• 미생물학회지
• /
• 제38권4호
• /
• pp.247-253
• /
• 2002

환경오염원으로서 폭약 2,4,6-trinitrotoluene (TNT)에 대한 TNT 분해세균 Stenotrophomonas sp. OK-5의 세포반응에 대하여 조사하였다. 아치사조건의 TNT농도와 노출시간에 따른 균주 OK-5의 생존율을 분석한 결과, 이 세균의 생존율은 스트레스 충격 단백질의 생성과 비례하였다. 총세포 지방산 조성분석에서 균주 OK-5는 tryp-ticase soy agar에서 자랄 때보다 TNT 배지에서 자랄 때 여러 가지 종류의 지방산이 생성되거나 사라지는 것이 밝혀졌다. 주사전자현미경하에서 TNT에 노출된 세포는 쭈글쭈글하고 불규칙적인 간상형으로 나타났다. Anti-DnaK와 anti-GroEL을 이용하여 SDS-PAGE와 Western blot을 통한 분석으로 균주 OK-5는 70 kDa DanK와 60 kDa GroEL을 포함하는 몇가지 스트레스충격단백질을 생성하는 것으로 밝혀졌다. TNT에 노출된 OK-5 배양에서 수용성 단백질 분획에 대하여 2-D PAGE를 실시하였으며, pH 3에서 pH 10의 범위에서 약 300여 개 spot들이 silver로 염색된 gel상에서 관찰되었다. 이들 가운데 TNT의 반응으로 현저하게 유도되고 발현된 10개의 spot들을 확인하였으며, 2개의 단백질, spot #1과 spot #10에 대한 내부아미노산 서열을 ESI-Q TOF로 분석한 결과, Xylella fastidiosa의 DnaK protein XF2340와 Mesorhizobium loti의 스트레스 유도단백질로 각각 밝혀졌다.

• 한국식품과학회지
• /
• 제53권6호
• /
• pp.756-760
• /
• 2021

현재 시판되는 항생제 및 항결핵약제 대부분은 균의 성장을 저해하는 기전에 맞추어 개발되었고, 지금도 새로운 약제들이 개발되고 있다. 이러한 약제들은 사용빈도가 증가함에 따라 자연발생적으로 약제내성이 출현하게 되고, 약제 내성 결핵의 경우 치료가 더 어려울 뿐만 아니라 치료에 소요되는 시간과 비용이 기하급수적으로 증가한다. 이런 관점에서 볼 때, 인체 면역을 활성화시켜 결핵을 치료하는 방법이 결핵균을 직접적으로 사멸시키는 방법보다 더 안전하고 효과적인 방법이 될 수 있을 것이다. 본 연구에서는 오랜 시간 동안 민간요법으로 사용되어지고 있으면서 탐식구 활성화를 촉진시키는 것으로 알려져 있지만, 항결핵 효과는 검증되어 있지 않은 FRM에 의한 항결핵효과를 검증하였다. 유향의 세포독성효과를 확인하기 위해 MTT reduction assay를 실시하여 31.3, 62.5 ㎍/mL의 농도에서 독성효과를 나타내지 않는 것을 확인하였다. 또한 형태학적 관찰을 통해서도 세포독성 효과를 나타내지 않은 것을 확인하였다. 대식세포의 면역 작용기전을 측정하는 중요한 지표인 cytokine TNF-α, IL-1β의 발현양의 변화를 통해서 면역반응을 확인하였다. 결핵균이 감염된 대조군에서는 이 cytokine들의 발현이 증가한 반면, FRM을 처리한 세포에서는 발현이 감소하였고, 결핵균 또한 감소하였다. 이러한 현상은 FRM이 necrosis를 감소시키면서 apoptosis로 면역반응을 유도하여 발생하는 것으로 가정할 수 있으며, 결과적으로 결핵균 제거 효과를 보여주었다. 현재 치료가 힘든 다제내성결핵, 광범위내성결핵에서 유향이 대식세포의 면역력 활성화를 통한 항결핵효과를 나타내는 치료제로서의 잠재적인 가능성을 제시하였다. 다만, 추가적인 항결핵효과의 검증 및 면역기전에 대한 연구가 이루어져야 할 것으로 사료된다.

• 미생물학회지
• /
• 제54권4호
• /
• pp.343-353
• /
• 2018

이전 연구에서 우리는 RDX (hexahydro-1,3,5-trinitro-1,3,5-triazine) 분해세균 Pseudomonas sp. HK-6에서 xenobiotic reductase B를 암호화하는 xenB 유전자의 돌연변이 균주를 이용하여 RDX 스트레스에 대한 xenB 유전자의 역할에 관하여 연구를 보고하였다[Lee et al. (2015) Curr. Microbiol. 70(1): 119-127]. 본 연구에서는 Pseudomonas sp. HK-6 xenA 돌연변이 균주로 연구 범위를 확대하여 RDX 스트레스 조건에서 세포반응과 프로테옴 프로필의 변화를 분석하였다. RDX 첨가 배지에서 xenA 돌연변이 균주는 야생균주와 비교하여 RDX를 약 2배 정도 느리게 분해하였으며, RDX 스트레스 하에서 xenA 돌연변이 균주의 생장률과 생존율은 야생균주와 비교하여 낮았다. RDX 스트레스에 의한 심한 형태적 손상이 xenA 돌연변이 균주의 세포 표면에 발생하는 것이 주사전자현미경을 통해서 확인되었다. RDX 스트레스 하에서 야생균주에서 발현된 충격단백질인 DnaK 및 GroEL의 양은 배양 초기 혹은 상대적으로 낮은 RDX 농도에서는 증가하였으나, 배양시간이 길어지거나 높은 RDX 농도에서는 다소 감소하였다. 그러나 xenA 돌연변이 균주에서는 DnaK와 GroEL의 발현양은 RDX 농도가 증가함에 따라 점차 감소되었다. RT-qPCR에 의해 측정된 야생균주에서 dnaA와 groEL의 전사 수준은 RDX 스트레스가 증가된 상태에서 잘 유지되었으나, xenA 돌연변이 균주에서는 점차 감소되어 결국에는 소멸되었다. RDX 스트레스에서 xenA의 돌연변이에 의한 프로테옴 프로필의 변화를 2-DE PAGE를 통해서 관찰한 결과에 따르면 27개 단백질이 감소하고 3개가 증가한 것으로 나타났다. 이들 결과로 보아, 정상적인 xenA 유전자는 RDX 스트레스 하에서 세포의 온전한 형태 유지와 효율적인 RDX 분해 과정을 수행하기 위해서 필요하다는 것을 의미하였다.

• Animal cells and systems
• /
• 제5권1호
• /
• pp.77-83
• /
• 2001

The present study was conducted to elucidate whether the expression level of poly(ADP-ribose) polymerase (PARP) is related to the ultraviolet radiation (UV)-induced apoptosis. After treatment of the mammalian cell lines HeLa S3 and Chinese hamster ovary (CHO) with 50 J/m2 UV, induction of apoptosis was determined by several means during 24 h post-incubation. Incidence of apoptosis was much lower in CHO than HeLa S3 cells based on the percentage of apoptotic cells in terms of morphological changes in nucleus or direct counting of viable cells and qualitative or quantitative DNA fragmentation. Interestingly, when the expression level of PARP was measured by western blotting, the amounts of PARP that was retained at each time point inversely correlated with the incidences of apoptosis in these cells. Concomitant with generation of the 85 kDa fragment, 116 kDa PARP disappeared in HeLa S3 within 6 h after UV treatment, whereas a fair amounts of 116 kDa band was still retained in CHO cells at 36 h post-incubation. This inverse relationship was also observed in the adaptive response system, in which cells weve treated with a high dose of UV after pretreatment with a low dose. As expected, typical adaptive responses appeared in CHO cells but not in HeLa cells, showing greater cell viability and lesser DNA fragmentation. During the adaptive response in CHO cells, PARP was expressed at much higher level compared to the single, high dose-treated cells. Interestingly, even though PARP was induced at 6 h post-incubation In both cell types, its expression was more prominent in CHO cells. Thus, our data indicate that the retained level of intact PARP against UV damage inversely correlates with incidence of apoptosis in mammalian cells, and also suggest that a machinery to protect the PARP degradation against UV damage exists in CHO but not in HeLa S3 cells.

• 원예과학기술지
• /
• 제33권6호
• /
• pp.845-853
• /
• 2015

The objective of this study was to predict the timing of nitrogen (N) demand through analyzing chlorophyll fluorescence (ChlF), soil-plant analysis development (SPAD), and normalized difference vegetation index (NDVI), which are positively correlated with foliar N concentration in star cluster (Pentas lanceolata). The plants were grown in potting soil under optimal conditions for 30 d, followed by weekly irrigation with five concentrations (0, 4, 8, 16, and 24 mM) of N for an additional 30 d. These five N application levels corresponded to leaf N concentrations of 2.62, 3.48, 4.00, 4.23, and 4.69%, respectively. We measured 13 morphological and physiological parameters, as well as the responses of these parameters to various N-fertilizer treatments. The general increases in Dickson's quality index (DQI), above-ground dry weight (DW), total DW, flowering rate, ${\Delta}F/Fm$', and qP in response to treatment with 0 to 8 mM N were similar to those of SPAD, NDVI, and Fv/Fm. Consistent and strong correlations ($R^2$= 0.60 to 0.85) were observed between leaf N concentration (%) and SPAD, NDVI, ${\Delta}F/Fm$', and above-ground DW. Validation of leaf S PAD, NDVI, and ${\Delta}F/Fm$' revealed that these vegetation indices are accurate predictors of leaf N concentration that can be used for non-destructive estimation of the proper timing for N-solution irrigation of P. lanceolata. Moreover, irrigation with 8 mM N-fertilizer i s recommended w hen leaf N concentration, SPAD, NVDI, and ${\Delta}F/Fm$' ratios are reduced from their saturation values of 4.00, 50.68, 0.64, and 0.137%, respectively.

• Molecular & Cellular Toxicology
• /
• 제4권1호
• /
• pp.66-71
• /
• 2008

Long term exposure of Jurkat cells to 2 ATA pressure resulted in the inhibition of cell growth. Under a 2 ATA pressure, the morphological changes in the cells were visualized by electron microscopy. The cells exhibited significant inhibitory responses after three passages. However, short-term exposure study was carried out, 2 ATA pressure may have beneficial effects. The Jurkat cells were exposed to $H_2O_2$ (25 and $50{\mu}M$) in order to induce DNA damage, and then incubated under at either normal pressure or 2 ATA for 1 or 2 hours in order to recover the DNA damage. The extent of DNA damage was determined via Comet assay. More recovery from DNA damage was observed at 2 ATA than at normal pressure. The activity of the DNA repair enzymes, DNA polymerase-$\beta$, was also evaluated at both normal pressure and 2 ATA. The activity of DNA polymerase-$\beta$ was observed to have increased significantly at the 2 ATA than at normal pressure. In conclusion, the effects of hyperbaric pressure from 1 ATA to 2 ATA on biochemical systems can be either beneficial or harmful. Long term exposure to hyperbaric pressure clearly inhibited cell proliferation and caused genotoxic effects, but short-term exposure to hyperbaric pressure proved to be beneficial in terms of bolstering the DNA repair system. The results of the present study have clinical therapeutic application, and might prove to be an useful tool in the study of genotoxicity in the future.

• Journal of Periodontal and Implant Science
• /
• 제41권2호
• /
• pp.67-72
• /
• 2011

Purpose: The aim of this study is to determine whether certain biomaterials have the potential to support cell attachment. After seeding bone marrow stromal cells onto the biomaterials, we investigated their responses to each material in vitro. Methods: Rat bone marrow derived stromal cells were used. The biomaterials were deproteinized bovine bone mineral (DBBM), DBBM coated with fibronectin (FN), synthetic hydroxyapatite (HA), HA coated with FN, HA coated with $\beta$-tricalcium phosphate (TCP), and pure $\beta$-TCP. With confocal laser scanning microscopy, actin filaments and vinculin were observed after 6, 12, and 24 hours of cell seeding. The morphological features of cells on each biomaterial were observed using scanning electron microscopy at day 1 and 7. Results: The cells on HA/FN and HA spread widely and showed better defined actin cytoskeletons than those on the other biomaterials. At the initial phase, FN seemed to have a favorable effect on cell adhesion. In DBBM, very few cells adhered to the surface. Conclusions: Within the limitations of this study, we can conclude that in contrast with DBBM not supporting cell attachment, HA provided a more favorable environment with respect to cell attachment.

• 대한전기학회:학술대회논문집
• /
• 대한전기학회 2009년도 제40회 하계학술대회
• /
• pp.1232_1233
• /
• 2009

Paraquat is well-known to cause hepatotoxic responses in human and other mammal species. In solution, it forms free radicals and charge-transfer complex of which formation plays an important role in determination of its biological activity in the presence of various anions. The HepG2 cells were cultured onto a quartz crystal sensor which is possible to detect the density and a viscosity changes using the resonance frequency (F) and the resonance resistance (R). The plot of F-R diagram is able to explain the rheological change of cells onto the surface of the quartz crystal sensor. In this paper, we investigated the physical properties of the HepG2 cells cultured onto a ITO electrode of the quartz crystal sensor according to the paraquat injection at various concentrations (100 mM, 10 mM, 1 mM). We also observed the morphological changes with a micro CCD camera, simultaneously. The HepG2 cells were cultured onto the ITO electrode surface of the quartz crystal modified a collagen film in $CO_2$ incubator. After the paraquat injection, we observed the changes of the morphologies by the micro CCD camera depending on time and analyzed the physical changes of cells on the electrode surface of quartz crystal using F-R diagram. From all results, we proved the effect of paraquat at various concentrations which is led to an apoptosis such as weakening and death of the cells by oxidation and reduction reaction that were produced the superoxide anions and other free radicals.

• 한국환경농학회지
• /
• 제17권4호
• /
• pp.335-340
• /
• 1998

This study was conducted to determine the morphological responses of barley seedlings to NaCl stress and to investigate histological changes of cells with transmission electron microscope(TEM) after NaCl stress. Plant height and root length of 10-day old barley seedlings with NaCl stress were reduced and inhibition level was found to be more severe in the plant height than in the root length. The leaf length, leaf width and leaf area were shorter as well with NaCl stress than without NaCl stress. However, there was no difference in the number of roots between NaCl treatments. The weight of dry matter decreased at higher NaCl concentrations, especially at 100mM NaCl. The water content of shoots tend to decrease at higher NaCl concentrations, but there was no difference in the water content of roots, The reduced sugar content was greatly increase than starch. Cellulose content was higher in NaCl stressed-plant than control, and tended to decreased at higher NaCl concentrations. Lignin content also decreased NaCl stressed-plant but there was no tendency at NaCl stress concentrations. Electric conductivity of cell sap with seedlings was high with NaCl stressed-plant. Amount of cell sap gradually increased with time in the roots than in the shoots, The grana of chloroplasts was changed by 150mM NaCl concentration. The christe of mitochondria in root meristematic sells ruined in structure and cell wall of leaf and root was also ruined by NaCl stress.

• Journal of Microbiology and Biotechnology
• /
• 제13권4호
• /
• pp.529-536
• /
• 2003

The growth-inhibitory activity of Cassia tora seed-derived materials against seven intestinal bacteria was examined in vitro, and compared with that of anthraquinone, anthraflavine, anthrarufin, and 1-hydroxyanthraquinone. The active constituent of C. tore seeds was characterized as quinizarin, using various spectroscopic analyses. The growth responses varied depending on the compound, dose, and bacterial strain tested. At 1 mg/disk, quinizarin exhibited a strong inhibition of Clostridium perfringens and moderate inhibition of Staphylococcus aureus without any adverse effects on the growth of Bifidobacterium adolescentis, B. bifidum, B. longum, and Lactobacillus casei. Furthermore, the isolate at 0.1 mg/disk showed moderate and no activity against C. perfringens and S. aureus. The structure-activity relationship revealed that anthrarufin, anthraflavine, and quinizarin moderately inhibited the growth of S. aureus. However. anthraquinone and 1-hydroxyanthraquinone did not inhibit the human intestinal bacteria tested. As for the morphological effect of 1 mg/disk quinizarin, most strains of C. perfringens were damaged and disappeared, indicating that the strong activity of quinizarin was morphologically exhibited against C. perfringens. The inhibitory effect on aflatoxin $B_1$ biotransformation by anthraquinones revealed that anthrarufin ($IC_50,\;11.49\mu\textrm{M}$) anthraflavine ($IC_50,\;26.94\mu\textrm{M}$), and quinizarin ($IC_50,\;4.12\mu\textrm{M}$), were potent inhibitors of aflatoxin ${B_1}-8,9-epoxide$ formation. However, anthraquinone and 1-hydroxyanthraquinone did not inhibit the mouse liver microsomal sample to convert aflatoxin $B_1$ to aflatoxin ${B_1}-8,9-epoxide$. These results indicate that the two hydroxyl groups on A ring of anthraquinones may be essential for inhibiting the formation of aflatoxin ${B_1}-8,9-epoxide$. Accordingly, as naturally occurring inhibitory agents, the C. tora seed-derived materials described could be useful as a preventive agent against diseases caused by harmful intestinal bacteria, such as clostridia, and as an inhibitory agent for the mouse liver microsomal conversion of aflatoxin $B_1$ to aflatoxin ${B_1}-8,9-epoxide$.