PhD theses from IBERS

Xanthophylls as metabolic precursors

2012-01-31T13:52:13Z

Xanthophylls as metabolic precursors Davies, Bethan Wyn

Effect of progesterone priming on ovarian angiogenic factors during late follicular and early luteal development

2010-11-14T02:00:23Z

Effect of progesterone priming on ovarian angiogenic factors during late follicular and early luteal development Christensen, Ana Carolina Martinelli Infertility is a major issue in both human and animal medicine with a great economic impact on reproduction. In order to better understand the common causes of infertility it is necessary to understand the basic physiology underlying the complex process of folliculogenesis and luteogenesis (Campbell et al., 2003). The lack of extensive understanding of the factors involved in regulating follicular and oocyte maturation has resulted in the slow process of developing in vitro systems for the study of folliculogenesis (Campbell et al., 2004). However, domestic ruminants represent a valuable in vivo model system for the study of the endocrine and local mechanisms regulating follicular development not only in ruminants but also in humans (Campbell et al., 2003) and have thus been extensively used in reproduction research. Improved understanding of the basic cellular mechanisms of ovarian physiology may improve biotechnology strategies in livestock reproduction and enhance fertility protocols for endangered species. Similarly, a better understanding of these reproductive processes could lead to innovative strategies to improve reproductive health in women.

A Comparison of Ovarian Function in Juvenile and Adult Ewes Using In Vitro Culture and Proteomics

2011-01-17T11:50:21Z

A Comparison of Ovarian Function in Juvenile and Adult Ewes Using In Vitro Culture and Proteomics Younes, Mohammed A. This work was carried out to compare the endocrine function of ovarian tissue isolated in vitro, in an identical environment between ewes and ewe lambs. Furthermore, to determine whether the differences in endocrine and reproductive function of ewes and ewe lambs are related to differences in the proteomics of corpora lutea, follicles and oocytes. Oestradiol concentrations in tissue cultured in TCM-199 were similar for ewe and ewe lamb follicles collected post slaughter on day 9 to 12 of the oestrous cycle and cultured for different incubation times but increased with increase in follicular size. Oestradiol secretion was greater (P<0.001) for ewe and ewe lamb follicles cultured in media with FCS. Media progesterone concentrations were higher (P<0.001) for ewe than ewe lamb follicles. Progesterone in media and in follicular fluid was increased with increased follicles size. Ewe CL, collected on day 9 to 12 of the oestrous cycle, produced more progesterone than ewe lamb CL when cultured in TCM-199 with or without FCS, PVA, BSA. Proteomics indicated more large spots, in ewe follicular and CL tissue average gels compared with ewe lamb average gels. The protein spots were estimated to be between 45 to 97 kD, in both tissue and age groups, this range of molecular weight could have affected steroid hormone synthesis. (Chapter.3). Ewe and ewe lamb follicles cultured with FSH and LH produced more oestradiol than without, furthermore, oestradiol concentration increased with follicle size. There was no difference in media oestradiol concentration between age groups after 24 h of culture. However, for follicles cultured for 2, 4 or 6 h, concentrations were greater after 4 and 6 h, in ewes than in ewe lambs. Overall ewe lamb follicles produced more progesterone (P<0.001) than ewe follicles when cultured with FSH and LH when cultured for 24 h, but no difference was observed after 2, 4 and 6 hours between ewes and ewe lambs Overall ewe follicles produced more (P<0.002) progesterone than ewe lambs when cultured with different concentration of hCG although there was no difference between ages with respect to oestradiol concentrations. Ewe CL secreted more progesterone (P<0.002) than ewe lamb CL, when cultured for 0 or 24 h. Furthermore, tissue concentrations were greater in ewe CL than ewe lamb CL after incubation in TCM-199, TCM-199 plus BSA, TCM-199 plus FCS and TCM-199 plus PVA. Ewe lamb CL produced more progesterone than ewe CL in medium containing LH when cultured for 2, 4 and 6 hours, but ewes produced more progesterone than lambs when CL were cultured with different concentrations of hCG. Relative to untreated shells, the protein profiles of the ewe follicular shells treated with FSH and LH changed to a greater extent than that of the ewe lambs treated gels in both ages contained more protein spots than control gels. The largest spots were estimated to be between 30 and 97 kDa (Chapter.4). There was no difference between age groups for follicles from ewes and ewe lambs treated with ovagen in oestradiol and progesterone concentrations observed after 2,4, 6 and 8 h of incubation in TCM-199. However, treatment with ovagen plus hCG resulted in higher oestradiol and progesterone concentrations in the media from ewe follicles compared to ewe lambs. Furthermore, there were more protein spots in the range 30 to 66 kDa marker in gels from ewes treated with either ovagen or ovagen plus hCG than for ewe lambs (Chapter 5). Lamb oocytes were smaller than ewe oocytes and developed to a lesser extent in culture. Furthermore, the addition of FCS to TCM-199 caused greater cytoplasmic and nuclear maturation than other media used in this experiment and ewe lamb oocytes have a similar 1D protein bands compared with ewe oocytes, but contained less protein (Chapter 6).

'Identification of Protein Biomarkers Underlying Phenotypic Plasticity and Survival in the Environmental Sentinel Caenorhabditis elegans'

2011-06-01T08:19:51Z

'Identification of Protein Biomarkers Underlying Phenotypic Plasticity and Survival in the Environmental Sentinel Caenorhabditis elegans' Jones, Laura M. Global public health is threatened by increasing risk of disease outbreaks, epidemics, industrial accidents, natural disasters and other health emergencies. Effective management of global public health is reliant upon understanding the response of an organism to its environment and in turn its development and evolution. An environmentally induced change in phenotype, termed phenotypic plasticity, allows survival in heterogeneous environments. Due to its ease of laboratory cultivation and close similarity to parasitic nematodes the genome verified free-living nematode Caenorhabditis elegans provides an ideal animal model in which phenotypic plasticity may be investigated. The developmentally arrested and environmentally resistant dauer stage in C. elegans represents a model in which the ageing process can be investigated. Numerous biochemical, metabolic and transcriptomic analyses have supported a diversion of energy consumption away from anabolic processes and towards enhanced cellular maintenance and detoxification processes. For the first time, this study confirms some of these findings on the protein level. Important components of this enhanced longevity system include the alpha-crystallin family of small heat shock proteins (HSP-12.3 and -12.6), as well as anti-ROS defence systems (including SOD-1 and glutathione peroxidise) and activity of cellular phase II detoxification processes. The multi-gene family of ubiquitous multifunctional proteins GSTs are able to detoxify a broad range of exogenous and endogenous toxins in Phase II cellular detoxification. Comparative investigations of GST complements in dauers and in long-lived daf-2 adults (in comparison to their respective L3 and wild-type controls) have found an up-regulation in daf-2 adults and in dauer larvae, respectively of Pi-class members GST-1 (CE00302) and GST-10 (CE21937) and Nu-Class members GST-5 (CE01613) and GST-7 (CE07055). The use of RNAi and recombinant technology along with synthetically available model and natural substrates has enabled functional and biochemical characterisation to begin on the most abundant of these differently expressed GSTs, GST-1 (CE00302), GST-5 (CE01613) and GST-7 (CE07055). 2DE sub- proteomics revealed that GST-1 and -7 were reduced by approximately 70-80 % and GST-5 by approximately 45 %. Furthermore, in response to RNAi knock-down of GST-1 (CE00302) GST-26 (CE22416) and GST-27 (CE22417) were significantly up-regulated (p < 0.01). Biochemical characterisation of recombinant CE00302 (GST-1), CE01613 (GST-5) and CE07055 (GST-7) has suggested a role in both detoxification of secondary metabolites of lipid peroxidation and in the transport or modification of eicanosoid signals. Furthermore, metabolic analyses of daf-2 and wild-type adults have shown possible evidence of reduced lipid peroxidation in daf-2 adults and also possibly a reduced rate of cell growth and apoptosis. Oxidative damage, as a result of both exogenous and endogenous toxins, is believed to play a role in drug-induced toxicity and in a wide array of clinical disorders (Szeto, 2005). Therefore, these GSTs may provide a useful application as biomarkers for disease and environmental contamination, as well as chemotherapeutic targets for clinical disease treatment and parasitic nematode control.