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13:36 min
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October 20, 2023
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The scope of the glycoimmunology group research is studying the role of glycan immunology in human health and disease. This research aims to disentangle biological mechanisms modulated by glycosylations, and to discover novel glycan-based therapies for cancer and congenital disorders of glycosylation. Our focus has been on sialylated glycans that affect the function of relevant molecules, particularly during response.
We developed technologies to alter sialic acid contents and a multimodal platform for high-throughput dissolvement of glycan binding proteins with potential clinical applications. To advance the research in the field, we use several technologies such as glycan profiling, selecting or antibody staining, mass spectrometry, and immunohistochemistry, in vitro assays with cell lines as disease models, sialic acid content manipulations through enzymatic and metabolic engineering. The current experimental challenges relate to understanding the complex interactions between sialylated glycans and lectins.
There are mechanisms and functional consequences in several immune responses. The sialic acid modulates the potency of dendritic cells and MHC1 turnover. This discovery pinpointed new biological and pathological mechanisms and developed innovative therapies.
It also identified novel immune pathways and approaches for their therapeutic modulation. One of the issues in ex vivo production of human monocyte derived dendritic cells for therapeutic uses is their inability to mature fully. This protocol shows that DCs maturation can be achieved by enzymatically treating the cell surface with sialidases while maintaining cell viability.
This method is cost effective and time saving. Compared to sialic acid metabolic inhibitors, sialidase treatment offers a rapid effective and method of removing cell surface sialic acids while maintaining cell viability. Understanding the role of sialic acid content in glycans generates some precedent awareness of the importance of this glycan in cancer and will foster the identification of new disease mechanisms and neuropath strategies.
A unique, comprehensive protocol to generate de-sialylated human monocyte-derived dendritic cells (mo-DCs) from isolated peripheral blood mononuclear cells (PBMCs) using a sialidase treatment is presented. Further, methods to assess the phenotypic and functional characterization of mo-DCs and evaluate how sialidase treatment improves the maturation level of mo-DCs are described.
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Cite this Article
Luz, V. C. C., Silva, Z., Sobral, P., Tanwar, A., Paterson, R. L., Videira, P. A. Generation of Monocyte-Derived Dendritic Cells with Differing Sialylated Phenotypes. J. Vis. Exp. (200), e65525, doi:10.3791/65525 (2023).
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