166 Assessment of Chronic Spontaneous Urticaria by Serum-Induced TNF & ALPHA; and MMP-9 Release

BACKGROUND: Previous studies from our group have demonstrated that IgE-mediated basophil activation leads to release of TNFα that in turn can induce matrix metallo-proteinase-9 (MMP-9) release from monocytes. We wished to investigate if serum from chronic spontaneous urticaria-patients with auto-antibodies against IgE/IgE-receptor could induce TNFα and MMP-9 release from donor PBMCs, and if release levels could be used to assess severity and activity of chronic spontaneous urticaria (CSU). METHODS: Peripheral blood mononuclear cells (PBMCs) were isolated from whole blood from healthy donors and basophils isolated with MACS Basophil Isolation Kit to 97 to 99% purity. Cells were pulsed 1 hour with or without anti-IgE or with sera from CSU-patients/healthy controls and incubated for a total of 21 h before protein analysis of supernatants. MMP-9 and TNFα in supernatants were measured with commercial ELISAs (R & D), and histamine release determined with HR-test from RefLab ApS. RESULTS: Stimulations with serum-pools demonstrated that CSU-serum, in contrast to healthy controls, was able to induce TNFα-release from isolated basophils. 10 sera from healthy controls and 22 sera from CSU-patients were tested for serum-induced histamine, TNFα and MMP-9 release. The CSU sera were grouped by reaction/no reaction in the autologous skin serum test (ASST), each group consisting of 11 sera. Nine of the 22 CSU-sera were found positive in the HR-test, 6 sera from ASST+ and 3 from ASST- patients. Sera from ASST+ and ASST- patients were observed to induce highly significant MMP-9 and TNFα release from donor PBMCs when compared to sera from healthy controls (P < 0.001). Urticaria assessment score (UAS) did not appear to correlate with release levels for histamine, TNFα or MMP-9 in either group but in the ASST+ group, the ASST score appeared to be positive correlated to histamine and TNFα release and to a smaller degree to MMP-9 release. CONCLUSIONS: We have shown for the first time that serum from CSU-patients, in contrast to serum from healthy controls, can induce TNFα release from isolated basophils, as well as TNFα and MMP-9 from donor PBMCS. Release levels appeared to be positive correlated to ASST reaction in ASST+ patients but not to disease severity for CSU patients in general.