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. 2020 Jun 24;11:1122. doi: 10.3389/fimmu.2020.01122

Figure 2.

Figure 2

Disulphide-HMGB1 derives from non-myogenic cells in injured muscle. (A) Western blot probed with anti-CD45 (upper panel) and anti-HMGB1 (lower panel) antibodies in reducing conditions on tibialis anterior (TA) muscle lysates from WT or HMGB1 mKO mice at indicated time points after cardiotoxin (CTX) injection. Ctrl, control uninjured muscles. (B,C) Quantification of CD45 (B) and HMGB1 (C) protein expression, normalized on Ponceau staining, before (Ctrl) and after CTX injection (CTX at 1, 2, and 7 d) in TA and triceps muscle lysates (n ≥ 4 muscles/time point, n = 3 mice/genotype). A.U. = arbitrary unit. (D,E) Western blot probed with anti-HMGB1 antibody in non-reducing conditions (D) on TA muscle lysates from WT or HMGB1 mKO mice at indicated time points after CTX injection. The upper band corresponds to the fully reduced-HMGB1 (frHMGB1) and the lower band to the disulphide-HMGB1 (dsHMGB1). (E) Percentage of HMGB1 redox isoforms expression from WT or HMGB1 mKO mice before (Ctrl) and after CTX injection (CTX at 1, 2, 5, and 7 d) in TA and triceps muscle lysates (n ≥ 3 muscles/time point; n ≥ 4 mice/genotype). Data represent the means ± SEM and statistical significance was calculated by Student T-test (B,C) and Two-way ANOVA (E). **P < 0.01; ***P < 0.001; ##P < 0.01 (Ctrl vs. CTX).