Figure 6.

Metformin did not change the redox states or molecular weight of HMGB1. A, HMGB1 was incubated at 4 °C in the absence or presence of 100 mm metformin for the indicated time periods and analyzed by SDS-PAGE without or with β-mercaptoethanol (2-ME) treatment at 95 °C for 5 min. HMGB1 was detected by immunoblotting with anti-HMGB1 antibody. It is noted that disulfide and fully-reduced HMGB1 can be separated by mobility in SDS-PAGE without 2-ME treatment. Bands in the marker (M) lane indicated 30, 40, and 50 kDa from the bottom. B, mass spectrometry analysis of HMGB1 incubated without (upper diagram) or with 100 mm metformin (lower diagram) at 4 °C for 24 h.