Chasing electrons : exploring the redox balance of platelet functions
The platelet redox environment consists of the blood plasma and within it the lowmolecular weight thiols glutathione and cysteine. Both glutathione and cysteine exist as independent redox couples comprising carefully controlled ratios of the reduced (GSH, Cys) and oxidised (GSSG, CySS) form of each thiol. These ratios yield a GSH/GSSG redox potential and a Cys/CySS redox potential in the plasma. Changes in the redox status of the plasma are manifested by alterations in either the glutathione or cysteine redox potential and are associated with the progression of a large number of disease states including cancer, atherosclerosis and schizophrenia. In addition, oxidative changes in either of the plasma redox potentials have been directly linked with risk factors for cardiovascular disease. The post-translational modification of critical cysteine thiol groups on proteins is linked to such alterations in both the GSH/GSSG and Cys/CySS redox potentials and occurs extracellularly and intracellularly. Platelet integrins in particular are prime targets for redox modification due to their high cysteine content. Although the role of thiol/ disulphide bond exchange in platelet activation is established the effects of a changing redox environment on platelet reactivity are unclear. The aims of this thesis therefore, are to explore the impact and consequences of such changes on platelet function.
Platelet activation was significantly diminished in the presence of either reducing GSH/GSSG or Cys/CySS redox potentials. Moreover, this response was most notable when platelets were activated with collagen only. The redox modulated effects were shown to be specific to the integrin ɑ2β1, through a mechanism involving disulphide bond reduction. An analysis of the platelet integrin β subunits revealed the presence of the greatest number of potentially reactive cysteine residues within the Pi subunit when compared to the other platelet integrin beta subunit β3.
Furthermore, platelet proteins were S-glutathionylated in a reducing extracellular redox environment. The findings of this thesis reveal for the first time how exquisitely sensitive collagen-induced platelet activation is to alterations in the external redox environment. This, in fact, is potentially a novel demonstration of reductive stress regulating platelet reactivity and provides a novel avenue for therapeutic intervention using physiological redox reagents or their precursors.