Biomedical subjects

Jeong M Lim

Publications and source records attributed to Jeong M Lim.

9 recordsLinked to original sources

PubMed2006

Preimplantation and fetal development of mouse embryos cultured in a protein-free, chemically defined medium.

Mouse two-cell embryos of block (ICR) or nonblock (F1 of C57BL x DBA) strains were cultured in modified Chatot, Ziomek, and Bavister medium, supplemented with bovine serum albumin or polyvinyl alcohol, a synthetic macromolecule. The supplementation did not influence morula compaction and blastocyst formation, mean cell numbers of total blastomeres, inner cell mass (ICM) cells and trophectodermal (TE) cells in blastocysts, rates of pregnancy and delivery, mean litter size, number of implantation spots in pregnant females, and the expression of several genes related to pluripotency, organogenesis, and implantation.

Animals↗

PubMed2006

Influence of ovarian hyperstimulation and ovulation induction on the cytoskeletal dynamics and developmental competence of oocytes.

This study was undertaken to determine the effects of gonadotrophin on cytoskeletal dynamics and embryo development and its role in improving the retrieval of developmentally competent oocytes. Female golden hamsters were injected with human chorionic gonadotrophin (hCG; 5-, 7.5- or 15-IU) on the day 4 of estrus, pregnant mare serum gonadotrophin (PMSG; 5-, 7.5- or 15-IU) on the day 1 of estrus, or 15-IU hCG at 56 hr post-15-IU PMSG injection in any cycle except estrus. Increasing the hCG dose decreased not only retrieval rate of 2-cell embryo but development to blastocyst after subsequent in vitro culture. Whereas, although increasing the PMSG dose induced increasing the number of 2-cell embryo and blastocyst, 15-IU PMSG injection caused retardation of development to blastocyst. No 2-cell embryos were retrieved by injecting both PMSG and hCG. The injections of 15-IU hCG and 7.5- or 15-IU PMSG inhibited the proliferation of trophectodermal and inner cell mass cells, respectively. Gonadotrophin injection didn't influence microtubular spindle formation, but 5- or 15-IU hCG, 15-IU PMSG, or PMSG and hCG injections induced aberrant cortical granule (CG) and microfilament distribution. After 15-IU hCG or PMSG and hCG injections, fewer oocytes had enriched cortical actin domains, and the expression of alpha-, beta- and gamma-actin genes was greatly increased. In conclusion, a high dose of gonadotrophins alters the microfilament and CG distribution, which in turn reduces the developmental competence of oocytes. Injecting a reduced dose of PMSG to initiate ovarian hyperstimulation without triggering ovulation contributes to the efficient retrieval of developmentally competent oocytes.

Actin Cytoskeleton↗

PubMed2005

Selective decrease of chick embryonic primordial germ cells in vivo and in vitro by soft X-ray irradiation.

The feasibility of soft (low-energy) X-ray irradiation as a means of depleting the endogenous primordial germ cell(s) (PGC) of chicken embryos, to improve the efficiency of germ cell-mediated transgenesis, was investigated. Eggs were subjected to a non-irradiated control treatment and embryos were exposed for 40s to soft X-ray at 15, 16.5, or 18 kV ( approximately 1.5, 1.65, and 1.8 Gy, respectively). Exposure of stage X embryos to each dose of X-ray resulted in a reduction of approximately 50% in the number of PGC apparent at stage 28, whereas the total number of gonadal cells was unaffected. Irradiation (16.5 kV) of embryos at stage 9 or 14 also resulted in similar decreases in the number of PGC with no effect on the total number of gonadal cells. Irradiation did not affect embryo hatchability, compared with the non-irradiated control treatment, although the hatch rate increased with the age of embryos at the time of irradiation. Exposure of gonadal cells isolated from stage 28 embryos to X-ray (16.5 kV, approximately 0.8 Gy) prevented the increase in PGC number during subsequent culture for 10 days; the increase in the total number of gonadal cells was not affected. In conclusion, exposure of chicken embryos to a low dose of soft X-rays is effective for depleting the endogenous PGC population without affecting embryo hatchability or somatic cell viability.

Animals↗

PubMed2005

Development of a hamster superovulation program and adverse effects of gonadotropins on microfilament formation during oocyte development.

OBJECTIVE: To establish a superovulation procedure for the golden hamster (Mesocricetus auratus) by elucidating gonadotropin effects on oocyte development. DESIGN: Randomized, prospective study. SETTING: University laboratory of embryology and gamete biotechnology. ANIMAL(S): Twelve- to 15-week-old female and sexually mature male hamsters. INTERVENTION(S): Different doses of pregnant mare serum gonadotropin (PMSG) were injected into female hamsters in metestrus, diestrus, or proestrus. The same dose of hCG was injected 56 hours later. MAIN OUTCOME MEASURE(S): Embryo development and oocyte morphology after treatment. RESULT(S): First, 10 IU or 15 IU each of PMSG and hCG was injected into 10 hamsters weighing <110 or 110-130 g, respectively. All hamsters were mated, but none delivered live young after injection. Second, the doses of 15 IU, 7.5 IU, 5 IU, or 0 IU of each gonadotropin were injected into each hamster (regardless of body weight, 5 per each group). Increasing numbers of embryos were retrieved as the dosage was increased (11.2 to 46.6 embryos per hamster), whereas the percentage of two-cell embryos at retrieval was significantly decreased (100% to 3%, P<.05). In subsequent culture, none developed to blastocysts after 15-IU injection, whereas 47%, 55%, and 70% of two-cell embryos developed after 7.5-IU, 5-IU, and 0-IU treatments, respectively. As a result, females injected with 5 IU yielded more blastocysts than did females without injection (67 vs. 39). The number of inner cell mass cells per blastocyst was greatly increased in the control groups compared with the 5-IU and 7.5-IU treatment groups (22 vs. 14.3-14.7 cells per blastocyst). Third, the ultrastructure of oocytes was examined after injecting 5 IU each of PMSG and hCG (regardless of body weight). Superovulation did not affect oocyte maturation, but different patterns in microfilament formation were detected after the treatment. CONCLUSION(S): Female hamsters can be superovulated effectively by injecting equal amounts of PMSG and hCG, 56 hours apart. However, embryo development was adversely affected in a dose-dependent manner at all doses of gonadotropins, and microfilament distribution was affected by such treatment.

Actin Cytoskeleton↗

PubMed2004

Promoting effect of amino acids added to a chemically defined medium on blastocyst formation and blastomere proliferation of bovine embryos cultured in vitro.

This study was conducted to elucidate the role of amino acids added singly or in groups to a chemically defined culture medium in blastocyst formation and blastomere proliferation of bovine embryos. Embryos were generated by in vitro fertilization, and blastocyst formation and hatching, and blastomere number of blastocysts were subsequently monitored after the culture of embryos in synthetic oviduct fluid medium (SOFM). First, one of four non-essential amino acids (asparagine, aspartate, glutamate or serine) was added to SOFM and, compared with no addition, a significant (P <0.05) increase in blastocyst formation was found after the addition of asparagine, aspartate, or glutamate (35-42% versus 22%). Second, one of four essential amino acids (arginine, cystine, isoleucine or leucine) was added and arginine or isoleucine greatly improved blastocyst formation (30-36% versus 16%). Third, the addition of five stimulatory amino acids (aspartate, asparagine, glutamate, arginine and isoleucine) to SOFM significantly improved blastocyst formation compared with no addition (12% versus 21%) and such value was similar to that obtained after the addition of 19 amino acids consisting of MEM amino acid solutions (21-27%). However, five amino acids yielded fewer hatched blastocysts than 19 amino acids. Finally, although five amino acids yielded more cell number of blastocysts than no addition (93 versus 74 cells per blastocyst), it was lower than that from 19 amino acids (131 cells per blastocyst). In conclusion, either single or combined addition of asparagine, aspartate, glutamate, arginine and isoleucine stimulated blastocyst formation, while other amino acids might be necessary for further stimulating blastomere proliferation and blastocyst hatching.

Amino Acids↗

PubMed2004

Development of a nonmechanical enucleation method using x-ray irradiation in somatic cell nuclear transfer.

Irradiation of in-vitro-matured bovine oocytes with x-rays of different durations was performed to develop an alternative to conventional mechanical enucleation methods in somatic cell nuclear transfer. No significant difference in embryo development to the blastocyst stage was detected between nonmechanical and mechanical methods, and cytologic analyses of karyotype and microtubule formation showed the potential availability of x-ray irradiation.

Animals↗

PubMed2004

An optimized protocol of a human-to-cattle interspecies somatic cell nuclear transfer.

Human somatic cells were transferred into cattle enucleated oocytes, and a prospective, randomized study was designed to optimize donor cell preparation, fusion medium, and culture method. As a result, improved development of interspecies embryos was achieved by employing serum-starved cord fibroblasts, with Ca(2+)/Mg(2+)-free fusion medium and a serum-free medium being used for bovine embryos.

Animals↗

PubMed2004

An interclass nuclear transfer between fowl and mammal: in vitro development of chicken-to-cattle interclass embryos and the detection of chicken genetic complements.

An attempt was made to develop an interclass somatic cell nuclear transfer method as an alternative means of establishing chicken embryonic stem cells. Chicken-to-cattle interclass embryos that activated calcium ionophore, cycloheximide, and cytochalasin D were developed into blastocysts, and the developing interclass embryos had chicken genetic complements.

Animals↗

PubMed2003

Blastocyst formation, karyotype, and mitochondrial DNA of interspecies embryos derived from nuclear transfer of human cord fibroblasts into enucleated bovine oocytes.

OBJECTIVE: To establish an interspecies somatic cell nuclear transfer (iSCNT) technique for deriving blastocysts having human chromosome complements without sacrificing human oocytes. DESIGN: Prospective, randomized study undertaken in vitro. SETTING: University-affiliated hospital and laboratory, Seoul National University. PATIENT(S): Postpartum women with natural spontaneous vaginal delivery. INTERVENTION(S): Human cord fibroblasts were retrieved from five postpartum women from whom informed consent was obtained. After subculture and cryopreservation, serum-starved cells were transferred into enucleated bovine oocytes. MAIN OUTCOME MEASURE(S): Embryo development, karyotype, and the presence of mitochondrial DNA (mtDNA). RESULT(S): A total 1,742 oocytes were provided for iSCNT and results showed that both fibroblast batch and reconstruction method significantly affected iSCNT outcome. An iSCNT using a single DC pulse of 1.9-2.1 kV/cm for 20 microseconds yielded better rates of fusion (30%-56%) and cleavage (36%) than the other iSCNT protocols. Four to 9% interspecies embryos produced with the optimized method developed to morulae or blastocysts after cultured in a serum-free medium. Results from karyotyping demonstrated that 56% of interspecies embryos evaluated had human chromosome complements. In polymerase chain reaction (PCR) analysis of a single embryo, both human and bovine mtDNAs were detected until the 16-cell stage, whereas only the bovine mtDNA was found beyond the morula stage. CONCLUSION(S): An iSCNT using human cord fibroblasts and bovine oocytes can yield blastocysts and the results of karyotyping and mtDNA analysis confirmed the feasibility of the iSCNT technique.

Adult↗