MicroRNA-132 is a physiological regulator of hematopoietic stem cell function and B-cell development

MicroRNAs are a class of small non-coding RNAs that negatively regulate gene expression. Several microRNAs have been implicated in altering hematopoietic cell fate decisions. Importantly, deregulation of many microRNAs can lead to deleterious consequences in the hematopoietic system, including the onset of cancer, autoimmunity, or a failure to respond effectively to infection. As such, microRNAs fine-tune the balance between normal hematopoietic output and pathologic consequences. In this work, we explore the role of two microRNAs, miR-132 and miR-125b, in regulating hematopoietic stem cell (HSC) function and B cell development. In particular, we uncover the role of miR-132 in maintaining the appropriate balance between self-renewal, differentiation, and survival in aging HSCs by buffering the expression of a critical transcription factor, FOXO3. By maintain this balance, miR-132 may play a critical role in preventing aging-associated hematopoietic conditions such as autoimmune disease and cancer. We also find that miR-132 plays a critical role in B cell development by targeting a key transcription factor, Sox4, that is responsible for the differentiation of pro-B cells into pre-B cells. We find that miR-132 regulates B cell apoptosis, and by delivering miR-132 to mice that are predisposed to developing B cell cancers, we can inhibit the formation of these cancers and improve the survival of these mice. In addition to miR-132, we uncovered the role of another critical microRNA, miR-125b, that potentiates hematopoietic stem cell function. We found that enforced expression of miR-125b causes an aggressive myeloid leukemia by downregulation of its target Lin28a. Importantly, miR-125b also plays a critical role in inhibiting the formation of pro-B cells. Thus, we have discovered two microRNAs with important roles in regulating normal hematopoiesis, and whose dregulation can lead to deleterious consequences such as cancer in the aging hematopoietic system. Both miR-132 and miR-125b may therefore be targeted for therapeutics to inhibit age-related immune diseases associated with the loss of HSC function and cancer progression.

Investigations into the structure and function of the exocrine pancreas of lampreys. [Translation from: Morph.Jb. 110 245-269, 1967.]

As representatives of the most primitive of recent vertebrate groups, lampreys show fundamental differences in different features of organisation to the species of the remaining classes of vertebrates. The topical distinction between exocrine and endocrine pancreas is also considered among the morphological peculiarities of Petromyzontida. This study aims to contribute to a further explanation of this phenomenon. 50 brook lampreys were histologically examined.

Structure-Function Studies of Nicotinic Acetylcholine Receptors Using Selective Agonists and Positive Allosteric Modulators

This dissertation primarily describes chemical-scale studies of nicotinic acetylcholine receptors (nAChRs) in order to better understand ligand-receptor selectivity and allosteric modulation influences during receptor activation. Electrophysiology coupled with canonical and non-canonical amino acids mutagenesis is used to probe subtle changes in receptor function.

The first half of this dissertation focuses on differential agonist selectivity of α4β2-containing nAChRs. The α4β2 nAChR can assemble in alternative stoichiometries as well as assemble with other accessory subunits. Chapter 2 identifies key structural residues that dictate binding and activation of three stoichiometry-dependent α4β2 receptor ligands: sazetidine-A, cytisine, and NS9283. These do not follow previously suggested hydrogen-bonding patterns of selectivity. Instead, three residues on the complementary subunit strongly influence binding ability of a ligand and receptor activation. Chapter 3 involves isolation of a α5α4β2 receptor-enriched population to test for a potential alternative agonist binding location at the α5 α4 interface. Results strongly suggest that agonist occupation of this site is not necessary for receptor activation and that the α5 subunit only incorporates at the accessory subunit location.

The second half of this dissertation seeks to identify residue interactions with positive allosteric modulators (PAMs) of the α7 nAChR. Chapter 4 focuses on methods development to study loss of potentiation of Type I PAMs, which indicate residues vital to propagation of PAM effects and/or binding. Chapter 5 investigates α7 receptor modulation by a Type II PAM (PNU 120596). These results show that PNU 120596 does not alter the agonist binding site, thus is relegated to influencing only the gating component of activation. From this, we were able to map a potential network of residues from the agonist binding site to the proposed PNU 120596 binding site that are essential for receptor potentiation.

Comparative investigations on the respiratory physiology of euryhaline gammarids with special reference to salinity adaptation. [Translation from: Helgolaender Wissenschaftliche Meeresuntersuchungen 23(4) 485-534, 1972.]

Members of the family Gammaridae are very closely interrelated. There arises the question as to how far they also differ amongst themselves through physiological characteristics. Comparative respiratory and physiological experiments were made on the five euryhaline species Gammarus locusta, G. oceanicus, G. salinus, G. zaddachi and G. duebeni. The respiratory measurements carried out within the framework of this experiment were occupied with the relationships between oxygen consumption and body size depending on salinity. They also had the object of determing the variations in metabolic intensity after an abrupt change in the salt content of the external medium, and to establish the period of time for the process of adaptation. As the experiments were carried out polarographically in a testing plant with continuous flow-through, and the method which was applied permitted continuous recording over prolonged intervals, there could also be carried out comparisons between metabolism at rest and under activity, and the alterations of oxygen consumption during the process of moulting could be measured.

Optical interconnection for neural networks by use of a self-imaging function

A grating-lens combination unit is developed to form a scaling self-transform function that can self-image on scale. Then an array of many such grating-lens units is used for the optical interconnection of a two-dimensional neural network, and experiments are carried out. We find that our idea is feasible, the optical interconnection system is simple, and optical adjustment is easy. (C) 1998 Optical Society of America.

Computation of the ambiguity function for circularly symmetric pupils

A method of computing the ambiguity function (AF) for a circularly symmetric pupil function is presented. The AFs of a clear aperture and two shaded apertures are considered in detail and an explicit expression for the first of these AFs is given. We explain these results in the context of the well-known optical transfer function theory and show a primary application of these computations. A good analytic approximation is also introduced, providing an alternative method for calculating the AF, in a simpler way.

Organization and function of the phage Lambda chromosome

The process of prophage integration by phage λ and the function and structure of the chromosomal elements required for λ integration have been studied with the use of λ deletion mutants. Since att^φ, the substrate of the integration enzymes, is not essential for λ growth, and since att^φ resides in a portion of the λ chromosome which is not necessary for vegetative growth, viable λ deletion mutants were isolated and examined to dissect the structure of att^φ.

Deletion mutants were selected from wild type populations by treating the phage under conditions where phage are inactivated at a rate dependent on the DNA content of the particles. A number of deletion mutants were obtained in this way, and many of these mutants proved to have defects in integration. These defects were defined by analyzing the properties of Int-promoted recombination in these att mutants.

The types of mutants found and their properties indicated that att^φ has three components: a cross-over point which is bordered on either side by recognition elements whose sequence is specifically required for normal integration. The interactions of the recognition elements in Int-promoted recombination between att mutants was examined and proved to be quite complex. In general, however, it appears that the λ integration system can function with a diverse array of mutant att sites.

The structure of att^φ was examined by comparing the genetic properties of various att mutants with their location in the λ chromosome. To map these mutants, the techniques of heteroduplex DNA formation and electron microscopy were employed. It was found that integration cross-overs occur at only one point in att^φ and that the recognition sequences that direct the integration enzymes to their site of action are quite small, less than 2000 nucleotides each. Furthermore, no base pair homology was detected between att^φ and its bacterial analog, att^B. This result clearly demonstrates that λ integration can occur between chromosomes which have little, if any, homology. In this respect, λ integration is unique as a system of recombination since most forms of generalized recombination require extensive base pair homology.

An additional study on the genetic and physical distances in the left arm of the λ genome was described. Here, a large number of conditional lethal nonsense mutants were isolated and mapped, and a genetic map of the entire left arm, comprising a total of 18 genes, was constructed. Four of these genes were discovered in this study. A series of λdg transducing phages was mapped by heteroduplex electron microscopy and the relationship between physical and genetic distances in the left arm was determined. The results indicate that recombination frequency in the left arm is an accurate reflection of physical distances, and moreover, there do not appear to be any undiscovered genes in this segment of the genome.

Studies on the role of histones in the structure and function of chromatin

Studies on the dissociation of histones from chromatin by increasing concentrations of sodium deoxycholate (DOC) have shown that histrone II is removed at lowest concentrations of DOC, while slightly higher concentrations remove histones III and IV. Still higher concentrations remove histone I.

The complete separation of chromatin and ¹⁴C-DOC by sucrose sedimentation indicated that the binding of DOC to chromatin is readily and completely reversible.

The dissociation of histones from chromatin by increasing concentrations of related cholanic acids and some of their conjugated derivatives were studied. The results suggested that the driving force for the interaction between the cholanic acid anion and histones is the lowering of the activity coefficient of the cholanic acid anion which occurs when it is partially removed from solution by interaction with hydrophobic regions of the positively charged histones.

The role of histones in the structure of chromatin has been studied by comparing the effects of selective removal of histones from chromatin by increasing concentrations of DOC with those caused by NaCl (removes histone I at lowest concentrations, while higher concentrations remove histones II, III, and IV). Properties studied included thermal denaturation, sedimentation velocity, flow dichroism, relaxation times of molecules oriented in a flow field, and the irreversible disruption of a 130 S, cross-linked component of sheared chromatin. The data indicated that none of the structural or chemical parameters with which these properties are correlated show a dependence on the presence of one particular histone fraction.

The template activity (ability to prime a 0.2 M KC1 DNA-dependent RNA synthesis system catalyzed by E. coli RNA polymerase) increases from that of native chromatin (approximately 25 per cent of that pure DNA) to that of pure DNA in a fashion which shows a nearly linear relationship to the amount of histone coverage of the template. The precipitability of partially dehistonized chromatin samples in 0.15 M NaCl shows a large dependence on the presence of histone I.

Part I: Latent heat of vaporization of n-decane. Part II: Heat and mass transfer from a cylinder placed in a turbulent air stream - Sherwood and Frössling numbers as a function of free stream turbulence level and Reynolds number

Part I

The latent heat of vaporization of n-decane is measured calorimetrically at temperatures between 160° and 340°F. The internal energy change upon vaporization, and the specific volume of the vapor at its dew point are calculated from these data and are included in this work. The measurements are in excellent agreement with available data at 77° and also at 345°F, and are presented in graphical and tabular form.

Part II

Simultaneous material and energy transport from a one-inch adiabatic porous cylinder is studied as a function of free stream Reynolds Number and turbulence level. Experimental data is presented for Reynolds Numbers between 1600 and 15,000 based on the cylinder diameter, and for apparent turbulence levels between 1.3 and 25.0 per cent. n-heptane and n-octane are the evaporating fluids used in this investigation.

Gross Sherwood Numbers are calculated from the data and are in substantial agreement with existing correlations of the results of other workers. The Sherwood Numbers, characterizing mass transfer rates, increase approximately as the 0.55 power of the Reynolds Number. At a free stream Reynolds Number of 3700 the Sherwood Number showed a 40% increase as the apparent turbulence level of the free stream was raised from 1.3 to 25 per cent.

Within the uncertainties involved in the diffusion coefficients used for n-heptane and n-octane, the Sherwood Numbers are comparable for both materials. A dimensionless Frössling Number is computed which characterizes either heat or mass transfer rates for cylinders on a comparable basis. The calculated Frössling Numbers based on mass transfer measurements are in substantial agreement with Frössling Numbers calculated from the data of other workers in heat transfer.

Analysis of optical systems with extended depth of field using the Wigner distribution function

We describe the use of a Wigner distribution function approach for exploring the problem of extending the depth of field in a hybrid imaging system. The Wigner distribution function, in connection with the phase-space curve that formulates a joint phase-space description of an optical field, is employed as a tool to display and characterize the evolving behavior of the amplitude point spread function as a wave propagating along the optical axis. It provides a comprehensive exhibition of the characteristics for the hybrid imaging system in extending the depth of field from both wave optics and geometrical optics. We use it to analyze several well-known optical designs in extending the depth of field from a new viewpoint. The relationships between this approach and the earlier ambiguity function approach are also briefly investigated. (c) 2006 Optical Society of America.

Connectivity and Function of the Primate Insula

The insula is a mammalian cortical structure that has been implicated in a wide range of low- and high-level functions governing one’s sensory, emotional, and cognitive experiences. One particular role of this region is considered to be processing of olfactory stimuli. The ability to detect and evaluate odors has significant effects on an organism’s eating behavior and survival and, in case of humans, on complex decision making. Despite such importance of this function, the mechanism in which olfactory information is processed in the insula has not been thoroughly studied. Moreover, due to the structure’s close spatial relationship with the neighboring claustrum, it is not entirely clear whether the connectivity and olfactory functions attributed to the insula are truly those of the insula, rather than of the claustrum. My graduate work, consisting of two studies, seeks to help fill these gaps. In the first, the structural connectivity patterns of the insula and the claustrum in a non-human primate brain is assayed using an ultra-high-quality diffusion magnetic resonance image, and the results suggest dissociation of connectivity — and hence function — between the two structures. In the second study, a functional neuroimaging experiment investigates the insular activity during odor evaluation tasks in humans, and uncovers a potential spatial organization within the anterior portion of the insula for processing different aspects of odor characteristics.

Fast Lattice Green's Function Methods for Viscous Incompressible Flows on Unbounded Domains

In this thesis, a collection of novel numerical techniques culminating in a fast, parallel method for the direct numerical simulation of incompressible viscous flows around surfaces immersed in unbounded fluid domains is presented. At the core of all these techniques is the use of the fundamental solutions, or lattice Green’s functions, of discrete operators to solve inhomogeneous elliptic difference equations arising in the discretization of the three-dimensional incompressible Navier-Stokes equations on unbounded regular grids. In addition to automatically enforcing the natural free-space boundary conditions, these new lattice Green’s function techniques facilitate the implementation of robust staggered-Cartesian-grid flow solvers with efficient nodal distributions and fast multipole methods. The provable conservation and stability properties of the appropriately combined discretization and solution techniques ensure robust numerical solutions. Numerical experiments on thin vortex rings, low-aspect-ratio flat plates, and spheres are used verify the accuracy, physical fidelity, and computational efficiency of the present formulations.

The ecological impact of different mechanisms of chronic sub-lethal toxicity on feeding and respiratory physiology

Sub-lethal toxicity tests, such as the scope-for-growth test, reveal simple relationships between measures of contaminant concentration and effect on respiratory and feeding physiology. Simple models are presented to investigate the potential impact of different mechanisms of chronic sub-lethal toxicity on these physiological processes. Since environmental quality is variable, even in unimpacted environments, toxicants may have differentially greater impacts in poor compared to higher quality environments. The models illustrate the implications of different degrees and mechanisms of toxicity in response to variability in the quality of the feeding environment, and variability in standard metabolic rate. The models suggest that the relationships between measured degrees of toxic stress, and the maintenance ration required to maintain zero scope-for-growth, may be highly nonlinear. In addition it may be possible to define critical levels of sub-lethal toxic effect above which no environment is of sufficient quality to permit prolonged survival.