Fig 1.
Number of trophoblast nuclei in each zone of the mouse placenta as determined using stereology.
(A) Absolute number of trophoblast cell nuclei and total placenta volume. (B) Distribution of trophoblast cell nuclei among different layers of the placenta. (C) Number of trophoblast nuclei within the labyrinth layer (blue bars), as well as labyrinth volume (purple line) and trophoblast cell volume (orange line) from E12.5 to E18.5. (D) Trophoblast cell volume calculated as total trophoblast cell volume in the labyrinth (from C) divided by the number of trophoblast nuclei from E12.5 to E18.5. Different letter labels indicate significant differences between days (p<0.05) and are valid for both volume and total trophoblast number (n = 3).
Table 1.
Percentage of trophoblast cells undergoing the cell cycle between E8.5 and 18.5.
Data represent mean and SEM is in parentheses.
Fig 2.
Ki67 and Phospho-histone H3 immunofluorescence staining in P-TGCs, junctional zone and labyrinth.
Histological sections were dual stained for Ki67/K18 (red/green respectively) or phospho-histone H3/K18 (red/green respectively) and nuclei are counterstained with DAPI (blue). Phospho-histone H3 was detectable at E12.5 in sinusoidal-TGCs within the labyrinth (white arrow; x400 magnification in the inset) but not in SynT cells (red arrow). MBS–maternal blood space; FBS–fetal blood space.
Fig 3.
Weights of fetuses and placentas, and fetal/placental weight ratios, in control and dietary protein restricted pregnancies.
(A) Fetal weights separated by sex and embryonic day (E). (B) Placental weights separated by sex and embryonic day. (C) Fetal to placental weight ratio (F:P) separated by sex and embryonic day. Explanatory variables–diet, sex, litter; Co-variates–maternal weight, litter size. Letter labels denote statistically significant main effects of diet within days (p<0.05). Main effects of sex (if present) are denoted by symbols at respective days (p<0.05). No diet by sex interactions were present. Number of conceptuses in each group (from left to right): E13.5–28, 32, 41, 39; E16.5–24, 37, 26, 24; E18.5–23, 42, 19, 32.
Fig 4.
Labyrinth growth in low protein pregnancies is mediated by trophoblast hypertrophy and nutrient transport function.
(A) Total labyrinth area separated by diet and sex (n = 4–5). (B) Glucose and amino acid transporter expression in male and female placentas. Fold change in protein restricted placentas (bars) relative to control diet placental expression (dotted line). * p<0.05; ** p < 0.01 (n = 3).
Table 2.
Labyrinth composition expressed as a percent area.
(mean; SEM in parenthesis).
Fig 5.
RNA sequencing analysis of Con and LP placentas at E16.5.
(A) Principal component analysis with different points representing different litter (number) and sex within litter (male versus female). (B) Heat map of differentially expressed genes. (C) Gene network diagrams produced by GeneMania program, showing relationships between genes that are up- (red) or down-regulated (blue) in LP placentas (n = 5).
Fig 6.
Spongiotrophoblast (SpT) cells contribute to the reduction in junctional zone size due to changes in cell size and number.
(A) Junctional zone area. (B) Total SpT cell area was measured in Image J using automated thresholding, distinguishing SpT cells by expression of Prl8a8 mRNA. (C) Spongiotrophoblast cell number (n = 4–5).
Fig 7.
Sinusoidal trophoblast giant cell (S-TGC) and spongiotrophoblast (SpT) cell ploidy in control and protein-restricted pregnancies.
(A) Frequency of S-TGCs at each ploidy level across gestation. (B) Frequency of SpT cells at each ploidy level across gestation (n = 6).