteria. In five to ten years, we will be able to discover new signal molecules, some of which may have applications in food preservation to inhibit the growth of pathogens on foods.

• 대한의생명과학회지
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• 제16권2호
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• pp.119-122
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• 2010

Interleukin-1${\beta}$ $(IL-1{\beta})$ is one of the key proinflammatory cytokines and it plays an important role for the antimycobacterial host defense mechanisms. In this study, we examined Mycobacterium tuberculosis (MTB)-stimulated induction of IL-1${\beta}$ and evaluated the associated signal transduction pathways. In PMA-differentiated THP-1 cells, MTB infection increased mRNA expression of IL-$1{\beta}$ in a dose-dependent manner. The expression of IL-1${\beta}$ mRNA began to be induced at 1.5 h after infection, and induced expression of IL-1${\beta}$ was retained for 48 h after MTB infection. The increase in expression of IL-1${\beta}$ caused by MTB was reduced in cells treated with Ro-31-8425 (an inhibitor of PK$C{\alpha}$, ${\beta}I$, ${\beta}II$, ${\gamma}$, ${\varepsilon}$) or PD98059 (an inhibitor of MEK1), meanwhile, pre-treatment with $G\ddot{o}6976$ (an inhibitor of $Ca^{2+}$ dependent PK$C{\alpha}$ and PK$C{\beta}I$) or Rottlerin (an inhibitor of PK$C{\delta}$) has no effect on MTB-induced expression of $IL-1{\beta}$ mRNA. These results show that the expression of $IL-1{\beta}$ mRNA caused by MTB may be mediated via MEK1 and PKC isoforms including PK$C{\beta}II$, $PKC{\gamma}$, or $PKC{\varepsilon}$. Further studies are required to determine whether other PKC isoforms $(PKC {\eta},\;{\theta},\;{\varepsilon},\;and\;{\lambda}/{\iota})$, except $PKC{\delta}$, $PKC{\alpha}$, and $PKC{\beta}I$, are also involved in $IL-1{\beta}$ mRNA expression after mycobacterial infection.

• 생명과학회지
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• 제19권7호
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• pp.892-898
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• 2009

Mifepristone (MIF)은 프로게스테론 유사체이며, 강한 항프로게스테론 효과 때문에 전립선암 치료에 사용되고 있다. 본 연구에서는 MIF의 세포독성효과가 세포 내 칼슘농도 조절에 의한 것임을 밝힌다. 5-40 $\mu$M의 MIF를 처리 시 LNCaP 전립선암세포의 성장이 농도와 시간의존적으로 감소하였다. 반대로, 세포 내 칼슘의 레벨은 MIF의 처리시간과 농도도 의존적으로 증가하였다. MIF를 처리한 세포를 PI 혹은 Hoechst로 염색한 결과, 전형적인 세포자살의 징후인 응축된 염색질과 핵 조각단편들이 관찰되었다. 이들 세포자살징후 역시 MIF의 처리시간과 농도가 증가 할수록 심화되었다. 세포자살에 직접적으로 관여하는 중요한 단백질인 Bcl-2 그룹 단백질의 발현을 조사해 본 결과, 세포자살 억제단백질인 Bcl-2의 발현은 MIF처리시 치명적으로감소하였고, 대신에 세포자살 촉진단백질인 Bax의 발현은 2배로 증대되었다. 이상의 결과로 보아 MIF의 세포독성효과는 세포 내의 칼슘조절에 따른 세포자살에 의한 것으로 생각된다.

• 생명과학회지
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• 제24권9호
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• pp.927-934
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• 2014

혈근초(Sanguinaria canadensis) 뿌리에서 유래된 benzophenanthridine alkaloid의 일종인 sanguinarine은 항균, 항산화 및 항암작용 등 다양한 생리활성을 지니고 있는 것으로 알려져 있다. 비록 TRAIL이 암세포에서는 apoptosis를 유도하지만 정상세포에서는 세포독성을 나타내지 않는다는 큰 장점으로 제 2 임상 단계에서 유의적인 성과를 이루었지만, TRAIL 저항성을 극복해야 하는 큰 어려움이 남아있다. 본 연구실에서는 TRAIL이 세포독성을 나타내지 않는 범위의 sanguinarine과 혼합처리에 의하여 TRAIL 저항성 AGS 위암세포에서 apoptosis를 유발하였음을 보고한 바 있으며, 본 연구에서는 sanguinarine의 TRAIL 저항성 관련 극복에 대한 추가적인 기전 연구를 실시하였다. 본 연구의 결과에 의하면, sanguinarine과 TRAIL의 혼합처리는 각각의 단독 처리에 비하여 AGS 세포의 증식억제 및 apoptosis 유도의 상승 효과가 있었으며, 이를 MTT assay, agarode gel 전기영동, 염색질 응축 현상 및 flow cytometry 분석을 통하여 확인하였다. 또한 sanguinarine과 TRAIL의 혼합처리는 DR5의 발현을 증가시켰으며, ROS의 생성을 촉진시켰다. 그러나 동일 조건에서 MAPKs 신호 전달계에는 큰 영향을 주지 않았다. 아울러 ROS 생성을 인위적으로 차단하였을 경우, sanguinarine과 TRAIL의 혼합처리에 의한 생존도 저하가 유의적으로 회복되었다. 이러한 결과는 sanguinarine과 TRAIL이 DR5의 발현 증가와 ROS의 생성을 촉진시킴으로서 apoptosis 신호를 활성화하였음을 의미하는 결과로서 TRAIL 저항성 극복을 위한 sanguinarine 활용의 유용성을 보여주는 것이다.

• 한국발생생물학회지:발생과생식
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• 제21권4호
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• pp.407-415
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• 2017

In the present study, we investigated the role of binding immunoglobulin protein/glucose-regulated protein, 78-kDa (BIP/GRP78)-regulated endoplasmic reticulum (ER)-stress on meiotic maturation and cumulus cells expansion in porcine cumulus-oocyte complexes (COCs). Previously, it has been demonstrated that unfolded protein response (UPR)-related genes, such as molecules involved in ER-stress defense mechanisms, were expressed in matured oocytes and cumulus cells during in vitro maturation (IVM) of porcine oocytes. However, BIP/GRP78-mediated regulation of ER stress in porcine oocytes has not been reported. Firstly, we observed the effects of knockdown of BIP/GRP78 (an UPR initiation marker) using porcine-specific siRNAs (#909, #693, and #1570) on oocyte maturation. Among all siRNAs, siRNA #693 significantly reduced the protein levels of UPR marker proteins (BIP/GRP78, ATF4, and P90ATF6) in porcine COCs observed by Western blotting and immunofluorescence analysis. We also observed that the reduction of BIP/GRP78 levels by siRNA#693 significantly inhibited the meiotic maturation of oocytes (siRNA #693: $32.5{\pm}10.1%$ vs control: $77.8{\pm}5.3%$). In addition, we also checked the effect of ER-stress inhibitors, tauroursodeoxycholic acid (TUDCA, $200{\mu}M$) and melatonin ($0.1{\mu}M$), in BIP/GRP78-knockdown oocytes. TUDCA and melatonin treatment could restore the expression levels of ER-stress marker proteins (BIP/GRP78, $p-eIF2{\alpha}$, $eIF2{\alpha}$, ATF4, and P90ATF6) in siRNA #693-transfected matured COCs. In conclusion, these results demonstrated that BIP/GRP78-mediated regulation of UPR signaling and ER stress plays an important role in in vitro maturation of porcine oocytes.

• Journal of Microbiology and Biotechnology
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• 제27권9호
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• pp.1628-1638
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• 2017

Viola tianshanica Maxim, belonging to the Violaceae plant family, is traditionally used in Uighur medicine for treating pneumonia, headache, and fever. There is, however, a lack of basic understanding of its pharmacological activities. This study was designed to observe the effects of the ethanol extract (TSM) from Viola tianshanica Maxim on the inflammation response in acute lung injury (ALI) induced by LPS and the possible underlying mechanisms. We found that TSM (200 and 500 mg/kg) significantly decreased inflammatory cytokine production and the number of inflammatory cells, including macrophages and neutrophils, in bronchoalveolar lavage fluid. TSM also markedly inhibited the lung wet-to-dry ratio and alleviated pathological changes in lung tissues. In vitro, after TSM ($12.5-100{\mu}g/ml$) treatment to RAW 264.7 cells for 1 h, LPS ($1{\mu}g/ml$) was added and the cells were further incubated for 24 h. TSM dose-dependently inhibited the levels of proinflammatory cytokines, such as NO, $PGE_2$, $TNF-{\alpha}$, IL-6, and $IL-1{\beta}$, and remarkably decreased the protein and mRNA expression of $TNF-{\alpha}$ and IL-6 in LPS-stimulated RAW 264.7 cells. TSM also suppressed protein expression of $p-I{\kappa}Ba$ and p-ERK1/2 and blocked nuclear translocation of $NF-{\kappa}B$ p65. The results indicate that TSM exerts anti-inflammatory effects related with inhibition on $NF-{\kappa}B$ and MAPK (p-ERK1/2) signaling pathways. In conclusion, our data demonstrate that TSM might be a potential agent for the treatment of ALI.

• Journal of Plant Biotechnology
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• 제41권4호
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• pp.194-200
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• 2014

Monodehydroascorbate reductase (MDHAR)는 활성산소종 제거에 중요한 효소이다. 본 연구에서는 MDHAR 유전자를 현사시나무(Populus alba ${\times}$ P. glandulosa)에서 분리하여 이를 PagMDHAR1이라 명명하고 유전자의 구조와 발현특성을 조사 하였다. PagMDHAR1 유전자는 434개의 아미노산으로 구성된 단백질을 암호화하며 3개의 FAD/NAD(P)H 결합 영역이 보존되어 있다. PagMDHAR1은 현사시나무의 염색체에 1 ~ 2 copy가 존재하며, 배양세포와 꽃에서 높게 발현하였다. 현탁배양세포의 생장주기에서는 유도기와 초기 지수생장기에서 높게 발현하였다. 또한 PagMDHAR1은 건조와 염, 저온, 상처 및 ABA 처리에 의해서 발현이 증가하는 것으로 나타났다. 따라서 PagMDHAR1은 ABA를 경유한 신호전달경로를 따라 다양한 스트레스에 반응하며, PagMDHAR1의 기능이 활성산소종에 의해 유도되는 산화 스트레스 방어기작에서 중요한 역할을 할 뿐만 아니라 스트레스 내성에도 기여할 것으로 생각된다. 이는 향후 PagMDHAR1 형질전환 식물체 생산 등 생명공학적 기술을 이용한 유전자 기능에 대한 연구를 비롯하여 신기능성 임목의 개발에 활용 가능할 것으로 기대된다.

• Molecules and Cells
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• 제26권4호
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• pp.338-343
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• 2008

An embryonic stem cell is a powerful tool for investigation of early development in vitro. The study of embryonic stem cell mediated neuronal differentiation allows for improved understanding of the mechanisms involved in embryonic neuronal development. We investigated expression profile changes using time course cDNA microarray to identify clues for the signaling network of neuronal differentiation. For the short time course microarray data, pattern analysis based on the quadratic regression method is an effective approach for identification and classification of a variety of expressed genes that have biological relevance. We studied the expression patterns, at each of 5 stages, after neuronal induction at the mRNA level of embryonic stem cells using the quadratic regression method for pattern analysis. As a result, a total of 316 genes (3.1%) including 166 (1.7%) informative genes in 8 possible expression patterns were identified by pattern analysis. Among the selected genes associated with neurological system, all three genes showing linearly increasing pattern over time, and one gene showing decreasing pattern over time, were verified by RT-PCR. Therefore, an increase in gene expression over time, in a linear pattern, may be associated with embryonic development. The genes: Tcfap2c, Ttr, Wnt3a, Btg2 and Foxk1 detected by pattern analysis, and verified by RT-PCR simultaneously, may be candidate markers associated with the development of the nervous system. Our study shows that pattern analysis, using the quadratic regression method, is very useful for investigation of time course cDNA microarray data. The pattern analysis used in this study has biological significance for the study of embryonic stem cells.