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Effect of interleukin-5 exposure during in vitro eosinophilopiesis on MAC-1 adhesion molecule expression and function
K J Hamann1, S P Neeley, T L Dowling
1Department of Medicine, University of Chicago, IL 60637, USA.
Abstract:
We examined the selective effects of interleukin (IL-5) in regulating the maturational expression of surface adhesion molecules on human eosinophils and adhesion to endothelial cells during eosinophiiopolesis in vitro. Expression of the beta 2 integrins (CD11/CD18) and the beta 1 integrin, VLA-4 (CD49d/ CD29), was assessed during development in culture with IL-3, IL-5, and granulocyte-macrophage colony stimulating factor in cultures of human umbilical cord blood-derived eosinophil (CDE) precursor cells. Expression of both CD11b and CD18 subunits of Mac-1 was lower on CDE which were continuously (= chronically) exposed to IL-5 than on CDE which were cultured without IL-5 for the final week of culture. CD11b expression on cells grown without IL-5 was 71.3 +/- 5.92 (mean specific fluorescence value [MSF] as measured by flow cytometry) versus 52.5 +/- 4.48 MSF for Mac-1 alpha (CD11b) on CDE grown in the continued presence of 2 x 10 - 11 mol/L IL-5 (P < .01). Although expression of VLA-4 decreased as CDE matured, expression of CD29 and CD49d were similar regardless of cytokine exposure for the final week of culture. For eosinophils cultured without IL-5, acute stimulation with 10 - 8 mol/L IL-5 increased CD11b surface expression and increased the number of cells adhering to unstimulated human umbilical vein endothelial cells (HUVEC) from 4,570 +/- 780 cells (9.14 +/- 1.56% adhesion) to 8,385 +/- 515 cells (16.8 +/- 1.03% adhesion) (P < .01). Basal adhesion to unstimulated HUVEC of CDE cultured continuously with IL-5 was comparable (8.62 +/- 1.12% adhesion; P = NS), but neither CD11b expression (50.3 +/- 11.8 MSF; P = NS v control) nor adhesion to HUVEC (6.77 +/- 1.35%; P = NS) was enhanced in these eosinophils after acute stimulation with IL-5. Blockade of adhesion to IL-1-stimulated HUVEC caused by the anti-CD49d monoclonal antibody (MoAb), HP2/1, was comparable for cells cultured with IL-5 and without IL-5. However, the anti-CD18 MoAb, R15.7, caused 47.6 +/- 5.08% inhibition of adhesion of eosinophils cultured without IL-5 and only 25.8 +/- 5.20% for cells cultured continuously with IL-5 (P < .01), and failed to block significantly the adhesion of only the latter cells to IL-4-stimulated HUVEC. Our data show that continuous, chronic exposure to low concentrations of IL-5 causes decreased expression of Mac-1 and refractoriness to acute stimulation with IL-5 of adhesion to HUVEC. These data further demonstrate that CDE maturing in the continued presence of IL-5 adhere to HUVEC predominantly through VLA-4 ligation.
Insights
Chronic exposure to interleukin-5 (IL-5) reduces Mac-1 expression on eosinophils, making them less responsive to IL-5 stimulation. These eosinophils primarily adhere to endothelial cells via VLA-4, not Mac-1.
Area of Science:
- Immunology
- Cell Biology
- Hematology
Background:
- Eosinophils play a key role in allergic inflammation and host defense.
- Interleukin-5 (IL-5) is a critical cytokine for eosinophil development, survival, and activation.
- Understanding eosinophil adhesion to endothelial cells is crucial for elucidating inflammatory processes.
Purpose of the Study:
- To investigate the selective effects of IL-5 on the expression of surface adhesion molecules in human eosinophils.
- To determine how IL-5 influences eosinophil adhesion to endothelial cells during in vitro eosinophilopoiesis.
- To elucidate the specific adhesion molecules involved in IL-5-mediated eosinophil-endothelial cell interactions.
Main Methods:
- Human umbilical cord blood-derived eosinophil (CDE) precursor cells were cultured with IL-3, IL-5, or granulocyte-macrophage colony-stimulating factor.
- Expression of beta 2 integrins (CD11/CD18) and beta 1 integrin VLA-4 (CD49d/CD29) was assessed using flow cytometry.
- Eosinophil adhesion to human umbilical vein endothelial cells (HUVEC) was measured, with and without cytokine stimulation and antibody blockade.
Main Results:
- Chronic IL-5 exposure decreased the expression of Mac-1 (CD11b/CD18) on CDE compared to cells cultured without IL-5.
- Acute IL-5 stimulation enhanced Mac-1 expression and adhesion to HUVEC in eosinophils not chronically exposed to IL-5.
- Eosinophils chronically exposed to IL-5 showed reduced Mac-1 expression and were refractory to acute IL-5-induced adhesion enhancement.
- Adhesion of chronically IL-5-exposed eosinophils to HUVEC was predominantly mediated by VLA-4, as indicated by antibody blockade experiments.
Conclusions:
- Continuous, chronic exposure to IL-5 downregulates Mac-1 expression on maturing eosinophils.
- Chronic IL-5 exposure leads to refractoriness to acute IL-5-mediated increases in eosinophil adhesion.
- Eosinophils maturing in the presence of IL-5 preferentially utilize VLA-4 for adhesion to endothelial cells.