STRUCTURAL AND FUNCTIONAL-CHARACTERIZATION OF PLASMA FIBRONECTIN IN PATIENTS WITH ESSENTIAL MIXED CRYOGLOBULINEMIA

Experimental studies suggest that plasma fibronectin may be involved in the cryoprecipitation of cryoglobulins in essential mixed cryoglobulinaemia; reduced plasma concentrations of the glycoprotein have been shown in the disease. The present work was undertaken in order to verify this latter finding and to detect a possible structural alteration of plasma fibronectin as result of enzymatic digestion of the molecule in vivo. This could, in turn, induce a decreased reactivity of the protein in immunometric assays and a reduced opsonic activity, which is normally due to the affinity of fibronectin to the Clq component of complement. Moreover, since a polymorphic variant of fibronectin has been described in plasma during experimental vascular injury and in patients with autoimmune vascular diseases, the aim of this study was also to verify the presence of a polymorphism of the glycoprotein in cryoglobulinaemic vasculitis. Twenty seven patients with essential mixed cryoglobulinaemia and 26 normal subjects were included in the study. Significantly reduced concentrations of plasma fibronectin, as assessed by ELISA, were found in patients when compared with controls (231.7 +/- 15.3 vs 316.1 +/- 16.6 mg/l, P < 0.0002). In contrast, when affinity-purified plasma fibronectin from 10 patients with essential mixed cryoglobulinaemia and 8 healthy subjects were analysed by western blotting, employing a panel of five monoclonal antibodies to different regions of the molecule, no differences were observed between patients and controls, suggesting integrity of the glycoprotein in the disease. Moreover, plasma fibronectin from cryoglobulinaemic patients and normal subjects bound to solid phase C1q in a dose-dependent manner with identical efficiency in the two groups, further suggesting that the molecule is functionally and structurally unaltered in the disease. The production of an abnormally glycosylated form of fibronectin in patients with essential mixed cryoglobulinaemia also seems to be excluded, as SDS-PAGE revealed no differences in electrophoretic mobility and apparent molecular weight between fibronectin from patients and controls. Taken together these data are consistent with the hypothesis that plasma fibronectin concentrations are actually reduced in essential mixed cryoglobulinaemia possibly by consumption during cryoprecipitate formation, and that a polymorphic form of the protein is not released into the circulation during cryoglobulinaemic vasculitis.