한우에서 반복 과배란처리에 의한 수정란 채란 가능성: 사례보고

공일근,정세환,손병훈,이정우,정수룡,오인석,정경일,황금영,배인휴 韓國受精卵移植學會 2002 한국동물생명공학회지 Vol.17 No.1

본 연구는 고능력 한우 공란우의 이용효율을 극대화하기 위하여 반복 과배란처리가 수정란회수율에 미치는 영향을 알아보고자 수행하였다 공란우의 선발은 DNA marker에 의한 고급육을 선발하였다. 선발된 공란우는 Folltropin-V와 PG를 병용 처리하여 과배란을 유도하였고, FSH 처리 6회째 PG를 처리하여 발정을 유도하였다. 발정유도된 공란우의 수정은 12시간 간격으로 3회, 1회당 2 straw식 인공수정을 실시하였다. 수정 후 7~8일째 비외과 The purpose of this study was carried out to determine the possibility of continued embryo collection by repeatable superovulation from a Hanwoo elite cow, who was already estimated to an elite cow by DNA analysis. The donor cow was superovulated with Folltropin-V and PG combination method for 5 times, 2 months interval during 10 months. The estrus cows were inseminated with frozen semen 3 times 12 hrs interval. Embryo collection from superovulated cows was flushed non-surgical method at 7 to 8 days after artificial insemination. The superovulated cow was collected from 1 to 5 times as total (14, 4, 10, 6, 9 embryos; total 30 embryos) and transferable embryos (7, 4, 10, 3, 6; MeanS.E : 6.02.7 embryos). Seventeen recipients were transferred 24 embryos and then detected to pregnant 10 recipients after embryo transfer (58.8%; 10/17 recipients) The results indicated that the possibility of repeatable superovulation from elite donor cow for embryo transfer could be confirmed in this study, although the replicated number was not enough.

개 인공수정

공일근 한국수정란이식학회 2002 한국수정란이식학회 학술대회 Vol.2002 No.1

Royal Jelly 첨가가 동결융해 후 개 정자의 활력도 및 생존성에 미치는 영향

공일근,조성균 韓國受精卵移植學會 2001 한국동물생명공학회지 Vol.16 No.1

This study was conducted to evaluate whether \"Royal jelly\" (RJ) added to Tris-buffer dilute contributed to supporting post-thaw viability and longevity of frozen canine spermatozoa. Two Japanese spitzs (2 to 4 years of age) were used as a semen donor. Semen was collected by manual masturbation and separated into 3 fractions. Only the sperm-rich fraction having sperm motility of more than 70%, containing sperm concentration of 2~410 cells/ml and having dead or abnormal spermatozoa of less than 15% was used for the experiment. Each ejaculated semen was centrifuged at 400 g for 5 min and then diluted in a Tris-buffer supplemented with 20 ml egg yolk (Ext I), 4% glycero1 and 1% Equex STM Paste (Ext II) or g1ycero1, Equex STM paste and RJ of various concentrations (Ext II-RJ). After freezing and thawing, viability of spermatozoa in Ext II -RJ containing 1% RJ immediately after thawing (67.59.6) was significantly lower than that of Ext II , Ext II -RJ containing 0.01 or 0.1% RJ (77.512.5, 78.78.2 and 80.06.3). However, Ext II-RJ containing 0.1% RJ yielded higher viability than Ext II, Ext II-RJ containing 0.01% at or 1% 1 h after thawing (69.58.1 vs. 55.012.9, 57.59.6 and 41.512.6; P<0.05). At 1 h after thawing, the viability of spermatozoa thawed in 7 (68.812.5) was significantly higher than that of spermatozoa thawed in 38 (48.816.3), although there was no difference in the viability between both groups immediately after thawing (77.59.6 and 81.38.1). Post-thaw viability and longevity of post-thaw spermatozoa in Ext II-RJ containing 0.1% RJ was higher in those in Ext II at 1 h (65.012.9 vs. 42.512.6), 2 h (52.512.6 vs. 27.517.1) and 3 h (40.014.1 vs. 20.012.1) after thawing. These results indicated that addition of 0.1% af to Tris-buffer enhanced post-thaw viability and longevity of canine spermatozoa and this additive can be used for increasing the possibility of collision between spermatozoa and ova during insemination.emination.

S-allylcysteine의 항암효과

공일근(Il-Keun Kong),김현희(Hyun Hee Kim),민계식(Gyesik Min) 한국생명과학회 2015 생명과학회지 Vol.25 No.11

S-allylcysteine (SAC)은 숙성된 마늘로부터 유래된 수용성 유기황화합물로서, 여러 유형의 암세포에 대한 항암효과를 갖는 것으로 제시되어왔다. 본 논문은 in vitro 및 in vivo 연구결과에 기초하여 SAC가 세포증식, 세포사멸, 세포주기 및 전이에 미치는 세포신호전달경로와 분자적 메커니즘을 정리하였다. SAC는 Bax와 caspase-3을 포함하는 세포사멸촉진 단백질을 활성화하고 Bcl-2 세포사멸억제 단백질군을 억제하여 미토콘드리아-매개 내인성 경로를 통한 세포사멸을 초래한다. SAC는 또한 PI3K/Akt/mTOR 및 MAPK/ERK 신호전달경로를 억제하여 NF-κB, cyclins, Cdks, PCNA 및 c-Jun의 발현과 활성을 감소시키고, 세포주기 억제단백질인 p16 및 p21의 발현을 증가시킴으로써 세포주기 억제를 유도하여 세포증식을 억제한다. 뿐만 아니라, SAC는 glutathione-s-transferase (GST)와 같은 항산화효소의 활성을 유도하여 독성물질에 의해 유도된 발암작용을 방지한다. 그리고, SAC는 MAPK/ERK 및 PI3K/Akt/mTOR/NF-κB 신호경로의 억제를 통한 전사억제조절인자 Id-1 및 SLUG의 발현억제를 통하여 초래된 COX-2의 발현감소와 E-cadherin의 발현증가에 의해 신생혈관생성과 MET의 억제를 유도함으로써 암세포의 침투와 전이를 억제한다. 따라서, SAC는 암의 예방과 치료를 위한 하나의 잠재적 화학요법제로 간주될 수 있다. S-allylcysteine (SAC) is an aged garlic derived water soluble organosulfur compound and has been suggested to have anticarcinogenic activity against diverse types of cancer cells. This review summarizes the cellular signaling pathways and molecular mechanisms whereby SAC exerts its effects on cellular proliferation, apoptosis, cell cycle progression and metastasis based on the results from both in vitro and in vivo studies. SAC activates proapoptotic proteins including Bax and caspase-3, but suppresses antiapoptotic Bcl-2 family proteins to bring about cancer cell death through mitochondria-mediated intrinsic pathway. SAC also inhibits cellular proliferation by inducing cell cycle arrest in which SAC reduces expression and activation of NF-κB, cyclins, Cdks, PCNA and c-Jun, but elevates expression of cell cycle inhibitor proteins p16 and p21 through suppression of both PI3K/Akt/mTOR and MAPK/ERK signaling pathways. And, SAC inhibits invasion and metastasis of cancer cells by inducing suppression of both angiogenesis and epithelial-mesenchymal transition (EMT) through decreased cyclooxygenase (COX)-2 expression and increased E-cadherin expression which were then caused by suppression of inhibitory transcription factors Id-1 and SLUG from SAC-mediated inactivation of both MAPK/ERK and PI3K/Akt/mTOR/NF-κB signaling pathways. Furthermore, SAC prevents toxic compound-induced carcinogenesis by inducing antioxidant enzymes such as glutathione-s-transferase (GST). Thus, SAC can be considered as a potential chemotherapeutic agent for the prevention and treatment of cancer.

Enhanced tyrosine hydroxylase expression in PC12 cells co-cultured with feline mesenchymal stem cells

공일근,Guang-Zhen Jin,Xi-Jun Yin,Xian-Feng Yu,Su-Jin Cho,Hyo-Sang Lee,이효종 대한수의학회 2007 Journal of Veterinary Science Vol.8 No.4

Mesenchymal stem cells (MSCs) secrete a variety of neuroregulatory molecules, such as nerve growth factor, brain-derived neurotrophic factor, and glial cell-derived neurotrophic factor, which upregulate tyrosine hydroxylase (TH) gene expression in PC12 cells. Enhancing TH gene expression is a critical step for treatment of Parkinson's disease (PD). The objective of this study was to assess the effects of co-culturing PC12 cells with MSCs from feline bone marrow on TH protein expression. We divided the study into three groups: an MSC group, a PC12 cell group, and the combined MSC + PC12 cell group (the co-culture group). All cells were cultured in DMEM-HG medium supplemented with 10% fetal bovine serum for three days. Thereafter, the cells were examined using western blot analysis and immunocytochemistry. In western blots, the co-culture group demonstrated a stronger signal at 60 kDa than the PC12 cell group (p < 0.001). TH was not expressed in the MSC group, either in western blot or immunocytochemistry. Thus, the MSCs of feline bone marrow can up-regulate TH expression in PC12 cells. This implies a new role for MSCs in the neurodegenerative disease process.

개 자궁내 인공수정기에 의한 인공수정 후 산자생산

공일근,조성균,임용택,이상인,위성하 한국임상수의학회 1999 한국임상수의학회지 Vol.16 No.2

This study was conducted to develop an intrauterine inseminator (IUI) to deposit of frozen semen into uterus and to evaluate the results obtained after artificial insemination by IUI. Two Japanese spitzs (2 to 4 years of age) were used as semen donors. Semen was collected by manual masturbation into sterile glass collection tubes and separated into 3 fractions with only the sperm-rich fractions retained for further examination. Sperm motility >70%, sperm concentration of 200 to $400{\times}10^6 cells/ml$</TEX., and up to 15% abnormal and dead spermatozoa were used. Each ejaculate was centrifuged at 400$\times$g for 5 min and poured out the suspended solution, and then diluted with 2 ml Tris-buffer which was consisted of 2.4 g Tris, 1.4 g citric acid, 0.8 g glucose, 0.1 $\mu\textrm{g}$/ml streptomycin, 100 IU/ml penicillin, 20 ml egg yolk to 100 ml mili-Q water (Ext I) or supplemented with 8 ml glycerol and 1 ml Equex STM paste to 100 rnl (Ext II). The diluted semen was cooled to 5$^{\circ}C$ in cold room, where the temperature in the sample reached 5$^{\circ}C$. Two h after beginning the cooling procedure, 2 ml of Ext II, also at 5$^{\circ}C$, was added and mixed by gently reversing the tubes several times during 1 h. The final sperm concentration for freezing was approximately $50{\times}10^6 cells/ml$. After equilibration, the semen was loaded into 0.5 ml straw and frozen on the liquid nitrogen vapour in styrofoam box. The straws were thawed at 7$0^{\circ}C$ for precisely 6 sec. After thawing of each straw, the frozen semen can survived over 50% motility. All the females were inseminated twice with 1 ml of $25{\times}10^6 cells/ml$</TEX. concentration at 48 and 96 h after optimal insemination days estimated by viginal epithelial cells staining. The semen was deposited into uterine body by IUI. The catheter of IUI can passed the cervix and deposited frozen semen in intrauterine body instead of vagina. Four out of five females inseminated were pregnant and three out of four pregnant female were delivered 11 pups. The results obtained indicated that Tris-buffer contained Equex STM paste could be frozen the canine semen successfully, and IUI can also be improved the pregnancy rate and reduced the total semen concentration per AI rather than that of vagina insemination.

한우 체외수정란의 동결보존시 평형시간과 배 발달단계가 생존성에 미치는 영향

공일근,주영국,이은봉,김용권,박충생 韓國受精卵移植學會 1994 한국동물생명공학회지 Vol.9 No.1

The present experiments on cryopreservation were designed to examine the effects of solution toxicity, equilibration time and cell stages on the post-thaw survival of bovine IVF embryos. The oocytes were matured in vitro(IVM) for 24 hrs. in TCM-199 supplemented with 35 g /ml FSH, 10 g /ml LH, 1 g /ml estradiol-17 and granulosa cells at 39 under 5% in air. They were fertilized in vitro(IVF) by epididymal spermatozoa treated with heparin for 24 hrs., and then the zygotes were co-cultured in vitro(IVC) with bovine oviductal epithelial cells for 7 to 9 days. The bovine IVF embryos were exposed to the EFS solution in one step at room temperature, kept in the EFS solution during different period for toxicity test, vitrified in liquid nitrogen, and thawed rapidly. 1. after the bovine blastocysts were exposed to EFS solution for 2 min. at room temperature and then they were washed in 0.5 M sucrose solution and TCM-199, they were cultured to examined cryoprotectant induced injury during exposure, Most of the embryos(95.0%) developed to reexpanded blastocoels. However, when the exposure time was extended to 5 and 10 min, these development rates dropped dramatically in 5 min. (69.5%) and 10 min. (47.4%), respectively, 2. When the bovine IVF embryos were vitrified in EFS solution after the equilibration for 1 and 2 min. exposure, The embryos to have reexpanded blastocoels following thawing, washing and culture processes were found to he 82.6 and 73.9%, respectively. However, when the exposure time was extended to 3 min, this survival rate dropped to 18.2%. The optimal time for equilibration of bovine IVF blastocysts in EFS solution seemed to he 1~2 min. 3. When the bovine IVF embryos were equilibrated for 1 min. the significantly (P<0. 05) higher post-thaw survival rates were obtained from the embryos of blastocyst stage(81.3%) than morulae stage(5. 1%). The optimal cell stage for viterification with EFS solution proven to he blastocyst stage in bovine IVF embryos. 4. The number of blastomeres of blastocyst stage was examined with nuclear staining with Hoechst 33342 during 7 to 9 days post-insemination. The cell counts of frozen bovine IVF embryos were found significantly(P7.5 and those of the fresh embryos 76.67. 1, which were cultured in the sarne period and conditions as frozen embryos.

돼지의 자궁내 인공수정기술개발에 관한 연구

공일근,정금택,이정우,정수룡,오인석,유대중,이효상,김기수,배인휴 韓國受精卵移植學會 2002 한국동물생명공학회지 Vol.17 No.1

본 연구는 2001년 5월과 7월에 순천지역의 한 농장을 대상으로 종모돈 64두와 미국의 SGI회사로부터 직수입된 동결정액을 가지고 인공수정시 정액의 형태와 방법이 종모돈의 번식성적에 미치는 영향을 구명하고자 실시한 바, 그 얻어진 결과는 다음과 같았다. 1. 일반적인 액상정액을 이용한 인공수정과 자궁내 이식기구를 이용한 동결정액의 이용이 번식성적에 미치는 영향을 조사한 결과는, 분만율에서는 액상정액을 이용한 처리구 (86.4%)가 동결정액을 이용한 처 This study was carried out to investigate the possibility of porcine artificial insemination (A·I) on fertilizing capacity using intrauterine inseminator (IUI) method and conventional A·I (CAI) method. Number of sows used in this study was 15 far IUI and 59 fur (CAI), respectively. The results obtained are as fellows: 1 . The frozen and liquid semen used for A·I showed the higher farrowing rate in liquid semen (86.4%) than frozen semen (67%). Number of pigs born per semen type showed the higher values of number of piglets with no statistical significance using frozen semen (9.7) than liquid semen (9.3). 2. The farrowing rate per parity was highest in the 3∼5th parities (100%), f311owe4 by 0∼ 2th parities (60%), and was the smallest in 6 ∼ 10th parities (25%). Number of pigs born per litter was highest in 0∼2th parities (11.3), followed by 3 ∼ 5th parities (9.2) and lowest in 6∼ 10th parities. In the number of pigs bort per litter, the sow s in the high parities delivered lower number of piglets than those in low parities with no significant difference. These results indicated that fertilizing capacity could be improved by using IUI method.

정액종류 및 배양조건에 따른 체외수정란의 생산 및 동결보존의 효율에 미치는 영향

공일근,이상인 韓國受精卵移植學會 1999 한국동물생명공학회지 Vol.14 No.1

The objective of this study was to optimize the selection of sperm sources, optimal culture systems and vitrification method depends on sperm sources. The oocytes were inseminated with either KPN 105, 114, 191, SNU 101, 102, 103 or epididymis and then embryos inseminated were cultured in oviductal cell co-culture or HECM-6 as defined me dium. The blastocysts produced were pooled according to sperm sources as KPN, SNU or epididymis and then vitrified by OPP vitrification method. The results obtained were as follows: 1. The cleavage(86.2 or 84.7%) and development rates to blastocyst (30.6 or 32.0%) were not significantly different between oviductal cell co-culture or HECM-6 culture systems(P<0.05). 2. To determine the optimal sperm sources for using IVF in this system, cleavage rates in KPN 191 and SNU 101 (74.2, 55.8%) were significantly lower rather than those in KPN 105, 114, SNU 102, 103 or epididymis (86.7, 85.1, 89.8, 85.5 or 81.2%), but development rates to blastocyst in KPN 114, SNU 103 or epididymis sperm (30.0, 33.0 or 28.6%) were significantly higher rater than those in KPN 105, 191, SNU 101, 102(21.4, 15.4, 14.9 or 25.4%), respectively (P<0.05). 3. The blastocysts produced were pooled according to sperm sources as KPN, SNU or epididymis and then vitrified by OPP vitrification method. The survival rates were not significantly different among sperm sources (89.6%: 43/48 ; 90.1%: 46/51 ; 83.3% : 20/24). These results obtained indicate that the defined medium, HECM-6, could be use to produce of IVP bovine embryos. Since the frozen semen must be required to maintain of unvariation data in IVP embryo production system, KPN 114 and SNU 103 produced in our laboratory were useful for this purpose. The blastocysts produced by different sperm sources as KPN, SNU or epididymis were vitrified by OPP vitrification method and survived very high rates. The OPP vitrification method could be susceptibility to use of IVP bovine blastocyst embryos.

생쥐 수정란의 유리화 동결보존에 있어서 동결전 처리방법에 관한 연구

공일근,이은봉,강대진,박충생,Gong, Il-Geun,Lee, Eun-Bong,Gang, Dae-Jin,Park, Chung-Saeng 한국동물번식학회 1991 Reproductive & developmental biology Vol.15 No.1

This study was carried out to investigate the effect of equilibration time, precooling and straw loading method on the post-thaw survival rates of mouse embryos cryopreserved by vitrification method. The effect of the vitrification procedure on the damage of the embryos was also examined by the straining of nuclei using Hoechst 33342. The results obtained were summarized as follows; 1. The equilibration in Medium-1 for 10 minutes was considered to be the optimum equilibration time. Embryos equilibrated in Medium-1 for 10 minutes(81.0%) showed significantly(P<0.05) higher survival rates than those equilibrated for 5 minutes(40.0%) or 15 minutes(74.1%). 2. The survival rate of embryos cryopreserved using the double Medium-2 column(81.0%) was significantly(P<0.01) higher than that using the single Medium-2 column, whish was considered to be due to the double Medium-2 column method being more reliable for preventing contamination of diluent solution of 1M sucrose. 3. The survival rate of compacted morula stage embryos cryopreserved with the precooled and non-precooled Medium-2 was not significantly(P<0.05) different. 4. The number of blastomeres of embryos at late blastocyst stage after culture of mouse morulae for 24 to 28hours was significantly(P<0.05) decreased by freezing embryos using vitrification(53.3${\pm}$1.6 vs 41.4${\pm}$1.5), which was considered to be due to the damage of embryos during vitrification and the delay of embryo development by handling in vitro. 5. The vitrification procedure is considered to be a very simple and efficient method for cryopreservation of embryos at early developmental stage.