Reticuloendotheliosis virus (REV) is a retrovirus that causes a neoplastic disease in a wide range of avian hosts including chickens, turkeys, and ducks. In 1993, REV was detected in the endangered Attwater's prairie chicken (Tympanachus cupido attwateri), a subspecies of Tympanachus cupido. Subsequent infections of this prairie chicken have been identified at captive breeding facilities throughout Texas. The implications of these infections have severely hindered repopulation efforts by these facilities. This study focused on investigating REV infection of captive Attwater'€™s prairie chicken in order to better understand the disease affecting these endangered birds. The overall objective was to develop a means of eliminating this threat to the repopulation of the Attwater's prairie chicken. Several aspects of virus infection were investigated. Reagents capable of recognizing prairie chicken IgY and viral gag polypeptides were developed for use in assays for detection of antibody responses and titration of viral concentrations. Sequencing data of genomes collected from isolates of Texas prairie chickens and domestic chickens, as well as three REV prototype viruses, were compared to determine relationships among strains and identify the potential origin of the REV infecting Attwater'€™s prairie chicken. Additionally, a flow cytometry technique of segregating the lymphocyte population from peripheral blood mononuclear cells (PBMC) using a pan leukocyte monoclonal antibody was developed to more accurately measure changes within lymphocyte populations. This technique combined with intracellular labeling was used to deduce the target cells of REV infection. A nested polymerase chain reaction (PCR) test was developed for greater sensitivity in detecting infection in birds than the previous method of single amplification PCR. This greater sensitivity results in earlier identification of the virus in infected birds, which allows for earlier removal of infected birds to minimize transmission of the virus throughout the flock. The sensitivity of the nested PCR diagnostic test was determined in a dose response pathogenesis study, which was conducted on hybrid greater/Attwater's prairie chicken to observe the experimental development of disease in these birds. Finally, a vaccine was developed using plasmid DNA with REV encoded genes and tested on naturally infected prairie chickens to determine its efficacy in reducing viral load. Although no reduction in viral load was detected, the vaccine may be effective in providing prophylactic protection in future studies.

IKK[alpha] is a 85KD serine/threonine protein kinase and a subunit of the IKK complex, which contains IKK[alpha], IKK[beta], and IKK[gamma]. IKK[alpha] and IKK[beta] are highly conserved and they contain three functional domains of kinase domain, leucine zipper (LZ), and helix-loop-helix (HLH). Although IKK[alpha] and IKK[beta] can phosphorylate IκB proteins in vitro, IKK[alpha] and IKK[beta] have distinct physiological functions during mouse development. Genetic studies showed that IKK[alpha] is essential for embryonic skin development in mice. Mice deficient in IKK[alpha] display a hyperplastic epidermis that lacks terminal differentiation, resulting a death soon after birth because of the severely impaired skin. Recently, we reported a reduction in IKK[alpha] expression and identified somatic Ikk[alpha] mutations in a high proportion of poorly differentiated human squamous cell carcinomas (SCCs) (Liu et al., 2006). The aim of this study is to investigate the novel role of IKK[alpha] in skin carcinogenesis. We firstly examined IKK[alpha] expression and Ikk[alpha] mutations in human SCCs and found a reduction of IKK[alpha] in poorly differentiated human SCCs and identified somatic Ikk[alpha] mutations in exon 15 of Ikk[alpha] in human SCCs. We then examined the susceptibility of Ikk[alpha] hemizygotes to chemical carcinogeninduced skin carcinogenesis. In this chemical carcinogen-induced skin carcinogenesis setting, 7,12-dimethylbenz[a]anthracene (DMBA) induces Ras mutations and 12-Otetradecanoyl-phorbol-13-acetate (TPA) promotes Ras-initiated cell proliferation. We found two times more papillomas and eleven times more carcinomas in Ikk[alpha superscript +/-] mice than in Ikk [alpha] superscript +/+] mice induced by DMBA/TPA. Ikk[alpha superscript +/-] mice developed larger and earlier tumors than did Ikk[alpha superscript +/+] mice. Poorly differentiated carcinomas expressed low levels of IKK[alpha]. Ninety five percent of the Ikk[alpha superscript +/-] carcinomas and 44% of the Ikk[alpha superscript +/-] papillomas lost the remaining wild type Ikk[alpha] allele. This result indicates that the remaining one wild type Ikk[alpha] allele is important for preventing malignant carcinoma conversion. Also Ikk[alpha] mutations were detected in these skin tumors. Reduced IKK[alpha] was found to enhance TPA-induced mitogenic and angiogenic activities in mouse skin. Taken together, these results suggest that reduction of IKK[alpha] expression provides a selective growth advantage, which cooperates with DMBA-initiated Ras activity to promote skin carcinogenesis. In addition, we observed a small group of FVB female Ikk [alpha superscript +/-] mice for 1.5 years and found that 12/ 24 mice developed various spontaneous tumors including mammary gland carcinomas, uterine and ovary tumors, and dermal fibrosacomas. Somatic Ikk[alpha] mutations, elevated IKK/ NF[subscript -k]B and extracellular signal-regulated kinases (ERK) activities and elevated cyclin D1 levels were detected in these spontaneous tumors. These results suggest that these molecular alterations may contribute to the development of these tumors although the precise role of the down-regulation of IKK in the development of the tumors remains to be determined. Overall, our data and other published results suggest that IKK[alpha] is a new tumor suppressor in men and mice. / text

published_or_final_version / Medicine / Doctoral / Doctor of Philosophy

published_or_final_version / Paediatrics and Adolescent Medicine / Master / Master of Research in Medicine

Diabetic retinopathy (DR) was characterized by numerous hyperglycemia-dependent cellular and pathological changes in the retina, including retinal ischemia/reperfusion (I/R) injury. To determine the role of the 2nd enzyme of polyol pathway in relation with the pathogenesis in ischemic retinopathy, SDH deficient mice, C57BL/LiA, that lacked SDH activity, was used to study the pathogenesis of diabetic retinopathy, which also included I/R injury. Wild type and SDH-deficient mice were subjected to I/R injury by transiently occluding middle cerebral artery for two hours and twenty-two hour of reperfusion. The rationale of this study was to investigate the effect by blocking the conversion of sorbitol to fructose by SDH null mutation (SDH -/-), leading to accumulation of sorbitol level and reduction of oxidative stress, as demonstrated by the polyol pathway. Results: After induction with transient MCAO, there was increase in the thickness of OLM to ILM ipsilateral SDH+/+ compared with contralateral SDH+/+ (from 84 +/- 1 to 96 +/- 2 μm) while that of ipsilateral SDH-/- compared with contralateral SDH -/- (from 77 +/- 2 to 90 +/- 2 μm) suggested that there was edema after ischemic reperfusion injury. The result showed that there was increased cellular edema in ipsilateral retina of both SDH +/+ and SDH -/- retina after transient MCAO. The level of immunoreactivity against Aquaporin-4 and nitrotyrosine in studying the presence of oxidative stress; glutamine synthetase and glutamate in studying the toxicity of astrocyte glutamate; sarco-endoplasmic reticulum Ca2+-ATPase (SERCA) in studying the regulation Ca2+ homeostasis was determined using immunohistochemistry. For all the antibodies, there was similar immunoreactivity level between the contralateral side of both SDH+/+ and SDH -/- mice. For the SDH+/+ group, there was increase in signal in the ipsilateral retina in comparison with the contralateral one. On the other hand, for the SDH-/- group, similar result was observed. There was increase in signal and it was found more in the ipsilateral retina in comparison with the contralateral retina. Finally, in the ipsilateral retina of both SDH +/+ and SDH -/- mice, increased immunoreactivity was found in both but their difference was not statistically significant. This concluded that SDH deletion and subsequent accumulation of sorbitol metabolites did not contribute significantly in the role of pathogenesis of ischemic retinopathy especially in mice after I/R injury. / published_or_final_version / Anatomy / Master / Master of Medical Sciences

<p><italic>Cryptococcus neoformans </italic>is a human fungal pathogen that is also ubiquitous in the environment. To cause disease inside a human host, <italic>C. neoformans</italic> must be able to sense and respond to a multitude of stresses. One of the major responses to the host is the induction of a polysaccharide capsule, which allows the fungus to resist damage and evade the host immune response. This capsule is regulated by a number of signal transduction cascades, but a major contributor is the conserved cAMP/PKA pathway. </p><p> Using genetic and molecular biology techniques, I identified Gcn5 and Rim101 as key transcriptional regulators of capsule within the host. I determined that <italic>C. neoformans</italic> Rim101 is activated by a combination of the canonical pH sensing pathway and the cAMP/PKA pathway. This novel connection potentially gives the pathogen greater flexibility in responding to environmental stimuli, thus allowing for a greater capacity for disease. </p><p> I determined that the Rim101 transcription factor regulates cell wall remodeling in the context of the host by deep mRNA sequencing, electron microscopy, and biochemical assays. Using chromatin immunoprecipitation, I confirmed that these cell wall changes are under direct control of Rim101. I then confirmed the importance of cell wall changes in the host by nanoString profiling of fungal RNA in the context of a murine lung infection. I also examined the lungs of infected mice for cytokine and immune cell infiltrate and determined that <italic>C. neoformans</italic> cell wall changes are important in avoiding triggering an aberrant host response. I hypothesize that this cell wall remodeling via Rim101 activation is required for full capsule attachment and for masking immunogenic molecules from the host immune system.</p> / Dissertation

No description available.

No description available.

No abstract available. / Thesis (Ph.D.)-University of KwaZulu-Natal, Durban, 2011.

In this study we investigated the impact of exogenous reinfection on genetically resistant and genetically sensitive sub populations. We qualitatively analysed the dynamics of TB by assuming that TB is transmitted in two ways namely homogeneous and heterogeneous modes of transmission. Analytically, we computed the fundamental thresholds used to measure disease persistence; the basic reproduction number R₀; and found that the exogenous reinfection parameters do not appear in the basic reproduction number. Hence, basic reproduction number derived in presence of exogenous reinfection does not adequately predict the course of a TB epidemic. We obtained the exogenous reinfection threshold which indicated that exogenous reinfection complicates TB dynamics. Both analytical and simulation results disclosed that when exogenous reinfection is above exogenous reinfection threshold TB dynamics were governed by a backward bifurcation implying TB may continue to invade the population despite basic reproduction number being less than one. We computed critical value of basic reproduction numbers Rᴄ and found that TB can only be eradicated if basic reproduction number is reduced below critical value Rc. Furthermore, we incorporated TB therapy in heterogeneous model among individuals with clinically active TB and performed sensitivity and uncertainty analysis using Latin Hypercube Sampling. The sensitivity and uncertainty results showed that transmission rates, reactivation rates and proportion that is genetically resistant greatly infuenced outcome variables of our TB model. / Thesis (M.Sc.)-University of KwaZulu-Natal, Pietermaritzburg, 2013.