Limits...
Fractalkine preferentially mediates arrest and migration of CD16+ monocytes.

Ancuta P, Rao R, Moses A, Mehle A, Shaw SK, Luscinskas FW, Gabuzda D - J. Exp. Med. (2003)

Bottom Line: CD16+ monocytes produce high levels of proinflammatory cytokines and may represent dendritic cell precursors in vivo.In contrast to CD16- monocytes, CD16+ monocytes expressed high CX3CR1 and CXCR4 but low CCR2 and CD62L levels and underwent efficient transendothelial migration in response to fractalkine (FKN; FKN/CX3CL1) and stromal-derived factor 1 alpha (CXCL12) but not monocyte chemoattractant protein 1 (CCL2).These results demonstrate that FKN preferentially mediates arrest and migration of CD16+ monocytes and suggest that recruitment of this proinflammatory monocyte subset to vessel walls via the CX3CR1-FKN pathway may contribute to vascular and tissue injury during pathological conditions.

View Article: PubMed Central - PubMed

Affiliation: Department of Cancer Immunology and AIDS, Dana-Farber Cancer Institute, Harvard Medical School, Boston, MA 02115, USA.

ABSTRACT
CD16+ monocytes represent 5-10% of peripheral blood monocytes in normal individuals and are dramatically expanded in several pathological conditions including sepsis, human immunodeficiency virus 1 infection, and cancer. CD16+ monocytes produce high levels of proinflammatory cytokines and may represent dendritic cell precursors in vivo. The mechanisms that mediate the recruitment of CD16+ monocytes into tissues remain unknown. Here we investigate molecular mechanisms of CD16+ monocyte trafficking and show that migration of CD16+ and CD16- monocytes is mediated by distinct combinations of adhesion molecules and chemokine receptors. In contrast to CD16- monocytes, CD16+ monocytes expressed high CX3CR1 and CXCR4 but low CCR2 and CD62L levels and underwent efficient transendothelial migration in response to fractalkine (FKN; FKN/CX3CL1) and stromal-derived factor 1 alpha (CXCL12) but not monocyte chemoattractant protein 1 (CCL2). CD16+ monocytes arrested on cell surface-expressed FKN under flow with higher frequency compared with CD16- monocytes. These results demonstrate that FKN preferentially mediates arrest and migration of CD16+ monocytes and suggest that recruitment of this proinflammatory monocyte subset to vessel walls via the CX3CR1-FKN pathway may contribute to vascular and tissue injury during pathological conditions.

Show MeSH

Related in: MedlinePlus

Phenotypic analysis of human monocyte subsets. (A) PBMCs were stained with FITC anti-CD14 and PC5 anti-CD16 mAbs. Monocytes were gated according to size, granularity, and CD14 expression. Three subsets of monocytes were identified: CD14high CD16−, CD14high CD16+, and CD14low CD16+. Results are representative of experiments performed with cells from 20 different donors. (B) PBMCs were stained with FITC anti-CD14, PE anti-CD56, and PC5 anti-CD16 mAbs, and CD14+ monocytes were analyzed for CD16 and CD56 expression. Results are representative of four experiments performed with cells from different donors. (C and D) PBMCs were stained with FITC anti-CD14, PC5 anti-CD16, and the indicated mAbs, and the phenotype of each monocyte subset was analyzed by flow cytometry. Values represent the percentage of positive cells (mean ± SD, n = 9). *, P < 0.05, Student's t test (CD16+ vs. CD16− monocytes).
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2193954&req=5

fig1: Phenotypic analysis of human monocyte subsets. (A) PBMCs were stained with FITC anti-CD14 and PC5 anti-CD16 mAbs. Monocytes were gated according to size, granularity, and CD14 expression. Three subsets of monocytes were identified: CD14high CD16−, CD14high CD16+, and CD14low CD16+. Results are representative of experiments performed with cells from 20 different donors. (B) PBMCs were stained with FITC anti-CD14, PE anti-CD56, and PC5 anti-CD16 mAbs, and CD14+ monocytes were analyzed for CD16 and CD56 expression. Results are representative of four experiments performed with cells from different donors. (C and D) PBMCs were stained with FITC anti-CD14, PC5 anti-CD16, and the indicated mAbs, and the phenotype of each monocyte subset was analyzed by flow cytometry. Values represent the percentage of positive cells (mean ± SD, n = 9). *, P < 0.05, Student's t test (CD16+ vs. CD16− monocytes).

Mentions: The expression of a large panel of chemokine receptors and adhesion molecules was assessed on monocyte subsets. Staining with anti-CD14 and anti-CD16 mAbs distinguished three monocyte subsets: CD14high CD16−, CD14high CD16+, and CD14low CD16+, representing 85.5 ± 6.2, 3.6 ± 1.5, and 6.7 ± 3.0% of total monocytes, respectively (mean ± SD, n = 8; Fig. 1 A), consistent with previous reports (1). The high expression of HLA-DR on CD14low CD16+ monocytes, together with the absence of neutrophil (CD16b, expressed on neutrophils but not monocytes nor NK cells; 19), NK (CD56), and T cell (CD3) markers (Fig. 1, B and D, and unpublished data), demonstrated the monocyte identity of these cells. The major CD14high CD16− monocyte subset expressed high CCR1, CCR2, CXCR2, CXCR4, PSGL-1, CD62L, CD18, CD11a, CD11b, CD11c, very late antigen 4 (VLA-4), ICAM-1, CD31, CD44, CD32, CD64, and HLA-DR, intermediate CXCR1, and low or undetectable CCR3, CCR5, CXCR5, and CX3CR1 levels (Fig. 1, C and D, and unpublished data). Compared with CD16− monocytes, CD14low CD16+ monocytes expressed significantly lower levels of CCR1, CCR2, CXCR1, and CXCR2, similar levels of CXCR4, and higher levels of CX3CR1 (Fig. 1 C). The expression of PSGL-1 was high in both monocyte subsets, whereas CD62L expression was high on CD16− monocytes but low or undetectable on CD14low CD16+ monocytes (Fig. 1 D). The adhesion molecules CD18, CD11a, CD11b, CD11c, VLA-4, ICAM-1, CD31, and CD44 were expressed on 95–100% of both CD16− and CD14low CD16+ monocytes (Fig. 1 D and unpublished data). The mean fluorescence intensity for CD18, CD11a, CD11c, VLA-4, and CD31 expression was higher on CD16+ monocytes as compared with CD16− monocytes (unpublished data). The phenotype of CD14high CD16+ monocytes was intermediate between that of CD14low CD16+ and CD16− monocytes (Fig. 1, C and D). Similar to CD14low CD16+ monocytes, a significant decrease in CCR2, CD62L, and CD64, and increase in CX3CR1 expression was observed on CD14high CD16+ compared with CD16− monocytes. CD14high CD16+ monocytes could be distinguished from the other two monocyte subsets by high CCR5 expression (Fig. 1 C). The pattern of CCR1, CCR2, CX3CR1, and CD62L expression on the three monocyte subsets was similar when staining was performed on whole blood and PBMCs, indicating that expression of these markers was not altered by Ficoll separation (unpublished data).


Fractalkine preferentially mediates arrest and migration of CD16+ monocytes.

Ancuta P, Rao R, Moses A, Mehle A, Shaw SK, Luscinskas FW, Gabuzda D - J. Exp. Med. (2003)

Phenotypic analysis of human monocyte subsets. (A) PBMCs were stained with FITC anti-CD14 and PC5 anti-CD16 mAbs. Monocytes were gated according to size, granularity, and CD14 expression. Three subsets of monocytes were identified: CD14high CD16−, CD14high CD16+, and CD14low CD16+. Results are representative of experiments performed with cells from 20 different donors. (B) PBMCs were stained with FITC anti-CD14, PE anti-CD56, and PC5 anti-CD16 mAbs, and CD14+ monocytes were analyzed for CD16 and CD56 expression. Results are representative of four experiments performed with cells from different donors. (C and D) PBMCs were stained with FITC anti-CD14, PC5 anti-CD16, and the indicated mAbs, and the phenotype of each monocyte subset was analyzed by flow cytometry. Values represent the percentage of positive cells (mean ± SD, n = 9). *, P < 0.05, Student's t test (CD16+ vs. CD16− monocytes).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2193954&req=5

fig1: Phenotypic analysis of human monocyte subsets. (A) PBMCs were stained with FITC anti-CD14 and PC5 anti-CD16 mAbs. Monocytes were gated according to size, granularity, and CD14 expression. Three subsets of monocytes were identified: CD14high CD16−, CD14high CD16+, and CD14low CD16+. Results are representative of experiments performed with cells from 20 different donors. (B) PBMCs were stained with FITC anti-CD14, PE anti-CD56, and PC5 anti-CD16 mAbs, and CD14+ monocytes were analyzed for CD16 and CD56 expression. Results are representative of four experiments performed with cells from different donors. (C and D) PBMCs were stained with FITC anti-CD14, PC5 anti-CD16, and the indicated mAbs, and the phenotype of each monocyte subset was analyzed by flow cytometry. Values represent the percentage of positive cells (mean ± SD, n = 9). *, P < 0.05, Student's t test (CD16+ vs. CD16− monocytes).
Mentions: The expression of a large panel of chemokine receptors and adhesion molecules was assessed on monocyte subsets. Staining with anti-CD14 and anti-CD16 mAbs distinguished three monocyte subsets: CD14high CD16−, CD14high CD16+, and CD14low CD16+, representing 85.5 ± 6.2, 3.6 ± 1.5, and 6.7 ± 3.0% of total monocytes, respectively (mean ± SD, n = 8; Fig. 1 A), consistent with previous reports (1). The high expression of HLA-DR on CD14low CD16+ monocytes, together with the absence of neutrophil (CD16b, expressed on neutrophils but not monocytes nor NK cells; 19), NK (CD56), and T cell (CD3) markers (Fig. 1, B and D, and unpublished data), demonstrated the monocyte identity of these cells. The major CD14high CD16− monocyte subset expressed high CCR1, CCR2, CXCR2, CXCR4, PSGL-1, CD62L, CD18, CD11a, CD11b, CD11c, very late antigen 4 (VLA-4), ICAM-1, CD31, CD44, CD32, CD64, and HLA-DR, intermediate CXCR1, and low or undetectable CCR3, CCR5, CXCR5, and CX3CR1 levels (Fig. 1, C and D, and unpublished data). Compared with CD16− monocytes, CD14low CD16+ monocytes expressed significantly lower levels of CCR1, CCR2, CXCR1, and CXCR2, similar levels of CXCR4, and higher levels of CX3CR1 (Fig. 1 C). The expression of PSGL-1 was high in both monocyte subsets, whereas CD62L expression was high on CD16− monocytes but low or undetectable on CD14low CD16+ monocytes (Fig. 1 D). The adhesion molecules CD18, CD11a, CD11b, CD11c, VLA-4, ICAM-1, CD31, and CD44 were expressed on 95–100% of both CD16− and CD14low CD16+ monocytes (Fig. 1 D and unpublished data). The mean fluorescence intensity for CD18, CD11a, CD11c, VLA-4, and CD31 expression was higher on CD16+ monocytes as compared with CD16− monocytes (unpublished data). The phenotype of CD14high CD16+ monocytes was intermediate between that of CD14low CD16+ and CD16− monocytes (Fig. 1, C and D). Similar to CD14low CD16+ monocytes, a significant decrease in CCR2, CD62L, and CD64, and increase in CX3CR1 expression was observed on CD14high CD16+ compared with CD16− monocytes. CD14high CD16+ monocytes could be distinguished from the other two monocyte subsets by high CCR5 expression (Fig. 1 C). The pattern of CCR1, CCR2, CX3CR1, and CD62L expression on the three monocyte subsets was similar when staining was performed on whole blood and PBMCs, indicating that expression of these markers was not altered by Ficoll separation (unpublished data).

Bottom Line: CD16+ monocytes produce high levels of proinflammatory cytokines and may represent dendritic cell precursors in vivo.In contrast to CD16- monocytes, CD16+ monocytes expressed high CX3CR1 and CXCR4 but low CCR2 and CD62L levels and underwent efficient transendothelial migration in response to fractalkine (FKN; FKN/CX3CL1) and stromal-derived factor 1 alpha (CXCL12) but not monocyte chemoattractant protein 1 (CCL2).These results demonstrate that FKN preferentially mediates arrest and migration of CD16+ monocytes and suggest that recruitment of this proinflammatory monocyte subset to vessel walls via the CX3CR1-FKN pathway may contribute to vascular and tissue injury during pathological conditions.

View Article: PubMed Central - PubMed

Affiliation: Department of Cancer Immunology and AIDS, Dana-Farber Cancer Institute, Harvard Medical School, Boston, MA 02115, USA.

ABSTRACT
CD16+ monocytes represent 5-10% of peripheral blood monocytes in normal individuals and are dramatically expanded in several pathological conditions including sepsis, human immunodeficiency virus 1 infection, and cancer. CD16+ monocytes produce high levels of proinflammatory cytokines and may represent dendritic cell precursors in vivo. The mechanisms that mediate the recruitment of CD16+ monocytes into tissues remain unknown. Here we investigate molecular mechanisms of CD16+ monocyte trafficking and show that migration of CD16+ and CD16- monocytes is mediated by distinct combinations of adhesion molecules and chemokine receptors. In contrast to CD16- monocytes, CD16+ monocytes expressed high CX3CR1 and CXCR4 but low CCR2 and CD62L levels and underwent efficient transendothelial migration in response to fractalkine (FKN; FKN/CX3CL1) and stromal-derived factor 1 alpha (CXCL12) but not monocyte chemoattractant protein 1 (CCL2). CD16+ monocytes arrested on cell surface-expressed FKN under flow with higher frequency compared with CD16- monocytes. These results demonstrate that FKN preferentially mediates arrest and migration of CD16+ monocytes and suggest that recruitment of this proinflammatory monocyte subset to vessel walls via the CX3CR1-FKN pathway may contribute to vascular and tissue injury during pathological conditions.

Show MeSH
Related in: MedlinePlus