Published online before print August 7, 2008

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(American Journal of Pathology. 2008;173:728-740.)
© 2008 American Society for Investigative Pathology
DOI: 10.2353/ajpath.2008.080047

Enhanced Nitrosative Stress during Trypanosoma cruzi Infection Causes Nitrotyrosine Modification of Host Proteins

Implications in Chagas’ Disease

Monisha Dhiman^*, Ernesto Satoshi Nakayasu, Yashoda Hosakote Madaiah, Brobey K. Reynolds, Jian-jun Wen^*, Igor Correia Almeida and Nisha Jain Garg^*¶||

From the Departments of Microbiology and Immunology,^* Pediatric Child Health Research, Internal Medicine–Gastroenterology, Pathology,^¶ and the Center for Biodefense and Emerging Infectious Diseases and the Sealy Center for Vaccine Development,|| University of Texas Medical Branch, Galveston; and the Department of Biological Sciences, The Border Biomedical Research Center, University of Texas at El Paso, El Paso, Texas

Oxidative/nitrosative stress may be important in the pathology of Chagas’ disease. Experimental animals infected by Trypanosoma cruzi showed an early rise in myocardial and peripheral protein-3-nitrotyrosine (3NT) and protein-carbonyl formation that persisted during the chronic stage of disease. In comparison, experimental chronic ethanol-induced cardiomyopathy was slow to develop and presented with a moderate increase in oxidative stress and minimal to no nitrosative stress after long-term alcohol feeding of animals. The oxidative stress in both chagasic animals and animals with ethanol-induced cardiomyopathy correlated with the persistence of reactive oxygen species-producing inflammatory intermediates. Protein-3NT formation in T. cruzi-infected animals was associated with enhanced nitric oxide expression (inferred by nitrite/nitrate levels) and myeloperoxidase activity, suggesting that both peroxynitrite- and myeloperoxidase-mediated pathways contribute to increased protein nitration in Chagas’ disease. We used one- and two-dimensional gel electrophoresis and Western blot analysis to identify disease-specific plasma proteins that were 3NT-modified in T. cruzi-infected animals. Nitrated protein spots (56 in total) were sequenced by matrix-assisted laser desorption ionization/time of flight mass spectrometry and liquid chromatography–tandem mass spectrometry and identified by a homology search of public databases. Clustering of 3NT-modified proteins according to their functional characteristics revealed that the nitration of immunoglobulins, apolipoprotein isoforms, and other proteins might perturb their functions and be important in the pathology of Chagas’ disease. We also showed that nitrated peptides derived from titin and -actin were released into the plasma of patients with Chagas’ disease. Such modified proteins may be useful biomarkers of Chagas’ disease.