Our previous study demonstrated that Coprisin, a peptide from Copris tripartitus infected with bacterial pathogens, has an antibacterial activity. We assessed in this study whether Coprisin caused cellular toxicity in various mammalian cell lines. Coprisin selectively caused a marked drop of cell viability in Jurkat T cells, U937 cells and AML-2 cells belonging to the human leukemia cells but not in Caki cells and Hela cells. Fragmentation of DNA, a maker of apoptosis, was also confirmed in theleukemia cell lines but not in other cells. The Coprisin-induced apoptosis in leukemia cells was mediated by AIF (apoptosis inducing factor), a caspase -independent pathway.

Human mesenchymal stem cell (hMSCs) isolated from human adult bone marrow have self-renewal capacity and can differentiate into multiple cell types in vitro and in vivo. A number of studies have now demonstrated that MSCs can differentiate into various neuronal populations. Due to their autologous characteristics, replacement therapy using MSCs is considered to be safe and does not involve immunological complications. The basic helix-loop-helix (bHLH) transcription factor Olig2 is necessary for the specification of both oligodendrocytes and motor neurons during vertebrate embryogenesis. To develop an efficient method for inducing neuronal differentiation from MSCs, we attempted to optimize the culture conditions and combination with Olig2 gene overexpression. We observed neuron-like morphological changes in the hMSCs under these induction conditions and examined neuronal marker expression in these cells by RTPCR and immunocytochemistry. Our data demonstrate that the combination of Olig2 overexpression and neuron-specific conditioned medium facilitates the neuronal differentiation of hMSCs in vitro. These results will advance the development of an efficient stem cell-mediated cell therapy for human neurodegenerative diseases.

COPRISIN is an antibiotic substance extracted from Copris tripartitus. This study is intended to identify various cell biological stimuli that COPRISIN, widely known as an antibacterial substance, has on human cells and to identify its molecule mechanism. A variety of human cell lines were divided into epithelial cells including kidney cells or womb cells, and immunocyte including T cells or macrophages and, after their being cultivated and maintained, cell biological changes of the respective cells according to COPRISIN treatment were compared. As a result, it was confirmed that, different from other experiment cells, COPRISIN specifically caused cell kill in T cells and macrophages. That is, fragmentation of DNA, typical characteristics observed in the process of apoptosis, was confirmed in the nucleus of cells dying owing to COPRISIN treatment. An Apoptosis process is one dependent upon activity of caspase family protein, it was proved that COPRISIN medium cell kill process was one through a caspase-independent route such as AIF. Though it was found out that transcription of TNF-α and extracellular TNF-α secretion increased in blood cells stimulated by COPRISIN, it was also confirmed that TNF-α was a major medium factor in a COPRISIN induced cell kill process from the fact that a cell kill process by COPRISIN was not inhibited at all with TNF-α inhibiting antibody treatment. Above results revealed that COPRISIN, different from other tissue origin cells including kidney cells, can specifically induce apoptosis in immunocyte, which is caused by a caspase-independent cell signal transmission route.

The effects of additions/deletions in glycosylated residues of recombinant human EPO (rhEPO) produced in CHO-K1 on their secretion were examined. hEPO cDNA was amplified from human liver mRNA and cloned into the pCR2.1 TOPO. Using overlapping-extension site-directed mutagenesis method, glycosylation sites at 24th, 38th, 83rd, and 126th were respectively or accumulatively removed by substituting its asparagine (or serine) with glutamine. To add novel glycosylation sites, 69 and 105th leucine was mutated to asparagine. Mutant and wild type rhEPO constructs were cloned into the pcDNA3 expression vector with CMV promoter and transfected into CHO cell line, CHO-K1, to produce mutant rhEPO mutant rhEPO proteins. Enzyme-linked immunosorbant assay (ELISA) and Western analysis with monoclonal anti-EPO antibody were performed using supernatants of the cultures showing transient and stable expressions respectively. Addition of novel glycosylation reduced rhEPO secretion dramatically while deletion mutants had little effect except some double deletion mutants (△24/83 and △38/83) and triple mutant (△24/38/83). This fact suggests that not single but combination of changes in glycosyl groups affect secretion of rhEPO in cell culture, possibly via changes in their conformations.

Gonadotropin-releasing hormone (GnRH) is a key neuropeptide regulating reproduction in humans and other vertebrates. Here, we evaluated the reproductive control system mediated by GnRH in the Pacific abalone Haliotis discus hannai. We cloned a prepro-GnRH cDNA (Hdh-GnRH) from the pleural-pedal ganglion (PPG) in H. discus hannai, and analyzed its spatiotemporal gene expression pattern. The open reading frame of Hdh-GnRH encodes a protein of 101 amino acids, consisting of a signal peptide, a GnRH dodecapeptide, a cleavage site, and a GnRH-associated peptide. This structure and sequence are highly similar to GnRH-like peptides reported for mollusks and other invertebrates. Quantitative polymerase chain reaction demonstrated that Hdh-GnRH mRNA was more strongly expressed in the ganglions (PPG and cerebral ganglion [CG]) than in other tissues (gonads, gills, intestine, hemocytes, muscle, and mantle) in both sexes. In females, the expression levels of Hdh-GnRH mRNA in the PPG and branchial ganglion (BG) were significantly higher at the ripe and partial spent stages than at the early and late active stages. In males, Hdh-GnRH mRNA levels in the BG showed a significant increase in the partial spent stage. Unexpectedly, Hdh-GnRH levels in the CG were not significantly different among the examined stages in both sexes. These results suggest that Hdh-GnRH mRNA expression profiles in the BG and possibly the PPG are tightly correlated with abalone reproductive activities.

Background : 1-Deoxynojirimycin (DNJ) is a representative compound of the antidiabetic constituent in mulberry leaves (Morus alba L.). The hot water extracts of mulberry leaves were fermented with lactic acid bacteria in order to analysis the changes of the DNJ contents and α-glucosidase inhibition. Methods and Results : The mulberry leaves were extracted with hot water (121℃, 3 hr). The extracts were fermented with nine strains of lactic acid bacteria such as Lactobacillus acidophilus, in order to increase the contents of DNJ and α-glucosidase inhibition. DNJ contents in the fermented extracts were determinated by HPLC analysis. The α-glucosidase inhibition was mesured by comparing dose-inhibition curves of α-glucosidase and IC50 value. The DNJ contents after fermentation have increased in the almost fermented extracts. Especially, DNJ of the extracts fermented with L. acidophilus was increased from 8.38 ㎍ ㎖ -1 to 21.77 ㎍ ㎖-1. IC50 values of the α-glucosidase inhibition were shown to be decreased in the fermented extracts. The extracts fermented with L. casei KCTC 3109 was determined at 290.04 ㎍ ㎖-1, resulting it is lower about 140 ㎍ ㎖-1 than 429.76 ㎍ ㎖-1 of the control. Conclusion : From the above results, we may suggest that the lactic acid fermentation of mulberry leaves extracts can more enhance the hypoglycemic activities such as DNJ contents.

Background : The hypoglycemic effects of mulberry leaves extract were evaluated by comparing the abilities on glucose uptake in C2C12 myotubes and 3T3-L1 adipocyte. Glucose uptake of the extracts were identified to be enhanced by bio-conversion using cellulolytic enzyme like Viscozyme. Methods and Results : The mulberry (Morus alba L) leaves were extracted with 30% ethanol or hot water. The hypoglycemic compounds such as Moracin C and, Quercetin and 1-Deoxynojirimycin were identified from the extracts of mulberry leaves. The extracts were fermented using kinds of celluolytic enzymes, which were vicozyme, pectinase, β-glucosidase and xylanase, in order to increase the contents of hypoglycemic constituents in the extracts. The hypoglycemic effects of the fermented extracts were evaluated by comparing the abilities on glucose uptake in C2C12 myotubes and 3T3-L1 adipocyte. The extracts of mulberry leaves fermented with only Viscozyme were identified to increase glucose uptake in C2C12 myotubes and 3T3-L1 adipocyte by supplement of the concentration of 10 μM extracts, compared to insulin as control. However, bio-conversion effects by other enzymes were not shown in the treatments, suggesting hypoglycemic constituents in the extracts of mulberry leaves can be conversed to more active compounds by cellulolytic enzyme treatment like viscozyme. Conclusion : From the above results, we may suggest that the hypoglycemic constituents in mulberry leaves extracts can be conversed to more active compounds by cellulolytic enzymes.

Background : Recently, many studies to seek for medicinal herb compounds from natural products have been attempted capable of being free from harmful side effects and reducing the economic burden. This study was conducted to investigate whether Morus alba L.(MAL) water extracts has a biological safety, and an inhibitory potential against the mutagenicity induced by cigarette smoke condensates(CSC). Methods and Results : MAL was extracted with 70% ethanol and the yield of the extract was 35.1%. The 70% ethanol MAL extract was fractionated sequentially by diethylether, chloroform, dichloromethane, and water, respectively. Crude MAL 70% ethanol extract itself and its solvent fractions did not show any mutagenic effect up to 1 mg/plate toward Salmonella typhimurium TA 98 with or without the microsomal S-9 mixture of liver metabolic activations. On the other hand, the crude MAL extract showed an inhibitory activity against the mutagenicity of CSC with S-9 mixture. Diethyl ether layer among five solvent fractions showed the highest inhibitory activity at the same dose. The inhibitory activity of diethyl ether layer was also increased dose-dependently and the inhibitory rate was about 97.1% at the concentration of 1 mg/plate. Conclusion : In this study, we concluded that all of the crude MAL 70% ethanol extract and its solvent layers are potentially safe for mutagenicity and the diethyl ether layer among the crude MAL 70% ethanol extract has an inhibitory effect against the mutagenicity of CSC.

Background : Dioscorea quinqueloba(DQ) is a medicinal herb that is used as an alternative therapy for cardiovascular disease and various medical conditions. The objective of this study was to characterize the antioxidant activities of DQ. Methods and Results : The samples were extracted with Distilled water and analyzed for total flavonoid contents, polyphenol contents, DPPH radical scavenging activity, and ABTS radical scavenging activity. H9c2 cardiomyoblast cells were subjected to H2O2, to study the protective effect of DQ on cell viability, and ROS production. The total amounts of polyphenols and flavonoids, which indicate the antioxidant capabillity of water extracts from DQ were 27.21mg/g and 22.95mg/g, respectively. The DQ water extract showed highest antioxidant activity by DPPH and ABTS scavenging activities. The DQ water extract was protected cells against H₂O₂-induced cell death without any cytotoxicity, as determined by the MTT assay. The DQ water extract also was inhibiting production of intracellular ROS. Conclusion : These observations suggest that DQ can use potentially good natural antioxidant in daily life for possible health benefits.

Sorghum is the fifth most important cereal in the world as one of the staple food. For the use of natural dye, we have done some researches about sorghum red pigments extracted from stalk and leaves on its physiochemical properties, extracting methods and applications. The researches involved maximum extraction of sorghum pigment and analysis of its processing condition. Total polyphenol and tannin contents were measured by varieties and different plant parts. The stabilities of pigment by irradiation and heat treatment for processing were measured by colorimeter and thermal gravimetric analysis (TGA). In addition, hybrid nano-silica composites with sorghum pigment were made by combining with polyvinyl alcohol, polyvinyl acetate and sodium silicate. Water silica hybrids with sorghum pigment were performed by emulsion treatment. Nano-silica particles were identified and measured their size to be about 200 ~ 400 nm by SEM analysis.

One objective of soybean breeders is to develop cultivars with elevated oleic acid content. Testing soybean seeds for oleic acid content is possible using gas chromatography (GC), however it is time-consuming and requires destructing the seeds. Using single seeds, we developed a near-infrared reflectance spectroscopy (NIR) calibration equation relating oleic acid measured with GC to oleic acid predicted by NIR. The slope of the regression line of oleic acid measured with GC on oleic acid predicted by NIR and the intercept was not different from zero. A set of 300 soybean seeds was used to calculate the NIR equation and an independent set of 100 soybean seeds was used for validation. This NIRS equation showed significant correlation between reference values and NIRS estimated values based on the SEP, r2, and the RSP of reference data to SEP. This research shows that NIR prediction of oleic acid using intact soybean seeds is accurate and rapid and should be especially useful for early generation screening.

“Jecy Gold” is a new kiwifruit variety developed at the National Institute of Subtropical Agriculture (NISA), RDA in 2003, which was for the utilization as fruit with yellow flesh and high soluble solids. This variety was selected from seeds obtained from the cross breeding between Actinidia chinensis cv. “Golden Yellow” and A. chinensis cv. “Songongu” with yellow flesh in 1997. Seedling and line selections were conducted from 1998 to 1999, and characteristic trials were carried out from 2000 to 2002. After developed, it was also conducted to the test of adaptability in kiwifruit orchards of Jeju island from 2003 to 2006. The branch of “Jecy Gold” sprouts strongly and the color of leaf is slightly dark green. The shape of fruit is obovoid, and the color of peel is yellowish-brown without hair. The average weight of fruit is 128.4 g. The core is middle and soft, the percicarp is golden yellow, and the texture is fragile and juicy. Soluble solids concentration is 14.8 oBrix. The total acid content is 0.69% and the fruit yield is higher about 30% than that of A. deliciosa cv. “Hayward”. The content of vitamin C is higher than that of A. deliciosa cv. “Hayward”. The harvesting time is from late October to early November and the fruit can be preserved about 90 days at 2°C. This variety can be planted bellow 100 m sea level and should be grown under shelter to prevent from disease and cold. This variety will be cultivated about 100 ha by 2011 in the southern part of Korea and has been supplied about 10 ha in 2007.