Promise of new imaging technologies for assessing ovarian function

Abstract

Advancements in imaging technologies over the last two decades have ushered a quiet revolution in research approaches to the study of ovarian structure and function. The most significant changes in our understanding of the ovary have resulted from the use of ultrasonography which has enabled sequential analyses in live animals. Computer-assisted image analysis and mathematical modeling of the dynamic changes within the ovary has permitted exciting new avenues of research with readily quantifiable endpoints. Spectral, color-flow and power Doppler imaging now facilitate physiologic interpretations of vascular dynamics over time. Similarly, magnetic resonance imaging (MRI) is emerging as a research tool in ovarian imaging. New technologies, such as three-dimensional ultrasonography and MRI, ultrasound-based biomicroscopy and synchrotron-based techniques each have the potential to enhance our real-time picture of ovarian function to the near-cellular level. Collectively, information available in ultrasonography, MRI, computer-assisted image analysis and mathematical modeling heralds a new era in our understanding of the basic processes of female and male reproduction.

Fig. 1

Demographics of publications on ultrasound-based ovarian research in human, cattle, horse and other domestic species over the last 8 years. Data were obtained by searching for keywords—(ultrasound or ultrasonography) and (ovary or follicle or corpus luteum) in Agricola, Medline and CAB databases for human, cattle and other species (sheep, goat, mare, camlids, pig). Based on the title and year of publication, number of publications for each species were categorized into (i) folliculodynamics and luteal development, (ii) disease, (iii) assisted reproduction/IVF/superstimulation and (iv) other studies.

Fig. 2

Ultrasound-guided follicle aspiration technique for calves. (a) Demonstrates the topography of the genital organs and position of the transducer for transvaginal ultrasonography and oocyte aspiration in 6 week-old calves in dorsal recumbency. (b) Ultrasound image of an ovarian follicle during the aspiration procedure. The image of the needle entering the anechoic follicle is clearly observed (modified with permission from Brogliatti and Adams, 1996).

Fig. 3

(A) To illustrate the sensitivity of computer-assisted data analysis, an excised bovine ovary containing a preovulatory (left) and a regressing (right) follicle was scanned in a degassed water bath using a high resolution ultrasound instrument. The two large follicles imaged appear visually indistinguishable (B) Clear differences in the follicle walls and antra can be detected by computer-assisted analyses. In a three-dimensional representation of the brightness of the pixels comprising the ultrasound image, the echotexture of the antrum appears more heterogeneous and the pixels of the follicular wall are brighter and drop off more sharply in the follicle on the right. Image attributes were consistent with the notion that the follicle on the left was the dominant follicle of a new wave, while the follicle on the right was the dominant follicle of a previous wave (modified with permission from Adams and Pierson, 1995).

Fig. 4

(A, B) Computer-assisted analysis of ultrasound image of an ovarian follicle. Portion of the follicle wall from part A is enlarged in part B to display the picture-elements (pixels) forming the ultrasound image. Spot-analysis of the antrum is performed to measure the pixel value (black = 0, white = 255) and pixel heterogeneity by placing the measuring circle at four different locations (a, b, c, d) over the follicle antrum to cover approximately 20% of the area in each quadrant. (C) Line-analysis of the peripheral antrum, follicle wall and stroma was performed by placing a computer-generated line on the ultrasonographic image at 10 and 2 o’clock positions (e, f on part A, e on part B) and plotting the gray-scale values of pixels along the line. The pixel value graph was divided into three segments (peripheral antrum, follicle wall, stroma). The antrum-wall interface was used as a reference point. (D) Pixel value (PV), pixel heterogeneity (Hetero), and the intercept and slope of the regression line of each segment is recorded separately (modified with permission from Singh et al., 1998).

Fig. 5

Regional analysis of a single human preovulatory follicle. (A) Image of the follicle with follicle wall identified (black line). (B) Wire-frame model of the pixel by pixel mesh created from the image of the follicle. (C) Computer generated “skin” stretched over the wire-frame model. (D) Height shaded color algorithm added to the image to enhance visual appreciation and allow comparison of different zones with images of different follicles or images of the same follicle on different days.

Fig. 6

Time-series analysis of a dominant anovulatory follicle. Individual follicles are identified (top three panels) on a daily basis and a graphic line representing their numerical values created (second three panels). The numerical values are concatenated and a wire-mesh framework created (lower left panel). The X dimension represents the diameter of the follicle including the follicle wall, the Y dimension represents the numerical pixel value, and the Z dimension represents the days of the cycle on which the follicle was uniquely identified. A surface rendering (skin) is placed over the framework to yield a conceptualization of the physiologic status of the follicle (lower right panel) from the time that the follicle was first identified, completes its growth phase, enters and passes through a static phase, and then regressed until it could no longer be identified. Addition of color shading algorithms adds to greatly enhanced visual perception.

Fig. 7

Three dimensional parametric space model of ovarian follicular development. Maturity, m, of the follicles was equated to diameter. The rate of growth of the follicle z was determined by using central finite differences from the ultrasonographic time series data. The maturation rate (pg E₂ per hour calculated based on daily blood samples, ξ reflects circulating E₂ and ξ_i reflects maturity. SF represents the spatial area of subordinant follicles, DOF that occupied by dominant ovulatory follicles and DAF the area associated with dominant anovulatory follicles. The data were then plotted in the three dimensional (z, m, E₂) parametric space and interpolated to yield a mathematical maturation surface. We are interested in modifying this model to include image attributes, like echotexture and follicle size, and the effects of pathology and pharmacologic intervention which will yield interpretive models. On the surface framework, the line to the left illustrates the developmental history of a dominant ovulatory follicle while the line to the right illustrates the history of an dominant anovulatory follicle (modified from Sarty and Pierson, 1998).

Fig. 8

(a) Intercept (circles) and slope of regression lines obtained by line-analysis of the follicle wall of the dominant and subordinate bovine follicles. Values with no common letters indicate significantly different values (P < 0.05) in intercept (A, B, C, D, E) and slope (U, V, W, X, Y, Z) (modified from Singh et al., 1998). (b) Diameter profiles and associated echotexture characteristics of dominant anovulatory and ovulatory follicles in cattle (mean ± S.E.M.; n = 15 observations per day). The growing (days 0–6) and regressing phases (more than day 12) of the dominant anovulatory follicle are separated by a static phase (days 6–12). Values with no common superscripts indicate significant difference (P = 0.05) (modified from Tom et al., 1998).

Fig. 9

Reconstructed 3D ultrasonographic images of a bovine ovary with cutout portions (a–d) to expose the topographic locations of ovarian follicles and corpus luteum. Different transparency levels (d–f) allow the ‘dissolution’ of various structures.

Fig. 10

Color flow (A, C) and power flow Doppler ultrasound images of imminently preovulatory follicles (A, B) demonstrating directional peri-follicular flow and recent sites of ovulation (C, D) demonstrating the vascular patterns around walls of the recently collapsed follicle/developing corpus luteum. In color flow images, blood flow toward the transducer is displayed in red, while flow away from the transducer is displayed in blue. In power flow images, all flow is displayed in a graded variant of yellow to orange colors.

Fig. 11

A. High resolution image of an ovarian follicle of an anesthetized mouse obtained by transabdominal scanning with the ultrasound biomicroscope. The resolution obtained is far greater than that obtained in Fig. 3A. Compared to the image of bovine follicle (Fig. 3A), the mouse follicle was much smaller (2.4 mm vs. 15 mm) and was imaged in situ (live) rather than in vitro (excised ovary in a water bath). The cumulus-oocyte-complex (arrowhead) is visible in the center of the follicle. (B) Region analysis of the ultrasound image in A was performed and never-before seen details of the follicle wall (arrows) and the cumulus–oocyte-complex (arrowhead) are clearly evident.

Fig. 12

MRI images of an ex vivo bovine ovary in two A, B and three C, D dimensions. Two-dimensional T₁- (A) and T₂-weighted images demonstrating follicles in different stages of atresia (a, A, b, B), the physiologically dominant follicle (c, C) and a corpus albicans (d, D). Small follicle of 2 mm diameter were easily visualized. Three dimensional images created using FISP and PFISP imaging. Follicles and luteal structures are clearly identifiable in all views. The images may be rotated 360° in the computer. Adapted from Hilton et al., 2000 and Sarty et al., 2000.

Fig. 13

Diffraction enhanced images (DEI) of a bovine ovary produced with X-rays on beamline ×15A of the National Synchrotron Light Source, Brookhaven National Laboratory, Upton, NY. Three antral follicles approximately 4 mm in diameter are indicated on the left, and a corpus luteum is outlined on the right. The images were acquired on opposite sides of the diffraction rocking curve, resulting in opposing contrast. Artifact was produced by a paper clip (at top) used to suspend the ovary in a plexiglass water bath (vertical striations).