• Clinical and Experimental Reproductive Medicine
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• v.39 no.3
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• pp.95-106
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• 2012

It has been revealed that multiple cohorts of tertiary follicles develop during some animal estrous cycle and the human menstrual cycle. To reach developmental competence, oocytes need the support of somatic cells. During embryogenesis, the primordial germ cells appear, travel to the gonadal rudiments, and form follicles. The female germ cells develop within the somatic cells of the ovary, granulosa cells, and theca cells. How the oocyte and follicle cells support each other has been seriously studied. The latest technologies in genes and proteins and genetic engineering have allowed us to collect a great deal of information about folliculogenesis. For example, a few web pages (http://www.ncbi.nlm. nih.gov; http://mrg.genetics.washington.edu) provide access to databases of genomes, sequences of transcriptomes, and various tools for analyzing and discovering genes important in ovarian development. Formation of the antrum (tertiary follicle) is the final phase of folliculogenesis and the transition from intraovarian to extraovian regulation. This final step coordinates with the hypothalamic-pituitary-ovarian axis. On the other hand, currently, follicle physiology is under intense investigation, as little is known about how to overcome women's ovarian problems or how to develop competent oocytes from in vitro follicle culture or transplantation. In this review, some of the known roles of hormones and some of the genes involved in tertiary follicle growth and the general characteristics of tertiary follicles are summarized. In addition, in vitro culture of tertiary follicles is also discussed as a study model and an assisted reproductive technology model.

• Biomedical Science Letters
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• v.12 no.2
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• pp.105-112
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• 2006

This study was undertaken to investigate the morphological changes between normal and atretic follicle after gamma irradiation and treatment of follicle stimulating hormone (FSH). The ovaries of each group of treated immature mice were prepared the paraffin sections after 0, 6, 12, and 24 hours (hrs) of those treatment. Hematoxylin-eosin (HE) stain, reticulin stain, and terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick end labeling (TUNEL) immunohistochemical stain were performed on the each paraffin sections. As the results of HE staining, the condensed nuclei of oocytes were observed in the atretic primordial follicles, on the other hand the condensations of granulosa cell nuclei were prominent in the atretic primary, preantral, and antral follicles. Only the granulosa cells of atretic follicle were stained specifically with TUNEL staining but not stained in the theca cells, which suggested granulosa cells degenerated through apoptosis. In the reticulin staining, the basement membranes of atretic follicle which was stained weakly showed irregular structure and detachment from the follicles. The ratio of normal to atretic follicle in control and FSH treated group was about 33% but this ratio increased rapidly over 90% in the 6, 12, and 24 hrs group after the irradiation. It could be suggested that the gamma irradiation is the useful tool far the induction of follicle atresia and immunohistochemical staining methods are essential in the study of follicle atresia.

• 대한생식의학회:학술대회논문집
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• 2004.11a
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• pp.56-57
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• 2004

• Journal of Radiation Protection and Research
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• v.24 no.2
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• pp.87-92
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• 1999

The present study was carried out to evaluate the morphological changes in the degenerating primordial follicles induced by $\gamma$-radiation. The prepubertal female mice of three weeks old ICR strain were whole-body irradiated with a dose of $LD_{80(30)}$ (8.3 Gy). The ovaries were collected at 0 h, 3 h, 6 h, and 12 h post-irradiation. The largest cross sections were prepared with histological semi-thin sections and then observed microscopically. The ratio of normal to atretic follicles was reduced significantly after 6th post-irradiation. At 6 h post-irradiation, the number of degenerated primordial follicles increased. Germinal vesicles disappeared, and lipid droplets increased. No more ooplasmic membranes were seen. Granulosa cells became round in shape, and apoptotic cells started to appear. The ratio of normal to atretic follicles in the control group was 62.50%. The ratio decreased with time after irradiation. The ratio decreased down to 51.61 %, 48.97 %, 11.11 %, and 7.14 % at 0 h, 3 h, 6 h, and 12 h, respectively. Taken together, ionizing radiation acutely induced the degeneration of primordial follicles. The patterns of degeneration are 1) apoptosis of one or more granulosa cells with relatively intact oocyte, 2) apoptosis of oocyte with intact follicle cells, or 3) apoptotic degenerations of both cells. The Present study can provide morphological clues for the identification of degenerating primordial follicles.

• Proceedings of the Korean Society of Embryo Transfer Conference
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• 2002.11a
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• pp.98-98
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• 2002

Recruitment of primordial follicles(PMF) is crucial for female fertility. however, factors and mechanisms that regulate this process is poorly understood. The present study was conducted to obtain an inclusive view of the gene expression and to identify novel factors and their pathways of regulating PMF arrest and/or growth initiation. Ovaries from one-day neonatal(consists of oocyte and PMF) and five-day old(consists of PMF and primary follicles, PRIF) mice were collected, either total RNA or mRNA was isolated, and suppression subtractive hybridization(SSH) was used to isolate and clone genes that differentially expressed in day 1 and day 5 ovaries. Confirmation that some of these genes are differentially expressed in PMF and/or in PRIF was accomplished by using laser captured microdissection(LCM), RT-PCR. in situ hybridization(ISH) and/or immunohistochemistry(IHC). In toto, 357 clones were sequenced and analyzed by BLAST and RIKEN program. Sequences of 330 clones significantly matched database entries while 27 clones were novel. Forty-two and 47 different genes were identified as differentially expressed in day 1 and day 5 ovaries, respectively, while 7 genes were expressed in both stages of ovaries. Day 5-subtracted library included several genes known as markers far growing follicles, such as ZP2, MATER, and fetuin. Among the genes with assigned functions, 23.8% was associated with cell cycle/apoptosis regulation, 7.1% with cellular structure, 11.9% with metabolism, 26.2% with signal transduction, and 31.0% with gene/protein expression in day 1; while 10.6%, 17.0%, 23.5%, 25.5%, and 23.4% in day 5, respectively. Genes such as GDF-8, Lats2, Septin2, and Weel were the highly expressed genes in PMF, while HSP84, Laminin2, MATER, MTi7, PTP, and Wrn were highly expressed genes in PRIF. We have successfully discovered list of genes expressed in day 1 and day 5 ovaries and confirmed that some of them are differentially expressed in PMF and/or PRIF. Gene expression profile from the present study would provide insight for the future study on the mechanism(s) involved in primordial-primary follicular transition. This work was Supported by Korean Health 21 RND Project, Ministry of Health and Welfare, Korea (01-PJ10-PG6-01GN13-0002).

• Korean Journal of Clinical Laboratory Science
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• v.40 no.1
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• pp.31-40
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• 2008

Despite of the importance of the primordial follicle (PMF) recruitment, factors and mechanisms for process are poorly understood. To evaluate expression and role of the follicular transition from PMF to PMF/primary follicles (PMIF) in the present study, we evaluated expression of lats1, lats2, cyclin A1, and cyclin A2 mRNA and protein, and elucidated and role of lats1-cyclin A in the follicular transition from PMF to PRIF. To analysis of differential expression in PMF and PMIF, each stage follicles were collected by day1 and day5 of immuno-compromised rats (ICR) and analyzed by real-time PCR for the genes. For localization of mRNAs and proteins of the genes, in situ hybridization and immunohistochemistry were performed. We confirmed that the lats1, lats2, cyclin A1, and cyclin A2 mRNA were more expressed in PMF than PMIF. Localization of the four genes expression were observed in nuclei of oocytes from the arrested primordial, and in the surrounding granulosa cells of the growing follicles. The mRNA expressions were gradually decreased with follicular development. From immunohistochemistry studies, Cyclin A1 protein expression were observed in oocyte cytoplasmas of early stage follicles, while observed in granulose cells and oocyte nucleoli during growing follicles. This study suggested that the presence of lats gene family might perform negatively regulation of cell proliferation by modulation of the CDC2/Cyclin A complex activity. lats-cyclin A genes in oocytes of the early stage follicles might play a role in the meiotic cell cycle arrest of the primary oocytes at the primordial follicle stage as well as the follicular growth.

• Development and Reproduction
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• v.25 no.3
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• pp.173-183
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• 2021

The incidence of infertility among individuals of reproductive age has been growing due to genetic and environmental factors, and considerable research efforts are focused on solving this issue. Ovarian development is an overly complex process in the body, involving the interaction between primordial germ cells and gonad somatic cells. However, follicles located in the center of the in vitro ovary are poorly formed owing to ovarian complexity, nutrient deficiency, and signaling deficiency. In the present study, we optimized methods for dissociating gonads and culture conditions for the in vitro generation of miniaturized ovaries. The gonads from embryos were dissociated into cell masses and cultured on a Transwell-COL membrane for 3-5 weeks. Approximately 12 follicles were present per in vitro ovary. We observed that miniaturized ovaries successfully matured to MII oocytes in vitro from 150 to 100 ㎛ gonad masses. This method will be useful for investigating follicle development and oocyte production.

• 대한생식의학회:학술대회논문집
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• 2002.11a
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• pp.87.1-87.1
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• 2002

• The Journal of Korean Medicine
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• v.18 no.2
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• pp.273-282
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• 1997

To observe the effect of Heulbuchuketang on endometriosis, surgically induced endometriosis in rats was treated with Heulbuchuketang for l00days. The result are as follows: 1. Fragments of right uterine horn were implanted at the serosal wall of small intestine and its adjacent aterial suppling branch. After 4 weeks later endometriosis was observed at the area of implantation of small intestine. 2. After l00-day-treatment, the size of the endometriosis of treated group was much smaller compared to those of the untreated group. Especially the result of the later 60-day-treatment was much better than that of the early 40-day-treatment. 3. In case of inflammation or pustula induced from endometriosis, the size of abnormal endometriosis tissue showed no change after the total treatment and fibrotic change was often founded there, which was not responding to Heulbuchuketang. 4. In the ovary of untreated group, primordial follicle, primary follicle, maturing primary follicle were easily founded and there was no difference in shape and size between the ovary of Heulbuchuketang treated group. From the upper 1 to 4 laboratory result it can be suggested that Heulbuchuketang can be specially effective to mild endometriosis with conservating the ability of fertility.

• Korean Journal of Animal Reproduction
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• v.24 no.4
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• pp.347-356
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• 2000

포유동물의 난소 내에는 많은 수의 primordial follicles 과 preantral follicles 이 존재하고 있으며, 이것은 수정란을 체외생산하기 위한 잠재적인 난자의 공급원이 될 수 있다. 생쥐 preantral follicles 내에 존재하는 난자의 체외성장과 발달을 위하여 몇몇 배양체계가 개발이 되었으며, 적당한 배양조건에서 감수분열 능력이 없는 preantral follicles 내의 난자가 체외배양을 통하여 난자 직경이 증가하고 완전한 핵성숙을 하는 것으로 나타났다. 또한, 체외성장 및 성숙된 난자로부터 생쥐 산자의 성공적인 생산은 preantral follicles 내의 난자가 체외배양을 통해서도 완전한 발달능력을 얻을 수 있음을 입증하였다. 그렇지만, 생쥐 preantral follicle로부터 수정란의 체외생산능력은 매우 낮은 것으로 보고되고 있다. 한편 사람을 비롯한 돼지, 소와 같은 중ㆍ대가축의 경우에는 pre antral follicle의 체외배양을 통하여 감수분열능이 있는 단계의 난자로까지의 발달이 아직 보고가 되지 않고 있다. 따라서 지금까지의 연구결과들을 종합해 볼 때, preantral follicles의 체외배양조건을 개선하거나 새로운 배양체계를 개발에 대한 많은 연구가 요구되고 있다. 사람과 중ㆍ대가축의 preantral follicles의 체외배양체계의 확립은 우수한 형질을 가진 동물의 확장, 희귀동물 혹은 멸종위기 동물의 보존에 이용될 수 있을 것이며, 그리고 암치료를 위하여 화학적, 방사선 치료를 받아야 하는 여성에게서 향후에 불입치료를 위한 방법으로 활용될 수 있을 것으로 기대된다.