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Phenotypic and functional analysis of monocyte populations in cattle peripheral blood identifies a subset with high endocytic and allogeneic T-cell stimulatory capacity.

Corripio-Miyar Y, Hope J, McInnes CJ, Wattegedera SR, Jensen K, Pang Y, Entrican G, Glass EJ - Vet. Res. (2015)

Bottom Line: The bovine subsets expressed similar levels of CD80, CD40 and CD11c molecules and mRNA encoding CD115.The more diffuse CD14(+)CD16(+) population generally expressed intermediate levels of these molecules.All three populations responded to stimulation with phenol-extracted lipopolysaccharide (LPS) by producing interleukin (IL)-1β, with the CD16(++) subset expressing higher levels of IL-12 and lower levels of IL-10.

View Article: PubMed Central - PubMed

Affiliation: Division of Infection & Immunity, The Roslin Institute and Royal (Dick) School of Veterinary Studies, University of Edinburgh, Easter Bush, Midlothian, EH25 9RG, UK. Yolanda.Corripio-Miyar@moredun.ac.uk.

ABSTRACT
Circulating monocytes in several mammalian species can be subdivided into functionally distinct subpopulations based on differential expression of surface molecules. We confirm that bovine monocytes express CD172a and MHC class II with two distinct populations of CD14(+)CD16(low/-)CD163(+) and CD14(-)CD16(++)CD163(low-) cells, and a more diffuse population of CD14(+)CD16(+)CD163(+) cells. In contrast, ovine monocytes consisted of only a major CD14(+)CD16(+) subset and a very low percentage of CD14(-)CD16(++)cells. The bovine subsets expressed similar levels of CD80, CD40 and CD11c molecules and mRNA encoding CD115. However, further mRNA analyses revealed that the CD14(-)CD16(++) monocytes were CX3CR1(high)CCR2(low) whereas the major CD14(+) subset was CX3CR1(low)CCR2(high). The former were positive for CD1b and had lower levels of CD11b and CD86 than the CD14(+) monocytes. The more diffuse CD14(+)CD16(+) population generally expressed intermediate levels of these molecules. All three populations responded to stimulation with phenol-extracted lipopolysaccharide (LPS) by producing interleukin (IL)-1β, with the CD16(++) subset expressing higher levels of IL-12 and lower levels of IL-10. The CD14(-)CD16(++) cells were more endocytic and induced greater allogeneic T cell responses compared to the other monocyte populations. Taken together the data show both similarities and differences between the classical, intermediate and non-classical definitions of monocytes as described for other mammalian species, with additional potential subpopulations. Further functional analyses of these monocyte populations may help explain inter-animal and inter-species variations to infection, inflammation and vaccination in ruminant livestock.

No MeSH data available.


Related in: MedlinePlus

Differential endocytosis of OVA and Dextran by subpopulations of myeloid cells. PBMC were isolated and incubated for 30 min with 100 μg/mL of TexasRed-OVA (A, B) or TexasRed-Dextran (C, D) at 37 °C or on ice (to assess background uptake). The cells were subsequently washed and stained with conjugated CD14 and CD16 antibodies and analysed by flow cytometry. Live, single cells were gated as in Figure 1 and the MFI was calculated for the CD14−CD16++ and CD14+ gated populations (A, C). The results shown (B, D) are mean values with error bars showing ± SE for four animals and represent the TexasRed MFI at 37 °C minus MFI on ice (4 °C).
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Fig8: Differential endocytosis of OVA and Dextran by subpopulations of myeloid cells. PBMC were isolated and incubated for 30 min with 100 μg/mL of TexasRed-OVA (A, B) or TexasRed-Dextran (C, D) at 37 °C or on ice (to assess background uptake). The cells were subsequently washed and stained with conjugated CD14 and CD16 antibodies and analysed by flow cytometry. Live, single cells were gated as in Figure 1 and the MFI was calculated for the CD14−CD16++ and CD14+ gated populations (A, C). The results shown (B, D) are mean values with error bars showing ± SE for four animals and represent the TexasRed MFI at 37 °C minus MFI on ice (4 °C).

Mentions: To assess the capacity for antigen uptake by the CD14+ and CD14−CD16++ cell populations, PBMC from four animals were incubated with either Dextran (DX), a complex glucan taken up via the mannose receptor (MR; CD206) by macropinocytosis [43] or Ovalbumin (OVA) a protein taken up by clathrin-coated pits [44], both fluorescently labelled with TexasRed (TR). The cell populations were then identified within PBMC as CD14+ or CD14−CD16++ after incubation with CD14 and CD16 mAbs. Low level internalisation was observed when cells were incubated on ice for both TR-OVA (Figure 8A) and TR-DX (Figure 8C); while significantly higher levels of uptake were observed after incubation at 37 °C. Comparing uptake between the cell populations revealed that the CD14−CD16++ cells internalised significantly higher levels of TR-OVA when compared to the CD14+ cells (p = 0.03, Figure 8B).Figure 8


Phenotypic and functional analysis of monocyte populations in cattle peripheral blood identifies a subset with high endocytic and allogeneic T-cell stimulatory capacity.

Corripio-Miyar Y, Hope J, McInnes CJ, Wattegedera SR, Jensen K, Pang Y, Entrican G, Glass EJ - Vet. Res. (2015)

Differential endocytosis of OVA and Dextran by subpopulations of myeloid cells. PBMC were isolated and incubated for 30 min with 100 μg/mL of TexasRed-OVA (A, B) or TexasRed-Dextran (C, D) at 37 °C or on ice (to assess background uptake). The cells were subsequently washed and stained with conjugated CD14 and CD16 antibodies and analysed by flow cytometry. Live, single cells were gated as in Figure 1 and the MFI was calculated for the CD14−CD16++ and CD14+ gated populations (A, C). The results shown (B, D) are mean values with error bars showing ± SE for four animals and represent the TexasRed MFI at 37 °C minus MFI on ice (4 °C).
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4582714&req=5

Fig8: Differential endocytosis of OVA and Dextran by subpopulations of myeloid cells. PBMC were isolated and incubated for 30 min with 100 μg/mL of TexasRed-OVA (A, B) or TexasRed-Dextran (C, D) at 37 °C or on ice (to assess background uptake). The cells were subsequently washed and stained with conjugated CD14 and CD16 antibodies and analysed by flow cytometry. Live, single cells were gated as in Figure 1 and the MFI was calculated for the CD14−CD16++ and CD14+ gated populations (A, C). The results shown (B, D) are mean values with error bars showing ± SE for four animals and represent the TexasRed MFI at 37 °C minus MFI on ice (4 °C).
Mentions: To assess the capacity for antigen uptake by the CD14+ and CD14−CD16++ cell populations, PBMC from four animals were incubated with either Dextran (DX), a complex glucan taken up via the mannose receptor (MR; CD206) by macropinocytosis [43] or Ovalbumin (OVA) a protein taken up by clathrin-coated pits [44], both fluorescently labelled with TexasRed (TR). The cell populations were then identified within PBMC as CD14+ or CD14−CD16++ after incubation with CD14 and CD16 mAbs. Low level internalisation was observed when cells were incubated on ice for both TR-OVA (Figure 8A) and TR-DX (Figure 8C); while significantly higher levels of uptake were observed after incubation at 37 °C. Comparing uptake between the cell populations revealed that the CD14−CD16++ cells internalised significantly higher levels of TR-OVA when compared to the CD14+ cells (p = 0.03, Figure 8B).Figure 8

Bottom Line: The bovine subsets expressed similar levels of CD80, CD40 and CD11c molecules and mRNA encoding CD115.The more diffuse CD14(+)CD16(+) population generally expressed intermediate levels of these molecules.All three populations responded to stimulation with phenol-extracted lipopolysaccharide (LPS) by producing interleukin (IL)-1β, with the CD16(++) subset expressing higher levels of IL-12 and lower levels of IL-10.

View Article: PubMed Central - PubMed

Affiliation: Division of Infection & Immunity, The Roslin Institute and Royal (Dick) School of Veterinary Studies, University of Edinburgh, Easter Bush, Midlothian, EH25 9RG, UK. Yolanda.Corripio-Miyar@moredun.ac.uk.

ABSTRACT
Circulating monocytes in several mammalian species can be subdivided into functionally distinct subpopulations based on differential expression of surface molecules. We confirm that bovine monocytes express CD172a and MHC class II with two distinct populations of CD14(+)CD16(low/-)CD163(+) and CD14(-)CD16(++)CD163(low-) cells, and a more diffuse population of CD14(+)CD16(+)CD163(+) cells. In contrast, ovine monocytes consisted of only a major CD14(+)CD16(+) subset and a very low percentage of CD14(-)CD16(++)cells. The bovine subsets expressed similar levels of CD80, CD40 and CD11c molecules and mRNA encoding CD115. However, further mRNA analyses revealed that the CD14(-)CD16(++) monocytes were CX3CR1(high)CCR2(low) whereas the major CD14(+) subset was CX3CR1(low)CCR2(high). The former were positive for CD1b and had lower levels of CD11b and CD86 than the CD14(+) monocytes. The more diffuse CD14(+)CD16(+) population generally expressed intermediate levels of these molecules. All three populations responded to stimulation with phenol-extracted lipopolysaccharide (LPS) by producing interleukin (IL)-1β, with the CD16(++) subset expressing higher levels of IL-12 and lower levels of IL-10. The CD14(-)CD16(++) cells were more endocytic and induced greater allogeneic T cell responses compared to the other monocyte populations. Taken together the data show both similarities and differences between the classical, intermediate and non-classical definitions of monocytes as described for other mammalian species, with additional potential subpopulations. Further functional analyses of these monocyte populations may help explain inter-animal and inter-species variations to infection, inflammation and vaccination in ruminant livestock.

No MeSH data available.


Related in: MedlinePlus