IL-1β (Interleukin-1 beta) is a proinflammatory and immunoregulatory cytokine involved in a variety of cellular activities. It has been elucidated that IL-1β stimulation of cells activates MMP-9 (matrix metalloproteinases-9) secretion by the activation of the dual signalling pathways. Thus, a stimulation assay of IL-1β was conducted using 3T3 cell line, and the MMP-9 activity was detected through gel zymography. Briefly, 1×106 3T3 cells were cultured overnight in 5%DMEM, washed with serum-free medium and then stimulated with different concentrations of IL-1β for 20h, and cell lysates were collected to measure MMP-9 activity. Cell lysates samples were denatured by SDS loading buffer, electrophoresed through sodium dodecyl sulphate–polyacrylamide gel (SDS–PAGE; 10% gels) containing gelatin (1mg/mL) with nonreducing conditions. After renaturation, incubation and coomassiebrilliant blue (CBB)-stained, MMPs hydrolyzed gelatin nearby, indicated by the white binds on the gel.
Result： Increased MMP-9 activity in 3T3 cells due to the stimulation of IL-1β was shown in Figure 1.
Figure. Activation of MMP-9 by IL-1β.