(P30-037-25) Marginal Iron Depletion Impairs Mitochondrial Complex I Activity and Reduces NDUFB8 and ISCU Protein Levels in Mouse C2C12 Myoblast Cells

Objectives: Mammalian cell models have been used to identify biological functions of iron, including mitochondrial energy metabolism. Models of iron deficiency often use acute high-dose iron chelators to induce iron depletion which leads to severe impairments in mitochondrial energy metabolism. These models do not address marginal stages of iron depletion and have yet to define the dose at which declines in oxidative phosphorylation and iron-dependent pathways begin. The objective of this research is to establish the impact of marginal iron depletion on mitochondrial electron transport chain (ETC) complexes and the mitochondrial iron-dependent synthesis pathways that may contribute to declines in oxidative phosphorylation.

Methods: Mouse C2C12 myoblast and Hepa1-6 hepatocyte cells were plated in high-glucose DMEM medium; once adhered, medium was replaced with high-glucose DMEM medium containing 4 doses of deferoxamine (DFO), ranging from 0-7.5µM, to induce a range of iron depletion. After 72 hours, transferrin receptor, ETC complex activity, ETC protein levels, and mitochondrial membrane potential were evaluated.

Results: Both cell types exhibited a positive linear relationship between DFO concentration and transferrin receptor. Marginal iron depletion impaired mitochondrial complex activities in both C2C12 (complex I, II, IV) and Hepa1-6 (complex I, IV) cell lines, however, complex II activity was not impaired in Hepa1-6 cells. NDUFB8 (Complex I) and ISCU (iron-sulfur cluster assembly enzyme) protein concentrations decreased in a dose-dependent manner in C2C12 cells, while MT-CO1 (Complex IV) and FECH (ferrochelatase) protein concentrations did not change. Furthermore, in response to DFO treatment, C2C12 cells exhibited an increase in mitochondrial membrane potential while Hepa1-6 cells exhibited a decrease.

Conclusions: Exposure to low dose iron chelators impaired mitochondrial complex activity in both cell lines and decreased proteins involved in complex I and iron-sulfur cluster assembly in C2C12 cells. This suggests impaired mitochondrial complex activity in marginally iron depleted skeletal muscle cells may result from compromised iron-sulfur cluster assembly and complex I proteins.

Funding Sources: National Institutes of Health