Orientador: Iracilda Zeppone Carlos / Banca: Daniele Cardoso Geraldo Maia / Banca: Clelia Akiko Hiruma Lima / Banca: Cleverton Roberto de Andrade / Banca: Edson Rodrgues Filho / Resumo: Nos últimos vinte anos, os quimioterápicos naturais foram introduzidos no tratamento do câncer e com estas novas perspectivas, vem aumentando o interesse das indústrias farmacêuticas em produtos de origem natural. Além disso, existe um grande interesse na identificação de novos agentes antineoplásicos, que possuam ação citotóxica seletiva associada à atividade imunomodulatória. Neste contexto, o presente estudo investiga possíveis efeitos farmacológicos de Qualea multiflora Mart (Vochysiaceae), utilizada na medicina popular do cerrado para o tratamento de úlceras, gastrites, amebíase, diarréia com sangue, cólicas intestinais e inflamações. A partir de resultados in vitro, propôs-se avaliar um tratamento in vivo em modelo tumoral mamário murino empregando-se uma fração terpênica (FT) derivada de Qualea multiflora (Q. multiflora) e as substâncias isoladas presentes nesta fração, lupeol (LP), lupenona (LP) e friedelina (FR), concomitantemente à avaliação dos efeitos imunológicos das substâncias testadas nos animais portadores de tumor. Os resultados parciais do presente estudo mostraram que as substâncias isoladas LP, LN e FR e a fração FT, na maioria das concentrações testadas, não apresentaram efeito tóxico in vitro sobre as células imunes, macrófagos e linfócitos . No entanto, LP, LN, FR e FT apresentaram excelente atividade citotóxica contra a linhagem tumoral LM3. AE não mostrou efeito tóxico in vitro contra a linhagem LM3. Por outro lado, a droga padrão Taxol (TX), apresentou pouco efeito tóxico contra a linhagem tumoral LM3, e acentuado efeito citotóxico contra macrófagos, indispensáveis à contenção tumoral. A partir destes resultados foram selecionadas as concentrações ideais para tratamento in vivo. Os resultados mostraram expressiva inibição do volume tumoral quando os animais foram tratados... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: In the last twenty years, natural chemotherapeutic treatment were introduced into treatment of cancer and with these new perspectives, it has been increasing the interest of pharmaceutical products from natural origin. Moreover, there is great interest in identifying new anticancer agents that have selective cytotoxic activity associated with immunomodulatory activity. In this context, this study investigates possible pharmacological effects of Qualea multiflora (Q. multiflora) Vochysiaceae family, used in folk medicine of Brazilisn cerrado in the treatment of ulcers, gastritis, amebiasis, bloody diarrhea, intestinal pain and inflammation. From in vitro results, this study proposed to evaluate an alternative form of treatment for breast cancer employing a terpenic fraction (FT) derived from the chloroform extract of Q. multiflora and the isolated compounds present in this fraction, Ellagic Acid (EA), lupeol (LP), Lupenona (LP) and friedelin (FR) in an in vivo model of murine breast cancer concurrently to assess the immunological effects of the substances tested in animals with tumor. Partial results of this study showed that the isolated compounds AE, LP, LN and FR and FT fraction, most of the concentrations tested, showed no toxic effect in vitro on immune cells, macrophages and lymphocytes. However, LP, LN, FR and FT showed excellent cytotoxic activity against tumor line LM3. AE showed no toxic effect in vitro against strain LM3. Moreover, the standard drug Taxol (TX) showed little toxic effect against LM3 type of tumor, and marked effect against cytotoxic macrophages necessary for the containment of the tumor. From these results we selected the optimal concentrations for in vivo treatment. The results showed significant inhibition of tumor volume when the animals were treated... (Complete abstract click electronic access below) / Doutor

O conhecimento científico da doença meningocócica cresceu significativamente desde que a natureza epidêmica da enfermidade foi pioneiramente descrita por Vieusseux no começo do século dezenove. De fato, dois séculos depois, houve avanços revolucionários nas esferas de saúde pública, técnicas diagnósticas, antibioticoterapia, assim como nas terapias adjuvantes envolvendo modulação das cascatas de coagulação e inflamação. Além disso, terapia de suporte para manter a homeostase via monitorização do status hemodinâmico, empregos de vasopressores, transfusão de plasma e suporte respiratório. Contudo, as taxas de letalidade e morbidade desta infecção não se alteraram significativamente nas últimas três décadas. Anualmente, a Organização Mundial de Saúde estima que ocorram 1.2 milhões de novos casos da doença, resultando em 135.000 mortes. No Brasil, anualmente, há 3500 casos da doença com uma taxa de incidência igual a 2 casos por 100.000 habitantes e uma taxa de letalidade igual a 20%. No Instituto de Infectologia Emílio Ribas (IIER) há 100 casos por ano. Assim, nós observamos um amplo espectro clínico da infecção, incluindo manifestações dramáticas como purpura fulminans e complicações atípicas como peritonite. O propósito deste estudo foi avaliar as características clínicas e laboratoriais, assim como a severidade da doença nos pacientes hospitalizados no IIER entre 2003 e 2004. Assim, pacientes previamente hígidos com diagnóstico de infecção meningocócica foram incluídos neste estudo prospectivo. Durante o período de estudo um total de 91 (53 homens e 38 mulheres) pacientes foram identificados como casos confirmados de infecção meningocócica como previamente descrito em materiais e métodos. Culturas de sangue, líquor, ou biópsia de pele de 39 pacientes (43%) foram positivas para N. meningitidis. Todos pacientes tiveram o exame de contraimunoeletroforese ou teste de aglutinação do látex positivo. A idade mediana dos pacientes foi igual a 6 anos (variação, 2 a 16 anos). A letalidade foi igual a 14% (13/91) e seqüelas ou complicações clínicas ocorreram em 63% (49/78) dos sobreviventes. Todos pacientes morreram de choque séptico e o tempo mediano entre hospitalização e óbito foi igual a 24 horas (variação, 18-72 horas). Concluindo, os resultados deste estudo prospectivo criam um elo entre infecção meningocócica e uma constelação de fenômenos fisiopatológicos: hipocalemia, hipomagnesemia, hipocalcemia, hiponatremia, hipoglicemia, hiperglicemia, leucopenia, coagulopatia e plaquetopenia. Os níveis de IL-6 e IL-10 corresponderam bem com a classificação baseada em parâmetros clínicos. A letalidade foi igual a 14% (13/91) e seqüelas ou complicações clínicas ocorreram em 63% (49/78) dos sobreviventes. Uma complexa interação entre mediadores é responsável por determinar a severidade da infecção e tais observações podem ser utilizadas para identificar fatores associados com a gravidade na fase aguda da infecção meningococócica / Scientific knowledge of meningococcal infection has increased greatly since the epidemic nature of the illness was first described by Vieusseux at the dawn of the nineteenth century. In fact, two centuries later, revolutionary advances have been made in public health measures, diagnostic procedures, antimicrobial therapy, and adjunctive therapies involving modulation of the inflammatory and coagulation cascades. In addition, supportive care facilities can maintain homeostasis by monitoring hemodynamic status, administering vasoactive agents and/or plasma exchange, and providing respiratory support. However, the prognosis of and case fatality rate among patients affected by meningococcal disease have not changed significantly over the past 3 decades. The World Health Organization estimates that there are 1.2 million cases of meningococcal disease and 135,000 related deaths annually. In Brazil, 3500 cases are reported annually, with a median incidence of 2 cases per 100,000 population and a case-fatality rate of 20%. At the Emílio Ribas Institute of Infectology (ERII), the incidence of meningococcal disease is approximately 100 cases per year. Therefore, we have observed a broad range of clinical presentations of N. meningitidis infection, including dramatic manifestations as purpura fulminans and atypical complications such as peritonitis. The purpose of this study was to evaluate the clinical features, laboratory features, and the severity of meningococcal disease in patients admitted to the ERII between 2003 and 2004. Therefore, consecutive previously healthy patients with a diagnosis of meningococcal disease were included in a prospective cohort study. During the study period a total of 91 (53 males and 38 females) patients were identified as confirmed cases of meningococcal infection as previously described in material and methods. Cultures of blood, CSF, or skin biopsy specimens from 39 patients (43%) yielded N. meningitidis. All patients had positive counterimmunoelectrophoresis or latex agglutination test in blood or CSF. The median age of the patients was 6 years (range, 2 years to 16 years). The case fatality rate was 14% (13/91) and sequelae or clinical complications occurred in 63% (49/78) of the survivors. All patients died of irreversible septic shock and the median duration from the hospitalization until death was 24 hours (range, 18-72 hours). In conclusion, results from this prospective cohort study support a link between meningococcal infection and a constellation of pathophysiological phenomena: hypokalemia, hypomagnesemia, hypocalcemia, hyponatremia, hypoglycemia, hyperglycemia, leukopenia, coagulopathy, and thrombocytopenia. Both pro-inflammatory IL-6 and anti-inflammatory IL-10 corresponded well with a classification based on clinical parameters. The case fatality rate was 14% (13/91) and sequelae or clinical complications occurred in 63% (49/78) of the survivors. A complex interplay of mediators is responsible for determining severity of disease and these observations can be used to identify factors associated with severity in the acute phase of meningococcal infection

INTRODUÇÃO: Lúpus eritematoso (LE) é uma doença inflamatória crônica, autoimune. A presença de citocinas do tipo 1 nas lesões cutâneas discoides sugere que estas sejam críticas para a indução, desenvolvimento e manutenção destas manifestações. As citocinas do tipo 2 em combinação com Interferon- local podem estar relacionadas com a fisiopatologia do lúpus cutâneo. O perfil das citocinas ainda é desconhecido nas lesões orais do LE. MÉTODOS: Foram investigadas e comparadas as expressões das citocinas Th1 e Th2, representadas por IL-4, IL-5, IL-6, IL-10, IL-12, fator de necrose tumoral alfa (TNF-) e interferon gama (IFN-), de 29 biopsias de LE de mucosa intraoral (área não exposta ao sol) e semimucosa labial (área exposta) por meio da técnica de imuno-histoquímica. RESULTADOS: O infiltrado inflamatório das lesões de LE foi fortemente positivo para IFN- (97%) e TNF- (90%), ambas citocinas Th1. Interleucina 10 (IL-10), citocina Th2, foi fortemente expressa. Outras citocinas foram apenas moderadamente positivas. O padrão de citocinas foi semelhante nas lesões orais do LE na mucosa intraoral (área não exposta) e na semimucosa labial (área exposta). CONCLUSÕES: As lesões orais de LE estão associadas tanto a citocinas Th1 quanto a citocinas Th2, caracterizadas por expressão forte de IFN-, TNF- e IL-10. Além disso, os achados sugerem que apesar da radiação ultravioleta estar envolvida na indução das lesões de LE, os mecanismos de formação das lesões podem ser similares nas áreas expostas e não expostas ao sol. / Lupus erythematosus (LE) is a chronic inflammatory disease, autoimune. Presence of type 1 cytokines in cutaneous discoid lesions suggests that they may be critical for induction, development and maintenance of these manifestations. Type 2 cytokines in combination with local interferon (IFN-) are thought to be related to the physiopathology of cutaneous LE. Cytokines profiles are still unknown in oral LE lesions. Material and Methods: Expression of Th1 and Th2 cytokines (including IL-4, IL-5, IL-6, IL-10, IL-12, tumor necrosis factor (TNF-) and IFN- was investigated and compared in 29 biopsies of intra-oral (sun-protected) and labial lesions (sun-exposed) of LE using immunohistochemistry. Results: Inflammatory infiltrate of LE lesions was strongly positive for IFN- (97%) and TNF- (90%), both Th1 type cytokines. Interleukin-10, a Th2 cytokine was also strongly expressed. Other cytokines were only mildly positive. Cytokines patterns were similar in intra-oral (sun-covered) and labial (sun-exposed) LE lesions Conclusions: Oral LE lesions are associated with both type 1 and type 2 cytokines, characterized by stronger expression of IFN-, TNF- and IL-10. These findings suggest that although ultraviolet light is involved in the induction of LE lesions, mechanisms of lesions formation may be similar in sun-exposed as well as sun-covered areas.

Background: Lewy body dementia (LBD), consisting of Parkinson's disease dementia (PDD) and dementia with Lewy bodies (DLB), is known to make up more than 15% of dementia cases at autopsy, however the clinical prevalence rate is reported to be much lower at around 5-6%. Difficulties with diagnosis and/or lack of specific treatments may contribute to this difference. This study investigated the diagnosis and management pathways of LBD and whether inflammation could play a role in the pathophysiology and hence provide a route for future diagnostic and treatment pathways. Methods: Clinical diagnostic rates of LBD in clinics across several NHS trusts in East Anglia were reviewed, followed by an in-depth notes review of patients identified with LBD together with age and gender matched controls. A literature review of the current evidence for inflammation in LBD, preceded a case control study to investigate further. Nineteen DLB patients together with 16 age and gender matched healthy controls underwent [11C]PK11195 PET imaging, and the same cohorts, plus an additional 10 matched control subjects underwent peripheral cytokine analysis. Results: The clinical prevalence rate of LBD was low compared to the known pathology rates, with delays identified in the diagnosis of DLB compared to other dementia subtypes. Delays were also seen between the onset of dementia symptoms and the clinical diagnosis of dementia in Parkinson's disease (PD). The literature review identified studies providing evidence of inflammation in PD but few studies had been carried out in DLB. PET imaging revealed microglial activation negatively correlated with disease severity in DLB, suggesting inflammation occurs early in the disease. DLB patients also showed evidence of differences in cytokine levels compared to healthy controls. Conclusion: The study showed evidence of inflammatory changes in DLB, providing a potential target for treatment and/or biomarkers, that could assist in increasing clinical diagnostic rates.

Clinical and preclinical studies revealed that heart failure induces depression. Injury of myocardial tissue initiates an inflammation cascade that extends to the CNS and might contribute to depression following myocardial infarction (MI). Sex differences were noticed in the progression of heart failure and depression in clinical studies. We hypothesized that depression-like behavior induced by HF post-MI is influenced by sex through modulation of inflammation pathway. First, we evaluated sex differences in depression-like behavior in male and female rats at 8-10 weeks post-MI, as well as circulating cytokines, the extent of inflammation in the PFC, PVN, and amygdala, and mBDNF levels in the PFC and amygdala that are affected by neuroinflammation. Then we evaluated the effect of ovariectomy (OVX) and whether estrogen replacement with 17β-estradiol (E2) prevents post-MI induced depression-like behavior through inhibiting neuroinflammation. Thirdly we evaluated the role of inflammation for cardiac dysfunction and development of depression-like behavior in OVX female rats post-MI by oral treatment with pentoxifylline (PTX). Male rats developed depression-like behavior by ten weeks post-MI but not females as assessed by sucrose preference test (SPT) and forced swim test (FST). Both developed similar cardiac dysfunction and a comparable increase in plasma and PVN cytokine levels, but cytokine levels increased and mBDNF levels decreased only in the PFC of male rats post-MI. OVX per se decreased sucrose consumption and induced passive behavior assessed by SPT and FST, respectively.The combination of MI and OVX aggravated depression-like behvaiour. E2 treatment prevented the development of mild depression-like behavior in OVX and severe symptoms in OVX female rats post-MI. E2 had no effect on cardiac dysfunction in OVX female rats at 10 weeks post-MI. Despite the similar increase in circulating cytokines in OVX ± E2 at 10 weeks post-MI, E2 decreased the proinflammatory cytokines and increased IL-10 (anti-inflammatory cytokine) in the PFC. Finally, we evaluated the role of neuroinflammation in depression-like behavior in OVX female rats post-MI through inhibiting cytokine production by administering oral PTX. PTX prevented depression-like behavior in OVX female rats post-MI and reduced cytokines levels in plasma, PVN and PFC. However, PTX did not affect the progression of cardiac dysfunction at 10 weeks post-MI. Sex determines the development of depression-like behavior in HF post-MI and neuroinflammation appears to play a critical pathway that is affected by sex and can be inhibited by hormonal replacement or anti-inflammatory agents.

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Abstract not available

Since its discovery 15 years ago there has been an explosion of research in the field of DNA immunisation. Unfortunately despite early promises that DNA immunisation had the potential to cure almost any infectious disease, autoimmune disease or even cancer, progress towards clinical trials has been slow. This has been due in part to the huge range of permutations possible in delivering the DNA. One approach is to deliver the DNA by gene gun. Gene gun delivery is a very efficient way of transfecting cells however also has a number of possible disadvantages. These drawbacks include a weak immunogenicity in larger animals as well as the tendency to bias towards the development of a strong type 2 response. In an effort to enhance antigen-specific immune responses and counter the type 2 polarisation of gene gun delivery, a series of DNA vaccines were created where the extracellular portion of the hemagglutinin (HA) gene from influenza A/PR8/34 virus was genetically fused the type 1 cytokines IFNγ, IL-12 and IL-23. Interleukin-23 has been recently discovered and even though both IL-12 and IL-23 contain the p40 subunit they seem to have dissimilar functions. The vaccine constructs were first tested in cellular assays in vitro to ensure correct production and biological activity of the attached cytokines. They were then delivered in various combinations to groups of BALB/c mice to test development of immune responses and the effect of different delivery regimes. Finally mice were immunised then challenged with live influenza virus to determine the different DNA vaccines� protective efficacy. DNA vaccines containing the HA gene alone (pHA) or fused to IFNγ (pIFNγHA), IL-12 (pIL-12HA) or IL-23 (pIL-23HA) were successfully constructed. The fusion of the HA gene to the genes for IFNγ, IL-12 or IL-23 did not significantly disturb the structure of the antigen or prevent the biological actions of the cytokines. Mice immunised three times with pHA had high titres of serum IgG1 antibody and their splenocytes produced approximately equal amounts of IFNγ and IL-5. Co-delivery of IFNγ was unable to alter immune responses regardless of whether it was delivered at the first, last or during all immunisations. Surprisingly co-delivery of IL-12 acted to suppress both antibody and cellular immune responses, possibly through an IFNγ/nitric oxide feedback loop. On the other hand co-delivery of IL-23 tended to enhanced immune responses and, while it did not significantly alter the type 1 to type 2 balance, it was able to increase the ability of mice to clear live influenza virus from their lungs when they were challenged 26 weeks after immunisation. This protection was associated with increased levels of neutralising antibody in the serum of pIL-23HA immunised mice. This research has illuminated several of the pitfalls in the development of DNA vaccines and the use of cytokine as adjuvants. However it has also broadened our understanding of IL-23 and implies that IL-23 could be effectively used to increase the development of longterm immunity after immunisation.

The pressure-tension hypothesis is the governing dogma of orthodontic tooth movement. This theory proposes that the application of loads to the crown of a tooth during orthodontic mechano-therapy results in differential site-specific reactionary strains in the para-dental tissues. Briefly, following the application of orthodontic load the bone and periodontal ligament (PDL) on one side of the tooth is placed in compression favoring bone resorption, while on the other side of the tooth they are placed in tension favoring osteogenesis The present in vitro model provides a surrogate for the PDL on the tension side of the tooth during orthodontic tooth movement and aims to identify mechanically induced changes in the expression of osteo-regulatory cytokines in human PDL cell cultures in response to tensile mechanical strain. Materials and Methods: PDL explants were obtained from pathology free bicuspids of two human subjects following extraction of the teeth for orthodontic purposes. Following serial passage, cells were plated on Uniflex� plates and consigned to either the experimental or control groups. Experimental cells were exposed to a cyclic uniaxial tensile mechanical strain for 6,12 or 24 hours using the Flexercell FX 4000 strain unit. Total RNA was extracted using a two-step procedure and samples were analysed using real-time RT-PCR assays for a range of osteo-regulatory cytokines. Results: Human PDL cells expressed mRNA for a range of cytokines of known significance to osteogenesis and osteoclastogenesis in response to mechanical stimulation. Conclusions: The production of osteo-regulatory cytokines by PDL cells in response to mechanical strain suggests that these cells have the potential to contribute to the osseous modeling of orthodontic tooth movement. The presence of osteogenic signalling drive in response to tensile strain tends to support the basic assertions of the pressure-tension hypothesis.

Bibliography: leaves 151-179. / xx, 179 leaves : ill. (some col.) ; 30 cm. / Title page, contents and abstract only. The complete thesis in print form is available from the University Library. / Examines aspects of the distribution of leukocyte subpopulation in human corpus luteum, cytokine determination in human preovulatory follicular fluid, as well as the effects of cytokines on human granulosalutein cells; with the aim of investigating one of the ovarian regulatory systems, which may be controlled by immune cell-derived cytokines. / Thesis (Ph.D.)--University of Adelaide, Dept. of Obstetrics and Gynaecology, 1993