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Estrogens produced by livestock can be released into soils when their manure is spread onto agricultural land. This is the first study to determine the sorption of a range of estrogens in a wide range of soils at the regional scale, including the sorption of the phytoestrogen equol which had never been previously studied. Sorption increased in the order of 17β-estradiol=estriol <estrone<equol in surface soils collected from 41 agricultural fields in Alberta and was significantly positively correlated with soil organic carbon content (SOC) for all estrogens. 17β-estradiol was further investigated and its mineralization in non-amended and manure-amended soils never exceeded 30% at 90 days, which suggest that even under optimum environmental conditions for mineralization, 17β-estradiol or its metabolites estrone and/or estriol appear to have a relatively long persistence in Alberta soils. Maximum 17β-estradiol mineralization was significantly positively correlated with sorption and hence increased in soils with greater SOC such as those used in this study with a long-term history of solid beef manure applications. Two ELISAs were developed using rabbit polyclonal antibodies for future field experiments and environmental monitoring. Of these, a developed 17β-estradiol+estrone+estriol ELISA could detect estriol in water from an edge of field experiment at concentrations as low as 1 ng mL-1.

In birds, estrogen is a strong stimulator of gene programs that regulate the formation of very low density lipoproteins (VLDL). Apolipoprotein-B (ApoB) is an integral part of very low density lipoproteins. In mammals, the rate of ApoB synthesis is controlled by post-translational means. In contrast, estrogen treated birds show changes in ApoB transcript level; in a natural setting, the bird?s metabolism and transcription are in great flux due to yolk formation. Besides the ApoB gene, the entire complement of genes that is necessary to form a VLDL is not known. To determine the genes that play a role in the formation of VLDL 7-10d old chicks were injected with estrogen at several time points over a 24hr period. Following exsanguinations by cardiac puncture, livers were removed and RNA was extracted. The RNA was quantified and hybridized to microarrays using a dual-dye system. Slides were scanned and analyzed, and features were extracted. To qualify microarray results, quantitative real time PCR (q-RTPCR) was done on a selection of genes. Previous studies had shown that approximately 200 genes are upregulated by the treatment of hormone naive chickens with estrogen. As a result of our liver transcriptional profiling, we identified 1,528 genes at 1.5hrs, 1,931 genes at 3hrs, 2,398 genes at 6hrs, 2,356 at 12hrs, and 1,713 genes at 24hrs following estrogen exposure. We determined that these regulated genes include those responsible for the transcription of RNA used to create the gene products that serve as components of VLDL itself or that act in VLDL assembly. These include genes encoding structural proteins, like ApoB, and genes encoding assembly-related proteins. Of the differentially expressed genes as compared to time 0, there were approximately 30% which were unannotated with regards to function limiting conclusions. We hope to determine the function of these genes and to annotate them based on this information.

Estrogens produced by livestock can be released into soils when their manure is spread onto agricultural land. This is the first study to determine the sorption of a range of estrogens in a wide range of soils at the regional scale, including the sorption of the phytoestrogen equol which had never been previously studied. Sorption increased in the order of 17β-estradiol=estriol <estrone<equol in surface soils collected from 41 agricultural fields in Alberta and was significantly positively correlated with soil organic carbon content (SOC) for all estrogens. 17β-estradiol was further investigated and its mineralization in non-amended and manure-amended soils never exceeded 30% at 90 days, which suggest that even under optimum environmental conditions for mineralization, 17β-estradiol or its metabolites estrone and/or estriol appear to have a relatively long persistence in Alberta soils. Maximum 17β-estradiol mineralization was significantly positively correlated with sorption and hence increased in soils with greater SOC such as those used in this study with a long-term history of solid beef manure applications. Two ELISAs were developed using rabbit polyclonal antibodies for future field experiments and environmental monitoring. Of these, a developed 17β-estradiol+estrone+estriol ELISA could detect estriol in water from an edge of field experiment at concentrations as low as 1 ng mL-1.

The physiological mechanisms that control energy homeostasis are reciprocally linked to reproduction. However, the neuroendocrine circuitry that registers endocrine cues to direct homeostatic responses in energy balance and reproduction remain unknown. Neuropeptide Y (NPY) neurons have emerged as a key central target of estrogen and leptin that are capable of modulating both reproduction and energy balance. The hypothesis was generated that NPY neuronal subpopulations act as an integration centre to regulate the effects of estrogen and leptin on these important physiological processes through specific signaling pathways. Using hypothalamic cell lines that express the leptin receptor (Ob-R), estrogen receptor (ER) and NPY, this hypothesis was tested in three aims. 17β-estradiol (E2) was previously demonstrated to biphasically regulate NPY mRNA in the mHypoE-38 neuronal cell line; where 24 h E2 exposure induced NPY gene expression that our group proposed may be involved in the gonadotropin-releasing hormone (GnRH) preovulatory surge. E2 also acts as an anorexigenic hormone through unknown hypothalamic targets. E2 directly decreased NPY secretion in the mHypoE-42 and mHypoA-2/12 neuronal cell lines through ER-α. The anorexigenic action of E2 was mediated through the energy sensing 5’ AMP-activated protein kinase (AMPK) and the phosphoinositide-3-kinase (PI3K) pathway. NPY secretion was also decreased by leptin in mHypoA-59 and NPY-GFP cell models through AMPK- and PI3K-dependent mechanisms. Prolonged exposure to leptin in NPY-GFP cell lines prevented AMPK signaling and the leptin-mediated reduction in NPY secretion, indicating NPY neuronal resistance with prolonged leptin exposure. Leptin also stimulated NPY secretion in mHypoE-38 neurons, which was blocked by pharmacological inhibitors of the mitogen-activated protein kinase (MAPK) and PI3K pathways. Importantly, conditioned medium from the mHypoE-38 NPY neuronal cells induced GnRH transcripts in GT1-7 neurons, which was inhibited by Y1-receptor antagonists. Pharmacological inhibitors of the MAPK and PKA signal transduction pathways attenuated the NPY-mediated increase in GnRH transcription. Based upon these findings, I propose NPY neurons in the hypothalamus consist of a heterogeneous population of neurons, and provide the first evidence of intrinsically different responses to function as physiological integrators for two different systems: NPY secretion can be suppressed to decrease food intake and induced to stimulate GnRH neurons.

Nitric oxide (NO) alters chondrocyte metabolism, and is thought to be a key catabolic mediator in osteoarthritis. NO is also an important modifier of bone metabolism, and may partially mediate the bone-sparing effects of oestrogen. Oestrogen has also been linked to the modulation of osteoarthritis, though its role is not clear. The aim of this study was to examine the structural and metabolic effects of ovariectomy and the NO donor glyceryl trinitrate (GTN) on (1) normal ovine femoro-tibial joint tissues, and (2) the progression of joint lesions in the established ovine meniscectomy model of osteoarthritis. Preliminary investigations tested a novel computer-assisted histomorphometric method of assessing osteochondral changes post-meniscectomy, in a trial of a putative disease-modifying osteoarthritis compound. Quantitative assessment revealed a subtle protective effect not evident by qualitative methods. These techniques were then used to test the experimental hypotheses in a combined trial involving 48 aged ewes, variously subjected to ovariectomy, bilateral lateral meniscectomy, and/or topical GTN therapy. At six months, joint tissues were analysed using histology, histomorphometry, dynamic biomechanical testing, serum markers, bone densitometry, and tissue culture of synovial fibroblasts and explants of cartilage and bone. Ovariectomy modified cartilage structure and chondrocyte metabolism, and induced subchondral bone remodelling. Prior ovariectomy altered the development of OA lesions post-meniscectomy, producing thicker but biomechanically inferior cartilage and elevated metabolic activity in subchondral bone. GTN treatment of normal sheep induced thinner, structurally-altered cartilage in normal sheep, and accentuated cartilage and subchondral bone lesions post-meniscectomy. These results support an important homeostatic role for oestrogen in joint tissues, and show that GTN, a commonly used angina therapy, can induce structural alterations in joint tissues and potentially accelerate the progression of concurrent OA. Results also advance understanding of the role of synovial and subchondral bone changes in the pathogenesis of this OA model.

Thesis (M. Res. (Med.))--University of Hong Kong, 2007.

Diss. (sammanfattning) Stockholm : Karol. inst., 2003. / Härtill 4 uppsatser.