Analysis of a preimplantation genetic test for aneuploidies in 893 screened blastocysts using KaryoLite BoBs: a single-centre experience.

Introduction: Does euploidy of trophectoderm (TE) biopsies correlate with conventional blastocyst morphological, maternal age and implantation potential?

Methods: This is a one-centre, retrospective, observational study.

Results: Eight hundred and ninety-three blastocysts were biopsied; 57.73% were euploid.

Live birth from vitrified-warmed human oocytes fertilized with frozen-thawed testicular spermatozoa.

A couple with male infertility due to non-obstructive azoospermia were referred to the fertility centre for treatment. Testicular biopsy was performed on the male partner and testicular samples were frozen. The female partner underwent ovarian stimulation and 31 mature oocytes were recovered by ultrasound-guided vaginal aspiration.

Cumulus-specific genes are transcriptionally silent following somatic cell nuclear transfer in a mouse model.

This study investigated whether four cumulus-specific genes: follicular stimulating hormone receptor (FSHr), hyaluronan synthase 2 (Has2), prostaglandin synthase 2 (Ptgs2) and steroidogenic acute regulator protein (Star), were correctly reprogrammed to be transcriptionally silent following somatic cell nuclear transfer (SCNT) in a murine model. Cumulus cells of C57xCBA F1 female mouse were injected into enucleated oocytes, followed by activation in 10 micromol/L strontium chloride for 5 h and subsequent in vitro culture up to the blastocyst stage. Expression of cumulus-specific genes in SCNT-derived embryos at 2-cell, 4-cell and day 4.

Dynamic changes in microtubules and early development of reconstructed embryos after somatic cell nuclear transfer in a non-human primate.

In order to improve somatic cell nuclear transfer (SCNT) efficiency and to understand cellular changes in SCNT, the dynamic changes in microtubules/DNA and early development of SCNT embryos with single or multiple pronuclei were investigated, along with activation timing on efficiency of SCNT, were studied in the Cynomolgus monkey. The confocal images showed that microtubules assembled around condensed DNA at 1h after cell injection; normal or abnormal reconstructed spindle formed at 2 h after cell injection; and reconstructed spindle separated at 2 h after activation. The results of nuclear formation showed that 61.

Exposure of mouse cumulus cell nuclei to porcine ooplasmic extract eliminates TATA box protein binding to chromatin, but has no effect on DNA methylation.

Purpose: The low cloning efficiency with SCNT is due to incomplete or partial reprogramming of the donor somatic cell nuclei after microinjection into the enucleated oocyte. A possible solution may be to initiate nuclear reprogramming prior to SCNT.

Methods: Pre-exposure of donor somatic cell nuclei to a novel porcine ooplasmic extract prior to microinjection could possibly extend the duration of exposure to ooplamic nuclear reprogramming factors.

Early development of reconstructed embryos after somatic cell nuclear transfer in a non-human primate.

To improve efficiency and assess variation in nuclear transfer techniques in non-human primates, we investigated the following factors: type of donor cell, interval between enucleation and cell injection, activation after electrical pulsing and cytokinesis inhibitors. An average of 16.4 oocytes were recovered from 91 retrievals; however, 15 (14%) additional retrieval attempts yielded no oocytes due to a failure of follicular stimulation.

Aberrant profile of gene expression in cloned mouse embryos derived from donor cumulus nuclei.

Somatic cell nuclear transfer has successfully been used to clone several mammalian species including the mouse, albeit with extremely low efficiency. This study investigated gene expression in cloned mouse embryos derived from cumulus cell donor nuclei, in comparison with in vivo fertilized mouse embryos, at progressive developmental stages. Enucleation was carried out by the conventional puncture method rather than by the piezo-actuated technique, whereas nuclear transfer was achieved by direct cumulus nuclear injection.

Developmental competence of transported in-vitro matured macaque oocytes.

This study examines in-vitro maturation (IVM) in a non-human primate model, Macaca fascicularis. The animals had hormonal injections and laparoscopic oocyte retrieval (OR)) at 12- and 24- h after human chorionic gonadotrophin (HCG). The immature oocytes were placed in tightly capped tubes containing pre-equilibrated IVM medium and transported for 5 h in a dry portable 37 degrees C incubator without CO2 supplement.

fertilization outcome in severe ovarian hyperstimulation syndrome: An age-matched contemporaneous control study.

Ovarian hyperstimulation syndrome (OHSS) is a potentially life-threatening, iatrogenic complication of assisted reproduction and has been associated with poor fertilization outcome. The aim of the present study was to evaluate the pregnancy rate and outcome following severe OHSS, at a single center over a three-year period. The incidence of severe OHSS at the IVF Center, National University of Singapore, in Singapore, was 4% (48 cases over 1200 cycles) during the period of 1997-2000.

Reprogramming autologous skeletal myoblasts to express cardiomyogenic function. Challenges and possible approaches.

Cell transplantation therapy is emerging as a promising mode of treatment following myocardial infarction. Of the various cell types that can potentially be used for transplantation, autologous skeletal myoblasts appear particularly attractive, because this would avoid issues of immunogenicity, tumorigenesis, ethics and donor availability. Additionally, skeletal myoblasts display much higher levels of ischemic tolerance and graft survival compared to other cell types.

Influence of cysteamine supplementation and culture in portable dry-incubator on the in vitro maturation, fertilization and subsequent development of mouse oocytes.

The need to transport oocytes and embryos between two laboratories have prompted us to evaluate the effects of in vitro maturation of immature mouse oocytes in a CO2-deficient dry heat portable incubator and subsequent in vitro development of these fertilized mouse oocytes in a standard CO2 incubator. In addition, the effects of cysteamine supplementation on maturation rate and embryonic development during in vitro maturation (IVM) and culture of embryos in the portable incubator were also investigated. Germinal vesicle stage mouse oocytes, recovered at 40-h post-FSH from 6- to 8-week-old C57BL/6xCBA F1 healthy female mice, were matured in vitro in a modified TCM-199 supplemented with or without 100 microM cysteamine in a standard incubator (5% CO2; 37 degrees C) or cultured in a CO2-deficient dry heat portable incubator for 5 h at 37 degrees C and thereafter transferred to a standard incubator for further culture.

Comments about possible use of human embryonic stem cell-derived cardiomyocytes to direct autologous adult stem cells into the cardiomyogenic lineage.

Several studies have shown that cell-transplantation therapy following myocardial infarction has some efficacy in aiding myocardial repair and subsequent recovery of heart function. Large-scale production of human embryonic stem cell-derived cardiomyocytes can potentially provide an abundant supply of donor cells for myocardial transplantation. There are, however, immunological barriers to their use in human clinical therapy.

"Waste" follicular aspirate from fertility treatment--a potential source of human germline stem cells?

Fertility clinics worldwide routinely produce a large volume of 'waste' follicular aspirate, which is potentially an abundant source of immature ovarian follicles. Current attempts to cultivate these further in vitro to yield viable mature oocytes for fertility treatment have not yet achieved much success. Instead, recent lines of evidence have emerged that are suggestive of a potential stem cell niche within such immature ovarian follicles.

Potential utility of cell-permeable transcription factors to direct stem cell differentiation.

Application of embryonic and adult stem cells in regenerative medicine will require efficient protocols for directing stem cell differentiation into well-defined lineages. Differentiation induced by exogenous cytokines, growth factors, or extracellular matrix components will require extended in vitro culture that would delay autologous transplantation and may well alter the immunogenicity of cultured cells. Genetic modulation to direct stem cell differentiation may obviate prolonged culture, but safety concerns preclude clinical application of genetically altered cells in the foreseeable future A novel alternative would be to incorporate protein transduction domains (PTDs) into recombinant transcription factors that play important roles in somatic differentiation.

Effects of granulosa coculture on in-vitro oocyte meiotic maturation within a putatively less competent murine model.

A less competent murine in vitro maturation (IVM) model was achieved by shortening the standard duration of in vivo PMSG stimulation from 48 to 24 h and selecting only naked/partially naked GV oocytes from a mixture of large and small follicles. Porcine granulosa coculture enhanced meiotic maturation within such a less competent model (37.3% versus 23.

The first cell cycle after transfer of somatic cell nuclei in a non-human primate.

Production of genetically identical non-human primates through somatic cell nuclear transfer (SCNT) can provide diseased genotypes for research and clarify embryonic stem cell potentials. Understanding the cellular and molecular changes in SCNT is crucial to its success. Thus the changes in the first cell cycle of reconstructed zygotes after nuclear transfer (NT) of somatic cells in the Long-tailed Macaque (Macaca fascicularis) were studied.

Strategies for directing the differentiation of stem cells into the cardiomyogenic lineage in vitro.

Most studies on stem cell transplantation therapy on myocardially infarcted animal models and phase-I human clinical trials have focused on the use of undifferentiated stem cells. There is a strong possibility that some degree of cardiomyogenic differentiation of stem cells in vitro prior to transplantation would result in higher engraftment efficiency, as well as enhanced myocardial regeneration and recovery of heart function. Additionally, this may also alleviate the probability of spontaneous differentiation of stem cells into undesired lineages and reduces the risk of teratoma formation, in the case of embryonic stem cells.

An overview and synopsis of techniques for directing stem cell differentiation in vitro.

The majority of studies on stem cell differentiation have so far been based in vivo, on live animal models. The usefulness of such models is limited, since it is much more technically challenging to conduct molecular studies and genetic manipulation on live animal models compared to in vitro cell culture. Hence, it is imperative that efficient protocols for directing stem cell differentiation into well-defined lineages in vitro are developed.

Strategies for the cryopreservation of microencapsulated cells.

The major challenge in developing cryopreservation protocols for microencapsulated cells is that the relatively large size (300-400 microm) and the fragile semipermeable membrane of microcapsules makes them particularly prone to cryodamage. Rapid-cooling cryopreservation protocols with high DMSO concentrations (3.5M, 25% v/v) resulted in low post-thaw cell viability (<10%), which did not improve with higher concentrations (4.

Ovarian hyperstimulation syndrome is associated with reversible impairment of vascular reactivity.

Objective: To determine if there is a loss of normal peripheral arteriolar vasoconstrictor reactivity in women with severe ovarian hyperstimulation syndrome (OHSS).

Setting: Prospective controlled study.

Design: National University Hospital, Singapore.

Intracytoplasmic injection of frozen-thawed epididymal spermatozoa in a nonhuman primate model, the cynomolgus monkey (Macaca fascicularis).

Intracytoplasmic sperm injection (ICSI) with frozen-thawed epididymal spermatozoa was performed in the cynomolgus monkey (Macacafascicularis) to produce embryos in vitro. Eleven sexually mature females were hyperstimulated with an GnRH agonist (1.8 mg active triptorelin per 2 kg body weight), followed (2 weeks later) by rFSH (37.