• 생명과학회지
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• 제27권11호
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• pp.1349-1356
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• 2017

$K^+$ 통로 개방제들은 심근, 뇌, 골격근 등에서 세포막 혹은 미토콘드리아 내막에 존재하는 큰 전도성의 $Ca^{2+}$-의 존성 $K^+$ (BK) 통로 및 ATP-조절성 $K^+$ 통로(ATP-sensitive $K^+$ channels, $K_{ATP}$)에 작용하여 허혈성 혹은 산화성 세포 손상을 완화하는 효과가 있는 것으로 보고되어 있다. 본 연구에서는 망막 색소 상피세포주인 ARPE-19 세포를 실험 모델로 하여 큰 전도성의 BK 통로 개방제인 NS 1619가 유사한 보호 효과를 나타낼 수 있는지, 또한 그 작용기전이 무엇인지를 확인하고자 하였다. AREE-19 세포를 여러 형태의 산화 스트레스에 노출시켜 세포 손상을 유발하고 그 손상의 정도 및 이에 미치는 NS 1619의 효과를 trypan blue 배출능, Tunel 염색 분석을 통하여 측정하였다. NS 1619는 여러 형태의 산화 스트레스에 의한 괴사성 및 apoptosis에 의한 세포 손상을 효과적으로 방지하였으며 그 보호 효과는 BK 통로 봉쇄제인 paxilline 의해 차단되었다. NS 1619는 $H_2O_2$에 의한 세포내 ATP 고갈을 현저히 완화시켰으며, 또한 MTT 환원능으로 측정한 미토콘드리아의 기능을 보호하는 효과를 보였다. 유세포형광 분석법을 이용한 실험에서 NS 1619는 $H_2O_2$에 의한 미토콘드리아 막전압의 소실을 유의하게 방지하였다. 이상의 결과들을 종합하면 NS 1619는 망막 색소 상피세포에서 산화성 세포 손상을 방지하는 효과를 나타내며 그 기전에 미토콘드리아 기능에 대한 보호 작용이 연관되어 있는 것으로 사료된다.

• Journal of Preventive Medicine and Public Health
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• 제39권3호
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• pp.199-204
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• 2006

Objectives: Mercury is a hazardous organ-specific environmental contaminant. It exists in a wide variety of physical and chemical states, each of which has unique characteristics for the target organ specificity. Exposure to mercury vapor and to organic mercury compounds specifically affects the CNS, while the kidney is the target organ for inorganic Hg compounds. Methods: In this study, mercury chloride $(HgCl_2)$ was studied in a renal derived cell system, i.e., the tubular epithelial Madin-Darby canine kidney (MDCK) cell line, which has specific sensitivity to the toxic effect of mercury. MDCK cells were cultured for 6-24 hr in vitro in various concentrations (0.1-100 M) of $HgCl_2$, and the markers of apoptosis or cell death were assayed, including DNA fragmentation, caspase-3 activity andwestern blotting of cytochrome c. The influence of the metal on cell proliferation and viability were evaluated by the conventional MTT test. Results: The cell viability was decreased in a time and concentration dependent fashion: decreases were noted at 6, 12 and 24 hr after $HgCl_2$, exposure. The increases of DNA fragmentation were also observed in the concentrations from 0.1 to 10 M of $HgCl_2$ at 6 hr after exposure. However, we could not observe DNA fragmentation in the concentrations more than 25 M because the cells rapidly proceeded to necrotic cell death. The activation of caspase-3 was also observed at 6 hr exposure in the $HgCl_2$ concentrations from 0.1 to 10 M. The release of cytochrome c from the mitochondria into the cytosol, which is an initiator of the activation of the caspase cascade, was also observed in the $HgCl_2-treated$ MDCK cells. Conclusions: These results suggest that the activation of caspase-3 was involved in $HgCl_2-induced$ apoptosis. The release of cytochrome c from the mitochondria into the cytosol was also observed in the $HgCl_2-treated$ MDCK cells. These findings indicate that in MDCK cells, $HgCl_2$ is a potent inducer of apoptosis via cytochrome c release from the mitochondria.

• Radiation Oncology Journal
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• 제31권2호
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• pp.57-65
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• 2013

Beta-lapachone (${\beta}$-Lap; 3,4-dihydro-2, 2-dimethyl-2H-naphthol[1, 2-b]pyran-5,6-dione) is a novel anti-cancer drug under phase I/II clinical trials. ${\beta}$-Lap has been demonstrated to cause apoptotic and necrotic death in a variety of human cancer cells in vitro and in vivo. The mechanisms underlying the ${\beta}$-Lap toxicity against cancer cells has been controversial. The most recent view is that ${\beta}$-Lap, which is a quinone compound, undergoes two-electron reduction to hydroquinone form utilizing NAD(P)H or NADH as electron source. This two-electron reduction of ${\beta}$-Lap is mediated by NAD(P)H:quinone oxidoreductase (NQO1), which is known to mediate the reduction of many quinone compounds. The hydroquinone forms of ${\beta}$-Lap then spontaneously oxidizes back to the original oxidized ${\beta}$-Lap, creating futile cycling between the oxidized and reduced forms of ${\beta}$-Lap. It is proposed that the futile recycling between oxidized and reduced forms of ${\beta}$-Lap leads to two distinct cell death pathways. First one is that the two-electron reduced ${\beta}$-Lap is converted first to one-electron reduced ${\beta}$-Lap, i.e., semiquinone ${\beta}$-Lap $(SQ)^{{\cdot}-}$ causing production of reactive oxygen species (ROS), which then causes apoptotic cell death. The second mechanism is that severe depletion of NAD(P)H and NADH as a result of futile cycling between the quinone and hydroquinone forms of ${\beta}$-Lap causes severe disturbance in cellular metabolism leading to apoptosis and necrosis. The relative importance of the aforementioned two mechanisms, i.e., generation of ROS or depletion of NAD(P)H/NADH, may vary depending on cell type and environment. Importantly, the NQO1 level in cancer cells has been found to be higher than that in normal cells indicating that ${\beta}$-Lap may be preferentially toxic to cancer cells relative to non-cancer cells. The cellular level of NQO1 has been found to be significantly increased by divergent physical and chemical stresses including ionizing radiation. Recent reports clearly demonstrated that ${\beta}$-Lap and ionizing radiation kill cancer cells in a synergistic manner. Indications are that irradiation of cancer cells causes long-lasting elevation of NQO1, thereby sensitizing the cells to ${\beta}$-Lap. In addition, ${\beta}$-Lap has been shown to inhibit the repair of sublethal radiation damage. Treating experimental tumors growing in the legs of mice with irradiation and intraperitoneal injection of ${\beta}$-Lap suppressed the growth of the tumors in a manner more than additive. Collectively, ${\beta}$-Lap is a potentially useful anti-cancer drug, particularly in combination with radiotherapy.

• Journal of Microbiology and Biotechnology
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• 제31권10호
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• pp.1343-1349
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• 2021

Cockroaches live in places where various pathogens exist and thus are more likely to use antimicrobial compounds to defend against pathogen intrusions. We previously performed an in silico analysis of the Periplaneta americana transcriptome and detected periplanetasin-5 using an in silico antimicrobial peptide prediction method. In this study, we investigated whether periplanetasin-5 has anticancer activity against the human leukemia cell line K562. Cell growth and survival of K562 cells treated with periplanetasin-5 were decreased in a dose-dependent manner. By using flow cytometric analysis, acridine orange/ethidium bromide (AO/EB) staining and DNA fragmentation, we found that periplanetasin-5 induced apoptotic and necrotic cell death in leukemia cells. In addition, these events were associated with increased levels of the pro-apoptotic proteins Fas and cytochrome c and reduced levels of the anti-apoptotic protein Bcl-2. Periplanetasin-5 induces the cleavage of pro-caspase-9, pro-caspase-8, pro-caspase-3, and poly (ADP-ribose) polymerase (PARP). The above data suggest that periplanetasin-5 induces apoptosis via both the intrinsic and extrinsic pathways. Moreover, caspase-related apoptosis was further confirmed by using the caspase inhibitor carbobenzoxy-valyl-alanyl-aspartyl-[O-methyl]-fluoromethylketone (Z-VAD-FMK), which reversed the periplanetasin-5-induced reduction in cell viability. In conclusion, periplanetasin-5 caused apoptosis in leukemia cells, suggesting its potential utility as an anticancer therapeutic agent.

• The Korean Journal of Physiology and Pharmacology
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• 제27권3호
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• pp.231-240
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• 2023

Fabry disease is a lysosomal storage disorder characterized by the lysosomal accumulations of glycosphingolipids in a variety of cytotypes, which include endothelial cells. The disease is inherited and originates from an error in glycosphingolipid catabolism caused by insufficient α-galactosidase A activity, which causes uncontrolled progressive storage of intracellular globotriaosylceramide (Gb3) in the vasculature and extracellular accumulation of lyso-Gb3 (a deacetylated soluble form of Gb3). Necrosis can lead to inflammation, which exacerbates necrosis and creates a positive feedback loop that triggers necroinflammation. However, the role played by necroptosis, a form of programmed necrotic cell death, in the cell-to-cell inflammatory reaction between epithelial and endothelial cells is unclear. Thus, the present study was undertaken to determine whether lyso-Gb3 induces necroptosis and whether necroptosis inhibition protects endothelial dysfunction against lyso-Gb3 inflamed retinal pigment epithelial cells. We found lyso-Gb3 induced necroptosis of a retinal pigment epithelial cell line (ARPE-19) in an autophagy-dependent manner and that conditioned media (CM) from ARPE-19 cells treated with lyso-Gb3 induced the necroptosis, inflammation, and senescence of human umbilical vein endothelial cells. In addition, a pharmacological study showed CM from lyso-Gb3 treated ARPE-19 cells induced endothelial necroptosis, inflammation, and senescence were significantly inhibited by an autophagy inhibitor (3-MA) and by two necroptosis inhibitors (necrostatin and GSK-872), respectively. These results demonstrate lyso-Gb3 induces necroptosis via autophagy and suggest that lyso-Gb3 inflamed retinal pigment epithelial cells trigger endothelial dysfunction via the autophagy-dependent necroptosis pathway. This study suggests the involvement of a novel autophagy-dependent necroptosis pathway in the regulation of endothelial dysfunction in Fabry disease.

• Clinical and Experimental Pediatrics
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• 제48권5호
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• pp.545-550
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• 2005

목 적: 저산소성 허혈성 손상을 받은 신생아의 뇌조직에서 유세포 방법을 통해 세포의 사멸을 분석하기 위해서는 단일세포의 분리가 이루어져야 한다. 본 연구는 세포 분리에 있어서 세포막의 소실을 최소화하고 항원성을 유지하기 위하여 물리적인 방법과 효소 처리를 통한 세포 분리방법의 효율성에 대해 알아보고자 하였다. 방 법 : 생후 7일된 10마리의 SD 신생백서에 우측 경동맥 결찰 후, 8% 산소에 노출시켜 저산소성 허혈증의 손상을 유발하였으며 48시간이 지난 후 뇌조직을 얻어 같은 수의 정상 대조군과 비교하였다. 세포 분리는 물리적인 방법(pipette)과 효소 처리(trypsin 및 collagenase) 방법을 통하여 이루어 졌으며, 세포막의 손상 정도와 범위에 대해서는 annexin V 및 propidium iodide의 형광 염색을 통한 유세포 분석방법을 이용하였다. 결 과 : 정상 대조군에서, 물리적인 방법을 통한 세포 분리가 양반구 모두에서 효소 처리를 한 경우에 비해서 세포의 사멸과 괴사가 통계적으로 유의하게 증가하였다. 저산소성 허혈증을 유발한 군 중, collagenase를 이용하여 세포 분리를 시행한 경우에서 우측 반구의 세포 사멸과 괴사의 비율이 좌측 반구 및 정상 대조군 보다 유의하게 증가하였다. 효소 처리를 통한 세포 분리에서는 서로 유사한 경향을 보였으나, trypsin을 이용한 경우가 collagenase를 이용한 경우에 비해 세포 변화의 정도가 유의하게 감소하였다. 결 론 : 신생아의 뇌조직에서 collagenase를 이용한 단일 세포 분리방법이 세포막의 손상을 최소화하면서 세포막의 성상을 보존할 수 있는 가장 유용한 방법이었다.

• The Plant Pathology Journal
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• 제32권4호
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• pp.371-376
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• 2016

Grapevine Algerian latent virus (GALV) is a member of the genus Tombusvirus in the Tombusviridae and infects not only woody perennial grapevine plant but also herbaceous Nicotiana benthamiana plant. In this study, we developed GALV-based gene expression and virus-induced gene silencing (VIGS) vectors in N. benthamiana. The GALV coat protein deletion vector, pGMG, was applied to express the reporter gene, green fluorescence protein (GFP), but the expression of GFP was not detected due to the necrotic cell death on the infiltrated leaves. The p19 silencing suppressor of GALV was engineered to inactivate its expression and GFP was successfully expressed with unrelated silencing suppressor, HC-Pro, from soybean mosaic virus. The pGMG vector was used to knock down magnesium chelatase (ChlH) gene in N. benthamaina and the silencing phenotype was clearly observed on systemic leaves. Altogether, the GALV-derived vector is expected to be an attractive tool for useful gene expression and VIGS vectors in grapevine as well as N. benthamiana.

• 대한수의학회지
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• 제33권2호
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• pp.301-308
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• 1993

Three or 7day old piglets were infected experimentally with different encephalomyocarditis virus isolates to detect the viral antigen by the immunoperoxidase technique and to observe strain difference in their pathogenecity in newborn pigs by comparing clinical signs and pathologic lesions. Clinical signs of the infected pigs were different depending on the virus strain, pig age and infection route. Encephalomyocarditis virus(EMCV) NVSL-PR isolate was more pathogenic than MN-25 and MN-30 isolate. Three day old piglets showed more severe illness than 7 day old piglets. Predominant clinical signs were sudden death without noticeable clinical signs and dyspnea manifested as heavy abdominal breathing. Contact-infection from infected piglets to controls was observed in the oro-nasally infected group but not the intramuscular group. Common necropsy findings of dead piglets in both age groups infected with MN-25 and NVSL-PR were accumulation of excessive fluid in the body cavities and mild to diffuse necrotic areas observed in the hearts and occasionally in the livers. Microscopically, myocarditis with inflammatory cell infiltration, necrosis of the myocardial muscle fibers and occasional mineralization were observed along with interstitial pneumonia and centrolobular necrosis in the liver. Using an immunoperoxidase technique, viral antigen was detected in myocardial muscle fibers of piglets infected with EMCV.

• Journal of Yeungnam Medical Science
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• 제40권4호
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• pp.343-351
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• 2023

Diabetic foot is one of the most devastating consequences of diabetes, resulting in amputation and possibly death. Therefore, early detection and vigorous treatment of infections in patients with diabetic foot are critical. This review seeks to provide guidelines for the therapy and rehabilitation of patients with moderate-to-severe diabetic foot. If a diabetic foot infection is suspected, bacterial cultures should be initially obtained. Numerous imaging studies can be used to identify diabetic foot, and recent research has shown that white blood cell single-photon emission computed tomography/computed tomography has comparable diagnostic specificity and sensitivity to magnetic resonance imaging. Surgery is performed when a diabetic foot ulcer is deep and is accompanied by bone and soft tissue infections. Patients should be taught preoperative rehabilitation before undergoing stressful surgery. During surgical procedures, it is critical to remove all necrotic tissue and drain the inflammatory area. It is critical to treat wounds with suitable dressings after surgery. Wet dressings promote the formation of granulation tissues and new blood vessels. Walking should begin as soon as the patient's general condition allows it, regardless of the wound status or prior walking capacity. Adequate treatment of comorbidities, including hypertension and dyslipidemia, and smoking cessation are necessary. Additionally, broad-spectrum antibiotics are required to treat diabetic foot infections.

• 한국진공학회:학술대회논문집
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• 한국진공학회 2015년도 제49회 하계 정기학술대회 초록집
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• pp.51.2-51.2
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• 2015

Understanding interfacial phenomena has been one of the main research issues not only in semiconductors but only in life sciences. I have been trying to meet the atomic scale surface and interface analysis challenges from semiconductor industries and furthermore to extend the application scope to biomedical areas. Optical imaing has been most widely and successfully used for biomedical imaging but complementary ion beam imaging techniques based on mass spectrometry and ion scattering can provide more detailed molecular specific and nanoscale information In this presentation, I will review the 27 years history of medium energy ion scattering (MEIS) development at KRISS and DGIST for nanoanalysis. A electrostatic MEIS system constructed at KRISS after the FOM, Netherland design had been successfully applied for the gate oxide analysis and quantitative surface analysis. Recenlty, we developed time-of-flight (TOF) MEIS system, for the first time in the world. With TOF-MEIS, we reported quantitative compositional profiling with single atomic layer resolution for 0.5~3 nm CdSe/ZnS conjugated QDs and ultra shallow junctions and FINFET's of As implanted Si. With this new TOF-MEIS nano analysis technique, details of nano-structured materials could be measured quantitatively. Progresses in TOF-MEIS analysis in various nano & bio technology will be discussed. For last 10 years, I have been trying to develop multimodal nanobio imaging techniques for cardiovascular and brain tissues. Firstly, in atherosclerotic plaque imaging, using, coherent anti-stokes raman scattering (CARS) and time-of-flight secondary ion mass spectrometry (TOF-SIMS) multimodal analysis showed that increased cholesterol palmitate may contribute to the formation of a necrotic core by increasing cell death. Secondly, surface plasmon resonance imaging ellipsometry (SPRIE) was developed for cell biointerface imaging of cell adhesion, migration, and infiltration dynamics for HUVEC, CASMC, and T cells. Thirdly, we developed an ambient mass spectrometric imaging system for live cells and tissues. Preliminary results on mouse brain hippocampus and hypotahlamus will be presented. In conclusions, multimodal optical and mass spectrometric imaging privides overall structural and morphological information with complementary molecular specific information, which can be a useful methodology for biomedical studies. Future challenges in optical and mass spectrometric imaging for new biomedical applications will be discussed.