• Reproductive and Developmental Biology
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• 제28권4호
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• pp.247-252
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• 2004

Human embryonic stem (ES) cells are derived from the inner cell mass of the preimplantation embryo. Human ES cells have the capacity to differentiate into various types of cells in the body. Human ES cells are indefinite source of cells for cell therapy in various degenerative disorders including neuronal disorders. Directed differentiation of human ES cells is a prerequisite for their clinical application. The objective of this study is to develop the culture condition for the derivation of neural precursor cells from human ES cells. Neural precursor cells were derived from human ES cells in a stepwise culture condition. Neural precursor cells in the form of neural rosette structures developed into neurospheres when cultured in suspension. Suspension culture of neurospheres has been maintained over 4 months. Expressions of nestin, soxl, sox2, pax3 and pax6 transcripts were upregulated during differentiation into neural precursor cells by RT-PCR analysis. In contrast, expression of oct4 was dramatically downregulated in neural precursor cells. Immunocytochemical analyses of neural precursor cells demonstrated expression of nestin and SOX1. When induced to differentiate on an adhesive substrate, neuro-spheres were able to differentiate into three lineages of neural systems, including neurons, astrocytes and oligo-dendrocytes. Transcripts of sox1 and pax6 were downregulated during differentiation of neural precursor cells into neurons. In contrast, expression of map2ab was elevated in the differentiated cells, relative to those in neural precursor cells. Neurons derived from neural precursor cells expressed NCAM, Tuj1, MAP2ab, NeuN and NF200 in immunocytochemical analyses. Presence of astrocytes was confirmed by expression of GFAP immuno-cytochemically. Oligodendrocytes were also observed by positive immuno-reactivities against oligodendrocyte marker O1. Results of this study demonstrate that a stepwise culture condition is developed for the derivation of neural precursor cells from human ES cells.

• Journal of Microbiology and Biotechnology
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• 제16권10호
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• pp.1613-1621
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• 2006

Isolation and identification of algal lytic bacteria were carried out. Nine strains of algal lytic bacteria were isolated by the double-layer method using Anabaena flos-aquae as a sole nutrient. The isolate, AFK-13, showing the highest algal lytic activity was identified as Sinorhizobium kostiense based on the l6S rDNA sequence. The algal lytic experiments of the culture supernatants of AFK-13 demonstrated that the bacterial cell growth reached a maximum at 36-h culture, but the supernatant of 72-h culture exhibited the highest activity. Components among the extracellular products in the crude enzyme of the supernatant from S. kostiense AFK-13 culture were responsible for degradation of cell walls of Anabaena flos-aquae. Algal lytic assay tests of the culture supernatants suggest that the main substances for algal lytic activity could be proteinaceous. The activity of glucosidase was observed highly by polysaccharolytic analysis using the crude enzyme from S. kostiense AFK-13, whereas activities of galactosidase, mannosidase, rhamnosidase, and arabinosidase were also detected in low levels. The molecular weights (MW) of ${\alpha}-\;and\;{\beta}$-glucosidases were estimated to be approximately 50-100 kDa by the ultrafiltration method.

• Journal of Microbiology and Biotechnology
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• 제22권5호
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• pp.659-667
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• 2012

2,3-Butanediol (2,3-BDO) is an organic compound with a wide range of industrial applications. Although Escherichia coli is often used for the production of organic compounds, the wild-type E. coli does not contain two essential genes in the 2,3-BDO biosynthesis pathway, and cannot ferment 2,3-BDO. Therefore, a 2,3-BDO biosynthesis mutant strain of Escherichia coli was constructed and cultured. To determine the optimum culture factors for 2,3-BDO production, experiments were conducted under different culture environments ranging from strongly acidic to neutral pH. The extracellular metabolite profiles were obtained using high-performance liquid chromatography (HPLC), and the intracellular metabolite profiles were analyzed by ultra-performance liquid chromatography and quadruple time-of-flight mass spectrometry (UPLC/Q-TOF-MS). Metabolic flux analysis (MFA) was used to integrate these profiles. The metabolite profiles showed that 2,3-BDO production favors an acidic environment (pH 5), whereas cell mass favors a neutral environment. Furthermore, when the pH of the culture fell below 5, both the cell growth and 2,3-BDO production were inhibited.

• Journal of Microbiology and Biotechnology
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• 제34권7호
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• pp.1425-1432
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• 2024

This study analyzed the effects of Ca²⁺ metal ions among culture medium components on the Chlorella sorokiniana strain DSCG150 strain cell growth. The C. sorokiniana strain DSCG150 grew based on a multiple fission cell cycle and growth became stagnant in the absence of metal ions in the medium, particularly Ca²⁺. Flow cytometry and confocal microscopic image analysis results showed that in the absence of Ca²⁺, cell growth became stagnant as the cells accumulated into four autospores and could not transform into daughter cells. Genetic analysis showed that the absence of Ca²⁺ caused upregulation of calmodulin (calA) and cell division control protein 2 (CDC2_1) genes, and downregulation of origin of replication complex subunit 6 (ORC6) and dual specificity protein phosphatase CDC14A (CDC14A) genes. Analysis of gene expression patterns by qRT-PCR showed that the absence of Ca²⁺ did not affect cell cycle progression up to 4n autospore, but it inhibited Chlorella cell fission (liberation of autospores). The addition of Ca²⁺ to cells cultivated in the absence of Ca²⁺ resulted in an increase in n cell population, leading to the resumption of C. sorokiniana growth. These findings suggest that Ca²⁺ plays a crucial role in the fission process in Chlorella.

• 한국생물공학회:학술대회논문집
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• 한국생물공학회 2003년도 생물공학의 동향(XII)
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• pp.398-401
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• 2003

Bacillus species predominantly outgrown in a night soil treatment system were isolated and characterized. Cell interactions took place among them and cell population changed under various culture conditions. Maximum removal of $NH_4\;^+-N$ and cell production occurred under the conditions of 30% DO and C/N ratio of 8. Additions of 0.8% peptone and 0.3% yeast extract to a basal medium influenced the growth of isolates and the removal of $NH_4\;^+-N$ in flask culture, and metal ions such as Ca, Fe and Mg also did. During the flask experiment of nitrogen removal under an aerobic condition, active nitrification by the isolates occurred largely in 1 h with the decrease of COD and alkalinity destructed was only 74.6% of theoretical value. From the nitrogen balance, the percentage of nitrogen lost in the flask culture was estimated to be 29.0%. This conversion of ammonia to $N_2$ under an aerobic condition was confirmed by GC analysis. The B3 process using the Bacillus species seemed to have some economic advantage.

• Asian Pacific Journal of Cancer Prevention
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• 제17권2호
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• pp.581-590
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• 2016

Resistance to anoikis, a cell-detachment induced apoptosis, is one of the malignant phenotypes which support tumor metastasis. Molecular mechanisms underlying the establishment of this phenotype require further investigation. This study aimed at exploring protein expression profiles associated with anoikis resistance of a metastatic breast cancer cell. Cell survival of suspension cultures of non-metastatic MCF-7 and metastatic MDA-MB-231 cells were compared with their adherent cultures. Trypan blue exclusion assays demonstrated a significantly higher percentage of viable cells in MDA-MB-231 than MCF-7 cell cultures, consistent with analysis of annexin V-7-AAD stained cells indicating that MDA-MB-231 possess anti-apoptotic ability 1.7 fold higher than MCF-7 cells. GeLC-MS/MS analysis of protein lysates of MDA-MB-231 and MCF-7 cells grown under both culture conditions identified 925 proteins which are differentially expressed, 54 of which were expressed only in suspended and adherent MDA-MB-231 but not in MCF-7 cells. These proteins have been implicated in various cellular processes, including DNA replication and repair, transcription, translation, protein modification, cytoskeleton, transport and cell signaling. Analysis based on the STITCH database predicted the interaction of phospholipases, PLC and PLD, and 14-3-3 beta/alpha, YWHAB, with the intrinsic and extrinsic apoptotic signaling network, suggesting putative roles in controlling anti-anoikis ability. MDA-MB-231 cells grown in the presence of inhibitors of phospholipase C, U73122, and phospholipase D, FIPI, demonstrated reduced ability to survive in suspension culture, indicating functional roles of PLC and PLD in the process of anti-anoikis. Our study identified intracellular mediators potentially associated with establishment of anoikis resistance of metastatic cells. These proteins require further clarification as prognostic and therapeutic targets for advanced breast cancer.

• International Journal of Advanced Culture Technology
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• 제10권3호
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• pp.339-345
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• 2022

In this paper, the effect of porosity on the acoustic phase velocity of the 3D printed Kelvin closed-cell structure was investigated using the spectral phase analysis. Since Kelvin cells bring about the large amount of scattering, acoustic pulses in ultrasonic measurements undergoes a distortion of waveforms due to the dispersion effect. In order to take account on the dispersion, mathematical expressions for calculating the phase velocity of longitudinal waves propagating normal to the plane of the Kelvin structure are suggested by introducing a complex wave number based on Fourier transform. 3D Kelvin structure composed of identical unit-cells, a polyhedron of 14 faces with 6 quadrilateral and 8 hexagonal faces, was developed and fabricated by 3D CAD and 3D printer to represent the micro-structure of porous materials such as aluminum foam and cancellous bone. Total nine samples of 3D Kelvin structure with different porosity were made by changing the thickness of polyhedron. Ultrasonic pulse of 1MHz center frequency was applied to the Kelvin structures for the measurement of the phase velocity of ultrasound using the TOF(time-of-flight) and the phase spectral method. From the experimental results, it was found that the acoustic phase velocity decreased linearly with the porosity.

• The Korean Journal of Physiology and Pharmacology
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• 제6권5호
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• pp.269-274
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• 2002

A large number of factors such as osteotropic hormones, cytokines, or growth factors are related to the bone remodeling which is characterized by the coupling of osteoclast-mediated bone resorption and osteoblast-mediated bone formation. Recent investigations have indicated that cytokines such as $interleukin-1{\beta}\;(IL-1{\beta})$ and tumor necrosis $factor-{\alpha}\;(TNF-{\alpha})$ play a potential role in the bone resorption associated with a variety of pathological conditions such as inflammatory osteolytic disease. Collagen is the most abundant protein of the extracellular matrix of bone, and the participation of collagenase in bone resorption has been widely investigated. In this study, effects of $IL-1{\beta}$ and $TNF-{\alpha}$ on the release of collagenase from osteoblastic cells were measured. The gelatinase activity was also measured by gel substrate analysis (zymography) after electrophoresis of conditioned media of osteoblastic cell culture. $IL-1{\beta}$ increased the collagenase activity in ROS17/2.8 and HOS cell culture. $TNF-{\alpha}$ also increased the collagenase activity of osteoblastic cells. When two kinds of cytokines were treated simultaneously in the culture of osteoblastic cells, synergistic increase of collagenase activity was seen in ROS17/2.8 cells. $IL-1{\beta}$ and $TNF-{\alpha}$ significantly increased the collagenase activity after 6 hour treatment in the osteoblastic cell culture, and there was no additional increase according to the culture period. Osteoblastic cells released the gelatinase and molecular weight of this enzyme was measured about 70 KDa as assessed by zymogram. $IL-1{\beta}$ and $TNF-{\alpha}$ showed increase of the gelatinase activity produced by ROS17/2.8 and HOS cells. Taken together, this study suggested that $IL-1{\beta}$ and $TNF-{\alpha}$ can modulate bone metabolism, at least in part, by increased release of collagenase and gelatinase from osteoblasts.

• Biotechnology and Bioprocess Engineering:BBE
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• 제7권3호
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• pp.171-177
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• 2002

The metabolic flux pattern for L-histidine production was analyzed when glucose and/or acetate were used as carbon sources. Total L-histidine production was enhanced when mixed substrate (glucose and acetate) was used, compared wish that when either glucose or acetate was used as the sole carbon source. Theoretical maximum carbon fluxes through the main pathways for L-histldine production, cell growth, and ATP consumption for cell maintenance were obtained by the linear programming (LP) method. By comparison of the theoretical maximum carbon fluxes tilth actual ones, it was found that a large amount of glucose was actually used for maintenance of cell viability. On the other hand, acetate was used for cell growth. After depletion of acetate in the mixed substrate culture, the flux for glucose to L-histldine synthesis was markedly enhanced. A strategy for effective L-histidine production using both carbon sources was proposed.

• Journal of Microbiology and Biotechnology
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• 제5권5호
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• pp.264-268
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• 1995

Sulforhodamine B(SRB) assay was performed on the human lung carcinoma, A549 cell line to screen soil microorganisms for production of anti-cancer agent. Among 4, 265 microorganisms, 45 isolates were selected for their cytotoxicity and tested for their effects on the expression of c-myc by RNA slot blot and Northern blot analysis resulting in selection of No.2303 isolate. This No.2303 was identified as Streptomyces sp. by ISP classification and the chemotaxonomic analysis method. NO.2303 inhibited the expression of cmyc in Col0320 DM and A549 cell lines. The culture extract of No. 2303 also inhibited the progression of the cell cycle of Go in NIH 313 cells, implying that the extract also inhibited the expression of c-myc in NIH 313 cell.