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RNA isolation from Peyer's patch lymphocytes and mononuclear phagocytes to determine gene expression profiles using NanoString technology.

ABSTRACT: Sampling and immune surveillance within gut-associated lymphoid tissues such as the intestinal Peyer's patch (PP) occurs by an elegantly orchestrated effort that involves the epithelial barrier, B and T lymphocytes, and an extensive network of mononuclear phagocytes. Although we now understand more about the dynamics of antigen and microbial sampling within PPs, the gene expression changes that occur in individual cell subsets during sampling are not well characterized. This protocol describes the isolation of high-quality RNA from sorted PP, B and T-lymphocytes, and CD11c+ phagocytes for use with nCounter-NanoString technology. This method allows investigators to study gene expression changes within PPs in response to antigens, microbes, and oral vaccine delivery vehicles of interest that are sampled.

SUBMITTER: Singh N

PROVIDER: S-EPMC6706143 | biostudies-literature | 2018

REPOSITORIES: biostudies-literature

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RNA isolation from Peyer's patch lymphocytes and mononuclear phagocytes to determine gene expression profiles using NanoString technology.

Singh Navjot N Gallagher Heather C HC Song Renjie R Dhinsa Jaskiran K JK Ostroff Gary R GR De Jesus Magdia M

Journal of biological methods 20180702 3

Sampling and immune surveillance within gut-associated lymphoid tissues such as the intestinal Peyer's patch (PP) occurs by an elegantly orchestrated effort that involves the epithelial barrier, B and T lymphocytes, and an extensive network of mononuclear phagocytes. Although we now understand more about the dynamics of antigen and microbial sampling within PPs, the gene expression changes that occur in individual cell subsets during sampling are not well characterized. This protocol describes t ...[more]

PMID: 31453245

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Project description:The gut represents a potential entry site for a wide range of pathogens including protozoa, bacteria, viruses, or fungi. Consequently, it is protected by one of the largest and most diversified population of immune cells of the body. Its surveillance requires the constant sampling of its encounters by dedicated sentinels composed of follicles and their associated epithelium located in specialized area. In the small intestine, Peyer's patches (PPs) are the most important of these mucosal immune response inductive sites. Through several mechanisms including transcytosis by specialized epithelial cells called M-cells, access to the gut lumen is facilitated in PPs. Although antigen sampling is critical to the initiation of the mucosal immune response, pathogens have evolved strategies to take advantage of this permissive gateway to enter the host and disseminate. It is, therefore, critical to decipher the mechanisms that underlie both host defense and pathogen subversive strategies in order to develop new mucosal-based therapeutic approaches. Whereas penetration of pathogens through M cells has been well described, their fate once they have reached the subepithelial dome (SED) remains less well understood. Nevertheless, it is clear that the mononuclear phagocyte system (MPS) plays a critical role in handling these pathogens. MPS members, including both dendritic cells and macrophages, are indeed strongly enriched in the SED, interact with M cells, and are necessary for antigen presentation to immune effector cells. This review focuses on recent advances, which have allowed distinguishing the different PP mononuclear phagocyte subsets. It gives an overview of their diversity, specificity, location, and functions. Interaction of PP phagocytes with the microbiota and the follicle-associated epithelium as well as PP infection studies are described in the light of these new criteria of PP phagocyte identification. Finally, known alterations affecting the different phagocyte subsets during PP stimulation or infection are discussed.

Dataset Information

RNA isolation from Peyer's patch lymphocytes and mononuclear phagocytes to determine gene expression profiles using NanoString technology.

Publications

RNA isolation from Peyer's patch lymphocytes and mononuclear phagocytes to determine gene expression profiles using NanoString technology.

Similar Datasets

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