• Korean Journal of Animal Reproduction
• /
• v.21 no.3
• /
• pp.293-302
• /
• 1997

Follicular oocytes of Grade I and II were collected from 2~6 mm ovarian follicles and matured in vitro (IVM) for 24 hrs in TCM-199 su, pp.emented with 35$\mu\textrm{g}$/ml FSH, 10$\mu\textrm{g}$/ml LH, and 1$\mu\textrm{g}$/ml estradiol-17$\beta$ at 39$^{\circ}C$ under 5% CO2 in air. They were fretilized in vitro (IVF) by epididymal spermatozoa capacitated with heparin for 12 hrs. The zygotes were then co-cultured in vitro with bovine oviducted epithelial cells (BOEC) for 7 to 9 days. The optimal time for IVM, the successful enucleation of IVM oocytes by micromanipulation at different oocyte ages after IVM, and the ideal culture system for IVM for effective IVF and in vitro development of IVM-IVF embryos was examined for in vitro production of nuclear recipient oocytes and nuclear donor embryos. To improve the efficiency of nuclear transplantation (NT) of IVF embryo into IVM follicular oocytes, this study evaluated the optimal electric condition and oocytes age for activation of IVM oocytes and in vitro development of NT embryos. In vitro development of NT embryos with preactivation or non-preactivation in enucleation oocytes, cell number of IVN-IVF embryos, and NT embryos wre also examined. The results obtained were as follows; 1. The most suitable enucleation time was at 24 hpm (83.3%) rather than that of 28 hpm(69.6%) and 32 hpm(50.0%). 2. There was no difference among the fusion rates of NT embryos at the voltages of 0.75, 1.0 and 1.5 kV/cm, but the in vitro development rates to morule and blastocyst were significantly (P<0.05) higher at the voltage of 0.75(12.5%) and 1.0kV/cm (12.6%) compared to 1.5kV/cm(0%). 3. No significant difference in activation rates were seen in NT embryos stimulated for 30, 60 and 120 $\mu$sec (71.7, 85.2 and 71.9%, respectively), but the in vitro development rates to morulae and blastocyst were significantly (P<0.05) higher in the oocytes stimulated for 30 $\mu$sec (11.6%) and 60 $\mu$sec(10.7%) than 120 $\mu$sec(0.0%). 4. The fusion rates (71.0 and 87.3%) and the in vitro development rates (9.1 and 12.7%) to morula and blastocyst were seen in the NT embryos stimulated at 28 and 32 hpm under the condition of 1.0 kV/ml, 60 $\mu$sec. However, at 24 hpm the fusion rates were 64.8% and the in vitro development to morula and blastocyst were not seen. 5. The fusion rates between the 8~12, 13~17 and 18~22-cell stage of IVM-IVF embryos were not significantly different. The in vitro development rates of the fused embryos to morula and blastocyst which were received from a blastomere of 8~12, 13~17 and 18~22-cell stages of IVM-IVF embryos were 14.9, 8.3 and 6.5%, respectively. 6. The in vitro development rate of the enucleated recipient oocytes with preactivation (24.2%) to morula and blastocyst was significantly (P<0.05) higher than that of non-preactivation (12.8%). 7. The cell numbers of NT blastocyst and IVM-IVF blastocyst cultured during 7~9 days were 63$\pm$11 and 119$\pm$23, and then their the mean cell cycle number were 5.98 and 6.89, respectively.

• Clinical and Experimental Reproductive Medicine
• /
• v.13 no.1
• /
• pp.11-19
• /
• 1986

Follicular flxid (FF) and their matched oocytes were obtained from 58 follicles of 27 women who underwent an in vitro fertilization (IVF) procedure with ovarian hyperstimulation by clomiphene citrate(n=8), hMG(n=9),FSH/hMG(n=10). Follicular aspiration was performed 36 hours after human chorionic gonadotropin administration. The concentcation of androstendione (ADD), testosterone (T) was correlated with hyperstimulation regimens, the morphology of the oocyte-corona-cumulus complex (OCCC), oocyte fertilization, and the incidence of pregnancy after embryo transfer. The results were as follows. 1. According to hyperstimulation regimens, there was no significant differance in FF ADD and T concentrations of the similar morphology of OCCC. 2. In clomiphene-treated and FSH/hMG-treated cycles, FF ADD concentrations of preovulatory oocytes were 43.09${\pm}$9.53 ng/ml and 59.46${\pm}$9.09 ng/ml, those of immature occytes were 96.98${\pm}$16.55 ng/ml and 116.13${\pm}$36.81 ng/ml, those of atretic oocytes were 246.5 ${\pm}$9.25 ng/ml and 634.25${\pm}$9.25 ng/ml respectively, reflecting the significant relationship between FF ADD level and morphologic maturity of OCCC (p<0.05). But in hMG-treated cycles, such relationship was not found (p>0.1). In clomiphene-treated and FSH/hMG-treated cycles, FF T concentrations of preovulatory oocytes were 11.37${\pm}$2.38 ng/ml and 11.68${\pm}$1.73 ng/ml respectively which were significantly lower than those of atretic oocytes (25.1${\pm}$7.50 ng/ml and 23.25${\pm}$0.95 ng/ml respectively) (p<0.05). But in all cycles, FF T concentrations of immature oocytes were not significantly different from those of preovulatory oocytes, artetic oocytes (p>0.1). 3. In hMG-treated and FSH/hMG-treated cycles, FF ADD concentrations of fertilized oocytes were 32.43${\pm}$4.09 ng/ml and 42.61${\pm}$4.82 ng/ml respectively which were significantly lower than those of non-fertilized oocytes (72.18${\pm}$17.31 ng/ml and 108.09${\pm}$17.32 ng/ml respectively) (p<0.05), but in clomiphene-treated cycles there was no significant difference (p>0.1). In FSH/hMG-treated cycles, FF T concentration of fertilized oocytes was 7.33${\pm}$1.06 ng/ml which was significantly lower than that of non-fertilized oocytes (20.3${\pm}$6.21 ng/ml) (p>0.02), but in clomiphne-treated and hMG-treated cycles there was no significant difference (p>0.1). 4. In all cycles FF ADD and T concentrations did not correlated with the success of pregnancy after embryo transfer. Above results suggested that FF ADD and T may play an important role in oocyte maturation and fertilization, but their relationship with the success of psegnancy was not found.

• Korean Journal of Animal Reproduction
• /
• v.6 no.1
• /
• pp.19-30
• /
• 1982

1. The cows slaughtered at age of 3, 4, 6, 7, 8, and 9 years old were 1.5, 1.5, 15.0, 62.5 and 4.4% respectively. 2. The cows slaughtered at 351-450kg and more than 500kg were 60 and 28% respectively. 3. Best, very good, good and bad cows in nutritional condition were 1.6, 25.8, 62.9, and 9.7% respectively. Among the six cows which were bad nutrition, the two were with severe endometritis, the three were normal in genital function and one was on 70 days of pregnancy. 4. Holstein cows(55.2%) showed higher reproductive failure than the Korean cows(33.3%). 5. The slaughted ratio of the Korean cattle and Holstein cows was 36 and 64% respectively. 6. Pregnant cows were about 16% among the slaughtered one. 7. Reproductive failures were composed of 46% in uterus, 32% in ovaries, 8% in udder, 6% in oviduct, 4% in cervix of uterine, 2% in vagina and 2% inmummified fetus. 8. Forty six percentages of uterine diseases were as follows; horn, 13%, body of uterus, 32% and ovary diseases were 32%, that is, 12% of ovary atrophy, 8% of ovarycyst and 6% of lutealcyst. 9. The cows of reproductive failures were commonly infected with 1.6 kinds of diseases. 10. According to classification, six type of ovaries were as follows; normal, 58%, ovary-cyst, 11%, luteum cyst, 4%, coexistence of follicles and corpus luteum, 16%, weak function of ovaries, 10% and ovarian atrophy, 1%. 11. Major axis, minor axis and thickness of right ovary were larger than those of left one both in Korean cattle and Holstein cows. Holstein cow had generally larger size of ovary than these of the Korean cattle.. 12. The left and right oviducts showed no difference in length, but Holstein had longer oviduct than Korean cow. 13. There was no difference in the length of uterine horn between right and left in the Korean cows, but the right was longer than the left in Holstein cows. 14. Holstein had longer horn and body of uterine than the Korean cows. 15. The weight of right ovary was heavier than that of left in both breeds, but there was no differences in weight of left ovary between two breeds and right ovary of Holstein breed was heavier than that of the Korean cow. 16. The weight of right oviduct and uterine born was heavier than that of the left, and Holstein had heavier oviducts and uterine horns than the Korean cows. 17. Holstein had heavier uterine body and cervix of uterine than the Korean cows. 18. The length of reproductive systems of Korean cow is as follows; Major and minor diameter and thickness ofovary are 3.6${\pm}$0.7, 2.3${\pm}$0.4 and 1.6${\pm}$1.4 cm in left and 3.7${\pm}$0.6, 2.5${\pm}$0.5 and 1.8${\pm}$0.5 cm in right. Oviduct is 28.4${\pm}$3.1 cm in left and 27.8${\pm}$3.3 cm in right. Uterine horn is 27.4${\pm}$4.5 cm in left and 27.7${\pm}$4.9 cm in right. Uterine body and cervix are 3.4${\pm}$1.1 and 6.5${\pm}$1.7 cm. 19. The length of female reproductive systems ofHolstein cow is as follows; Major and minor diameter and thickness of ovary are 3.9${\pm}$1.3, 2.3${\pm}$0.5, and 1.5${\pm}$0.6 cm in left and 4.0${\pm}$0.8, 2.8${\pm}$0.6 and 1.8${\pm}$0.6 cm in right. Oviduct is 29.4${\pm}$4.2 cm in left and 29.3${\pm}$4.1 cm in right. Uterine horn is 30.2${\pm}$7.4 cm in left and 32.6${\pm}$8.4 cm in right. Uterine body and cervix are 4.5${\pm}$2.5 and 7.8${\pm}$2.9 cm. 20. The weight of reproductive systems of Korean cow is as follows; Ovary is 8.4${\pm}$4.1 g in left and 9.3${\pm}$3.6g in right. Oviduct is 1.5${\pm}$0.5 g in left and 1.6${\pm}$0.5 g in right. Uterine horn is 109${\pm}$27 g left and 118${\pm}$32 g in right. Uterine body and cervix are 30.4${\pm}$14.1 and 76.7${\pm}$38.4g. 21. The weight of reproductive systems of Holstein cow is as follows; Ovary is 8.2${\pm}$3.1 g in left and 12.5${\pm}$5.6 g in right. Oviduct is 1.7${\pm}$0.6 g in left and 1.9${\pm}$0.9 g in right. Uterine horn is 199${\pm}$14.2 g in left and 221${\pm}$111.2g in right. Uterine body and cervix are 58.2${\pm}$46.5 and 126.7${\pm}$103.3 g.

• Korean Journal of Fisheries and Aquatic Sciences
• /
• v.31 no.4
• /
• pp.599-607
• /
• 1998

To clarify annual reproductive cycle of Korean dark sleeper, Odontobutis platycephala (Iwata et Jeon), we examined the seasonal changes of gonadosomatic index (GSI), the proportional frequency of oocyte development stages in the ovary and the changes of sex steroid hormone levels in blood from December 1995 to November 1997. In July and August, GSI was 0.35 to 0.72 and most oocytes in the ovary were chromatin-nucleolus stage and perinucleolar stage (proportional frequency: $87\%\~96\%$). In September, GSI was 1.20 $\pm$ 0.12, some oocytes in the ovary were yolk vesifle stage (proportional frequency: $22.8\%$) and vitellogenic stage which appeared very rarely(proportional frequency: $2.2\%$). GSI increased gradually from October and reached 4.59± 0.61 to December. During this period, oocytes of vitellogenic stage increased slightly (proportional frequency in December: $22.1\%$). In January, GSI was 4.32 $\pm$ 0.72 but the proportional frequency of oocytes in vitellogenic stage increased (proportional frequency: $51.2\%$). from February, GSI was increased sharply and reached to 10.51 $\pm$ 1.04 in March, the highest value throughout the year and the proportional frequency of oocytes in vitellogenic stage also reached the highest levels (proportional frequency: $60\%$). From April, GSI was gradually decreased and fell down to 1.11 $\pm$ 0.35 in June. During this period, the proportional frequency of mature oocytes was the highest in April (proportional frequency of mature oocyte stage: $40\%$ in April, $12\%$ May, $5\%$ June) throughout the year, and atretic ovarian follicles were appeared. The blood level of estradiol-17$\beta$ ($E_2$), which stimulates the hepatic synthesis and secretion of vitellogenin, was $0.84{\pm}0.20\;ng/m{\ell}$ in August, and thereafter was not changed until December. from January, it increased sharply and reached the highest level of $ 2.85{\pm}0.35\;ng/m{\ell}$ in March throughout the year, but fell to $0.14{\pm}0.02\;ng/m{\ell}$ in July(P<0.05), 17$\alpha$-hydroxprogesterone(17$\alpha$-OHP) was the peak $13.37{\pm}0.52ng/m{\ell}$ in March, but no significant changes in other period(below $3ng/m{\ell}$, P<0.05). 17$\alpha$, 20$\beta$-dihydroxy-4-pregnen-3-one(17$\alpha$, 20$\beta$-P), which was known as the final maturation inducing hormone in teleost, was $0.74{\pm}0.09ng/m{\ell}$ in April and $0.54{\pm}0.07ng/m{\ell}$ in May, but no significant changes in other period (below $0.26\;ng/m{\ell}$, p<0.05). Taken together these results, the annual reproductive cycle of O. platycephala divided into 4 periods as follows: 1) ripe and spawning period from April to June, main spawning period was from April to May, 2) Resting period from July to August, 3) Growing period from September to December, 4) Maturing period from January to March. Moreover, It was showed that the changes of sex steroid hormone in blood played a important roles in the annual reproductive cycle of O. platycephala.

• Clinical and Experimental Reproductive Medicine
• /
• v.23 no.3
• /
• pp.247-268
• /
• 1996

It has been known for a long time that gonadotropins and steroid hormones play a pivotal role in a series of reproductive biological phenomena including the maturation of ovarian follicles and oocytes, ovulation and implantation, maintenance of pregnancy and fetal growth & development, parturition and mammary development and lactation. Recent investigations, however, have elucidated that in addition to these classic hormones, multiple growth factors also are involved in these phenomena. Most growth factors in reproductive organs mediate the actions of gonadotropins and steroid hormones or synergize with them in an autocrine/paracrine manner. The insulin-like growth factor(IGF) system, which is one of the most actively investigated areas lately in the reproductive organs, has been found to have important roles in a wide gamut of reproductive phenomena. In the present communication, published literature pertaining to the intrauterine IGF system will be reviewed preceded by general information of the IGF system. The IGF family comprises of IGF-I & IGF-II ligands, two types of IGF receptors and six classes of IGF-binding proteins(IGFBPs) that are known to date. IGF-I and IGF-II peptides, which are structurally homologous to proinsulin, possess the insulin-like activity including the stimulatory effect of glucose and amino acid transport. Besides, IGFs as mitogens stimulate cell division, and also play a role in cellular differentiation and functions in a variety of cell lines. IGFs are expressed mainly in the liver and messenchymal cells, and act on almost all types of tissues in an autocrine/paracrine as well as endocrine mode. There are two types of IGF receptors. Type I IGF receptors, which are tyrosine kinase receptors having high-affinity for IGF-I and IGF-II, mediate almost all the IGF actions that are described above. Type II IGF receptors or IGF-II/mannose-6-phosphate receptors have two distinct binding sites; the IGF-II binding site exhibits a high affinity only for IGF-II. The principal role of the type II IGF receptor is to destroy IGF-II by targeting the ligand to the lysosome. IGFs in biological fluids are mostly bound to IGFBP. IGFBPs, in general, are IGF storage/carrier proteins or modulators of IGF actions; however, as for distinct roles for individual IGFBPs, only limited information is available. IGFBPs inhibit IGF actions under most in vitro situations, seemingly because affinities of IGFBPs for IGFs are greater than those of IGF receptors. How IGF is released from IGFBP to reach IGF receptors is not known; however, various IGFBP protease activities that are present in blood and interstitial fluids are believed to play an important role in the process of IGF release from the IGFBP. According to latest reports, there is evidence that under certain in vitro circumstances, IGFBP-1, -3, -5 have their own biological activities independent of the IGF. This may add another dimension of complexity of the already complicated IGF system. Messenger ribonucleic acids and proteins of the IGF family members are expressed in the uterine tissue and conceptus of the primates, rodents and farm animals to play important roles in growth and development of the uterus and fetus. Expression of the uterine IGF system is regulated by gonadal hormones and local regulatory substances with temporal and spatial specificities. Locally expressed IGFs and IGFBPs act on the uterine tissue in an autocrine/paracrine manner, or are secreted into the uterine lumen to participate in conceptus growth and development. Conceptus also expresses the IGF system beginning from the peri-implantation period. When an IGF family member is expressed in the conceptus, however, is determined by the presence or absence of maternally inherited mRNAs, genetic programming of the conceptus itself and an interaction with the maternal tissue. The site of IGF action also follows temporal (physiological status) and spatial specificities. These facts that expression of the IGF system is temporally and spatially regulated support indirectly a hypothesis that IGFs play a role in conceptus growth and development. Uterine and conceptus-derived IGFs stimulate cell division and differentiation, glucose and amino acid transport, general protein synthesis and the biosynthesis of mammotropic hormones including placental lactogen and prolactin, and also play a role in steroidogenesis. The suggested role for IGFs in conceptus growth and development has been proven by the result of IGF-I, IGF-II or IGF receptor gene disruption(targeting) of murine embryos by the homologous recombination technique. Mice carrying a null mutation for IGF-I and/or IGF-II or type I IGF receptor undergo delayed prenatal and postnatal growth and development with 30-60% normal weights at birth. Moreover, mice lacking the type I IGF receptor or IGF-I plus IGF-II die soon after birth. Intrauterine IGFBPs generally are believed to sequester IGF ligands within the uterus or to play a role of negative regulators of IGF actions by inhibiting IGF binding to cognate receptors. However, when it is taken into account that IGFBP-1 is expressed and secreted in primate uteri in amounts assessedly far exceeding those of local IGFs and that IGFBP-1 is one of the major secretory proteins of the primate decidua, the possibility that this IGFBP may have its own biological activity independent of IGF cannot be excluded. Evidently, elucidating the exact role of each IGFBP is an essential step into understanding the whole IGF system. As such, further research in this area is awaited with a lot of anticipation and attention.