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No / This paper investigates the seasonal variations of effective earth radius factor, k, in the Arabian Gulf region where non-standard propagation phenomena are commonly experienced. The k-factor is derived from the vertical profile of refractivity in the area under study. The vertical refractivity gradient, δN, in the first kilometre of the atmosphere above the ground surface is analysed using long term meteorological data. Long-term radiosonde data gathered in the United Arab Emirates (UAE), are used to calculate the variations of refractivity gradient and k-factor parameter.

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<p>Hemophilia A and B are X-linked recessive bleeding disorders caused by the deficiency of coagulation factor VIII (FVIII) and Factor IX (FIX), respectively. Current treatment involves life-long protein replacement therapy which is invasive, expensive and inaccessible to the majority of hemophiliacs worldwide. Treatment is further compromised by the development of neutralizing antibodies. Thus, the development of an alternative treatment that is safer, cost effective and non-invasive that circumvents immune response induction is desirable.</p> <p>To this end, a chitosan-mediated gene therapy strategy delivered orally was developed to provide clinically relevant plasma expression of FVIII or FIX. Hemophilia A mice that ingested chitosan nanoparticles containing FVIII DNA transiently expressed canine FVIII reaching >100 mU one day post treatment, together with partial phenotypic correction. Residual FVIII activity was detected for several days. Repeated administration of nanoparticles restored FVIII expression for 4 weeks and reduced clotting time in treated mice. Interestingly, inhibitors and non-neutralizing antibodies were not detectable throughout the experiment.</p> <p>The immunomodulatory effects of chitosan-mediated oral gene delivery was investigated in naive hemophilia A mice and mice with pre-existing inhibitors. Administration of nanoparticles containing human FVIII DNA in naive mice suppressed systemic antibody responses and provided long-term tolerance to rhFVIII protein immunizations for at least 8 weeks. This tolerance was transferable to naive mice, suggesting development of regulatory T cells. In contrast, repeated oral nanoparticle administration was unable to suppress FVIII-specific antibody responses in hemophilia A mice with pre-existing inhibitors.</p> <p>Treatment of hemophilia B is challenged by a 25-50 fold higher therapeutic threshold. Nevertheless, hemophilia B mice fed chitosan nanoparticles containing CpG-FIXi plasmid transiently expressed therapeutically relevant human FIX >14mU/mL plasma.</p> <p>Chitosan nanoparticle formulation was optimized <em>in vitro</em> for improved transfection efficiency. Nanoparticles formulated at a chitosan:DNA charge ratio of >2:1 (N:P) provided DNA protection against proton and enzymatic degradation that mimic conditions of the stomach and intestine, respectively. The inclusion of 25 mM sodium acetate-acetic acid decreased transfection of HEK 293 cells 4-fold, while 50 mM sodium sulphate increased uptake by ~40%. Optimal transfection was achieved with chitosan chloride (CL 213) formulated at a charge ratio of 3:1 in 50 mM sodium sulphate.</p> <p>These findings suggest chitosan nanoparticles can provide clinically relevant FVIII and FIX transgene expression, which is amenable to a one-tablet-a-day dosing strategy. Taken together, chitosan-mediate gene therapy delivered orally is proposed as a potential non-invasive alternative strategy for hemophilia treatment and without inducing neutralizing and non-neutralizing antibody production.</p> / Doctor of Philosophy (PhD)

Development of the mammary gland is likely mediated by locally produced growth factors acting in concert with circulating mitogens. To investigate the importance of mammary synthesis of insulin-like growth factor-I (IGF-I) and IGF-binding proteins (IGFBP), the initial objective was to evaluate the physiological effects of recombinant IGF-I synthesis in the mouse mammary gland. Expression of recombinant IGF-I was targeted by the mouse mammary tumor virus - long terminal repeat (MMTV-LTR) to the mammary glands of two lines (15 and 29) of transgenic mice. Mammary synthesis of recombinant IGF-I increased the frequency of appearance of mammary alveolar buds (71% vs. 21%) in transgenic compared with non-transgenic CD-1 mice. During lactation, mammary synthesis of recombinant IGF-I reduced the amount of endogenous native IGF-I secreted into milk of transgenic mice. Regardless of transgenesis, a shift in the milk IGFBP profile from predominantly IGFBP-3 to a lower molecular weight IGFBP occurred between d 8 and d 12 of lactation. The altered composition of milk from transgenic line 29 dams reduced by 27% the average daily gain of suckling litters, compared with CD-1 dams. Moreover, mammary glands of transgenic mice were less regressed after weaning than controls and were characterized by the presence of more organized secretory lobules. The second overall objective was to evaluate the regulation and physiological effects of mammary IGF-I and IGFBP synthesis in prepubertal heifers. Serum and extracts of mammary tissue at 5% concentration in media stimulated DNA synthesis 545% and 28%, respectively, in primary mammary epithelial organoids in collagen gel culture. Addition of IGFBP-3 strongly inhibited this growth response. High feeding level tended to increase IGFBP-3 levels in mammary tissue and reduced by 30% the growth response to mammary tissue extracts. Somatotropin increased the mitogenic response to mammary extracts at high feeding level and increased the tissue content of IGF-I by 46%. In summary, local synthesis of IGF-I and IGFBP is influenced by feeding level and exogenous somatotropin and contributes substantially to effects on mammary cell proliferation. Interactions of locally produced IGFBP-3 with IGF-I and other growth factors appear to be especially important when mammary growth is modulated by feeding level. / Ph. D.

The primary aim of this text is to gain insight on how cellular activation by a insulin-like growth factor (IGF-I), in the presence of insulin-like growth factor binding protein-3 (IGFBP-3), is influenced by heparan sulfate proteoglycans (HSPG). Initial research will be presented, assumptions and hypotheses that were included in the development of mathematical models will be discussed, and the future enhancements of the models will be explored. There are many potential scenarios for how each component might influence the others. Mathematical modeling techniques will highlight the contributions made by numerous extracellular parameters on IGF-I cell surface binding. Tentative assumptions can be applied to modeling techniques and predictions may aid in the direction of future experiments. Experimentally, it was found that IGFBP-3 inhibited IGF-I Bovine Aortic Endothelial (BAE) cell surface binding while p9 HS slightly increased IGF-I BAE cell surface binding. IGFBP-3 has a higher binding affinity for IGF-I (3 x 10-9 M) than p9 HS has for IGF-I (1.5 x 10-8 M) as determined with cell-free binding assays. The presence of p9 HS countered the inhibiting effect of IGFBP-3 on IGF-I BAE cell surface binding. Although preliminary experiments with labeled p9 HS and IGFBP-3 indicated little to no cell surface binding, later experiments indicated that both IGFBP-3 and p9 HS do bind to the BAE cell surface. Pre-incubation of BAE cells with either IGFBP-3 or p9 HS resulted in an increase of IGF-I BAE cell surface binding . There was a more substantial increase of IGF-I surface binding when cells were pre-incubated with IGFBP- 3 than p9 HS. There was a larger increase of IGF-I BAE cell surface binding when cells were pre-incubated with p9 HS than when p9 HS and IGF-I were added simultaneously. This suggests that IGFBP-3 and p9 HS surface binding plays key role in IGF-I surface binding, however, p9 HS surface binding does not alter IGF-I surface binding as much as IGFBP-3 surface binding seems to. Experimental work helps further the understanding of IGF-I cellular activation as regulated by IGFBP-3 and p9 HS. Developing mathematical models allows the researcher to focus on individual elements in a complex systems and gain insight on how the real system will respond to individual changes. Discrepancies between the model results and the experimental data presented indicate that soluble receptor inhibition is not sufficient to account for experimental results. The alliance of engineering analysis and molecular biology helps to clarify significant principles relevant to the conveyance of growth factors into tissue. Awareness of the effects of individual parameters in the delivery system, made possible with mathematical models, will provide guidance and save time in the design of future therapeutics involving growth factors. / Master of Science

Throughout gestation, the blood vessel network of the placenta is formed sequentially by processes known as vasculogenesis and angiogenesis, which together meet the needs of the growing fetus. Normal placental angiogenesis is critical to support adequate fetal growth and assure the health of the offspring. Proper angiogenesis requires precise regulation of expression of agents that modulate this process; otherwise, pathologies of pregnancy such as preeclampsia may occur. The placenta is composed of different layers of tissue, including the lamellar (LZ), junctional, and glandular zones, each with a vascular morphology attuned to its function. We hypothesized that higher expression of pro-angiogenic factors is associated with increased morphological metrics in the LZ, the major vascularized zone. Thus, we aimed to characterize the major changes in morphology and vascular development in the placenta throughout pregnancy in cats, alongside a compressive analysis of the expression of major angiogenic factors and their receptors in the placenta, with an emphasis on the identification and interaction of different isoforms of the VEGF family. Microscopic analysis of tissue specimens from different stages of pregnancy revealed increased thickness of the LZ, especially during early to mid-gestation, at which time the tissue is composed of abundant materno-fetal interdigitations that appears rich in capillaries. VEGF proteins were detected in placental tissue in both fetal and maternal cells of the placenta, suggesting stimulatory interactions between different cell types to promote growth and angiogenesis. Gene expression analysis of placenta revealed upregulation of the pro-angiogenic factor VEGF-A in mid-pregnancy, followed by a steady decline toward term, consistent with morphologic changes in the LZ. In contrast, another pro-angiogenic factor, PlGF, showed a marked increase toward term; Flt-1, which acts as a receptor or reservoir for PLGF and VEGF A, was also upregulated at late pregnancy. Increased ratios of PLGF:VEGF-A may contribute to LZ proliferation in the last trimester. These findings are consistent with the creation of a proangiogenic placental state during gestation. Overall, we expect that this research will help elucidate mechanisms of placental vascularization, which can be applied to the design of improved strategies to treat vascular complications of pregnancy. Lastly, we applied the tools developed for placental studies to investigate pathologic angiogenesis in cutaneous squamous cell carcinoma (CSCC), a common skin cancer with major economic and medical impacts in humans and veterinary species. The creation of a new blood supply is essential for growth and metastasis of many tumor types. The goal of this study was to measure expression of variants of proteins that stimulate angiogenesis or transmit an angiogenic stimulus in feline CSCC. The results were mixed, with differences detected in expression of some regulatory agents and, for others, unexpectedly lower expression in CSSC compared to controls. Interestingly, the expression of VEGF-A relative to the protein that transmits its signal (KDR) was elevated in CSCC, suggestive of an altered signaling relationship. This finding supports our hypothesis and is consistent with human SCC studies. Our results encourage further studies on angiogenic factor variants in feline CSCC. / PHD