Quantification of placental change in mouse models of preeclampsia using magnetic resonance microscopy

Abstract

Abnormal development of the placenta is postulated to be central to the aetiology of preeclampsia. This study investigates changes in placental histopathology in mouse models of preeclampsia compared to the morphology using magnetic resonance microscopy (MRM) (11.7 T) of intact ex vivo tissue followed by 3D analysis of the image data. Here, C57BL/6JArc pregnant mice were subject to either normal pregnancy (n=3), or to one of two experimental models of preeclampsia; TNF-α infusion (n=3) or reduced uterine perfusion pressure(RUPP) (n=3). Placental tissue was collected at gestational day (gd) 17, fixed in formalin and incubated with Magnavist™ contrast agent, and high resolution images (50 μm × 50 μm × 50 μm voxels) obtained by magnetic resonance imaging at 11.74 T. Visual segmentation into placental subregions and three dimensional (3D) reconstruction followed by volume analysis was performed with Amira™ 3D analysis software. The significance of differences between treatment groups in total and regional volumes was assessed. In a single placenta the volumes measure by standard histology were compared. Three placentas from each animal were imaged, segmented into anatomical regions and 3D reconstructions generated. Total placental volume, labyrinth and decidual volume were not significantly different between groups. The junctional zone volume was found to be significantly larger in the RUPP animals (18.5±1.5 mm3) compared to TNF-α infused animals (15.8±1.5) or control animals (15.0±0.7, P<0.01). However, the decidual/junctional zone volume was smaller in the TNF-a compared to control animals (P<0.05). Placental structural change in experimental models of preeclampsia is able to be visualized and quantified using MRM and 3-D analysis. These techniques could prove to be a powerful tool in examining changes in placental morphology.

Keywords: 3-D reconstruction.; MRI; Magnetic Resonance Microscopy; Placental volume; preeclampsia.

Figure 1.

A) Representative high resolution (50 × 50 × 50 μm voxels) GEFI image of a formalin fixed and Magnevist contrast agent infused gestational day 17 control placenta. B) Representative histological section immunostained with cytokeratin showing trophoblasts (brown) (100x magnification) (reproduced with permission from Bobek et al., PLoS One 2013;8:e59971). Regions identified in MRI image correspond to those in histological section; Labyrinth (L), Junctional zone (Jz) comprising spongiotrophoblast and glycogen trophoblast cells, the decidua (D) and the mesometrial triangle (mesometrium, Mt). Other features observed are central canal (CC) and spiral arteries (SA). Note that shrinkage of ~1.5 for the tissue processed for histology and the distortions of tissue especially the mesometrium.

Figure 2.

Representative GEFI image (A) of a single ‘virtual slice’ of a placenta from a TNF- infused animal and (B) overlay showing segmentation into placental regions as indicated. Mt, mesometrium; D, decidua; Jz, junctional zone; L, labyrinth; A, amnion. Segmentation was performed using the visualisation software (Amira™).

Figure 3.

Representative GEFI image of a normal (A,B), TNF- infused (C,D) and RUPP (E,F) mouse placenta showing overlay (B,D,F of the segmented labelled regions; mesometrium (yellow), decidua (blue), junctional zone (green), labyrinth (red), amnion (cyan).

Figure 4.

Representative 3D reconstruction of a placenta using segmented label data. Volren of the placenta, showing A) maternal surface view; B) foetal surface view; C) clipped sagittal view; D) clipped sagittal view with solid colours showing mesometrium (yellow), decidua (blue), junctional zone (green), amnion (blue) and the labyrinth remaining in the red density rendering. 3D volume rendering was performed using the visualisation software (Amira™).

Figure 5.

Comparison of ratios of placental region volumes in animals from different treatment groups. A) Labyrinth volume/ Junctional zone volume B) Decidua volume/ Junctional zone volume. Generalized Estimating Equation Modelling clustering placenta within animals (n=3) and animals within treatment groups (n=3) was used to evaluate the statistical differences in ratios. A logarithmic transformation was carried out on ratios prior to statistical analysis. *P<0.01.

Figure 6.

Validation of MRI approach compared to conventional stereology. Four placentas from the same animal were imaged using MRI and then processed into paraffin blocks and sectioned. Volumes estimated by conventional stereology are shown in solid black and volumes measured from the MRI images are shown with diagonal hatched lines. Measured volumes in paired placenta were compared using a paired samples t-test (*P<0.05).