Maternal choline supplementation during murine pregnancy modulates placental markers of inflammation, apoptosis and vascularization in a fetal sex-dependent.

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Maternal choline supplementation during murine pregnancy modulates placental markers of inflammation, apoptosis and vascularization in a fetal sex-dependent manner Sze Ting (Cecilia) Kwan, Julia H. King, Jian Yan, Xinyin Jiang, Emily Wei, Vladislav G. Fomin, Mark S. Roberson, Marie A. Caudill Placenta Volume 53, Pages 57-65 (May 2017) DOI: 10.1016/j.placenta.2017.03.019 Copyright © 2017 Elsevier Ltd Terms and Conditions

Fig. 1 Maternal hepatic concentrations of A) choline, B) betaine, C) dimethylglycine and D) trimethylamine N-oxide at E18.5 in response to three different choline treatments (1X, 2X and 4X). Data were analyzed using ANOVA followed by post-hoc Bonferroni corrections. Values are presented as mean ± SEM. *P ≤ 0.05, **P ≤ 0.001. # Punadjusted ≤ 0.05, Padjusted > 0.05. Placenta 2017 53, 57-65DOI: (10.1016/j.placenta.2017.03.019) Copyright © 2017 Elsevier Ltd Terms and Conditions

Fig. 2 A) Transcript and B-C) protein abundance of IL-1β in the female placentas obtained from dams receiving 1X, 2X or 4X choline treatments at E10.5, E12.5, E15.5 and E18.5. The transcript data are expressed as fold-change relative to the housekeeping gene Tbp and the protein data are expressed relative to β-actin. After normalization, the mean value of the control group at E10.5 was assigned a value of 1 and the mean values of the other groups were presented as a fraction of this value. Statistical analysis was done using the mixed linear model followed by post-hoc Bonferroni corrections. Values are given as mean ± SEM. *P ≤ 0.05. #Punadjusted ≤ 0.05, Padjusted > 0.05. Placenta 2017 53, 57-65DOI: (10.1016/j.placenta.2017.03.019) Copyright © 2017 Elsevier Ltd Terms and Conditions

Fig. 3 A) Transcript and B) protein expression of NF-κB in the E18.5 female placentas obtained from dams receiving 1X, 2X or 4X choline treatments. The transcript data are expressed as fold-change relative to the housekeeping gene Tbp and the protein data are expressed relative to β-actin. After normalization, the mean value of the control group was assigned a value of 1 and the mean values of the treatment groups were presented as a fraction of this value. Statistical analysis was done using the mixed linear model followed by post-hoc Bonferroni corrections. Values are given as mean ± SEM. *P ≤ 0.05. Placenta 2017 53, 57-65DOI: (10.1016/j.placenta.2017.03.019) Copyright © 2017 Elsevier Ltd Terms and Conditions

Fig. 4 mRNA and protein abundance of A-B) IL-1β and C-D) TNF-a in the male placentas obtained from dams receiving 1X, 2X or 4X choline treatments. The transcript data are expressed as fold-change relative to the housekeeping gene Tbp and the protein data are expressed relative to β-actin. After normalization, the mean value of the control group at E10.5 (for mRNA data) or the mean value of the control group (for protein data) was assigned a value of 1 and the mean values of the other groups were presented as a fraction of this value. Statistical analysis was done using the mixed linear model followed by post-hoc Bonferroni corrections. Values are given as mean ± SEM. *P ≤ 0.05. #Punadjusted ≤ 0.05, Padjusted > 0.05. Placenta 2017 53, 57-65DOI: (10.1016/j.placenta.2017.03.019) Copyright © 2017 Elsevier Ltd Terms and Conditions

Fig. 5 mRNA abundance of Vegfa in the female placentas obtained from dams receiving 1X, 2X or 4X choline treatments at E10.5, E12.5, E15.5 and E18.5. Data are expressed as fold-change relative to the housekeeping gene Tbp. After normalization, the mean value of the control group at E10.5 was assigned a value of 1 and the mean values of the other groups were presented as a fraction of this value. Statistical analysis was done using the mixed linear model followed by post-hoc Bonferroni corrections. Values are given as mean ± SEM. *P ≤ 0.05. Placenta 2017 53, 57-65DOI: (10.1016/j.placenta.2017.03.019) Copyright © 2017 Elsevier Ltd Terms and Conditions

Fig. 6 A) The percentage of TUNEL-positive cells in the placenta from dams receiving 1X, 2X or 4X choline treatments at E10.5, E12.5, E15.5 and E18.5. B) Representative images of the apoptotic nuclei within the E10.5 placentas are shown. C) Arterial luminal area in the maternal decidua from dams receiving 1X, 2X or 4X choline treatments at E10.5, E12.5, E15.5 and E18.5. D) Representative images of the smooth muscle actin staining within the maternal decidua are shown. Data were analyzed using the mixed linear model followed by post-hoc Bonferroni corrections. Values are presented as mean ± SEM. *P ≤ 0.05, **P ≤ 0.001. Placenta 2017 53, 57-65DOI: (10.1016/j.placenta.2017.03.019) Copyright © 2017 Elsevier Ltd Terms and Conditions