Fibronectin fragments cause an underestimation of plasma fibronectin levels in severe pre-eclampsia

Abstract

Elevated fibronectin (Fn) concentrations in plasma from pregnant women have been suggested to be predictive for the development of pre-eclampsia. However, reported correlations between Fn concentration with gestational age or disease severity appear to be confounded by several variables. Our finding of degradation products of Fn in plasma from patients with severe pre-eclampsia led to a study of whether their presence could influence immunological and functional measurements of intact plasma Fn. Plasma samples (10 control pregnant and 10 severe pre-eclampsia patients) were studied for the presence of intact Fn and fragments by SDS-PAGE and Western blotting. Percentages of immunoreactive Fn fragmentation in control plasma (9.37 ± 6.7%) and severe pre-eclampsia plasma (62.57 ± 7.0%) were determined by densitometric scanning. Immunoassays by ELTSA were performed on normal plasma samples and on purified Fn in the absence or presence of plasma digests of pure Fn. Artefactual underestimations of Fn concentration occurred in the presence of Fn fragments. The percentage underestimation increased with increasing digestion times (1 h, 46.52 ± 3.69%; 3 h, 77.5 ± 7.18%; 4 h, 10%) and with increasing concentrations of Fn digest products (1h) added to normal plasma samples (n=10) (40 μg, 23 ± 8.1%; 125 μg, 36.33 ± 5.1%; 250 μg, 43.66 ± 6.5%). The underestimation of Fn concentration in the presence of Fn fragments suggests that ELISAs for Fn may be unreliable and thus lose their predictive value in these patients. Opsonic activities of control pregnant and severe pre-eclamptic plasma were determined by gelatin-latex agglutination assay. The ratios of opsonic activity to Fn concentration in plasma from patients with pre-eclampsia were significantly lower than those of control pregnant plasma samples (p<0.008). The impaired gelatin-latex agglutination of Fn in pre-eclamptic plasma suggests that Fn fragmentation could have biological sequelae in vivo.

Key Words:

• ELISA
• fibronectin concentration
• gelatin—latex agglutination
• opsonic activity