Thesis (Ph. D.)--Case Western Reserve University, 2008. / [School of Medicine] Department of Pathology. Includes bibliographical references.

Thesis (M.S.)--University of Texas at El Paso, 2009. / Title from title screen. Vita. CD-ROM. Includes bibliographical references. Also available online.

Thesis (Ph. D.)--University of Hong Kong, 2005. / Title proper from title frame. Also available in printed format.

Thesis (Ph.D.) - University of Queensland, 2005. / Includes bibliography.

Inflammatory myeloid dendritic cells (DCs) are critical in the pathogenesis and maintenance of psoriasis vulgaris, a chronic inflammatory skin disease of unknown etiology. New ways to define these cells, and their precursors, may allow us to better understand their role in inflammation. Immunohistochemistry was performed on frozen tissue sections of normal and psoriasis biopsies to examine the dermal expression of potential markers of inflammatory DCs, namely CLEC9A, CD103, SlanDC, and TREM-1. The allostimulatory capacity of DC subsets (of SlanDC+ and CD1c+) was compared in a mixed leukocyte assay (MLR). Potential precursors of inflammatory DCs were FACS-sorted for transcriptomic profiling and functional assays. CLEC9A, CD103, and SlanDC did not prove useful in uniquely identifying myeloid dendritic cells in normal skin, and inflammatory dendritic cells in inflammation. TREM-1 was highly upregulated in psoriasis lesional skin as compared to non-lesional, and its activation may be critical in the maintenance of inflammation. Contrary to published findings, CD1c+ DCs possessed a higher allo-stimulatory capacity than SlanDCs, and induced greater IL-17 in T cells. TREM-1 may provide a novel therapeutic target for psoriasis treatment. The six circulating monocyte and dendritic cell populations in human peripheral blood were obtained via FACS sorting, and their genomic profiles will be examined. By comparing the genomic profiles of the six circulating monocyte and dendritic cell populations in human blood, and examining their allo- and autostimulatory capacities in a peptidoglycan (PGN) stimulated in vitro model of inflammation, the source of these inflammatory dendritic cells can be identified, and provide future targets of therapy for this psoriasis.

No description available.

Ng, Sin Man. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2011. / Includes bibliographical references (leaves 96-104). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstract also in Chinese.

Follicular dendritic cells (FDCs) are key elements of secondary lymphoid organs where they form the stromal component of B-cell follicles. FDCs possess extensive dendritic process that trap intact antigen via Fc and complement receptors on the cell surface. The antigen is displayed to B-cells, providing a basis for selection of high affinity B cells. FDC also have important roles in facilitating the clearance of apoptotic B cells by the secretion of the opsonising factor MFGE8. It is well established that lymphotoxin signalling is required for FDC maturation but the specific details of the molecular mechanisms that regulate FDC development and differentiation are not fully understood. MicroRNAs (miRNAs) are non-coding RNAs of approximately 18-25 nucleotides in length that regulate gene expression at the post-transcriptional level. MiRNAs bind to their target gene transcripts as part of the RNA induced silencing complex and repress translation of the target gene product. The objective of this study was to identify miRNAs that play a role in the development and function of FDCs. An in vivo murine model of FDC de-differentiation was used to provide material for miRNA analysis. By comparison of miRNA profiles from spleen tissue with FDC at different stages of de-differentiation, we would be able to obtain a miRNA signature for mature FDC. Spleens were collected at various time points over a 28 day period following transient blockade of lymphotoxin signalling. A variety of methods were used to profile the miRNAs expressed at different time points during the suppression and recovery of the FDC network. Comparison of the miRNA profiles of spleens containing mature, partially de-differentiated, and fully de-differentiated FDC identified a number of miRNAs that were differentially expressed during FDC de-differentiation. To assess the role of specific miRNAs in FDC development, the mouse FDC-like cell line, FL-YB, was used as an in vitro model system. FL-YB cells were used to perform gain-offunction and loss-of-function studies on selected miRNAs and to assess the effects of various stimuli/conditions on miRNA expression. The effects of different treatments on cell proliferation, morphology and adhesion, and on gene expression by FL-YB, were monitored. Loss-of-function studies for one of the selected miRNA (miR-100-5p) revealed a significant effect on a number of gene transcripts involved in mediation of the germinal centre response (Il-6, Tlr4, Ptgs1/2). These data indicate that miR-100-5p has a role in regulating Il-6, Tlr4 and Ptgs1/2 transcripts. None of these transcripts contain predicted target sites for miR-100-5p and so the effect of miR-100-5p on these transcripts is likely to be indirect. Further studies on these miRNA: target interactions are required to elucidate the mechanisms and biological consequences of miRNA regulation in FDCs.

Dendritic cells (DCs) are leukocytes specialized in antigen presentation. As competent stimulators of naive T lymphocytes, they link the innate and adaptive immune responses of vertebrates. The RAG-mediated adaptive immune system appeared approximately 500 million years ago in jawed fish and a number of studies suggest that DCs exist in bony and cartilaginous fish. However, the exact role of DCs in the fish immune system is not determined and questions remain as to whether a cell type truly homologous to DCs in homeotherms does exist. My project aimed to identify potential DCs surface markers (CD209A and LAMP3) in rainbow trout (Oncorhynchus mykiss) leukocytes for evaluation of the expression patterns by qRT-PCR under different conditions and stimuli, in vitro and in vivo. Another goal was to validate and evaluate the specificity of a produced anti-trout CD209A polyclonal antibody to further characterise antigen presenting cells (APCs) in fish. The methodology was to look for up-regulation of the predicted markers together with other markers known to be expressed by DCs in mammals and to evaluate at the mRNA and protein expression level after in vitro stimulation of trout primary leukocytes with trout rIL-4/13. Trout CD209A and LAMP3 mRNA was expressed in the main lymphoid organs of fish and could be modulated with microbial mimics. Upon in vitro stimulation of trout primary leukocytes with trout rIL-4/13, trout CD209A mRNA expression was up-regulated together with both CD83 and the MHC class II chain known to be expressed by mammalian DCs. In addition, CD209A protein expression was highly induced by trout rIL-4/13. Taken together, these results suggests that the characterisation of DCs in trout with tools such as transcript evaluation of surface markers and the anti-trout CD209A antibody, could help to more precisely define these leukocyte subsets. These findings could have further impact on fish vaccine improvements and be of importance for the aquaculture industry, by optimising stimulation of adaptive immunity.

DCs are critical to the early events of HIV-1 infection. They are the first cells that HIV-1 encounters at mucosal surfaces and as sentinel antigen-presenting cells of the immune system these should alarm the immune system and activate innate immune defences to recruit effective adaptive immunity and viral clearance. A peculiar characteristic of HIV – in contrast to other ssRNA viruses – is its ability to completely evade host innate recognition pathways. Additionally, it has the unique ability to manipulate the endo-lysosomal system of DCs and promote transmission via trans-infection to CD4+ T cells across virological synapses. However, it is largely unknown how HIV-1 is sensed by the innate immune system. Here, a multipronged experimental approach based on phosphoproteomics, transcriptomics and custom RNAi screen was developed to characterize the early signaling complex induced by HIV-1 in DCs. A novel method of phosphoproteomics to identify the HIV-1 phosphoproteome in DCs showed that 342 proteins were differentially phosphorylated following 10 min of HIV-1 infection compared to time-matched mock-infected DCs. Functional analysis of these phosphoproteins showed enrichments in several cellular pathways, including vesicular trafficking, cytoskeletal rearrangements and the secretory pathway and a relative paucity of signaling molecules involved in inflammatory pathways. Proteomics analysis of HIV-1 virions was undertaken to identify host molecules hijacked by HIV-1 during viral replication and revealed a close interaction between the virus and the endo-lysosomal system. Transcriptomics analysis of HIV-1 infected DCs showed a muted immune response with no detectable differentially regulated genes. The results of the phoshoproteomic screen provided the basis for a custom RNAi screen to identify host proteins that are differentially phosphorylated by the virus and required for efficient trans-infection from DCs to CD4+ lymphocytes. The results of this screen showed that 54 of the 120 host factors tested were required for efficient viral transfer to CD4+ T cells and characterize the compartment that HIV-1 is internalized in on a molecular level. Two host factors identified within the HIV-1 phosphoproteome were chosen for further studies. Studies of BLOC-1 (biogenesis of lysosome-related organelles complex-1) and its subunits identified a role for snapin in HIV-1 trans-infection and HIV-1 and TLR8 sensing. Snapin may act as determinant of sorting of HIV-1 intraluminal vesicles to non-degradative, non-immunogenic compartments by activating mammalian target of rapamycin, mTOR, and inhibiting autophagy. Furthermore, HIV-1 triggered dephosphorylation of the cytosolic tyrosine phosphatase possibly via the interaction of host CD47 incorporated in the virion and the transmembrane glycoprotein SIRPα expressed on DCs. Blocking of this interaction with an inhibitory CD47 antibody resulted in a reduction of HIV-1 replication. Taken together, this multipronged approach reveals the complexity of the interaction of HIV-1 with the host cell machinery and identifies novel mechanism of the immune evasion tactics usurped by HIV-1.