Oxidative Modification of Fibrinogen Is Associated With Altered Function and Structure in the Subacute Phase of Myocardial Infarction

Abstract

Objective-Among plasma proteins, fibrinogen represents a major target of oxidative modifications. In patients with postacute myocardial infarction (6 months after the acute event), fibrinogen oxidation-induced carbonyls and fibrinogen function were estimated using in vitro and ex vivo approaches. Fibrinogen structural features and clot architecture were also explored. Approach and Results-In 39 patients with post-acute myocardial infarction and 28 age-, sex-, and risk factor-matched controls, oxidative stress markers (in plasma and in purified fibrinogen fractions), thrombin-catalyzed fibrin polymerization, and plasmin-induced fibrin lysis were estimated. Circular dichroism spectra of purified fibrinogen extracts, electron microscopy, and differential interference contrast microscopy analyses of fibrin clots were also performed. Marked signs of oxidative stress in plasma (P<0.01 versus controls) and, correspondingly, an increased extent of fibrinogen carbonylation (3.5-fold over control values; P<0.01 versus controls) were observed in patients. Furthermore, fibrinogen fractions purified from patients exhibited significantly reduced clotting ability and decreased susceptibility to plasmininduced lysis (P<0.01 versus controls). Alterations in fibrinogen secondary structure, as suggested by circular dichroism spectroscopy, and in fibrin clot architecture, as analyzed by electron and differential interference contrast microscopy, were also identified. Conclusions-Here, we report for the first time that patients with post-acute myocardial infarction present with an overall imbalance in redox status and marked fibrinogen carbonylation associated with altered fibrinogen function, thus suggesting a role for carbonylation as a direct mechanism of fibrinogen function. The observed features occur along with modifications in protein structure and in clot architecture. Results SubjectsDemographic and clinical characteristics of the population studied are summarized in Table 1. Oxidative Stress Markers in Plasma and in FibrinogenAs reported in Table 2, patient plasma displayed significantly higher total PC and thiobarbituric acid reactive substances levels, and lower total antioxidant capacity, than healthy controls (P<0.05 versus controls). Plasma total PC was positively and significantly correlated with fibrinogen PC (P<0.05; R=0.555; Figure 1A) and with thiobarbituric acid reactive substances levels (P<0.05; R=0.444; Figure 1B). Purified fibrinogen from patients displayed significantly increased carbonylation (3.5-fold) in comparison with healthy controls (P<0.01; Figure 1C). PC amounts obtained in human, 2,2′-azobis(2-amidinopropane) dihydrochloride (AAPH)-challenged purified fibrinogen (SIGMA, Milan, Italy) were comparable with those observed in fibrinogen from patients with post-AMI and resulted significantly increased compared with untreated fibrinogen (Table 3). Circular Dichroism Spectra: Analysis of Secondary StructureSecondary protein structure was analyzed by far-UV circular dichroism spectroscopy (Figure 2). In ...

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