Figure 3

Inflammatory cytokines were induced by human IL-32β in mice. (a) Raw 267.4 was cultured with the supernatant of human IL-32β (h IL-32β) or mock-transfected mammalian cells (293T) for 24 hours. Left, relative expression of mouse tumor necrosis factor alpha (mTNFα), compared with β-actin; right, secreted TNFα protein level measured by ELISA. (b) We generated hIL-32β overexpressed mice by transplantation of hIL-32β-transduced bone marrow cells. The expression of green fluorescent protein, was analyzed by flow cytometry 6–9 weeks after transplantation. (c) Expression of mTNFα, mIL-1β and mIL-6 in the cultured splenocytes of the control group (white bars; n = 3), or bone-marrow chimeric mice of the mock group (BM-Mock mice) (gray bars; n = 4), or hIL-32β (BM-hIL-32) (black bars; n = 4) with or without 1 μg/ml lipopolysaccharide (LPS). Concentrations of indicated cytokines of the cultured supernatants are shown in the right-hand figures. (d) Serum concentration of mTNFα determined in control mice (n = 4), in BM-Mock mice (n = 8), and in BM-hIL-32 mice (n = 8). (e) Expression of mTNFα in splenic F4/80⁺ CD11c^- macrophages of BM-Mock mice (gray bars; n = 4) and in BM-hIL-32 mice (black bars; n = 4). (f) Expression of mTNFα, mIL-1β, and mIL-6 in LPS-stimulated splenic F4/80⁺ CD11c^- macrophages and CD11c⁺, CD3^-, and CD19^- dendritic cells in BM-Mock mice (gray bars; n = 4), and in BM-hIL-32 mice (black bars; n = 4). Data are representative of at least three independent studies. *P < 0.05, **P < 0.01, BM-hIL-32 mice versus BM-Mock mice or control mice.