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Figure 5 | Journal of Inflammation

Figure 5

From: Cross-linking of IgGs bound on circulating neutrophils leads to an activation of endothelial cells: possible role of rheumatoid factors in rheumatoid arthritis-associated vascular dysfunction

Figure 5

ICAM -1 expression and IL- 8 production in HUVECs. HUVECs were incubated for 48 hrs with supernatants of control (Ctrl) or IgGs cross-linked human neutrophils (Stimulated) prepared as described in Material and Methods. A: The presence of ICAM-1 on HUVECs was measured by flow cytometry. HUVECs were washed with PBS and detached by 5 minutes incubation with accutase at 37°C. Cells were centrifuged, resuspended in HBSS/0.005% BSA and incubated with anti-ICAM-1 antibody (10 μg/ml) for 1 hour at 4°C before wash. They were then incubated with the secondary anti-mouse FITC-conjugated antibody (1/100) for 30 minutes at 4°C, washed and fluorescence was analysed. B: IL-8 release by HUVECs was measured by ELISA. The supernatants of HUVECs were collected and the remaining cells were discarded by centrifugation. The amount of IL-8 in the supernatants was measured in duplicates using the human IL-8 CytoSet from Invitrogen (catalog no.CHC1303) following the manufacturer’s instructions. IL-8 contained in neutrophils supernatants was substracted. Results are expressed as mean +/- SEM of three independent experiments. * = P < 0.05 (one-sample t test).

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