In these studies we sought to determine the role of Fyn kinase and ADAM10 in B cell biology. A disintegrin and metalloproteinase 10 (ADAM10) is a zinc dependent proteinase related to matrix metalloproteinases. ADAM10 has emerged as a key regulator of cellular processes by cleaving and shedding extracellular domains of multiple transmembrane receptors and ligands. In particular, ADAM10 has been identified as a key regulator of lymphocyte development. Here we report that ADAM10 is dispensable for early B cell development within the bone marrow. However, deletion of ADAM10 from all peripheral B cells or in post-switch cells leads to severe impairments in humoral responses. When ADAM10 was deleted from all peripheral B cells a decrease in antigen specific IgG production was seen both with respect to serum levels and IgG ASCs, indicating that plasma cell (PC) differentiation is influenced. Cells producing high affinity antigen specific antibodies were particularly affected, consistent with defects in germinal center (GC) reactions. Moreover, changes in lymphoid architecture were also observed. Consistent with these findings, follicular dendritic cell (FDC)-reticula was undetectable following immunization. On the other hand, when ADAM10 was deleted in post-switch B cells, GC formation and lymphoid architecture were not impaired. Despite normal architecture, however, antibody production was still affected, likely due to abnormal gene expression in ADAM10-deficient PCs. Consistent with this hypothesis, PCs isolated from ADAM10Δ/ΔIgG1-cre+/- showed decreased expression of genes that facilitate plasma cell differentiation and function and increased expression of Bcl6, an inhibitor of PC differentiation. Fyn kinase is a member of the Src protein tyrosine kinase. Fyn is widely expressed in many cell types, including lymphocytes. Fyn has been shown to interact with both the B cell and T cell receptor (BCR and TCR, respectively). While Fyn-deletion did not impair the development of immature T cells and B cells, TCR signaling was altered in mature T cells. Our results demonstrate that Fyn-KO mice have significantly low basal levels of IgG1 and IgG2a. Additionally, these mice displayed delayed kinetics in the production of NP-specific IgG1 and IgG2b, and significantly low NP-specific IgG2a after a T-dependent immunization protocol. Defects in antibody production correlated with significantly reduced numbers of GC B cells, TFH cells and splenic PCs. Moreover, Fyn-KO B cells showed decreased production antibody following in vitro activation. Our results thus demonstrate that Fyn-mediated signaling and B cell ADAM10 expression are necessary for optimal humoral responses.

Influenza A virus (IAV) infection is a serious respiratory disease associated with significant morbidity and mortality worldwide. Annual vaccination is the most effective way to prevent infection and its potentially severe complications; however, the vaccines currently offered have several drawbacks that limit its availability and protective efficacy. Influenza virus-like particles (VLPs), which lack viral genetic material and are non-infectious, represent a promising vaccine candidate. Previous reports have shown VLPs are more immunogenic than subunit or recombinant proteins, and confer protection upon lethal challenge. A critical component of this protection is mediated by influenza HA-specific neutralizing Abs produced by memory B cells and plasma cells, the cellular products of the germinal center (GC) reaction. While preliminary studies have examined the humoral immune response to VLP vaccination, the current study is the first to characterize the GC response in secondary and tertiary lymphoid tissues. Mice were vaccinated with influenza VLPs using three immunization routes: subcutaneous (s.c.), intramuscular (i.m.), and intranasal (i.n.) and the GC response was assessed over time. Robust GC reactions were induced in the dLNs regardless of vaccination route, though the largest response was generated with VLPs s.c. The pattern of isotype expression was remarkably similar between routes, with IgM+ and IgG2+ B cells representing the majority of the GC B cell population. Mucosal immune responses in the upper (nasal) and lower (lung) airway were measured in mice vaccinated i.n. Marked GC reactions were induced in the nasal-associated lymphoid tissue (NALT), while the pulmonary response was relatively modest and short-lived compared to infection with IAV. Within the GC B cell population, IgM+ and IgG2+ B cells made up the majority, similar to the dLN response. Importantly, the pattern of isotype expression induced by VLPs mimicked the response induced by natural IAV infection, and suggests that VLPs contain the necessary innate immune agonists to induce a TH1 biased response.

<p>Uppsatsen behandlar den ekonomiska situationen i "Germinal" av Émile Zola. Det är mycket i romanen som handlar om ekonomi, t.ex. gruvarbetarnas ekonomiska situation och krisen i industrin. Jag har undersökt gruvarbetarnas ekonomiska situation, deras arbetsförhållanden och boendeförhållanden, liksom borgarnas situation. Uppsatsen behandlar också den övergripande ekonomiska situationen och hur denna påverkar industri, gruvarbetare och borgare. Bokens händelser och gruvarbetarnas och borgarnas reaktioner på dessa granskas i uppsatsen, liksom gruvbolagets ekonomi och organisation. Jag har också studerat motsvarande situationer i verkligheten och gjort en jämförelse.</p><p>Le mémoire traite la situation économique dans «Germinal» par Émile Zola. Une grande partie du roman tourne autour de l’économie, soit l’économie des mineurs, soit la crise industrielle. J’ai examiné la situation économique des mineurs, leurs conditions de travail et leur mode de logement aussi bien que la situation des employeurs, la situation économique en </p><p>gros et comment celle-ci influence l’industrie, la bourgeoise et les mineurs. La série d’évènements et la réaction des employeurs et des mineurs est prise en considération ainsi que l’organisation et la situation économique de la Compagnie des Mines de Montsou. J’ai étudié les situations correspondantes dans la réalité aussi, par exemple la situation économique des mineurs, les conditions de travail et la situation des mines de charbon. Une comparaison entre roman et réalité a également été faite.</p>

Uppsatsen behandlar den ekonomiska situationen i "Germinal" av Émile Zola. Det är mycket i romanen som handlar om ekonomi, t.ex. gruvarbetarnas ekonomiska situation och krisen i industrin. Jag har undersökt gruvarbetarnas ekonomiska situation, deras arbetsförhållanden och boendeförhållanden, liksom borgarnas situation. Uppsatsen behandlar också den övergripande ekonomiska situationen och hur denna påverkar industri, gruvarbetare och borgare. Bokens händelser och gruvarbetarnas och borgarnas reaktioner på dessa granskas i uppsatsen, liksom gruvbolagets ekonomi och organisation. Jag har också studerat motsvarande situationer i verkligheten och gjort en jämförelse. Le mémoire traite la situation économique dans «Germinal» par Émile Zola. Une grande partie du roman tourne autour de l’économie, soit l’économie des mineurs, soit la crise industrielle. J’ai examiné la situation économique des mineurs, leurs conditions de travail et leur mode de logement aussi bien que la situation des employeurs, la situation économique en gros et comment celle-ci influence l’industrie, la bourgeoise et les mineurs. La série d’évènements et la réaction des employeurs et des mineurs est prise en considération ainsi que l’organisation et la situation économique de la Compagnie des Mines de Montsou. J’ai étudié les situations correspondantes dans la réalité aussi, par exemple la situation économique des mineurs, les conditions de travail et la situation des mines de charbon. Une comparaison entre roman et réalité a également été faite.

No description available.

Diss. (sammanfattning) Göteborg : Univ., 2008. / Härtill 4 uppsatser.

The studies presented in this dissertation establish the dynamics of Eleven nineteen Lysine-rich leukemia (ELL) family of elongation factors during B cell differentiation and provide a description of ELL3 function in B cells. The transition from a mature naïve B cells into an activated B cell is dependent on a large increase in transcriptional output, which is followed by focused expression on secreted immunoglobulin upon terminal differentiation into plasma cell. While ELL family members have previously been implicated in alternative splicing at the immunoglobulin heavy chain locus in plasma cells, their presence and function prior to differentiation is currently not known. However, the use of elongation factors has been implied by the finding of mostly paused RNA polymerase II in the genome of naïve B cells. In the first study, the expression of transcriptional elongation factor ELL3 is shown to be restricted to activated B cells and B cell lymphomas. All three family members were characterized in B cell lymphoma cell lines, genome wide expression, microarray analysis and primary B cell stimulus. The expression of ELL3 was induced upon activation of B cells concurrently with family member ELL. In addition, the abundant expression of ELL3 was restricted to GC derived B cell lymphoma cell lines. While the expression of ELL is maintained, the expression of ELL3 is diminished and ELL2 is up-regulated in terminally differentiated plasma cells. The expression of master regulator of terminal plasma cell differentiation PRDM1 was inverse correlated with that of ELL3. To further establish PRDM1s role in regulating the ELL family member dynamics, global binding was assessed in plasma cell lines. Chromatin immunoprecipitation followed by quantitative PCR was utilized to identify direct association of PRDM1 at exclusively the ELL3 loci. Ectopic expression of PRDM1 in B cells down regulated the expression of ELL3. Furthermore, two consensus PRDM1 binding sites were defined at the ELL3 loci, which mediate significant repression of the promoter activity. Collectively, these experiments indicate that PRDM1 mediates the switch from ELL3 in B cells to ELL2 in plasma cells. The data presented in the final chapter aimed at defining a function for ELL3 in the cells that express it most abundantly, which are B cell lymphoma cell lines. Transient depletion of ELL3 in a Burkitt’s lymphoma cell line resulted in a diminished proliferation rate due to a severe disruption of DNA replication and its regulators minichromosome maintenance proteins. Additionally, compromised cell division and mitotic regulators were observed along with increased DNA damage and cell death. The data presented here demonstrate a key role for ELL3 in the proliferation and survival of B cell lymphomas and positions ELL3 as an attractive therapeutic target against B cell lymphoma’s with a germinal center origin.

Made available in DSpace on 2014-06-11T19:33:33Z (GMT). No. of bitstreams: 0 Previous issue date: 2001-12-11Bitstream added on 2014-06-13T18:45:26Z : No. of bitstreams: 1 jacomini_jo_dr_jabo.pdf: 3623737 bytes, checksum: 3b6e9aca9b253bde7a9aa506de4fa862 (MD5) / Este estudo teve como objetivo acompanhar o processo de desenvolvimento testicular desde a fase indiferenciada até sua completa formação. Embriões e fetos de vacas da raça Nelore (Bos taurus indicus) foram obtidos em frigoríficos. Imediatamente após o abate das fêmeas, o útero era aberto e os embriões e fetos coletados. As gônadas e os embriões menores foram fixados em Bouin e processados para microscópica óptica convencional. Foi realizada a PCR (Reação em Cadeia da Polimerase) dos embriões cujo sexo não pode ser identificado macroscopicamente. A crista gonádica foi observada, primeiramente, em um embrião de 1,0 cm de comprimento. Em embriões com 2,5 cm de comprimento, a presença da albugínea permitiu a identificação do sexo. A espessura média da albugínea variou de 29,1 a 558,5 mm. O mediastino encontra-se localizado centralmente. Houve uma diminuição no espaço ocupado pelos cordões testiculares de 63,7 para 42,0% do volume total dos testículos. O seu diâmetro variou de 31,7 a 48,8 mm. O diâmetro das células germinativas (e dos seus núcleos) foi de 12,3 (6,6) a 16,9 (14,2) mm. A quantidade de células germinativas, por corte transversal de cordão, diminuiu de um máximo de 2,8 para 0,8. O número total de células germinativas aumentou de 16, no princípio da colonização da gônada, para 18,3 x 106 no final do estudo. O número de células de Sertoli, por corte transversal de cordão, variou de 10,0 a 16,2. Os resultados obtidos mostram que a origem e a formação dos testículos nos embriões e fetos de vacas da raça Nelore (Bos taurus indicus), ocorrem de forma muito semelhante à descrita para Bos taurus taurus. / The aim of this study was to accompany the process of testicular development from the indifferentiable phase to its complete formation. Embryos and fetuses of Nelore breed cows (Bos taurus indicus) were obtained in slaughterhouses near the Uberlandia city, Minas Gerais. As soon as the females were slaughtered, the uterus was opened and the embryos and fetuses gathered. The gonads and the smaller embryos were fixed in Bouin's fixative and afterwards processed for conventional optical microscopy. It was carried out the PCR (Polymerase Chain Reaction) from parts of embryos whose sex could not be identified macroscopically. The gonadal ridge was observed firstly in a 1.0 cm long embryo. In 2.5 cm long embryos the presence of the albuginea allowed the sex identification. The mean thickness of the albuginea ranged from 29.1 to 558.5 mm. Gradually increase of vascularization of the albuginea and parenchyma was observed. The mediastinum was located centrally. There was a decrease in the space occupied by the testicular cords, from 63.7 to 42.0 % of the total testes volume. Its diameter ranged from 31.7 to 48.8 mm. The diameter of germinal cells (and their nuclei) ranged from 12.3 (6.6) to 16.9 914.2) mm. The quantity of germinal cells by cross section of cord decreased from a maximum of 2.8 to 0.86. The total number of germinal cells was from 16 at the beginning of colonization of the gonad to 18.3 x 106 at the end of the study. The number of Sertoli's cells by cross section of cord ranged from 10.0 to 16.2. The results showed that the origin and formation of testes in embryos and fetuses from Nelore breed cows (Bos taurus indicus) does occur in a very similar way to what is described for Bos taurus taurus.

Indiana University-Purdue University Indianapolis (IUPUI) / Follicular T helper (TFH) and regulatory (TFR) cells are two key classes of CD4+ T cells found in germinal centers (GCs). The primary role of TFH cells is to help B cells form GCs to produce high-affinity antibodies during an infection while the role of TFR cells remains controversial. The transcriptional repressor Bcl6 is essential for the differentiation of TFH, TFR and GCB cells and understanding signaling pathways that induce Bcl6 and TFH cell differentiation are important. We observed that Bcl6 is highly up-regulated in activated CD4 T cells following glucose deprivation by a pathway involving the metabolic sensor AMP kinase. The transcription factor Blimp1 represses both TFH cell differentiation and Bcl6 expression, and we show the major role of Blimp1 on TFH cell differentiation is to repress Bcl6 expression and not other genes in the TFH differentiation pathway. We also found Bcl6 positively regulates expression of the key TFH cell receptor PD-1 by inhibiting the repression of PD-1 by the transcription factor Tbet. The roles of TFH and TFR cells in controlling allergen-specific IgE were investigated using a peanut allergy model and strains of mice with alterations in the TFH and TFR pathways. We found TFR cells unexpectedly play an essential role in promoting and maintaining IgE production and anaphylaxis, as well as the GC reaction. Compared to control mice, TFR-deficient mice lacked circulating peanut-specific IgE and anaphylactic responses were significantly weakened. Mechanistically, TFR cells require Blimp1 controlled IL-10 to promote GCB cell survival and IgE production. Blocking IL-10 signals mimicked the loss of IgE levels in TFR-deficient mice and rescued mice from anaphylaxis. Overall, these studies have defined novel roles of Bcl6, TFH and TFR cells in regulating antibody production by the GC reaction, and provide greater understanding of how allergic immune responses are controlled. / 2019-11-21

Orientador: Irani Quagio-Grassiotto / Dissertação (mestrado) - Universidade Estadual de Campinas, Instituto de Biologia / Made available in DSpace on 2018-08-14T18:28:03Z (GMT). No. of bitstreams: 1 Mazzoni_TalitaSarah_M.pdf: 22131608 bytes, checksum: df7814a1f55f5088400f2b70e4520216 (MD5) Previous issue date: 2009 / Resumo: Numa nova visão da morfogênese gonadal, sua descrição em Cyprinus carpio, mostra como a proliferação e diferenciação de células germinativas e somáticas a partir do primórdio gonadal levam à formação das diferentes estruturas ovarianas e testiculares e à constituição do epitélio germinativo que margeia as lamelas ovígeras e os túbulos testiculares. Em C. carpio, o primórdio gonadal é formado por células germinativas primordiais (CGPs) rodeadas por células somáticas. Após sucessivas divisões mitóticas das células somáticas, o tecido gonadal aumenta em comprimento e espessura. As CGPs isoladas entre células somáticas se dividem mitoticamente formando grupos de células germinativas, que se organizam em cordões contínuos, os quais são invadidos por células somáticas, levando à uma reorganização estrutural e diferenciação gonadal. Nas gônadas femininas, as oogônias são envolvidas por expansões citoplasmáticas das agora células pré-foliculares, formando cistos, delimitados por uma membrana basal em formação. Cada oogônia divide-se por mitose, formando novas oogônias ou entra em meiose originando os oócitos. Com a entrada e permanência em diplóteno, os oócitos, ainda no interior dos cistos, iniciam seu desenvolvimento que completar-se-á no interior dos folículos ovarianos. As células pré-foliculares progressivamente interpenetram nos cistos e envolvem cada oócito individualizando-os. Estas gradativamente sintetizam a membrana basal envolvendo progressivamente o folículo em formação. As células foliculares assentadas em parte na membrana basal do próprio epitélio germinativo mantêm uma região de contato entre o folículo ovariano e o epitélio germinativo, que compartilham uma mesma membrana basal. Ao término da foliculogênese, o oócito inicia seu crescimento primário. No tecido gonadal, células mesenquimais se interconectam e se diferenciam, dando origem ao estroma ovariano, isolado do compartimento germinativo pela membrana basal. Células indiferenciadas do estroma emitem prolongamentos que contatam os folículos em formação, constituindo a teca. Células somáticas periféricas por migração e invaginação no tecido gonadal, formam as lamelas ovígeras. Lâminas teciduais de células somáticas formam-se em ambos os lados do ovário, projetando-se até se contatarem, formando o lúmen ovariano. Nas gônadas masculinas, as células somáticas, pré- Sertoli, invadem os cordões contínuos de CGPs. Estas, agora espermatogônias, são envolvidas por expansões citoplasmáticas das células de Sertoli, formando cistos. Estes formam conjuntos celulares que se distribuem ao longo da gônada, cada qual circundado por células somáticas. Ao redor de cada conjunto celular, inicia-se a formação da membrana basal, porém de forma incompleta. Gradativamente, os cistos de espermatogônias que constituem um mesmo conjunto, afastam-se uns dos outros, criando um espaço central. Células somáticas de conjuntos celulares adjacentes afastam-se, permitindo fusão entre os dois conjuntos celulares e aumento do espaço central formando um único compartimento luminal, delimitado por cistos de espermatogônias, estes, apoiados na membrana basal. Formam-se os túbulos testiculares e o epitélio germinativo masculino é estabelecido. O compartimento germinativo encontra-se agora separado pela membrana basal dos demais componentes celulares, que irão se diferenciar no compartimento intersticial. Inicia-se a espermatogênese no interior de cada cisto, de forma sincrônica. Após espermiogênese, os espermatozóides são liberados no lúmen, e os túbulos testiculares se anastomosam, culminando com a formação do ducto espermático na porção dorsal do testículo / Abstract: The description of gonadal morphogenesis in Cyprinus carpio provides a new vision of proliferation and differentiation of germ and somatic cells from the gonadal primordium leading to the formation of different ovarian and testicular structures, and the constitution of germinal epithelium which borders the ovigerous lamellae and the seminiferous tubules. In C. carpio the gonadal primordium is an elongated structure with individual PGCs scattered among somatic cells. The PGCs divide and organize into continuous cords that are delimited by the somatic cells. Then, in female gonad, somatic cells move into the cords, wrap around and individualize the PGCs that subsequently differentiate in oogonia. Each oogonium is wrapped by the now prefollicle cells giving rise to a cyst. Prefollicle cells rest upon a forming basement membrane. Inside the cysts oogonium proliferates by mitosis, originating new oogonia. Or, they enter into meiosis, becoming oocytes. Oocytes advance to diplotene where meiosis is arrested. Still inside the cysts oocytes enter primary growth, they are subsequently surrounded by prefollicle cells, and they become ovarian follicles. The differentiating gonad maintains a compact structure, continues elongating and becomes larger. Invaginations appear in the ventral region of the ovary; these form the ovigerous lamellae. Mesenchymal cells scattered inside the lamellae move away from one another giving rise to the extra-vascular space in the developing stroma where the cellular processes of these cells connect to one another and form a cellular net. Meanwhile, epithelial cells coming from the gonad periphery, and present in the ventral invaginations, associate with oogonia forming germinal epithelium. These also interact with the follicles that become connected to the epithelium sharing some extension of the basement membrane. Mesenchymal cells surround the ovarian follicles, becoming theca and, include the follicle, forms the follicle complex. On either side of the developing ovary, a coelomic epithelial cell proliferation forms a laminar tissue that grows ventrally, then extending beneath the developing ovary and fusing to form the central lumen of the carp cystovarian ovary. In male gonad, somatic cells move into the cords, wrap around and individualize the PGCs that subsequently differentiate in spermagonia. Each spermatogonium wrapped by the now pre-Sertoli cells giving rise to a cyst. The cysts join one another forming clusters. A basement membrane is synthesized around each cluster. Pre-Sertoli cells rest upon the forming basement membrane. In the center of the clusters a space is created when pre-Sertoli cells move away from one another. Then, nearby clusters fuse to one another that become connected by the same luminal space. The progressive fusion of the clusters gives rise to the seminiferous tubules that are bordered by the new formed germinal epithelium constituted by the cysts that rest upon the basement membrane. Mesenchymal cells surround the seminiferous tubules give rise to the cellular components of the interstitial compartment. Inside the cysts spermatogenesis starts. As the final spermatic cells are released into the luminal compartment, the anostomosis of the testicular tubules occurs forming the spermatic duct on the dorsal region of the testis / Mestrado / Biologia Celular / Mestre em Biologia Celular e Estrutural