• 대한한방내과학회지
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• 제38권1호
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• pp.1-9
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• 2017

Objective: This study investigated the effect of purified Carthamus tinctorius (C. tinctorius) extracted with a hot water and ethanol method on adipogenic differentiation of mouse bone marrow-derived mesenchymal stromal stem cells (mBMSCs). Methods: The C. tinctorius was extracted using hot water and ethanol. The samples were concentrated by a rotary evaporator and were then dried using a freeze-dryer. The mBMSCs were cultured and maintained in a minimum essential medium eagle alpha (${\alpha}-MEM$) supplemented with 10% FBS and 1% antibiotic antimycotic solution. To induce adipogenic differentiation, the cells were treated with Dulbecco's modified eagle's medium-low glucose (DMEM-LG) containing 1 mg/mL insulin, 1 mM dexamethasone, and 0.5 mM 3-isobutyl-1-methylxanthine. To evaluate the adipogenic differentiation ability, oil-red O staining was performed after adipogenic differentiation for 21 days. The mRNA expression and protein level of adipogenic-related genes were quantified by quantitative real-time PCR and western blotting, respectively. Results: In the results of the MTT assay, no concentrations of C. tinctorius extracts showed toxicity on mBMSCs, so we fixed the treatment concentration of the extract at 100 ng/mL. In oil-red O staining, the water-C. tinctorius extract treatment significantly decreased adipogenic differentiation compared with the control and ethanol extract groups. The water-C. tinctorius extract group in particular showed reduced mRNA and protein expression of Peroxisome proliferator-activated receptor gamma ($Ppar{\gamma}$) and CCAAT/enhancer-binding protein alpha ($C/ebp{\alpha}$), which are adipogenic-related transcription factors. Conclusion: These data suggest that extract of C. tinctorius decreased the adipogenic differentiation of mBMSCs, while only water-C. tinctorius extract had an effect on different adipogenesis in mBMSCs. The C. tinctorius will be a useful therapeutic reagent for the prevention of obesity-related diseases such as diabetes, hyperlipidemia, coronary artery disease, and osteoporosis.

• Clinical and Experimental Reproductive Medicine
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• 제21권1호
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• pp.99-110
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• 1994

One consequence of fertilization in mammals is an increased resistance of the zona pellucida (ZP) to proteases and various chemical reagents. This phenomenon has been called 'zona pellucida hardening' (ZPH), and it is generally accepted that it is caused by the secretory products of cortical granules released by the egg at fertilization. ZP of mouse oocytes maturing in vitro in a chemically defined medium becomes progressively more resistant to solubilization by chymotrypsin ("Spontaneous" ZP hardening). In the present study, it was aimed to find the specificity of spontaneous ZPH in relation to its possible relevance to the cortical reaction and the physiological block to polyspermy. When a maturation inhibitors, cAMP analog(dbcAMP) and phosphodiesterase inhibitor (IBMX) was added to culture medium, it prevent spontaneous ZPH of mouse oocyte during in vitro culture. Thus spontaneous ZPH requires GVBD, since it is prevented by those agents, which inhibit GVBD in vitro. However, culture for 3 hours in the presence of PMA(lOng/ml), a protein kinase C activator, resulted in ZPH without GVBD, thus suggesting that ZPH may be regulated independently apart from the event of GVBD. Pretreatment of mouse oocyte with FBS result in partially inhibitory effect on subsequent spontaneous ZPH. Induction of GVBD in vivo had a inhibitory effect on the spontaneous ZPH, but subsequent spontaneous ZPH. Induction of GVBD in vivo had a inhinbitory effect on the spontaneous ZPII, but had no inhibitory effect on PMA-induced ZPH. Treatment with a microfilament formation blocker(cytochalasin-B) at 1${\mu}g$/ml concentration, resulted in the excellent inhibitory effect on spontaneous ZPH. However cytochalasin-B did not inhibit PMA-induced ZPH. Thus this suggesting that spontaneuse ZPH had a different mechanism from PMA-induced ZPH.

• 한국어병학회지
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• 제17권2호
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• pp.75-82
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• 2004

인터페론은 바이러스에 감염된 세포에서 생성되어 다른 세포로 하여금 바이러스의 증식을 억제하는 단백질을 합성하게 하는 사이토카인의 일종으로서 바이러스에 대한 방어작용에서 중요한 역할을 담당한다. 본 연구 결과 강력한 인터페론 유도물질로 알려져 있는 poly inosinic : cytidylic acid (poly I:C)를 잉어에 주사한 결과 SVCV에 대한 항바이러스 활성을 확인할 수 있었다. 즉, Poly I:C 주사구에서 대조구에 비해 누적폐사율이 감소하였으며, 또한 두신백혈구를 분리하여, poly I:C를 처리한 결과 interferon-like cytokine (ILC)이 생성되었다. Crude ILC의 항바이러스 활성을 cytopathic effect reduction (CPER) assay로 조사한 결과, 적정 HKLs 농도는 1×$10^6$cells/ml으로 나타났으며, 20~50$\mu{g}$/ml 농도의 poly I:C를 처리했을 때 유의적으로 증가하였다. ILC 생성을 위한 적정온도 및 FBS의 농도는 각각 20$20^\circ{C}$와 5%로 나타났다.

• 한국수정란이식학회지
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• 제24권2호
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• pp.121-125
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• 2009

In the present study, effects of concentration of cryoprotectant solutions on the nuclear maturation of vitrifiedthawed bovine oocytes were examined. Also, the developmental capacity of vitrified-thawed immature oocytes following ICSI was investigated. Oocytes were cultured in TCM-199 medium supplemented with 5% FBS at $38^{\circ}$C in 5% $CO_2$ and air. The in vitro maturation rate of vitrified oocytes was 24.5 ${\pm}$ 4.2%. The in vitro maturation rate of vitrified oocytes was lower than that of the control (72.0 ${\pm}$ 3.5%, p<0.05). The in vitro maturation rate of vitrified${\sim}$thawed oocytes incubated in TCM-199 medium supplemented with 1.0${\sim}$5.0 ug CB were 26.7 ${\pm}$ 3.2%, 35.7 ${\pm}$ 3.2%, 54.0 ${\pm}$ 3.0%, 42.5 ${\pm}$ 3.6%, respectively. The in vitro maturation rate (57.0 ${\pm}$ 3.0%) of the vitrified-thawed oocytes treated with 3.0 ${\mu}$g CB for 20 min was the highest of all vitrification groups, although the maturation rate were significantly (p<0.05) lower than those of fresh oocytes. The in vitro maturation rates of the vitrified-thawed (with EDS and EDT) oocytes were 53.8 ${\pm}$ 3.4%, 51.1 ${\pm}$ 3.5%, respectively. This results were lower than the control group (72.0 ${\pm}$ 3.0%). The in vitro developmental rates of the vitrified-thawed oocytes following ICSI were 28.6 ${\pm}$ 4.5%, 25.6 ${\pm}$ 4.3%, respectively. This results were lower than the control group (40.0 ${\pm}$ 4.0%).

• 한국수정란이식학회지
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• 제24권2호
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• pp.115-119
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• 2009

In the present study, effects of concentration of cryoprotectant solutions on the nuclear maturation of vitrifiedthawed porcine oocytes were examined. Oocytes were cultured in TCM-199 medium supplemented with 5% FBS at $38^{\circ}$C in 5% $C0_2$ and air. The percentage of monospermy in the toxicity group and vitrification group (22.0 ${\pm}$ 3.0% and 31.5 ${\pm}$ 3.5%) was decreased compared with that of the control group (44.0 ${\pm}$ 4.0%). The percentage of in vitro development to blastocyst in the toxicity group and vitrification group (12.0 ${\pm}$ 2.5% and 14.8 ${\pm}$ 2.8%) was decreased compared with that of the control group (28.0 ${\pm}$ 3.0%, p<0.05). The survival and in vitro developmental rate of oocytes vitrification-thawed with EDS and EDT + TCM-199 medium supplemented with 0.1% PVA were 46.3 ${\pm}$ 3.0%, 54.5 ${\pm}$ 3.8% and 14.8 ${\pm}$ 2.5%, 16.4 ${\pm}$ 2.7%, respectively. This results were lower than the control group (28.0 ${\pm}$ 3.5%). The in vitro developmental rate of embryos vitrified with EDS and EDT supplemented PVA did not have a significant difference. The survival and in vitro developmental rate of vitrified-thawed morula and blastocyst embryos were 44.2 ${\pm}$ 3.5%, 17.3 ${\pm}$ 3.0% and 48.1 ${\pm}$ 4.2%, 18.5 ${\pm}$ 3.5%, respectively. Vitrified morulae and blastcyst embryos had a lower survival and developmental rates than their control counterparts.

• 대한한의학회지
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• 제24권3호
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• pp.35-44
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• 2003

Objective : This study was performed to evaluate the effects of Palmijihwang-hwan (Baweidehuang-wan) and Obaeja (Galla Rhois) on the regeneration of periodontal tissue. Methods : In this study, we used MC3T3-El cells, such as osteoblast-like cell lines and human periodontal ligament fibroblasts, for experimental material. We separated each type of cells into a control group and an experimental group. In the control group, the cells were cultivated for 48 hours with distilled water and media which contained 10% fetal bovine serum (FBS) and penicillin (l00unit/ml)-streptomycin ($l00{\mu\textrm{g}}/ml$) at $37^{\circ}$ in 5% $CO_2$ gas. In the experimental group, the cells were cultivated for 48 hours with Palmijihwang-hwan extract and Obaeja extract (concentrations $1{\mu\textrm{g}}/ml,{\;}25{\mu\textrm{g}}/ml,{\;}50{\mu\textrm{g}}/ml$) under the same conditions as the control group. Investigating the regeneration of periodontal tissue was performed by evaluating proliferation, the activity of alkaline phosphatase and the synthetic ability of proteins using those cultivated cells by means of microculture tetrazolium (MTT) assay, alkaline phosphatase substrate kit and protein assay kit. Results : 1. In vitro, Palmijihwang-hwan extract increased the proliferation of MC3T3-El cells. 2. In vitro, Obaeja extract increased the activity of alkaline phosphatase and the synthetic ability of protein in MC3T3-El cells and human periodontal ligament fibroblasts depending on Obaeja extract's concentration. Conclusion : Obaeja extract can be developed as a subsidiary medicine for the regeneration of periodontal tissue. Further studies to evaluate the different concentrations the Obaeja extract and clinical trials in vivo are suggested.

• 한국식품과학회지
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• 제23권3호
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• pp.370-374
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• 1991

마늘의 돌연변이유발 억제효과와 사람의 HT-29 결장암 세포의 성장저해 효과에 대하여 검토하였다. 마늘의 메탄올 추출물은 김치 제조시 사용되는 첨가농도 2%에서 아플라톡신 $B_1(AFB_1)$과 MNNG에 대해 항돌연변이적 효과를 나타내었다. 이 메탄올 추출물의 농도를 증가시켰을 때 Salmonella typhimurium TA98과 TA100에서 모두 $AFB_1$에 대하여 돌연변이유발 억제효과가 증가되었다. 이를 다시 분획하였을 때 비극성 분획인 클로르포름 분획에서 더 큰 돌연변이유발 억제효과를 관찰할 수 있었다. 또한 마늘의 클로르포름 분획은 사람의 HT-29 결장암 세포의 성장을 FBS 농도 1%와 5%에서 각각 저해하였다.

• 한방비만학회지
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• 제18권1호
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• pp.10-18
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• 2018

Objectives: Metabolic syndrome is correlated with increased cardiovascular risk and characterized by several factors, including visceral obesity, hypertension, insulin resistance, and dyslipidemia. Several members of a large family of nonselective cation entry channels, e.g., transient receptor potential (TRP) melastatin 7 (TRPM7) channels have been associated with the development of cardiovascular diseases. The purpose of this study was to investigate the effects of Dangkwisoo-san, ginger and curcumin on TRPM7 channel. Methods: Human embryonic kidney (HEK) 293 cells stably transfected with the TRPM7 expression vectors were maintained in Dulbecco's modified Eagle's medium (DMEM) supplemented with 10% fetal bovine serum (FBS), 1% penicillin/streptomycin, $5{\mu}g/mL$ blasticidin, and 0.4 mg/mL zeocin in a humidified 20% $O_2$/10% $CO_2$ atmosphere at $37^{\circ}C$. Whole-cell patch clamp recordings were obtained using an Axopatch 700B amplifier and pClamp v.10.4 software, and signals were digitalized at 5 kHz using Digidata 1422A. Results: Dangkwisoo-san extract (100, 200, 300, 400, and $500{\mu}g/mL$) inhibited the outward and inward TRPM7 whole-cell currents at dose dependent manner and the half maximal inhibitory concentration $(IC)_{50}$ of Dangkwisoo-san was $218.3{\mu}g/mL$. Also, ginger extract (100, 200, 300, 400, and $500{\mu}g/mL$) inhibited the outward and inward of TRPM7 whole-cell currents in a dose dependent manner and the $IC_{50}$ of ginger was $877.2{\mu}g/mL$. However, curcumin had no effects on TRPM7 whole-cell currents. Conclusions: These results suggest that both Dangkwisoo-san and ginger have good roles to inhibit the TRPM7 channel, suggesting that Dangkwisoo-san and ginger are considered one of the candidate agents for the treatment of metabolic syndrome such as cardiovascular disease.

• 한국수정란이식학회지
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• 제11권3호
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• pp.259-269
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• 1996

The present study was undertaken to examine the critical effect of $Ca^2$+ concentration on electrostimulation and post-electrostimulation media for electric activation of in vitro matured oocytes of Korean Native Cattle. Oocytes collected from slaughterhouse ovaries were matured in TCM 199 containing FSH, estradiol-17$\beta$ and FBS with granulosa cell monolayer for 24 hours and denuded with hyaluronidase. And then cumulus-free oocytes were submitted to a DC field of 1.0 kV/cm for 60 $\mu$sec in electroporation media(0.28 M mannito' and PBS) with different $Ca^2$+ concentations (0.00, 0.05, 0.10 and 0.15 mM). Stimulated oocytes were stained and examined for pronuclear formation after incuhation in SOF for 12 hours. The rates of pronuclear formation in hovine oocytes electrically stimulated in 0.28 M mannitol with 0.05, 0.10 and 0.15 mM $Ca^2$+(60.3, 82.2 and 75.0%) were significantly higher than without $Ca^2$+(6.3%) at 12 hours after an electric pulse(p<0.005). The activation rates of Korean Native Cattle oocytes stimulated in PBS supplemented with 0.05, 0.10 and 0.15 mM $Ca^2$+(71.0, 75.8 and 75.4%) were significantly higher than without $Ca^2$+(23.5%) after post-stimulation incubation(p<0.005). After incubation of oocytes in SOF with and without $Ca^2$+ following electric stimulation in 0.28 M mannitol with 0.10 mM $Ca^2$+, the rates of pronuclear formation of bovine oocytes in $Ca^2$+-free SOF(85.7%) was significantly higher than in SOF with 1.71 mM $Ca^2$+(62.5%, p<0.05). When oocytes were stimulated in two electrostimulation media supplemented with $Ca^2$+ and incubated in $Ca^2$+-free SOF, there were no significant differences in the rates of pronuclear formation hetween 0.28 M mannitol and PBS. These results indicate that a single electric pulse could induce activation of Korea Native Cattle oocytes in 0.28 M mannitol and PBS supplemented with $Ca^2$+. Furthermore, to improve the activation rates, it was hetter that stimulated oocytes were incubated in $Ca^2$+-free SOF after electric stimulation than in SOF with $Ca^2$+.

• Journal of Periodontal and Implant Science
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• 제37권1호
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• pp.115-124
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• 2007

Silica is known as a promising osteoconductive material, and polycaprolactone is a bioactive and degradable material. The purpose of this study was to monitor the differentiation of MC3T3-E1 cells cultured on the layer-built silica/poly caprolactone non-woven fabric produced by electrospinning. Non-woven fabric (silica, polycaprolactone, PSP, SPS) was made by electrospinning and they were inserted in the 48 well cell culture plate. MC3T3-E1 cells were prepared by subculture. Cells were seeded to each well $1{\times}10^5$ concentration per well. Dulbecco's modified eagle medium with 10% FBS and 1% antibiotic-antimycotic solution was used. Confocal laser scanning microscope was taken 4 hours after incubation (95% air. 5% $CO_2$, $37^{\circ}C$). Cell proliferation was monitored by spectrophotometer on 1, 7, 14 days, and the morphology of the growing cells was observed by field emission scanning electron microscope. To monitor the differentiation of osteoblasts on the materials, MC3T3-E1 cells were incubated in 48 well culture plate after seeding with the density of $1{\times}10^5$ concentration. Then ELISA kit & EIA kit were used on to assess osteocalcin and osteopontin expression respectively. The other conditions were the same as above. MC3T3-E1 cells were proliferated well on all of the materials. There were no statistical differences among them. The osteopontin expression of silica, PSP, SPS was significantly higher than other groups on day 3 (p/0,05), but after that time, there were no statistically signigicant differences. The osteocalcin expression was significantly higher in silica and PSP than other groups on day 14. These findings show that PSP was as good as silica on the effect of osteoblast differentiation. The PSP non-woven fabric may have the possibility as bone graft materials.