Glutathionyl hemoglobin in uremic patients undergoing hemodialysis and continuous ambulatory peritoneal dialysis

Abstract

To assess the redox state in hemodialysis (HD) and continuous ambulatory peritoneal dialysis (CAPD) patients, we focused on the formation of glutathionyl hemoglobin (Hb) because the ratio of oxidized glutathione disulfide (GSSG) to reduced glutathione (GSH) is increased in uremia, and GSSG is a source of glutathionyl Hb. Glutathionyl Hb levels were measured in 30 HD patients, 10 CAPD patients, and 20 healthy subjects by using liquid chromatography/electrospray ionization-mass spectrometry (LC/ESI-MS). Hbbeta showed a peak at 15,868 D in a deconvoluted ESI mass spectrum. Glutathionyl Hbbeta was detected at 16,173 D (15,868 + 305). The peak at 16,173 D was identified as glutathionyl Hbbeta based on the following findings: (1) the peak disappeared by reducing the sample with dithiothreitol, and (2) the peak could be detected at a high level by incubating Hb in vitro with GSH in water at 37 degrees C for seven days. Glutathionyl Hb levels expressed as the peak height ratios of glutathionyl Hbbeta to intact Hbbeta were significantly elevated in HD patients (8.0 +/- 3.6%, mean +/- SD, N = 30, P < 0.0001) and CAPD patients (5.9 +/- 2.7%, N = 10, P < 0.05) as compared with normal subjects (3.0 +/- 1.6%, N = 20). However, there were no significant differences in the glutathionyl Hb levels before (8.7 +/- 3.2%, N = 12) and after HD (8.7 +/- 2.8%, N = 12). Glutathionyl Hb levels were increased in HD and CAPD patients, probably because of enhanced oxidative stress. The measurement of glutathionyl Hb may be useful to assess oxidative stress in uremic patients.