Suitability of antral follicle counts and computer-assisted analysis of ultrasonographic and magnetic resonance images for estimating follicular reserve in porcine, ovine and bovine ovaries ex situ

Abstract

This study was conducted to determine if correlations exist between the numbers of microscopic follicles comprising ovarian follicular reserve (OFR) and antral follicle counts (AFCs), and to assess the usefulness of computerized analyses of ovarian ultrasonograms and magnetic resonance (MR) images for estimating OFR in excised porcine, ovine and bovine ovaries. As a pre-requisite to these analyses, we characterized and compared ovarian cortical histomorphology and follicle populations in the three species varying in prolificacy and overall reproductive longevity, and hence the total number of microscopic and antral follicles. Ultrasonographic and MR images were obtained at the scanner settings optimized to provide opposing contrasts between antral follicles and the ovarian stroma. Commercially available ImageProPlus® analytical software was used to calculate numerical pixel values (NPVs) and pixel heterogeneity (standard deviation of the pixel values) along the computer-generated lines (4-6) placed in the area corresponding to the ovarian cortex. The numbers of primordial (r = 0.38, P < 0.01) and intermediate follicles (r = 0.37, P < 0.01) were correlated with the numbers of antral follicles in bovine ovarian sections. The numbers of primordial (r = 0.28, P < 0.05), intermediate (r = 0.31, P < 0.01) and primary follicles (r = 0.27, P < 0.05) correlated directly with mean NPVs of the ultrasonographic ovarian images in cattle. There was a negative correlation between primary follicle numbers and NPVs of MR images (3D FAST-SPOILED GRADIENT ECHO) of the porcine ovarian cortex (r = -0.31, P < 0.05). To summarize, the numbers of primordial and intermediate follicles could only be estimated from AFCs in cows. Using ultrasound NPVs, the numbers of primordial, intermediate and primary follicles could be directly estimated in bovine ovaries and the quantitative image attributes of MR images were useful for quantifying porcine primary follicles. The bovine ovarian model is compatible with human situation and hence future studies should be undertaken to ascertain the usefulness of AFCs and ultrasonographic image analyses for estimating OFR in women.

Keywords: Ovarian follicle reserve; digital image analysis; magnetic resonance imaging; ovarian histology; ultrasonography.

Figure 1

(a) A diagram illustrating the topographic location of cross-sections (two from near-middle (NM) and two from near-pole (NP) regions of each ovary) used for comparative histological assessment and computer-assisted image analyses of the ovarian cortex; (b) demonstration of line placement (4–6 lines per cross-section) for image analysis using the Image ProPlus® analytical software. Computer-generated lines with the length corresponding to the area occupied by ovarian cortex (on the basis of earlier histological examinations of ovarian histograms) were drawn at approximately 10, 12, 2, 4, 6, and 8 o’clock positions. A special care was taken to avoid visible antral follicles and, in the ultrasonographic images, the lines at 12 and 6 o’clock positions were frequently omitted due to the presence of reflection artifacts

Figure 2

Histomorphological classification and photographic reproductions of porcine, ovine and bovine follicles at 100× (dark gray background) or 40× image magnification (light gray background).