Confocal laser endomicroscopy: exploring endometrial image quality in relation to the oestrous cycle of mares.

Overview

• This study investigated how the quality of images obtained from confocal laser endomicroscopy (CLE) of the mare’s endometrium varies with the phases of the oestrous cycle.
• The goal was to understand the best timing for CLE imaging to ensure optimal image quality for assessing uterine health in mares.

Background and Importance

• Endometrial assessment is a key part of evaluating breeding soundness in mares.
• The gold standard has been histopathology of a biopsy, but a single biopsy can lead to inconsistent results due to sampling limitations.
• Confocal laser endomicroscopy (CLE) offers a non-invasive way to visualize the uterine lining in real-time at multiple sites, potentially overcoming sampling issues.
• Previous research showed variability in CLE image quality between individual mares, but reasons for this variability were not clear.

Study Objective

• To compare and analyze the quality of CLE images obtained during different phases of the oestrous cycle—specifically dioestrus and oestrus—in the same mares.
• To understand whether the timing within the cycle affects the ability to get high-quality endometrial images using CLE.

Methods

• Subjects: 24 mares underwent CLE imaging twice—once during dioestrus and once during oestrus.
• Image Evaluation: A 4-point grading scale for image quality was created.
• Blinding: Four independent examiners, blinded to the cycle phase and mare identity, graded the images.
• Statistical Analysis: Agreement among raters was analyzed by calculating the intraclass correlation coefficient (ICC).
• Additional Analysis: The study assessed how image depth (luminal epithelium vs subluminal layers) and stage of the cycle affected image quality.
• Oedema scores, reflecting fluid accumulation associated with the cycle phase, were also measured to see their correlation with image quality.

Key Findings

• There was moderate to good agreement among the four examiners in image quality scoring (ICC = 0.702), confirming the grading system’s reliability.
• Image quality was statistically significantly lower when images were taken at the luminal epithelium layer compared to subluminal layers (average scores: 2.46 vs 2.68).
• CLE images obtained during oestrus had lower quality scores compared to images collected during dioestrus (2.47 vs 2.67).
• There was a negative correlation between image quality and endometrial oedema scores, meaning that higher oedema (common in oestrus) reduced image quality.

Interpretation and Implications

• Endometrial oedema, which peaks during oestrus, appears to reduce the clarity of CLE images, possibly due to increased fluid obscuring microscopic detail.
• The subluminal imaging depth yields clearer images, suggesting that targeting deeper tissue layers may improve diagnostic capability.
• Timing CLE examinations during phases when endometrial oedema is minimal (e.g., dioestrus) results in better image quality and potentially more accurate assessment of uterine health.
• This information can guide veterinarians on the optimal timing for using CLE in breeding soundness exams to avoid poor image quality caused by physiological changes during oestrus.

Conclusions

• CLE is a promising non-invasive tool for in-vivo uterine imaging in mares, but its image quality is influenced by physiological changes in the oestrous cycle.
• The phase of the cycle and image depth should be considered when performing CLE to maximize image clarity and diagnostic usefulness.
• Practically, scheduling CLE examinations outside periods of maximal endometrial oedema (i.e., avoiding peak oestrus) is recommended for improved outcomes.