Application of Resonance Raman spectroscopy for the direct detection of manganese porphyrins and their redox state in endothelial cells

MURGIDA D; TOVMASYAN A; CARBALLAL S; RADI R; FERRER SUETA G; VALEZ V; BATINIC-HABERLE I; ALVAREZ PAGGI D

Berlin

Congreso; OCC World Congress and Annual SFRR-E Conference 2017 Metabolic Stress and Redox Regulation Berlin; 2017

Oxygen Club of California

Manganese porphyrins (MnPs) can act as efficient catalytic antioxidants. MnPs can reach mitochondria and confer pharmacological protection in different models of disease where oxidants such as peroxynitrite participate. MnPs quantitation and subcellular distribution by LC-MS/MS has been reported; however, a direct method to evaluate their uptake and redox state in living cells has not been developed. Here, we applied Resonance Raman (RR) confocal microscopy and confirmed that the MnPs-based lipophilic SOD mimics, MnTnBuOE-2-PyP5+ and MnTnHex-2-PyP5+ were incorporated into endothelial cells, and that Mn(III)P can be reduced by intracellular components such as the mitochondrial electron transport chain. Experiments with isolated mitochondria revealed the reduction to Mn(II)P, which was in part affected by the inhibitors of electron transport chain, supporting the action of MnPs as efficient redox active compounds in mitochondria. We have also characterized the reaction kinetics of MnTnBuOE-2-PyP5+ with peroxynitrite and evaluated the cytoprotective capacity of MnPs by exposing the endothelial cells to nitro-oxidative stress induced by peroxynitrite. This data introduce a novel application of the RR spectroscopy for the direct detection of MnPs and their redox state in living cells, and helps to rationalize its antioxidant capacity in biological systems.