• KSBB Journal
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• v.27 no.3
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• pp.186-194
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• 2012

Various human host cell lines, which are more effective than the other original human cell lines, have been developed and used. Highly efficient human cell line can be obtained from the fusion between human embryonic kidney 293 (HEK293) and human Burkitt's lymphoma cells (Namalwa). Fused cell line has the advantages of both cell lines such as the high transfection efficacy of HEK293 cells and the constitutive expression of Epstein-Barr virus (EBV) genome which is related with high expression of target protein and anti-apoptotic growth of Namalwa cells. In this study, characterization of two original cell lines was performed by using design of experiment (DOE) considering cell maintenance, media development, optimization of culture condition, and scale-up. The formation of aggregates was apparent with high glutamine concentration at more than 6 mM. Supplementation of hydrolysates showed positive effects on the growth performances of HEK293 cells. On the contrary, Namalwa cells showed negative results. It was confirmed that Namalwa cells were more sensitive to lower temperature at $35^{\circ}C$ and hyperosmotic condition over 260 mOsm/kg. In addition, both cell lines showed limited growth in 3-L bioreactor due to shear stress.

• Journal of Microbiology and Biotechnology
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• v.29 no.9
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• pp.1453-1459
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• 2019

Zeaxanthin is an important pigment in the photo-protection mechanism of microalgae. However, zeaxanthin epoxidase, an enzyme involved in the accumulation and conversion of zeaxanthin, has not been extensively studied in microalgae. In this work, we report the expression pattern of zeaxanthin epoxidase in Dunaliella tertiolecta (DtZEP) at different light and diverse salinity conditions. To confirm the responsiveness to light conditions, the ZEP expression pattern was investigated in photoperiodic (16 h of light and 8 h of dark) and continuous (24 h of light and 0 h of dark) light conditions. mRNA expression levels in photoperiodic conditions fluctuated along with the light/dark cycle, whereas those in continuous light remained unchanged. In varying salinity conditions, the highest mRNA and protein levels were detected in cells cultured in 1.5 M NaCl, and ZEP expression levels in cells shifted from 0.6 M NaCl to 1.5 M NaCl increased gradually. These results show that mRNA expression of DtZEP responds rapidly to the light/dark cycle or increased salinity, whereas changes in protein synthesis do not occur within a short period. Taken together, we show that DtZEP gene expression responds rapidly to light irradiation and hyperosmotic stress. In addition, ZEP expression patterns in light or salinity conditions are similar to those of higher plants, even though the habitat of D. tertiolecta is different.

• KSBB Journal
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• v.16 no.4
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• pp.344-350
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• 2001

The effect of salinity stress of Brassica olearacea and Capsicum annuum were studied at various levels of salinity conditions(Na-gluconate, K-gluconate, NaCl, KCl). The effects of salinity stress were measured by seedling growth rates and secondary metabolites contents of the stressed plants. Each seedling studied on the response of different salinity stress. Seedling growth of Capsicum annuum was inhibited up to 200 mM salt tolerance and Brassica olearacea was inhibited up to 400 mM salt tolerance. The produced anthocyanin was separated to high value from 200 mM NaCl in case of Brassica olearana and 50 mM K-gluconate in case of Capsicum annuum. The BADH activity was very high in Brassica olearacea seedlings treated with 200 mM NaCl and in Capsicum annuum seedlings treated with 100 mM K-gluconate. The BADH activities were increased during the early culture days, it induced betaine synthesis. The salinity stress promoted BADH activiy, subsequently endogenous betaine contents were increased, and it seemed to be secure seedling from salinity stress. The salinity concentration of 200 mM was effective on the inhibition of seed germination and on the increase of proline accumulation in tissue. The inhibition of seedling growth and accumulation of secondary metabolites in seedling were caused osmotic hypersensitivity against salinity stress.

• Journal of Microbiology and Biotechnology
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• v.26 no.4
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• pp.700-709
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• 2016

The mitogen-activated protein kinase HOG1 (high-osmolarity glycerol response pathway) plays a crucial role in the response of yeast to hyperosmotic shock. Trichosporonoides oedocephalis produces large amounts of polyols (e.g., erythritol and glycerol) in a culture medium. However, the effects of HOG1 gene knockout and environmental stress on the production of these polyols have not yet been studied. In this study, a To-HOG1 null mutation was constructed in T. oedocephalis using the loxP-Kan-loxP/Cre system as replacement of the targeted genes, and the resultant mutants showed much smaller colonies than the wild-type controls. Interestingly, compared with the wild-type strains, the results of shake-flask culture showed that To-HOG1 null mutation increased erythritol production by 1.44-fold while decreasing glycerol production by 71.23%. In addition, this study investigated the effects of citric acid stress on the T. oedocephalis HOG1 null mutants and the wild-type strain. When the supplementation of citric acid in the fermentation medium was controlled at 0.3% (w/v), the concentration of erythritol produced from the wild-type and To-HOG1 knockout mutant strains improved by 18.21% and 21.65%, respectively.

• Korean Journal of Exercise Nutrition
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• v.23 no.4
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• pp.14-22
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• 2019

[Purpose] Here, we aimed to determine the effect of Polygonum cuspidatum stem extract (PSE) on exorbital lacrimal gland-excised rat models and hyperosmotic stress-stimulated human conjunctival cells (HCCs). [Methods] Seven week old male Wistar rats were divided into six groups. Only the rats in the control group (NOR, n=5) did not undergo surgery. Three days after the surgery, the exorbital lacrimal gland-excised rats were randomly allocated to five groups: (1) vehicle-treated dry-eyed rats (DED, n=5); (2) PSE (10 mg/kg) treated DED rats (PSE-10, n=5); (3) PSE (100 mg/kg) treated DED rats (PSE-100, n=5); and (4) PSE (250 mg/kg) treated DED rats (PSE-250, n=5). In addition, the HCC line was co-treated with hyperosmolar media (528 mOsm) and PSE (1-100 μg/ml). [Results] PSE treatment restored the tear volume and goblet cell density by inhibiting severe corneal irregularities and damage. The treatment with PSE significantly attenuated the hyperosmolar stress-induced inflammation and cell death through the suppression of mRNA expression levels of Tumor necrosis factor-α (TNF-α), Interleukin-6 (IL-6), Interleukin-1β (IL-1β), and Interferon-γ (IFN-γ), and the expression of Bcl-2-associated X protein (Bax) as well as the activation of caspase-3 in vitro. [Conclusion] The inhibitory effects of PSE treatment on dry eye disease indicate the potential of nutritional intervention by PES against inflammatory diseases without adverse effects.

• Journal of Microbiology and Biotechnology
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• v.17 no.2
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• pp.325-334
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• 2007

Recently, quorum sensing has been implicated as an important global regulator controlling the production of numerous virulence factors such as capsular polysaccharides in bacterial pathogens. The nucleotide and deduced amino acid sequences of smcR, a homolog of V. harveyi luxR identified from V. vulnificus ATCC29307, were analyzed. The amino acid sequence of SmcR from V. vulnificus was 72 to 92% similar to those of LuxR homologs from Vibrio spp. Functions of SmcR were assessed by the construction of an isogenic mutant, whose smcR gene was inactivated by allelic exchanges, and by evaluating its phenotype changes in vitro and in mice. The disruption of smcR resulted in a significant alteration in biofilm formation, in type of colony morphology, and in motility. When compared with the wild-type, the smcR mutant exhibited reduced survival under adverse conditions, such as acidic pH and hyperosmotic stress. The smcR mutant exhibited decreased cytotoxic activity toward INT 407 cells in vitro. Furthermore, the intraperitoneal $LD_{50}$ of the smcR mutant was approximately $10^2$ times higher than that of parental wild-type. Therefore, it appears that SmcR is a novel global regulator, controlling numerous genes contributing to the pathogenesis as well as survival of V. vulnificus.

• Journal of Microbiology and Biotechnology
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• v.12 no.2
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• pp.318-326
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• 2002

The pathogenic marine bacterium Vibrio vulnificus is the causative agent of food-borne diseases such as life-threatening septicemia. To better understand this organism's strategies to survive osmotic stress, a mutant that was more sensitive to high osmolarity was screened from a library of mutants constructed by a random transposon mutagenesis. By a transposon-tagging method, putAP genes encoding a proline dehydrogenase and a proline permease were identified and cloned from V. vulnificus. The amino acid sequences deduced from nucleotide sequences of putAP from V. vulnificus were 38 to $59\%$ similar to those of PutA and PutP reported from other Enterobacteriaceae. Functions of putAP genes were assessed by the construction of mutants, whose putAP genes were inactivated by allelic exchanges. When proline as the sole carbon or nitrogen source was used, the putA mutant was not able to grow to the substantial level, revealing the proline dehydrogenase is the only enzyme for metabolic conversion of proline into other amino acids. Although the growth rate of the putP mutant on proline as the sole carbon or nitrogen source was significantly reduced, the mutant still grew. This indicated that at least one more proline permease is produced by V. vulnificus. The putP mutant decreased approximately $2-log_10$ CFU/ml after a hyperosmotic challenge, while the parent strain decreased approximately $l-log_10$ CFU/ml. This result suggests that the gene product of putP contributes to the osmotic tolerance of V. vulnificus.