Detection of CD 14 on Migrated Monocytes by Specific Antibody: a Possible Quantification for Blood Monocyte Chemotaxis

Tekstra, Janneke; Tuk, Cornelis W.; Beelen, Robert H.J.

doi:10.1016/s0171-2985(96)80018-0

Cited by 2 publications

(1 citation statement)

References 20 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…4) effectively blocked chemotactic activity of these supernatants for monocytes. The percentage of monocytes that migrated in response to 30 ng/ml recombinant human MCP-1 was greater than 50% of that migrating in response to the reference chemoattractant 10 À8 fMLP (data not shown), con®rming previous results [21,30]. The anti-MCP-1 antibody completely abolished the increased monocyte migration towards recombinant human MCP-1, but had no effect on the fMLP-induced chemotaxis (not shown).…”

Section: Bioactivity Of Mcp-1: Monocyte Chemotaxissupporting

confidence: 87%

Infection of human endothelial cells withStaphylococcus aureusinduces the production of monocyte chemotactic protein-1 (MCP-1) and monocyte chemotaxis

Tekstra

Beekhuizen

Gevel

et al. 1999

Clinical and Experimental Immunology

View full text Add to dashboard Cite

Bacterial infection coincides with migration of leucocytes from the circulation into the bacterium-infected tissue. Recently, we have shown that endothelial cells, upon binding and ingestion of Staphylococcus aureus, exhibit proinflammatory properties including procoagulant activity and increased intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) expression on the cell surface, resulting in hyperadhesiveness, mainly for monocytes. The enhanced extravasation of monocytes to bacterium-infected sites is facilitated by the local production of chemotactic factors. From another study we concluded that the locally produced chemokine MCP-1 is important in the recruitment of monocytes to the peritoneal cavity in a model of bacterial peritonitis. In the present study we investigated whether cultured human endothelial cells after infection with bacteria produce and release MCP-1, which in turn stimulates monocyte chemotaxis. We observed that endothelial cells released significant amounts of MCP-1 within 48 h after ingestion of S. aureus. This was dependent on the number and the virulence of the bacteria used to infect the endothelial cells. The kinetics as well as the amount of MCP-1 released by S. aureus-infected endothelial cells differed markedly from that released by endothelial cells upon stimulation with IL-1beta. Supernatant from S. aureus-infected or IL-1beta-stimulated cells promoted monocyte chemotaxis which was almost entirely abrogated in the presence of neutralizing anti-MCP-1 antibody, indicating that most of the chemotactic activity was due to the release of MCP-1 into the supernatant. Our findings support the notion that endothelial cells can actively initiate and sustain an inflammatory response after an encounter with pathogenic microorganisms, without the intervention of macrophage-derived proinflammatory cytokines.

show abstract

Section: Bioactivity Of Mcp-1: Monocyte Chemotaxissupporting

confidence: 87%

Infection of human endothelial cells withStaphylococcus aureusinduces the production of monocyte chemotactic protein-1 (MCP-1) and monocyte chemotaxis

Tekstra

Beekhuizen

Gevel

et al. 1999

Clinical and Experimental Immunology

View full text Add to dashboard Cite

show abstract

Migration of Mononuclear Cells in the Modified Boyden Chamber as Evaluated by DNA Quantification and Flow Cytometry

et al. 2000

View full text Add to dashboard Cite

In vitro migration of mononuclear cells in the modified Boyden chamber was evaluated using flow cytometry and DNA quantification (Hoechst 33258) of all adherent and nonadherent cells. The effects of different membrane pore sizes, cell concentrations and incubation times were studied. Pore sizes of 3 and 5 microm resulted in a reduction in the number of nonadherent cells compared with a pore size of 8 microm. Reducing the incubation time from 60 to 40 and 20 min resulted in too few migrating monocytes for analysis by flow cytometry. Flow cytometry showed that both monocytes and lymphocytes migrated and adhered to the membrane when using peripheral blood mononuclear cells (PBMC) to study monocyte migration. Migration of lymphocytes under these conditions is a novel observation. A substantial number of migrated cells could be identified by flow cytometry and quantified by DNA measurement as nonadherent below the membrane. Samples of synovial fluid (n = 49) and plasma (n = 133) as chemoattractants analysed in triplicate resulted in mean coefficient of variation (CV) values of 11 and 9%, respectively. Variation from assay to assay on the same day, using N-formyl-methionyl-leucyl-phenylanine (fMLP) 10(-7) M as chemoattractant resulted in a CV of 13%. Day-to-day variation, using fMLP 10(-7) M as chemoattractant and the same well on three different days, resulted in a CV of 21%. These results were obtained using a pore size of 5 microm, a PBMC concentration of 3 x 10(6)/ml and 60 min of incubation. The combination of DNA quantification and flow cytometry thus allowed characterization and quantification of subsets of migrating adherent as well as nonadherent mononuclear cells.

show abstract

Detection of CD 14 on Migrated Monocytes by Specific Antibody: a Possible Quantification for Blood Monocyte Chemotaxis

Cited by 2 publications

References 20 publications

Infection of human endothelial cells withStaphylococcus aureusinduces the production of monocyte chemotactic protein-1 (MCP-1) and monocyte chemotaxis

Infection of human endothelial cells withStaphylococcus aureusinduces the production of monocyte chemotactic protein-1 (MCP-1) and monocyte chemotaxis

Migration of Mononuclear Cells in the Modified Boyden Chamber as Evaluated by DNA Quantification and Flow Cytometry

Contact Info

Product

Resources

About