Abstract
Characterisation of the physiologic equine amniotic fluid (AF) proteome is a prerequisite to study its changes during diseases and discover new biomarkers. The aim of this study was to identify by a proteomic approach the most abundant proteins of equine AF. AF samples were collected at parturition from 24 healthy mares that delivered healthy foals. All samples were subjected to sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) on 4–12% gels. A pool of the 24 samples, after SDS-PAGE, was cut in 25 slices, trypsin-digested and analysed by mass spectrometry (MS) for protein identification. Mean AF protein concentration was 1.96±1.12g/L. Thirty-four proteins were successfully identified by MS and subsequently categorised according to Gene Ontology (GO). Twelve proteins (e.g. fibronectin, lumican, thrombospondin and fibulin) belonged to or interacted with the extracellular matrix (ECM) playing an important role in the development of foetal tissues. Most of the remaining proteins were classified as transport (e.g. albumin, major allergen Equ c1 and alpha-fetoprotein) delivering nutrients, ions and lipids essential for foetal growth and development. Among these proteins, major allergen Equ c1 is widely studied in human medicine because it induces Ig-E mediated type I allergic reaction. The absence of immunoglobulins in equine AF was also confirmed.
Highlights
Amniotic fluid (AF) is a complex dynamic milieu that changes as pregnancy progresses
All samples presented a similar pattern characterised by two clusters of bands: the first with molecular weights (MW) higher than 62 kDa and the second with MW lower than 34 kDa
Out of the 25 slices cut from the gel (Figure 3), 20 yielded significant results leading to the unambiguous identification of 34 proteins (Table 2)
Summary
Amniotic fluid (AF) is a complex dynamic milieu that changes as pregnancy progresses. AF contains nutrients and growth factors that facilitate foetal growth, provides mechanical cushioning and antimicrobial effectors to protect the foetus and allows assessment of foetal maturity and disease (Underwood et al 2005). The physiology and pathophysiology of foetal fluids in domestic mammals are poorly understood (Canisso et al 2015). Some studies have investigated biochemical composition, enzymes and electrolytes, in AF collected by ultrasound-guided transabdominal amniocentesis, at delivery or at slaughter (Holdstock et al 1995; Lyle et al 2006; Williams et al 1993; Zanella et al 2014). AF was studied for evaluation of foal’s lung maturity at birth through lecithin/sphingomyelin ratio and lamellar body count (Castagnetti et al 2007).
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