• Journal of Animal Reproduction and Biotechnology
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• v.34 no.2
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• pp.117-122
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• 2019

The objective of this study was to establish an in vitro culture system for ovarian preantral follicles of B6D2F1. First, we optimized the in vitro preantral-follicle culture by culture duration, follicle stimulating hormone (FSH) type, and activin A concentration. Duration of in vitro culture for 9, 11, and 13 days was sufficient for the normal development of preantral follicles to antral follicles. Formation of cumulus cell-oocyte complex (COC) was induced by treatment with human chorionic gonadotropin (hCG; 2.5 IU/mL) and epidermal growth factor (EGF; 5 ng/mL). In addition, metaphase II (MII) oocytes formed during this in vitro culture of preantral follicles. In vitro preantralfollicle culture for 9 days showed higher rates of growth and maturation, thus yielding a greater number of antral follicles, and there were significant differences (p < 0.05) in the number of MII oocytes (that formed from these preantral follicles via differentiation) between the 9-day culture and 11-day or 13-day culture. The follicles cultured for 9 days contained a tightly packed well-defined COC, whereas in follicles cultured for 11 days, the COC was not well defined (spreading was observed in the culture dish); the follicles cultured for 13 days disintegrated and released the oocyte. Second, we compared the growth of the preantral follicles in vitro in the presence of various FSH types. There were no significant differences in the growth and maturation rates and in differentiation into MII oocytes during in vitro culture between preantral follicles supplemented with FSH from Merck and those supplemented with FSH from Sigma. To increase the efficiency of MII oocyte formation, the preantral follicles were cultured at different activin A concentrations (0 to 200 ng/mL). The control follicles, which were not treated with activin A, showed the highest rate of differentiation into antral follicles and into MII oocytes among all the groups (0 to 200 ng/mL). Therefore, activin A (50 to 200 ng/mL) had a negative effect on oocyte maturation. Thus, in this study, we propose an in vitro system of preantral-follicle culture that can serve as a therapeutic strategy for fertility preservation of human oocytes for assisted reproductive medicine, for conservation of endangered species, and for creation of superior breeds.

• Korean Journal of Animal Reproduction
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• v.15 no.2
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• pp.103-107
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• 1991

This study was carried out to investigate the survival rate in vitro culture after frozen-thawed to used DMSO(dimethyl sulfoxide), glycerol and ethylene glycol of cryoprotective agents at the zona pellucida removed and encased into alien bisected embryo of the mouse early embryos. The results obtained from this study were as follows : 1. The survival rate of in vitro culture after frozen-thawed to used cryoprotective agents of three kinds at the zona pellucida removed bisected morula was 46.6%, 35.8% and 27.3%, total or mean were 36.6%, respectively. 2. The survival rate of in vitro culture after frozen-thawed to used cryoprotective agents of three kinds at the encased into alien bisected morula was 70.6%, 65.3% and 66.4%, total or mean were 67.4%, respectively. 3. The survival rate of in vitro culture after frozen-thawed to used cryoprotective agents of three kinds at the zona pellucida removed bisected blastocysts was 50.4%, 36.7% and 30.4%, total of mean were 39.2%, respectively. 4. The survival rate of in vitro culture after frozen-thawed to used cryoprotective agents of three kinds at the encased into alien bisected blastocysts was 71.1%, 66.7% and 63.9%, total or mean were 67.2%, respectively.

• Korean Journal of Animal Reproduction
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• v.15 no.2
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• pp.97-101
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• 1991

This study was carried out to investigate the survival rate of in vitro culture after frozenthawed, to used DMSO(dimethyl sulfoxide), glycerol and ethylene glycol of cryorpotective agents at the zona pellucida removed and intact on the morulae and blastocysts. The results obtained from this study were as follows : 1. The survival rate of in vitro culture after frozen-thawed to used cryoprotective agents of three kinds at the morulae was 86.0%, 87.1% and 83.3%, total or mean were 85.5%, respectively. 2. The survival rate of in vitro culture after frozen-thawed to used cryoprotective agents of three kinds at the zona pellucida removed morulae was 53.2%, 42.3% and 37.5%, total or mean were 44.3%, respectively. 3. The survival rate of in vitro cultrue after frozen-thawed to used cryoprotective agents of three kinds at the blastocysts was 89.4%, 86.2%, total or mean were 86.7%, respectively. 4. The survival rate of in vitro culture after frozen-thawed to used cryoprotective agents of three kinds at the zona pellucida removed blastocysts was 55.8%, 51.6% and 40.6%, total or mean were 49.3%, respectively.

• Journal of the Korean Association of Oral and Maxillofacial Surgeons
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• v.27 no.5
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• pp.453-456
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• 2001

Background: Autogenous alveolar bone cell transplantation may be suitable for tissue engineering for alveolar bone reconstruction. This study aimed to isolate human alveolar bone-derived cells (HABDCs) and to evaluate the ability of collagen gels to support HABDC proliferation and differentiation for human alveolar bone tissue engineering applications. Method: Cultures of primary HABDCs were established from alveolar bone chips obtained from 10 persons undergoing tooth extraction. These cells were expanded in vitro until passage 3 and used for the in vitro characterization of HABDCs and the in vitro analysis of collagen gels for alveolar bone tissue engineering. Results: Of the 10 attempts made to obtain HABDC cultures, eight were successful. HABDCs expressed the osteoblastic phenotype characterized by alkaline phosphatase activity, osteocalcin expression and the mineralization of the extracellular matrix in vitro. When seeded on collagen gels, HABDCs penetrated into the collagen gel matrices and proliferated inside the gels. Significantly, when HABDCs were embedded into the gels, collagen fibers and mineralization were produced within the gels. Conclusion: This study demonstrates the feasibility of using cultured HABDCs and collagen gels for human alveolar bone tissue engineering applications.

• Journal of Embryo Transfer
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• v.23 no.2
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• pp.77-80
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• 2008

In the present study, effects of concentration of cryoprotectant solutions on the nuclear maturation of vitrified-thawed porcine oocytes were examined. Also, the developmental capacity of vitrified-thawed immature porcine oocytes following ICSI was investigated. Oocytes were cultured in NCSU-23 medium supplemented with 5% FBS at $38^{\circ}C$ in 5% $CO_2$ and air. The in vitro maturation rate of vitrified-thawed oocytes ($24.1{\pm}2.5%$) was lower than that of the control ($46.0{\pm}3.2%$, p<0.05). The in vitro maturation rate of vitrified-thawed oocytes treated with $1.0{\sim}5.0\;ug$ CB + NCSU- 23 medium were $22.2{\pm}3.0%$, $30.7{\pm}3.2$, $46.3{\pm}3.1%$, $38.5{\pm}3.2%$, respectively. The in vitro maturation rate ($46.3{\pm}3.4%$) of the vitrified-thawed oocytes treated with $3.0\;{\mu}g$ CB for 30 min was the highest of all vitrification groups. When the in vitro developmental rates of the vitrified-thawed (with EDS and EDT) oocytes following ICSI were $18.5{\pm}2.5%$, $16.4{\pm}2.1%$, respectively. This results were lower than the control group ($24.0{\pm}2.5%$).

• Journal of Korean Medical Ki-Gong Academy
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• v.7 no.1
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• pp.1-29
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• 2003

Research studies of Qigong therapy for cure for the past 20 years were reviewed from three different categories: clinical study on human patients, in-vitro study of abnormal cells, and in-vivo study of abnormal cell with Qigong therapy, in an attempt to understand the role Qigong therapy plays in many kinds of disease. There is a lot of evidence suggesting that Qigong therapy has an inhibitory effect on abnormal cell growth, both in vitro and in vivo studies, as well as in clinical observation (often there was room for improvement in these studies and some studies require replication in order to verify their findings). Qigong therapy is an area that is often neglected by mainstream medicine and research, and it should be seriously examined and considered as an important supplement to conventional treatment.

• Development and Reproduction
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• v.18 no.3
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• pp.153-160
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• 2014

Adaptive development of early stage embryo is well established and recently it is explored that the mammalian embryos also have adaptive ability to the stressful environment. However, the mechanisms are largely unknown. In this study, to evaluate the possible role of aquaporin in early embryo developmental adaptation, the expression of aquaporin (AQP) 5 gene which is detected during early development were examined by the environmental condition. To compare expression patterns between in vivo and in vitro, we conducted quantitative RT-PCR and analyzed localization of the AQP5 by whole mount immunofluorescence. At in vivo condition, Aqp5 expressed in oocyte and in all the stages of preimplantation embryo. It showed peak at 2-cell stage and decreased continuously until morula stage. At in vitro condition, Aqp5 expression pattern was similar with in vivo embryos. It expressed both at embryonic genome activation phase and second mid-preimplantation gene activation phase, but the fold changes were modified between in vivo embryos and in vitro embryos. During in vivo development, AQP5 was mainly localized in apical membrane of blastomeres of 4-cell and 8-cell stage embryos, and then it was localized in cytoplasm. However, the main localization area of AQP5 was dramatically shifted after 8-cell stage from cytoplasm to nucleus by in vitro development. Those results explore the modification of Aqp5 expression levels and location of its final products by in vitro culture. It suggests that expression of Aqp5 and the roles of AQP5 in homeostasis can be modulated by in vitro culture, and that early stage embryos can develop successfully by themselves adapting to their condition through modulation of the specific gene expression and localization.

• Reproductive and Developmental Biology
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• v.34 no.3
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• pp.223-227
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• 2010

These study was carried out to investigate the effects of the recovery time, diameter of oocytes on in vitro fertilization or intracytoplasmic sperm injection (ICSI). The in vitro maturation rates to MII stage of oocytes recovered at the inactive, follicular and luteal stages matured for 72 h were $1.4{\pm}0.0%$, $43.4{\pm}3.2%$ and $10.8{\pm}2.7%$, respectively. The fertilization rates of in vitro cultured oocytes recovered from ovaries at the in active, follicular and luteal stages were $0.0{\pm}0.0%$, $15.7{\pm}3.4%$ and $7.6{\pm}3.5%$, respectively. The in vitro maturation rate of oocytes recovered from ovaries at the follicular stage of the reproductive cycle was significantly higher than those at the inactive and luteal stages (p<0.05). The penetration rate determined that the percentages of oocytes with diameters in the < $100\;{\mu}m$, 100 to $100\;{\mu}m$ and 110 to $120\;{\mu}m$ ranges were $17.5{\pm}4.7%$, $43.9{\pm}4.5%$, $21.3{\pm}3.4%$, respectively. The penetration rate of oocytes with diameters between 100 to $100\;{\mu}m$ was significantly higher than that of oocytes whose diameters were < $100\;{\mu}m$ and $110{\sim}120\;{\mu}m$ (p<0.05). The penetration rate of oocytes determined that the percentages of ovaries with diameters between 1 to 5 mm and 6 to 10 mm were $32.9{\pm}3.2%$ and $17.5{\pm}3.7%$, respectively. Thus, the diameters of the ovaries were significantly higher at 1 to 5 mm (p<0.05). A total of 264 oocytes were fixed and stained after co-incubation with sperm, of which 72 had identifiable nuclear material. After in vitro fertilization for 20 hrs, 27.3% of oocytes were penetrated by spermatozoas. Oocytes were fixed and stained after ICSI, of which 38 oocytes contained identifiable nuclear material. After in vitro fertilization and ICSI for 20 hrs, to 27.3% and 67.9% of oocytes were penetrated by spermatozoas. The in vitro fertilization rates by ICSI was significantly higher than that in vitro fertilization method (p<0.05).

• Proceedings of the KSAR Conference
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• 2002.06a
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• pp.9-9
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• 2002

This study was to evaluate the in vitro survival of vitrified-thawed bovine MII enucleated (MIIe) oocytes according to activation timing and minimun volume cooling (MVC) method and their in vitro development after somatic cell nuclear transfer (SONT). Bovine oocytes were recovered from slaughtered bovine ovary and matured in TCM-199 supplemented with 10% FBS. (omitted)

• Journal of Animal Science and Technology
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• v.65 no.5
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• pp.1002-1013
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• 2023

The objectives of the present study were to determine the nutrient digestibility of fish meal, defatted black soldier fly larvae (BSFL), and adult flies and to develop equations for estimating in vitro nutrient digestibility of BSFL for pigs. In vitro digestion procedures were employed to mimic the digestion and absorption of nutrients in the pig intestine. Correlation coefficients between chemical composition and in vitro nutrient digestibility of BSFL were calculated. In Exp. 1, in vitro ileal digestibility (IVID) of dry matter (DM) and crude protein (CP) and in vitro total tract digestibility (IVTTD) of DM and organic matter in defatted BSFL meal were less (p < 0.05) than those in fish meal but were greater (p < 0.05) than those in adult flies. In Exp. 2, CP concentrations in BSFL were negatively correlated with ether extract (r = -0.91) concentration but positively correlated with acid detergent fiber (ADF; r = 0.98) and chitin (r = 0.95) concentrations. ADF and chitin concentrations in BSFL were negatively correlated with IVID of DM (r = -0.98 and -0.88) and IVTTD of DM (r = -1.00 and -0.94) and organic matter (r = -0.99 and -0.98). Prediction equations for in vitro nutrient digestibility of BSFL were developed: IVID of CP (%) = -0.95 × ADF (% DM) + 95 (r² = 0.75 and p = 0.058) and IVTTD of DM (%) = -2.09 × ADF + 113 (r² = 0.99 and p < 0.001). The present in vitro experiments suggest that defatted BSFL meal was less digestible than fish meal but was more digestible than adult flies, and nutrient digestibility of BSFL can be predicted using ADF as an independent variable.