LAMELLAR HISTOLOGICAL AND IMMUNOHISTOCHEMICAL EVALUATION OF THE HOOF WALL IN HORSES WITH CHRONIC LAMINITIS TREATED WITH MESENQUIMAL STEM CELLS DERIVED FROM ADIPOSE TISSUE BY REGIONAL PERFUSION

Abstract

Background The objective was to evaluate the histological and immunohistochemical effects of regional perfusion of characterized mesenchymal stem cells derived from adipose tissue (Ad-MSC) in biopsy hoof samples from horses with laminitis. Methods Six mares with stable chronic laminitis in standard treatment for distal phalangeal realignment were subjected to regional perfusion of the digital vein with Ad-MSC (20 million cells, 3 infusions every 15 days) in one forelimb compared to placebo infusion in the contralateral one. Evaluations occurred in two 30-day periods, before and after treatment (from the second infusion). Dorsal hoof lamellar biopsies were taken 30 days before and 15 days after the therapeutic protocols. Histological 5-μm cuts stained with H&E and PAS were classified based on a crescent 1-5 scale lesion score system. The sum of each lesion grade represented the global lesion score. Immunohistochemistry for p63 positive cells were also performed in formalin-fixed samples. Results In the placebo limbs, there were increases over time in following histological variables: global lesion (P=0.01), keratinized axis loss (P=0.0001), stratum medium hyperplasia (P=0.001), primary epidermal lamellae homogeneity loss (P=0.003), and hyperkeratosis (P=0.05). Only the stratum medium hyperplasia score was increased in the Ad-MSC-treated hooves. P63-positive cells could be detected in all formalin-fixed paraffin-embedded samples, but background unspecific stain made counts imprecise to be made. Conclusion In conclusion Ad-MSC regional vascular perfusion prevented histological lesion progression in the lamellar tissue of horses with chronic laminitis. Our results confirm the therapeutic potential of Ad-MSC in the treatment of horses with stable chronic laminitis.