In vitro comparison of hydroxocobalamin (B12a) and the mitochondrial directed therapy by a succinate prodrug in a cellular model of cyanide poisoning

Abstract

The objective of this study was to compare the use of hydroxocobalamin (B12a) and a succinate prodrug to evaluate for improvement in mitochondrial function in an in vitro model of cyanide poisoning. Peripheral blood mononuclear cells (PBMC) and human aortic smooth muscle cells (HASMC) incubated with 50 mM of sodium cyanide (CN) for five minutes serving as the CN group compared to controls. We investigated the following: (1) Mitochondrial respiration; (2) Superoxide and mitochondrial membrane potential with microscopy; (3) Citrate synthase protein expression. All experiments were performed with a cell concentration of 2−3 × 106 cells/ml for both PBMC and HASMC. There were four conditions: (1) Control (no exposure); (2) Cyanide (exposure only); (3) B12a (cyanide exposure followed by B12a treatment); (4) NV118 (cyanide followed by NV118 treatment). In this study the key findings include: (1) Improvement in key mitochondrial respiratory states with the succinate prodrug (NV118) but not B12a; (2) Attenuation of superoxide production with treatment of NV118 but not with B12a treatment; (3) The changes in respiration were not secondary to increased mitochondrial content as measured by citrate synthase; (4) The use of easily accessible human blood cells showed similar mitochondrial response to both cyanide and treatment to HASMC. The use of a succinate prodrug to circumvent partial CIV inhibition by cyanide with clear reversal of cellular respiration and superoxide production that was not attributed to changes in mitochondrial content not seen by the use of B12a.