Function of microRNA-146a and NF-κB in physiologic and pathologic hematopoiesis

During inflammation and infection, hematopoietic stem and progenitor cells (HSPCs) are stimulated to proliferate and differentiate into mature immune cells, especially of the myeloid lineage. MicroRNA-146a (miR-146a) is a critical negative regulator of inflammation. Deletion of the gene encoding miR-146a—expressed in all blood cell types—produces effects that appear as dysregulated inflammatory hematopoiesis, leading to a decline in the number and quality of hematopoietic stem cells (HSCs), excessive myeloproliferation, and, ultimately, to exhaustion of the HSCs and hematopoietic neoplasms. Six-week-old deleted mice are normal, with no effect on cell numbers, but by 4 months bone marrow hypercellularity can be seen, and by 8 months marrow exhaustion is becoming evident. The ability of HSCs to replenish the entire hematopoietic repertoire in a myelo-ablated mouse also declines precipitously as miR-146a-deficient mice age. In the absence of miR-146a, LPS-mediated serial inflammatory stimulation accelerates the effects of aging. This chronic inflammatory stress on HSCs in deleted mice involves a molecular axis consisting of upregulation of the signaling protein TRAF6 leading to excessive activity of the transcription factor NF-κB and overproduction of the cytokine IL-6. At the cellular level, transplant studies show that the defects are attributable to both an intrinsic problem in the miR-146a-deficient HSCs and extrinsic effects of miR-146a-deficient lymphocytes and non-hematopoietic cells. This study has identified a microRNA, miR-146a, to be a critical regulator of HSC homeostasis during chronic inflammatory challenge in mice and has provided a molecular connection between chronic inflammation and the development of bone marrow failure and myeloproliferative neoplasms. This may have implications for human hematopoietic malignancies, such as myelodysplastic syndrome, which frequently displays downregulated miR-146a expression.

Conformal geometry processing

This thesis introduces fundamental equations and numerical methods for manipulating surfaces in three dimensions via conformal transformations. Conformal transformations are valuable in applications because they naturally preserve the integrity of geometric data. To date, however, there has been no clearly stated and consistent theory of conformal transformations that can be used to develop general-purpose geometry processing algorithms: previous methods for computing conformal maps have been restricted to the flat two-dimensional plane, or other spaces of constant curvature. In contrast, our formulation can be used to produce---for the first time---general surface deformations that are perfectly conformal in the limit of refinement. It is for this reason that we commandeer the title Conformal Geometry Processing.

The main contribution of this thesis is analysis and discretization of a certain time-independent Dirac equation, which plays a central role in our theory. Given an immersed surface, we wish to construct new immersions that (i) induce a conformally equivalent metric and (ii) exhibit a prescribed change in extrinsic curvature. Curvature determines the potential in the Dirac equation; the solution of this equation determines the geometry of the new surface. We derive the precise conditions under which curvature is allowed to evolve, and develop efficient numerical algorithms for solving the Dirac equation on triangulated surfaces.

From a practical perspective, this theory has a variety of benefits: conformal maps are desirable in geometry processing because they do not exhibit shear, and therefore preserve textures as well as the quality of the mesh itself. Our discretization yields a sparse linear system that is simple to build and can be used to efficiently edit surfaces by manipulating curvature and boundary data, as demonstrated via several mesh processing applications. We also present a formulation of Willmore flow for triangulated surfaces that permits extraordinarily large time steps and apply this algorithm to surface fairing, geometric modeling, and construction of constant mean curvature (CMC) surfaces.

Theory of ozone isotopic effects and various electron transfer reactions

Three separate topics, each stimulated by experiments, are treated theoretically in this dessertation: isotopic effects of ozone, electron transfer at interfaces, and intramolecular directional electron transfer in a supramolecular system.

The strange mass-independent isotope effect for the enrichment of ozone, which has been a puzzle in the literature for some 20 years, and the equally puzzling unconventional strong mass-dependent effect of individual reaction rate constants are studied as different aspects of a symmetry-driven behavior. A statistical (RRKM-based) theory with a hindered-rotor transition state is used. The individual rate constant ratios of recombination reactions at low pressures are calculated using the theory involving (1) small deviation from the statistical density of states for symmetric isotopomers, and (2) weak collisions for deactivation of the vibrationally excited ozone molecules. The weak collision and partitioning among exit channels play major roles in producing the large unconventional isotope effect in "unscrambled" systems. The enrichment studies reflect instead the non-statistical effect in "scrambled" systems. The theoretical results of low-pressure ozone enrichments and individual rate constant ratios obtained from these calculations are consistent with the corresponding experimental results. The isotopic exchange rate constant for the reaction ^(16)O + ^(18)O ^(18)O→+ ^(16)O ^(18)O + ^(18)O provides information on the nature of a variationally determined hindered-rotor transition state using experimental data at 130 K and 300 K. Pressure effects on the recombination rate constant, on the individual rate constant ratios and on the enrichments are also investigated. The theoretical results are consistent with the experimental data. The temperature dependence of the enrichment and rate constant ratios is also discussed, and experimental tests are suggested. The desirability of a more accurate potential energy surface for ozone in the transition state region is also noted.

Electron transfer reactions at semiconductor /liquid interfaces are studied using a tight-binding model for the semiconductors. The slab method and a z-transform method are employed in obtaining the tight-binding electronic structures of semiconductors having surfaces. The maximum electron transfer rate constants at Si/viologen^(2-/+) and InP /Me_(2)Fc^(+/O) interfaces are computed using the tight-binding type calculations for the solid and the extended-Huckel for the coupling to the redox agent at the interface. These electron transfer reactions are also studied using a free electron model for the semiconductor and the redox molecule, where Bardeen's method is adapted to calculate the coupling matrix element between the molecular and semiconductor electronic states. The calculated results for maximum rate constant of the electron transfer from the semiconductor bulk states are compared with the experimentally measured values of Lewis and coworkers, and are in reasonable agreement, without adjusting parameters. In the case of InP /liquid interface, the unusual current vs applied potential behavior is additionally interpreted, in part, by the presence of surface states.

Photoinduced electron transfer reactions in small supramolecular systems, such as 4-aminonaphthalimide compounds, are interesting in that there are, in principle, two alternative pathways (directions) for the electron transfer. The electron transfer, however, is unidirectional, as deduced from pH-dependent fluorescence quenching studies on different compounds. The role of electronic coupling matrix element and the charges in protonation are considered to explain the directionality of the electron transfer and other various results. A related mechanism is proposed to interpret the fluorescence behavior of similar molecules as fluorescent sensors of metal ions.

Electrical resistance and thermoelectric power of the metastable phases in tellurium-base alloys

By using techniques of rapid quenching from the melt, metastable phases have been obtained in ternary alloys which contain tellurium as a major component and two of the three noble metals (Cu, Ag, Au) as minor components. The metastable phases found in this investigation are either simple cubic or amorphous. The formation of the simple cubic phase is discussed. The electrical resistance and the thermoelectric power of the simple cubic alloy (Au₃₀Te₇₀) have been measured and interpreted in terms of atomic bondings. The semiconducting properties of a metastable amorphous alloy (Au₅Cu₂₅Te₇₀) have been measured. The experimental results are discussed in connection with a theoretical consideration of the validity of band theory in an amorphous solid. The existence of extrinsic conduction in an amorphous semiconductor is suggested by the result of electrical resistance and thermoelectric power measurements.

The ionic basis of frequency selectivity in hair cells of the bullfrog's sacculus

Hair cells from the bull frog's sacculus, a vestibular organ responding to substrate-borne vibration, possess electrically resonant membrane properties which maximize the sensitivity of each cell to a particular frequency of mechanical input. The electrical resonance of these cells and its underlying ionic basis were studied by applying gigohm-seal recording techniques to solitary hair cells enzymatically dissociated from the sacculus. The contribution of electrical resonance to frequency selectivity was assessed from microelectrode recordings from hair cells in an excised preparation of the sacculus.

Electrical resonance in the hair cell is demonstrated by damped membrane-potential oscillations in response to extrinsic current pulses applied through the recording pipette. This response is analyzed as that of a damped harmonic oscillator. Oscillation frequency rises with membrane depolarization, from 80-160 Hz at resting potential to asymptotic values of 200-250 Hz. The sharpness of electrical tuning, denoted by the electrical quality factor, Q_e, is a bell-shaped function of membrane voltage, reaching a maximum value around eight at a membrane potential slightly positive to the resting potential.

In whole cells, three time-variant ionic currents are activated at voltages more positive than -60 to -50 mV; these are identified as a voltage-dependent, non-inactivating Ca current (I_ca), a voltage-dependent, transient K current (I_a), and a Ca-dependent K current (I_c). The C channel is identified in excised, inside-out membrane patches on the basis of its large conductance (130-200 pS), its selective permeability to Kover Na or Cl, and its activation by internal Ca ions and membrane depolarization. Analysis of open- and closed-lifetime distributions suggests that the C channel can assume at least two open and three closed kinetic states.

Exposing hair cells to external solutions that inhibit the Ca or C conductances degrades the electrical resonance properties measured under current-clamp conditions, while blocking the A conductance has no significant effect, providing evidence that only the Ca and C conductances participate in the resonance mechanism. To test the sufficiency of these two conductances to account for electrical resonance, a mathematical model is developed that describes I_ca, I_c, and intracellular Ca concentration during voltage-clamp steps. I_ca activation is approximated by a third-order Hodgkin-Huxley kinetic scheme. Ca entering the cell is assumed to be confined to a small submembrane compartment which contains an excess of Ca buffer; Ca leaves this space with first-order kinetics. The Ca- and voltage-dependent activation of C channels is described by a five-state kinetic scheme suggested by the results of single-channel observations. Parameter values in the model are adjusted to fit the waveforms of I_ca and I_c evoked by a series of voltage-clamp steps in a single cell. Having been thus constrained, the model correctly predicts the character of voltage oscillations produced by current-clamp steps, including the dependencies of oscillation frequency and Q_e on membrane voltage. The model shows quantitatively how the Ca and C conductances interact, via changes in intracellular Ca concentration, to produce electrical resonance in a vertebrate hair cell.