Burn Serum Stimulates Myoblast Cell Death Associated with IL-6-Induced Mitochondrial Fragmentation.

BACKGROUND

Burn patients suffer muscle mass loss associated with hyperinflammation and hypercatabolism. The mitochondria are affected by this metabolic alteration. Mitochondrial fission activates a caspase cascade that ultimately leads to cell death. We postulate that burn-induced muscle loss is associated with increased mitochondrial fission and subsequent functional impairment. Further, we investigated whether the cytokine IL-6 plays a major role in mitochondrial fission-associated cell death after burn.

METHODS

Murine myoblast C2C12 cells were treated with 10% serum isolated either from control rats or 40% total body surface area burned rats. Mitochondria were labeled with MitoTracker Green for live cell images. Mitochondrial function was assessed with an Enzo Mito-ID membrane potential cytotoxicity kit. Protein signals were detected by Western blot analysis. Moreover, recombinant IL-6 was applied to stimulate C2C12 to differentiate the role of cytokine IL-6; lastly, we treated burn serum-stimulated cells with IL-6 antibodies.

RESULTS

Caspase 3 activity increased in C2C12 cells with burn serum stimulation, suggesting increased cell death in skeletal muscle after burn. Mitochondrial morphology shortened and mitochondrial membrane potential decreased in cells treated with burn serum. Western blot data showed that mitofusion-1 expression significantly decreased in burn serum-treated cells, supporting the morphologic observation of mitochondrial fission. Mitochondrial fragmentation increased with IL-6 stimulation, and IL-6 antibody decreased caspase 3 activity and mitochondrial membrane potential improved in burn serum-stimulated cells.

CONCLUSION

Burn serum caused muscle cell death associated with increased mitochondrial fission and functional impairment. This alteration was alleviated with IL-6 antibody treatment, suggesting the cytokine plays a role in mitochondrial changes in muscle after systemic injury.