Development of extra-embryonic membranes and fluid compartments

An account of the development of extra-embryonic membranes in the embryo of poultry. The roles of these membranes in the transfer of water from albumen to yolk and to embryonic tissue is reviewed

Prospects for transgenesis in the chick

Research to develop a useful method for genetic modification of the chick has been on-going since the first demonstrations in the mouse in the 1980s that genetic modification is an invaluable tool for the study of gene function. Manipulation of the chick zygote is possible but inefficient. Considerable progress has been made in developing potentially pluripotent embryo stem cells and their contribution to somatic chimeric birds well-established. Germ line transmission of gametes derived from genetically modified embryo cells has not been described. Transfer of primordial germ cells from a donor embryo to a recipient and production of functional gametes from the donor-derived cells is possible. Genetic modification of primordial germ cells before transfer and their recovery through the germ line has not been achieved. The first transgenic birds described were generated using retroviral vectors. The use of lentiviral vectors may make this approach a feasible method for transgenic production, although there are limitations to the applications of these vectors. It is likely that a method will be developed in the next few years that will enable the use of transgenesis as a tool in the study of development in the chick and for many other applications in basic research and biotechnology

Disappearing women, vanishing ladies and property in embryos

Guidelines on embryo storage prioritise 'respect for the embryo' above the wishes of the women whose labour and tissue have gone into creating the embryo in the first place, effectively making women and the female body disappear. In this article I draw a parallel between this phenomenon relating to embryo storage and other instances of a similar phenomenon that I have called 'the lady vanishes', particularly in stem cell and 'mitochondrial transfer' research. I suggest that a modified property regime could protect women's interests in embryo storage, making a parallel with the Yearworth case, in which it was recognised that men had limited but actionable interests in their stored sperm.

Live birth after fresh embryo transfer vs elective embryo cryopreservation/frozen embryo transfer in women with polycystic ovary syndrome undergoing IVF (FreFro-PCOS): study protocol for a multicenter, prospective, randomized controlled clinical trial

BACKGROUND: Polycystic ovary syndrome (PCOS) patients are at increased risk of pregnancy complications, which may impair pregnancy outcome. Transfer of fresh embryos after superovulation may lead to abnormal implantation and placentation and further increase risk for pregnancy loss and complications. Some preliminary data suggest that elective embryo cryopreservation followed by frozen–thawed embryo transfer into a hormonally primed endometrium could result in a higher clinical pregnancy rate than that achieved by fresh embryo transfer. METHODS/DESIGN: This study is a multicenter, prospective, randomized controlled clinical trial (1:1 treatment ratio of fresh vs. elective frozen embryo transfers).. A total of 1,180 infertile PCOS patients undergoing the first cycle of in vitro fertilization (IVF) or intracytoplasmic sperm injection will be enrolled and randomized into two parallel groups. Participants in group A will undergo fresh embryo transfer on day 3 after oocyte retrieval, and participants in group B will undergo elective embryo cryopreservation after oocyte retrieval and frozen–thawed embryo transfer in programmed cycles. The primary outcome is the live birth rate. Our study is powered at 80 to detect an absolute difference of 10 at the significance level of 0.01 based on a two-sided test. DISCUSSION: We hypothesize that elective embryo cryopreservation and frozen–thawed embryo transfer will reduce the incidence of pregnancy complications and increase the live birth rate in PCOS patients who need IVF to achieve pregnancy. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT0184152

The physiological significance of insemination in programming pregnancy outcome

The cellular and molecular environment of the uterus during the pre - and peri - implantation period of early pregnancy is critical for implantation success and optimal fetal and placental development. Perturbations to this environment not only have consequences for the success of pregnancy and neonatal health and viability, but can also drive adverse health outcomes in the offspring after birth, particularly the development of metabolic disorders such as obesity, hypertension and insulin resistance. The influence of seminal plasma on the cytokine and immune uterine environment has been previously well characterised in mice, however the effects of disruption in uterine seminal plasma exposure for pregnancy outcome have not been investigated. The studies described in this thesis employed the use of surgical seminal vesicle ablation in males and embryo transfer experiments to investigate the physiological significance of uterine seminal plasma exposure on programming fetal and neonatal outcomes, and growth and metabolic status in adult offspring. We demonstrate that in the absence of seminal plasma, oocyte fertilisation and embryo implantation are reduced, showing that seminal plasma acts primarily to facilitate fertilisation, possibly by promoting sperm transport and survival in the reproductive tract. In addition we show that pregnancies initiated in the absence of seminal plasma give rise to offspring which display accelerated growth after birth and increased adiposity in adulthood, compared to those developed in a tract exposed to seminal plasma at the time of conception. Offspring conceived in the absence of seminal plasma also displayed alterations in serum leptin and adiponectin content, similar to those known to be associated with obesity in the mouse. Using embryo transfer experiments, we showed that some, but not all aspects of the perturbed postnatal development are recapitulated when embryos fertilised in the presence of what semen are transferred to a recipient tract which has not been exposed to seminal plasma. More severe perturbations were seen in 2 - cell transfer than in blastocyst transfer experiment. Additionally, there was a significant effect of the embryo transfer procedure, irrespective of seminal plasma exposure, on fetal and postnatal development that confounded interpretation of these experiments. In addition, we investigated the potential mechanisms by which the influence of seminal plasma is exerted. Mediators of pre - implantation embryo development, implantation and the modulation of the maternal immune response to pregnancy were all assessed for regulation by seminal plasma using QRT - PCR. It was demonstrated that seminal plasma exposure induces the up - regulation of key embryotrophic factors, LIF, GM - CSF and IL - 6, in the oviduct following insemination. Factors important in tissue remodelling required for implantation and angiogenesis, MMP - 2, MMP - 3 and VEGF - C, were also shown to be increased at the time of implantation after seminal plasma exposure. Additionally the generation of T - regulatory cells in uterine tissues, demonstrated by the up - regulation of the transcription factor FOXp3 was shown to be dependent on semen exposure. The influence of seminal plasma on embryonic development, implantation and modulation of the maternal immune response to pregnancy may therefore be mechanisms which contribute to the adverse outcomes seen in pregnancies initiated in the absence of seminal plasma. Together these experiments show a role for seminal plasma signalling at the time of insemination in influencing the pre - implantation embryo to program later fetal and neonatal development, thereby impacting on the metabolic health of offspring. We conclude that seminal plasma is not simply a transport medium for sperm, but acts also as a key regulator of a female tract environment providing optimal support for the developing embryo.Thesis (Ph.D.)--School of Paediatrics and Reproductive Health, 2006

Role of the Mitochondrial Genome During Early Development in Mice

The role of the mitochondrial genome in early development and differentiation was studied in mouse embryos cultured in vitro from the two to four cell stage to the blastocyst (about 100 cells). During this period the mitochondria undergo morphological differentiation: progressive enlargement followed by an increase in matrix density, in number of cristae, and in number of mitochondrial ribosomes. Mitochondrial ribosomal and transfer RNA synthesis occurs from the 8 to 16 cell stage on and contributes to the establishment of a mitochondrial protein-synthesizing system. Inhibition of mitochondrial RNA- and protein-synthesis by 0.1 µg/ml of ethidium bromide or 31.2 µg/ml of chloramphenicol permits essentially normal embryo development and cellular differentiation. Mitochondrial morphogenesis is also nearly normal except for the appearance of dilated and vesicular cristae in blastocyst mitochondria. Such blastocysts are capable of normal postimplantation development when transplanted into the uteri of foster mothers. Higher concentrations of these inhibitors have general toxic effects and arrest embryo development. It is concluded that mitochondrial differentiation in the early mouse embryo occurs through the progressive transformation of the preexisting mitochondria and is largely controlled by the nucleocytoplasmic system. Mitochondrial protein synthesis is required for the normal structural organization of the cristae in blastocyst mitochondria. Embryo development and cellular differentiation up to the blastocyst stage are not dependent on mitochondrial genetic activity

Frozen-thawed embryo transfer cycles

Objective: To review the outcomes of frozen-thawed embryo transfer cycles. Design: Retrospective review. Setting: Tertiary assisted reproduction centre, Hong Kong. Patients: Subfertile patients undergoing frozen-thawed embryo transfer between July 2005 and December 2007. Main outcome measures: Clinical and ongoing pregnancy rates. Results: A total of 983 frozen-thawed embryo transfer cycles performed during the study period were reviewed. The clinical pregnancy and ongoing pregnancy rates were 35% and 30%, respectively. Factors associated with successful outcome included younger maternal age (≤35 years) and 4 or more blastomeres at replacement, but not the method of insemination, the cause of subfertility, or the type of frozen-thawed embryo transfer cycle. The overall multiple pregnancy rate was 18%. For cycles with a single embryo replaced, embryos having 4-cell or higher stages at replacement gave an ongoing pregnancy rate of 25%, whereas those with less than 4 cells had a significantly lower ongoing pregnancy rate of 5% only. Blastomere lysis after thawing significantly reduced the clinical pregnancy and ongoing pregnancy rates of cycles with one embryo replaced. Conclusions Clinical pregnancy and ongoing pregnancy rates of frozen-thawed embryo transfer cycles were 35% and 30%, respectively. Higher pregnancy rates were associated with younger maternal age (≤35 years), blastomere numbers of 4 or more, and no blastomere lysis after thawing.published_or_final_versio

Enhancement of mononuclear cell oxidative burst by early post-transfer sera from human patients after unsuccessful embryo transfer

Early pregnancy sera were earlier shown to modulate T lymphocyte rosetting with sheep erythrocytes. We set out to investigate whether early pregnancysera also modulate the state of activation of another immunologically relevant cell type, the monocyte. As an index of monocyte activation, we measured phorbol ester-triggered oxidative burst activity by a highly sensitive chemiluminescence method. Contrary to our expectations, incubation of mononuclearcells with sera taken early after embryo transfer from patients with a successfully developing pregnancy had no effect. Unexpectedly, such sera from patients from a control group of patients in whom no pregnancy developed after embryo transfer caused enhancement of mononuclear cell chemiluminescence. Stimulatory activity of these sera appeared between days 4 and 6 and was maximal between days 7 and 9 post embryo transfer. Whether this phenomenon is causally related to, or represents a consequence of the failure of embryo transfer, can currently not be decide