Fig. 2.

Effects of maternal gut microbiome and B. breve supplementation during pregnancy on placental structure on day 16.5 of gestation. A Placenta weight. B Placental efficiency determined by dividing fetal by placental mass. C Placental regional analysis. Scale bar = 1 mm. D Representative staining of placental glycogen with PAS and glycogen abundance. Scale bar = 2.5 mm and 250 μm. E Representative image of lectin and cytokeratin staining for labyrinth zone structural quantification. Scale bar = 500 μm and 50 μm. F–I Stereological parameters determined in placental labyrinth zone. J Representative image of TUNEL staining for apoptosis quantification in labyrinth zone. Scale bar = 2.5 mm and 100 μm. All data were analyzed by a general linear mixed model, taking into account litter size as a covariate and taking each fetus as a repeated measure followed by Tukey multiple comparisons test. ROUT test was used for the identification of outlier values. Dots represent raw data (individual values). However, the statistical analysis and the mean ± SEM reported within the graphs were obtained with the general linear mixed model (further explanations can be found in the Materials and Methods statistical analysis section). Placental weight-efficiency was obtained from: SPF (49 fetuses/6 dams), GF (33 fetuses/5 dams), BIF (34 fetuses/6 dams). Laboratorial analysis was performed with: SPF (14–15 placentas from 6 dams), GF (10 placentas from 5 dams) and BIF (9–11 placentas from 6 dams). Only placentas collected on day 16.5 of gestation were analysed. One to three placentas per litter were randomly selected and used for assessment. Placentas were analysed blind to the experimental groups. (NS, not significant; *P < 0.05; ***P < 0.001). D decidua, Jz junctional zone, Lz labyrinth zone, TB trophoblasts, FC fetal capillaries, MBS maternal blood spaces