CD4+ T Cell Responses: A Complex Network of Activating and Tolerizing Signals as Revealed by Gene Expression Analysis: A Dissertation

Immunologic self-tolerance is maintained by both central and peripheral mechanisms. Furthermore, regulation of mature lymphocyte responses is governed by inhibitory as well as stimulatory signals. TCR recognition of cognate peptide bound to MHC molecules provides the initial stimulus leading to T lymphocyte activation and determines the antigen specificity of any subsequent response. However, lymphocytes must discriminate between foreign and self antigens presented by self-MHC molecules to maintain self tolerance and avoid pathological autoimmunity. Consequently, TCR ligation alone is reported to result in abortive activation, T cell anergy, apoptosis, and tolerance. Under normal physiological conditions, costimulatory signals modify lymphocyte responsiveness to TCR ligation to prevent autoimmunity while enabling robust responses to foreign antigen. Members of the CD28/B7 superfamily provide the critical secondary signals essential for normal immune cell function. CD28 is an essential positive costimulatory molecule with critical functions in thymic development, lineage commitment, and regulation of peripheral lymphocyte responses to antigenic stimuli. CD28 ligation by APC-expressed B7 molecules alters proximal signaling events subsequent to MHC/TCR interactions, and initiates unique signaling pathways that alter mRNA stability and gene transcription. Furthermore, CD28 signaling is required for regulatory T cell development and function. Thus, CD28 has a central role in both potentiating lymphocyte activation mediated by TCR engagement and regulating peripheral tolerance. In contrast, Ctla-4 mediates an inhibitory signal upon binding B7 molecules on an antigen-presenting cell. Its importance in governing lymphocyte responses is manifested in the fatal lymphoproliferative disorder seen in Ctla-4-/- mice. The lymphocyte proliferation is polyclonal, antigen and CD28 dependent, and arises from defects in peripheral CD4+T cell regulation. The high percentage of peripheral T lymphocytes expressing activation markers is accompanied by lymphocyte infiltration into numerous non-lymphoid tissues and results in death by 3-4 weeks. While still controversial, Ctla-4 signaling has been reported to be essential for induction of peripheral T lymphocyte tolerance in vivo and in some model systems is proposed to regulate both T lymphocyte anergy induction and the immune suppressive effects of some regulatory T cells in the prevention of autoimmunity. Signaling pathways activated by TCR ligation and CD28 costimulation have been extensively characterized. In contrast, the mechanisms mediating Ctla-4 maintenance of tolerance remain largely unknown. Ctla-4 gene expression is tightly controlled during T cell development and activation, and its intracellular localization and expression on the cell surface is regulated by numerous pathways and intermediates. While a tailless Ctla-4 mutant is capable of inhibiting T cell activation, recent studies have shown that a ligand independent form of Ctla-4 is also capable of providing an inhibitory signal to T lymphocytes. In conjunction with the strictly controlled expression kinetics and the perfect amino acid homology between the intracellular domains of mouse and human Ctla-4, this data suggests that Ctla-4 may participate in the modulation or initiation of intracellular signaling pathways. Positive and negative costimulatory receptors on the T cell modify lymphocyte responses by altering both quantitative and qualitative aspects of the lymphocyte response including threshold of activation, cytokine secretion, and memory responses. Positive costimulation augments T cell responses, in part, by downregulating the expression of genes that actively maintain the quiescent phenotype. This study was initiated to determine the role of Ctla-4 ligation in modifying the global gene expression profile of stimulated T cells and to determine if the Ctla-4 mediated maintenance of T cell tolerance was achieved, in part, by altering the transcription of quiescence genes necessary for the prevention of T cell activation subsequent to TCR and CD28 stimulation. Previous studies investigating the influence of Ctla-4 ligation on transcriptional profiles of activated lymphocytes detected only quantitative alterations in the transcriptional regulation initiated by CD28 signaling. In contrast, our data suggests that quantitative effects of Ctla-4 ligation that differentially influence pathways acting downstream of stimulatory receptors results in a stable and qualitatively unique phenotype detectable at the level of the transcriptome. Thus, the cumulative effect of Ctla-4 signaling is unique and not constrained to reversing alterations in expression initiated by CD28. In addition, Ctla-4 ligation can be shown to influence T lymphocyte responsiveness and the resulting global expression profile within 4 hours after stimulation and prior to detectable Ctla-4 surface expression. In a subpopulation of T cells, TCR stimulation activates pathways that result in commitment to activation with 2-6 hours. In contrast, CD28 signaling must be maintained for 12-16 hours to ensure maximal responses at the population level. The period of sensitivity to Ctla-4 inhibition of activation is more constrained and does not extend beyond 12 hours. Together, these data support a potential role for Ctla-4 in modification of the early transcriptional response and may explain various alterations in phenotype resulting from Ctla-4 ligation that have been reported in secondary responses. Identification of genes involved in lymphocyte activation, maintenance of selftolerance, and attenuation of immune responses opens the door to therapeutic manipulation of the pathways implicated. CD28 costimulation results in general amplification of TCR-initiated transcriptional responses, and specifically alters the expression profile of a subset of genes. In contrast, Ctla-4 ligation directly and specifically alters the expression of a select group of genes when ligated, and results in minimal suppression of the global CD28-mediated costimulatory transcriptional response. Ctla-4 regulated genes comprise a heterogeneous family, but include known quiescence factors, transcriptional regulators, and various determinants of cell cycle progression and senescence. The role of Ctla-4 in maintaining self-tolerance indicates that targeted manipulation of these gene products presents a novel therapeutic opportunity, and suggests that the mechanisms involved in Ctla-4-mediated maintenance of peripheral T cell tolerance and regulation of immune responsiveness is more nuanced than previously thought. In addition, this study provides the most comprehensive description of global gene expression during primary lymphocyte activation yet available. The integration of statistical and bioinfomatics analyses with large scale data mining tools identifies genes not previously characterized in lymphocytes and can direct future work by predicting potentially interacting gene products and pathways

Comparison of student perceptions of the learning environment during an academic calendar change

This is the second phase of a longitudinal research project to determine the effects on the learning environment of the change from a quarter system to a semester system form of academic calendar at Iowa State University. The null hypotheses for this part of the project were: (1) There will be no change in the student\u27s perception of the learning environment between Year 1 (quarter system) and Year 2 (semester system). (2) The degree of change as reflected by difference scores will not be related to the independent variables studied. (3) Within subgroups of the selected variables, there will be no change between Year 1 and Year 2. The independent variables studied were sex, college affiliation, G.P.A., residential location, full-time/part-time, work, attendance at another school under the semester system and classification;Hypothesis number one was rejected for two factors and many individual items in the survey. In Year 1, the respondents were undecided or slightly favorable toward the Semester Advantages factor. In Year 2, the respondents disagreed with the Semester Advantages factor. With the Quarter Advantages factor, the respondents were on the disagree side of the scale in both years, but the disagreement was less strong in Year 2;Hypothesis number two was rejected for the independent variables of classification, sex, and work. The Semester Advantages factor produced a significant change in all three variables;Hypothesis number three was rejected for sex, work, previous exposure to a semester system and classification. The Semester Advantages factor and Quarter Advantages factor indicated significant change for the first three variables while the Cultural/Community Activities factor was significant for graduate students;A prediction equation was attempted for the Semester Advantages factor. The best predictor was how the person scored on that factor in Year 1. However, only 28% of the variability could be explained

Pathogen propagation in cultured three-dimensional tissue mass

A process for propagating a pathogen in a three-dimensional tissue mass cultured at microgravity conditions in a culture vessel containing culture media and a culture matrix is provided. The three-dimensional tissue mass is inoculated with a pathogen and pathogen replication in the cells of the tissue mass achieved

H.P. Lovecraft & The French Connection: Translation, Pulps and Literary History

Weird fiction writer H. P. Lovecraft captured the zeitgeist of the modernist movement, despite his association with popular fiction. Lovecraft’s post-mortem climb from the margins of the American literary system to its center is indicative of his influence on “mass” and “elite” cultures alike in the second half of the twentieth century and onward. Lovecraft’s influence is not restricted to American culture, but it spread like an airborne virus to other cultures, and to France in particular. His imaginative weird fiction, a unique combination of horror and science fiction, has been translated into more than 25 languages from Bengali to Serbo-Croatian. The French were the first to translate Lovecraft and, according to S. T. Joshi, they hold provocative and insightful ideas and interpretations about him. Their interest in Lovecraft and their philosophical tradition have lead them to be the assumed champions of Lovecraft. Due to their unique differences, how does a comparison of Lovecraft’s image in French culture inform the modern American Lovecraft scholar and ehtusiast? This question is important because the modern American Lovecraft scholarship is primarily monolingual, and therefore cannot fully benefit from the excellent scholarship that is produced in the French language. The goal of this dissertation is to trace Lovecraft’s literary and cultural history in France, from its beginning to present-day. This lineage will be traced through an application of translation polysystems literary theory as described by Itamar Evan-Zohar and Gideon Toury. This system is effective in offering a conceptual model for the structures of literary systems on the micro (national) and the macro (world) level. Regarding the world literary system, the French have gained a particularly dominant position. Therefore, if the French have been leaders of Lovecraftian scholarship, how much of their interpretation of Lovecraft is visible in the American interpretation of Lovecraft? Ultimately, despite the literary influence of France on world cultural systems, polysystems literary theories explain the particular role that translation plays in the balance of literary domination

Power supply switching for a mm-wave asymmetric multilevel outphasing power amplifier system

Thesis (M. Eng.)--Massachusetts Institute of Technology, Dept. of Electrical Engineering and Computer Science, 2010.Cataloged from PDF version of thesis.This thesis demonstrates power switches to be used in our new Asymmetric Multilevel Outphasing (AMO) transmitter architecture at mm-wave frequencies. The AMO topology breaks the linearity vs. efficiency design objective in radio frequency power amplifiers (PAs) which has until now appeared to be fundamental. These power switches allow for the modulation of the PA supply rail between four discrete levels at a maximum sampling rate of 2 GHz. This modulation results in a higher average system efficiency by reducing the outphasing angle between the phase paths. This work was designed in a 130-nm SiGe BiCMOS process.by Jonathon David Spaulding.M.Eng

Short term doxycycline treatment induces sustained improvement in myocardial infarction border zone contractility.

Decreased contractility in the non-ischemic border zone surrounding a MI is in part due to degradation of cardiomyocyte sarcomeric components by intracellular matrix metalloproteinase-2 (MMP-2). We recently reported that MMP-2 levels were increased in the border zone after a MI and that treatment with doxycycline for two weeks after MI was associated with normalization of MMP-2 levels and improvement in ex-vivo contractile protein developed force in the myocardial border zone. The purpose of the current study was to determine if there is a sustained effect of short term treatment with doxycycline (Dox) on border zone function in a large animal model of antero-apical myocardial infarction (MI). Antero-apical MI was created in 14 sheep. Seven sheep received doxycycline 0.8 mg/kg/hr IV for two weeks. Cardiac MRI was performed two weeks before, and then two and six weeks after MI. Two sheep died prior to MRI at six weeks from surgical/anesthesia-related causes. The remaining 12 sheep completed the protocol. Doxycycline induced a sustained reduction in intracellular MMP-2 by Western blot (3649±643 MI+Dox vs 9236±114 MI relative intensity; p = 0.0009), an improvement in ex-vivo contractility (65.3±2.0 MI+Dox vs 39.7±0.8 MI mN/mm2; p<0.0001) and an increase in ventricular wall thickness at end-systole 1.0 cm from the infarct edge (12.4±0.6 MI+Dox vs 10.0±0.5 MI mm; p = 0.0095). Administration of doxycycline for a limited two week period is associated with a sustained improvement in ex-vivo contractility and an increase in wall thickness at end-systole in the border zone six weeks after MI. These findings were associated with a reduction in intracellular MMP-2 activity

Production of Normal Mammalian Organ Culture Using a Medium Containing Mem-Alpha, Leibovitz L 15, Glucose Galactose Fructose

Normal mammalian tissue and the culturing process has been developed for the three groups of organ, structural and blood tissue. The cells are grown in vitro under micro- gravity culture conditions and form three dimensional cells aggregates with normal cell function. The microgravity culture conditions may be microgravity or simulated microgravity created in a horizontal rotating wall culture vessel. The medium used for culturing the cells, especially a mixture of epithelial and mesenchymal cells contains a mixture of Mem-alpha and Leibovits L15 supplemented with glucose, galactose and fructose

Cultured normal mammalian tissue and process

Normal mammalian tissue and the culturing process has been developed for the three groups of organ, structural and blood tissue. The cells are grown in vitro under microgravity culture conditions and form three dimensional cell aggregates with normal cell function. The microgravity culture conditions may be microgravity or simulated microgravity created in a horizontal rotating wall culture vessel

A Call to Action: A Blueprint for Academic Health Sciences in the Era of Mass Incarceration

Over 100 million Americans have criminal records, and the U.S. incarcerates seven times more citizens than most developed countries. The burden of incarceration disproportionately affects people of color and ethnic minorities, and those living in poverty. While 95% of incarcerated people return to society, recidivism rates are high with nearly 75% arrested again within five years of release. Criminal records impede access to employment and other social services such as shelter and health care. Justice-involved people have higher rates of substance, mental health, and some chronic medical disorders than the general population; furthermore, the incarcerated population is rapidly aging. Only a minority of academic health science centers are engaged in health services research, workforce training, or correctional health care. This commentary provides rationale and a blueprint for engagement of academic health science institutions to harness their capabilities to tackle one of the country\u27s most vexing public health crises