Abstract
Natural killer (NK) cells are important for immune protection of the gut mucosa. Previous studies have shown that under pathologic conditions NK cells, T cells and dendritic cells are found co-localised in secondary lymphoid organs where their interaction coordinates immune responses. However, in the gut-associated lymphoid tissues (GALTs), there are few detailed reports on the distribution of NK cells. Sheep harbour several types of organised lymphoid tissues in the gut that have different functions. The ileal Peyer’s patch (IPP) functions as a primary lymphoid tissue for B cell generation, while the jejunal Peyer’s patches (JPPs) and colon patches (CPs) are considered secondary lymphoid tissues. In the present study, we analysed tissues from healthy lambs by flow cytometry and in situ multicolour immunofluorescence, using recently described NCR1 antibodies to identify ovine NK cells. Most NCR1+ cells isolated from all tissues were negative for the pan T cell marker CD3, and thus comply with the general definition of NK cells. The majority of NCR1+ cells in blood as well as secondary lymphoid organs expressed CD16, but in the GALT around half of the NCR1+ cells were negative for CD16. A semi-quantitative morphometric study on tissue sections was used to compare the density of NK cells in four compartments of the IPPs, JPP and CPs. NCR1+ cells were found in all gut segments. Statistical analysis revealed significant differences between compartments of the primary lymphoid organ IPP and the secondary lymphoid organs of the JPPs and CP. NK cells co-localised and made close contact with T cells, dendritic cells and other NK cells, but did not show signs of proliferation. We conclude that NK cells are present in all investigated segments of the sheep gut, but that presence of other innate lymphoid cells expressing NCR1 cannot be excluded.
Highlights
Natural killer (NK) cells are lymphocytes of the innate immune system traditionally known for their immediate cytotoxic activity against stressed, transformed or infected cells [1]
Quantification and characterization of NCR1+ cells by flow cytometry The quantity of NCR1+ cells was given as the number of positive cells in relation to the total number of mononuclear cells (MNC) gated in a FSC/SSC scatter plot
To assess whether NCR1 could be present on a subclass of T cells, we labelled cells isolated from various tissues collected from 5 older abattoir lambs using an antibody against the Cytoplasmic region of CD3 (cytCD3)
Summary
Natural killer (NK) cells are lymphocytes of the innate immune system traditionally known for their immediate cytotoxic activity against stressed, transformed or infected cells [1]. They have been shown to be present in lymphoid tissues, mucosal tissues and several other organ systems, where they exhibit direct effector functions as well as immunoregulatory actions on other cells through cytokine production [2,3,4,5]. A further compartment is the lamina propria, which is found along the whole length of the gut, and is present both within and beyond the borders of PPs. T- and B cells predominate in the IFA and follicle, respectively, of the sheep PPs [19,20,21]. In the lamb and sheep gut, DCs are mostly found in the dome, IFA and lamina propria and have been shown to express CD11c, CD205, and MHCII [22]
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