Pharmacologic ascorbate as a pro-drug for hydrogen peroxide release to kill mycobacteria

Background and purpose: Tuberculosis is one of the most highly fatal diseases worldwide, and one-third of the world's population has been infected with Mycobacterium tuberculosis (M. tuberculosis). A previous study showed that M. tuberculosis was highly susceptible to being killed by ascorbate (i.e. vitamin C, VC), but the molecular mechanisms of the bactericidal activity of VC against M. tuberculosis are still not well understood. Experimental approach: We assayed the effects of VC as an anti-tuberculosis drug against mycobacteria (i.e. M. bovis BCG or M. tuberculosis H37Rv) in macrophages (i.e. RAW 264.7 cells). Relative global protein expression changes in 5 mM VC-treated and control samples of H37Rv were investigated by Tandem mass tag (TMT)-based quantitative proteomic analysis. qRT-PCR was also used to measure the differential expression of six intracellular stress response mycobacteria genes. Key results: Quantitative proteomic analysis showed that 11 peptide components including rip3, fdxA, Rv2028c, mtp, LH57_00670, hspX, pfkB, Rv1824, Rv1813c, LH57_08410 and Rv2030c were up-regulated and 17 peptide components were down-regulated in 5 mM VC-treated H37Rv compared with the control samples. qRT-PCR also verified that VC could induce the expression of six genes (hsp, fdxD, furA, devR, hspX, and dnaB) in BCG and H37Rv. We also found that exosomes from RAW 264.7 cells treated with pharmacologic VC could kill M. bovis BCG in vitro. Conclusion and implications: Our results demonstrated that the bactericidal activity of VC against mycobacteria, as a pro-drug for hydrogen peroxide formation (H2O2), was dependent on reactive oxygen species production and the activated oxidative stress pathway, which suggested that pharmaceutical VC and exosomes from macrophages treated with VC could be used as potential anti-tuberculosis drugs.