• Title/Summary/Keyword: fluorescent protein

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Dieckol Attenuates Microglia-mediated Neuronal Cell Death via ERK, Akt and NADPH Oxidase-mediated Pathways

  • Cui, Yanji;Park, Jee-Yun;Wu, Jinji;Lee, Ji Hyung;Yang, Yoon-Sil;Kang, Moon-Seok;Jung, Sung-Cherl;Park, Joo Min;Yoo, Eun-Sook;Kim, Seong-Ho;Ahn Jo, Sangmee;Suk, Kyoungho;Eun, Su-Yong
    • The Korean Journal of Physiology and Pharmacology
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    • v.19 no.3
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    • pp.219-228
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    • 2015
  • Excessive microglial activation and subsequent neuroinflammation lead to synaptic loss and dysfunction as well as neuronal cell death, which are involved in the pathogenesis and progression of several neurodegenerative diseases. Thus, the regulation of microglial activation has been evaluated as effective therapeutic strategies. Although dieckol (DEK), one of the phlorotannins isolated from marine brown alga Ecklonia cava, has been previously reported to inhibit microglial activation, the molecular mechanism is still unclear. Therefore, we investigated here molecular mechanism of DEK via extracellular signal-regulated kinase (ERK), Akt and nicotinamide adenine dinuclelotide phosphate (NADPH) oxidase-mediated pathways. In addition, the neuroprotective mechanism of DEK was investigated in microglia-mediated neurotoxicity models such as neuron-microglia co-culture and microglial conditioned media system. Our results demonstrated that treatment of anti-oxidant DEK potently suppressed phosphorylation of ERK in lipopolysaccharide (LPS, $1{\mu}g/ml$)-stimulated BV-2 microglia. In addition, DEK markedly attenuated Akt phosphorylation and increased expression of $gp91^{phox}$, which is the catalytic component of NADPH oxidase complex responsible for microglial reactive oxygen species (ROS) generation. Finally, DEK significantly attenuated neuronal cell death that is induced by treatment of microglial conditioned media containing neurotoxic secretary molecules. These neuroprotective effects of DEK were also confirmed in a neuron-microglia co-culture system using enhanced green fluorescent protein (EGFP)-transfected B35 neuroblastoma cell line. Taken together, these results suggest that DEK suppresses excessive microglial activation and microglia-mediated neuronal cell death via downregulation of ERK, Akt and NADPH oxidase-mediated pathways.

Effect of Methoxy PEG-45 Thioctate (LA-PEG) against Oxidative Protein Damage and Anti-glycation (Methoxy PEG-45 Thioctate (LA-PEG)의 항노화 효과에 대한 연구)

  • Kim, Jin Hwa;Oh, Jung Young;Bae, Jun Tae;Lee, Geun Soo;Pyo, Hyeong Bae
    • Journal of the Society of Cosmetic Scientists of Korea
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    • v.43 no.3
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    • pp.239-245
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    • 2017
  • Aging is a physiological and irreversible, progressive process involving changes in the ability to maintain cellular functionality. It affects tissues, organs and the whole organism and thus finally cause to death. Oxidative stress has been postulated to contribute significantly to the accelerated accumulation of advanced glycation endproducts (AGEs) in collagen, which is implicated in the process of skin aging. In the present study, glycation inhibitory activity of methoxy PEG-45 thioctate (LA-PEG), and its inhibitory effect of cellular oxidation and senescence was investigated. Treatment of LA-PEG significantly showed lower fluorescent intensity induced by AGEs. In addition, LA-PEG was significantly reduced the formation of ROS induced by AGEs. High antioxidant and anti-glycation activities of LA-PEG in glycated collagen model indicated its contribution to anti-aging process. Cellular senescence leads to an increase in senescence-associated ${\beta}$-galactosidase ($SA-{\beta}-gal$) activity, which can be used as a biomarker to identify senescent cells. Treatment with LA-PEG showed a dose-dependent, statistically significant decreased in $SA-{\beta}-gal$ indicating reduced senescence. These results suggest that LA-PEG may have potent anti-aging effects and can be used as new functional materials against cellular accumulation of AGEs.

Identification and analysis of microRNAs in Candida albicans (Candida albicans의 마이크로RNA 동정과 분석)

  • Cho, Jin-Hyun;Lee, Heon-Jin
    • Journal of Life Science
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    • v.27 no.12
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    • pp.1494-1499
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    • 2017
  • Oral infection due to Candida albicans is a widely recognized and frequent cause of superficial infections of the oral mucosa (oral candidiasis). Although oral candidiasis is not a life-threatening fungemia, it can cause severe problems in individuals under certain conditions. MicroRNAs (miRNAs) are noncoding, small RNA molecules, which regulate the expression of other genes by inhibiting the translation of target mRNAs. The present study was designed to identify miRNAs in C. albicans and determine their possible roles in this organism. miRNA-sized small RNAs (msRNAs) were cloned in C. albicans by deep sequencing, and their secondary structures were analyzed. All the cloned msRNAs satisfied conditions required to qualify them as miRNAs. Bioinformatics analysis revealed that two of the most highly expressed C. albicans msRNAs, Ca-363 and Ca-2019, were located in the 3' untranslated region of the corticosteroid-binding protein 1 (CBP1) gene in a reverse orientation. miRNA mimics were transformed into C. albicans to investigate their RNA-inhibitory functions. RNA oligonucleotide-transformed C. albicans was then observed by fluorescent microscopy. Quantitative PCR analysis showed that these msRNAs did not inhibit CBP1 gene expression 4 hr and 8 hr after ectopic miRNA transformation. These results suggest that msRNAs in C. albicans possess an miRNA-triggered RNA interference gene-silencing function, which is distinct from that exhibited by other eukaryotic systems.

Spectroscopic Comparison of Photo-oxidation of Outside and Inside of Hair by UVB Irradiation (자외선B 조사에 의한 모발 외부와 내부의 광산화에 관한 분광학적 비교)

  • Ha, Byung-Jo
    • Applied Chemistry for Engineering
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    • v.31 no.2
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    • pp.220-225
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    • 2020
  • Hair is made of proteins containing various amino acids. Ultraviolet (UV) radiation is believed to be responsible for the most damaging effects of sunlight, and also plays an important role in hair aging. The purpose of this study was to investigate the changes in morphological and chemical structures after ultraviolet B (UVB) irradiation of human hair. The UVB-irradiated hair showed characteristic morphological and structural changes, compared to those of the normal hair. The result from a scanning electron microscope (SEM) equipped with an energy dispersive X-ray diffractometer (EDX) showed that the scale of UV-irradiated hair appeared to be rough and the amount of oxygen element was higher than that of the normal hair. Fluorescence and three dimensional (3D) topographical images were obtained by a confocal laser scanning microscope (CLSM). In 3D images, the green emission intensity of normal hair was much higher than that of fluorescing UVB-irradiated hair. The intensity of green emission reflects the intrinsic fluorescence of hair protein. Also, a fluorescent imaging method using fluorescamine reagent was used to identify the free amino groups resulting from a peptide bond breakage in UVB-irradiated hair. Strong blue fluorescence of UVB-irradiated hair, which indicates a very high level of amino groups, was observed by CLSM. Therefore, the fluorescamine as an extrinsic fluorescence could provide a useful tool to identify the peptide bond breakage in UVB-irradiated hair. Infrared image mapping was also employed to assess the cross-sections of normal and UVB-irradiated specimens to examine the oxidation of disulfide bonds. The degree of peak areas with strong absorbance for the disulfide mono-oxide was spread from the outside to the inside of hair. The spectroscopic techniques used alone, or in combination, launch new possibilities in the field of hair cosmetics.

Gender-independent efficacy of mesenchymal stem cell therapy in sex hormone-deficient bone loss via immunosuppression and resident stem cell recovery

  • Sui, Bing-Dong;Chen, Ji;Zhang, Xin-Yi;He, Tao;Zhao, Pan;Zheng, Chen-Xi;Li, Meng;Hu, Cheng-Hu;Jin, Yan
    • Experimental and Molecular Medicine
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    • v.50 no.12
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    • pp.12.1-12.14
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    • 2018
  • Osteoporosis develops with high prevalence in both postmenopausal women and hypogonadal men. Osteoporosis results in significant morbidity, but no cure has been established. Mesenchymal stem cells (MSCs) critically contribute to bone homeostasis and possess potent immunomodulatory/anti-inflammatory capability. Here, we investigated the therapeutic efficacy of using an infusion of MSCs to treat sex hormone-deficient bone loss and its underlying mechanisms. In particular, we compared the impacts of MSC cytotherapy in the two genders with the aim of examining potential gender differences. Using the gonadectomy (GNX) model, we confirmed that the osteoporotic phenotypes were substantially consistent between female and male mice. Importantly, systemic MSC transplantation (MSCT) not only rescued trabecular bone loss in GNX mice but also restored cortical bone mass and bone quality. Unexpectedly, no differences were detected between the genders. Furthermore, MSCT demonstrated an equal efficiency in rectifying the bone remodeling balance in both genders of GNX animals, as proven by the comparable recovery of bone formation and parallel normalization of bone resorption. Mechanistically, using green fluorescent protein (GFP)-based cell-tracing, we demonstrated rapid engraftment but poor inhabitation of donor MSCs in the GNX recipient bone marrow of each gender. Alternatively, MSCT uniformly reduced the $CD3^+T$-cell population and suppressed the serum levels of inflammatory cytokines in reversing female and male GNX osteoporosis, which was attributed to the ability of the MSC to induce T-cell apoptosis. Immunosuppression in the microenvironment eventually led to functional recovery of endogenous MSCs, which resulted in restored osteogenesis and normalized behavior to modulate osteoclastogenesis. Collectively, these data revealed recipient sexually monomorphic responses to MSC therapy in gonadal steroid deficiency-induced osteoporosis via immunosuppression/anti-inflammation and resident stem cell recovery.

Effects of Water-deficit Stress on Yield and Chlorophyll Fluorescence in Rice during the Early Tillering Stage (분얼기 수분 스트레스가 벼 생육 특성 및 엽록소 형광 반응에 미치는 영향)

  • Han, Chae-Min;Shin, Jong-Hee;Kwon, Jung-Bae;Won, Jong-Gun;Kim, Sang-Kuk
    • KOREAN JOURNAL OF CROP SCIENCE
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    • v.67 no.2
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    • pp.77-84
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    • 2022
  • This study aimed to evaluate the growth, yield, and grain quality characteristics of rice varieties that were subjected to water stress during the tillering stage. We also sought to identify whether water stress could be indicated via the plants' response to chlorophyll fluorescence. In this study, we induced water stress by intentionally cutting off water for 30 days, starting 15 days after transplanting the rice varieties to a greenhouse. We analyzed nine rice varieties, including Ilpum, which is the most frequently cultivated variety in Gyeongsangbuk-do, South Korea. The control group was planted in a paddy field where irrigation was entirely dependent on rainfall. Our results revealed that the heading stage of the nine studied varieties occurred approximately ten days earlier in the rain shelter than in the field. Moreover, the rice yield, head rice rate, and palatability score decreased by 18.6%, 17.1%, and 8.3%, respectively, while protein content increased by 20.2% compared with the control group. The Saenuri and Haimi varieties showed the lowest reduction in yield under the water stress conditions, while the Daebo and Samkwang varieties showed the highest reduction in yield. The chlorophyll fluorescence response after re-irrigation was measured between July 30th and August 17th. The ratio of variable fluorescence to maximum chlorophyll fluorescence (FV/FM) values failed to recover to their baseline values, resulting in either no change or a reduction in fluorescent response, even after re-irrigation of Daebo and Samkwang varieties. These results can be utilized as empirical data for drought-affected farms to select resistant varieties that can respond to spring drought in the southern plains of Gyeongsangbuk-do.

Suppression of PMA-induced Differentiation via Foam Cell Formation in THP-1 Cells by 7-Ketocholesterol (THP-1 세포에서 거품세포 형성과 단핵구 분화 및 활성화에서 7-ketocholesterol의 역할)

  • Lee, Mi Sun;Park, Si Eun;Kim, Koanhoi;Park, Young Chul
    • Journal of Life Science
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    • v.32 no.2
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    • pp.142-147
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    • 2022
  • Oxysterols are known to be involved in the physiopathology of atherosclerosis. Since 7-ketocholesterol (7-KC) is found in large amounts in oxysterols and in atherosclerotic plaque, the study on how 7-KC may affect monocyte differentiation induced by phorbol myristate acetate (PMA) in the monocytic cell line, THP-1, is essential. 7-KC induced a dose-dependent reduction in cell proliferation without inducing cytotoxicity, and the substantial staining of Nile red demonstrates the increased absorption of intracellular lipids. Although 7-KC itself did not increase cell adhesion, it markedly decreased the adhesion of cells treated with PMA. Furthermore, by observing the effect of 7-KC on phagocytosis using fluorescent-labeled latex beads, 7-KC's ability to abolish phagocytosis in PMA-stimulated macrophages was illustrated. The effect of 7-KC on the expression of selected protein markers on the process of differentiation induced by PMA in THP-1 cells was also examined. 7-KC inhibited expression of ICAM-1, CD11a, SR-A1, and SR-B2 (CD36) in PMA-stimulated THP-1 cells. Conversely, 7-KC drastically increased the expression of SR-D1 (CD68)in PMA-stimulated THP-1 cells. In conclusion, these results suggest that 7-KC modulates monocyte differentiation and activation via the intracellular accumulation of lipid droplets.

Neuro-Restorative Effect of Nimodipine and Calcitriol in 1-Methyl 4-Phenyl 1,2,3,6 Tetrahydropyridine-Induced Zebrafish Parkinson's Disease Model

  • Myung Ji Kim; Su Hee Cho; Yongbo Seo; Sang-Dae Kim; Hae-Chul Park; Bum-Joon Kim
    • Journal of Korean Neurosurgical Society
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    • v.67 no.5
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    • pp.510-520
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    • 2024
  • Objective : Parkinson's disease (PD) is one of the most prevalent neurodegenerative diseases, characterized by the loss of dopaminergic neurons in the substantia nigra pars compacta. The treatment of PD aims to alleviate motor symptoms by replacing the reduced endogenous dopamine. Currently, there are no disease-modifying agents for the treatment of PD. Zebrafish (Danio rerio) have emerged as an effective tool for new drug discovery and screening in the age of translational research. The neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) is known to cause a similar loss of dopaminergic neurons in the human midbrain, with corresponding Parkinsonian symptoms. L-type calcium channels (LTCCs) have been implicated in the generation of mitochondrial oxidative stress, which underlies the pathogenesis of PD. Therefore, we investigated the neuro-restorative effect of LTCC inhibition in an MPTP-induced zebrafish PD model and suggested a possible drug candidate that might modify the progression of PD. Methods : All experiments were conducted using a line of transgenic zebrafish, Tg(dat:EGFP), in which green fluorescent protein (GFP) is expressed in dopaminergic neurons. The experimental groups were exposed to 500 μmol MPTP from 1 to 3 days post fertilization (dpf). The drug candidates : levodopa 1 mmol, nifedipine 10 μmol, nimodipine 3.5 μmol, diethylstilbestrol 0.3 μmol, luteolin 100 μmol, and calcitriol 0.25 μmol were exposed from 3 to 5 dpf. Locomotor activity was assessed by automated tracking and dopaminergic neurons were visualized in vivo by confocal microscopy. Results : Levodopa, nimodipine, diethylstilbestrol, and calcitriol had significant positive effects on the restoration of motor behavior, which was damaged by MPTP. Nimodipine and calcitriol have significant positive effects on the restoration of dopaminergic neurons, which were reduced by MPTP. Through locomotor analysis and dopaminergic neuron quantification, we identified the neuro-restorative effects of nimodipine and calcitriol in zebrafish MPTP-induced PD model. Conclusion : The present study identified the neuro-restorative effects of nimodipine and calcitriol in an MPTP-induced zebrafish model of PD. They restored dopaminergic neurons which were damaged due to the effects of MPTP and normalized the locomotor activity. LTCCs have potential pathological roles in neurodevelopmental and neurodegenerative disorders. Zebrafish are highly amenable to high-throughput drug screening and might, therefore, be a useful tool to work towards the identification of disease-modifying treatment for PD. Further studies including zebrafish genetic models to elucidate the mechanism of action of the disease-modifying candidate by investigating Ca2+ influx and mitochondrial function in dopaminergic neurons, are needed to reveal the pathogenesis of PD and develop disease-modifying treatments for PD.

Nutritional Studies for Improvement of Feeding on Korean Native Goat - Absorption of Nutrients in Rumen - (한국재래산양(韓國在來山羊)의 사양개선(飼養改善)에 관(關)한 연구(硏究) - 제일위((第一胃)에서의 영양소(營養素) 흡수(吸收)에 대(對)하여 -)

  • Kwon, Soon Ki
    • Korean Journal of Agricultural Science
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    • v.9 no.1
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    • pp.284-302
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    • 1982
  • Development of protein resources as food has been a big issue especially in Southeast Asia region, and intake of protein is also insufficient in Korea. To cope with this shortage of protein resources and its improvement together with increased production of high nutritive animal protein, studies were carried out on feeding of Korean native goats. In the experiments were made absorption of carbohydrate and volatile fatty acid in miniature rumen, and absorption of amino acid in rumen as in vivo were conducted as part of studies on nutritional absorption in rumen. Those nutritional for improvement of feeding and management as described above are summarized as following. 1. According to the result of test on the nutritional absorption of native goat by means of miniature rumen method, absorption ratio of VFA measured at 0.5, 1 and 2 hours after injection of nutrition showed propionic acid 70-86%, acetic acid 74-87%, and lactic acid 76-89%. In the absorption of organic substances, ethyl alcohol of 0.5% showed 29-87% and lactic acid of 0.1M showed 12-27% of absorption ratio. 2. Result of absorption measurement in rumen from L-type free amino acid injection in the content of rumen vein showed lower rate at menthionine-free group compared to whole-egg amino acid injection in the content of rumen vein showed lower rate at methioine-free group compared to whole-egg amino acid group, and high absorption ratio was observed at methionine 3 times group and urea added group. Deficiency of methionine caused no change of the content in mucous membranes. 3. Absorption of amino acid in rumen muscular layer showed equal tendency as in the mucous membrane without exerting any influence of methionine deficiency. At the methionine3-times group, content of methionine and glutamine were increased by 14.7 and 4.4 times as compared to whole-egg amino acid group, an absorption ratio of glutamine, proline and valine were increased at urea added group. 4. In general, concentration of amino acid in rumen vein plasma was lower than in rumen mucous membrane and muscular layer. Absorption ratio of amino acid is decreased due to methionine deficiency, and tripling of methionine or urea adding caused increment of amino acid. Absorption pattern is thus varied depending on the composition of amino acid. 5. At the urea added group, content of ammonia-N, amino-N and urea were increased in rumen muscular layer. As the inside of goat's rumen was unable to clean thoroughly, investigation was made on remaining bacteria, however, variation of ammonia-N was affected by these bacterial content. 6. Variation in rumen structure by differential absorption of amino acid was observed by general microscope and fluorescent microscope. According to the result of observation in the methionine 3 times group, single cylinder epithelium of mucous membrane showed rather thin, and it was thick at urea added group though no significant differences existed among test groups in submucous membrane and muscular layer.

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Elimination of Lily Symptomless Virus by In Vitro Scaling and Reinfection Rates under Various Culture Conditions in Korean Native Lilies (한국 자생나리의 기내 인편삽에 의한 Lily Symptomless Virus 제거 및 구근 재배조건에 따른 재감염 분석)

  • Kim, Min Hui;Park, In Sook;Park, Kyeung Il;Oh, Wook;Kim, Kiu Weon
    • Horticultural Science & Technology
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    • v.33 no.6
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    • pp.891-899
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    • 2015
  • The lily symptomless virus (LSV) is the most common virus in Korean native lilies and causes various types of damage to overall plant growth. This study was carried out to investigate the elimination rate of the LSV by the in vitro scale culture (scaling) method in Korean native lilies and to test reinfection rates of the LSV under several field culture conditions of bulb production. Four Korean native lilies (Lilium dauricum, L. distichum, L. lancifolium, and L. maximowitzii) were used and their scales were cultured in vitro for micro-scale formation. The micro-scales were subcultured repeatedly using MS culture medium supplemented with 30 or $90g{\cdot}L^{-1}$ sucrose. The culture conditions were $24{\mu}mol{\cdot}m^{-2}{\cdot}s^{-1}$ PPFD with 16 hour daylength using fluorescent lamps and maintained at $22{\pm}2^{\circ}C$. The virus-free bulblets were grown for one to three years in the greenhouse and transplanted to the field in October or March. Virus infection rates were investigated by direct tissue blotting immunobinding assays and measurement of chlorophyll and protein contents. Virus-free plants could be obtained from the 5th subculture of micro-scales in L. lancifolium and L. maximowitzii or from primary culture in L. dauricum and L. distichum. LSV-free plants were reinfected during bulb production in the field. Reinfection rates were higher at older bulb ages and under higher planting density. The plants planted in October and at inland Gyeongsan had higher infection rates than those planted in March and at coastal area Pohang. The reinfection rate of L. maximowitzii was higher than those of L. dauricum and L. lancifolium. The LSV-infected plants had lower chlorophyll contents and unchanged protein contents compared to virus-free plants.