OBJECTIVE: To investigate whether extracellular phosphorylation of plasma proteins takes place in vivo in patients with systemic lupus erythematosus (SLE), to determine possible correlations between phosphate levels and clinical and/or laboratory parameters, and to identify individual phosphorylated plasma proteins.
METHODS: Sera from SLE patients were analyzed for total amounts of protein-bound phosphate by a colorimetric technique, and for levels of beta-thromboglobulin by radioimmunoassay. In addition, the ability of these sera to activate platelets, resulting in the release of protein kinase, was tested using an assay in which platelet-rich plasma from healthy blood donors was incubated with sera or immune complexes from SLE patients. In this assay, [gamma-32P]ATP was added, and 32P-labeled C3 was quantified. Phosphate in individual proteins was detected by Western blot analysis.
RESULTS: 32P-labeled, activated platelets were able to phosphorylate exogenously added proteins, without the addition of ATP or cations. Platelet-rich plasma from healthy blood donors became activated by sera or by polyethylene glycol-precipitated immune complexes from patients with SLE, which led to the extracellular phosphorylation of plasma proteins, exemplified in the C3 assay. The phosphate content in plasma proteins was increased in SLE patients with previous thrombosis. The degree of phosphorylation increased up to 3-fold in serial samples obtained from 2 SLE patients during periods of disease exacerbation. Substantial phosphate increases were seen in C3 and fibrinogen. The changes were linked to platelet activation because of the observed covariation with the levels of beta-thromboglobulin.
CONCLUSION: In SLE patients, the phosphate content in plasma proteins (including C3 and fibrinogen) increases due to platelet activation.
1997. Vol. 40, no 12, 2178-2186 p.