Thesis (M. Sc.)--Université Laval, 1999. / Includes bibliographical references.

Thesis (doctoral)--Rijksuniversiteit Groningen, 1989. / Includes bibliographical references.

Thesis (doctoral)--Rijksuniversiteit Groningen, 1989. / Includes bibliographical references.

Thesis (Ph. D.)--Medical University of South Carolina, 1998. / Includes bibliographical references.

Thesis (M.S.)--University of Florida, 2005. / Typescript. Title from title page of source document. Document formatted into pages; contains 57 pages. Includes Vita. Includes bibliographical references.

A presença de Candida albicans nos biofilmes microbianos aderidos na superfície interna das próteses removíveis, principalmente totais superiores, está relacionada com uma doença inflamatória no palato, conhecida como estomatite protética (EP). Assim, torna-se fundamental a realização de novos estudos sobre alternativas terapêuticas, direcionados à prótese e não somente à mucosa, que sejam simultaneamente antimicrobianas, anti-inflamatórias, não tóxicas para os tecidos bucais e que produzam menos danos à prótese que os métodos convencionais. Os fitoterápicos podem representar uma destas alternativas. Objetivos: O objetivo deste trabalho foi estudar a ação fitoterápica do Equisetum giganteum, nas concentrações de 50, 25, 16, 8 e 4 mg/mL, sobre C. albicans e descartar sua ação citotóxica sobre o palato humano bem como sobre monócitos humanos. Material e Métodos: Após coleta, obtenção e identificação de compostos por espectrometria de massas do extrato hidroalcoólico de E. giganteum, sua atividade antimicrobiana foi determinada pela concentração inibitória mínima em meio líquido, contra as cepas clínicas Candida albicans SC 5314 e Escherichia coli O:124, e a cepa padrão Staphylococcus aureus ATCC 6538. Propriedades antiaderentes do extrato, sobre biofilmes de C. albicans induzidos sobre corpos de prova de resina acrílica, foram determinadas por imunofluorescência (LIVE/DEAD) e pela análise em microscópio de varredura confocal a laser. A atividade anti-inflamatória do fitoterápico foi averiguada através da análise da produção de espécies reativas de oxigênio (ROS) por monócitos humanos estimulados por C. albicans e LPS, por meio da marcação fluorescente utilizando o reagente Cell Rox Deep Red®. Avaliação de citotoxicidade foi realizada in vitro com células epiteliais de palato humano e monócitos humanos, por meio do ensaio colorimétrico MTT. Os resultados foram expressos como média ± desvio padrão e submetidos aos testes estatísticos Teste de Kruskal-Wallis; ANOVA one-Way; ANOVA two-Way, Teste de Miller; Teste de Tukey e Dunnet; Teste de Fisher, sendo p<0,05 considerado significante. Resultados: O estudo da composição química do extrato EtOH 70% de E. giganteum evidenciou a presença de compostos fenólicos derivados dos ácidos cafeico e ferúlico, heterosídeos de flavonoides derivados de quercetina e kaempferol, além de estirilpironas. A atividade bactericida/fungicida foi comprovada em todas as concentrações testadas do extrato e contra todas as cepas avaliadas. Todas as concentrações do extrato resultaram em redução significativa dos biovolumes de células viáveis em relação a resina não tratada, o que indica interferência do fitoterápico na aderência de C. albicans à resina termopolimerizável. A atividade anti-inflamatória foi determinada pelo retorno da produção de espécies reativas de oxigênio aos níveis basais, pelos monócitos humanos ativados, quando tratados com todas as concentrações do extrato. Houve manutenção da viabilidade celular de monócitos humanos e das células epiteliais de palato humano, após contato com o fitoterápico nos períodos de 1, 12 e 24 horas. Conclusão: Estes resultados sugerem que o fitoterápico E. giganteum possui propriedades: microbicida frente a C. albicans, E. coli e S. aureus, antiaderente para C. albicans sobre a superfície da resina acrílica termopolimerizável e anti-inflamatória sobre monócitos humanos ativados por C. albicans. Além disso, o fitoterápico não comprometeu a viabilidade de monócitos humanos e de células epiteliais de palato humano. / The presence of Candida albicans in the microbial biofilms adhered to the internal surface of the removable denture, mainly the full upper ones, is related to an inflammatory palate disease known as denture stomatitis (DS). Thus, it is essential that new studies are done about therapeutic alternatives directed to the dentures, not only to the buccal mucosa, and which are, at the same time, antimicrobial, antiinflammatory, non-poisoning to the buccal tissues and that they produce less harm to the denture than the current methods. The phytotherapeutic (herbal) remedies may represent a good alternative. Objectives: The aim of this paper is to study the phytotherapeutic action of Equisetum giganteum in the concentrations of 50, 25, 16, 8 and 4 mg/mL on C.albicans and discard the cytotoxic action on the human palate, as well as on human monocytes. Material and Methods: After collecting, obtaining and identifying the compounds by means of mass spectrometry of the hydroalcoholic extract of E. giganteum, its antimicrobial activity was determined by the inhibitory minimum concentration in liquid media, against clinic strains of Candida albicans SC 5314 and Escherichia coli O:124, and standard Staphylococcus aureus ATCC 6538 strains. The antiadherent, properties of the extract on biofilms of C. albicans over acrylic resin proof specimens were determined by immunofluorescence test (LIVE/DEAD) and by the analysis in a Confocal Laser Microscope Scanning. The anti-inflammatory activity of the phytotherapeutic (herbal) remedy was assessed through the analysis of the production of reactive oxygen species (ROS) to human monocytes stimulated by C. albicans and LPS, through fluorescent lighting using the reagent Cell Rox Deep Red®. The evaluation of cytotoxicity was done in vitro with epithelial cells of human palate and human monocytes, through colorimetric MTT assay. The results were expressed in means ± standard deviation and submitted to statistics Kruskal-Wallis Test; ANOVA Two-way, Miller Test; Tukey and Dunnet Test; Fisher Test, where p<0,05 was considered significant. Results: The study of the chemical composition of the extract EtOH 70% of E. giganteum has shown a clear presence of phenolic compounds derived from caffeic and ferulic acids, flavonoid heterosides derived from quercitin and kaempferol, in addition to estirilpirones. Its bactericidal/fungicide activity was proved in all extract concentrations tested and against all evaluated strains. All extract concentrations resulted in significant reduction of bio volumes of viable cells in relation to non-treated resin, which indicates interference of the phytotherapeutic in the adherence of C. albicans to the thermopolymerizable acrylic. The antiinflammatory activity was determined by the production return of reactive oxygen specimens to basal levels, by activated human monocytes, when treated with all extract concentrations. There was cell viability of human monocytes and of human palate epithelial cells, after the contact of with the phytoterapeutic in the periods of 1, 12 and 24 hours. Conclusion: These results suggest that the phytotherapeutic E. giganteum has properties: microbicide in relation to C. albicans, E.coli and S. aureus, antiadherent to C. albicans on thermopolymerizable acrylic and antiinflammatory on human monocytes activated by C. albicans. In addition to this, the phytotherapeutic did not compromise either human monocytes viability or human palate epithelial cells.

A presença de Candida albicans nos biofilmes microbianos aderidos na superfície interna das próteses removíveis, principalmente totais superiores, está relacionada com uma doença inflamatória no palato, conhecida como estomatite protética (EP). Assim, torna-se fundamental a realização de novos estudos sobre alternativas terapêuticas, direcionados à prótese e não somente à mucosa, que sejam simultaneamente antimicrobianas, anti-inflamatórias, não tóxicas para os tecidos bucais e que produzam menos danos à prótese que os métodos convencionais. Os fitoterápicos podem representar uma destas alternativas. Objetivos: O objetivo deste trabalho foi estudar a ação fitoterápica do Equisetum giganteum, nas concentrações de 50, 25, 16, 8 e 4 mg/mL, sobre C. albicans e descartar sua ação citotóxica sobre o palato humano bem como sobre monócitos humanos. Material e Métodos: Após coleta, obtenção e identificação de compostos por espectrometria de massas do extrato hidroalcoólico de E. giganteum, sua atividade antimicrobiana foi determinada pela concentração inibitória mínima em meio líquido, contra as cepas clínicas Candida albicans SC 5314 e Escherichia coli O:124, e a cepa padrão Staphylococcus aureus ATCC 6538. Propriedades antiaderentes do extrato, sobre biofilmes de C. albicans induzidos sobre corpos de prova de resina acrílica, foram determinadas por imunofluorescência (LIVE/DEAD) e pela análise em microscópio de varredura confocal a laser. A atividade anti-inflamatória do fitoterápico foi averiguada através da análise da produção de espécies reativas de oxigênio (ROS) por monócitos humanos estimulados por C. albicans e LPS, por meio da marcação fluorescente utilizando o reagente Cell Rox Deep Red®. Avaliação de citotoxicidade foi realizada in vitro com células epiteliais de palato humano e monócitos humanos, por meio do ensaio colorimétrico MTT. Os resultados foram expressos como média ± desvio padrão e submetidos aos testes estatísticos Teste de Kruskal-Wallis; ANOVA one-Way; ANOVA two-Way, Teste de Miller; Teste de Tukey e Dunnet; Teste de Fisher, sendo p<0,05 considerado significante. Resultados: O estudo da composição química do extrato EtOH 70% de E. giganteum evidenciou a presença de compostos fenólicos derivados dos ácidos cafeico e ferúlico, heterosídeos de flavonoides derivados de quercetina e kaempferol, além de estirilpironas. A atividade bactericida/fungicida foi comprovada em todas as concentrações testadas do extrato e contra todas as cepas avaliadas. Todas as concentrações do extrato resultaram em redução significativa dos biovolumes de células viáveis em relação a resina não tratada, o que indica interferência do fitoterápico na aderência de C. albicans à resina termopolimerizável. A atividade anti-inflamatória foi determinada pelo retorno da produção de espécies reativas de oxigênio aos níveis basais, pelos monócitos humanos ativados, quando tratados com todas as concentrações do extrato. Houve manutenção da viabilidade celular de monócitos humanos e das células epiteliais de palato humano, após contato com o fitoterápico nos períodos de 1, 12 e 24 horas. Conclusão: Estes resultados sugerem que o fitoterápico E. giganteum possui propriedades: microbicida frente a C. albicans, E. coli e S. aureus, antiaderente para C. albicans sobre a superfície da resina acrílica termopolimerizável e anti-inflamatória sobre monócitos humanos ativados por C. albicans. Além disso, o fitoterápico não comprometeu a viabilidade de monócitos humanos e de células epiteliais de palato humano. / The presence of Candida albicans in the microbial biofilms adhered to the internal surface of the removable denture, mainly the full upper ones, is related to an inflammatory palate disease known as denture stomatitis (DS). Thus, it is essential that new studies are done about therapeutic alternatives directed to the dentures, not only to the buccal mucosa, and which are, at the same time, antimicrobial, antiinflammatory, non-poisoning to the buccal tissues and that they produce less harm to the denture than the current methods. The phytotherapeutic (herbal) remedies may represent a good alternative. Objectives: The aim of this paper is to study the phytotherapeutic action of Equisetum giganteum in the concentrations of 50, 25, 16, 8 and 4 mg/mL on C.albicans and discard the cytotoxic action on the human palate, as well as on human monocytes. Material and Methods: After collecting, obtaining and identifying the compounds by means of mass spectrometry of the hydroalcoholic extract of E. giganteum, its antimicrobial activity was determined by the inhibitory minimum concentration in liquid media, against clinic strains of Candida albicans SC 5314 and Escherichia coli O:124, and standard Staphylococcus aureus ATCC 6538 strains. The antiadherent, properties of the extract on biofilms of C. albicans over acrylic resin proof specimens were determined by immunofluorescence test (LIVE/DEAD) and by the analysis in a Confocal Laser Microscope Scanning. The anti-inflammatory activity of the phytotherapeutic (herbal) remedy was assessed through the analysis of the production of reactive oxygen species (ROS) to human monocytes stimulated by C. albicans and LPS, through fluorescent lighting using the reagent Cell Rox Deep Red®. The evaluation of cytotoxicity was done in vitro with epithelial cells of human palate and human monocytes, through colorimetric MTT assay. The results were expressed in means ± standard deviation and submitted to statistics Kruskal-Wallis Test; ANOVA Two-way, Miller Test; Tukey and Dunnet Test; Fisher Test, where p<0,05 was considered significant. Results: The study of the chemical composition of the extract EtOH 70% of E. giganteum has shown a clear presence of phenolic compounds derived from caffeic and ferulic acids, flavonoid heterosides derived from quercitin and kaempferol, in addition to estirilpirones. Its bactericidal/fungicide activity was proved in all extract concentrations tested and against all evaluated strains. All extract concentrations resulted in significant reduction of bio volumes of viable cells in relation to non-treated resin, which indicates interference of the phytotherapeutic in the adherence of C. albicans to the thermopolymerizable acrylic. The antiinflammatory activity was determined by the production return of reactive oxygen specimens to basal levels, by activated human monocytes, when treated with all extract concentrations. There was cell viability of human monocytes and of human palate epithelial cells, after the contact of with the phytoterapeutic in the periods of 1, 12 and 24 hours. Conclusion: These results suggest that the phytotherapeutic E. giganteum has properties: microbicide in relation to C. albicans, E.coli and S. aureus, antiadherent to C. albicans on thermopolymerizable acrylic and antiinflammatory on human monocytes activated by C. albicans. In addition to this, the phytotherapeutic did not compromise either human monocytes viability or human palate epithelial cells.

Approximately 26% of the U.S. population between the ages of 65 and 74 years are completely edentulous. Of the different proposed predictors and risk factors, low income and education levels have the highest correlation with tooth loss. While the incidence of complete edentulism in the United States has progressively declined over the past decade, the continued growth of the population strongly suggests that edentulism prevalence will likely remain constant or increase over the next few decades. In patients wearing complete prosthetic appliances, several post-treatment complications may arise, including denture associated Candida species infections and mucosal stomatitis. These type of fungal infections are associated with patient-reported symptoms (e.g. pain or discomfort) and may impede normal oral function. In this study the activity of 11 (antimicrobial and/or antifungal) agents in a dental adhesive carrier against two strains of C. albicans was assessed. In conventional minimal inhibitory concentration (MIC) assays, C. albicans were resistant to histatin 5 and lactoferricin B, yet very susceptible to SMAP28; susceptible to long chain bases sphingosine, dihydrosphingosine, and phytosphingosine; and susceptible to anti-fungal agents amphotericin B, chlorhexidine dihydrochloride, chlorhexidine gluconate, fluconazole, and nystatin. However, in 1% dental adhesive (final concentration) C. albicans were resistant to histatin 5, lactoferricin B, SMAP28, sphingosine, dihydrosphingosine, and phytosphingosine suggesting that the components in denture adhesive may inactivate local innate immune factors in the oral cavity possibly predisposing users to fungal infections in relation to their dental prostheses. In MIC assays in 1% dental adhesive (final concentration) C. albicans were susceptible (p value < 0.05) to amphotericin B, chlorhexidine, dihydrochloride, chlorhexidine gluconate, fluconazole, and nystatin strongly suggesting that these anti-fungal agents could be candidates for inclusion in denture adhesive formulations, and also be used as a prescribed topical treatment in individuals with fungal infections of the oral mucosa.

Doctor of Philosophy / This thesis describes experiments which were carried out at the Institute of Dental Research in Sydney and within the Department of Prosthetic Dentistry at the United Dental Hospital of Sydney between February 1991 and May 1996. The study is concerned with finding practical means of treating chronic atrophic candidosis, also referred to as Candida-associated denture stomatitis and to this purpose two methods of denture disinfection are investigated, namely, sodium hypoclorite denture soak and microwave irradiation. Although the aetiology of denture stomatitis is generally considered to be multifactorial, there is sufficient evidence that Candida species and in particular C. albicans play an important role in the aetiology of the condition. In Chapter 1, therefore, the literature review, which provides relevant background information for the experiments to be described in later chapters, is primarily concerned with Candida species. The characteristics and distribution of Candida species are described and factors affecting the distribution of or Candida are discussed. The literature relating to the cause of chronic atrophic candidosis is vast and consequently a detailed description is given of Candida-associated denture stomatitis in the section concerned with oral diseases caused by Candida and their treatment. Each of the subsequent chapters, contains a brief literature review of material relevant to the subject of the particular chapter. Chapter 2 describes laboratory work to assess the effect of sodium hypochlorite on the adhesion of Candida species to oral surfaces and the ability of Candida to coaggregate with oral streptococci. The results showed that sodium hypochlorite decreased the ability of Candida species to adhere to both inert surfaces and BECs. However, coaggregation of Candida with streptococci was increased. Thus, hypochlorite if used as a denture soak may initially reduce the ability of Candida species to adhere to the denture surface and may therefore assist the treatment of denture stomatitis. The effects of hypochlorite on the characteristics of Candida species that are associated with tissue invasion are described in Chapter 3. The production of acid proteinase, the formation of germ tubes and presence of major cell wall proteins at 43 and 27 kDa are demonstrated. The ability of the whole cells of certain species of Candida to aggregate human platelets was assessed. The results showed that sodium hypochlorite did not affect proteinase production by Candida species but the rate of germ tube formation and the production of Candida cell wall proteins were increased. Hypochlorite did not affect the ability of certain Candida species to aggregate human platelets. Mechanisms to defend the host against candidal invasion are discussed and include platelet aggregation where aggregated platelets release antimicrobial factors that are active against Candida. Chapter 4 describes an in vitro study to test the effects of sodium hypochlorite and microwave irradiation on the survival of Candida species and oral streptococci on denture surfaces. The results showed that 0.02% sodium hypochlorite denture soak for 8 h will eliminate Candida species and reduce the growth of streptococci. However, microwaving of dentures at medium setting for 6 min will eliminate both Candida and streptococci. This information servers as baseline data for clinical assessments described in Chapters 7 and 8. Denture hygiene is an important factor in the prevention and treatment of Candida-associated denture stomatitis. Hence, a clinical study to assess the microbiology of denture plaque is described in Chapter 5. The results showed that denture plaque was composed mainly of Gram-positive streptococci with varying proportions of Gram-positive rods, Gram-negative cocci and rods and is similar to dental plaque. Candida was not always isolated and when detected constituted a very small proportion (< 1%) of the total aerobic bacterial count. The results of an investigation to test the effect of soft denture liners in lower dentures on the colonization of denture surfaces by Candida species and aerobic bacteria are given in Chapter 6. There was no significant difference in Candida /bacterial colonization of dentures with soft denture liners and those without liners. Chapter 7 describes a clinical study to test the efficiency of sodium hypochlorite (0.02%) over-night denture soak as an effective denture disinfecting agent. Treatment of dentures with hypochlorite over a trial period resulted in reductions of Candida and aerobic bacteria and although the reductions were not significant the effect over the trial period could be assessed. A significant finding was that for the palate, treatment with hypochlorite over the trial period prevented an increase in candidal load. Thus, sodium hypochlorite may function as an effective disinfecting agent when used as 0.02% denture soak for a prolonged period. A pilot study to assess the effectiveness of microwaving dentures for ten min (350 W, 240 MHz) as a potential method of denture disinfection is described in Chapter 8. For practical reasons the dentures were microwaved only once only and therefore the effect over a trial period could not be assessed. However, one treatment resulted in significant reductions in the levels of Candida and aerobic bacteria. These findings have indicated that future research should be carried out to test the effect of daily consecutive microwave treatments on candidal and bacterial growth. The general discussion in Chapter 9 summarizes the data presented in the previous chapters and from the findings conclusions are made concerning the prevention and treatment of Candida-associated denture stomatitis. The limitations of this thesis are recognized and some important aspects of the study are recommended for future research.

Eukaryotic initiation factor 2B (eIF2B) is a heteropentameric guanine nucleotide exchange factor (GEF) that converts inactive eIF2 GDP-bound binary complexes into active eIF2 GTP-bound complexes that can bind initiator t-RNA molecules and ribosomes to begin translation. eIF2B is functionally divided into two subcomplexes: the catalytic core comprised of eIF2B epsilon and eIF2B gamma, and the regulatory core comprised of eIF2B alpha, eIF2B beta and eIF2B delta. While the catalytic subunits are responsible for exerting GEF activity, the regulatory subunits recognize eIF2 and respond to eIF2 alpha phosphorylation. Cellular stress, such as virus infection, inhibits host protein synthesis by activating specific kinases that are capable of phosphorylating the alpha subunit of eIF2, which can then sequester eIF2B to stall guanine nucleotide exchange by a currently unresolved mechanism. Importantly, we demonstrate that loss of eIF2B alpha or expression of a variant of the human eIF2B alpha subunit harboring a single point mutation (T41A) is sufficient to neutralize the consequences of eIF2 alpha phosphorylation, and render primary MEFs significantly more susceptible to vesicular stomatitis virus infection. To extend this analysis, we further exhibit the vital function of eIF2B alpha in protein synthesis through phenotypic studies in yeast. Here, we report that this subunit can sufficiently substitute for its yeast counterpart, GCN3, and reproduce similar growth phenotypes under normal and amino acid deprived conditions. In addition, the human eIF2B alpha-T41A variant was unable derepress GCN4 translation in response to an inhibitor of amino acid biosynthesis in yeast, an activity that requires sensitivity to phosphorylation of the yeast eIF2 alpha homolog, SUI2. Previously, we have demonstrated that vesicular stomatitis virus can infect and replicate to high levels in tumor cells. Moreover, these cells appear to contain defects in eIF2 alpha-mediated translational control, plausibly due to disregulation of eIF2B activity, which overcomes the inhibitory effects of eIF2 alpha phosphorylation. Our data suggest a role for eIF2B, specifically eIF2B alpha, in suppression of translation following virus infection, and imply that this complex may contribute to oncogenic transformation. These results emphasize the importance of eIF2B alpha in mediating eIF2 kinase translation inhibitory activity and may provide insight into the complex nature of viral oncolysis and cellular transformation.