• Biomolecules & Therapeutics
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• 제20권2호
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• pp.165-170
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• 2012

Cell migration plays a role in many physiological and pathological processes. Reactive oxygen species (ROS) produced in mammalian cells influence intracellular signaling processes which in turn regulate various biological activities. Here, we investigated whether melanoma cell migration could be controlled by ROS production under normoxia condition. Cell migration was measured by wound healing assay after scratching confluent monolayer of B16F10 mouse melanoma cells. Cell migration was enhanced over 12 h after scratching cells. In addition, we found that ROS production was increased by scratching cells. ERK phosphorylation was also increased by scratching cells but it was decreased by the treatment with ROS scavengers, N-acetylcysteine (NAC). Tumor cell migration was inhibited by the treatment with PD98059, ERK inhibitor, NAC or DPI, well-known ROS scavengers. Tumor cell growth as judged by succinate dehydrogenase activity was inhibited by NAC treatment. When mice were intraperitoneally administered with NAC, the intracellular ROS production was reduced in peripheral blood mononuclear cells. In addition, B16F10 tumor growth was significantly inhibited by in vivo treatment with NAC. Collectively, these findings suggest that tumor cell migration and growth could be controlled by ROS production and its downstream signaling pathways, in vitro and in vivo.

• KSBB Journal
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• 제25권3호
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• pp.257-264
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• 2010

Bone morphogenetic protein-7 (BMP-7) is one of important growth factors for skeletal development and bone growth. In this work, BMP-7 was efficiently expressed in recombinant Bacillus subtilis. The mature BMP-7 protein indicated molecular weight of 15.4 kDa by Western blot assay and was secreted into culture medium with 0.35 ng/mL. The extracellular and intracellular rhBMP-7 proteins were purified by using a FPLC system with an ion exchange column and a gel filtration column. The extracellular and intracellular rhBMP-7 proteins had finally a 57.1% purity and a 36.2% purity, respectively. The purified rhBMP-7 proteins showed an intact biological activity which stimulated alkaline phophatase (ALP) activity in MC3T3-E1 cells.

• 한국응용약물학회:학술대회논문집
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• 한국응용약물학회 2003년도 Annual Meeting of KSAP : International Symposium on Pharmaceutical and Biomedical Sciences on Obesity
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• pp.66-66
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• 2003

Flavonoids are phenolic compounds widely distributed in wide variety of edible plants including leafy vegetables, fruits, beverages. Quercetin is one of bioflavonoid compounds and has anti-tumor effect by suppressing tumor growth in vitro and in vivo, including multiple biological effects by antioxidant and effective anti-inflammatory agent. The present study investigated whether quercetin can enhance antioxidant enzyme activities (glutathione peroxidase: GPX, superoxide dismutase : SOD, catalase: CAT) and intracellular reactive oxygen intermediate (ROI) levels in the presence of taurine on B16F10 murine melanoma cells. From this result, the antioxidant enzyme activities of quercetin in the presence of taurine was enhanced. In addition, the same treatments decreased intracellular reactive oxygen intermediate levels on B16F10 murine melanoma cells. Taken together, these results demonstrate that the antioxidant effect of quercetin can enhance in the presence of taurine and it might play an important role in anti-tumor effect.

• Bulletin of the Korean Chemical Society
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• 제23권10호
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• pp.1451-1489
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• 2002

Sphingosine is known to regulate a wide range of cell physiology including growth, differentiation, and apoptosis. In this study, we examined the effect of sphingosine on the phospholipase D (PLD) activity in rat thymocytes. Sphingosine potently stimulated PLD in the absence of extracellular calcium, while depletion of intracellular calcium by BAPTA/AM treatment completely blocked activation of PLD by sphingosine. Sphingosine-induced increase of the intracellular calcium concentration was confirmed using a fluorescent calcium indicator Fluo-3/AM. A phosphoinositide-specific phospholipase C inhibitor U73122 partially inhibited the stimulation of PLD by sphingosine. When mouse PLD2 gene was transfected into mouse thymoma EL4 cells, which lack intrinsic PLD activity, sphingosine could stimulate PLD2 significantly while overexpression of human PLD1 had no effect. Taken together, the sphingosine-stimulated PLD activity in rat thymocytes is dependent on the mobilization of intracellular calcium and appears to be due to the PLD2 isoform.

• Animal cells and systems
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• 제4권3호
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• pp.273-278
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• 2000

Transforming growth factor-$\beta$ (IGF-$\beta$)s are multifunctional small polypeptides synthesized in most cell types. TGF-$\beta$ exerts pivotal effects on both bone formation and resorption. In addition, increasing lines of evidence implicate TGF-$\beta$ as a potential coupling factor between these two processes during bone remodeling. In the present study, the expression form and the activation mechanism of latent-TGF-$\beta$ were investigated using specific antibodies for each isoform. TGF-$\beta$s were observed to be synthesized and accumulated in a large amount in cultured osteoblastic cells. The estimated molecular weights of intracellular TGF-$\beta$2 and -$\beta$3 were 49 and 55 kDa, respectively. Based on proteolytic digestion study and immunofluorescence observation, these precursor forms seemed to be accumulated in distinct intracellular compartments. To examine whether the internal pool of TGF-$\beta$ was possiblely regulated by external signals, their biological activites were examined in a conditioned media of this cell. Although the intact conditioned media did not contain detectable TGF-$\beta$ activity, heat-treatment or acid-activation of the conditioned media revealed significant TGF-$\beta$ activity. Furthermore, in the presence of estrogen, this activity was dramatically diminished. It is known that activation of latent TGF-$\beta$ can be achieved by different chemical and enzymatic treatments, or by incubation with certain cell types. This extracellular activation was suggested as a key step in the regulation of TGF-$\beta$ activity. In addition to these extracellular activation, this study suggests that the synthesis and intracellular processing are important regulation steps for TGF-$\beta$ action. In addition, this regulation Is specific for TGF-$\beta$ type 2, because the change was not observed in TGF-$\beta$3 in osteoblastic cell line.

• 한국방사성폐기물학회:학술대회논문집
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• 한국방사성폐기물학회 2004년도 학술대회지
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• pp.212-212
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• 2004

• 대한한방부인과학회지
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• 제25권3호
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• pp.27-39
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• 2012

Objectives: Recently Ciprofloxacin, used in the treatment of mastitis, showed many serious side effects. The object of this study was to recognize whether TNS and KWT can be used in the treatment of mastitis by observing the in vitro antibacterial effects of TNS and KWT aqueous herbal extracts against S. aureus. Methods: Antibacterial activities of TNS and KWT aqueous extracts against S. aureus ATCC 25923 were detected using standard agar microdilution methods. In addition, the effects on the bacterial growth curve were monitored at MIC and $MIC{\times}2$ levels. The effects on the intracellular killing and bacterial invasion of individual test materials were also observed using Raw 264.6 and MCF-7. The results were compared with Ciprofloxacin, a second generation of quinolone antibiotics in the present study. Results: MIC of aqueous extracts of TNS and KWT against S. aureus were detected as ($0.313{\pm}0.107$) and ($0.137{\pm}0.053$) mg/ml, respectively. MIC of Ciprofloxacin was detected as ($0.469{\pm}0.297$) ${\mu}g/ml$ at same conditions. In addition, TNS, KWT aqueous herbal extracts and Ciprofloxacin were also showed marked dosage-dependent inhibition of bacterial growth, and dramatical inhibitions on the both intracellular killing assays and bacterial invasion using Raw 264.6 and MCF-7 cells were detected. Conclusions: The results obtained in this study suggest that TNS and KWT aqueous herbal extracts showed antibacterial effects against S. aureus, and they also showed dosage-dependent inhibitory effects on the bacterial growth. And they showed the significant intracellular killing and bacterial invasion effects. It means, KWT and TNS may show more potent anti-infectious effects against S. aureus in vivo.

• Applied Biological Chemistry
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• 제41권1호
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• pp.53-59
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• 1998

수종의 두과작물과 수도에서 분리한 Rhizobium 및 Azospirillum들의 내염성을 조사하고 salt stress에 의하여 유도되는 균주의 생리적 특성 및 식물이나 미생물에서 osmoprotectant 로 작용하는 proline, glycine betaine 및 glutamate가 질소고정균의 생육이나 질소고정력에 미치는 영향을 연구하였다. 분리된 대부분의 질소고정균들은 0.6 M NaCl 농도에서 생육이 현저히 감소되었지만 Acacia rhizobia sp86 은 1.4 M NaCl 농도에서도 생육이 가능하였다. Osmotic stress에 의하여 증가되는 intracellular 유리아미노산은 Rhizobium 및 Azospirillum에서 glutamate 였으며, 특히 Acacia rhizobia sp86은 salt stress에 의하여 5배정도 glutamate를 축적하였다. Osmoprotectant (proline, glycine betaine, glutamate)를 배지내에 1 mM 첨가함으로서 salt stress에 의하여 감소되는 질소고정균의 생육과 질소고정력을 방지하였으며, glycine betaine이 가장 효과적이었다.

• Asian-Australasian Journal of Animal Sciences
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• 제15권7호
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• pp.1045-1050
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• 2002

The aims of this study were to detect spontaneously occurring apoptosis in cultured porcine ovarian cells, to examine the role of growth hormone (GH), tyrosine kinase (TK), protein kinase G (PKG) and cyclin-dependent kinase (CDK) in the control of this process, and to determine whether the effect of GH on apoptosis is mediated by TK-, PKG- and cdc2-dependent intracellular mechanisms. We studied the action of pGH (10 ng/ml), blockers of TK (genistein, lavendustin, both 100 ng/ml), PKG (Rp-Br-PET-cGMPS, 50 nM; KT5823, 100 ng/ml) and CDK (olomoucine, $1{\mu}g/ml$), as well as combinations of GH with these blockers, on the onset of apoptosis in cultured granulosa cells isolated from antral (3-6 mm) porcine follicles. The functional characteristics of an early apoptotic event, DNA fragmentation, were determined using terminal deoxynucleotidyltransferase (TdT)-mediated dUTP nick end labelling (TUNEL), whilst morphological signs of advanced apoptosis such as pyknosis, chromatin marginalization, shrinkage and fragmentation of nucleus, were detected using routine light microscopy. After culture, some ovarian granulosa cells exhibited DNA fragmentation, which in some cases was associated with morphological apoptosis-related changes (pyknosis, shrinkage and fragmentation of the nucleus). GH significantly reduced the proportion of TUNEL-positive cells. Neither TK nor CDK blockers when given alone, significantly affected the percentage of TUNEL-positive cells although both PKG blockers significantly increased this index. Furthermore, TK and PKG blockers given together with GH, prevented or reversed the inhibitory effect of GH on apoptosis, whilst the CDK blocker olomoucine promoted it. These observations demonstrate apoptosis in porcine ovaries and suggest the involvement of GH, TK, PKG and CDK in the control of this process. They also suggest that the effect of GH on ovarian apoptosis is mediated or regulated by multiple signalling pathways including TK-, PKG- and CDK-dependent intracellular mechanisms.

• Journal of Microbiology and Biotechnology
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• 제28권10호
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• pp.1736-1748
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• 2018

Brucella abortus can survive and replicate within host macrophages, and great efforts have been made to demonstrate the genes involved in pathogenicity, such as internalization, in Brucella research. Here, intracellular responses were compared between THP-1 macrophage cells stimulated with B. abortus wild-type and four mutants (C1, C10, C27, and C32) using microarray to demonstrate the role of genes related to intracellular survival and replication. These mutants were generated by deleting genes encoding BAB_RS13225 (4-hydrobenzoate 3-monooxygenase, PHBH), BAB_RS00455 (heme exporter protein cytochrome C, CcmC), BAB_RS03675 (exopolyphosphatase, PPX), and BAB_RS13225 (peptidase M24). The results showed that mutants C1 and C10 induced significant suppression of survival levels and cytokine expression relative to wild-type in the THP-1 macrophage cells. These findings suggest that the BAB_RS13225 and BAB_RS00455 genes play important roles in survival within human macrophages. Conversely, mutants C27 and C32 induced significantly higher survival level than wild-type in the cells inhibiting cellular signal transduction. It is assumed that the BAB_RS03675 and BAB_RS13225 genes play a role in cellular resistance to B. abortus. Therefore, the disrupted genes are involved in B. abortus intracellular growth, and especially in its survival, and they could be effective targets for understanding the intracellular bacterium, B. abortus.