• Reproductive and Developmental Biology
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• v.32 no.2
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• pp.81-87
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• 2008

Production of transgenic animals for studying specific gene has been limited due to a low efficiency, lack of skilled researchers and the need for expensive equipment. Currently, the boar spermatozoa as a vector to deliver exogenous DNA into the oocyte were used to improve the efficiency of transfection rate. In this study, we revealed that the optimal conditions for DNA uptake in spermatozoa by liposome were to 90 min of incubation, $17^{\circ}C$, $10^5$ spermatozoa, 4 ng/ml of exogenous DNA and 0.5% (v/v) liposome, without damage to fertility. In addition, the developmental rate to the blastocyst stage of embryo in control group was significantly higher than those embryos with exogenous DNA and liposome, whereas there were no significant differences in embryo development between the liposome and type of DNA. The transfection rates of embryo using treated spermatozoa with both liposome and circular DNA were higher than those using linear DNA. These findings raise the possibility thattreated spermatozoa with liposome/DNA complexes could be used in in vitro fertilization, and the exogenous DNA transferred into the oocytes. Taken together, we demonstrated that liposome a vector for the uptake of exogenous DNA in boar spermatozoa could improve the efficiency of sperm-mediated gene transfer in creating transgenic pig and the other domestic transgenic animals.

• The Korean Journal of Physiology and Pharmacology
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• v.19 no.4
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• pp.319-325
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• 2015

Among solute carrier proteins, the organic anion transporters (OATs) play an important role for the elimination or reabsorption of endogenous and exogenous negatively charged anionic compounds. Among OATs, SLC22A9 (hOAT7) transports estrone sulfate with high affinity. The net decrease of estrogen, especially in post-menopausal women induces rapid bone loss. The present study was performed to search the SNP within exon regions of SLC22A9 in Korean females with osteoporosis. Fifty healthy controls and 50 osteoporosis patients were screened for the genetic polymorphism in the coding region of SLC22A9 using GC-clamped PCR and denaturing gradient gel electrophoresis (DGGE). Six SNPs were found on the SLC22A9 gene from Korean women with/without osteoporosis. The SNPs were located as follows: two SNPs in the osteoporosis group (A645G and T1277C), three SNPs in the control group (G1449T, C1467T and C1487T) and one SNP in both the osteoporosis and control groups (G767A). The G767A, T1277C and C1487T SNPs result in an amino acid substitution, from synonymous vs nonsynonymous substitution arginine to glutamine (R256Q), phenylalanine to serine (F426S) and proline to leucine (P496L), respectively. The Km values and Vmax of the wild type, R256Q, P496L and F426S were 8.84, 8.87, 9.83 and $12.74{\mu}M$, and 1.97, 1.96, 2.06 and 1.55 pmol/oocyte/h, respectively. The present study demonstrates that the SLC22A9 variant F426S is causing inter-individual variation that is leading to the differences in transport of the steroid sulfate conjugate (estrone sulfate) and, therefore this could be used as a marker for certain disease including osteoporosis.

• Reproductive and Developmental Biology
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• v.35 no.3
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• pp.221-225
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• 2011

This study was undertaken to evaluate the relationship between in vitro maturation and plasminogen activators (PAs) activity on porcine cumulus-oocytes complexes (COCs) exposed to oxidative stress. When COCs were cultured in maturation medium with hydrogen peroxide ($H_2O_2$), the proportion of the germinal vesicle breakdown (GVBD) and oocytes maturation were decrease with addition of $H_2O_2$, and were significantly (p<0.05) lower in medium with 0.1 mM $H_2O_2$ than control group. Also, the rate of degenerated oocytes was increased in as $H_2O_2$ concentration in eased. When COCs were cultured for 48 h, three plasminogen-dependent lytic bands were observed: tissue-type PA (tPA); urokinase-type PA (uPA); and tPA-PA inhibitor (tPA-PAI). PA activity was quantified using SDS-PAGE and zymography. When $H_2O_2$ concentration was increased, tPA and tPA-PAI activities also increased in porcine oocytes cultured for 48 h, but not uPA. In other experiment, embryos were divided into three groups and cultured in (1) control medium, (2) control medium with 1.0 mM $H_2O_2$ and (3) control medium with 1.0 mM $H_2O_2$ along with catalase in concentrations of 0.01, 0.1, and 1.0 mg/ml, respectively. $H_2O_2$ decreased the rate of GVBD and maturation in porcine COCs but catalase revealed protective activity, against oxidative stress caused by $H_2O_2$. In this experiment, tPA and tPA-PAI activities were higher in media with 1.0 mM $H_2O_2$ alone. Increasing concentration of catalase decreased tPA and tPA-PAI activities in porcine oocytes. These results indicate that the exposure of porcine follicular oocytes to ROS inhibits oocytes maturation to metaphase-II stage and increase the oocytes degeneration. Also, we speculated that increased ROS level may trigger tPA and tPA-PAI activities in porcine oocytes matured in vitro.

• Journal of the Korean Society of Food Science and Nutrition
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• v.34 no.9
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• pp.1367-1374
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• 2005

Amino acid transporters play an important role in supplying nutrition to normal and cancer cells for cell proliferation. Amino acid transport system L is a major nutrient transport system responsible for the $Na^+$-independent transport of neutral amino acids including several essential amino acids. The system L is divided into two major subgroups, the L-tyre amino acid transporter 1 (LAT1) and the L-type amino acid transporter 2 (LAT2). In the present study, we have examined the expression and functional characterization of system L amino acid transporters in FOB human osteoblast cells. RT-PCR and western blot analysis have revealed that the FOB cells expressed LAT1, LAT2 together with their associating protein 4F2hc. The uptakes of $[^{14}C]_L$-leucine by FOB cells are $Na^+$-independent and almost completely inhibited by system L amino acid transporter selective inhibitor, 2-aminobicyclo-(2,2,1)-heptane-2-carboxylic acid (BCH). These results suggest that the transport of neutral amino acids including several essential amino acids for cellular nutrition into the FOB human osteoblast cells is mediated by system L amino acid transporters.

• Journal of Aquaculture
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• v.11 no.4
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• pp.529-546
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• 1998

Gonadal part that developed by indifferentiation period for 6 months after hatching is made as gonad and fat body. These gonad are thin semi-transparant and undistinguished germ cell. Germinal epithelium is distinguished by development of gonad epithelial tissue from 7 months after hatching. Sex differentiation is begun by oogonia develoment at 8 months after hatching. Primary oocytes grow over germinal epithelium of gonadal cavity, at 9 months after hatching, gonadal cavity become ovarian cavity as they increasing. As soon as oocytes at 13 months after hatching are filled with the whole part of gonad, degeneration of oocyte is begun. And then, gonad has cavity tissue, a small number of oocyte are located in gonadal cavity. At 15 months after hatching, new primary oocyte develop and cavity of ovarian tissue in the central of ovarian cavity. Spermatogonia multiplicate and cavity tissue consist of testicular tissue. These gonad become hermaphrodite and then ditermine the sex of female and male. These results show the red sea bream is juvenile hermaphrodite and undif-ferentiated gonochoristic teleost. Male and female differentiation type of gonad is divided in undifferentiation stage, oogonia-like stage, ovary-like stage, ovary development stage, hermaphroditic testis stage, hermaphroditic ovary stage, and testis development stage. Undifferentiation stage is continued total lenth 18cm at 13 months after hatching. ovary-like stage is continued total length 11~18cm at 13 months after hatching. Ovary-like stage is continued total length 14~26cm at 10~14 months after hatching. Ovary development stage begins from total length 20cm, 14 months after hatching. At 20 months after hatching, 44 percent of total sampled individuals had ovary. Hermaphroditic ovary stage first begins total length 19~20 cm at 15 months after hatching, but it is not observed total length 28~29cm at 20months after hatching. Hermaphroditic testis stage first begins total length 21~22cm at 20months after hatching and is continued for 20months. Testis development stage first begins total length 20~21cm at 20 months after hatching, and is occupied 33 percent total length 28~29cm at 20 months. The beginning of sex differentiation more than 50 percent is from total length 16cm at 11 months after hatching. Sex determination begins total length 20cm, 14months after hatching in female and total length 20cm, 15 months after hatching in male. Sex determination more than 50 percent begins total length 23cm,, 17 months after hatching. Undifferentiated gonadal part of red sea bream consist gonad and fat body. As differentiation is going on and gonad is growing, fat body shrinks. This appearence is showed the same tendency in 3-year old red sea bream. 1.9mm larvae after hatching grow about 19mm larvae for 47 days. The relationship between the total length and body weight of larvae and juveniles in $BW=4.45{\times}10^{-6}TL^{3.4718}$ r=0.9820. Fishes in cage culture grow to maximum total length 28.4cm. The relationship between the total length and body weight of these fishes is $BW=2.36{\times}10^{-2}TL^{2.9180}$, r=0.9971. Undifferentiated gonadal part of red sea bream consist gonad and fat body. As differentiation is going on and gonad is growing, fat body shrinks.

• Journal of Aquaculture
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• v.8 no.1
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• pp.31-46
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• 1995

Gonadal maturation and annual reproductive cycle in ovoviviparous oblong rockfish, Sebastes oblangus on the basis of monthly gonadosomatic indices (GSI), hepatosomatic indices (HSI) and histological observations of gonadal tissues. GSI values of female were in a wide range from $0.l5\pm0.0l\;(July)\;to\;58.54\pm3.86$ (December) and began to increase in August and reached the maxium in December, then decreased rapidly thereafter. Male GSI values were in a range from $0.08\pm0.03$ (July) to $1.55\pm0.27$ (September) and began to increase rapidly in July and reached the maximum in September, then decreased gradually, thereafter. Female HSI was in a range from $0.89\pm0.12$ (December) to $3.73\pm0.15$ (October), and male's was from $2.09\pm0.76$ (October) to $3.62\pm0.48$ (August). HSI reached the maximum values one or two months before GSI reached their maxium values in both sex, and then decreased rapidly thereafter. Mature oocytes began to appear in late October as being oocytes began to mature in August, and the type of oocyte development is categorized in the roup-synchronous oocyte development'. Ovulation and fertilization of ripe oocytes occurred in November, and hatched larvae were born from December to January. Maturation of testis was progressed in short term from August to October and spermatozoa were released in October. Sperm balls consisted of many spermatozoa were preserved in ovarian cavity of female after copulation. These results may suggest that the annual reproductive cycle of oblong rockfish could be divided into the following successive stages: growing (August and September), mature (September and October), gestation (November and December), parturition (December and January) and resting (February to July) in female, and growing (August and September), mature (September and October), copulation (October) and resting (November to July) in male.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.30 no.4
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• pp.627-633
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• 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 $(February\~May)$.

• Development and Reproduction
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• v.20 no.4
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• pp.289-296
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• 2016

This report describes the sex differentiation of the Korean rose bitterling, Rhodeus uyekii, from hatching to 170 days post-hatch (DPH) in relation to total length (TL), body weight (BW), and integral water temperature (IWT). The growth curve of TL from just hatching to 83 DPH was $5.144e^{0.045t}$ ($R^2=0.961$; t, time), and that of BW was $2.398e^{0.086t}$ ($R^2=0.725$). Primordial germ cells (PGCs) were observed at 17 DPH (7.9 mm TL, 3.74 mg BW, $374^{\circ}C$ IWT), and thereafter began to protrude into the peritoneal cavity. At 21 DPH ($9.2{\pm}0.14mm$ TL, $4.8{\pm}0.07mg$ BW, $462^{\circ}C$ IWT), some PGCs contained condensed chromatin and oocyte were observed in meiotic prophase. In contrast to the ovaries, which grew gradually after sexual differentiation, testes began multiplying at 25 DPH (10.1 mm TL, 5.42 mg BW, $550^{\circ}C$ IWT), when testicular differentiation was first identified, and multiplied continuously thereafter. At 33 DPH (11.2 mm TL, 10.5 mg BW, $726^{\circ}C$ IWT), the developing testes contained spermatogonia that exhibited mitotic activity. No spermatocyte or sperm cell was observed until 83 DPH (18.9 TL, 48.2 mg BW, $1,826^{\circ}C$ IWT). At 170 DPH (32.5 mm TL, 270.1 mg BW, $3,740^{\circ}C$ IWT), which was the end point of this study, the mature ovaries showed germinal vesicle breakdown, while the mature testes contained observable spermatocytes and sperm cells. These results allow us to identify the sex differentiation type of the Korean rose bitterling as differentiated gonochoristic.

• Applied Microscopy
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• v.37 no.2
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• pp.65-72
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• 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
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• v.31 no.1
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• pp.8-16
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• 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 ($April\~August$).