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Fig. 1 | Molecular Medicine

Fig. 1

From: Inhibition of HMGB1/RAGE-mediated endocytosis by HMGB1 antagonist box A, anti-HMGB1 antibodies, and cholinergic agonists suppresses inflammation

Fig. 1

Anti-HMGB1 antibodies attenuated LPS-induced TNF release in vitro and in vivo. a. Upper: Male BALB/c mice received IP injection of polyclonal rabbit anti-HMGB1 antibody or rabbit control IgG as indicated at time point − 16 h, LPS (7.5 mg/kg) at 0 h (IP injection). Mice were euthanized 2 h post LPS administration, and serum TNF was assayed. N = 6–8 mice per group. *: P < 0.05 vs. IgG group. Lower: Male BALB/c mice received IP injection (500 μg/mouse) of polyclonal rabbit anti-HMGB1 antibody at indicated time points, and were challenged with LPS (7.5 mg/kg, IP injection). Mice were euthanized 2 h post LPS injection, serum TNF levels were measured by ELISA. N = 5–10 per group. *: P < 0.05 vs. LPS alone. b. Male BALB/c mice had IP injection of m2G7 and LPS and were euthanized 2 h later. Serum TNF was measured. N = 10 mice per group. *: P < 0.05 vs. LPS alone. c. Primary human macrophages were seeded at a density of 105 cells/well in 24-well culture plates and were stimulated with LPS (2 ng/ml) in the presence or absence of m2G7 for 16 h in serum-free Opti-MEM medium. TNF released in the supernatant was measured. N = 3–4 separate experiments each from different donors. *: P < 0.05 vs. LPS alone. d. Primary human macrophages were plated in 60 mm plates (106 cells/well) and stimulated with LPS in the presence or absence of m2G7 for 60 min. Total RNA was isolated, TNF gene expression was examined and expressed as fold change from the GAPDH level. N = 4–5 experiments each from different donors. Data are expressed as percent of LPS stimulation alone. *: P < 0.05 vs. LPS alone. e. Primary human macrophages in 96-well plate (5 × 104 cells/well) were activated with LPS in the presence or absence of box A for 2 h at 37 °C. TNF (upper) and IL-6 (lower) released were measured. N = 4 separate experiments each from different donors. *: P < 0.05 vs. LPS alone

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