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Fusobacterium nucleatum is an invasive anaerobic bacterium, which is ubiquitous in human dental plaque and is frequently associated with periodontal diseases. Previous studies have focused on the virulence factors produced during infection of F. nucleatum and disease progression, but early recognition by the immune system and the role of the inflammasome in response to bacteria is not well characterized. Furthermore, although an inflammasome in gingival epithelial cells (GECs) can be stimulated by danger-associated molecular pattern molecules (DAMPs) such as ATP, inflammasome activation by this periodontal pathogen has yet to be described in these cells. We aimed to examine the effects of F. nucleatum infection on pro-inflammatory cytokine expression and inflammasome activation in GECs. Our results indicate that infection induces translocation of NF-κB into the nucleus, resulting in cytokine gene expression. In addition, infection activates the NLRP3 inflammasome, which in turn activates caspase-1 and release of mature IL-1β. Unlike other pathogens studied until now, F. nucleatum activates the inflammasome in GECs in the absence of exogenous DAMPs. Finally, infection promotes release of other DAMPs mediating inflammation, such as high-mobility group protein B1 (HMGB1) and apoptosis-associated speck-like protein, with a similar time-course as caspase-1 activation. Thus, F. nucleatum expresses the pathogen-associated molecular patterns (PAMP) necessary to activate NF- κB, and also provides an endogenous DAMP to stimulate the inflammasome and further amplify inflammation through secretion of secondary DAMPs. Our findings can provide novel pathway of F. nucleatum invasion and improve our understanding of its roles in periodontal infection. Thus, research on pathogenic mechanisms and the hosts’ immune system can postulate the relationship between oral pathogens and emerging systemic diseases.

Disulfide bonds are an important post-translational modification that provides stability for many proteins. In the bacterium Escherichia coli, oxidative protein folding is carried out in the periplasm by the thiol-disulfide oxidoreductase proteins. The thiol disulfide oxidoreductase DsbA is responsible for the initial generation of disulfide bonds into oxidatively folding proteins. DsbA is maintained in a catalytic oxidized state by the inner membrane protein DsbB, which channels electrons from DsbA to the electron transport chain through quinone reduction. My plan was to investigate if mutation and selection/screening could generate possible substitutes for the normal disulfide formation pathway. This may provide insight into what minimal requirements define a thiol disulfide oxidase. First, we took the disulfide reductase thioredoxin, a distantly related homologue to DsbA, and selected for mutants in the active site CXXC motif that when exported to the periplasm through the twin-arginine pathway could complement a DeltadsbB strain. We were able to show that mutation of the active site from CGPC to either CACC or CPCC resulted in variants that were capable of partial complementation of the defect in disulfide formation seen in the DeltadsbB strain; surprisingly, the mutant thioredoxin had acquired an iron sulfur cluster. These results demonstrated that an iron sulfur-containing thioredoxin could function to complement the entire disulfide formation pathway. Second, I discovered mutations in DsbB that bypassed the need for DsbA by utilizing DsbC as a catalyst for forming disulfide bonds. DsbC is one of the two putative disulfide isomerases in the periplasm. This work suggests that in addition to functioning as an isomerase, DsbC can also function as an oxidase. Finally, we have been able to isolate a chromosomal mutant that is able to bypass the need for both DsbA and DsbB. The strain suppresses all tested disulfide phenotypes, and is not dependent on any Dsb protein except DsbC. Initial mapping locates the source of suppression to an area on the chromosome corresponding to Complex I of the electron transport chain. These results reveal a surprising variety of ways that the cell can compensate for the loss of the normal disulfide catalytic machinery. In addition, this work may point to novel functions for the thiol disulfide isomerase DsbC.

An efficient and accurate hybrid modeling is presented for the analysis of electromagnetic interference (EMI) effects on RF/microwave components enclosed within multilayer cavities. The method of moments (MoM) is employed in conjunction with rigorous modal Green's function (GF) (the MoM-GF technique) to account for EM interactions within rectangular cavity enclosure containing typical microwave structures. We considered structures with wire configurations and multilayer printed circuit boards (PCBs) that support possible devices such as filters and amplifiers. For the analysis of enclosed cable and wire configurations, a challenge was the convergence of the associated moment method matrices and two approaches are presented. One is based on the Kummer's transformation technique to accelerate the sum associated with the double modal series used in the representation of the cavity GFs. The other is a hybrid method utilizing a two-step decomposition to account for the interactions between enclosures and internal conductors. Rigorous analysis of multilayer printed circuits within cavity enclosures with multiple apertures and penetrating cables is considered. As part of this study, a process is developed to account for the interactions of vertical and planar currents within a multilayered substrate configuration enclosed within the structure using efficient and rather fast matrix element evaluations. Using this procedure, EMI effects through multiple apertures are investigated via a parametric study for different circuit geometries and lumped loads. For the analysis of PCBs within a multilayered and possibly cascaded multi-cavity configuration, a simple aperture coupling method is developed. This approach allows for the concurrent analysis of a large number of vertical connections and planar metallization within each layer interface. An important component of the proposed EMI investigation is the evaluation of coupling on the PCBs in the presence of active RF circuits such as power amplifiers. This is done through a decomposition approach that integrates the MoM-GF technique with a SPICE-like circuit solver. The validity and flexibility of the proposed modeling technique is demonstrated for various cavity geometries and internal printed circuit components subject to external EM illuminations.

Wnt pathways are highly conserved pathways that regulate key processes such as cell type specification and migration of cells during animal development. The C. elegans vulva displays symmetry such that the posterior half of the vulva forms a mirror image of the anterior half. We observed that nuclear levels of POP-1 also display similar mirror symmetry in the developing vulva. We are studying the processes that establish this mirror symmetric pattern of POP-1 localization. Because mutations in the putative WNT receptors lin-17/frizzled and lin-18/Ryk were known to cause defects in the posterior half of the vulva, we examined POP-1 localization patterns in these mutants. We observed that POP-1 localization is reversed in the posterior most part of the vulva with respect to wild-type, in these mutants. lin-17/lin-18 double mutants show a more penetrant reversal of POP-1 localization, suggesting that LIN-17 and LIN-18 act in a semi-redundant manner to reorient POP-1 localization in the posterior half of the vulva. We have investigated the mechanisms by which two other P ectoblasts, P11 and P12 are specified. Prior studies have shown that lin-44/Wnt, lin-17/Frizzled and bar-1/canonical b-catenin function in P12 specification. We have obtained additional evidence that a canonical Wnt pathway functions in P12 specification. We have demonstrated that activated forms of BAR-1 (deltaNBAR-1) are sufficient to cause a P11 to P12 transformation. Interestingly, time-course studies with deltaNBAR-1 expression suggest that Wnts may act at the same time as the EGF pathway, suggesting the requirement of two parallel pathways. We predict a function for POP-1 in P12 specification as BAR-1 and b-catenins in other organisms are known to act primarily through TCFs to activate transcription. We found that the pop-1 hypomorphs, hu9 and q645 that should reduce bar-1 activity cause a low rate of P12 to P11 transformations. This supports the hypothesis that POP-1 acts in P12 specification in a BAR-1 dependent manner. Surprisingly, we find that pop-1 RNAi causes a P11 to P12 transformation, revealing a second function of POP-1 in P11 specification. This antagonistic function of POP-1 appears to be independent of BAR-1 as observed by epistasis experiments. We find that multiple Wnts act in P12 specification. lin-44(n1792/Wnt) and egl-20(n585)/Wnt double mutants display an enhanced rate of P12 to P11 transformations. Genetic analysis using other Wnts and Wnt receptors possibly functioning with LIN-44 and LIN-17 in P12 specification will be discussed.

Canonical correlation analysis is an extremely useful technique, especially in biomedical investigations where a large number of variables of distinct types, e.g., physical and psychological, are studied simultaneously. Discriminant analysis and regression analysis are particular cases of this general technique. In this dissertation the following aspects of canonical correlation analysis are investigated: (i) Anderson's classification statistic, (ii) assignment of optimum scores to the categories of a multidimensional contingency table, by using a stepwise method for completing canonical analysis for more than two vector variables, and (iii) the predictive ability of scores in contingency tables. Anderson's classification statistic is expressed in a simpler form by using a series of random orthogonal transformations. Its first two exact moments are derived which are then utilized in obtaining a normal approximation to its distribution. The usefulness of this statistic in estimation of relative potency of a new drug or test preparation is illustrated. Assignment of scores to the categories of a contingency table by use of a stepwise method of canonical correlations is proposed and illustrated for the well known blood serological data of Taylor, first analyzed by Fisher. The efficiency of this method is investigated using redundancy analysis indices. The stepwise method also provides a technique for eliminating categories that do not contribute significantly to the relationship. The predictive ability of the optimum scores obtained by using the canonical variables is investigated for the Taylor data, wherein predicted and observed values agreed in almost 80% of the cases.

In the first part of this Dissertation we define and study certain almost complex structures (a.c.s.) in tangent bundles of Riemannian manifolds, which we call isotropic and are integrable, i.e., complex structures, precisely when ${\cal M}$ has constant sectional curvatures.(cf. (1)) We characterize those isotropic integrable a.c.s. that are invariant by the full isometry group of ${\cal M}$, acting by tangent maps, in terms of a two parameter family of real functions of the energy function. We show that in the tangent bundle of hyperbolic space, there are isotropic integrable almost complex structures that are not invariant in the above sense. In the second part of this dissertation we define the real analogue of the Lempert-Szoke-Guillemin-Stenzel canonical adapted complex structure on tubular neighborhoods of the zero section in the tangent bundle of a real analytic Riemannian manifold introduced in (2) (the Grauert tube of ${\cal M}$.) This dual is the adapted product structure and is geometrically interpreted in terms of the shape operator of geodesic spheres. We define a CR analogue of the Anosov splitting, as a limit of a rescaled version of the complex structure. This CR limit, when it exists, is a geodesic flow invariant CR structure in the unit tangent bundle of ${\cal M}$. We show that among symmetric spaces of compact type and a certain family of metrics of revolution on the two sphere, such CR limit exists precisely for the rank symmetric cases and relate this to the thickening of the Gauss map for such metrics of revolution. We generalize, as follows, Stenzel's reduction of the equation for a Ricci-flat Kahler metric in the Grauert tubes of rank 1 symmetric spaces of compact type to an ODE found in (4). We establish a real analogue of a Kahler metric where the complex structure is replaced by a product structure P, and a non-degenerate symmetric two tensor, anti-invariant by P. The equation Einstein pseudo-Riemannian metrics on tubes containing the zero section in the tangent bundle reduces to an ODE when ${\cal M}$ is an harmonic manifold. Upon complexification, the equation for a Kahler-Einstein metric on the Grauert tubes of harmonic Riemannian manifolds reduces to an ODE and we study the completeness of the metrics obtained.

A data-based procedure is proposed to guide exploratory analysis to determine which categories of a multiway contingency table may be grouped. This method is based on the application of canonical analysis for several sets of variables to the multiway contingency table. The extension of this approach to several groups of multiway tables with similar canonical scores is considered. Next, a comparison with Fisher's approach in assigning canonical scores to the non-numerical observations in a two-way contingency table is completed. It is found that when the data are displayed in an incomplete two-way table with non-numerical observations, the proposed method is more appropriate than Fisher's. By grouping categories in two-way contingency tables, it is shown that the non-zero canonical correlations for the original table need to be the same as those for the collapsed table. The asymptotic joint distribution of the canonical correlations for the original and the collapsed tables are derived under multinomial, Possion, product multinomial and hypergeometric sampling schemes. A Wald's test is developed to test the equality of the squared canonical correlations for the original and collapsed tables. Similarly, the asymptotic distributions of the test statistics for quasi-independence or quasi-homogeneity of the two-way contingency table making use of Pearson's chi-squared statistics for the collapsed tables are also derived. Finally, for normal data with several groups, the conditional joint distribution of the squared canonical correlations for the original data and the data when some groups are collapsed is derived. Tests are proposed to evaluate a hypothetical discriminant function and to assess departure from collinearity of the group means when homogenous groups are merged.