This doctoral research explores the ways in which UK law engages with embryonic processes, namely under the Human Fertilisation and Embryology Act 1990 (as amended). The research offers a fuller understanding of these elusive and evolving biological processes, and in particular, how they can, in turn, allow us to understand legal process and legal regulation more deeply. To do so, the thesis employs an anthropological concept - liminality - coined by Arnold van Gennep, which is itself concerned with revealing the dynamics of process. Liminality may be described as being concerned with the spaces in between distinct stages of human experience or with the process of transition between such stages. With this framing of liminality in mind - which is often characterised as a three-stage process of human experience - the research is divided into three parts, broadly reflecting the three parts of van Gennep's liminal schema: into, through, and out of liminality. It is argued herein that in regulating the embryo - that is, a processual liminal entity in itself - the law is regulating for uncertainty. Tracing the legal governance of the early stages of human life, from its inception to today's regulatory frameworks, the research diagnoses a 'legal gap' between the conceptual basis for regulation, and practical 'realities' of the 1990 Act (as amended). In particular, this 'gap' is typified by uncertainty surrounding embryos in vitro, and what this thesis diagnoses as 'legal stasis'. In order to situate this novel liminal analysis within existing paradigms, however, the thesis first frames embryos in vitro as 'gothic', building upon emergent analytical responses to postmodern forms of categorisation. This framing helps to articulate the nature of, and the reasons for, the above-mentioned 'legal gap'. This framing is nonetheless incomplete without a liminal lens, as it draws our attention to the dynamics of the processes occurring within this 'gap'. It is argued that considering the 'problem' in this manner enables us to move beyond conceptualisation, towards realisation. The gothic, and the liminal are thus used to critically assess legal representations of the embryo, and suggests that there are ways in which the law might better embrace the multiplicity of environments through which the embryo in vitro can travel, that is, either towards reproductive or research ends. It is argued that full recognition of these variable, relational liminal states of the embryo is important for the future of artificial reproduction and embryo research, and that this does not currently happen. In order for the law to reflect better the uncertain nature of embryonic processes, and the technologies that create them, the thesis posits a nuanced, contextual reframing of the embryo that captures the multiplicity of embryonic 'pathways' available within the 1990 Act (as amended). The overarching objective of this work is to consider a more coherent and robust intellectual defence of the ways in which we justify different treatments of in vitro embryos. It thus proposes a 'context-based approach' that embraces the variable, relational pathways already facilitated by the 1990 Act (as amended) in order to lead the embryo (and itself) into, through and out of liminality.

Genes that are associated with the response of cells to growth arrest and DNA damage include the growth arrest specific (<I>gas</I>) genes, the growth arrest and DNA damage (<I>gadd</I>) genes and the tumour suppressor gene <I>p53</I>. This study was undertaken to characterise the changes in gene expression as the embryo responds to growth arrest. The different types of stress that may be encountered by the embryo in culture, such as amino acid deficiency, DNA damaging agents (MMS and sodium arsenite) and metabolic inhibitors (tunicamycin and PALA), induced different patterns of gene expression. Two different pathways implicated with the negative regulation of growth were identified; one involving the transcriptional activator CHOP-10; the other mediated by arrest in the G₁ phase involving <I>p53</I> and <I>gadd45. </I>Both <I>gadd45</I> and <I>p53</I> were expressed in the mouse embryo at the blastocyst-stage, suggesting a role for these genes in a system that arrests growth when embryos are exposed to DNA damage. CHOP-10 was expressed at a constant level from the 8-cell stage onwards and was induced when blastocysts were treated with MMS, the metabolic inhibitor sodium arsenite or an inhibitor or protein glycosylation tunicamycin, but not in blastocysts treated with the inhibitor of nucleotide synthesis PALA. The overexpression of CHOP-10 may be a marker of one of the pathways that lead to apoptosis in the blastocyst. Overall these findings suggest that there is more than one control system regulating growth arrest in the blastocyst and the fetal outcome may differ depending on the type of stress encountered in culture.

Diss. (sammanfattning) Stockholm : Karol. inst. / Härtill 5 uppsatser och 2 appendix.

Diss. (sammanfattning) Stockholm : Karol. inst. / Härtill 5 uppsatser.

In vitro produced (IVP) bovine embryos are useful for investigating the mechanisms affecting early embryonic failure. The work in this thesis explored how interleukin-6 (IL6), an embryokine that increases inner cell mass (ICM) influences post-transfer embryonic survival and development of the embryo-proper and fetus. Four replicates of slaughterhouse-derived cumulus oocyte complexes underwent in vitro maturation and fertilization. On day 5 post-fertilization, embryos were treated with either 1% Bovine Serum Albumin (BSA) (CONT) or 100ng/mL recombinant bovine IL6 with 1% BSA (TRT). On day 7.5 post-fertilization, individual blastocysts were loaded into transfer straws. Beef and dairy cow recipients were synchronized with the day of in vitro fertilization using a 7-d CO-Synch protocol. A subset of cows from each group underwent fixed-time artificial insemination (AI) (n=37). The remaining cows underwent embryo transfer (ET) in the uterine horn ipsilateral to a corpus luteum (CL) (IL6 n=35; CONT n=51). Embryo and fetal measurements were performed via transrectal ultrasonography weekly from days 28-56 post-insemination, respectively. Overall pregnancy rates were 40.0% IL6; 19.6% CONT; and 32.4% AI. Crown-rump lengths (CRL) were reduced (P<0.05) in CONT pregnancies when compared with IL6 and AI at days 28, 35, 42, and 56. A tendency (P=0.057) for larger abdominal diameters was detected between IL6 and CONT groups. Also, IL6 had larger crown-nose lengths than CONT (P<0.05) and tended to be larger than AI (P=0.07). In summary, IL6 treatment produced pregnancies resembling AI-generated pregnancies more so than conventionally cultured embryos, supporting the hypothesis that IL6 improves developmental competency of IVP embryos. / Master of Science / The incidences of pregnancy loss in both beef and dairy cattle industries are profound and are upwards of 60 percent. Financial stability for both of these industries revolves closely around the ability of cattle to give birth to a live calf annually. While artificial insemination (AI) has been heavily adopted and utilized widely in the dairy industry, the use of in vitro produced (IVP) bovine embryos has shown promise in lessening some of the stresses placed on impregnating cattle. The IVP of bovine embryos serves as a strong model to understand how pregnancy losses occur. Briefly, IVP involves the collection of eggs from donor animals, and subsequent fertilization to mimic what occurs within the animal naturally. A disadvantage of in vitro produced embryos is their reduced likelihood to establish pregnancy after transfer into recipient animals. Interleukin-6 (IL6) was recently identified as a pro-developmental factor that may improve the quality and post-transfer competency of in vitro produced embryos. The objective of this work was to determine if IL6 supplementation during in vitro culture improves post-transfer fetal development. Oocytes (i.e. eggs) were retrieved from slaughterhouse-derived ovaries and subjected to in vitro maturation and fertilization. On day 5 post-fertilization, embryos were treated with either 0 (CONT; 1% BSA) or 100ng/mL recombinant bovine IL6. On day 7.5 post-fertilization, individual embryos (blastocyst stage) were loaded into transfer straws. Estrous synchronized beef (n = ) and dairy (n = ) cow recipients were allocated into treatment groups in the following manner. A subset of cows from each group underwent fixed-time AI (n=37). Remaining cows underwent embryo transfer (ET) in the uterine horn ipsilateral to a corpus luteum; 51 of these cows received a CONT embryo and the remaining 35 cows received an IL6 embryo. Thus, there were three treatment groups: AI, CONT, and IL6. Embryo and fetal measurements were performed via transrectal ultrasonography weekly from day 28 to 56, these included crown-rump length, crown-nose length, abdominal diameter, and amniotic vesicle. Pregnancies that remained throughout the entirety of the experiment 40.0% for IL6 (14/35); 19.6% for CONT (10/51); and 32.4% for AI (12/37). In summary, IL6 treatment of embryos produced pregnancies with characteristics more similar to the current industry standard of AI, rather than conventionally cultured embryos (CONT), supporting the hypothesis that IL6 supplementation to bovine embryos on day 5 post-fertilization improves developmental competency of in vitro produced embryos.

Thesis (Ph. D.)--Cornell University, May, 2008. / Vita. Includes bibliographical references.

The focus of this research is to gain insight into the role of dispersion for synchronized development and yield of mature embryos on solid and in liquid culture medium. It is hypothesized that dispersion and synchronous development of embryos will result in high yield of plants. The increase in yield would be helpful in practical implementation of somatic embryogenesis for large-scale clonal propagation of plants and agricultural goods. This doctoral research investigates the yield and the synchronized development of mature embryos, if immature embryos in aggregates of proembryogenic masses (PEMs) have access to same nutritional environment. In order to explore this, a dispersion mechanism was automated and used to provide the same nutritional environment to all PEMs. The effectiveness of the dispersion system was investigated by culturing the PEMs of Norway spruce (Picea abies) on solid medium and in a well-functioning liquid culture system, i.e., bioreactor that could be used for large-scale clonal propagation of plants. Distribution of nutrient concentrations at different locations in an aggregate of PEMs during the culture period was studied using a mathematical model. Results have indicated that dispersion of aggregates of PEMs of Norway spruce has a favorable effect on the rate of proliferation of PEMs and subsequent development of mature somatic embryos. Compared to non-dispersed aggregates of PEMs with dispersed aggregates of PEMs, embryo development increased two folds on solid medium and three to five folds in liquid medium in bioreactors in this study. Bioreactor culture has shown significantly higher yield of mature embryos compared to that on solid culture. The effect of dispersion on synchronized development of mature embryos appears to be cell line dependent. Dispersion has improved synchronization of embryo development in one of two cell lines used in liquid medium experiments and two of four cell lines examined on solid medium. Cell line 11:12:02 has shown more synchronized development of embryos in dispersed PEMs in both medium. To further investigate the details to understand the association between development of mature embryos and nutrient uptake by cells and tissues, a nutrient diffusion model was developed using the volume averaging technique. It estimated the concentration distributions of nutrients, e.g. sugars, in a PEMs cluster on solid medium over a period of culture. The Michaelis-Menten enzyme kinetics was used in the model for uptake of nutrients by cells and tissues. Enzymatic assaying of soluble sugars was performed to determine concentrations of sugars (glucose, fructose, and sucrose) at different locations in tissue clusters. In both experiments and model simulation, sharp decline in concentrations of sucrose and fructose was observed in the first 12 hours after inoculation in glucose-containing ½ LP gel medium. A significant match between predicted and experimental outputs was observed over the culture period. Both experiments and simulation of the model showed a rapid uptake of glucose from the medium and saturation of PEMs cluster within 12 hours. Hence in a PEMs cluster there was no scarcity of nutrients that would inhibit growth and development of somatic embryos. Though dispersion resulted in a significant increase in development of mature somatic embryos, it might not play the decisive role in synchronized development of embryos. It seems that the major factor in synchronization is the initial developmental stage of immature embryos in a culture and genetic characteristics.

No description available.

No description available.

No description available.