• Clinical and Experimental Reproductive Medicine
• /
• v.47 no.3
• /
• pp.180-185
• /
• 2020

Objective: Tyrosine phosphorylation is an essential process in many biological systems, including the male reproductive system. The presence of tyrosine-phosphorylated (TyrPho) proteins has been well documented in male reproductive organs, but research in fertile females is still limited. Methods: The ovary, oviduct, and uterus of adult female Sprague-Dawley rats in the estrus phase were used to localize TyrPho proteins using an immunohistochemical technique. These proteins were separated and their expression patterns were examined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blot analysis, respectively. Results: TyrPho proteins were localized in the cytoplasm of the oocyte except the antral fluid; in the granulosa cells, theca cells, and stromal cells of the ovary; at the apical surface of oviductal epithelial cells; and in the basal epithelium and submucosa of the uterine wall. Moreover, we found that 72-, 43-, and 28-kDa TyrPho proteins were localized in the ovary, while 170-, 55-, and 43-kDa proteins were localized in the oviduct. In the uterus, we detected four major bands, corresponding to 61-, 55-, 54-, and 43-kDa TyrPho proteins. Conclusion: Given that these TyrPho proteins were found in major reproductive organs in the estrus phase, these proteins may play important roles in female fertility.

• Journal of Animal Reproduction and Biotechnology
• /
• v.38 no.2
• /
• pp.54-61
• /
• 2023

Background: Germ cells undergo towards male or female pathways to produce spermatozoa or oocyte respectively which is essential for sexual reproduction. Mesenchymal stem cells (MSCs) have the potential of trans-differentiation to the multiple cell lineages. Methods: Herein, rat MSCs were isolated from bone marrow and characterized by their morphological features, expression of MSC surface markers, and in vitro differentiation capability. Results: Thereafter, we induced these cells only by retinoic acid supplementation in MSC medium and, could able to show that bone marrow derived MSCs are capable to trans-differentiate into male germ cell-like cells in vitro. We characterized these cells by morphological changes, the expressions of germ cell specific markers by immunophenotyping and molecular biology tools. Further, we quantified these differentiated cells. Conclusions: This study suggests that only Retinoic acid in culture medium could induce bone marrow MSCs to differentiate germ cell-like cells in vitro. This basic method of germ cell generation might be helpful in the prospective applications of this technology.

• Korean Journal of Ichthyology
• /
• v.26 no.3
• /
• pp.179-184
• /
• 2014

A histological study on the egg envelope and oogenesis of Gobiobotia brevibarba (Pisces, Cyprinidae) was carried out by a light microscope and a scanning electron microscope. Various developmental cells appeared in the ovary caught during May 2014, spawning season. For the relative area of oocyte, the ovary consisted of mature stage (74.5%), a vitellogenic stage (yolk granule stage, 16.6% and yolk vesicle stage, 6.6%) and previtellogenic stage (perinucleolus stage 2.2%), which means its spawning season. The cytoplasm of the perinucleolus oocyte is acidic and many nucleoli are located at the inner side of the nuclear membrane. The yolk vesicles, an early vitellogenic stage, has a follicular layer and a zona radiata clearly. Numerous villi, called an egg envelope, begin to form on the zona radiata. The yolk granules, an another vitellogenic stage, proceeds and they show a strong eosinophilic nature. Such yolk granules appeared between the yolk vesicles occupying most cytoplasm, and as the stage proceeds, there are some yolk masses fused with each other. Egg envelope is covered with plenty of villi ($2{\sim}3{\mu}m$ in the length) over the entire egg surface.

• Journal of Embryo Transfer
• /
• v.15 no.3
• /
• pp.295-302
• /
• 2000

The objective of this study was to compare different superovulation treatments using PMSG or PG600$^{ }$ and to determine the optimal time of oocyte recovery after hCG administration. A total of 90 prepubertal Yorkshire x Landrace gilts crossed with Duroc, 6~7 months old and 100~120 kg of body weight, were used. PMSG (1,500 IU/head) or 5~7.5 ml of PG600$^{ }$(400 IU of PMSG and 200 IU of hCG) were administrated subcutaneously, and then 1,000 IU of hCG were administered intramuscularly at 72 hours after PMSG or PG600$^{ }$ injection. At carious time of 44, 46, 48 and 50 hours after hCG injection, superovulated gilts were slaughtered in a local abattoir. Ovaries together with oviducts were excised from the body immediately after slaughtered and transported to laboratory in 39$^{\circ}C$ saline. Ovaries were examined fur the number of corpus hemorrhagicum and unovulated follicles present in the surface of ovary. The unovulated follicles were categorized into small (1~3 mm in diameter) and large (4~8 mm) groups according to their diameter. Oocytes were recovered by flushing both oviducts with micropipette tip (1~100 $\mu$l) attached to a 10-ml disposable syringe. The number of CH on ovary and recovered oocytes at 46, 48 and 50 hr after hCG injection in PG600$^{ }$ treated groups were significantly higher than the other group. Group of phCG 50 hr among PMSG treated groups had a greater number of CH and recovered oocytes(P<0.05). The number of CH on ovary and recovered oocytes at 50 hr after hCG injection in 1$\frac{1}{2}$ vial(7.5 ml) of PG600$^{ }$ treated groups was significantly higher than 1 vial(5 ml) of PG600$^{ }$ treated group(P<0.05). In conclusions, considering a number of corpus hemorrhagicum and recovered oocytes after superovulation in gilts, effective time of oocyte recovery by treatment with PMSG and hCG was post-hCG 50 hr and with PG600$^{ }$ plus hCG was post-hCG 46, 48 and 50 hr. Also, admini-stration of 1$\frac{1}{2}$ vial(7.5 ml) of PG600$^{ }$ treated group had a great number of CH and recovered oocytes.covered oocytes.

• Asian-Australasian Journal of Animal Sciences
• /
• v.17 no.5
• /
• pp.612-616
• /
• 2004

This study was carried out to compare the semen characteristics, frozen-thawed sperm viability and testosterone concentration and in vitro fertilization (IVF) and development of in vitro matured pig oocytes between two Yorkshire boars. Semen and blood samples were collected once per week from October to November 2002 from two adult Yorkshire boars at 18 months of age with 170 kg body weight. Sperm were deep frozen in 5 ml maxi-straws with lactose-egg yolk and N-acetyl-D-glucosamine (LEN) diluent and stored in liquid nitrogen. Blood samples were obtained at 10 a.m. by inserting a 21 gauge, hypodermic needle attached to 10 ml syringe into surface veins in the ear. The concentration of testosterone was determined by Competitive Enzyme Immunoassay. Ovaries were collected from prepubertal gilts at a local slaughter house. Cumulus oocyte complexes were aspirated from antral follicles (3 to 6 mm in diameter). The medium used for oocyte maturation was modified TCM 199. After about 22 h of culture, oocytes were cultured without cysteamine and hormones for 22 h at $38.5^{\circ}C$, 5% $CO_2$ in air. For IVF, one frozen 5 ml straw was thawed at $52^{\circ}C$in 40 sec and was diluted with 20 ml Beltsville thawing solution at room temperature. Sperm were washed 2 times in mTLP-PVA and inseminated without preincubation after thawing. Oocytes were inseminated with $2{\times}10^7$/ml sperm concentration. Oocytes were coincubated for 6 h in 500 ${\mu}$l mTBM fertilization medium. At 6 h after IVF, oocytes were transferred into 500 ${\mu}$l NCSU-23 culture medium for further culture of 48 and 144 h. There were no significant differences in the semen volume, motility, normal acrosome morphology and sperm concentration of raw semen between A and B of Yorkshire boar. However, motility and normal acrosome of boar A were higher than those of boar B at 0.5, 2, 3, 4, 5 and 6 h incubations of frozen-thawed sperm. Testosterone concentration (3.75 ng/ml) of boar A was higher than that (2.34 ng/ml) of boar B. The rate of blastocyst formation (15.1%) of boar A was higher than that (10.4%) of boar B. In conclusion, serum testosterone concentration of boar showed very important role for the frozen-thawed sperm viability and the blastocyst formation of pig oocytes matured in vitro.

• Korean Journal of Fisheries and Aquatic Sciences
• /
• v.37 no.5
• /
• pp.385-392
• /
• 2004

Gonadal development, gametogenesis, reproductive cycle, gonad index, and flesh weight rate of the murex shell (Ocenebra japonica) collected from the rocky intertidal zone of Buan-gun, Jeollabuk-do, Korea were investigated by means of histological method from January to December 2002. O. japonica had separate sexes, and was oviparous. The gonad was widely situated on the surface of the digestive gland located in the rear of the spiral flesh part in the shell. The male penis was located near the two tentacles. The ovary was composed of a number of oogenic follicles, and the testis was composed of several spermatogenic tubules. The size of ripe oocyte was approximately $140{\mu}m$ in diameter. The gonad index (GI) began to increase in March $(33.24{\pm}2.33)$ and reached the maximum in June $(47.77{\pm}1.90)$ Thereafter, the values decreased from July $(45.12{\pm}3.60)$ to October $(19.32{\pm}2.91)$. The flesh weight rate (FWR) began to increase in January $(25.93{\pm}1.32)$ and reached the maxium in May $(31.78{\pm}1.09)$ Thereafter, the values decreased from June $(31.50{\pm}0.66)$ to October $(24.09{\pm}1.60)$. The reproductive cycle could be classified into five successive stages: early active (October to April), late active (January to June), ripe (May to September), spawning (July to September) and recovery (September to February). The reproductive cycle was closely related to the seawater temperature.

• Korean Journal of Animal Reproduction
• /
• v.17 no.4
• /
• pp.339-346
• /
• 1994

An understanding of the structure and function of mammalian spermatozoa requires the iso-lation of these components. In this study, frozen-thawed bovine spermatozoa were treated by physical treatments (vortexing, 26 gauge needle, strained 26 gauge needles and freezing-thawing) or chemical treatments (trypsin, dithiothreitol, sodium dodecylsulfate and $\beta$-mercaptoethanoJ) to yield free heads and tails. The most effective treatment was repeated pumping of sperm suspension through a strained 26 gauge needle conneted to a syringe. Spermatozoa by this treatment were mainly broken at the junction of the head and the tail, resulting in 90-100% yields. Also, sperm head surface did not modify during strained 26 gauge needle treatment when either spermatozoa or sperm heads were incubated in 250${\mu}\textrm{g}$/ml of FITC-UEA 1 for 1 h at room temperature to detect the modification of sperm surface components. Other physical treatments were less efficient for the breakdown of spermatozoa. The effects of chemical treatments on bovine spermatozoa are not noticeable. Dissected sperm heads and tails should be fractional leading to nearly pure components by sucrose gradient centrifugation at 1,000 rpm for 15 min. The result suggest that the established method may be useful for the biochemical study of spermatozoal components, and the understanding of oocyte activation mechanism either by spermatozoal components during fertilization or microinjection of isolated components.

• Korean Journal of Fisheries and Aquatic Sciences
• /
• v.30 no.4
• /
• pp.627-633
• /
• 1997

Gonad structure, germ cell development and reproductive cycle of the smallmouth scorpionfish, Scorpaena miostoma were investigated based on histological method. Samples were collected monthly in the vicinity of Suyoung Bay, Pusan, Korea from November 1995 to October 1996. The testis is seminiferous tubule type in internal structure. Seminiferous tubule consists of numerous testicular cysts which contain numerous germ cells in same developmental stage. The ovary consists of several ovarian lamellae originated from ovarian outer membrane. Oogonia originated from the inner surface of the ovarian lamella protrude to the ovarian cavity in oocyte stage, and they are suspended by the egg stalk. Biological minimum size of female and male were 12.5cm in total length. Gonadosomatic index (GSI) of female (3.81) and male (0.23) were the highest in October. Reproductive cycle was classified into the following successive stages: in female, growing stage $(May\~August)$, maturation stage $(September\~October)$, ripe and spawning stage $(November\~December)$, recovery and resting stage $(January\~April)$, and in male, growing stage $(June\~August)$, maturation stage $(September\~October)$, ripe and spent stage $(November\~January)$ and recovery and resting stage $(February\~May)$.

• Applied Microscopy
• /
• v.37 no.2
• /
• pp.65-72
• /
• 2007

The oogenesis and ultrastructure of fertilized egg envelope of false dace were investigated by light and electron microscope. The cytoplasm of false dace oogonia was basophilic and many nucleoli were located at inner side of nuclear membrane. In primary oocytes, yolk vesicles were distributed in marginal area only and egg envelope was not formed on egg outside. In secondary oocyte, the egg envelope was formed and yolk vesicles were increased than that of early stage in cytoplasm. The amount of basophilic substance was decreased. In case of matured egg, thickness of egg envelope and site of egg were increased, basophilic substance was distributed in egg envelope around only. The yolk vesicles were changed to yolk mass in accordance with development. The fertilized egg was of ellipsoidal, adhesive type and yellowish, have a single micropyle in the area of the animal pole. The fertilized egg envelope consisted of three layers, an outer adhesive layer, a middle layer consisting of 6 lamellae alternating layers and an inner electron dense layer. An outer surface of the fertilized egg envelope was arranged by adhesive fibrous structures. In conclusion, it is summarized that the oogenesis of false dace were the increase of cell size, the formation and accumulation of yolk, and decrease of basophilic intensity in cytoplasm. These ultrastructural characteristics of fertilized egg envelope from false dace can be utilized in taxonomy of teleost.

• Korean Journal of Fisheries and Aquatic Sciences
• /
• v.31 no.1
• /
• pp.8-16
• /
• 1998

Sexual maturation and reproductive cycle of the goldeye rockfish, Sebastes thompsoni were investigated under photomicroscopy. Samples were collected monthly in the coastal water of Samcheonpo ($34^{\circ}55'N$ ), Korea from November 1995 to October 1996, The ovary consists of several ovarian lamellae originated from ovarian outer membrane. Oogonia which are originated from the inner surface of the ovarian lamella protrude to the ovarian cavity in oocyte stage, and they ave suspended by the egg stalk. The testis is seminiferous tubule type in internal structure. Seminiferous tubule consists of many testicular cysts which contain numerous germ cells in same developmental stage. Biological minimum size of female and male were 19.5 cm and 21.5 cm in total length, respectively. Gonadosomatic index (GSI) of female was the highest (9.56) in March and the lowest (0.15) in August. GSI of male was the highest (0.25) in February and the lowest (0.04) in July. Reproductive cycle was classified into the following successive stages: in female, growing (October and November), maturation ( $December\~February$), gestation (March), parturition and recovery ($April\~June$) and resting ($July\~September$), and in male, growing ($September\~November$), maturation ( December and January), ripe and copulation ( February and March) and degeneration and resting ($April\~August$).