Tyrosine nitration in plasma proteins from patients undergoing hemodialysis

A growing body of evidence suggesting that oxidative stress might be one of the most important complications occurring during hemodialysis (HD) has accumulated. However, although the role of reactive oxygen species has been investigated extensively, little is known about the involvement of reactive nitrogen species. In the present investigation, levels of protein modifications in the form of tyrosine nitration in patients undergoing long-term HD therapy were evaluated.

Tyrosine nitration of plasma proteins was detected by means of Western blotting using a specific nitrotyrosine-recognizing monoclonal antibody, and band intensity was evaluated by using image analysis software. Immunoprecipitation of plasma proteins by antinitrotyrosine-agarose--conjugated antibodies, followed by Western blotting, was used in an attempt to identify the nitrated proteins.

Although several proteins with nitrated tyrosine residues were observed in plasma of healthy individuals, increased nitration levels were observed in some specific proteins in all patients tested (n = 25) compared with controls (n = 6). At least 6 apparent bands appeared to be more nitrated than their counterparts in plasma from controls. Ceruloplasmin was identified as 1 of the proteins with significantly increased nitration in patients.

Results of the present investigation show that specific plasma proteins of HD patients are post-translationally modified by nitration of their tyrosine residues. The nature of these proteins, as well as the exact molecular mechanisms and consequences of these modifications, warrant additional investigation.