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. 2018 Mar 1;13(3):e0193482.
doi: 10.1371/journal.pone.0193482. eCollection 2018.

The effect of increasing the sulfation level of chondroitin sulfate on anticoagulant specific activity and activation of the kinin system

J Hogwood  1   2 , A Naggi  3 , G Torri  3 , C Page  2 , P Rigsby  1 , B Mulloy  2 , E Gray  1   2
Affiliations

The effect of increasing the sulfation level of chondroitin sulfate on anticoagulant specific activity and activation of the kinin system

J Hogwood et al. PLoS One. .

Abstract

Oversulfated chondroitin sulfate (OSCS) was identified as a contaminant in certain heparin preparations as the cause of adverse reactions in patients. OSCS was found to possess both plasma anticoagulant activity and the ability to activate prekallikrein to kallikrein. Differentially sulfated chondroitin sulfates were prepared by synthetic modification of chondroitin sulfate and were compared to the activity of OSCS purified from contaminated heparin. Whilst chondroitin sulfate was found to have minimal anticoagulant activity, increasing sulfation levels produced an anticoagulant response which we directly show for the first time is mediated through heparin cofactor II. However, the tetra-sulfated preparations did not possess any higher anticoagulant activity than several tri-sulfated variants, and also had lower heparin cofactor II mediated activity. Activation of prekallikrein was concentration dependent for all samples, and broadly increased with the degree of sulfation, though the di-sulfated preparation was able to form more kallikrein than some of the tri-sulfated preparations. The ability of the samples to activate the kinin system, as measured by bradykinin, was observed to be through kallikrein generation. These results show that whilst an increase in sulfation of chondroitin sulfate did cause an increase in anticoagulant activity and activation of the kinin system, there may be subtler structural interactions other than sulfation at play given the different responses observed.

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Conflict of interest statement

Competing Interests: The authors have declared that no competing interests exist.

Figures

Fig 1
Fig 1. Kallikrein generation in normal pooled plasma by synthetic and purified OSCSs, native chondroitin sulfate, dextran sulfate and unfractionated heparin.
Error bars = standard deviations (SD), n = 6, note lines for native CS, unfractionated heparin and plasma alone are overlapping. Inset chart shows line (….) indicating Δ0.2 OD above baseline used to select concentration of samples for the bradykinin assay.
Fig 2
Fig 2. Kallikrein and Bradykinin levels obtained from a defined quantity of synthetic and purified OSCS, native chondroitin sulfate, dextran sulfate and unfractionated heparin.
Different amounts of each material, predicted to generate the same level of (A) kallikrein as determined by optical density increase and (B) bradykinin, were used (see Table 4). No statistical difference was observed between the samples (oneway ANOVA, p = 0.883 and 0.568 respectively). CS-A and UFH were included at 100μg/ml, the highest level used. Error bars = standard deviation; n = 6.
Fig 3
Fig 3. Antithrombin titration response for synthetic OSCS, native chondroitin sulfate and a heparin sample.
The increase in fluorescence (ex280nm em340nm) response measured from antithrombin with increasing concentrations of each sample. Error bar is SD, n = 6.
See this image and copyright information in PMC

References

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