• Altered platelet reactivity in peripheral vascular disease complicated with elevated plasma homocysteine levels.

      Riba, Rocio; Nicolaou, Anna; Troxler, M.; Homer-Vanniasinkam, Shervanthi; Naseem, Khalid M. (2004)
      Elevated plasma concentrations of the sulphur-containing amino acid homocysteine (Hcy) is associated with increased risk of atherosclerosis and arterial thrombosis. The mechanism by which Hcy exerts these effects has yet to be fully elucidated, although a variety of possible mechanisms have been proposed, including endothelial dysfunction or haemostatic abnormalities. However, the influence of Hcy on platelets, cells central to the atherothrombotic process, has never been addressed directly in patient studies. Here, the influence of mild hyperhomocysteinaemia (hHcy) on platelet function was explored in patients with peripheral occlusive arterial disease as evidence by intermittent claudication. Claudicants (n=39) were assigned to one of two subgroups depending on their plasma Hcy concentrations. hHcy claudicants had plasma Hcy concentrations of 18.9±1.0 ¿M (n=24), compared to 11.3±0.5 ¿M for normohomocysteinemic (nHcy) claudicants (n=15) and 12.6±0.7 ¿M for age-matched controls (n=15). Platelet function was evaluated ex vivo in both groups and compared to age-matched controls. Platelet activation and sensitivity to nitric oxide-mediated inhibition was assessed by platelet fibrinogen binding and P-selectin expression. At low concentrations of adenosine diphosphate (ADP; 0.1 ¿M) and thrombin (0.02 U/ml), platelets from hHcy claudicants were more reactive than those from age-matched controls, but not nHcy claudicants. Agonist-induced P-selectin expression was significantly raised in hHcy claudicants compared to all other groups. Interestingly no differences were observed between nHcy claudicants and age-matched controls, indicating that claudication per se did not affect platelet function. Since platelet activity in vivo is determined by the exposure to both agonists and antagonists, we subsequently tested the sensitivity of platelets to inhibition by nitric oxide (NO), using the same platelet markers. Platelets from hHcy claudicants were significantly less sensitive to GSNO (1¿100 ¿M)-mediated inhibition than all other groups. GSNO (1 ¿M) induced 42.6±10 and 39±11.5% inhibition of ADP-induced fibrinogen binding for the nHcy claudicants and age-matched controls, respectively. However, in hHcy claudicants only 16.4±9.7% inhibition was observed, significantly less than the other groups (P<0.01). Again no differences between nHCy claudicants and controls were observed. These results suggest the presence of claudication alone does not influence platelet function but if complicated with mild hyperhomocysteinemia, the sensitivity to agonists is increased, and more importantly, their sensitivity to inhibition is greatly reduced. The overall effect would be an increased propensity for platelet activation. The presence of even mildly elevated plasma Hcy could dramatically increase thrombotic risk.
    • Increased nitrotyrosine production in patients undergoing abdominal aortic aneurysm repair

      Troxler, M.; Naseem, Khalid M.; Homer-Vanniasinkam, Shervanthi (Wiley, 2004)
      Vascular inflammation is implicated in the pathogenesis of atherosclerosis and abdominal aortic aneurysm (AAA), and is thought to involve reactive species such as the nitric oxide-derived oxidant peroxynitrite. In the present study nitrotyrosine was measured as a stable marker of peroxynitrite production in vivo. Perioperative blood samples were obtained from patients undergoing elective open or endovascular repair of an AAA and from patients with intermittent claudication, smoking aged-matched controls, non-smoking aged-matched controls and non-smoking young healthy controls. Plasma nitrotyrosine was measured by an enzyme-linked immunosorbent assay. The median plasma nitrotyrosine concentration in patients with an AAA (0·46 nmol nitrated bovine serum albumin equivalents per mg protein) was significantly higher than that in patients with intermittent claudication (0·35 nmol; P = 0·002), smoking controls (0·36 nmol; P = 0·001), non-smoking controls (0·35 nmol; P = 0·002) and young healthy controls (0·27 nmol; P < 0·001). Nitrotyrosine concentrations increased during early reperfusion in open AAA repair, but not during endovascular repair. AAA exclusion from the circulation reduced levels to control values (P = 0·001). Patients with an AAA had raised levels of circulating nitrated proteins compared with patients with claudication and controls, suggesting a greater degree of ongoing inflammation that was not related to smoking. Copyright
    • Ischaemic skeletal muscle increases serum ischaemia modified albumin.

      Troxler, M.; Thompson, D.; Homer-Vanniasinkam, Shervanthi (2009-11-02)
      Objectives Ischaemia modified albumin (IMA) has been used as a marker of myocardial ischaemia but little is known about its production during ischaemia of other tissues. The clinical models of patients with intermittent claudication and major arterial surgery were used to investigate IMA production from ischaemic skeletal muscle. Materials and methods IMA was measured pre-operatively, at end ischaemia, and 5min, 4, 24, 48, 72 and 144h post-surgery in patients undergoing (a) revascularisation for intermittent claudication (IC, n=15), (b) abdominal aortic aneurysm repair (AAA, n=12) and controls (n=16). Results The median pre-operative IMA concentration in IC patients was significantly higher than the AAA group (88.3 versus 83.5U/ml, p=0.036) and controls (88.3 versus 80.3U/ml, p=0.031). IMA concentrations increased significantly during arterial clamping in both IC and AAA groups (88.3 versus 120.0U/ml, p=0.001; 83.5 versus 118.8U/ml, p=0.002, respectively) consistent with increased skeletal muscle ischaemia. In contrast, there was only a mild perioperative increase in the controls (80.3 versus 91.6U/ml, p=0.012). Conclusions Patients with intermittent claudication have significantly elevated IMA and skeletal muscle ischaemia during arterial surgery results in significantly increased circulating IMA. When IMA is used to detect myocardial ischaemia, ischaemic skeletal muscle must be excluded.