Earticle

Adult stem cells, which have characteristic of self-renewal and multipotency, are specialized cell types, responsible for the tissue regeneration of the damaged tissue. Recent studies suggest that stem cells senescence (or stem cells’ ageing) is closely associated with the variety of ageing-related phenotypes such as tissue atrophy, degenerative diseases and onset of cancers. During ageing, declining of stem cells function and subsequently occurring mal-differentiation of stem cells would be important to understand the biological process of development of ageing-related phenotypes such as tissue degenerations and cancers. This review focuses on the DNA damage stress as a cause of senescence of stem cells and their maldifferentiation, which is closely link to defect of regeneration potentials and neoplastic transformation. Understanding of molecular mechanisms governingsuch events is likely to have important implications for developing novel avenues for balancing tissue homeostasis longer period of time, further leading to ‘Healthy ageing’.

Much interest has been recently focused on the production of large quantities of hydrogen, due to its potential importance in our economy and needs in the petroleum and chemical industries. Formate dehydrogenase H (FDH-H) from Escherichia coli containing selenocysteine that oxidizes formate to carbon dioxide with the release of hydrogen is a component of the anaerobic formate hydrogen-lyase complex of E. coli. To make full use of FDH-H, we need effective expression condition. In this approach, we investigated the effect of pH on FDH-H stability and observed the effect of selenite and formate concentration on the activity of FDH-H. Additionally, coexpression of selenocysteine insertion genes were tried to improve the expression of FDH-H. The highest level of FDH-H expression was achieved by coexpression of selenocysteine insertion genes (pSUABC) as well as by the addition of 10 μM selenite and 10 mM formate. At this optimized condition, a 2.6 fold elevation of expression of FDH-H was achieved.

Polyester was prepared through the esterification reaction between watsepaper liquefied by ethylene glycol and carboxylic acid. Liquefaction was carried out at the previously determined condition of 100 minutes, 160℃, and 3% sulfuric acid, and the hydroxyl value of the liquefied product was 411 mg KOH/g. In order to remove bubbles produced during the curing step, the method to introduce a slight nitrogen stream into reaction vessel and/or the method to preheat a polyester film at 85℃ before curing step were used alone or in combination. But if curing temperature was 130℃, simple method to cure a film for 5 hours at 130℃ without using both methods was found to be most effective. The polyesters prepared with various carboxylic acids showed significant different physical properties, and maleic acid was best among them. Also, the effect of reaction time and temperature, C/H (carboxyl group/hydroxyl group) ratio, and type of additive on the crosslinkage of polyester was investigated. Lithium hydroxide or citric acid as additive was used to enhance the crosslinkage of polyester and citric acid was proved to be much more effective than lithium hydroxide. The effect of reaction temperature on the crosslinkage was marginal, but the crosslinkage decreased above 130℃. The crosslinkage was 86% when the polyester was prepared at an optimum condition such as 130℃ and 15 minutes of reaction condition, 1.5 of C/H ratio, 130℃ and 5 hours of curing condition, and 10% addition of citric acid.

Using tetrameric β-galactosidase as a model protein, anchoring motives were screened in Bacillus subtilis spore display system. Eleven spore coat proteins were selected considering their expression levels and the location in the spore coat layer. After chromosomal single-copy homologous integration in the amyE site of Bacillus subtilis chromosome, cotE and cotG were chosen as possible spore surface anchoring motives with their higher whole cell β-galactosidase activity. PAGE and Wester blot of extracted fraction of outer layer of purified spore, which express CotE-LacZ or CotG-LacZ fusion verified the existence of exact size of fusion protein and its location in outer coat layer of purified spore. β-galactosidase activity of spore with CotE-LacZ or CotG-LacZ fusion reached its highest value around 16~20 h of culture time in terms of whole cell and purified spore. After intensive spore purification with lysozyme treatment and renografin treatment, spore of BJH135, which expresses CotE-LacZ, retained only 1~2% of its whole cell β- galactosidase activity. Whereas spore of BJH136, which has cotG-lacZ cassette in the chromosome, retained 10~15% of its whole cell β-galactosidase activity, proving minor perturbation of CotG-LacZ, when incorporated in the spore coat layer of Bacillus subtilis compared to CotE-LacZ. Usage of Bacillus subtilis WB700, of which 7 proteases are knocked-out and thereby resulting in 99.7 % decrease in protease activity of the host, did not prevent the proteolytic degradation of spore surface expressed CotG-LacZ fusion protein.

The growth effect of Nannochloropsis oculata (N. oculata), unicellular microalgae, by alginate was investigated. Alginate was depolymerized with sulfuric acid (H2SO4) and heat (121℃), simultaneously. Addition of 0.75% alginate oligomer depolymerized with 0.2 N H2SO4 showed the maximum yield and the growth rate of N. oculata. Chlorophyll content and reducing sugar was increased by alginate oligomer in a dose-dependent manner. Alginate oligomer promoted the growth of N. oculata, whereas the original alginate polysaccharides had no significant effect. Laminaria japonica (L. japonica) extract containing high level of alginate was also increased growth rate and chlorophyll content. CO2 supply addition to L. japonica extract showed no change the growth rate, although addition to alginate oligomer showed prominently increased. N. oculata could use more saccharides in presence of CO2 according to reducing sugar determination. From these results, it is useful to establish optimal condition for high cell density cultivation of N. oculata.

To study the effects of ethanol extract of the Rhodiola Sachalinensis on the anti-oxidation and blood pressure and skeletal muscles contractility in rats. The ethanol extracts and fractions of Rhodiola Sachalinensis were determinated the anti-oxidation effects by DPPH (1,1-dipenyl- 2-picrylhydrazyl) method and comparing with the BHA (Butylated hydroxyanisole) and AA (Ascorbic acid). The gastrocnemius and soleus muscle contractility were observed by stimulating the sciatic nerve with electricity after affusing stomach with ethanol extract of the 200 mg/kg dosage of Rhodiola Sachalinensis for 4 weeks. The ethanol extract of the Rhodiola Sachalinensis could shorten latent period, increase maximal contractive extent and time and relaxative time at the twitch and complete tetanus of gastrocnemius and soleus muscles. The ethanol extract of the Rhodiola Sachalinensis shows high anti-oxidation effect and can enhance skeletal muscles contractility in rats.

Itraconazole is a triazole antifungal agent to inhibit most fungal pathogens. To improve the oral absorption and dissolution of poorly water-soluble itraconazole, microsponge system composed of Eudragit® E100 and polyvinyl alcohol (PVA) formulated by quasi-emulsion solvent diffusion method, and its physicochemical properties and pharmacokinetic parameters of itraconazole were studied. The microsponge of itraconazole were discrete free flowing microsized particles with perforated orange peel like morphology as visualized by scanning electron microscope (SEM). Results showed that the drug loading efficiency, production yield, and particle size of itraconazole microsponge were affected by drug to polymer ratio, the volume of internal phase containing methylene chloride, stirring rate and the concentration of PVA used. Also, the results showed that the dissolution rate of itraconazole from the microsponges was affected by drug to polymer ratio. In other words, the release rate of itraconazole from micropsonges was increased from at least 27.43% to 64.72% after 2 h. The kinetics of dissolution mechanism showed that the dissolution data followed Korsmeyer-Peppas model. Therefore, these results suggest that microsponge system can be useful for the oral delivery of itraconazole by manipulating the release profile.

The effects of pH, glycerol concentration and salt on cell growth and ethanol production using Enterobacter aerogenes KCTC 2190 were evaluated in the anaerobic culture condition. In condition of initial pH 5, cell growth and ethanol production were highest. An initial concentration of 10 g/L of pure glycerol gave the highest cell growth and ethanol production. However, in case of over 15 g/L of pure glycerol, they decreased. The cell growth and ethanol production decreased with the increase of salt concentration. When 10 g/L of crude glycerol was used as the carbon source, the cell growth and ethanol production were 1.32 OD600 and 3.95 g/L, respectively, which were about 94.4% and 88.5% compared to those of pure glycerol. These result indicates that the crude glycerol produced in the biodiesel manufacturing process may be useful as a potential carbon source for ethanol production form Enterobacter aerogenes KCTC 2190.

An efficient screening method was developed for rapid selection of a few overproducers of itaconic acid (IA) among the great many mutants derived from mother strains of Aspergillus terreus. For this purpose, an attempt was made to reveal the relationships of the growth rate and sporulation of each mutant on PDA solid medium with its IA productivity in the final liquid production-culture. As a result, it was possible to classify the mutated strains into 5 groups (from [A] to [E] group) according to theirmorphologies (i.e., growth rate and sporulation extent) on the PDA slants. Notably, most of the high-yielding mutants of IA were observed to belong to [A]group which had the properties of the highest growth rate and sporulation among the 5 groups, whereas the mutant groups of [C], [D] and [E] with the contrasting morphological features showed significant reductions in their IA productivities. From these results, it was concluded that the probability of selecting IA overproducing mutants could be remarkably enhanced when the mutated colonies showing faster growth rates are firstly selected on the PDA plate, and then further screening process is performed on the basis of the sporulation extents of the mutants selected. Consequently, through the application of the strategy developed in this study, costs and time involvedin the labor-intensive task of strain improvement could be reduced to a great extent, because the time-consuming liquid culture processes did not need to performed for the unfavorable mutants belonging to the groups other than group [A].

As a number of archaea are ubiquitously found in non-extreme habitats, elucidation of their functional roles becomes currently an emerging issue. However, most of them are unable to grow in pure culture and so it remains to be established. In order to find genes lacking in the genome of an ammonia-oxidizing archaeon (AOA), we here report on the comparative analyses of an AOA genome with those of experimentally or theoretically established minimal genomes for independent growth. We assessed the genes lacking in AOA using logic of clusters of orthologous groups (COG), remote homology, consensus sequence weight matrix, function-based motif or domain, and then further excluded genes encoding hypothetical orarchaea-specific proteins. The results of these combination analyses revealed 19 candidate genes lacking in the genome of an AOA. Thus, our results provide a possibility of inducing independent growth of AOA when supplemented with product (s) of the lacking gene (s), and also give a chance for finding new proteins with novel sequence or structure space even if the predicted lacking-genes will be found using another algorithms or biochemical studies.

Using Bacillus subtilis spore display system, with cotG as an anchoring motif, levansucrase from Zymomonas mobilis, was displayed on the outer surface of Bacillus subtilis spore. Flow cytometry of DB104 (pSDJH-cotG-levU) spore, proved the surface localization of CotG-LevU fusion protein on the spore compared to that of DB104. Enzymatic activity of DB104 (pSDJH-cotG-levU) spore showed more than 1.5 times higher levansucrase specific activity compared to that of the host spore, which is a remarkable increase of enzymatic activity considering the existence of sacA (sucrase) and sacB (levansucrase) in the Bacillus subtilis chromosome. The spore integrity, revealed by sporulation frequency test after heat and lysozyme treatment of spore, did not changed at all in spite of the CotG-LevU fusion protein incorporation into the spore coat layer during spore formation process. These data prove again that Bacillus subtilis spore could be considered as good live immobilization vehicle for efficient bioconversion process.

Various biometerials have been researched and have been developed for treatment of some disease through transplantation to body. They have been evaluated by in vitro cytotoxicity test using some skin-derived cell lines for prediction of their biocompatibility in vivo. However, the results of experiments using mesenchymal or epithelial cells could not be considered in vivo immune reaction. In this study, we evaluated the biomaterial-elution (elute from high density polyethylene film) using some cell lines (L929, Jurkat, U937) in vitro, and then that results were compared with in vivo results from guinea pig sensitization test. In sensitization test, saline and elution of syringe could not induce erythema, but only DNCB (hypersensitive chemical) induce erythema at guinea pig sensitization test. In cell experiment, the cytotoxicity results of inflammatory cells (Jurkat; T lymphocyte, U937; monocyte) was no difference with L929 (fibroblast) in the overall trend. However, inflammatory cell lines were only secreted inflammatory cytokine (TNF-α, INF-γ) in some materials (biomateriallution, FAC, DNCB). And the biomaterial-elution did not have toxicity to the cells, but it induced the inflammatory cytokines in inflammatory cell lines only. So, we were predicted inflammatory reaction through the cytokine resultes of inflammatory cell lines, and it was more correlated with in vivo results than cytotoxicity test. Therefore, we suggested that the inflammatory cytokine assay using inflammatory cell lines are more effective method in vitro for evaluation of biocompatibility of biomaterials or chemicals.

This study was to suggest the method for wrinkle improvement using Mugil cephalus extract. In order to evaluate the cosmetic product made of M. cephalus extract, the experiments were conducted with subjects and skin states for 4 weeks. Effect of cosmetics containing M. cephalus extract on the water holding capacity and evaporation of the women skin were investigated. The water holding capacity was increased after 1 h of application. But after 2 h, it was decreased. In the case of the water evaporation, it was increased with the increase of time. On the clinical trial, it was also found that water, oil, and rough level on the women skin were increased and wrinkles were improved. Side effects were also not detected during the application and treatment of cosmetics. Based on this result, the application of M. cephalus extract had positive effects on the improvement of wrinkles and skin in women. Therefore, it will be used to develop the wrinkle improvement therapy for women.

This study aimed to examine the treatment effect of Muraenesox cinereus extract product combined with high frequency on men’s damaged skin. The subjects were treated by a product containing M. cinereus extract combined with high frequency therapy for six weeks and tested in skin condition. First, for moisture level change, there was no significant difference between the control group treated only by high frequency therapy and the experimental group treated by a M. cinereus extract product combined with high frequency therapy. But the subjects who increased consistently in moisture level in the cheek and chin were more common in the experimental group. Second, for an oil level, prominently high increase was found in both the control group and the experimental group. Finally, for a rough level, while there was little difference in the control group between immediately after peeling and after six weeks, high improvement effect on a curve was found in the experimental group. Based on this result, cosmetics using M. cinereus extract may have a positive effect on men’s skin which is exposed to several stress factors. Accordingly, this result will contribute to developing men’s functional cosmetics.