Taken together, we conclude that CTLA-4-Ig affects the level of cytokines and chemokines in the affected tissue by significantly reducing IL-4, IL-1β, MIP-2 and IP-10. To analyse the effect of CTLA-4-Ig on systemic inflammation, serum samples taken 24 and 48 h after challenge were analysed by ELISA for the acute-phase proteins

SAP and haptoglobin. These factors have been shown to be reliable click here markers of inflammation in this model as their serum levels correspond to ear swelling (A.D.C. and C.H., data not shown). Furthermore, increased serum concentration of these components indicates systemic inflammation with involvement of the liver [18]. Figure 6b,d shows that serum levels of SAP and haptoglobin were reduced significantly following treatment with CTLA-4-Ig compared to control treatment at both 24 and 48 h after challenge in the DNFB-induced model, and in the oxazolone-induced Metformin solubility dmso model serum concentrations of haptoglobin were suppressed significantly after both 24 and 48 h (Fig. 6c). Similarly, SAP was reduced significantly after 48 h but not at 24 h (Fig. 6a). Based on these findings, we conclude that CTLA-4-Ig inhibits systemic inflammation as measured by circulating levels of SAP and haptoglobin. In the CHS model, it is not known whether CTLA-4-Ig exerts its effect in the sensitization

phase alone or whether the presence of CTLA-4-Ig is also important in the effector phase. To test this, we set up an adoptive Carnitine dehydrogenase transfer system in which donor mice were sensitized in the presence or absence of CTLA-4-Ig. After 5 days, cells from the draining lymph node were transferred to recipient mice which had been treated with CTLA-4-Ig 24 h earlier or left untreated. Recipient mice were subsequently challenged with DNFB and ear swelling was measured 24, 48 and 72 h after challenge. As shown in Fig. 7, mice transferred

with cells exposed to CTLA-4-Ig during both the sensitization phase and the challenge phase or during the sensitization phase alone (labelled +/+ and +/−, respectively) exhibited a significantly suppressed ear-swelling response compared to the untreated control group (labelled −/−). In contrast, the mice which were treated only with CTLA-4-Ig during the challenge phase (labelled −/+) exhibited ear swelling similar to the untreated mice. Taken together, these results indicate that CTLA-4-Ig exerts its immunosuppressive effect primarily during the sensitization phase. We next tested whether regulation of cytokines and chemokines in the inflamed tissue followed the same pattern as ear swelling by comparing levels of IL-1β, IL-4, IP-10 and MIP-2 in the adoptive transfer model treated with CTLA-4-Ig in the sensitization or challenge phase only.