Transferrin receptor expression in serum exosomes as a marker of regenerative anaemia in the horse

Abstract

Evaluation of erythrocyte regeneration in horses is challenging, as they do not release reticulocytes into the peripheral blood. This study investigated transferrin receptor 1 (TfR1) expression in exosomes as a noninvasive method of characterising the regenerative response in anaemic horses. To quantify TfR1 in ultraprecipitate of serum in horses before and after phlebotomy-induced anaemia, and to identify exosomes as the source of TfR1. The hypothesis was that serum exosomal TfR1 expression would increase during a regenerative response. Experimental model of anaemia. Six horses were phlebotomised to achieve a 25% decrease in packed cell volume. Transferrin receptor 1 quantity in exosomes was determined by western blot and relative densitometry before and after phlebotomy. The size and density of the TfR1-associated particles were confirmed by transmission electron microscopy and density gradient centrifugation, respectively. Regenerative anaemia was confirmed by decreased packed cell volumes and decreased myeloid:erythroid ratios in the bone marrow. In all 6 horses, TfR1 expression increased between Days 7 and 10. Mean TfR1 levels peaked on Day 10 and at 3-fold higher than levels on Day 0. Appropriately sized particles were evident on transmission electron microscopy and sucrose density gradient fractions expected to contain exosomes also contained TfR1. These data indicate that TfR1 expression in serum exosomes may provide a marker for regeneration in anaemic horses.