• 제목/요약/키워드: frozen cell

검색결과 328건 처리시간 0.021초

Effects of Antioxidants Supplement in Porcine Sperm Freezing on in vitro Fertilization and the Glutathione and Reactive Oxygen Species Level of Presumptive Zygotes

  • Park, Sang-Hyoun;Jeon, Yubyoel;Yu, Il-Jeoung
    • 한국수정란이식학회지
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    • 제32권4호
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    • pp.337-342
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    • 2017
  • The present study was aimed to determine the effects of green tea extract (GTE) and beta-mercaptoethanol (${\beta}-ME$) supplementation in boar sperm freezing extender on in vitro fertilization (IVF) and reactive oxygen species (ROS) and glutathione (GSH) levels of presumptive zygotes (PZs). Experimental groups were allocated into lactose egg yolk (LEY) without antioxidant (control), GTE (1,000 mg/l in LEY) and ${\beta}-ME$ ($50{\mu}M$ in LEY). In freezing, spermatozoa extended with LEY were cooled to $5^{\circ}C$ for 3 h and then kept at $5^{\circ}C$ for 30 min following dilution with LEY containing 9% glycerol and 1.5% Equex STM. The final sperm concentration was $1{\times}10^8/ml$. Spermatozoa were loaded into straws and frozen in nitrogen vapor for 20 min. For IVF, oocytes were matured in NCSU-23 medium and co-cultured with spermatozoa following thawing at $37^{\circ}C$ for 25 sec. At 12 h following IVF, IVF parameters (sperm penetration and monospermy) were evaluated. In addition, GSH and ROS levels of PZs were determined by Cell Tracker Blue CMF2HC and DCHFDA, respectively. IVF parameters did not show any significant difference among the experimental groups. GSH and ROS levels of PZs were not significantly different between groups. In conclusion, antioxidant supplementation in boar sperm freezing could not influence IVF parameters, ROS and GSH levels of PZs.

정자 미세주입에 의한 소 난포란의 체외수정 (In Vitro Fertilization of Bovine Oocytes Matured In Vitro by Microinjection of Spermatozoa)

  • 김선구;곽대오;박충생;쿠란티;메틀러
    • 한국가축번식학회지
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    • 제16권3호
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    • pp.239-246
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    • 1992
  • 미성숙 나포란을 20% FCS가 첨가된 TCM-199으로써 24시간 동안 5% CO2, 8% O2의 공기 조건하에서 체외성숙시킨 다음 난자의 위란강내로 정자를 미세주입하여 체외수정을 실시하였다. 미세주입 하기전 정액을 0.75% BSA가 첨가된 Ham's F-10 배양액으로 2시간 동안 5% CO2, 8% O2의 공기 조건하에서 배양함으로써 수정능 획득을 하도록하였으며 이어 30분 동안 12mM의 dbcGMP와 10mM의 imidazol이 함유된 Ham's F-10 배양액으로 배양함으로써 첨체반응을 유도하였다. 본 실험에서 정자의 미세주입 후 제 2극체와 전핵형성이 일어난 난자의 비율은 각각 9.5, 5.4%였으며 2세포기 이후로 발달한 난자의 비율은 4.1%이었다.

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체외성숙 조건이 소 체외수정란의 체외발달에 미치는 영향 (Effects of In vitro Maturation Condition on Bovine IVF Embryos Development)

  • 최선호;류일선;김일화;박수봉;연성흠;진현수;서상욱;이충섭;손동수
    • 한국수정란이식학회지
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    • 제14권2호
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    • pp.113-119
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    • 1999
  • This study was performed to improve the development of the in vitro fertilized bovine embryos by the condition of in vitro maturation. COCs were matured in TCM 199 supplemented with 0.1% PVA, 10ng/ml EGF, Hormones (5$\mu\textrm{g}$/ml FSH, 10 IU hCG, 1 $\mu\textrm{g}$/ml estradiol 17-$\beta$) or granulsa cell+Hormones atmosphere 39$^{\circ}C$, 5% CO2, 95% air for 24hrs. Matured oocytes were fertilized with frozen-thawed semen capacitated with 5mM caffein in BO medium for 20 hrs. IVF embryos were cultured in TCM 199 containing with hormones(same as matured medium), 10% FBS and co-culture with bovine oviduct epitherial cells. Maturation rates of COCs were showed 73.8%, 78.5%, 83.2% and 87.6% respectively, and were significant differences between PVA, EGF, and Hormones, GC+Hormones(p<0.05). The cleavage rates of IVF embryos were revealed 72.5%, 78.4%, 82.3% and 84.2% and showed same tendency as maturation rates(p<0.05). The blastocysts matured by above maturation condition and cultured for 7~10 days after fertilization had 34.4, 43.6, 52.3 and 59.3 cells had no differences among the treatments. These results suggest that high molecules as a substitutes of serum and growth factor may induce nuclear resumption of COCs but we need more study to produce transferable IVF blastocysts by use of that agents.

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Crystallization and Preliminary X-Ray Crystallographic Analysis of UDP-N-Acetylglucosamine Enolpyruvyl Transferase from Haemophilus influenzae in Complex with UDP-N-Acetylglucosamine and Fosfomycin

  • Yoon, Hye-Jin;Ku, Min-Je;Ahn, Hyung Jun;Lee, Byung Il;Mikami, Bunzo;Suh, Se Won
    • Molecules and Cells
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    • 제19권3호
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    • pp.398-401
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    • 2005
  • The bacterial enzyme UDP-N-acetylglucosamine enolpyruvyl transferase catalyzes the first committed step of peptidoglycan biosynthesis, i.e., transfer of enolpyruvate from phosphoenolpyruvate to UDP-N-acetyl-glucosamine. We have overexpressed the enzyme from Haemophilus influenzae in Escherichia coli and crystallized it in the apo-form, as well as in a complex with UDP-N-acetylglucosamine and fosfomycin using ammonium sulfate as the precipitant. X-ray diffraction data from a crystal of the apo-form were collected to $2.8{\AA}$ resolution at 293 K. The crystal quality was improved by co-crystallization with UDP-N-acetylglucosamine and fosfomycin. X-ray data to $2.2{\AA}$ have been collected at 100 K from a flash-frozen crystal of the complex. The complex crystals belong to the orthorhombic space group I222 (or $I2_12_12_1$) with unit-cell parameters of a = 63.7, b = 124.5, and $c=126.3{\AA}$. Assuming a monomer of the recombinant enzyme in the crystallographic asymmetric unit, the calculated Matthews parameter ($V_M$) is $2.71{\AA}^3Da^{-1}$ and solvent content is 54.6%.

냉풍건조에 의한 전갱이 조미 반건품(半乾品)의 제조 및 빙결점 동결 저장 중 품질안정성 (Preparation of Seasoned and Semi-dried Horse Mackerel by Cold Air Drying and Quality of Its Product during Partially Frozen Storage)

  • 양승택
    • 한국식품과학회지
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    • 제29권5호
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    • pp.925-931
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    • 1997
  • 전갱이의 조미 반건품을 대량으로 생산하기 위한 기초자료를 얻기 위해, 전갱이를 조미 반건조한 후 나일론+폴리에틸렌+선형 저밀도폴리에틸렌$(0.015/0.045/0.04\;{\mu}m)$의 적층필름으로 진공포장하여 빙결점 동결$(-3{\pm}0.5^{\circ}C)$ 저장하면서 저장 중의 품질변화를 비교 검토하였다. 전갱이 조미 반건품을 제조하기 위한 가장 적합한 건조온도 및 시간은 상대습도 $50{\pm}5%$, 풍속 2m/sec 조건에서 각각 $20^{\circ}C$ 및 4시간이었으며 진공포장제품의 저장기한은 42일이었고 비포장제품의 경우는 28일이었다.

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Production and Transfer of In Vitro fertilized Hanwoo Embryos with Serum-free Media

  • Yoon, Do-Joong;Kim, Gye-Woong;Kim, Kon-Joong;Park, Byung-Kwon;Cui, Xiang-Shun;Kim, Nam-Hyung;Lee, Jong-Wan
    • 한국수정란이식학회지
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    • 제24권4호
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    • pp.281-287
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    • 2009
  • As a simple and economical method for in vitro produced embryos, we have used BSA instead of serum for the production and embryo transfer of Hanwoo in vitro fertilized (IVF) embryos and obtained the following results: 1) When using serum (FBS; fetal bovine serum) or BSA-containing culture media as the initial culture media for immature oocytes, it is regarded as inappropriate to add only BSA to the culture solutions from maturation of the immature oocytes to development stage culture, but serum still needs be added though there is no significant difference in the concentration, with a change from 5% to 10%. 2) The results of culturing IVF embryos after development (4 cell stage) in the Medium199 solutions containing BSA instead of serum (FBS) showed that 0.3% BSA concentration is not optimal and 0.5% or higher BSA concentration has no significant difference among 0.5%, 0.7%, 1% and 2% (p > 0.05). 3) The post-freezing survival ratio after development in 5% FBS-Medium199 showed that 1% BSA concentration of the culture solution is the most suitable in the BSA concentrations of 0.3% (51%), 0.5% (67%), 0.7% (69%), 1% (77%) and 2% (75%). 4) The pregnancy rates of the transplanted fresh(not frozen) blastocyst had no significant concentration dependency (p > 0.5), and the average pregnancy rate was 63.8%. 14% of overweight calves were found among the calves given birth to by the transfer of IVF blastocysts cultured in the serum-added culture solution, but none was found in the experimental groups in which BSA was added instead of serum.

동결보존액에 울금에서 추출한 Curcumin의 첨가가 정자의 운동성과 활성산소 생성에 미치는 영향 (Effects of Curcumin from Turmeric Supplementation in Freezing Buffer on Sperm Motilities and Reactive Oxygen Species Generation)

  • 이은주;김대영
    • 한국수정란이식학회지
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    • 제32권3호
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    • pp.249-255
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    • 2017
  • In this experiment, we determined the effect of curcumin supplementation in freezing buffer for miniature pig sperm cryopreservation. Each ejaculate was diluted with modified Modena B extender and mixed with lactose-egg yolk (LEY extender, 80% v/v lactose solution [310 mM], 20% v/v egg yolk, and $100{\mu}g/mL$ kanamycin sulfate) and LEY-glycerol Orvus ES Paste (LEYGO, 89.5% v/v LEY, 5% v/v glycerol, 1.5% v/v Orvus ES Paste), 100 mM trehalose supplemented with 0, 10, 50, 100, and $500{\mu}M$ of curcumin from turmeric, respectively. Following equilibration, the 0.5 mL French straws were frozen and plunged into $LN_2$ tank for 7 days at least. Sperm parameter and oxidative byproducts were determined by the computer assisted sperm motility analysis (CASA) and fluorescence-activated cell sorting (FACS) as compared with each groups. Supplementation of curcumin had no effect on sperm motility, progressive motility and curvilinear velocity. However, average-path velocity and straight-line velocity were significantly higher in $10{\mu}M$ curcumin group ($100.9{\pm}8.8{\mu}m/s$, $61.7{\pm}2.9{\mu}m/s$, respectively) than control group ($77.8{\pm}3.9{\mu}m/s$, $46.4{\pm}3.0{\mu}m/s$, respectively) (p < 0.05). In addition, the level of the O2 radical and H2O2 were comparatively decreased in curcumin groups by evaluation of ethidium and DCF fluorescence. According to the results, curcumin can improve sperm kinetic variables and alleviate ROS induced cryoinjury to pig sperm.

한우에 있어서 체외수정된 난포란의 배발생에 관한 연구 (Study on Development in vitro of Follicular Oocytes fertilized in vitro in Korean Native Cattle)

  • 서태광;박항균
    • 한국가축번식학회지
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    • 제14권4호
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    • pp.253-261
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    • 1990
  • This study was carried out to investigate the factors affecting development in vitro of follicular oocytes fertilized in vitro in Korean Native Cattle. The bovine ovaries were obtained at a slaughter house and the follicular oocytes were recovered by aspirating the follicular fluid from the visible follciles of 3~6mm. The bovine oocytes were matured in vitro for 20~24 hours in TCM0-199 containing 10% FCS and hormones (0.02AU/ml FSH, 10$\mu\textrm{g}$/ml LH, 1$\mu\textrm{g}$/ml estradiol-17$\beta$). The matured oocytes were fertilized in vitro using Percoll-separated frozen-thawed spermatozoa in BO solution containing caffeine(5mM) and heparin(10$\mu\textrm{g}$/ml). Twenty-four hours after insemination, the oocytes were cultured in vitro and then the effects of cumulus cell layer, co-culture with cumulus cells, bovine oviduct epithelial cells from ampulla or isthmus on development of ova, were studied. The results obtained are summarized as follows : 1. The in vitro development degree of oocytes attached with compact and dense layered cumulus cells was higher than that with 3~4 layered cumulus cells to be 9~16cells(P<0.01). 2. When the in vitro fertilized oocytes were co-cultured with bovine oviduct epithelial cells or cumulus cells, the development rate to be morula was 20.2% and 12.7%, respectively and the rates were higher than that of control, 2.1%(P<0.05). 3. The development rate to be morula was 15.8% and 23.8%, respectively when the in vitro fertilized oocytes were co-cultured with bovine oviduct epithelial cells from ampulla or isthmus, and the rates were higher than that of control, 0%(P<0.05%).

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히말라야의 토양 곰팡이, Preussia sp. 배양액으로부터 추출된 화학 성분들 및 항 염증 활성 (Chemical constituents from the culture filtrate of a Himalayan soil fungus, Preussia sp. and their anti-inflammatory activity)

  • 윤의중;서승석;임정한;김일찬;한세종
    • 미생물학회지
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    • 제54권1호
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    • pp.18-23
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    • 2018
  • 히말라야산의 동토에서 발견된 Preussia sp.로부터 신규 벤조산 유도체 benzyl 2,4-di(benzyloxy)benzoate (1)와 6개의 알려진 화합물들(2-7)이 분리되었다. 신규물질 및 다른 화합물들의 구조는 1D-와 2D NMR실험 및 문헌값과의 비교에 의하여 결정되었다. 화합물 2-7은 동 속 및 Sporormiaceae과에서 처음으로 얻어졌다. 분리된 물질들은 쥐의 대식세포 RAW 264.7 cells에서 lipopolysaccharide (LPS) 처리에 의하여 유도된 nitric oxide (NO)의 생성량 억제를 측정하였다. 화합물 1번과 2번은 100 mg/ml의 농도로 처리했을 때 LPS에 의하여 유도된 NO 생성량을 각각 50.7% 및 88.5%까지 억제하는 것을 보였다.

Optimum Combination of Thermoplastic Formability and Electrical Conductivity in Al-Ni-Y Metallic Glass

  • Na, Min Young;Park, Sung Hyun;Kim, Kang Cheol;Kim, Won Tae;Kim, Do Hyang
    • Metals and materials international
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    • 제24권6호
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    • pp.1256-1261
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    • 2018
  • Both thermoplastic formability and electrical conductivity of Al-Ni-Y metallic glass with 12 different compositions have been investigated in the present study with an aim to apply as a functional material, i.e. as a binder of Ag powders in Ag paste for silicon solar cell. The thermoplastic formability is basically influenced by thermal stability and fragility of supercooled liquid which can be reflected by the temperature range for the supercooled liquid region (${\Delta}T_x$) and the difference in specific heat between the frozen glass state and the supercooled liquid state (${\Delta}C_p$). The measured ${\Delta}T_x$ and ${\Delta}C_p$ values show a strong composition dependence. However, the composition showing the highest ${\Delta}T_x$ and ${\Delta}C_p$ does not correspond to the composition with the highest amount of Ni and Y. It is considered that higher ${\Delta}T_x$ and ${\Delta}C_p$ may be related to enhancement of icosahedral SRO near $T_g$ during cooling. On the other hand, electrical resistivity varies with the change of Al contents as well as with the change of the volume fraction of each phase after crystallization. The composition range with the optimum combination of thermoplastic formability and electrical conductivity in Al-Ni-Y system located inside the composition triangle whose vertices compositions are $Al_{87}Ni_3Y_{10}$, $Al_{85}Ni_5Y_{10}$, and $Al_{86}Ni_5Y_9$.