Thesis (MSc)--University of Stellenbosch, 2006. / ENGLISH ABSTRACT: The present study aims to determine the phylogenetic relationships among the sand lizards, Pedioplanis. In addition, a single mitochondrial gene is used to investigate the geographic genetic structure of the widey distributed P. burchelli. With 11 species, Pedioplanis is the most speciose genus among the southern African genera of the family Lacertidae. All the species are restricted to the subcontinent with the exception of three (P. namaquensis, P. undata and P. benguellensis), which extend their range northwards into Angola. A total of 2200 nucleotide positions derived from two mitochondrial markers (ND2 and 16S rRNA) and one nuclear gene (RAG-1) are used to determine the phylogenetic relationships among ten of the eleven Pedioplanis species. The first well resolved gene tree for the genus, drawn from 100 individuals, is presented and this is largely congruent with a phylogeny derived from morphology. Contrary to some previous suggestions, Pedioplanis forms a monophyletic assemblage with Heliobolus and Nucras. The genus Pedioplanis is monophyletic with P. burchelli/P. laticeps forming a sister clade to all the remaining congeners. Two distinct geographic lineages can be identified within the widespread P. namaquensis; one occurs in Namibia, while the other occurs in South Africa. The “P. undata” species complex is monophyletic, but one of its constituent species, P. inornata, is paraphyletic. Relationships among the subspecies of P. lineoocellata are much more complex than previously documented. An isolated population previously assigned to P. l. pulchella is paraphyletic and sister to the three named subspecies. The phylogeny identifies two biogeographical groupings that probably diverged during the mid-Miocene. The development of the Benguella Current could have initiated isolation mechanisms associated with changes in habitat that could have generated barriers and played a role in the evolution of this group. At the lower taxonomic level, the mtDNA phylogeographic structure of the wide spread P. burchelli in South Africa reveal at least six distinct clades that are geographically partitioned. The first one is restricted to the eastern mountains along the Great Escarpment (GE). The next three are found along the Cape Fold Mountains (CFM): the north-west CFM, central CFM and eastern CFM. The fifth one shares samples from central CFM and GE. The last clade is restricted to the eastern central mountains of the GE. These six geographic groupings are genetically divergent from each other and they started separating in the early Pliocene period. Phylogeographic studies on other taxa in the region have found different levels of genetic structuring among or within taxa. The fact that P. burchelli is restricted to high altitude areas could have resulted in limited dispersal and consequently contributed to its geographic structure. However, the exact cause of the pattern obtained is not readily apparent. Habitat fragmentation in the past is probably one of the most influential factors shaping the genetic distribution of the species across South Africa. The inclusion of nuclear markers will shed more light on the evolutionary history of P. burchelli in South Africa. / AFRIKAANSE OPSOMMING: Die huidige studie stel ten doel om ‘n filogenie daar te stel vir die Sand akkedisse, Pedioplanis. ‘n Enkele mitochondriale geen is ook gebruik om die geografiese genetiese struktuur van die wydverspreide P. burchelli vas te stel. Met 11 spesies is Pedioplanis die mees spesieryke genus onder die suidelike Afrika genera wat aan die Lacertidae familie behoort. Al die spesies is beperk tot die subkontinent met die uitsondering van drie (P. namaquensis, P. undata en P. benguellensis), wat ‘n uitgebreide verspreiding het noordwaarts tot in Angola. ‘n Totaal van 2200 nukleotied posisies wat afkomstig is van twee mitochondriale merkers (ND2 en 16S rRNA) en een nukluêre geen (RAG-1) is gebruik om die filogenetiese verwantskappe tussen 10 van die 11 Pedioplanis spesies vas te stel. Die eerste goed geondersteunde geen boom vir die genus, gebasseer op 100 individue, is verkry en dit is meestal ooreenstemmend met ‘n filogenie gebasseer op morfologie. In teenstelling met sekere voorstelle van die verlede vorm Pedioplanis ‘n monofiletiese groep tesame met Heliobolus en Nucras. Die genus Pedioplanis is monofileties met P. burchelli/P. laticeps wat ‘n suster groep vorm van al die oorblywende lede van die genus. Twee herkenbare geografiese lyne kan geidentifiseer word in die wydverspreide P. namaquensis; een kom in Namibia voor, terwyl die ander een in Suid Afrika voorkom. Die “P. undata” spesies kompleks is monofileties, maar een van die spesies wat deel uitmaak van die groep, P. inornata, is parafileties. Verwantskappe tussen die subspesies van P. lineoocellata is meer kompleks as wat aanvanklik aanvaar is. ‘n Geisoleerde bevolkimg wat voorheen toegesê is aan P. l. pulchella is parafileties en verteenwoordig ‘n suster groep van die benaamde subspesies. Die filogenie identifiseer twee biogeografiese groeperings wat moontlik gedivergeer het gedurende die middel-Miocene. Die ontwikkeling van die Benguella stroom het dalk versperrings geinisiëer as gevolg van die gesamentlike veranderinge in habitat wat dalk ook ‘n rol gespeel het in die evolusie van die groep. Op die laer taksonomiese vlak het die mtDNA filogeografiese struktuur van die wydverspreide P. burchelli in Suid Afrika ten minste ses groepe aangetoon wat geografies van mekaar geskei is. Die eerste een is beperk tot die oostelike berge wat aan die Groot Eskarpement (GE) behoort. Die volgende drie word gevind in die Kaapse Vouberge (KVB): die noord-westelike KVB, sentrale KVB en oostelike KVB. Die vyfde een deel eksemplare van beide die GE en die KVB. Die laaste groep is beperk tot die oostelike en sentrale berge van die GE. Hierdie ses geografiese groepe is geneties geskei van mekaar en hulle het begin om apart te ontwikkel gedurende die vroë Pliocene periode. Ander filogeografiese studies in die area het verskillende vlakke van genetiese struktuur vertoon tussen en binne taksa. Die feit dat P. burchelli beperk is tot hoogliggende dele kon moontlik bygedrae het tot die geografiese struktuur. Die presiese oorsaak van die patroon wat verkry is, is nie ooglopend nie. Habitat fragmentasie in die verlede is moontlik een van die mees invloedrykste faktore wat die genetiese verspreiding van die spesie in Suid Afrika beinvloed het. Die insluiting van nukluêre merkers sal meer lig warp op die evolusionêre geskiedenis van P. burchelli in Suid Afrika.

This study determined the impact of vitamin C dose on genistein-induced apoptosis in LNCaP cancer cells at various treatment regimens in vitro. Although the linear regression of viability assay (MTT) indicated a p-value = 0.11; NBT assay reveal a declining SOD activity during cell death. Apoptosis induction was the main mode of treatment induced cell death. The overall data showed the trend of treatment efficacy as;(Gen 10uM + Vit C 40uM) > (Gen 30uM + Vit C 40uM) > (Gen 70uM + Vit C 40uM) > 10uM genistein > 70uM genistein. The chi-square test for comparing necrosis, apoptosis and life cells showed that Vitamin C could impact genistein-induced apoptosis in LNCaP cells (p = 0.0003). This study forms the basis for in vivo studies of the impact of vitamin C on genistein-induced apoptosis in LNCaP prostate cancer cells. / Includes bibliography. / Thesis (M.S.)--Florida Atlantic University, 2015. / FAU Electronic Theses and Dissertations Collection

An estimated 220,800 new prostate cancer cases and 27,540 deaths are expected to occur in US men by the end of 2015. Despite the increased treatment modes for prostate cancer, there is still no definite cure, and prognosis remains, at best, cautiously optimistic. The explicit amalgamation of two or more cancer therapeutic modalities such as surgery, radiation, and chemotherapy, has been one of the main interests of clinical investigation for several decades. Genistein (GN) and Beta-lapachone (BL) are two of the most promising anticancer phytochemical compounds. However, the anticancer activities of BL have been correlated with the enzyme activity of NQO1. The aim of this study was to investigate the enhancing effects of VLDR derived from a portable pyroelectric crystal generator on the chemopreventive and/or chemotherapeutic effects of GN and BL in NQO1+ PC3 and NQO1± (deficient) LNCaP prostate cancer cells (PCa) in vitro. The combination treat ment-induced cytotoxicity was investigated via MTT and Trypan blue exclusion assays. Dicoumarol (an NQO1 inhibitor) was co-administered to assess the effect of VLDR on NQO1 modulation. Nitro-blue tetrazolium assay was used to assess the intracellular ROS levels. Fluorescence microscopy was also used to assess the mode of cell death. In this study, a novel quantitative modeling approach was employed to comparably assess the cytotoxic effects of specific drugs used alone or in combinations with VLDR and to predict the potential synergistic therapeutic combinations. The data suggests that VLDR induced a rise in ROS levels, followed by upregulation in NQO1 levels. Pharmacodynamic indices were developed to quantify and characterize the combination treatment as synergistic, additive or antagonistic per dose or time-interval. Synergism was found to be dose and time-interval dependent. The major mode of cell death by this combination therapeutic regimen was found to be via apoptosis . In conclusion, our results confirm that VLDR enhanced cytotoxicity effects of both drugs dose- and time-dependently. / Includes bibliography. / Thesis (M.S.)--Florida Atlantic University, 2015. / FAU Electronic Theses and Dissertations Collection

Huntington's Disease (HD) is a devastating neurodegenerative disorder caused by an expanded polyglutamine repeat within the Huntingtin gene IT15. In this study we demonstrated that Bcl-2 interacting mediator of cell death Extra Long (BimEL) protein expression was significantly increased in cells expressing mutant Huntingtin (mHtt). Moreover, striatal BimEL expression remained high in an R6/2 HD mouse model throughout the disease progression. Utilizing novel BimEL phospho-mutants we demonstrated the phosphorylation of Ser65 to be important for the stabilization of BimEL. We provided evidence that impaired proteasome function, increased JNK activity and reduced striatal BDNF lead to changes in the phosphorylation of BimEL, thereby promoting its stabilization specifically within the striatum of R6/2 mice. Furthermore, knocking down BimEL expression prevented mHtt-induced cell death in a HD cell culture. Taken together, these findings suggest that BimEL may contribute to the selective neurodegeneration and pathogenesis of HD. / by Rebecca Leon. / Thesis (Ph.D.)--Florida Atlantic University, 2012. / Includes bibliography. / Mode of access: World Wide Web. / System requirements: Adobe Reader.

Cytomegaloviruses (CMVs) are ubiquitous in nature, having evolved over many millenia with their hosts. While in healthy hosts most infections with CMV are asymptomatic, the virus can cause severe disease in immunocompromised hosts. Thus, the increase in organ transplantation and the HIV/AIDS pandemic have established human CMV (HCMV) as a clinically important pathogen. Indeed, HCMV infections are now the major cause of morbidity and mortality among immunocompromised patients, which has led to more research targeting CMV for effective anti-viral treatment. The discovery that cytomegaloviruses encode several genes which are involved in immune escape has prompted a new area of research, aimed at understanding immune escape mechanisms for exploitation as potential anti-viral therapeutics. By targeting the viral proteins directly, or their receptors in the host, it may be possible to treat CMV disease by agonistic/antagonistic therapy. The first part of this thesis describes the first demonstration of anti-NK1.1 staining in situ to identify NK cells using a modified in vivo perfusion/fixation method. Using this method, we have compared the acute NK1.1+ cellular response to wild-type MCMV infection in the visceral organs of genetically susceptible intra-NK complex recombinant BALB.B6-CT6 (Cmv1s, NK1.1+) mice with resistant C57B⁄J (Cmv1r, NK1.1+) and BALB.B6-Cmv1r mice (Cmv1r, NK1.1+). Expression of viral antigens and the consequences of infection on other cellular subsets, were also analyzed in this study. The data show that in susceptible mice (Cmv1s) MCMV infection is predominent in the marginal zone of splenic white pulp, resulting in local changes in various cellular constituents, including macrophages, NK cells and DC. In the liver, distinct foci of infection were comprised of large numbers of macrophages and NK1.1+ cells surrounding infected cytomegalic cells. In resistant mice (Cmv1r), 6 MCMV infection predominantly affected the red-pulp of the spleen and was associated with increased accumulation of NK1.1+ cells and macrophages at sites of viral infection

[Truncated abstract] Ovarian cancer is currently the leading cause of death from gynaecological malignancies in women from developed countries. Serous ovarian cancer is the most prevalent type of all ovarian cancers, with the majority diagnosed in an advanced stage where treatment efficacy is reduced and patient survival is poor. Because of this fact, the development of improved detection and treatment strategies are necessary, with much research focussing on the complex molecular pathways involved in ovarian tumour growth as one potential avenue for intervention. Apoptosis, or programmed cell death, is one such area of investigation because currently successful cancer treatments induce apoptosis in tumour cells. Molecular analysis of apoptosis in both normal tissue and tumours has established a positive relationship between increased expression of secreted frizzled-related protein 4 (SFRP4) and apoptosis, however to date, very little research has focussed on the role of this gene in the ovary . . . An examination of SFRP4 and β-catenin expression in 163 primary serous ovarian carcinomas revealed high SFRP4 expression was associated with low β-catenin expression and conversely, low SFRP4 was associated with high β-catenin expression in the majority of the ovarian tumours analysed, reinforcing the inverse relationship observed in the ovarian cell lines. A positive trend was observed between cancer stage and the expression level of these proteins, with increased SFRP4 expression and reduced β-catenin expression as cancer stage increased. Additionally, patient survival revealed a trend towards increased survival among ovarian cancer patients who had tumours expressing low levels of SFRP4. Taken together, the novel findings of this study indicate that the increased expression of SFRP4 observed in a large proportion of serous ovarian cancers is a cellular response to down-regulate the level of β-catenin, and thus an attempt to maintain cellular homeostasis by counteracting the excessive proliferating signals present in these tumour cells.

No description available.

Prostate cancer is one of the leading causes of death in men aged 40-55. Genistein isoflavone (4', 5', 7-trihydroxyisoflavone) is a dietary phytochemical with demonstrated anti-tumor activities in a variety of cancers. Topotecan Hydrochloride (Hycamtin) is an FDA-approved chemotherapy drug, primarily used for secondary treatment of ovarian,cervical and small cell lung cancers. This study was to demonstrate the potential anticancer activities and synergy of topotecan-genistein combination in LNCaP prostate cancer cells. The potential efficacy and mechanism of topotecan/genistein-induced cell death was investigated... Results: The overall data indicated that i) both genistein and topotecan induce cellular death in LNCaP cells, ii) topotecan-genistein combination was significantly more efficacious in reducing LNCaP cell viabiligy compared to either genistein or topotecan alone, iii) in all cases, cell death was primarily through apoptosis, via the activation of the intrinsic pathway, iv) ROS levels were increased and VEGF expression was diminished significantly with the topotecan-genistein combination treatment, v) genetic analysis of topotecan-genistein treatment groups showed changes in genetic expression levels in pathway specific apoptotic genes.... Conclusion: Treatments involving topotecan-genistein combination may prove to be an attractive alternative phytotherapy of adjuvant therapy for prostate cancer. / by Vanessa P. Hèormann. / Thesis (Ph.D.)--Florida Atlantic University, 2012. / Includes bibliography. / Mode of access: World Wide Web. / System requirements: Adobe Reader.

Indiana University-Purdue University Indianapolis (IUPUI) / Lyme disease, the most prevalent vector-borne illness in the United States, is a multisystem inflammatory disorder caused by infection with the spirochete Borrelia burgdorferi (Bb). This spirochete is maintained in nature through an enzootic cycle involving ticks and small mammals. The Bb genome encodes a large number of surface lipoproteins, many of which are expressed during mammalian infection. One of these lipoproteins is the major outer surface protein C (OspC) whose production is induced during transmission as spirochetes transition from ticks to mammals. OspC is required for Bb to establish infection in mice and has been proposed to facilitate evasion of innate immunity. However, the exact biological function of OspC remains elusive. Our studies show the ospC-deficient spirochete could not establish infection in NOD-scid IL2rγnull mice that lack B cells, T cells, NK cells, and lytic complement, whereas the wild-type spirochete was fully infectious in these mice. The ospC mutant also could not establish infection in SCID and C3H mice that were transiently neutropenic during the first 48 h post-challenge. However, depletion of F4/80+ phagocytes at the skin-site of inoculation in SCID mice allowed the ospC mutant to establish infection in vivo. In phagocyte-depleted SCID mice, the ospC mutant was capable to colonize the joints and triggered neutrophilia during dissemination in a similar pattern as wild-type bacteria. We then constructed GFP-expressing Bb strains to evaluate the interaction of the ospC mutant with phagocytes. Using flow cytometry and fluorometric assay for phagocytosis, we found that phagocytosis of GFP-expressing ospC mutant spirochetes by murine peritoneal macrophages and human THP-1 cells was significantly higher than parental wild-type Bb strains, suggesting that OspC has an anti-phagocytic property. This enhancement in phagocytosis was not mediated by MARCO and CD36 scavenger receptors and was not associated with changes in mRNA levels of TNFα, IL-1β, and IL-10. Phagocytosis assays with HL60 neutrophil-like cells showed that uptake of Bb strains was independent to OspC. Together, our findings reveal that F4/80+ phagocytes are important for clearance of the ospC mutant, and suggest that OspC promotes spirochetes' evasion of macrophages in the skin of mice during early Lyme borreliosis.

Thesis (MSc)--Stellenbosch University, 2000. / ENGLISH ABSTRACT: Progressive familial heart block type I (PFHBI) is a cardiac ventricular conduction disorder of unknown cause associated with risk of sudden death, which has been described in several South African families. Clinically, PFHBI is characterised by right bundle branch block on ECG, which may progress to complete heart block, necessitating pacemaker implantation. The disease shows an autosomal dominant pattern of inheritance with evidence of genetic anticipation. Using genetic linkage analysis, the PFHBI-causative gene was mapped to a 10 eentimorgan (cM) gene-rich area of chromosome (C) 19q13.3, which has, subsequently, been reduced to 7cM by fine mapping with polymorphic dinucleotide (CA)n short tandem repeat (STR) markers. Several attractive candidate genes, including muscle glycogen synthase (GSY 1) and histidine-rich calcium binding protein (HRC), lie within this region. The aim of the present study was two-fold: 1) to identify and characterise tetranucleotide (AAAT)n STRs within the PFHBI critical region that could be developed as polymorphic markers for use in genetic fine mapping and 2) to screen selected regions of GSY 1and HRC, positional candidate genes, for the presence ofPFHBI-causing mutation(s). Cosmids harbouring CI9q13.3 insert DNA were screened for the presence of (AAAT)n STRs by dot blot and Southern blot hybridisation using a radiolabelled (AAAT)lO oligonucleotide probe. To characterise the harboured (AAAT)n STRs, the positively hybridising fragments identified by Southern blot were sub-cloned, sequenced and primers designed from the unique repeat-flanking sequences. These primers were used to genotype the (AAAT)n repeat locus to assess its polymorphic nature in a panel of unrelated individuals. Alternatively, vectorette PCR, a rapid method of identifying repeat sequences and obtaining the flanking sequences in large inserts, was employed to develop polymorphic markers from the positively hybridising clones. Selected exons of GSY1 and HRC were screened for the presence of potentially disease-causing mutations by PCR-SSCP analysis and direct sequencing, respectively, in PFHBI-affected and unaffected family members. Of the available cosmid clones that gave strong signals on dot blot and Southern blot hybridisation, three, 29395, 24493 and 20381, were located within the critical PFHBI area and were used for marker development. An interrupted (AAAT)n repeat motif (n less than 5) was identified in cosmid 29395, however, the repeat locus was not polymorphic in the tested population. No (AAAT)n motif, single or repeated was observed in the partial sequence of the sub-cloned fragment of cosmid 24493. Using vectorette peR, no repeated (AAAT)n motif was identified on sequencing the generated products in either cosmid 24493 or 2038l. However, diffuse single AAAT motifs were detected in both cosmids. Exons 4, 5, 11, 12 and 16 of GSY 1, containing domains that are conserved across species, and the conserved eterminus- encoding exons 2-6 of HRC were selected for screening for potential PFHBI-causing mutation(s). However, no sequence variations were detected. The interrupted (AAAT)n repeat identified in cosmid 29395 was not polymorphic, which confirmed reports that complex repeats, especially those containing AAAT motifs of less than 6 repeats, are not polymorphic. One possible explanation for the absence of a repeated AAAT motif in cosmids 24493 and 20381, which both gave positive hybridisation signals, is that the low annealing temperature of the AfT -rich repeat-anchored primers used in vectorette peR may have resulted in transient annealing to the diffuse single AAAT motifs detected on sequencing. The screened regions of candidate genes GSYI and HRC were excluded from carrying the disease-causing mutation(s). The availability of new sequence data generated by the Human Genome Project will influence future strategies to identify the PFHBI gene. Electronic searches will allow identification of STR sequences for development of polymorphic markers and gene annotation will allow selection of new candidate genes for mutation screening. / AFRIKAANSE OPSOMMING: Sien volteks vir opsomming