Histomorphometric characterization of the endometrium in mules (Equus mulus): An approach to endometritis/endometrosis.

Overview

• This study investigates the structure and gene expression of the uterine lining (endometrium) in adult mules, which are sterile hybrids of mares and donkeys.
• It uses detailed tissue analysis and molecular methods to understand unique aspects of mule uterine biology and early signs of uterine disease.

Background and Objectives

• Mules are sterile hybrids resulting from mating a horse mare and a donkey stallion, combining physiological traits of both species.
• The reproductive organs of mules, specifically the endometrium, are not well understood despite their unique hybrid nature.
• This study aims to characterize the mule endometrium by:
  • Examining the tissue structure in detail (histomorphometry).
  • Measuring gene expression related to immune response and fibrosis (scarring).
  • Classifying tissue health using a standard veterinary grading system (Kenney-Doig system).

Methods

• Endometrial biopsies were collected from adult mules for analysis.
• Quantitative stereology was used to measure the volume proportions of different layers of the uterine lining:
  • Stratum spongiosum (a stromal layer)
  • Epithelium (lining cells)
  • Compact layer
• Histological classification following the Kenney-Doig system graded tissue health and degenerative changes.
• Quantitative PCR (qPCR) assessed the expression of immune-related genes (IL1B, IL6, TNFα) and fibrosis-related genes to detect inflammation and tissue remodeling.

Key Findings

• Structural Characteristics:
  • The stratum spongiosum constituted the majority (~84.7%) of the mule endometrium volume, which is higher than in mares.
  • This suggests increased stromal expansion or fluid accumulation (edema) in mules.
  • The proportion of epithelial and compact layers was smaller compared to horses, indicating a distinct endometrial architecture.
  • Endometrial glands were abundant, contributing to an intermediate phenotype with more features resembling donkeys (“asinine bias”).
• Tissue Health and Degeneration:
  • Most samples were graded as I, IIA, or IIB, indicating mild to moderate endometrial remodeling but no severe pathology (grade III was absent).
  • Tissue architecture was largely preserved despite signs of early degeneration.
  • Expression of pro-inflammatory cytokine genes (IL1B, IL6, TNFα) increased with higher degeneration grades, supporting presence of mild inflammation.
• Implications for Hybrid Physiology:
  • Mules have limited hormonal cycling compared to mares and donkeys, which may reduce progression from early inflammation and degeneration to severe fibrosis.
  • The unique hybrid nature leads to distinct uterine tissue features that differ somewhat from parent species but are still comparable.

Conclusions and Significance

• The study confirms that the Kenney-Doig system is useful for evaluating mule endometrial health but suggests refinement (especially for grade IIC) to better reflect hybrid-specific changes.
• The mule endometrium has identifiable structural and molecular differences that reflect its hybrid origin, including greater stromal dominance and intermediate glandular patterns.
• These findings provide a baseline for understanding uterine health in mules, which can guide clinical evaluations and interpretations of fertility-related issues in this hybrid species.
• The mule uterus may serve as a valuable comparative model for studying reproductive biology and endometrial disease in equids, offering insights into hybrid fertility and uterine pathology.
• Future research can build on this foundation to explore mechanisms underlying metabolic and immune adaptation in hybrid reproductive tissues and to improve veterinary care.