• Journal of Microbiology
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• 제33권4호
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• pp.328-333
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• 1995

$\^$31/ P-NMR experiment was performed to detect phophonates (Pn) utilization and degradation in the several different C-P lyase mutants of E. coli and in E. aerogenes and the recombinants. The relative peak intensity (RPI) for the standard samples of 0.5 mM methylphosphonate (MPn) and 1.0 mM aminoethylphosphonate in glucose-MOPS medium showed 0.5 : 1.0 ratio. In the case of BW14329 (.DELTA.phnC-P, .delta.phoA), RPI did not change significantly after 24 hrs culturing, which means it nearly could not utilize Pn. In vivo $\^$31/ P-NMR spectrum of E. aerogens (BWKL 16627) during 3 hrs starvation showed two intense peaks at 0-2 ppm and at near-10 ppm which indicate intracellular orthophosphate (Pi) and pyrophosphate (PPi), respectively. Both of them might be released by degradation of inorganic polyphosphate pool. When MPn is supplied to the medium as an unique P source, Pi content in the cell has the constant, but PPi seems to be slightly decreased. Recombinants (BWKL 16954) grew slower than E. aerogenes in the glucose-MOPS media with various P sources. In vivo $\^$31/ P-NMR spectrum of recombinant did not show any intense signal in the cell. Surprisingly, under the cultivation adding with MPn, a few intense peaks in the region of Pi AND phospate monoester were detected.

• Journal of Microbiology and Biotechnology
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• 제32권8호
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• pp.982-988
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• 2022

Licorice (Glycyrrhiza) has been used as preventive and therapeutic material for hyperpigmentation disorders. Previously, we isolated noble compounds including dehydroglyasperin C (DGC), dehydroglyasperin D (DGD) and isoangustone A (IAA) from licorice hexane/ethanol extracts. However, their anti-melanogenic effects and underlying molecular mechanisms are unknown. The present study compared effects of DGC, DGD and IAA on pigmentation in melan-a melanocytes and human epidermal melanocytes (HEMn). DGD exerted the most excellent anti-melanogenic effect, followed by DGC and IAA at non-cytotoxic concentrations. In addition, DGD significantly inhibited tyrosinase activity in vitro cell-free system and cell system. Western blot result showed that DGD decreased expression of microphthalmia-associated transcription factor (MITF), tyrosinase and tyrosinase-related protein-1 (TRP-1) in melan-a cells and HEMn cells. DGD induced phosphorylation of MITF, ERK and Akt signal pathway promoting MITF degradation system. However, DGD did not influence p38 and cAMP-dependent protein kinase (PKA)/CREB signal pathway in melan-a cells. These result indicated that DGD inhibited melanogenesis not only direct regulation of tyrosinase but also modulating intracellular signaling related with MITF level. Collectively, these results suggested a protective role for DGD against melanogenesis.

• The Korean Journal of Physiology and Pharmacology
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• 제27권5호
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• pp.449-456
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• 2023

N-methyl-D-aspartate (NMDA) receptors are ionic glutamine receptors involved in brain development and functions such as learning and memory formation. NMDA receptor inhibition is associated with autophagy activation. In this study, we investigated whether the NMDA receptor antagonists, memantine and ifenprodil, induce autophagy in human retinal pigment epithelial cells (ARPE-19) to remove N-retinylidene-N-retinylethanolamine (A2E), an intracellular lipofuscin component. Fluorometric analysis using labeled A2E (A2E-BDP) and confocal microscopic examination revealed that low concentrations of NMDA receptor antagonists, which did not induce cytotoxicity, significantly reduced A2E accumulation in ARPE-19 cells. In addition, memantine and ifenprodil activated autophagy in ARPE-19 cells as measured by microtubule-associated protein 1A/1B-light chain3-II formation and phosphorylated p62 protein levels. Further, to understand the correlation between memantine- and ifenprodil-mediated A2E degradation and autophagy, autophagy-related 5 (ATG5) was depleted using RNA interference. Memantine and ifenprodil failed to degrade A2E in ARPE-19 cells lacking ATG5. Taken together, our study indicates that the NMDA receptor antagonists, memantine and ifenprodil, can remove A2E accumulated in cells via autophagy activation in ARPE-19 cells.

• Preventive Nutrition and Food Science
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• 제5권4호
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• pp.184-188
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• 2000

HMG-CoA reductase is th rate-limiting enzyme of cholesterol biosynthesis. As intracellular levels of cholesterol should be regulated elaborately in response to external stimuli an internal needs, the expression of the HMG-CoA reductase gene is regulated intricately at several different levels from transcription to post-translational modification. In this study, we investigated the regulatory mechanism of HMG-CoA reductase gene expression at the post-transcriptional/pre-translational levels in a baby hamster kidney cell line, C100. when 25-hydroxycholesterol was added to cells cultured in medium containing 5% delipidized fetal bovine serum and 25$\mu$M lovastatin, the levels of HMG-CoA reductase mRNA decreased rapidly, which seemed to be due to the increased degradation of reductase mRNA. These suppressive effects of 25-hydroxycholesterol on MG-CoA reductase mRNA levels were blocked by a translation inhibitor, cycloheximide. Similarly, actinomycin D and 5,6-dichloro-1-$\beta$-D-ribofuranosylbenzimidazole, transcription inhibitors, blocked the 25-hydroxycholesterol-mediated degradation of HMG-CoA reductase mRNA. These results indicate that new protein/RNA synthesis is required for the degradation of HMG-CoA reductase mRNA. In addition, data from the transfection experiments shows that cis-acting determinants, regulating the stability of reductase mRNA, were scattered in the sequence corresponding to 1766-4313 based on the sequence of Syrian hamster HMG-CoA reductase cDNA. Our data suggests that sterol-mediated destabilization of reductase mRNA might be one of the important regulatory mechanism of HMG-CoA reductase gene expression.

• Tuberculosis and Respiratory Diseases
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• 제85권2호
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• pp.147-154
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• 2022

Background: The expression of calcium signaling pathway molecules is altered in various carcinomas, which are related to the proliferation and altered characteristics of cancer cells. However, changes in calcium signaling in anti-cancer drug-resistant cells (bearing a T790M mutation in epidermal growth factor receptor [EGFR]) remain unclear. Methods: Afatinib-mediated changes in the level of store-operated Ca²⁺ entry (SOCE)-related proteins and intracellular Ca²⁺ level in non-small cell lung cancer cells with T790M mutation in the EGFR gene were analyzed using western blot and ratiometric assays, respectively. Afatinib-mediated autophagic flux was evaluated by measuring the cleavage of LC3B-II. Flow cytometry and cell proliferation assays were conducted to assess cell apoptosis and proliferation. Results: The levels of SOCE-mediating proteins (ORAI calcium release-activated calcium modulator 1 [ORAI1], stromal interaction molecule 1 [STIM1], and sarco/endoplasmic reticulum Ca²⁺ ATPase [SERCA2]) decreased after afatinib treatment in non-small cell lung cancer cells, whereas the levels of SOCE-related proteins did not change in gefitinib-resistant non-small cell lung cancer cells (PC-9/GR; bearing a T790M mutation in EGFR). Notably, the expression level of SOCE-related proteins in PC-9/GR cells was reduced also responding to afatinib in the absence of extracellular Ca²⁺. Moreover, extracellular Ca²⁺ influx through the SOCE was significantly reduced in PC-9 cells pre-treated with afatinib than in the control group. Additionally, afatinib was found to decrease the level of SOCE-related proteins through autophagic degradation, and the proliferation of PC-9GR cells was significantly inhibited by a lack of extracellular Ca²⁺. Conclusion: Extracellular Ca²⁺ plays important role in afatinib-mediated autophagic degradation of SOCE-related proteins in cells with T790M mutation in the EGFR gene and extracellular Ca²⁺ is essential for determining anti-cancer drug efficacy.

• Journal of Microbiology and Biotechnology
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• 제34권2호
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• pp.407-414
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• 2024

Phosphorus is an essential but non-renewable nutrient resource critical for agriculture. Luxury phosphorus uptake allows microalgae to synthesize polyphosphate and accumulate phosphorus, but, depending on the strain of algae, polyphosphate may be degraded within 4 hours of accumulation. We studied the recovery of phosphorus from wastewater through luxury uptake by an engineered strain of Synechocystis sp. with inhibited polyphosphate degradation and the effect of this engineered Synechocystis biomass on lettuce growth. First, a strain (∆phoU) lacking the phoU gene, which encodes a negative regulator of environmental phosphate concentrations, was generated to inhibit polyphosphate degradation in cells. Polyphosphate concentrations in the phoU knock-out strain were maintained for 24 h and then decreased slowly. In contrast, polyphosphate concentrations in the wild-type strain increased up to 4 h and then decreased rapidly. In addition, polyphosphate concentration in the phoU knockout strain cultured in semi-permeable membrane bioreactors with artificial wastewater medium was 2.5 times higher than that in the wild type and decreased to only 16% after 48 h. The biomass of lettuce treated with the phoU knockout strain (0.157 mg P/m²) was 38% higher than that of the lettuce treated with the control group. These results indicate that treating lettuce with this microalgal biomass can be beneficial to crop growth. These results suggest that the use of polyphosphate-accumulating microalgae as biofertilizers may alleviate the effects of a diminishing phosphorous supply. These findings can be used as a basis for additional genetic engineering to increase intracellular polyphosphate levels.

• 생명과학회지
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• 제29권8호
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• pp.888-894
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• 2019

류마티스관절염(rheumatoid arthritis)과 골관절염(osteoarthritis) 같은 관절질환은 연골세포(chondrocytes) 감소와 관절연골(articular cartilage)의 분해를 수반한다. 최근, 연골세포의 활성과 연골 항상성(cartilage homeostasis)에 염증성 ROS (reactive oxygen species) burst 및 나이와 관련된 산화적 스트레스(oxidative stress)의 증가와 관련된 연구가 활발히 진행되고 있다. 본 연구는 관절연골로부터 분리한 연골세포의 노화 과정과 나이대별 관절연골에서 항산화 인자들(antioxidants)의 변화를 조사함으로써, 연골세포와 관절연골의 노화 과정 동안 산화적 스트레스로부터 조직을 보호하는 항산화 인자들의 역할을 규명하는데 목적이 있다. 쥐의 관절연골로부터 분리한 연골세포의 연속 계대배양을 통한 노화 과정에서 산화적 스트레스가 증가함을 관찰하였다. 그리고, 노화 유도한 연골세포는 세포내 총 glutathione (GSSG/GSH) 양과 항산화 효소 superoxide dismutase (SOD)와 heme oxygenase-1 (HO-1)의 발현이 증가하였다. 다음으로, 나이대별 쥐로부터 분리한 관절연골에서 항산화 인자의 발현을 분석하였다. 항산화 인자 glutathione의 양은 40주령에서 발현이 가장 높게 관찰되었으며 72주령에 다소 감소하였고, SOD와 HO-1의 발현은 나이대별로 현저히 증가되는 경향을 보였다. 이를 종합해 볼 때, 세포내 항산화 인자들은 과도한 양의 ROS에 반응하여 연골세포의 노화와 나이와 관련된 관절연골의 퇴화 과정에서 중요한 역할을 하는 것으로 보인다.

• Archives of Pharmacal Research
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• 제27권4호
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• pp.361-370
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• 2004

The discovery of insulin receptor substrate (IRS) proteins and their role to link cell surface receptors to the intracellular signaling cascades is a key step to understanding insulin and insulin-like growth factor (IGF) action. Moreover, IRS-proteins coordinate signals from the insulin and IGF receptor tyrosine kinases with those generated by proinflammatory cytokines and nutrients. The IRS2-branch of the insulin/IGF signaling cascade has an important role in both peripheral insulin response and pancreatic $\beta$-cell growth and function. Dysregulation of IRS2 signaling in mice causes the failure of compensatory hyperinsulinemia during peripheral insulin resistance. IRS protein signaling is down regulated by serine phosphorylation or protea-some-mediated degradation, which might be an important mechanism of insulin resistance during acute injury and infection, or chronic stress associated with aging or obesity. Under-standing the regulation and signaling by IRS1 and IRS2 in cell growth, metabolism and survival will reveal new strategies to prevent or cure diabetes and other metabolic diseases.

• The Korean Journal of Physiology and Pharmacology
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• 제24권1호
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• pp.1-10
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• 2020

Autophagy is a highly conserved intracellular degradation and energy-recycling mechanism that contributes to the maintenance of cellular homeostasis. Extensive researches over the past decades have defined the role of autophagy innate immune cells. In this review, we describe the current state of knowledge regarding the role of autophagy in neutrophil biology and a picture of molecular mechanism underlying autophagy in neutrophils. Neutrophils are professional phagocytes that comprise the first line of defense against pathogen. Autophagy machineries are highly conserved in neutrophils. Autophagy is not only involved in generalized function of neutrophils such as differentiation in bone marrow but also plays crucial role effector functions of neutrophils such as granule formation, degranulation, neutrophil extracellular traps release, cytokine production, bactericidal activity and controlling inflammation. This review outlines the current understanding of autophagy in neutrophils and provides insight towards identification of novel therapeutics targeting autophagy in neutrophils.

• Toxicological Research
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• 제17권
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• pp.89-96
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• 2001

Recently, considerable attention has been focused on the role of cyclooxygenase-2 (COX-2) in the carcinogenesis as well as in inflammation. Improperly overexpressed COX-2 has been observed in many types of human cancers and transformed cells in culture. Thus, it is conceivable that targeted inhibition of abnormally or improperly up-regulated COX-2 provides one of the most effective and promising strategies for cancer prevention. A ubiquitous eukaryotic transcription factor, NF-kB is considered to be involved in regulation of COX-2 expression. Furthermore, extracellular-regulated protein kinase and p38 mitogen-activated protein (MAP) kinase appear to be key elements of the intracellular signaling cascades involved in NF-kB activation in response to a wide array of external stimuli. Certain chemopreventive phytochemicals suppress activation of NF-kB by blocking one or more of the MAP kinases, which may contribute to their inhibitory effects on COX-2 induction. One of the plausible mechanisms by which chemopreventive phytochemicals inhibit NF-kB activation involves suppression of degradation of the inhibitory unit I kB, which hampers subsequent translocation of p65, the functionally active subunit of NF-kB.