Abstract
Background: Elevated levels of iron impair immune defence mechanisms specifically cell mediated immunity and macrophage function, favors infection with Mycobacterium tuberculosis (M.tb), its replication, progression to clinical disease and death form tuberculosis (TB). Chelation of iron in individuals with an excessive iron burden may restore host defence mechanisms, decrease M.tb viability and replication and could find application in the prevention and treatment strategies in a setting where both iron overload and TB are prevalent.
Aim: The aim of this study was to investigate the effect of iron chelation on mycobacterial replication, host viability and defence mechanisms in iron-loaded monocyte-macrophages during M.tb infection.
Materials and Methods: Mycobacterial replication was monitored using the microplate AlamarBlueTM assay (MABA) for M.tb strains H37Ra and H37Rv, or detection of green fluorescent protein (GFP) expression by BCG (GFP-BCG). Mitochondrial membrane potential (MMP), phosphatidylserine (PS) exposure and plasma membrane integrity of premonocytic U937 cells differentiated by vitamin D3 served as indicators of host cell viability after treatment with 500M FeSO4.7H2O alone or in combination with 500 M desferrioxamine (DFO) or silybin, and infection with M.tb at 1:1 infection ratio. Superoxide anion radical (O2-•) and total nitrate/nitrite generation was monitored as host defence mechanisms by absorption spectroscopy and fluorimetry respectively.
Results: Iron supplementation enhanced intra- and extracellular growth of M.tb and BCG. Chelation of iron with DFO prevented the enhanced replication of mycobacteria promoted by iron. Iron overload increased host cell death of H37Ra-infected monocyte-macrophages through increased PS exposure and decreased MMP and plasma membrane integrity, while increasing O2-• production and decreasing NO production. DFO restored the iron-related increase in PS exposure, plasma membrane integrity, O2-• and NO production to levels similar in H37Ra-infected, iron-normal cells, while MMP remained suppressed. In contrast, infection of iron-loaded cells with H37Rv reduced the already suppressed monocyte-macrophage MMP while not affecting cell death or NO production. DFO restored the iron-related suppression of O2-• in H37Rv-infected cells, and induced the production of NO while not affecting host cell death.
Conclusion: These results confirm the replication enhancing effect of iron-loading on M.tb. Its suppressive influence on macrophage viability and defence mechanisms by increasing cell death and decreasing NO production during infection, is also highlighted. The beneficial effect of iron chelation by DFO through the inhibition of mycobacterial replication and restoration of host viability and defence mechanisms are suggested.
Prof. Liza Bornman