Doctor of Philosophy

dissertationDestabilization of the endothelial monolayer lining blood vessels has profound consequences on organismal homeostasis. Vascular instability plays a well-known role in the pathophysiology of diseases from sepsis to stroke. A variety of factors are known to influence endothelial function, including the extracellular milieu, biomechanical factors, various molecular pathways, as well as genetic elements. Many factors promoting vascular instability have been described; however, far fewer studies have identified factors promoting stability. No overarching theory of vascular stability has yet been proposed that takes into account extracellular, biomechanical, molecular, and genetic variables. In this dissertation, I detail studies of extracellular matrix cues, molecular pathways, and genetic factors in an attempt to identify commonalities associated with regulation of vascular stability. I first show that the extracellular matrix protein elastin normalizes endothelial cell function. Next, I demonstrate a critical role of the small GTPase ARF6 in mediating cytokine-induced endothelial instability. I also identify a crucial role of ARF6 in mediating transduction of mechanical signals in the endothelium. Finally, I describe a central role of various genes associated with a human disease, Cerebral Cavernous Malformation (CCM), in endothelial stability. During the course of these studies, I developed and utilized a variety of tools not often found in molecular biology labs. I built new apparatuses and wrote software programs to answer the questions I had, rather than relying on what had already been built by others. Perhaps my largest contribution to the Li laboratory has been to propagate the use of these new systems to molecular biologists who may have a better capability to ask important questions, and who now have the ability to answer their own questions more quickly, more efficiently, and without the inherent bias associated with the standard protocols used in the majority of molecular biology labs. Finally, during my last two years in the laboratory, I further refined and integrated the tools I developed with new tools available from the Broad Institute to quantitatively evaluate endothelial stability through measurement of both structural and functional phenotypes. Using quantification of structural and functional phenotypes, I identified multiple drugs that ameliorate in vitro CCM models, and found two drugs that significantly reduce the formation of lesions in murine models of CCM disease. Further, I found that one of these compounds was protective against diverse destabilizing cues in the endothelium. These discoveries are important for the study of CCM disease, for patients with CCM disease, and have implications for many other diseases in the future. Overall, my studies resulted in specific new mechanistic insights into a wide-variety of factors that promote endothelial stability. More important is the development of a multilayered strategy and platform that can be applied to study effects of extracellular, biomechanical, molecular, and genetic cues on the endothelium either alone or in any combination. In the immediate future, this platform will serve as a vehicle for creating an over-arching model of vascular stability. Beyond this application, this same platform can be rapidly scaled to answer broad questions about factors crucial in health and disease across a wide variety of other cell and organ types. I intend to use the approach I developed and describe in my dissertation to try to untangle how all genes, all proteins, all diseases, and all drugs interact - a lofty goal to be sure

The complex links between governance and biodiversity

We argue that two problems weaken the claims of those who link corruption and the exploitation of natural resources. The first is conceptual. Studies that use national level indicators of corruption fail to note that corruption comes in many forms, at multiple levels, and may or may not affect resource use. Without a clear causal model of the mechanism by which corruption affects resources, one should treat with caution any estimated relationship between corruption and the state of natural resources. The second problem is methodological: Simple models linking corruption measures and natural resource use typically do not account for other important causes and control variables pivotal to the relationship between humans and natural resources. By way of illustration of these two general concerns, we demonstrate that the findings of a well known recent study that posits a link between corruption and decreases in forests, elephants, and rhinoceros are fragile to simple conceptual and methodological refinements.Environmental Economics and Policy,

Signatures of hybridization in <i>Trypanosoma brucei</i>

Genetic exchange among disease-causing micro-organisms can generate progeny that combine different pathogenic traits. Though sexual reproduction has been described in trypanosomes, its impact on the epidemiology of Human African Trypanosomiasis (HAT) remains controversial. However, human infective and non-human infective strains of Trypanosoma brucei circulate in the same transmission cycles in HAT endemic areas in subsaharan Africa, providing the opportunity for mating during the developmental cycle in the tsetse fly vector. Here we investigated inheritance among progeny from a laboratory cross of T. brucei and then applied these insights to genomic analysis of field-collected isolates to identify signatures of past genetic exchange. Genomes of two parental and four hybrid progeny clones with a range of DNA contents were assembled and analysed by k-mer and single nucleotide polymorphism (SNP) frequencies to determine heterozygosity and chromosomal inheritance. Variant surface glycoprotein (VSG) genes and kinetoplast (mitochondrial) DNA maxi- and minicircles were extracted from each genome to examine how each of these components was inherited in the hybrid progeny. The same bioinformatic approaches were applied to an additional 37 genomes representing the diversity of T. brucei in subsaharan Africa and T. evansi. SNP analysis provided evidence of crossover events affecting all 11 pairs of megabase chromosomes and demonstrated that polyploid hybrids were formed post-meiotically and not by fusion of the parental diploid cells. VSGs and kinetoplast DNA minicircles were inherited biparentally, with approximately equal numbers from each parent, whereas maxicircles were inherited uniparentally. Extrapolation of these findings to field isolates allowed us to distinguish clonal descent from hybridization by comparing maxicircle genotype to VSG and minicircle repertoires. Discordance between maxicircle genotype and VSG and minicircle repertoires indicated inter-lineage hybridization. Significantly, some of the hybridization events we identified involved human infective and non-human infective trypanosomes circulating in the same geographic areas

μ2-Iodido-bis­{dimeth­yl[methyl­bis(quinolin-8-yl)silanyl-κ3 N,Si,N′]platinum(IV)} tetra­kis(penta­fluoro­phen­yl)borate dichloro­methane 0.66-solvate

The title complex, [Pt2(CH3)4(C19H15N2Si)2I][B(C6F5)4]·0.66CH2Cl2, resulted from an attempt to synthesize a stable five-coordinate platinum species via ligand abstraction of a six-coordinate platinum precursor. However, dimerization occurred after ligand abstraction, thereby yielding the compound described in this study. The cation is a dinuclear PtIV organometallic complex, in which the metal centers are bridged by an I− anion. Both metal centers display a coordination geometry close to octa­hedral, including cis-arranged quinoline ligands connected by Si atoms, which form Pt—Si bonds, two cis-methyl groups, and the bridging I− anion. In the crystal structure, voids between cations and anions are partially filled with an average of 0.66 mol­ecules of dichloro­methane solvent

Enhancement of macromolecular ice recrystallization inhibition activity by exploiting depletion forces

Antifreeze (glyco) proteins (AF(G)Ps) are potent inhibitors of ice recrystallization and may have biotechnological applications. The most potent AF(G)Ps function at concentrations a thousand times lower than synthetic mimics such as poly(vinyl alcohol), PVA. Here, we demonstrate that PVA’s ice recrystallization activity can be rescued at concentrations where it does not normally function, by the addition of noninteracting polymeric depletants, due to PVA forming colloids in the concentrated saline environment present between ice crystals. These depletants shift the equilibrium toward ice binding and, hence, enable PVA to inhibit ice growth at lower concentrations. Using theory and experiments, we show this effect requires polymeric depletants, not small molecules, to enhance activity. These results increase our understanding of how to design new ice growth inhibitors, but also offer opportunities to enhance activity by exploiting depletion forces, without re-engineering ice-binding materials. It also shows that when screening for IRI activity that polymer contaminants in buffers may give rise to false positive results

A Catalogue of Field Horizontal Branch Stars Aligned with High Velocity Clouds

We present a catalogue of 430 Field Horizontal Branch (FHB) stars, selected from the Hamburg/ESO Survey (HES), which fortuitously align with high column density neutral hydrogen (HI) High-Velocity Cloud (HVC) gas. These stars are ideal candidates for absorption-line studies of HVCs, attempts at which have been made for almost 40 years with little success. A parent sample of 8321 HES FHB stars was used to extract HI spectra along each line-of-sight, using the HI Parkes All-Sky Survey. All lines-of-sight aligned with high velocity HI emission with peak brightness temperatures greater than 120mK were examined. The HI spectra of these 430 probes were visually screened and cross-referenced with several HVC catalogues. In a forthcoming paper, we report on the results of high-resolution spectroscopic observations of a sample of stars drawn from this catalogue.Comment: 7 pages, 4 figures. ApJS accepted. Full catalogue and all online-only images available at http://astronomy.swin.edu.au/staff/cthom/catalogue/index.htm

High Metallicity Mg II Absorbers in the z < 1 Lyman alpha Forest of PKS 0454+039: Giant LSB Galaxies?

We report the discovery of two iron-group enhanced high-metallicity Mg II absorbers in a search through 28 Lyman Alpha forest clouds along the PKS 0454+039 sight line. Based upon our survey and the measured redshift number densities of W_r(MgII) <= 0.3 A absorbers and Lyman Alpha absorbers at z ~ 1, we suggest that roughly 5% of Lyman Alpha absorbers at z < 1 will exhibit "weak" Mg II absorption to a 5-sigma W_r(2796) detection limit of 0.02 A. The two discovered absorbers, at redshifts z = 0.6248 and z = 0.9315, have W_r(Lya) = 0.33 and 0.15 A, respectively. Based upon photoionization modeling, the H I column densities are inferred to be in the range 15.8 <= log N(HI) <= 16.8 cm^-2. For the z = 0.6428 absorber, if the abundance pattern is solar, then the cloud has [Fe/H] > -1; if its gas-phase abundance follows that of depleted clouds in our Galaxy, then [Fe/H] > 0 is inferred. For the z = 0.9315 absorber, the metallicity is [Fe/H] > 0, whether the abundance pattern is solar or suffers depletion. Imaging and spectroscopic studies of the PKS 0454+039 field reveal no candidate luminous objects at these redshifts. We discuss the possibility that these Mg II absorbers may arise in the class of "giant" low surface brightness galaxies, which have [Fe/H] >= -1, and even [Fe/H] >= 0, in their extended disks. We tentatively suggest that a substantial fraction of these "weak" Mg II absorbers may select low surface brightness galaxies out to z ~ 1.Comment: Accepted The Astrophysical Journal; 25 pages; 6 encapsulated figure

New Constraints on the Lyman Continuum Escape Fraction at z~1.3

We examine deep far-ultraviolet (1600 Angstrom) imaging of the Hubble Deep Field-North (HDFN) and the Hubble Ultra Deep Field (HUDF) to search for leaking Lyman continuum radiation from starburst galaxies at z~1.3. There are 21 (primarily sub-L*) galaxies with spectroscopic redshifts between 1.1<z<1.5 and none are detected in the far-UV. We fit stellar population templates to the galaxies' optical/near-infrared SEDs to determine the starburst age and level of dust attenuation, giving an accurate estimate of the intrinsic Lyman continuum ratio, f_1500/f_700, and allowing a conversion from f_700 limits to relative escape fractions. We show that previous high-redshift studies may have underestimated the amplitude of the Lyman Break, and thus the relative escape fraction, by a factor of ~2. Once the starburst age and intergalactic HI absorption are accounted for, 18 galaxies in our sample have limits to the relative escape fraction, f_esc,rel < 1.0 with some limits as low as f_esc,rel < 0.10 and a stacked limit of f_esc,rel < 0.08. This demonstrates, for the first time, that most sub-L* galaxies at high redshift do not have large escape fractions. When combined with a similar study of more luminous galaxies at the same redshift we show that, if all star-forming galaxies at z~1 have similar relative escape fractions, the value must be less than 0.14 (3 sigma). We also show that less than 20% (3 sigma) of star-forming galaxies at z~1 have relative escape fractions near unity. These limits contrast with the large escape fractions found at z~3 and suggest that the average escape fraction has decreased between z~3 and z~1. (Abridged)Comment: Accepted for publication in ApJ. aastex format. 39 pages, 11 figure