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Thesis (Ph. D.)--University of Texas at Austin, 2002. / Vita. Includes bibliographical references. Available also from UMI Company.

Thesis (M. Phil.)--University of Hong Kong, 2000. / Includes bibliographical references (leaves 104-132).

Thesis (Ph. D.)--University of Texas at Austin, 2001. / Vita. Includes bibliographical references. Available also in a digital version from UMI/Dissertation Abstracts International.

This thesis describes investigations into Placental Protein 14 (PP14), a immunomodulator involved in human reproduction. The studies included the development of a purification procedure and an investigation of the activity of the protein. In addition the cDNA coding for the protein was cloned and expressed as a recombinant fusion protein and the molecular structure of the protein was predicted and analysed using computer-assisted modelling. Finally the clinical significance of the protein was studied in a range of patient groups. The purification scheme consisted of ion exchange, hydrophobic interaction and gel filtration chromatography, and the pure protein obtained was analysed by SDS-PAGE and Western blotting. The results demonstrate that the purification procedure is a suitable method to obtain PP14 in large quantity and with high purity. PP14 purified by this method retained its activity and was shown to suppress, in a dose-dependent manner, the uptake of 3H-Thymidine by peripheral blood mononuclear cells stimulated with interleukin-2. Purified PP14 was also shown to suppress the uptake of 3H-Thymidine by the cell line U937, also in a dose-dependent manner. This suppression could be removed by the incubation of the PP14 sample with an immunoabsorbent gel linked to monoclonal antibodies against PP14, demonstrating that PP14 was the molecule responsible for the observed activity. Based on the suppression by PP14 of U937 cell growth a bioassay for PP14 was developed, this assay was used to express the specific activity of PPM in Units/ml. To obtain recombinant PP14, mRNA was purified from a tissue sample and reverse transcription used to prepare cDNA. Specific primers were used to amplify the portion of cDNA coding for PP14 which was then ligated into the plasmids pUC 18 and pGEX-KG. Recombinant PP14 was then expressed as a fusion protein with glutathione-S-transferase. The expression conditions were optimised and the fusion protein was purified using affinity chromatography. The structure of PPM was investigated using computer assisted modelling. PPM is a member of the lipocalin family of proteins which share the feature of binding small hydrophobic molecules. The X-ray coordinates of two lipocalins known to share sequence homology with PP14 were used as a basis to model a predicted structure for PP14. An analysis of the structural motifs of the protein was carried out, and it was established that PP14 shares many of the characteristic features of this family of proteins including the presence of a binding pocket. The model was then used to predict potential ligands for PP14.PP14 was measured by radioimmunoassay in uterine flushings from fertile women, women with unexplained infertility and women suffering from recurrent miscarriages, and in plasma samples from fertile and infertile women. The results from the uterine flushings from fertile women showed that PP14 levels rose during the second half of the menstrual cycle reaching ug/ml levels by the end of the cycle. These physiological concentrations are in the same range as the concentrations at which the immunomodulatory activity of PP14 was observed in vitro. The levels of PP14 measured in uterine flushings were lower in infertile women than in fertile women, indicating that a deficiency in PP14 may be associated with infertility. The levels measured in plasma samples from these two groups of women did not pick up this difference. These results suggests that the measurement of proteins such as PP14 in uterine flushings instead of plasma samples may be a more sensitive indicator of local uterine function. In women suffering from recurrent miscarriage a significant lack of secretion of PP14 was observed around the time of implantation. This may be conected with the failure of implantation in these patients. A correlation was observed between the PP14 levels measured in uterine flushings from recurrent miscarriage patients and the level of endometrial development.

Decidualization involves the differentiation of the endometrial tissue into the decidual tissue of the pregnant uterus in several species including humans and rodents. This differentiation occurs only after the onset of implantation in mice and can be artificially-induced causing the formation of deciduomal tissue. The purpose of this study was to identify a group of differentially expressed genes between the developing decidua and deciduoma and study their expression as it may relate to conceptus influenced changes in endometrial gene expression during decidualization. In this study we artificially induced decidualization by transferring blastocyst-sized ConA-coated agarose beads into the uterus on Day 2.5 of pregnancy as we had previously found this model to be more "physiological". Total RNA was isolated from implantation sites of the uteri of pregnant mice as well as pseudopregnant mice that received beads. This RNA was then used for microarray analysis using Mouse Illumina Beadarray chips. This revealed potential differential mRNA levels of over 1,000 genes between the decidua and bead-induced deciduoma tissues of Day 7.5 pregnant and pseudopregnant mice, respectively. Of these, the mRNA levels of 102 genes were 2-fold greater in the decidual tissue while almost twice as many were 2-fold greater in the deciduoma. The broad functions of the protein encoded by the mRNAs included protein binding (e.g. Copz1, Gjb2, Dctn1, Islr, Nisch, Wwc1, Cdc20, Rxrb, Klhl7, Adam10), calcium transport (e.g. Anxa6, Itga11, Clta, Smoc2, Vdr), hydrolase/peptidase activity (e.g. Klk5, Klk26, Klk24, Tmprss4, Ptpn14, Ddx3x, Atp1a2, Usp25, Smarca1), ligase activity (e.g. Iars, Farsla, Ube2v1, Cbll1, Rnf19, Mccc1), and transcription (e.g. Irf1, Hip1, Bhlha15, Supt6h, Scand1, Myocd, Sp3, Mitf, Papolg). We confirmed the differential mRNA levels of a number of gene transcripts using quantitative RT-PCR. Finally, the level and localization of some of the mRNA's identified by our microaray analysis were examined in the mouse uterus during decidualization in more detail and included: Aldh3a1, Bcmo1, Guca2b, GCC, and Inhbb. Localization of mRNA expression in the Day 7.5 implantation site occurred in the mesometrial region near the lumen (Aldh3a1), luminal and glandular epithelia (Guca2b), and endothelial cells lining the sinusoids (Inhbb). This study provides the identity and expression analysis of steady-state mRNA levels of genes whose expression may be influenced by the conceptus using a physiological model for implantation.

The systematic study of the formation of β-SiC formed by low energy carbon ion (C-)implantation into Si followed by high temperature annealing is presented. The research is performed to explore the optimal annealing conditions. The formation of crystalline β-SiC is clearly observed in the sample annealed at 1100 °C for a period of 1 hr. Quantitative analysis is performed in the formation of β-SiC by the process of implantation of different carbon ion fluences of 1×1017, 2×1017, 5×1017, and 8×1017 atoms /cm2 at an ion energy of 65 keV into Si. It is observed that the average size of β-SiC crystals decreased and the amount of β-SiC crystals increased with the increase in the implanted fluences when the samples were annealed at 1100°C for 1 hr. However, it is observed that the amount of β-SiC linearly increased with the implanted fluences up to 5×1017 atoms /cm2. Above this fluence the amount of β-SiC appears to saturate. The stability of graphitic C-C bonds at 1100°C limits the growth of SiC precipitates in the sample implanted at a fluence of 8×1017 atoms /cm2 which results in the saturation behavior of SiC formation in the present study. Secondly, the carbon cluster formation process in silica and the characterization of formed clusters is presented. Silicon dioxide layers ~500 nm thick are thermally grown on a Si (100) wafer. The SiO2 layers are then implanted with 70 keV carbon ions at a fluence of 5×1017 atoms/cm2. The implanted samples are annealed 1100 °C for different time periods of 10 min., 30 min., 60 min., 90 min., and 120 min., in the mixture of argon and hydrogen gas (96 % Ar + 4% hydrogen). Photoluminescence spectroscopy reveals UV to visible emission from the samples. A detail mechanism of the photoluminescence and its possible origin is discussed by correlating the structural and optical properties of the samples. Raman spectroscopy, X-ray photoelectron spectroscopy, X-ray diffraction, spectroscopy, photoluminescence spectroscopy, and transmission electron microscopy are used to characterize the samples.

Indiana University-Purdue University Indianapolis (IUPUI)

Silicon-on-insulator formed by high dose oxygen ion implantation and subsequent epitaxially grown silicon layers were studied and compared with silicon on sapphire materials. Czochralski grown, (100) silicon wafers were implanted with molecular oxygen ions, 0+2, to a total dose of 2.12 x 10^18 0+/cm^2 at an energy of 150 keV/atom.

In the last years, there is a rise of interest in investigation and fabrication of nanometer sized magnetic structures due to their various applications (e.g. for data storage or micro sensors). Over the last several decades ion beam implantation became an important tool for the modification of materials and in particular for the manipulation of magnetic properties. Nanopatterning and implantation can be done simultaneously using focused-ion beam (FIB) techniques. FIB implantation and standard ion implantation differ in their beam current densities by 7 orders of magnitude. This difference can strongly influence the structural and magnetic properties, e.g. due to a rise of the local temperature in the sample during ion implantation. In previous investigations both types of implantation techniques were studied separately. The aim of the current research was to compare both implantation techniques in terms of structural changes and changes in magnetic properties using the same material system. Moreover, to separate any possible annealing effects from implantation ones, the influence of temperature on the structural and magnetic properties were additionally investigated. For the current study a model material system which is widely used for industrial applications was chosen: a 50 nm thick non-ordered nano-crystalline permalloy (Ni81Fe19) film grown on a SiO2 buffer layer based onto a (100)-oriented Si substrate. The permalloy films were implanted with a 30 keV Ga+ ion beam; and also a series of as-deposited permalloy films were annealed in an ultra-high vacuum (UHV) chamber. Several investigation techniques were applied to study the film structure and composition, and were mostly based on non-destructive X-ray investigation techniques, which are the primary focus of this work. Besides X-ray diffraction (XRD), providing the long-range order crystal structural information, extended X-ray absorption fine structure (EXAFS) measurements to probe the local structure were performed. Moreover, the film thickness, surface roughness, and interface roughness were obtained from the X-ray reflectivity (XRR) measurements. Additionally cross-sectional transmission electron microscope (XTEM) imaging was used for local structural characterizations. The Ga depth distribution of the samples implanted with a standard ion implanter was measured by the use of Auger electron spectroscopy (AES) and Rutherford backscattering (RBS), and was compared with theoretical TRIDYN calculation. The magnetic properties were characterized via polar magneto-optic Kerr effect (MOKE) measurements at room temperature. It was shown that both implantation techniques lead to a further material crystallization of the partially amorphous permalloy material (i.e. to an increase of the amount of the crystalline material), to a crystallite growth and to a material texturing towards the (111) direction. For low ion fluences a strong increase of the amount of the crystalline material was observed, while for high ion fluences this rise is much weaker. At low ion fluences XTEM images show small isolated crystallites, while for high ones the crystallites start to grow through the entire film. The EXAFS analysis shows that both Ni and Ga atom surroundings have a perfect near-order coordination corresponding to an fcc symmetry. The lattice parameter for both implantation techniques increases with increasing ion fluence according to the same linear law. The lattice parameters obtained from the EXAFS measurements for both implantation types are in a good agreement with the results obtained from the XRD measurements. Grazing incidence XRD (GIXRD) measurements of the samples implanted with a standard ion implanter show an increasing value of microstrain with increasing ion fluence (i.e. the lattice parameter variation is increasing with fluence). Both types of implantation result in an increase of the surface and the interface roughness and demonstrate a decrease of the saturation polarization with increasing ion fluence. From the obtained results it follows that FIB and standard ion implantation influence structure and magnetic properties in a similar way: both lead to a material crystallization, crystallite growth, texturing and decrease of the saturation polarization with increasing ion fluence. A further crystallization of the highly defective nano-crystalline material can be simply understood as a result of exchange processes induced by the energy transferred to the system during the ion implantation. The decrease of the saturation polarization of the implanted samples is mainly attributed to the simple presence of the Ga atoms on the lattice sites of the permalloy film itself. For the annealed samples more complex results were found. The corresponding results can be separated into two temperature regimes: into low (≤400°C) and high (>400°C) temperatures. Similar to the implanted samples, annealing results in a material crystallization with large crystallites growing through the entire film and in a material texturing towards the (111) direction. The EXAFS analysis shows a perfect near-order coordination corresponding to an fcc symmetry. The lattice parameter of the annealed samples slightly decreases at low annealing temperatures, reaches its minimum at about ~400°C and slightly rises at higher ones. From the GIXRD measurements it can be observed that the permalloy material at temperatures above >400°C reaches its strain-free state. On the other hand, the film roughness increases with increasing annealing temperature and a de-wetting of the film is observed at high annealing temperatures. Regardless of the material crystallization and texturing, the samples annealed at low temperatures demonstrate no change in saturation polarization, while at high temperatures a rise by approximately ~15% at 800°C was observed. The rise of the saturation polarization at high annealing temperatures is attributed to the de-wetting effect.