• Microbiology and Biotechnology Letters
• /
• v.44 no.4
• /
• pp.452-460
• /
• 2016

This study was designed to develop a functional pharma-food using Salicornia europaea (SE). Tiny seeds from the mature SE were collected, and their biological activities were evaluated. The extraction yield of the seed in hot water was found to be 29.6% and the hot water extract (HWE) contained 25.7 mg/g total polyphenol (TP) and 11.5 mg/g total flavonoid (TF), which are similar to those contained in leaf and stem of SE. Among the subsequent organic solvent fractions, the ethylacetate (EA) fraction exhibited the highest content of TP (158.3 mg/g), TF (136.2 mg/g), and total sugar (228.3 mg/g). The EA fraction exhibited broad-range antibacterial activities against gram-positive bacteria, and the butanol fraction exhibited growth inhibitory effect against only Staphylococcus epidermidis. An antioxidation activity assay of the HWE and its fractions showed the EA fraction to have the highest radical scavenging activity with $RC_{50}$ values of 57.0, 29.0, and $28.9{\mu}g/ml$ against DPPH anion, ABTS cation, and nitrite, respectively. The $RC_{50}$ values of vitamin C against DPPH anion, ABTS cation, and nitrite were 10.7, 4.0, and $18.0{\mu}g/ml$, respectively, indicating that the EA fraction of SE has potent antioxidant compounds. In an anticoagulation assay, the EA fraction exhibited a 15-fold extended thrombin time at 5 mg/ml and activated partial thromboplastin time at 7 mg/ml, which are comparable to the activities of aspirin. The HWE and its fractions had no hemolysis activities against human RBCs at up to 1 mg/ml. These results suggest that the EA fraction from SE has a great potential as a new antibacterial and anticoagulation agent.

• IMMUNE NETWORK
• /
• v.15 no.2
• /
• pp.66-72
• /
• 2015

Currently, detecting biochemical differences before and after allogeneic stem cell transplantation (SCT) for improved prediction of acute graft-versus-host disease (aGVHD) is a major clinical challenge. In this pilot study, we analyzed the kinetics of circulating adipokine levels in patients with or without aGVHD before and after allogeneic SCT. Serum samples were obtained and stored at $-80^{\circ}C$ within 3 hours after collection, prior to conditioning and at engraftment after transplantation. A protein array system was used to measure the levels of 7 adipokines of patients with aGVHD (n=20) and without aGVHD (n=20). The resistin level at engraftment was significantly increased (p<0.001) after transplantation, regardless of aGVHD occurrence. In the non-aGVHD group, the concentrations of the hepatocyte growth factor (HGF) (mean values${\pm}$SD; $206.6{\pm}34.3$ vs. $432.3{\pm}108.9pg/ml$, p=0.040) and angiopoietin-2 (ANG-2) (mean values${\pm}$SD; $3,197.2{\pm}328.3$ vs. $4,471.8{\pm}568.4pg/ml$, p=0.037) at engraftment were significantly higher than those of the pre-transplant period, whereas in the aGVHD group, the levels of adipokines did not change after transplantation. Our study suggests that changes in serum HGF and ANG-2 levels could be considered helpful markers for the subsequent occurrence of aGVHD.

• Proceedings of the Korean Society of Developmental Biology Conference
• /
• 2003.10a
• /
• pp.101-101
• /
• 2003

Main strategy for a treatment of Parkinson's disease (PD), due to a progressive degeneration of dopaminergic neurons, is a pharmaceutical supplement of dopamine derivatives or ceil replacement therapy. Both of these protocols have pros and cons; former exhibiting a dramatic relief but causing a severe side effects on long-term prescription and latter also having a proven effectiveness but having availability and ethical problems Embryonic stem (ES) cells have several characteristics suitable for this purpose. To investigate a possibility of using ES cells as a carrier of therapeutic gene(s), human ES (hES, MB03) cells were transfected with cDNAs coding for tyrosine hydroxylase (TH) in pcDNA3.1 (+) and the transfectants were selected using neomycin (250 $\mu /ml$). Expression of TH being confirmed, two of the positive clone (MBTH2 & 8) were second transfected with GTP cyclohydrolase 1 (GTPCH 1) in pcDNA3.1 (+)-hyg followed by selection with hygromycin-B (150 $\mu /ml$) and RT-PCR confirmation. By immune-cytochemistry, these genetically modified but undifferentiated dual drug-resistant cells were found to express few of the neuronal markers, such as NF200, $\beta$-tubulin, and MAP2 as well as astroglial marker GFAP. This results suggest that over-production of BH4 by ectopically expressed GTPCH I may be involved in the induction of those markers. Transplantation of the cells into striatum of 6-OHDA- denervated PD animal model relieved symptomatic rotational behaviors of the animals. Immunohistochemical analyses showed the presence of human cells within the striatum of the recipients. These results suggest a possibility of using hES cells as a carrier of therapeutic gene(s).

• Korean Journal of Plant Resources
• /
• v.25 no.2
• /
• pp.193-199
• /
• 2012

This study was carried out to evaluate the agronomic characteristics and biological activities of $Sorghum$ $bicolor$ germplasms introduced from USA. Plant height, stem diameter, tiller number, leaf length, leaf width, leaf vein color, ear type, ear length and ear width have different from the cultivated accessions. Sweet-N-Sterile (#4) showed the tallest height, widest ear-width and stem diameter. Most of $Sorghum$ $bicolor$ plants may be available to use for bio-energy from these results showing big biomass. Antioxidant activities of 11 cultivars collected from USA were examined by DPPH assay and reducing power. Among the cultivars, Premium stock (#1), Early Sumac (#7), SS Silage (#9) and WGF Grain Sorghum (#11) showed a significantly higher antioxidant activity in comparison to others. Early Sumac (#7) and SS silage (#9) showed more strong reducing power activities than ${\alpha}$-tocopherol, a positive control. Premium stock (#1), Sweet-N-Sterile (#4), Early Sumac (#7) and SS Silage (#9) were also showed high antioxidant activities by DPPH assay and reducing power experiment. BMR Gold I (#3) displayed strong antimicrobial activity against $Escherichia$ $coli$ at minimum inhibitory concentrations (125 ${\mu}g$/ml).

• Clinical and Experimental Reproductive Medicine
• /
• v.29 no.2
• /
• pp.129-138
• /
• 2002

Objective: This study was to compare the characteristics between parthenogenetic mES (P-mES) cells and in vitro fertilization mES cells. Materials and Methods: Mouse oocytes were recovered from superovulated 4 wks hybrid F1 (C57BL/6xCBA/N) female mice. For parthenogenetic activation, oocytes were treated with 7% ethanol for 5 min and $5{\mu}g$/ml cytochalasin-B for 4 h. For IVF, oocytes were inseminated with epididymal sperm of hybrid F1 male mice ($1{times}10^6/ml$). IVF and parthenogenetic embryos were cultured in M16 medium for 4 days. Cell number count of blastocysts in those two groups was taken by differential labelling using propidium iodide (red) and bisbenzimide (blue). To establish ES cells, b1astocysts in IVF and parthenogenetic groups were treated by immunosurgery and recovered inner cell mass (ICM) cells were cultured in LIF added ES culture medium. To identify ES cells, the surface markers alkaline phosphatase, SSEA-1, 3,4 and Oct4 staining were examined in rep1ated ICM colonies. Chromosome numbers in P-mES and mES were checked. Also, in vitro differentiation potential of P-mES and mES was examined. Results: Although the cleavage rate (${\geq}$2-cell) was not different between IVF (76.3%) and parthenogenetic group (67.0%), in vitro development rate was significantly low in parthenogenetic group (24.0%) than IVF group (68.4%) (p<0.05). Cell number count of ICM and total cell in parthenogenetic b1astocysts ($9.6{\pm}3.1,\;35.1{\pm}5.2$) were signficantly lower than those of IVF blastocysts ($19.5{\pm}4.7,\;63.2{\pm}13.0$) (p<0.05). Through the serial treatment procedure such as immunosurgery, plating of ICM and colony formation, two ICM colonies in IVF group (mES, 10.0%) and three ICM colonies (P-mES, 42.9%) in parthenogenetic group were able to culture for extended duration (25 and 20 passages, respectively). Using surface markers, alkaline phosphatase, SSEA-l and Oct4 in P-mES and mES colony were positively stained. The number of chromosome was normal in ES colony from two groups. Also, in vitro neural and cardiac cell differentiation derived from mES or P-mES cells was confirmed. Conclusion: This study suggested that P-mES cells can be successfully established and that those cell lines have similar characteristics to mES cells.

• Asian-Australasian Journal of Animal Sciences
• /
• v.26 no.2
• /
• pp.178-188
• /
• 2013

The glycosaminoglycans (GAGs) present in the female reproductive tract promote sperm capacitation. When bovine sperm were exposed to 10 ${\mu}g/ml$ of one of four GAGs (Chondroitin sulfate, CS; Dermatan sulfate, DS; Hyaluronic acid, HA; Heparin, HP) for 5 h, the total motility (TM), straight-line velocity (VSL), and curvilinear velocity (VCL) were higher in the HP- or HA-treated sperm, relative to control and CS- or DS-treated sperm. HP and HA treatments increased the levels of capacitated and acrosome-reacted sperm over time, compared to other treatment groups (p<0.05). In addition, sperm exposed to HP or HA for 1 h before IVF exhibited significantly improved fertilizing ability, as assessed by 2 pronucleus (PN) formation and cleavage rates at d 2. Exposure to these GAGs also enhanced in vitro embryo development rates and embryo quality, and increased the ICM and total blastocyst cell numbers at d 8 after IVF (p<0.05). A real-time PCR analysis showed that the expression levels of pluripotency (Oct 4), cell growth (Glut 5), and anti-apoptosis (Bax inhibitor) genes were significantly higher in embryos derived from HA- or HP-treated sperm than in control or other treatment groups, while pro-apoptotic gene expression (caspase-3) was significantly lower in all GAG treatment groups (p<0.05). These results demonstrated that exposure of bovine sperm to HP or HA positively correlates with in vitro fertilizing ability, in vitro embryo developmental potential, and embryonic gene expression.

• Advances in Traditional Medicine
• /
• v.10 no.1
• /
• pp.7-12
• /
• 2010

Typha orientalis' stem (TOS) is traditionally used as an herbal medicine for difficulty in urination, galactophoritis purulenta, whooping cough, and allergic dermatitis. However, its effect in experimental models remains unknown. Here, we report the effect of TOS on the phorbol 12-myristate 13-acetate (PMA) plus calcium ionophore A23187-induced inflammatory cytokine production and extracellular signal-regulated kinase (ERK) activation in the human mast cell line, HMC-1. TOS inhibited PMA plus A23187-induced cytokines such as tumor necrosis factor-alpha (TNF-$\alpha$) and interleukin (IL)-6. Maximal inhibition rate of TNF-$\alpha$ and IL-6 production by TOS (1 mg/ml) was about 44.02%, and 45.20%, respectively (P < 0.05). In addition, TOS inhibited the expression of TNF-$\alpha$ and IL-6 mRNA under the same condition. Moreover, TOS partially blocked PMA plus A23187-induced ERK phosphorylation. These results suggested TOS could inhibit the cytokine production through blocking of ERK activity.

• Proceedings of the Korean Society of Developmental Biology Conference
• /
• 2003.10a
• /
• pp.104-104
• /
• 2003

As an effort to direct differentiation of human embryonic stem (hES, MB03) cells to dopamine-producing neuronal cells, Nurr1 was transfected using conventional transfection protocol into MB03 and examined the expression of tyrosine hydroylase (TH) after differentiation induced by retinoic acid (RA) and ascorbic acid (AA). Experimentally, cells were transfected with linearized Nurr1 cDNA in pcDNA3.1 (+)-hygovernight followed by selection in medium containing hygromycin-B (150 $\mu$/ml). Expression of Nurr1 mRNA was confirmed by RT-PCR and protein by immunocytochemistry in the drug resistant clones. In order to study the effect of Nurr1 protein on the differentiation pattern of ES cells, one of the positive clones (MBNr24) was allowed to form embryoid body (EB) for 2 days and were induced to differentiate for another 4 days using RA (1 $\mu M$) and AA (50 mM) (2-/4+ protocol) followed by selection in N2 medium for 10 or 20 days. After 10 days in N2 medium, cells immunoreactive to anti-GFAP, anti-TH, or anti-NF200 antibodies were 38.8%, 11%, and 20.5%, respectively. After 20 days in N2 medium, cells expressing GFAP, TH, or NF200 were 28%, 15% and 44.8%, respectively but approximately 9% of MB03 expressed TH protein when the cells were induced to differentiate using a similar prorocol, These results suggest that ectopic expression of Nurr1 enhances generation of TH+ cells as well as neuronal cells when hES cells were differentiated by 2-/4+ protocol.

• Proceedings of the Korean Society of Developmental Biology Conference
• /
• 2003.10a
• /
• pp.98-98
• /
• 2003

Recently, human embryonic stem (hES) cells have become very important resources for ES cell basic research, cell replacement therapy, and other medical applications; thus, efficient cryopreservation methods for these cells are needed. This study examined whether a newly developed minimum volume cooling (MVC) vitrification method, which was tested through cryopreservation of sensitive bovine oocytes, can be used for freezing hES cells. Feeder-free cultured hES cell (MB03) colonies were mechanically dissected into several small clumps following enzymatic treatment. We compared the freezing efficiency of a slow-cooling method using a cryo-module (0.4-0.6C/min, 20-30 clumps/vial) and MVC vitrification using a modified 0.5-ml French mini-straw designated as a MVC straw (>$20,000{\circ}C$/min, 10 clumps/straw) After thawing, in vitro survival of hES cell clumps was higher for MVC-vitrified cells (80.8%, 97/120) than for slow-cooled cells (38.2%, 39/102). Further, the proliferation rate of surviving MVC-vitrified cells was similar to that of control hES cells from 2 weeks after thawing. In addition, vitrified-thawed hES cells demonstrated a normal karyotype, were positively immunostained for surface marker antibodies (AP, SSEA-4 and TRA-1-60) and the Oct-4 antibody, and could differentiate into all three embryonic germ layer cells in vitro. This result demonstrates that hES cell clumps can be successfully cryopreserved by a newly developed MVC vitrification method without loss of human cell characteristics.

• Proceedings of the KSAR Conference
• /
• 2003.06a
• /
• pp.78-78
• /
• 2003

Embryonic stem (ES) cells, derived from preimplantation embryo, are able to differentiate into various types of cells consisting the whole body, or pluripotency. In contrast, terminally differentiated cells do not usually alter their nature but frequently die or transform if they are exposed to inappropriate external stimulations. In addition to the plasticity, ES cells are expected to be different from terminally differentiated cells in very many ways, such as patterns of gene expressions, ability and response of the cells in confronting environmental stimulations, metabolism, and growth rate. As a model system to differentiate these two types of cells, human ES cells (MB03) and terminally differentiated cells (HeLa), we examined the ability of these two types of cells in confronting a severe oxidative insult, that is $H_2O$$_2$. Approximately 1$\times$10$^4$ cells were plated in 96 well plate and serum starved for overnight. The conditioned cells were exposed to a various concentration of $H_2O$$_2$ fur 24 hrs and loaded with neutral red (50$\mu\textrm{g}$/ml) for 4 hrs, washed with PBS for 2 min three times, and entrapped dye was dissolved out using acetic ethanol. Cytotoxicity was determined by reading the amount of dye in the medium using microplate reader. equipped with 575 nm filter. Relative amount of the dye entrapped within MB03 or HeLa were not significantly different when cells were exposed up to 0.4 mM $H_2O$$_2$. However, this sharply decreased down to 0.12% in HeLa cells when the cells were exposed to 0.8 mM $H_2O$$_2$, while it was approximately 54% in MB03 suggesting that this concentration of $H_2O$$_2$ is the defensive threshold for HeLa cells. The resistance to oxidative stimulation reversed, however, when cells were co-treated with BSO (L-buthionine- 〔S, R〕-sulfoximine) which chelates intracellular GSH. This result suggests that cellular GSH is the major defensive mechanism of human ES cells. Induction of enzymes involved in GSH metabolism and type of cell death is currently being studied.