Differential effects of divalent cations on fibrin structure

Calcium, a divalent cation, serves as a cofactor for several reactions of the haemostatic cascade and causes concentration-dependent enhancement of fibrin fibre size and fibrin structure. We have evaluated the ability of magnesium and zinc to influence fibrin assembly and structure. 20 mM magnesium shortened the lag phase prior to turbidity development and increased the fibre mass/length ratio (0.28 to 1.23 × 10 Da/cm) in a purified fibrin-thrombin system. In plasma clotted with thrombin, 20 mM magnesium increased fibre mass/ length ratios from 0.60 to 1.65 × 10 Da/cm. Fibrinopeptide B release was not required for expression of magnesiumʼs effect since lag phase shortening and increased fibre mass/length ratios occurred when clotting was initiated with batroxobin. In all cases, magnesium effects were similar to effects seen with calcium. In the purified system, zinc exerted effects at concentrations as low as 5 μM. At 10 μM, zinc shortened the period prior to turbidity rise from 15.3 to 2.3 s. The final fibre size increased from 0.30 to 1.16 × 10 Da/cm over the same concentration range. Zinc caused similar alterations in gels clotted with batroxobin. The lag phase shortened from 38.3 to 15.3 s, and the fibre mass/length ratio increased from 0.49 to 1.47 × 10 Da/cm as zinc concentration increased from 0 to 10 μM. The addition of zinc to citrated plasma at concentrations as high as 100 μM had no appreciable impact on fibrin assembly or structure. To avoid possible effects of citrate, zinc was added to fresh non-anticoagulated plasma. At concentrations as high as 100 μM, zinc had no effect on thrombin-induced clot formation or structure. In citrated plasma, very high zinc concentrations (500 μM) increased the fibre mass/length ratio by 40% and shortened the lag phase by 50%. Thus, except at very high concentrations, zinc has minimal effects in plasma. This result may be mediated by binding of zinc to plasma proteins such as albumin.