Spontaneous Release of Human Serum Albumin S-Bound Homocysteine in a Thiol-Free Physiological Medium

Elevated plasma homocysteine (Hcy) concentrations independently predict cardiovascular disease. However, the transport of Hcy into pathological tissues such as atherosclerotic plaques, characterized by relatively low local thiol concentrations, is largely unknown. We sought to address if albumin-bound Hcy can be released in a thiol-free medium with or without previous incubation with Hcy and homocysteine thiolactone (HTL). We found that Hcy release was dependent on the baseline amount of albumin-bound Hcy. After 48 h incubation in a thiol- free medium the quantity of albumin-bound Hcy released from commercial human serum albumin (HSA) was 1.15 mmol/mol prot. If HSA was pre-incubated with 50 µmol/L reduced Hcy and then transferred into a free-thiol medium (HSA-S-Hcy), the amount of Hcy released after 48 h increased to 33.5 mmol/mol prot. If HSA was pre-incubated with 5 mmol/L HTL and then with 50 µmol/L of reduced Hcy (HSA-HTL-S-Hcy), the amount of Hcy released increased to 92.8 mmol/mol prot. Hcy release from HSA-HTL-S-Hcy further increased in presence of cysteine (Cys), glutathione (GSH), or Cys + GSH in the medium. Therefore, a significant amount of albumin-bound Hcy is released into thiol-free environments, similar to atherosclerotic plaques, with potential deleterious effects on vascular homeostasis, atherosclerosis and thrombosis.