2.9. Hematoxylin and eosin staining of intestinal tissue

The horses utilized in this study were primarily intended for meat production, hence, at the end of the experiment, they were humanely euthanized by the ranch owner. Post-mortem, intestinal tissues were promptly harvested within an hour. From each horse, a 10 × 10 × 0.5 mm tissue sample (sourced from the colon) was dissected and segmented into 55 mm-square fragments, which were then preserved in 4% formalin for a period between 24 to 72 h. This was followed by routine processing and embedding in paraffin wax. A standardized procedure was followed to ensure that all tissue samples were collected from the same anatomical location across all horses. Hematoxylin and Eosin (H&E) staining methods were employed, which were subsequently analyzed under a standard light microscope. These H&E-stained slides served dual purposes: the detection of intestinal lesions and the performance of morphometric analysis. All visual observations were digitally captured using a specialized image capture software coupled with a microscope-integrated camera (Olympus, Japan).