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. 2020 Apr 18;21(8):2834.
doi: 10.3390/ijms21082834.

Prenatal S-Adenosine Methionine (SAMe) Induces Changes in Gene Expression in the Brain of Newborn Mice That Are Prevented by Co-Administration of Valproic Acid (VPA)

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Prenatal S-Adenosine Methionine (SAMe) Induces Changes in Gene Expression in the Brain of Newborn Mice That Are Prevented by Co-Administration of Valproic Acid (VPA)

Liza Weinstein-Fudim et al. Int J Mol Sci. .
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Abstract

In previous studies, we produced changes in gene expression in the brain of mice by early postnatal administration of valproic acid (VPA), with distinct differences between genders. The addition of S-adenosine methionine (SAMe) normalized the expression of most genes in both genders, while SAMe alone induced no changes. We treated pregnant dams with a single injection of VPA on day 12.5 of gestation, or with SAMe during gestational days 12-14, or by a combination of VPA and SAMe. In the frontal half of the brain, we studied the expression of 770 genes of the pathways involved in neurophysiology and neuropathology using the NanoString nCounter method. SAMe, but not VPA, induced statistically significant changes in the expression of many genes, with differences between genders. The expression of 112 genes was changed in both sexes, and another 170 genes were changed only in females and 31 only in males. About 30% of the genes were changed by more than 50%. One of the most important pathways changed by SAMe in both sexes was the VEGF (vascular endothelial growth factor) pathway. Pretreatment with VPA prevented almost all the changes in gene expression induced by SAMe. We conclude that large doses of SAMe, if administered prenatally, may induce significant epigenetic changes in the offspring. Hence, SAMe and possibly other methyl donors may be epigenetic teratogens.

Keywords: ASD; NanoString nCounter; SAMe; VPA; epigenetics; gene expression.

Conflict of interest statement

The authors declare no conflict of interest.

Figures

Figure 1
Figure 1
(AC). Control versus S-adenosyl methionine (SAMe) offspring weight gain between post-natal day (PND)1 and PND30. Weight analysis of control versus SAMe pups between PND1 and PND15 days. Males and females combined. The number of pups in each group was 18. (B,C): Weight analysis of control vs. SAMe offspring between PND22 and PND30. (B) represents males and (C) represents females. Number of animals in each group: 6–9 in males, 9–12 in females.
Figure 2
Figure 2
Number of genes differently expressed in males, females, and both sexes.
Figure 3
Figure 3
VEGF signaling pathway in males (A) and females (B) exposed to SAMe during pregnancy. Ingenuity pathway analysis of differentially expressed genes in mice treated with SAMe administration. Ingenuity Pathway (IPA) Analysis identified the VEGF-a (vascular endothelial growth factor A) signaling pathway as being enriched. The network was generated through the use of IPA (Qiagen, Ingenuity Systems, www.ingenuity.com) on normalized mRNA values measured by Nanostring analysis. Nodes represent molecules in a pathway, while the biological relationship between nodes is represented by a line (edge). Edges are supported by at least one reference in the Ingenuity Knowledge Base. The brightness of color in a node indicates the degree of upregulation (red) or downregulation (green). Nodes are displayed using shapes that represent the functional class of a gene product (Circle = Other, Nested Circle = Group or Complex, Rhombus = Peptidase, Square = Cytokine, Triangle = Kinase, Vertical ellipse = Transmembrane receptor). Edges are marked with symbols to represent the relationship between nodes (Line only = Binding only, Flat line = inhibits, Solid arrow = Acts on, Solid arrow with flat line = inhibits and acts on, Open circle = leads to, Open arrow = translocates to).
Figure 3
Figure 3
VEGF signaling pathway in males (A) and females (B) exposed to SAMe during pregnancy. Ingenuity pathway analysis of differentially expressed genes in mice treated with SAMe administration. Ingenuity Pathway (IPA) Analysis identified the VEGF-a (vascular endothelial growth factor A) signaling pathway as being enriched. The network was generated through the use of IPA (Qiagen, Ingenuity Systems, www.ingenuity.com) on normalized mRNA values measured by Nanostring analysis. Nodes represent molecules in a pathway, while the biological relationship between nodes is represented by a line (edge). Edges are supported by at least one reference in the Ingenuity Knowledge Base. The brightness of color in a node indicates the degree of upregulation (red) or downregulation (green). Nodes are displayed using shapes that represent the functional class of a gene product (Circle = Other, Nested Circle = Group or Complex, Rhombus = Peptidase, Square = Cytokine, Triangle = Kinase, Vertical ellipse = Transmembrane receptor). Edges are marked with symbols to represent the relationship between nodes (Line only = Binding only, Flat line = inhibits, Solid arrow = Acts on, Solid arrow with flat line = inhibits and acts on, Open circle = leads to, Open arrow = translocates to).
Figure 4
Figure 4
(A,B). Effect of Valproic acid (VPA) and SAMe administration on gene expression in the frontal half of the brain. Heat map of the genes significantly changed by SAMe in comparison to controls. A. left column - females, B. right column - males.
Figure 5
Figure 5
MA Plot of log 2 fold-change (y-axis) versus base mean (normalized average expression (x-axis). Every dot represents one gene. Red dots: genes with a statistically significant change compared to controls. Grey dots: no significant change. Higher and lower 2-fold change red dots are marked by arrows. (A). Gene expression in males. (B). Gene expression in females.
Figure 6
Figure 6
(A,B) show the MA plot of a two-fold log change in the different genes studied in VPA-treated versus control mice in both genders No statistically significant changes were observed between the VPA and control groups in both male and female newborn mice.
Figure 7
Figure 7
(A,B) show the MA plot of a two-fold log change in the different genes studied in VPA+ SAMe-treated versus control mice in both genders. In females, only Slc2a1 (solute carrier family 2 member 1) was found to be changed in the VPA+ SAMe group. This gene changed by 445% by SAMe administration alone compared to the control and was reduced by VPA injection to 40% of the change (Table 7). In males, four genes were changed by co administration of VPA+ SAMe: Fus (fused in sarcoma), Pdgfr (platelet derived growth factor receptor, beta polypeptide), Prkcg (protein kinase C, gamma), and Vegfa (vascular endothelial growth factor A). In Vegfa, VPA administration corrected the upregulation caused by SAMe from 390% to 28%, but the change was still statistically significant. In the other three genes, VPA injection did not change, or slightly increased, the effect of SAMe alone, as reported in Table 7.

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