• Journal of Animal Science and Technology
• /
• 제48권3호
• /
• pp.353-360
• /
• 2006

본 연구는 쥐 난자에서 인위적 활성화 처리가 난자의 활성화, cyclin B1 단백질의 발현 및 난자의 활성화와 cyclin B1 단백질 발현간의 상관관계에 미치는 영향에 관하여 조사하고자 실시하였다. 난자의 활성화 처리는 7% ethanol(EtOH) or 10μg/ml Ca-ionophore with or without 10μg/ml cycloheximide (CH) 방법으로 단일 또는 복합처리 하였다. 난자의 활성화 비율은 단일처리(p<0.05)와 복합처리 (p<0.01)한 난자가 무처리에 비하여 유의하게 높았다. Cyclin B1 단백질의 발현이 EtOH+CH 처리한 난자를 제외한 다른 처리군에서는 무처리에 비하여 유의하게 감소하였다(p<0.05). 한편 EtOH+CH(r=0.61, p<0.05)와 Ca+CH(r=0.86, p<0.01) 처리그룹에서 cyclin B1 단백질의 발현과 난자의 활성화 간에 높은 역상관관계가 있음을 확인하였다. 하지만 단일처리 그룹에서는 두 요소간에 상관관계가 없음을 알 수 있었다. 따라서 단일(EtOH and Ca-ionophore) 또는 복합(EtOH+CH and Ca+CH) 활성화 처리가 난자의 활성화를 증가시키며, 이것은 난자의 활성화 처리에 따른 cyclin B1 단백질의 감소와 밀접한 연관이 있다고 사료된다.

• Clinical and Experimental Reproductive Medicine
• /
• 제41권3호
• /
• pp.137-139
• /
• 2014

The effect of artificial oocyte activation (AOA) with a calcium ionophore on intracytoplasmic morphologically selected sperm injection (IMSI) was examined in patients with histories of repeated failed implantation attempts. Four singleton pregnancies and one twin pregnancy were obtained after embryos transfer (5/14, 35.7%). Therefore, AOA combined with IMSI can be considered an option for cycles with a fertilization defect and recurrent implantation failures.

• 한국수정란이식학회:학술대회논문집
• /
• 한국수정란이식학회 2004년도 제4회 발생공학 국제심포지움 및 학술대회
• /
• pp.5-6
• /
• 2004

• Clinical and Experimental Reproductive Medicine
• /
• 제42권2호
• /
• pp.45-50
• /
• 2015

Objective: Artificial oocyte activation (AOA) is an effective method to avoid total fertilization failure in human in vitro fertilization-embryo transfer (IVF-ET) cycles. AOA performed using a calcium ionophore can induce calcium oscillation in oocytes and initiate the fertilization process. We evaluated the usefulness of AOA with a calcium ionophore in cases of total fertilization failure in previous cycles and in cases of severe male factor infertility patients with non-motile spermatozoa after pentoxifylline (PF) treatment. Methods: The present study describes 29 intracytoplasmic sperm injection (ICSI)-AOA cycles involving male factor infertility at Cheil General Hospital from January 2006 to June 2013. Patients were divided into two groups (control, n=480; AOA, n=29) depending on whether or not AOA using a calcium ionophore (A23187) was performed after testicular sperm extraction-ICSI (TESE-ICSI). The AOA group was further split into subgroups according to sperm motility after PF treatment: i.e., motile sperm-injected (n=12) and non-motile sperm-injected (n=17) groups (total n=29 cycles). Results: The good embryo rate (52.3% vs. 66.9%), pregnancy rate (20.7% vs. 52.1%), and delivery rate (10.3% vs. 40.8%) were lower in the PF/AOA group than in the control group. When evaluating the effects of restoration of sperm motility after PF treatment on clinical outcomes there was no difference in fertilization rate (66.6% vs. 64.7% in non-motile and motile sperm, respectively), pregnancy rate (17.6% vs. 33.3%), or delivery rate (5.9% vs. 16.7%) between the two groups. Conclusion: We suggest that oocyte activation is a useful method to ensure fertilization in TESE-ICSI cycles regardless of restoration of sperm motility after PF treatment. AOA may be useful in selected patients who have a low fertilization rate or total fertilization failure.

• BMB Reports
• /
• 제52권8호
• /
• pp.482-489
• /
• 2019

Reproductive biotechnology has developed rapidly and is now able to overcome many birth difficulties due to infertility or the transmission of genetic diseases. Here we introduce the next generation of assisted reproductive technologies (ART), such as mitochondrial replacement technique (MRT) or genetic correction in eggs with micromanipulation. Further, we suggest that the transmission of genetic information from somatic cells to subsequent generations without gametes should be useful for people who suffer from infertility or genetic diseases. Pluripotent stem cells (PSCs) can be converted into germ cells such as sperm or oocytes in the laboratory. Notably, germ cells derived from nuclear transfer embryonic stem cells (NT-ESCs) or induced pluripotent stem cells (iPSCs) inherit the full parental genome. The most important issue in this technique is the generation of a haploid chromosome from diploid somatic cells. We hereby examine current science and limitations underpinning these important developments and provide recommendations for moving forward.

• Asian-Australasian Journal of Animal Sciences
• /
• 제14권2호
• /
• pp.174-183
• /
• 2001

During the rapid phase of gonadal development of the freshwater teleost, the catfish (Silurus asotus), the influence of hCG upon the inducement of final oocyte maturation and spawning was investigated electrophoretically and ultrastructurally. The electrophoretic patterns obtained were different in the presence and absence of some of the major or minor zones, because of the hormone level in catfish. The vitellogenin of hormone-treated fish was stained more intensively than that of sham-treated fish. These proteins showed some minor or main bands of egg extracts which migrated at positions corresponding to molecular weights of approximately 90,000. However, the thickness of electrophoretic band in molecular weight for hCG-treated fish was slightly lower than that for saline control. It seemed the plasma protein with molecular weight of approximately 45,000 in hCG-treated fish disappeared. In contrast to the control fish, the ovaries in the catfish treated with hCG shows a marked ultrastructural change under the electron microscope. No dilated profiles were seen in the granulosa cells of the mature oocyte before ovulation. After germinal vesicle breakdown (GVBD), the zona radiata interna (ZRI) becomes more compact, and there is a loss of all the processes from the pore canals. There is a wide space between the vitelline membrane and zona radiata. Also, during final maturation, the microvillar processes from the oocyte are seen no longer to penetrate deeply into the extracellular spaces of the overlying granulosa cells, and the reticulate patterns of the zona radiata interna becomes occluded, giving the zona radiata a more solid appearance. It has been possible to initiate 100% oocyte maturation in yolk granules and follicles in vivo by treatment with hCG and a high water temperature ($27^{\circ}C$). In hCG-treated fish, the percentages of successful artificial fertilization and hatching were maximal at 15 h after a single injection. It seems clear that a long acting preparation containing hCG can be successfully used in prespawning fish to advance the final events of gonadal maturation and initiate spawning. Further studies are necessary to evaluate the potential of hCG to either stimulate or inhibit the reproductive development of fish at other stages of the seasonal reproductive cycle.

• Applied Microscopy
• /
• 제30권1호
• /
• pp.61-72
• /
• 2000

U. unicinctus 난자를 대상으로 인공수정 후 수정막 형성에 따른 물질적 기원과 첨체반응 등에 관련된 구조변화에 대하여 광학현미경과 전자현미경을 이용하여 관찰하였다. 관찰결과에 의하면 1차 수정막의 물질은 과잉의 정자에 대한 효과적 방어 기능을 보유한 난외막 표층 기원의 물질이었으며, 2차 수정막은 피질과립의 분비에 의하여 형성되었다. 3차 수정막은 광학현미경하에서 특이한 구조의 관찰은 없었으나 전자현미경에 의한 관찰시 많은 입자상 물질들로 구성되었으며, 수정 후 60분에는 이들 입자물질의 응집에 의해서 수정막은 경과되는 것으로 관찰되었다. 1차 수정막과 난외막 표층의 유사성 그리고 1차 수정막의 형성과 미세융모의 퇴화가 동시적인 사건이라는 것은 미세융모가 U. unicinctus에서 정자의 수용체로 작용할 수 있는 것으로 추정할 수 있었다. 첨체소관에 의한 난세포막 침투시 침체 단부에서 과립상의 물질이 표층(surface coat)에 확산되었다. 첨체소관은 첨체의 기부에서 시작되고, 첨체간격을 통하여 신장되었다.

• 한국수산과학회지
• /
• 제50권1호
• /
• pp.41-47
• /
• 2017

Perhaps the most common type of reproductive dysfunction in captive fish is failure of females to undergo final oocyte maturation and thus to ovulate and spawn. The success of aquaculture could therefore be improved by developing techniques to enhance natural spawning, artificial maturation, and/or to induce ovulation in farmed fish. This study aimed to investigate the effects of prostaglandin $E_2$ ($PGE_2$) and prostaglandin $F_{2{\alpha}}$ ($PGF_{2{\alpha}}$) on in vitro oocyte maturation (germinal vesicle breakdown, GVBD) and ovulation in the marine fish Chasmichthys dolichognathus. Post-vitellogenic follicles (0.80-0.94 mm diameter oocytes) were incubated with $PGE_2$ or $PGF_{2{\alpha}}$ at concentrations of 5, 50, or 500 ng/mL for 24 hours. A significant increase in GVBD was seen in 0.84 mm and 0.94 mm oocytes incubated with 50 ng/mL $PGE_2$ compared with the control. There was no significant increase in GVBD in any of the other experimental conditions (5 or 500 ng/mL $PGE_2$ or 5, 50, or 500 ng/mL $PGF_{2{\alpha}}$). Neither of the prostaglandins induced ovulation at the concentrations tested.These results suggest that GVBD was induced by incubation with 50 ng/mL $PGE_2$.

• 한국수정란이식학회지
• /
• 제11권1호
• /
• pp.71-80
• /
• 1996

The envelope of the rnannnalian oocyte plays crucial roles in sperm-oocyte interactions by providing sperm receptors, inducing acrosome reaction and preventing polyspermy. Understanding of properties of the zona pellucida (ZP) is essential for the artificial control of fertility in mammals. This study was carried out to produce and characterize monoclonal antibodies(MAbs) to porcine ZP proteins. Approximately 8,000 ZPs were obtained from follicular oocytes and dissolved in 40$\mu$l of double distilled water. Following immunization through foot-pad injections of Balb /c mice with a ZP solution, the popliteal lymph nodes were recovered at 2 weeks after the last injection. Hybridoma cell lines were established by fusing lymph node cells with P3X63 myeloma cells through selection using HAT medium and screening by immunofluorescence(IF) microscopy on the isolated ZP. Secreted MAbs were found to consist k chains and different heavy chains as evidenced by isotyping. Some of the MAbs demonstrated high specificity to the ZP in IF. The Mabs also showed positive cross reactivity with hamster and mouse eggs, while negative with bovine eggs. The results implicate that the MAbs can be used not only for identification of functional regions of the ZP, but also for elucidation of mechanisms involved in fertilization of mammals. The MAbs will provide basic information on biochemical anatomy of the ZP as well as can be candidates for the future contraceptive vaccines.

• Asian-Australasian Journal of Animal Sciences
• /
• 제17권7호
• /
• pp.885-891
• /
• 2004

To evaluate the feasibility of using sperm-mediated gene transfer (SMGT) for carrying foreign gene into chicken oocyte, a reporter gene, CX-EGFP, was used in this study. The reporter gene was first mixed with liposome or liposome-like compound and the mixtures were further combined with ejaculated cock spermatozoa. The spermatozoa treated with liposome and CX-EGFP mixture was subsequently coincubated with DNaseI to remove the extra DNA which insured the authenticity of positive signals. The treated sperms were then subjected to transgene (reporter gene) existence analysis and artificial insemination of laying hens. Obtained results indicated that the spermatozoa were able to take-in the foreign DNA; which was confirmed by polymerase chain reaction and Southern blot analysis. In the following experiment, fresh ejaculated sperms were mixed with CX-EGFP-liposome or CX-EGFP-liposome-like complex then used for artificial insemination of each of six laying hens. Eggs laid between day-3 and day-7 post insemination were collected. Newly hatched chicks, two out of 53 from CX-EGFP/liposome treated group and two out of 21 from CXEGFP/liposome-like treated group, were proven to be transgenic. This study suggests that SMGT is a powerful method for generating transgenic chickens.