• Journal of Environmental Health Sciences
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• v.21 no.1
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• pp.68-73
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• 1995

This study of culturing testicular cell types in vitro has potential to be an invaluable tool for assessing the mechanisms of testicular toxicity, especially those of intragonadal interaction and spermatogenesis. Combined with the Sertoli/germ cell cultures, Leydig cells provide comprehensive and detailed information on the action of testicular toxicants at the level of the testis. Sertoli/germ cell were isolated and incubated well in vitro from 20~30 g rats and Leydig cells from 250~300 g rats. The Sertoli cells isolated from the testis of the SD rats grew into monolayer on about the 2nd~3rd day of culture, an appreciable cell increment being observed between the 4th~5th day. The Leydig cells isolated from the testis of the SD rats grew into a monolayer on about the 3rd-4th day of culture, an appreciable cell increment being observed between the 5th-7th day. These results suggest that Sertoli and Leydig cells can be cultured as a male fertility evaluation method alternative to the in vivo/conventional fertility test method and further study for the physio-chemical determination is needed.

• Korean Journal of Animal Reproduction
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• v.24 no.4
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• pp.385-394
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• 2000

When porcine primordial germ cells (PGCs) isolated from the genital ridge and placed in culture to establish EG cells, a large proportion of PGCs are lost during the early period of culture. To characterize the in vitro death of porcine PGCs, PGCs were cultured in suspension, and apoptosis analyzed using a fluorescent activated cell sorter-based DNA fragmentation assay. The results from flow cytometric analysis showed an increase in apoptosis in cultured cells. However, the cells isolated from the genital ridges are a mixture of PGCs and somatic cells. To detect apoptotic signals specific from porcine PGCs, quantitative TUNEL assay was performed at different time of culture (0 ∼ 24 h). The proportion of apoptotic porcine PGCs determined by double staining with alkaline phosphatase activity and in situ TUNEL assay increased as the time of culture progressed and continued at least 24 h. These results demonstrate that one of the causes of loss of porcine PGCs in vitro is apoptosis.

• Radiation Oncology Journal
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• v.8 no.2
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• pp.145-150
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• 1990

Mesna has been used with ifosfamide to prevent urotoxicity in the treatment of testicular cancers. This drug also protected the toxicities of adriamycin without compromising cytostatic activity. With an idea of radioprotective role of sulfhydryl group of radioprotectors and of mesna decreasing the toxic effect of adriamycin which produces free radicals, mesna and radiation were administered to mice to study the protective effect of this drug and to identify the difference in regenerative capacity of the germ cells in the testis between radiation-treated and both mesna-and radiation-treated groups. The shape and numbers of spermatogenic cells in the seminiferous tubules were examined every week after irradiation. In both groups, initial reduction and later recovery in germ cell numbers and shape was observed. The lowest germ cell number was found around three weeks after irradiation. Mean germ cell number of the mesna-treated group was significantly higher than radiation-treated group at all observed periods (p<0.05). More competent regeneration was present in mesna-treated group. These results suggest that mesna protect the testis from radiation injury. Further study will be necessary to identify whether mesna protects other tissues from radiation and it does not hamper tumor control.

• Proceedings of the Korea Society of Poultry Science Conference
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• 2005.11a
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• pp.66-67
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• 2005

The domestic chicken (Gallus gallus) is an important model for research in developmental biology because its embryonic development occurs in ovo. To examine the mechanism of embryonic germ cell development, we constructed proteome map of gonadal primordial germ cells (gPGC) from chicken embryonic gonads. Embryonic gonads were collected from 500 embryos at 6 day of incubation, and the gPGC were cultured in vitro until colony formed. After 7-10 days in cultured gPGC colonies were separated from gonadal stroma cells (GSCs). Soluble extracts of cultured gPGCs were then fractionated by two-dimensional gel electrophoresis (pH 4-7). A number of protein spots, including those that displayed significant expression levels, were then identified by use of matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF) mass spectrometry and LC-MS/MS. Of the 89 gPGC spots examined, 50 yielded mass spectra that matched avian proteins found in on-line databases. Proteome map of thistype will serve as an important reference for germ cell biology and transgenic research.

• Maxillofacial Plastic and Reconstructive Surgery
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• v.29 no.4
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• pp.279-288
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• 2007

In the present study, we focused on stem cells in the dental papilla of the tooth germ. The tooth germ, sometimes called the tooth bud, is the primordial structure from which a tooth is formed. The tooth germ consists of the enamel organ, the dental papilla, and the dental follicle. The dental papilla lies below a cellular aggregation of the enamel organ. Mesenchymal cells within the dental papilla are responsible for formation of dentin and pulp of a tooth. Tooth germ disappears as a tooth is formed, but that of a third molar stays in the jawbone of a human until the age of 10 to 16, because third molars grow slowly. Impacted third molar tooth germs from young adults are sometimes extracted for orthodontic treatment. In the present study, we evaluated the osteogenic activity and mineralization of cultured human dental papilla-derived cells. Dental papillas were harvested from mandible during surgical extraction of lower impacted third molar from 3 patients aged 13-15 years. After passage 3, the dental papilla-derived cells were trypsinized and subsequently suspended in the osteogenic induction DMEM medium supplemented with 10% fetal bovine serum, 50 g/ml L-ascorbic acid 2-phosphate, 10 nM dexamethasone and 10 mM -glycerophosphate at a density of $1\;{\times}10^6\;cells/dish$ in a 100-mm culture dish. The dental papilla-derived cells were then cultured for 6 weeks and the medium was changes every 3 days during the incubation period. Dental papilla-derived cells showed positive alkaline phosphatase (ALP) staining during 42 days of culture period. The formation of ALP stain showed its maximal manifestation at day 7 of culture period, then decreased in intensity during the culture period. ALP mRNA level was largely elevated at 1 weeks and gradually decreased with culture time. Osteocalcin mRNA expression appeared at day 14 in culture, after that its expression continuously increased in a time-dependent manner up to day 28. The expression remained constant thereafter. Runx2 expression appeared at day 7 with no detection thereafter. Von Kossa-positive mineralization nodules were first present at day 14 in culture followed by an increased number of positive nodules during the entire duration of the culture period. Osteocalcin secretion was detectable in the culture medium from 1 week. The secretion of osteocalcin from dental papilla-derived cells into the medium greatly increased after 3 weeks although it showed a shallow increase by then. In conclusion, our study showed that cultured human dental papilla-derived cells differentiated into active osteoblastic cells that were involved in synthesis of bone matrix and the subsequent mineralization of the matrix.

• Asian-Australasian Journal of Animal Sciences
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• v.22 no.2
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• pp.187-193
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• 2009

The present study identified the favorable conditions for isolation, enrichment and in vitro culture of highly purified, undifferentiated pig spermatogonial stem cell (SSC) lines that proliferate for long periods of time in culture. The colonies displayed morphology similar to miceSSC and were positive for markers of SSC (PGP9.5), proliferating germ cell (PigVASA), pre-meiotic germ cell (DAZL) and pluripotency (OCT4, SSEA-1, NANOG, and SOX2) based on immuno-cytochemistry and RT-PCR. The purity of these colonies was confirmed by negative expression of markers for sertoli cell (GATA4 and SOX9), peritubular myoid cell (${\alpha}$-SMA), differentiating spermatogonial and germ cells (c-KIT). The colonies could be maintained with undifferentiated morphology for more than two months and passaged more than 8 times with doubling time between 6-7 days. Taken together, we conclude that pigSSC could be successfully isolated and cultured in vitro and they possess characteristics similar to miceSSC.

• Journal of Animal Science and Technology
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• v.63 no.5
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• pp.1194-1203
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• 2021

Preparation of recipient stallions is critical step to produce donor spermatogonial stem cell (SSC) derived sperm using transplantation technique. This study was conducted to evaluate the effects of intravenous busulfan infusion on germ cell depletion, semen production, and libido in stallions. Six Thoroughbred stallions were separated into two treatment groups: 1) a multiple low-dose (2.5 mg/kg bw for the first 4 weeks and 5 mg/kg bw for the 5th week); and 2) control group treated with PBS. Testicular samples were obtained at 11 weeks and classified into three different patterns of spermatogenesis, such as normal, Sertoli cell only, and destroyed. Semen collection and libido experiments were performed 1 week before treatment, and 4 and 8 weeks after treatment. For the sperm analysis, total spermatozoa and motility were measured using a light microscope with a motility analyzing system. In the multiple low-dose group, the numbers of tubules categorized as Sertoli cell only were significantly higher than those in the control as well as the total population and total/progressive motility of sperm were significantly decreased 8 weeks after the start of the treatment. The sperm production and motility in the multiple low-dose group appears to be reduced, while libido was maintained. In conclusion, multiple administration of 2.5 mg/kg bw busulfan depletes endogenous germ cells in the stallion recipients for SSC transplantation.

• 대한생식의학회:학술대회논문집
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• 2007.11a
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• pp.113-114
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• 2007

• Proceedings of the Korean Aquaculture Society Conference
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• 2004.05a
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• pp.495-497
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• 2004

• Proceedings of the Korean Society of Developmental Biology Conference
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• 2003.02a
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• pp.31-31
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• 2003

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