Browsing by Subject "Topography"

The topographical properties of a landscape and their time evolution are key features of the Earth's surface, regulating ecosystem functioning in terms of soil properties as well as water and energy budgets, and creating visually diverse and striking patterns across various spatial scales. Furthermore, the natural evolution of a topography under the influence of geologic erosion can be greatly altered by anthropogenic disturbances (e.g., through agriculture, mining, deforestation), with the potential of accelerating soil erosion, causing land degradation and soil fertility losses. Hence, understanding the geomorphological processes driving the evolution of landscapes under natural and disturbed conditions is key not only to define the main factors and feedbacks shaping the Earth's topography, but also to foresee the consequences of intensive land use and implement optimal strategies of land management and recovery.

This dissertation addresses some key aspects of landscape evolution and stability, with a focus on the statistical description and modeling of hillslope morphologies under natural and disturbed conditions, the theoretical definition of drainage area at regular and non-regular points of the watershed, and the formation of spatially organized ridge and valley patterns.

We start from the analysis of topographic slopes under natural and accelerated soil erosion. Using large topographic datasets from mountain ranges worldwide, we show that the approximate age of a landscape is fingerprinted in the tails of its slope distributions. We then explore the role of the different processes driving this dynamic smoothing over geologic time scales by means of numerical experiments, showing that the relaxation process is mainly dominated by diffusion. The effect of agricultural-driven soil erosion on hillslope morphology is then investigated, highlighting how the natural aging process can be altered by intensive land use which, at smaller scales, produces key differences in the slope distribution tails. Furthermore, theoretical solutions are derived for the hillslope profile and the associated soil creep and runoff erosion fluxes, and used to link the observed differences in the morphological features of disturbed and undisturbed areas to a disruption of the natural balance between soil creep and runoff erosion mechanisms.

We then move the analysis to the drainage area, an important nonlocal morphometric variable used in a large number of geomorphological and ecohydrological applications. A nonlinear differential equation whose validity is limited to regular points of the watershed is obtained from a continuity equation, and the theory is then extended to critical and singular points by means of both Gauss' theorem and dynamical systems concepts. Such a link between the drainage area and a continuity equation sets the basis for the subsequent analysis of organized ridge and valley patterns and channel forming instability. The formation of ridge/valley patterns is analyzed by means of numerical experiments in detachment limited conditions, with the identification of various regimes as a function of diffusive soil creep, runoff erosion, and tectonic uplift as well as the specific geomorphic transport law assumed. Lastly, a linear stability analysis of the coupled water and landscape evolution dynamics is outlined to investigate the critical conditions triggering channel formation and the emergence of characteristic valley spacings in relation to the main geomorphological processes involved.

Regenerative medicine holds the promise of providing relief for people suffering from diseases where treatment has been unattainable. The research is advancing rapidly; however, there are still many hurdles to overcome before the therapeutic potential of regenerative medicine and cell therapy can be realized. Low in frequency in all tissues, stem cell number is often a limiting factor. Approaches that can control the proliferation and direct the differentiation of stem cells would significantly impact the field. Developing an adequate environment that mimics in vivo conditions is an intensively studied topic for this purpose. Collaboratively, researchers have come close to incorporating nearly all biological cues representative of the human body. Arguably the most overlooked aspect is the influence of electrical stimulation. In this dissertation, we examined polyvinylidene fluoride (PVDF) as a new biomaterial and developed a 3D scaffold capable of providing mechanical and electrical stimuli to cells in vitro.

The fabrication of a 3D scaffold was performed using electrospinning. To obtain highly aligned fibers and scaffolds with controlled porosity, the set-up was modified by incorporating an auxiliary electrode to focus the electric field. Highly aligned fibers with diameters ranging from 500 nm to 15 µm were fabricated from colorless polyimide (CP2) and polyglycolic acid (PGA) and used to construct multilayer scaffolds. This experimental set-up was used to electrospin α-phase PVDF into the polar β-phase. We demonstrated the transition to the β-phase by examining the crystalline structure using x-ray diffraction (XRD), differential scanning calorimetry (DSC), fourier transform infrared spectroscopy (FTIR) and polarized light optical microscopy (PLOM). We confirmed these results by observing a polarization peak at 80°C using the thermally stimulated current (TSC) method. Our results proved the electrospinning process used in our investigation poled the PVDF polymer in situ.

TThe influence of architecture and topographical cues was examined on 3D scaffolds and films of CP2 polyimide and PVDF. Culture of human mesenchymal stem cells (hMSCs) for 7 and 14 days demonstrated a significant difference in gene expression. The fibers upregulated the neuronal marker microtubule associated protein (MAP2), while downregulation of this protein was observed on films. Gap junction formation was observed by the expression of connexin-43 after 7 days on PVDF films attributed to its inherent pyroelectric properties. Connexin-43 expression on fibers showed cell-cell contact across the fibers indicating good communication in our 3D scaffold.

A scaffold platform was designed using PVDF fibers that allowed us to apply electrical stimulation to the cells through the fibers. The electrically stimulated PVDF fibers resulted in enhanced proliferation compared to TCPS as evidenced by a 10% increase in the uptake of EdU. Protein expression revealed upregulation of neuronal marker MAP2. Our findings indicate this new platform capable of delivering mechanical, electrical, topographical and biochemical stimuli during in vitro culture holds promise for the advancement of stem cell differentiation and tissue engineering.

In this dissertation I compile modern mammalian faunal lists, as well as ecomorphological measurements on living marsupials and rodents, to relate the diversity of small mammals, specifically the distributions of their dental topographies, to the climates in which they are found. The emphasis of this dissertation is to demonstrate the potential of distributions of dental topography metrics as proxies for the reconstruction of paleoenvironments in the Miocene of South America.

In Chapter 2, I compile complete, non-volant mammalian species lists for 85 localities across South America as well as 17 localities across Australia and New Guinea. Climatic and habitat variables were also recorded at each locality using GIS spatial data. Additionally, basic ecological data was collected for each species, including: diet, body size, and mode of locomotion. Niche indices that describe the relative numbers of different ecologies were calculated for each locality. These indices then served as the predictor values in a handful of regression models, including regression trees, random forests, and Gaussian process regression. The Australian/New Guinean localities were used as a geographically and phylogenetically independent for the purposes of testing the models derived from South America.

As for the dental ecomorphological analysis, I use three separate measures of dental topography, each of which measures a different component of dental topography; relief (the Relief Index, or RFI), complexity (orientation patch count rotated, OPCR), and sharpness (Dirichlet normal energy, DNE). Together, these metrics quantify the shape of the tooth surface without regard for tooth size. They also do not depend on homologous features on the tooth surface for comparative analysis, allowing a broad taxonomic sample as I present here. After a methodological study of DNE in Chapter 3, I present correlative studies of dental topography and dietary ecology in marsupials and rodents in Chapters 4 and 5, respectively. Finally, using the same localities from Chapter 2, I analyze the distributions of dental topography metrics as they relate to climate and habitat.

Results suggest that sharpness and relief are positively correlated with a higher amount of tough foods—such as leaves or insects—in the diet of marsupials, and that relief is positively correlated with grass-eating in rodents. The distributions of all three metrics show some utility when used as a proxy for climatic variables, though the distributions of RFI in marsupials and OPCR in rodents demonstrate the best correlations.

Overall, this dissertation suggests that dental topography can be used to discriminate dietary categories in a wide variety of mammalian groups, and that the distributions of dental ecometrics can be used as proxies for paleoenvironment reconstruction. This may eliminate the need to reconstruct behavior in individual taxa in order to construct ecological indices for fossil mammalian communities, thus offering a more direct avenue to reconstructing past environments.

The recent discovery that fibroblasts can be reprogrammed directly to functional neurons with lentivirus has reinvigorated the belief that autologous human cell therapies against many neurodegenerative diseases may be achievable in the near future. To increase the clinical translatability of this approach, we have developed a technique to perform this direct conversion without the use of virus. We transfected fibroblasts with plasmids condensed into non-viral nanoparticulate carriers with a bioerodible peptidomimetic polymer, pCBA-ABOL. Gene delivery with pCBA-ABOL was exceptionally effective and nontoxic, producing high transfection efficiencies and enabling serial dosing of plasmid cocktails. We delivered plasmids encoding neural lineage-instructive transcription factors to primary mouse embryonic fibroblasts (PMEFs), eliciting: drastic morphological changes, activation of endogenous neuronal transcription factor expression, neuronal promoter activity, and the appearance of neuronal proteins. Serial dosing of pCBA-ABOL complexes produced increasingly higher yields of these non-virally induced neurons (NiNs). The majority of NiNs fired action potentials under patch clamp. This is the first description of a method capable of directly inducing functional neuronal cells from fibroblasts without the use of virus.

We then moved on to further increase the yield of NiN generation, in an effort to produce a sufficient quantity of neurons for cell therapies. Informed by neurodevelopmental cues and by precedents set by the induced pluripotent stem cell (iPSC) field, we performed non-viral neuronal reprogramming in the presence of soluble microenvironmental factors that either inhibited GSK-3beta; and SMAD signaling, or induced chronic membrane depolarization. Chronic depolarization doubled the number of cells expressing neuronal markers produced with glutamatergic as well as with dopaminergic transcription factor cocktails. Inhibition of GSK-3beta; and SMAD signaling similarly doubled the yield of glutamatergic NiNs, and enabled induction of neuronal markers and morphological transformation in human fibroblasts.

In addition to soluble microenvironmental factors, the physical microenvironment can also strongly influence various cellular phenotypes. This list includes many phenotypes related to endocytosis - the transit mechanism of nanoparticulate gene carriers entering cells during non-viral transfection. As such, we set out to determine if patterned physical substrate topography could be used to increase non-viral transfection. We employed a high-throughput screen of micropitted substrate topographies, and indeed found that larger, denser micropits could support significantly higher transfection efficiencies in fibroblasts, compared to smooth substrates. The same topographies produced higher reverse transfection efficiencies, and greater siRNA-mediated knockdown of a reporter gene. The control of transfection with substrate topography is a new design parameter that could find broad application in in vitro non-viral reprogramming strategies, as well as in the intelligent design of nucleic acid-eluting scaffolds in vivo.

Encouraged by our success with direct neuronal reprogramming, and armed with a greater understanding of some microenvironmental mediators thereof, we attempted to produce non-virally-induced oligodendroglial progenitor cells (NiOPCs), which has been historically challenging for other investigators. We discovered the fibroblastic intracellular microenvironment is a significant barrier to the function of Olig2 - a master regulator of OPC phenotype - in direct reprogramming. Fibroblasts do not express Olig2 chaperones which are normally expressed in OPCs, causing Olig2 to become sequestered in the cytoplasm of transfected PMEFs. We relieved this barrier through fusion of a strong nuclear localization sequence (NLS) to Olig2, which repartitioned Olig2-NLS from the cytoplasm to the nucleus in transfected fibroblasts. The use of Olig2-NLS in iOPC reprogramming cocktails resulted in a drastic improvement in the yield of OPC-specific marker expression. The improvement associated with Olig2-NLS was insufficient to elicit significant myelin protein expression with the non-viral system, but the yield of virally-induced oligodendrocyte-like cells (iOLs) was improved dramatically. Further enhancements will be required to generate fully-reprogrammed NiOPCs, but the increased efficiency of viral iOPC generation is immediately useful for disease modeling and potentially in cell replacement therapies if human cells can be converted for the first time using this technique. During direct reprogramming, CNS-specific transcription factors are delivered to foreign intracellular contexts as a rule, which may reduce their ability to function effectively; we have shown this can be a critical yet under-appreciated determinant of the success or failure of a direct reprogramming system.

Taken together, the technological and intellectual advancements we describe herein represent significant improvements to non-viral transfection and reprogramming systems. These techniques can find broad appeal to the many researchers and clinicians deploying these systems. More specifically, we present significant steps towards realization of the dream of safe and effective autologous cell therapies against devastating and currently-intractable neurodegenerative diseases.