CD69, CD18 and CD49d blockade resulted in reduced TNF-α in Tck co-cultures and rheumatoid synovium culture. (a) CD4+CD45RO+ cells were fixed after 8 days of cytokine stimulation with 1% paraformaldehyde and then co-incubated with blocking antibodies against CD18 and CD69, as described in Materials and methods. Cytokine-activated T (Tck) cells were co-cultured with fresh monocytes at a ratio of 8:1. Supernatants were harvested after 18 hours and TNF-a production measured by ELISA. This figure represents three experiments conducted in different donors. (b) Rheumatoid arthritis (RA) synovial cell mononuclear cells from RA donor 2 (see Table 3) were cultured in triplicate in the presence of antibodies against CD69, CD18, or CD49d for 24 hours, as described in Materials and methods. Supernatants were harvested and spontaneous TNF-α production was measured using ELISA. **P < 0.01 and *P < 0.05, as assessed by one-way analysis of variance with post hoc Bonferroni correction.