Cryopreservation of germ cells from genetically proven animals could be a source of restoration tools from the risk of extinction or disappearance of wanted characteristics. Using frozen semen, the genetic gains of Korean native cattle have been increased greatly for 70 years. The preservation of genetic resources as a form of frozen semen straw has limited availability due to the numbers. To circumvent this weakness of frozen semen, we tested two re-freezing methods with different initial thawing temperatures using frozen Korean proven semen and rare breed semen from albino, black and chikso breeders. It has been known that human sperm could resist to cryo-damages by repeated freeze-thaw cycles, but not for Korean proven bulls number (KPN) or for rare breeds. Total 7 frozen semem from brindled(2), black(1), Korean Albino(2) and KPN(1) bulls were used for our research. After thawing straws under 5°C/2min or 37°C/40sec with low temperature water bath and thermo jug, spermatozoa were re-diluted with triladyl diluents after first thawing and re-frozen. Sperm motilities were compared between animals and treated groups after re-thawing. Mean values of motility and viability of refrozen/thawed sperm for expansion of the number of straws were significantly higher in 5°C than in 37°C (P< < 0.05). However, the activity of viable sperm thawed at 5°C was significantly decreased after first and second thawing. It is estimated that re-freezing of frozen semen from rare Korean native cattle is possible with resistant properties of survived spermatozoa.

Multiple interferon tau (IFNT) genes exist in bovine. An antiluteolytic substance secreted by the bovine conceptus and primarily responsible for maternal recognition of pregnancy is bovine trophoblast protein 1 (bIFNT1), a new type I interferon tau (IFNT) genes. The objectives of this research were to investigate whether multiple, distinct gene encode bIFNT1 and other type I bIFNT gene in the bovine genome and to examine expression of bIFNT1 and other bIFNTc1 mRNAs during conceptus development. These transcrips could be regulated through caudalrelatedhomeobox-2 (CDX2) and ETS2 and/or AP1 (JUN) expression, a transcription factor implicated in the control of cell differentiation in the trophectoderm. The presence of mRNAs encoded by bIFNT1 and type I bIFNTc1 genes were examined quantitatively via reverse transcription-polymerase chain reaction (RT-PCR) analysis of total cellular RNA (tcRNA) extracted from on day 17, 20 and 22 bovine conceptuses. The expression level of bIFNT1 was higher on day 17 transcripts were gradually weakly detectable on day 20 and 22. However, the other bIFNTc1 gene examined transcripts was highly expressed on day 20 and transcripts were weakly detectable on day 17 and 22 bovine conceptuses. Furthermore, human choriocarcinoma JEG3 was co-transfected with an -1kb-bIFNT1/c1-Luc constructs and several transcription factor expression plasmids. Compared to each -1kb-bIFNT1/c1-Luc increased when this constructs were co-transfected with, ETS2, AP1(JUN), CREBBP and/or CDX2. Also, bIFNTc1 gene was had very effect on activity by alone ETS2, and AP1 (JUN) expression factors in choriocarcinoma JEG3 cell. However, bIFNT1 gene expression of the upstream region was not identified. We demonstrated that the activities of bIFN genes are regulated by differential, tissue-specific and developmental competence during pregnancy.

The present study was conducted to investigate the effect of caffeine and theophilline on sperm motility during in vitro fertilization (IVF). Briefly, commercial boar semen was centrifuged and resuspended (5x107 sperms/ml) with fertilization medium (mTBM) supplemented with 2 mM caffeine (Caf), 5 mM theophylline (The), 2 mM caffeine + 5mM theophylline (Caf + The), and not supplemented control. The semen parameters of the four groups were analyzed by computer-assisted semen analysis (CASA, Medical Supply Co. Ltd) system at 6 h as time for IVF at 38.5 C under 5 % CO2 in air. The groups were examined 11 semen parameters such as total motility (TM), curvilinier velocity (VCL), straight-line velocity (VSL), average-path velocity (VAP), and hyperactivated (HYP), etc. A total motility of control group (41.3 %) at 6 h showed significantly (P<0.05) higher than those of other groups (Caf, 37.2; The, 35.2; Caf + The, 30.1 %). Results from many other sperm parameters indicated that the theophylline and caffeine negatively affected on sperm motility during IVF. These results suggested that the supplementation of caffeine and/or theophylline was not essential for IVF in pigs. To prove this suggestion, further studies are needed to analyze fertility and embryonic development after IVF with or without the supplementation of caffeine and/or theophylline.

The periods of elevated temperature and high humidity has been longer since last ten years and cause problems in program of artificial insemination or at the efficiency of in vitro production of transferable embryos. The aims of this study were to evaluate the relationship between time of heat shock (0, 1, 2 and 4), during in vitro maturation and developmental competence of subsequent embryo after in vitro fertilization. The develpmentat rate and percetage of apoptotic cells were evaluated on matured oocyte and day 8. 41℃ Heat treatment after IVM culture significantly decreased the developmental capacity of IVF embryos. Also the number of apoptotic cell in COCs, morula and blatostcysts was started to increase at 2 hr heat treatment but did not affect on the rate of maturation. These results indicate that heat treatment for 2 to 4 hr at 41℃ have negative effects on maturation rate of COCs and lower the developmental competence of heat shocked oocyte derived embryos.

During the oocyte maturation, antioxidants may be beneficial for futher developmental competence against reactive oxygen species (ROS) because the media for oocytes lack boiomolecules that serve as scavengers. In this study, N-acetyl-L-cysteine (NAC), N-acetyl-L-cysteine amide (NACA), glutathione (GSH) and cysteamime were compared to determine the effects of protection for ROS from GV to MII stage when supplemented during in vitro maturation (IVM) and in vitro culture (IVC) of bovine oocytes. NAC is one of well known ROS scavanger and NACA is modified form of NAC to help permeation into cytosolic area of oocytes. Significant improvement on the development undergoing blastocysts (32%, vs 18%, 22%) were found when cysteamine (0.1mM) was added to the maturation medium than NAC (0.3 mM), NACA (0.2mM) or GSH (0.5 mM) as compared to control medium with antioxydents. However, the addition of NAC(18%) or NACA(21%) to media did not improve the proportion of oocytes undergoing development to morula and blastocysts than control (24%) and GSH (26%). Our study showed that medium supplementation with cysteame during IVM and in vitro culture (IVC) improved the rate of bovine embryo development, in contrast to extracellular antioxidants like NAC, NACA and GSH that caused no improvement.

Interferon tau (IFNT), produced by the mononuclear trophectoderm, signals the process of maternal recognition of pregnancy in ruminants, but its expression in vivo is not well characterized. Objectives of this study were to determine IFNT gene isoforms expressed in the bovine uterus, and to identify differences in promoter sequences of IFNT genes that differ in their expression. Through the RNA-seq analysis of bovine conceptuses on days 17, 20 and 22 (day 0 = day of estrus), the expression of only two IFNT transcripts, IFNT1 and IFNTc1, were found, which were indeed classified into the IFNT gene clade. IFNT mRNAs were highest on day 17, and then decreased on days 20, and 22, which were also supported by the results of quantitative RT-PCR. Bovine ear-derived fibroblast (EF) cells were then cotransfected with luciferase reporter constructs carrying 5‘-upstream (positions -1000 to +51) regions of IFNT1 or IFNTc1 and various transcription factor expression plasmids. CDX2, either alone or with other Ap-1, ETS2 and/or CREBBP transcription factors, was found to increase luciferase activity approximately 10 and 18 fold more than twice of those cotransfected with bIFNT1, c1-Luc construct. Furthermore, The degree of transcriptional activation by a combination of the AP1, ETS2, CREBBP and/or CDX2 expression vectors was similar to that of CDX2 along plasmid. However, expression patterns of these Luc activity differented. Whereas bIFNTc1-Luc showed lowest antivity had than bIFNT1-Luc reports. Although, lowest antivity had of the bIFNTc1 –Luc report, cotransfected with the bIFNTc1-Luc construct and AP1(JUN) or/and ETS2 expression plasmid, Luc activity was enhanced approximately 2 and 4-fold more than the bIFNT1-Luc. Furthermore, along CDX2 expression factor had high effect on activity of bIFNT1-Luc reporter than the c1 gene in EF cells. These results suggest that two forms of IFNT genes are expressed in utero and their transcriptional regulations differ.

Multiple interferon tau (IFNT) genes exist in bovine. An antiluteolytic substance secreted by the bovine conceptus and primarily responsible for maternal recognition of pregnancy is bovine trophoblast protein 1 (bIFNT1), one of new type I interferon tau (IFNT) genes. The objectives of this research were to investigate whether multiple, distinct gene encode bIFNT1 and other type I bIFNT gene in the bovine genome or not and to examine the expression of bIFNT1 and other bIFNTc1 mRNAs during conceptus development. The transcription of these genes could be regulated through caudal-related homeobox-2 (CDX2) and ETS2 and/or AP1(JUN) expression, a transcription factor implicated in the control of cell differentiation in the trophectoderm. The presence of mRNAs encoded by bIFNT1 and type I bIFNTc1 genes were examined quantitatively via reverse transcription-polymerase chain reaction (RT-PCR) analysis of total cellular RNA (tcRNA) extracted from on the days 17, 20 and 22 bovine conceptuses. bIFNT1 was highly expressed on the day 17 and transcripts were gradually and weakly detectable on the days 20 and 22. However, the other bIFNTc1 gene examined transcripts was highly expressed on the day 20 and transcripts were weakly detectable on the days 17 and 22 bovine conceptuses. Furthermore, human choriocarcinoma JEG3 was co-transfected with an -1kb-bIFNT1/c1-Luc constructs and several transcription factor expression plasmids. Compared to each -1kb-bIFNT1/c1-Luc increased when this constructs were co-transfected with, ETS2, AP1(JUN), CREBBP and/or CDX2. Also, bIFNTc1 gene was had higher effect on activity by alone ETS2, and AP1(JUN) expression factors in choriocarcinoma JEG3 cell. However, bIFNT1 gene expression of the upstream region was not idented. These results demonstrate that these genes display differential, tissue-specific expression and developmental regulation during pregnancy.

Historically, Korea old cattle had been consisted with various lines of coat color brindle, black and white-brown breeds or more. The two rare lines of black and white coat color are maintained for animal resources and preserved critically. The present study was carried out to evaluate potential usage of cysteamine supplementation during in vitro matration (IVM) and in vitro culture/production of embryo (IVP) by transvaginal ultrasound-guided follicle aspiration (Ovum Pick-Up: OPU) for the establishment of cryo-banking system. Immature slaughterhouse-derived cumulus-oocyte complexes (SL-COCs) were matured in IVM medium supplemented with 0, 0.1, 0.3 or 0.9 mM cysteamine, and then cultured in mSOF-BAS for 8 days after in vitro fertilization. The treatment of 0.1 mM cysteamine on SL-COCs showed higher rate of blastocyst, so OPU-derived COCs from rare breeds were matured in TCM media supplemented with or without 0.1 mM cysteamine, FSH and 5% FBS. The embryos were evaluated their developmental stages on day 8. During IVM, cysteamine treatment significantly increased the embryo production rate of slaughterhouse-derived COCs (19.6% vs. 30.5%). The presence of cysteamine during IVM of OPU-derived COCs from rare Korean cattle breeds (albino white and black line) also increased embryo production rates than those from SL-COCs (27.4% vs. 41.9% and 36.4%). With these results, cysteamine treatment during IVM is one of key factors IVP of blastocysts to establish banking system of endangered rare Koarean cattle with OPU derived transferable blastocysts.

During the ovary preservation in low temperature, the cumulus oocyte complexes(COCs) lose their developmental competences after in vitro fertilization. We used phosphate-buffered saline (PBS) as a basic solutions of at various temperatures (25, 15 or 5 ℃) and supplemented them with 1mM glucose and 0.5mM glutamine as a source of carbohydrate metabolites. After recovery of COCs and in vitro fertilization, a significantly higher number of oocytes developed into blastocysts. The developmental competence of embryos that were originated from ovaries preserved at 15 ℃ was increased compared to those of 25 or 5 ℃. The maturation rate of oocytes was not differed between 24 and 36 h at 15 ℃ but showed lower than control group (71% versus 78%). In vitro-fertilized oocytes from ovaries stored at 25 ℃ for 24 h or at 5 ℃ for 24 h had a significantly decreased developmental potentials, but at 15 ℃ did not (27% versus 29% of blastocysts to develop into day 8). With these results, bovine ovaries can be preserved at 15 ℃ for 36 h without decreasing developmental capacity of in vitro-fertilized oocyte at least to the blastocyst stage. This information provides valuable information of preserving ovaries for embryo transfer or in vitro embryo production.

The purpose of this study is to produce wanted sex progeny of genetically confined White Hanwoo (albinism) with preselected sex sperm. One bull of White Hanwoo was chosen for semen donor and X sperm was sorted by MoFlo XDP cell sorter. To compare the pregnancy and birth rates, KPN straw was used as control, total number of unsorted sperm was 20×106/straw. Sexed X frozen semen with 2×106 cells or 4×106 cells per straw were in seminated twice on Hanwoo heifers. The abnormality of the sexed X semen was 24.9 ± 7.31% and distal reflex abnormality of mid piece was significantly (p<0.05) higher (11.7%) compared with that of KPN 768 (5.6%). There were no differences on the pregnancy and birth rates between 2×106 cells or 4×106 cells of X-sperm but KPN semen showed significant differences (p<0.05). The pregnancy rates of KPN 768, 2×106 cells and 4×106 cells X-sperm of White Hanwoo cattle were 85.0%, 26.3% and 50%. The birth rates were 80.0%, 15.8% and 21.4%, respectively. The female offspring rates of KPN 768, 2×106 cells and 4×106 cells X-sperm of White Hanwoo cattle were 43.8%, 100% and 100% (p<0.05). These results indicated that sex sorted White Hanwoo could be used for the production of wanted progeny with 2×106 cells/straw for AI. To increase the efficiency of calf production, the sperm number of sex sorted semen will be optimized for sex selection of White Hanwoo progeny.

Ski protein is a nuclear transcription factor that does not bind DNA directly. Due to its unique binding properties with multiple factors, Ski could perform various roles in the regulation of both cellular proliferation and differentiation. We had previously reported that Ski protein is present in granulosa cells of atretic follicles, but not in preovulatory follicles, suggesting that Ski has a role in apoptosis of granulosa cells. The alternative fate of granulosa cells other than apoptosis is to differentiate to luteal cells; however, it is unknown whether Ski is expressed and has a role in granulosa cells undergoing luteinization. Thus, the aim of the present study was to examine whether the initiation of luteinization with luteinizing hormone (LH) directly regulates expression of Ski in the luteinized granulosa and luteal cells after ovulation by in vitro models. RT-PCR and real time PCR analysis respectively revealed that LH had no effect on c-Ski mRNA expression in the cultured granulosa cells regardless of LH treatment. Though Ski protein isabsent in granulosa cells of preovulatory follicle, its mRNA(c-Ski) was expressed and the level was unchanged even after LH surge. Taken together, these results demonstrated that Ski protein expression is induced in granulosa cells upon luteinization, and suggested that its expression is regulated post-transcriptionally. Moreover, expression of mRNA of Arkadia, an E3 ubiquitin ligases, in luteinizing granulosa cells in vivo was assessed by real time-PCR. The levels of Arkadia mRNA expression were unchanged during follicular growth and post ovulatory luteinization. These findings suggest that Ski protein level may be regulated during luteinization at translational and/or post-translational level but not by Arkadia.

Sloan-Kettering virus gene product of a cellular pro-oncogene c-Ski is an unique nuclear pro-onco protein and belongs to the Ski/Sno proto-oncogene family. Ski plays multiple roles in a variety of cell types, it can induce both oncogenic transformation and terminal muscle differentiation when expressed at high levels. Ski protein is implicated in proliferation/differentiation in a variety of cells. The alternative fate of granulosa cells other than apoptosis is to differentiate to luteal cells, however, it is unknown whether Ski is expressed and has a role in granulosa cells undergoing luteinization. Thus, the aim of this study was, by means of immunohistochemical methods, to locate Ski protein in the rat ovaries during ovulation and corpora lutea(CL) formation to predict the possible involvement of Ski in luteinization. In addition, to examine whether the initiation of luteinization with luteinizing hormone(LH) directly regulates expression of Ski in the luteinized granulosa and luteal cells after ovulation by in vivo models. In order to examine the expression pattern of Ski protein along with the progress of luteinization, follicular growth was induced by administration of equine chorionic gonadtropin to immature female rat, and luteinization was induced by human chorionic gonadtropin treatment to mimic luteinizing hormone(LH) surge. While no Ski-positive granulosa cells were present in preovulatory follicle, Ski protein expression was induced in response to LH surge, and was maintained after the formation of corpus luteum(CL). These results indicate that Ski is profoundly expressed in the luteinized granulosa cells and luteal cells of CL during luteinization, and suggest that Ski may play a role in luteinization of granulosa cells.

For reconstituting genetic resource(Korean Native Chicken: KNC) with grem-line chimeric chicken made with cryopreserved biastdermal cells, the experiments were carried out to optimize cryopreservating conditions. Stage X biastdemal cells were collected from KNC embryos and dissociated. Cells were susupended in medium containing cyopretectant and fetal bovine serum(FBS), and distributed into plastic ampules. Cell susupensions were seeded to induce ice formation at — 7℃ to — 35℃ at in the experiments, the effect of modification of dissociation way, concentration of FBS and cell density on the vaibility of frezen-thawed cells were investigated by trypan blue exclusion. Then change the way of cell dissociation from pipetting to short time vortexing, viability of frozen- thawed cell tended to be increaced from 29 % to 52 %. Increase concentraition of FBS in frozen medium from 20 % to 80 % made viability of thawed cell from 28 % to 35 %. The viability of thawed cells were 33.9% frozen at 2 embryos/ 0.5 ml, and 43.6 % frozen at 20embryos/0.5 ml. Furthermore, combination of three modifications make big improvement. The viability of frozen-thawed cell was 60 % for combinated method, and 41 % for general method. This result means the advance to practical cryoreservation of blastdermal cell of the KNC(Ogolgye breed).

Ski protein is implicated in proliferation/differentiation in a variety of cells. We had previously reported that Ski protein is present in granulosa cells of atretic follicles, but not in preovulatory follicles, suggesting that Ski has a role in apoptosis of granulosa cells. The alternative fate of granulosa cells other than apoptosis is to differentiate to luteal cells; however, it is unknown whether Ski is expressed and has a role in granulosa cells undergoing luteinization. Thus, the aim of the present study was to locate Ski protein in the rat ovary during luteinizationto predict the possible role of Ski. In order to examine the expression pattern of Ski protein along with the progress of luteinization, follicular growth was induced by administration of equine chorionic gonadtropin to immature female rats, and luteinization was induced by human chorionic gonadtropin treatment to mimic luteinizing hormone (LH) surge. While no Ski-positive granulosa cells were present in preovulatory follicle, Ski protein expression was induced in response to LH surge, and was maintained after the formation of the corpus luteum (CL). Though Ski protein is absent in granulosa cells of preovulatory follicle, its mRNA (c-Ski)was expressed and the level was unchanged even after LH surge. Taken together, these results demonstrated that Ski protein expression is induced in granulosa cells upon luteinization, and suggests that its expression is regulated post-transcriptionally.

Cryopreservation of avian semen is a useful tool to preserve genetic resource for aim of preventing extinction induced by infectious disease like avian influenza. Unlike those of mammals, data from chicken cryopreserved semen has not been showed feasible results. So, various cryoprotectants and diluents have been examined in many methods. In this report, as a major ingredient of avian seminal plasm, glutamine was substituted by alanyl glutamine to enhance physiological stability of chicken semen during freezing. We studied effect of glycerol and Dimethylacetamide(DMA) on motility and progressive motility of spermatozoa using glutamine diluent(EK-G) or alanyl glutamine diluent(EK-A) condition. The semen of Ogye was collected twice a week by the dorso-abdominimal massage method and diluted with same volume of EK-G or EK-A at 25℃ and stored for 10 min at 4℃ in cold chamber. Glycerol or DMA was added to diluted semen to reached 7% of final concentration at 4℃. After 3min of equilibration, the diluted semen was packed into 0.25ml straws and subjected to cryopreservation used freezing equipment. The packed straw were placed on height 5 cm above surface of liquid nitrogen(LN2) and held for 10min. After preserved for 2 weeks, the straw was thawed onto the 4℃ cooling bath. The images of motility and progressive motility spermatozoa were recorded by digital image recorder and analyzed by manual. The results showed 68.5% motility and 34.1% progressive motility in DMA/EKA diluent, 31.45% and 17.6% in glycerol/EKA, 45.4% and 8.6% in DMA/EKG, and 9.7% and 6.4% in glycerol/EKG. With these results, the alanyl glutamine and DMA could be used as a main composition of diluent and cryoprotectant for cryopreservation of chicken semen.

For reconstituting genetic resource(Korean Native Chicken: KNC) with grem-line chimeric chicken made with cryopreserved biastdermal cells, the experiments were carried out to optimize cryopreservating conditions. Stage X biastdemal cells were collected from KNC embryos and dissociated. Cells were susupended in medium containing cyopretectant and fetal bovine serum(FBS), and distributed into plastic ampules. Cell susupensions were seeded to induce ice formation at —7 ℃ to —35 ℃ at in the experiments, the effect of modification of dissociation way, concentration of FBS and cell density on the vaibility of frezen-thawed cells were investigated by trypan blue exclusion. Then change the way of cell dissociation from pipetting to short time vortexing, viability of frozen-thawed cell tended to be increaced from 29 % to 52 %. Increase concentraition of FBS in frozen medium from 20 % to 80 % made viability of thawed cell from 28 % to 35 %. The viability of thawed cells were 33.9% frozen at 2 embryos/ 0.5ml, and 43.6 % frozen at 20 embryos/0.5 ml. Furthermore, combination of three modifications make big improvement. The viability of frozen-thawed cell was 60 % for combinated method, and 41 % for general method. This result means the advance to practical cryoreservation of blastdermal cell of the KNC(Ogolgye breed).

The event that occur in sperm during chemotaxis are only partly known. As a essential step of fertilization, sperm cells should undergo capacitation process inside female genital tracts. To understand the molecular event of calcium signals on sperm cells, Fluo 4 loaded spermatozoa was treated with follicular fluid. The motility of sperm was reduced by follicular fluids. Simultaneously, level of calcium in head and tail was also reduced for 5 10 second. The inhibition of sperm motility was believed as a reversible event, so the follicular fluid in graffiaan follicles in vivo could act as a selector on active spermatozoa that recover motility and calcium signals during ovulation. This suggested that the normal levels of calcium in sperm was also critical for active state of sperm cells and the follicular fluids during ovulation could inhibit the motility of sperm cell via calcium signaling.

Ski protein is implicated in proliferation/differentiation in a variety of cells. We had previously reported that Ski protein is present in granulosa cells of atretic follicles, but not in preovulatory follicles, suggesting that Ski has a role in apoptosis of granulosa cells. The alternative fate of granulosa cells other than apoptosis is to differentiate to luteal cells, however, it is unknown whether Ski is expressed and has a role in granulosa cells undergoing luteinization. Thus, the aim of the present study was to locate Ski protein in the rat ovary during luteinization to predict the possible role of Ski. In order to examine the expression pattern of Ski protein along with the progress of luteinization, follicular growth was induced by administration of equine chorionic gonadtropin to immature female rat, and luteinization was induced by human chorionic gonadtropin treatment to mimic luteinizing hormone (LH) surge. While no Ski-positive granulosa cells were present in preovulatory follicle, Ski protein expression was induced in response to LH surge, and was maintained after the formation of corpus luteum (CL). Though Ski protein is absent in granulosa cells of preovulatory follicle, its mRNA (c-Ski) was expressed and the level was unchanged even after LH surge. Taken together, these results demonstrated that Ski protein expression is induced in granulosa cells upon luteinization, and suggested that its expression is regulated post-transcriptionally.

Sloan-Kettering virus gene product of a cellular protooncogene c-Ski is an unique nuclear pro-oncoprotein and belongs to the Ski/Sno proto-oncogene family. Ski plays multiple roles in a variety of cell types, it can induce both oncogenic transformation and terminal muscle differentiation when expressed at high levels. The aim of the present study was to locate Ski protein in rat ovaries in order to predict the possible involvement of Ski in follicular development and atresia. First, expression of c-Ski mRNA in the ovaries of adult female rats was confirmed by RT-PCR. Then, ovaries obtained on the day of estrus were subjected to immunohistochemical analysis for Ski and proliferating cell nuclear antigen (PCNA) in combination with terminal deoxynucleotidyl transferase- mediated dUTP nick end-labeling (TUNEL). Ski was expressed in granulosa cells that were positive for TUNEL, but negative for PCNA, regardless of the shape and size of follicles. Expression of Ski in TUNEL-positive granulosa cells, but not in PCNA-positive granulosa cells, was also verified in immature hypophysectomized rats having a single generation of developing and atretic follicles by treatment with equine chorionic gonadotropin (eCG). These results indicate that Ski is profoundly expressed in the granulosa cells of atretic follicles, but not in growing follicles, and suggest that Ski plays a role in apoptosis of granulosa cells during follicular atresia.