Macrophages are phagocytic cells of the innate immune system, which have a central role in immune surveillance, tissue homeostasis and the immune response to bacterial, viral, protozoan and helminth parasites. It is now appreciated that many tissue resident macrophage (resMΦ) populations, including those in the peritoneal and pleural cavity, are derived prenatally prior to the establishment of definitive haematopoiesis in the bone marrow. Once seeded, these resMΦ populations are long-lived and capable of self-renewal via in situ proliferation driven by CSF-1. An inflammatory insult, such as bacterial infection, results in the recruitment of bone marrow derived macrophages (BMDMΦ) and the disappearance of the resMΦ population. BMDMΦ recruited to the site of infection become classically activated upon engagement of pathogen recognition receptors and subsequent STAT1 induction. Classically activated macrophages (CAMΦ) are highly bactericidal through the production of inflammatory cytokines, which direct the TH1 immune response, and upregulation of iNOS to generate high concentrations of intracellular nitric oxide. During resolution of acute inflammation resMΦ undergo a CSF-1 driven proliferative burst to repopulate the tissue. In contrast to bacterial infection, helminth parasites drive a TH2 immune response characterised by CD4+ T cell production of IL-4, which induces proliferation and alternative activation of the resMΦ population, thereby overcoming the need for an inflammatory influx of BMDMΦ. Alternatively activated macrophages (AAMΦ) are generated through signalling from the IL-4Rα subunit and subsequent expression of the molecules RELMα, YM1 and arginase-1. While both BMDMΦ and resMΦ upregulate RELMα, YM1 and Arg-1 in response to IL-4Rα stimulation, microarray analysis revealed an otherwise diverse transcriptional and cell surface phenotype between these populations. It was hypothesised that the diverse modes of macrophage accumulation enlisted by bacterial and helminth parasites, combined with the distinct alternative activation phenotypes employed by BMDMΦ and resMΦ populations would translate into important functional differences as regards anti-parasitic immunity. Chapter 1 and 2 of this thesis addresses the importance of macrophage origin during infection with the filarial nematode Litomosoides sigmodontis, taking advantage of the naturally occurring resistant C57BL/6 and susceptible BALB/c strains. A large disparity in MΦ accumulation was observed throughout the infection time course, with significantly larger numbers present within the pleural cavity of resistant C57BL/6 mice. This difference in MΦ number was a reflection of enhanced F4/80hi resMΦ accumulation. Through Ki67hi staining and the use of CCR2-/- and partial bone marrow chimeric mice, the expanded F4/80hi population in resistant C57BL/6 mice was shown to be a result of proliferation of the local F4/80hiGATA6+CD102+ resMΦ population. A high degree of BMDMΦ incorporation into the resMΦ pool through assumption of an F4/80hiGATA6+CD102+ phenotype was observed in both naïve and infected bone marrow chimeric animals, supporting a recent publication showing gradual incorporation of these cells into the resMΦ niche with age. Importantly, the degree of BMDMΦ incorporation into the F4/80hi population was equivalent between naïve and infected animals, despite a 27-fold difference in cell number, illustrating that expansion is a result of proliferation of local resMΦ, independent of origin. Susceptibility was marked by reduced resMΦ proliferation and enhanced recruitment of bone marrow derived F4/80lo MΦ and monocytes. These recruited BMDMΦ displaced the resMΦ population, failed to integrate the resMΦ niche and were highly positive for PD-L2, a marker specific to BMD AAMΦ. Prevention of monocyte influx and subsequent resMΦ displacement resulted in increased worm killing and a stronger TH2 immune response in susceptible BALB/c mice, thereby confirming a detrimental role for BMD AAMΦ in worm killing. Conversely, in order to confirm a protective role for the expanded resMΦ in resistant C57BL/6 mice we attempted to deplete the resMΦ population through intrapleural delivery of clodronate-loaded liposomes. Due to technical issues we were unable generate statistically significant results when depleting the resMΦ population, however a trend toward decreased worm killing in the absence of resMΦ is evident. Previously generated microarrays in the lab identified the complement cascade as being highly upregulated by AAMΦ induced in response to Brugia malayi infection compared to BMDMΦ (thioglycollate elicited). To investigate the role of complement in resistance to L. sigmodontis, Chapter 3 briefly phenotypes the macrophage compartment of C3-/- C57BL/6 mice during infection. No differences in worm burden or macrophage phenotype could be detected in C3-/- mice compared to WT controls, however this may be explained through differences in strain or MΦ origin. This chapter provides an important foundation for future studies on complement and its role in worm killing during L. sigmodontis infection. The final chapter of the thesis focuses on examining the bactericidal capabilities of BMD and resMΦ populations. An in vitro system was utilised to assess the interaction of bone marrow derived macrophages (thioglycollate elicited) and ResMΦ (from naïve mice) with Salmonella enterica serovar Typhimurium SL3261. I found that in vitro BMDMΦ are infected by/ingest SL3261 to a much greater degree than resMΦ. The resMΦ population is less efficient at both controlling the spread and killing intracellular SL3261 overtime, compared to the BMDMΦ population. In vivo however, there appears to be no difference in the ability of the monocyte derived F4/80lo MΦ and the F4/80hi resident MΦ to be infected by/ ingest SL3261, nor was a difference in bactericidal ability detected. Ultimately my work highlights that the anti-parasitic functions of MΦ populations are not dictated by origin but rather the activation phenotype upon infection and ability to respond to local stimuli.

Durch Forschungsarbeiten der neueren Zeit ist die Bedeutung der Akut-Phase-Proteine als wichtiger Bestandteil der unspezifischen Abwehr im Organismus sichtbar geworden. Die möglichen Wechselwirkungen zwischen der physiologischen Magen-Darm-Flora und diesen Proteinen sind jedoch weitesgehend unbekannt. In dieser Arbeit wurde die Magen-Darm-Flora von einem LBP-/--Knock-out-Maus-Stamm und dem Wildstamm (jeweils 20 Tiere) untersucht. Dabei wurde die aerobe Gesamtkeimzahl, die aerobe und anaerobe Gram-negative Gesamtkeimzahl und die Gesamtkeimzahl der auf MRS-Agar gewachsenen Keime in Darminhaltsproben der Darmabschnitte Jejunum, Zäkum und Kolon von oben genannten Mäusen erfasst. Zusätzlich wurden sieben serologische Untersuchungen bei 40 Tieren durchgeführt, um mögliche parallele immunologische Reaktionen des Körpers auf Bestandteile der physiologischen Magen-Darm-Flora zu erfassen und vergleichend zu betrachten. Bei den Untersuchungen konnten hochsignifikante Unterschiede bezüglich der Höhe und Zusammensetzung der aeroben Gesamtkeimzahl, der aeroben Gram-negativen Gesamtkeimzahl, der anaeroben Gram-negativen Gesamtkeimzahl und der Laktobazillen-Gesamtkeimzahl festgestellt werden. Dabei waren bei der LBP-/--Gruppe, den gendeletierten Mäusen, die aerobe Gesamtkeimzahl signifikant erhöht, die anderen Keimzahlen, insbesondere die Keimzahlen der aeroben und anaeroben Gram-negativen Bakterien und der Laktobazillen signifikant erniedrigt. Zusätzlich konnten Abweichungen verschiedener Koloniemerkmale wie Hämolyseverhalten, Koloniemorphologie und Genausprägung zwischen der LBP-/-- und der LBP+/+-Gruppe festgestellt werden. Bei den serologischen Untersuchungen konnten die Ergebnisse der mikrobiologischen Untersuchungen bestätigt werden. Es wurden signifikante Erhöhungen der relativen Antikörperspiegel gegenüber ausgewählten bakteriellen Strukturen, von Mikroorganismen, welche auch signifikant häufiger von den jeweiligen Gruppen isoliert werden konnten, festgestellt. Diese Ergebnisse lassen auf einen möglichen Zusammenhang zwischen der Gendeletion, also der Nichtexpression von LBP, und der Zusammensetzung der Magen-Darm-Flora schließen. Inwiefern dies auf direkten oder indirekten Einfluss des LBP auf die Magen-Darm-Flora zurückzuführen ist, müssen weitere Untersuchungen zeigen. Dabei sollten noch andere Einflussfaktoren, wie z. B. die Varianz des LBP in seiner genotypischen Ausprägung, berücksichtigt werden.

Submitted by Alessandra Portugal (alessandradf@ioc.fiocruz.br) on 2013-09-17T16:37:38Z No. of bitstreams: 1 Dissertacao completa - Lenilton Silveira.pdf: 2646876 bytes, checksum: fbad76d9c78a2517359d01fc6fc96b45 (MD5) / Made available in DSpace on 2013-09-17T16:37:38Z (GMT). No. of bitstreams: 1 Dissertacao completa - Lenilton Silveira.pdf: 2646876 bytes, checksum: fbad76d9c78a2517359d01fc6fc96b45 (MD5) Previous issue date: 2011 / Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Rio de Janeiro, RJ, Brasil / Neste estudo nós realizamos um monitoramento cinético, ex vivo, da resposta imune do hospedeiro, induzida durante infecção murina por L. (L.) amazonensis. Fêmeas de camundongos BALB/c foram inoculadas subcutaneamente no coxim plantar esquerdo com 106 promastigotas de fase estacionária e o progresso da infecção foi monitorado por medição da área de lesão. Os linfócitos dos linfonodos poplíteos drenantes da lesão (LNs) foram analisados por citometria de fluxo (FACS - fluorescence-activated cell sorting) e por ressonância plasmônica de superfície (SPR - surface plasmon resonance). Os resultados de FACS indicaram que o perfil de linfócitos é definido na primeira semana de infecção e caracterizado por elevados níveis de linfócitos B e por uma diminuição de linfócitos T. O percentual de linfócitos B foi três vezes maior quando comparado com o dos camundongos não infectados e os linfócitos T eram 1,5 vezes mais baixo. Estes valores permaneceram constantes ao longo da infecção até o ultimo ponto analisado. Foi observado um discreto aumento (0.3 a 0.5) na relação CD8+/CD4+ nos animais infectados, enquanto nos não infectados esta relação permaneceu em torno de 0.35, até o último dia do estudo. Adicionalmente, nós realizamos ensaios de detecção de linfócitos T CD8+ dos LNs, em tempo real, por SPR, utilizando moléculas H-2 Ld:Ig ligadas à peptídeos da região COOH-terminal de cisteína-proteinase B de L. (L.) amazonensis (P6.3= EFCLGGGL, P6.4= CLGGGLCL, P6.5= EFCLGGGLC, P1.7= VMVEQVICF, P1.9= MVEQVICFD, P1.10= VEQVICFD). Os peptídeos testados indicaram valores de constante de associação (0.16x107 – 22.0x107) e dissociação (0.047-0.123) favoráveis a formar complexos com os dímeros H-2 Ld em pH neutro, com valores de energia livre de Gibbs < 0. De forma geral, a ligação das células ao complexo H-2 Ld:Ig/peptídeo, mostrou uma resposta (RU - Response unit) de dissociação maior em células de animais infectados (113 RU – 465 RU) que nas dos animais não infectados (71 RU – 304 RU). A resposta aos complexos H-2 Ld:Ig/peptídeo revelou diferenças de reconhecimento nas semanas avaliadas, como alta resposta ao P1.10 (227 RU) na 3ª semana e ausência de resposta ao P1.7 na 16ª semana. Em uma mesma semana de análise, vimos que o nível de reconhecimento dos peptídeos difere entre eles, uns apresentando alto valor de RU e outros um valor mais baixo: 3ª (P1.10>P6.3>P1.9>P1.7>P6.4>P6.5), 4ª (P6.3.>P6.5>P1.9>P6.4>P1.7), 6ª (P6.4>P1.10>P1.9>P1.7>P6.5), 12ª (P1.9.>P6.3>P1.10>P6.5), 18ª semana (P6.4=P1.9>P6.3>P1.10>P1.7). Embora, apenas, dois peptídeos tenham sido testados na 1ª semana, a maior resposta foi relacionada ao P1.7 (161 RU), seguido do P1.9 (115 RU). Os dados gerados indicaram uma flutuação na população dos linfócitos T e B, assim como o envolvimento de linfócitos T CD8+ direcionados a peptídeos da região COOH-terminal da CPB, ao longo da infecção / In this study, we performed a kinetic ex vivo monitoring of host immune responses during murine infection with Leishmania (Leishmania) amazonensis. Female BALB/c mice were injected subcutaneously in the left hind footpad with 106 promastigotes in stationary growth phase and the progression of infection was monitored by measuring lesion area expansion. Lymphocytes from lesion draining popliteal lymph nodes (LNs) were analyzed by fluorescence-activated cell sorting (FACS) and surface plasmon resonance (SPR). FACS results indicated that the lymphocytes profile is defined in the first week post infection and was characterized by high levels of B cells and by a decrease in T cell numbers. The percentage of B lymphocytes was three times higher when compared to the non-infected mice and T lymphocytes were up to 1.5 times lower. These values remained constant throughout infection until the last point analyzed. It was observed a slight increase (0.3 to 0.5) in the CD8+/CD4+ ratio in the infected mice, whereas in the not infected group this ratio remained around 0.35, until the last assessed infection day. Additionally, we have performed an assay to detect CD8+ T lymphocytes from LNs in real time, by SPR, utilizing H-2 Ld:Ig molecules bound to peptides derived from the COOH-terminal region of cysteine-proteinase B from L. (L.) amazonensis (P6.3= EFCLGGGL, P6.4= CLGGGLCL, P6.5= EFCLGGGLC, P1.7= VMVEQVICF, P1.9= MVEQVICFD, P1.10= VEQVICFD). The peptides tested presented values of association (0.16x107 – 22.0x107) and dissociation constant (0.047-0.123) favorable to form complexes with H-2 Ld at neutral pH, with values of Gibbs free energy < 0. In general, the binding of cells to the H-2 Ld:Ig/peptide complexes showed a response unit (RU) of dissociation higher in cell preparations from infected mice (113 RU - 465 RU) than the cells from uninfected mice (71 RU – 304 RU). The recognition response to the complex H-2 Ld:Ig/peptide were assessed at different weeks as high response to P1.10 (227 RU) at 3th week and absence of response to P1.7 at 16th week. In the same week of analysis, we found that the level of recognition of peptides differs among them, some presenting a high value of RU and others a lower value: 3th (P1.10 >P6.3>P1.9>P1.7>P6.4>P6.5), 4th (P6.3.>P6.5>P1.9>P6.4>P1.7), 6th (P6.4> P1.10>P1.9>P1.7>P6.5), 12th (P1.9.>P6.3>P1.10>P6.5), 18th week (P6.4=P1.9>P6.3>P1.10>P1.7). Although only two peptides were tested at 1th week, the highest response was related to P1.7 (165 RU), followed by P1.9 (115 RU). Generated data indicated a fluctuation in the population of T and B lymphocytes, as well as the involvement of CD8+ T cells directed to peptides of the COOH-terminal region of the CPB, throughout the infection

Made available in DSpace on 2015-07-22T20:50:07Z (GMT). No. of bitstreams: 0 Previous issue date: 2010-09-29 / A epilepsia acomete cerca de 1 a 2% da população mundial e carece de propostas terapêuticas mais eficazes. Aproximadamente 30% das epilepsias são sintomáticas e a epilepsia do lobo temporal (ELT) se destaca pela alta incidência e gravidade. Cerca de 30% de pacientes com ELT possuem crises refratárias ao tratamento medicamentoso, sendo necessário, muitas vezes, intervenção cirúrgica para o controle das mesmas. A investigação dos mecanismos fisiopatológicos empregando tecido humano extraído cirurgicamente, embora muito importante, tem a limitação de refletir apenas o processo crônico em que as alterações moleculares, bioquímicas e de neuroplasticidade resultantes do quadro epiléptico já se encontram estabelecidos. Nesse sentido, os modelos experimentais são fundamentais por permitir uma melhor abordagem dos mecanismos envolvidos com a epileptogênese, ou seja, aqueles ativados durante a indução até o estabelecimento do quadro epiléptico. O modelo lesional de epilepsia induzida por pilocarpina em pequenos roedores tem sido amplamente utilizado por reproduzir as principais características da ELT. Nesse modelo as crises epilépticas são acompanhadas por uma ação combinada entre o sistema imune, nervoso e endócrino, envolvendo a ativação de mediadores como citocinas, quimiocinas, neurotransmissores e seus receptores celulares. Estes processos têm sido associados com a etiologia da condição epiléptica. No presente projeto, camundongos imunodeficientes BALB/c nude foram submetidos ao modelo de epilepsia induzida por pilocarpina, para caracterização das alterações comportamentais, eletrográficas, neuroquímicas e histológicas comparadas ao seu controle BALB/c. Os resultados indicaram que a integridade do sistema imunológico é necessário para proteger o sistema nervoso central do insulto causado pelo status epilepticus (SE) induzido pela pilocarpina. Após a aplicação da pilocarpina, os animais BALB/c nude apresentaram um maior número de crises tônicoclônicas até atingirem o SE, apresentando um alto índice de mortalidade quando comparados ao BALB/c. Os camundongos BALB/c nude também apresentaram uma maior ativação de c-Fos em áreas relacionadas com a circuitaria das crises e uma menor expressão de parvalbumina, uma proteína ligadora de cálcio intracelular em relação ao BALB/c estudado na mesma condição experimental. Diferenças nas concentrações de neurotransmissores excitatórios e inibitórios também foram observados no córtex e hipocampo do BALB/c nude em relação ao BALB/c. Os camundongos BALB/c nude tiveram aumento na concentração de aminoácidos excitatórios sem a alteração compensatória dos aminoácidos inibitórios durante as crises, conforme ocorreu nos camundongos BALB/c. A morte neuronal avaliada pelas técnicas histológicas de Fluoro Jade-B e Nissl, também se mostrou aumentada em algumas áreas do sistema nervoso central dos camundongos BALB/c nude que apresentaram SE em relação ao BALB/c Os dados sugerem uma correlação positiva entre ativação de c-Fos, concentração de aminoácidos excitatórios e morte neuronal nos camundongos BALB/c nude, sugerindo que a deficiência imunológica possa ter contribuído para a hiperexcitabilidade e vulnerabilidade do sistema nervoso central aos danos causados pelas crises. / Epilepsy affects about 1-2% of world population and lack of effective therapeutic interventions more effective. Approximately 30% are symptomatic of epilepsy and temporal lobe epilepsy (TLE) is distinguished by the high incidence and severity. About 30% of patients with TLE seizures are refractory to treatment, if necessary, often, surgical intervention for control of them. The investigation of pathophysiology mechanism using human tissue extracted surgically, although very important, is limited by reflects only the chronic process in which the molecular, biochemical, and neuronal plasticity alterations resulting from epileptic effects already established. Thus, experimental models are essential for better approach of the mechanisms involved in epileptogenesis. The epilepsy induced by pilocarpine in small rodents has been widely used for reproducing the main features of TLE. In this model, the seizures are accompanied by a combined action between immune system, nervous and endocrine systems, involving the mediators activation such as cytokines, chemokines, neurotransmitters and their cell receptors. These processes have been associated with the etiology of the epileptic condition. In this project, immunodeficient BALB/c nude mice submitted to epilepsy model induced by pilocarpine to characterize behavior, electrographic, histological and neurochemical changes compared to their control BALB/c. The results indicated that integrity of the immune system is necessary to protect the central nervous system insult caused by status epilepticus. After administration of pilocarpine, the BALB/c nude mice showed a larger number of tonic-clonic seizures, with a high mortality rate when compared to BALB/c. The BALB/c nude also showed a greater activation of c-Fos in areas related to the circuitry of the crisis and a lower expression of parvalbumin, an intracellular calcium-binding protein compared to BALB/c studied in the same experimental condition. Differences concentrations of excitatory and inhibitory neurotransmitters were also observed in cortex and hippocampus of the BALB/c nude compared to BALB/c. The BALB/c nude showed increased of the excitatory amino acids concentrations without compensatory alteration in the inhibitory amino acids during seizure, as occurred in BALB/c. Neuronal death measured by Fluoro Jade- B and Nissl histological techniques, was also increased in some areas of the central nervous system of BALB/c nude SE presented relative to BALB/c. The data suggest a positive correlation between c-Fos activation, excitatory amino acids concentration and neuronal death in BALB/c nude mice, suggesting that immune deficiency may have contributed to the hyperexcitability and vulnerability of the central nervous system injury caused by seizure. / TEDE / BV UNIFESP: Teses e dissertações

Submitted by Hiolanda Rêgo (hiolandarego@gmail.com) on 2015-10-20T16:10:30Z No. of bitstreams: 1 Dissertação_ICS_ Gabriele Costa Rodrigues.pdf: 669924 bytes, checksum: 39da9a8e9734a6d7491ddbfaf0f55cd4 (MD5) / Made available in DSpace on 2015-10-20T16:10:30Z (GMT). No. of bitstreams: 1 Dissertação_ICS_ Gabriele Costa Rodrigues.pdf: 669924 bytes, checksum: 39da9a8e9734a6d7491ddbfaf0f55cd4 (MD5) / A linfadenite caseosa é uma doença infecciosa crônica que afeta ovinos e caprinos, causada por uma bactéria intracelular facultativa, Corynebacterium pseudotuberculosis. Pouco se conhece sobre a imunorreatividade induzida por diferentes linhagens deste microrganismo, bem como sobre o efeito no hospedeiro de seus antígenos secretados. O objetivo deste trabalho foi avaliar o reconhecimento antigênico e a ativação de células esplênicas de camundongos Balb/c infectados com duas linhagens de C. pseudotuberculosis, utilizando para tanto dois preparados antigênicos obtidos a partir da linhagem atenuada T1, denominados SeT1 e Q5, ao longo dos primeiros 35 dias após a inoculação. Os camundongos foram infectados com 102 CFU da linhagem VD57 ou da linhagem T1 e sacrificados nos dias 0, 15 e 35. A resposta humoral e celular foram avaliadas pela identificação de IgG total e subclasses, específicas contra estes antígenos secretados, bem como pela quantificação de citocinas produzidas “in vitro”, contra estas mencionadas biomoléculas. O grupo infectado com a linhagem selvagem VD57 apresentou diversas lesões granulomatosas, ao contrário dos camundongos infectados com a linhagem atenuada T1 que não desenvolveram lesões. A linhagem VD57 induziu expressiva produção de TNF-alfa, IFN-gama e IL-10, enquanto que a produção de IL-4 foi semelhante nos dois grupos, apesar de ter apresentado valores significativos com a estimulação pela fração Q5. Através do “Western Blotting” foi possível identificar quatro bandas na fração antigênica Q5, reativas com soros de camundongos infectados com a linhagem T1. Utilizando-se o ensaio imunoenzimático ELISA observou-se uma baixa resposta pelo grupo infectado com a linhagem T1 e uma resposta expressiva dos animais infectados com a linhagem VD57 apenas no dia 35. Neste último grupo, notou-se resposta predominante de IgG1 e IgG2a quando se utilizou como antígeno, respectivamente os preparados Q5 e SeT1. Os dados mostram que os dois preparados antigênicos foram capazes de estimular a imunidade humoral e celular, entretanto, ativando diferentemente os padrões de resposta Th1 e Th2.

No description available.

Soluble dietary fibers (SDF) are fermented more than insoluble dietary fibers (IDF), but their effect on colonic bacterial community structure and function remains unclear. Thus, bacterial community composition and function in the colon of BALB/c mice (n = 7) fed with a high level (approximately 20%) of typical SDF, oat-derived beta-glucan (G), microcrystalline cellulose (M) as IDF, or their mixture (GM), were compared. Mice in group G showed a lowest average feed intake (p < 0.05) but no change on the average body weight gain (p > 0.05) compared to other groups, which may be associated with the highest concentration of colonic propionate (p < 0.05) in these mice. The bacterial alpha-diversity of group G was significantly lower than other groups (p < 0.01). In group G, the relative abundance of bacteria belonging to the phylum Bacteroidetes was significantly increased, whereas bacteria from the phylum Firmicutes were significantly decreased (p < 0.01). The core bacteria for different treatments showed distinct differences. Bacteroides, Dehalobacterium, and Prevotella, including known acetogens and carbohydrate fermenting organisms, were significantly increased in relative abundance in group G. In contrast, Adlercreutzia, Odoribacter, and Coprococcus were significantly more abundant in group M, whereas Oscillospira, Desulfovibrio, and Ruminoccaceae, typical hydrogenotrophs equipped with multiple carbohydrate active enzymes, were remarkably enriched in group GM (p < 0.05). The relative abundance of bacteria from the three classes of Proteobacteria, Betaproteobacteria, Gammaproteobacteria (including Enterobacteriaceae) and Deltaproteobacteria, were significantly more abundant in group G, indicating a higher ratio of conditional pathogenic bacteria in mice fed dietary beta-glucan in current study. The predicted colonic microbial function showed an enrichment of "Energy metabolism" and "Carbohydrate metabolism" pathways in mice from group G and M, suggesting that the altered bacterial community in the colon of mice with the two dietary fibers probably resulted in a more efficient degradation of dietary polysaccharides. Our result suggests that the influence of dietary beta-glucan (SDF) on colonic bacterial community of mice was more extensively than MCC (IDF). Co-supplementation of the two fibers may help to increase the bacterial diversity and reduce the conditional pathogens in the colon of mice.

Atopic dermatitis (AD) typically develops in patients with a history of allergic ailments, and is characterised by an itchy, inflammatory skin condition with scaling, lichenification, papules, excoriations and pruritus. In AD patients a chronic relapsing inflammatory condition is seen, associated with IgE hyper production. AD flares are largely triggered by environmental factors. However, the exact etiology of AD is unclear and there is a pressing need for new treatment regimens as AD is a chronic condition and requires long term treatment. Historically Aloe has been used to treat skin conditions as well as a variety of other diseases. To further explore the pathogenesis and treatment of AD, Balb/c mice were sensitized and challenged with 2,4-dinitrochlorobenzene (DNCB) for atopic dermatitis induction. Thereafter, mice were treated with either Aloe ferox or Aloe vera applied daily on the dorsal skin for 10 consecutive days. A placebo gel was used for the control mice. Blood was collected at the end of the treatment period and serum IgE levels measured. Serum IgE levels were significantly lowered in the Aloe ferox group than in the Aloe vera group. This study demonstrated Aloe’s immunoregulatory potential for alleviating atopic dermatitis through influencing of Th2 cell activation. / Dissertation (MSc)--University of Pretoria, 2013. / gm2014 / Pharmacology / unrestricted

Durch Forschungsarbeiten der neueren Zeit ist die Bedeutung der Akut-Phase-Proteine als wichtiger Bestandteil der unspezifischen Abwehr im Organismus sichtbar geworden. Die möglichen Wechselwirkungen zwischen der physiologischen Magen-Darm-Flora und diesen Proteinen sind jedoch weitesgehend unbekannt. In dieser Arbeit wurde die Magen-Darm-Flora von einem LBP-/--Knock-out-Maus-Stamm und dem Wildstamm (jeweils 20 Tiere) untersucht. Dabei wurde die aerobe Gesamtkeimzahl, die aerobe und anaerobe Gram-negative Gesamtkeimzahl und die Gesamtkeimzahl der auf MRS-Agar gewachsenen Keime in Darminhaltsproben der Darmabschnitte Jejunum, Zäkum und Kolon von oben genannten Mäusen erfasst. Zusätzlich wurden sieben serologische Untersuchungen bei 40 Tieren durchgeführt, um mögliche parallele immunologische Reaktionen des Körpers auf Bestandteile der physiologischen Magen-Darm-Flora zu erfassen und vergleichend zu betrachten. Bei den Untersuchungen konnten hochsignifikante Unterschiede bezüglich der Höhe und Zusammensetzung der aeroben Gesamtkeimzahl, der aeroben Gram-negativen Gesamtkeimzahl, der anaeroben Gram-negativen Gesamtkeimzahl und der Laktobazillen-Gesamtkeimzahl festgestellt werden. Dabei waren bei der LBP-/--Gruppe, den gendeletierten Mäusen, die aerobe Gesamtkeimzahl signifikant erhöht, die anderen Keimzahlen, insbesondere die Keimzahlen der aeroben und anaeroben Gram-negativen Bakterien und der Laktobazillen signifikant erniedrigt. Zusätzlich konnten Abweichungen verschiedener Koloniemerkmale wie Hämolyseverhalten, Koloniemorphologie und Genausprägung zwischen der LBP-/-- und der LBP+/+-Gruppe festgestellt werden. Bei den serologischen Untersuchungen konnten die Ergebnisse der mikrobiologischen Untersuchungen bestätigt werden. Es wurden signifikante Erhöhungen der relativen Antikörperspiegel gegenüber ausgewählten bakteriellen Strukturen, von Mikroorganismen, welche auch signifikant häufiger von den jeweiligen Gruppen isoliert werden konnten, festgestellt. Diese Ergebnisse lassen auf einen möglichen Zusammenhang zwischen der Gendeletion, also der Nichtexpression von LBP, und der Zusammensetzung der Magen-Darm-Flora schließen. Inwiefern dies auf direkten oder indirekten Einfluss des LBP auf die Magen-Darm-Flora zurückzuführen ist, müssen weitere Untersuchungen zeigen. Dabei sollten noch andere Einflussfaktoren, wie z. B. die Varianz des LBP in seiner genotypischen Ausprägung, berücksichtigt werden.

Chikungunya virus (CHIKV) is classified as an alphavirus in the Togaviridae family. This virus is known to rely on Aedes arthropod vectors for its dissemination. Human infection is characterized by rash, high fever, and severe chronic polyarthritis that can last for years. Recently, efforts in developing animal models have been made in an attempt to better understand CHIKV pathogenesis. CHIKV infection starts with a 7 to 10 day long febrile acute phase, in which most of the symptoms occur (rash, fever, and incapacitating pain in joints and muscle). Once the immune system clears most of the viral infection, a chronic phase starts in as many as 70% of the infected patients. Long term virus-related polyarthralgia is the hallmark of the CHIKV chronic phase. It is believed that CHIKV-infected macrophages infiltrate the joints during the acute phase, and CHIKV infects joint tissue and persists in it. Research into the effects of CHIKV infection in human and murine macrophages revealed that CHIKV-infected human macrophages produce high amounts of virions as well as induce the production of pro-inflammatory cytokines and monocyte recruiting chemokines. This contrasts with murine macrophage infection where low quantities of the virus were detected as well as lower production of pro-inflammatory cytokines. This may contribute to the lack of polyarthritis in murine animal models. Current literature suggests that CHIKV’s viral proteins bind and interact with human host cell machinery promoting viral replication more efficiently in humans than in mice. CHIKV-related neuropathology is not the most common outcome of the disease. However, recent outbreaks suggest that this pathology is becoming more prevalent, affecting as many as 30% of confirmed patients. The role of adaptive and innate immunity in CHIKV disease amelioration has been extensively, yet separately, explored. A RAG2-/-γc-/- Balb/c mouse model was used to study the role of these immune pathways and their associated immune cells in CHIKV infection. The mice in this study developed local arthritis at the site of inoculation as well as showed signs of viral invasion in the brain. This study added to the hypothesis that both innate and adaptive immune responses are necessary to ameliorate the disease and that the lack of adequately matured lymphocytes and STAT6-activation deficient macrophages may result in more severe pathologies.