Maternal obesity is associated with a lipotoxic placental environment

Abstract

Maternal obesity is associated with placental lipotoxicity, oxidative stress, and inflammation, where MAPK activity may play a central role. Accordingly, we have previously shown that placenta from obese women have increased activation of MAPK-JNK. Here, we performed RNA-sequencing on term placenta from twenty-two subjects who were dichotomized based on pre-pregnancy BMI into lean (BMI 19-24 kg/m(2); n = 12) and obese groups (BMI, 32-43 kg/m(2); n = 12). RNA-seq revealed 288 genes to be significantly different in placenta from obese women by ≥ 1.4-fold. GO analysis identified genes related to lipid metabolism, angiogenesis, hormone activity, and cytokine activity to be altered in placenta from obese women. Indicative of a lipotoxic environment, increased placental lipid and CIDEA protein were associated with decreased AMPK and increased activation of NF-κB (p65) in placenta from obese women. Furthermore, we observed a 25% decrease in total antioxidant capacity and increased nuclear FOXO4 localization in placenta from obese women that was significantly associated with JNK activation, suggesting that maternal obesity may also be associated with increased oxidative stress in placenta. Maternal obesity was also associated with decreased HIF-1α protein expression, suggesting a potential link between increased inflammation/oxidative stress and decreased angiogenic factors. Together, these findings indicate that maternal obesity leads to a lipotoxic placental environment that is associated with decreased regulators of angiogenesis and increased markers of inflammation and oxidative stress.

Keywords: Developmental programming; FOXO; Gestational obesity; Vascular development.

Figure 1. The effects of maternal obesity on global gene expression in placenta

Differentially expressed genes (±1.4-fold, P < 0.05) identified by RNA-seq in placenta (9-lean and 11-obese) were used for gene ontology (GO) analysis of biological processes (A) and molecular functions (D). B, C, E, and F: Fold change of RPKM values for genes identified by gene-set enrichment analysis of biological process (B and C) and molecular functions (E and F) showing genes involved in lipid metabolism (B), angiogenesis (C), hormone activity (E) and cytokine activity (F). Values are expressed as mean fold change ± SE, where * indicates statistical significance (P < 0.05).

Figure 2. The effects of maternal obesity on placental lipid and regulators of lipid metabolism

A: Representative 10x and 40x images from 6-lean and 6-obese placental sections stained for neutral lipids. Staining of lipid in the villous stroma (asterisk) and syncytium (arrow). Negative control for Oil-Red-O staining (bottom panel, 40X). B: Quantification of placental lipid following lipid extraction and normalization to tissue weight (12-lean and 12-obese). C and D: Western blot (C) and densitometric analysis (D) of lipid metabolism and inflammatory regulators (CIDE-A, AMPK, and NF-κB(p65)). Values are expressed as mean ± SE. E: Correlation analysis between CIDE-A and AMPK densitometry values. r = Pearson’s correlation coefficient, where a negative value indicates an inverse relationship. Statistical significance was determined when P < 0.05 (*).

Figure 3. The effects of maternal obesity on TAC and FoxO4 cellular localization in placenta

A: Total antioxidant capacity (TAC) analysis of term placenta from lean and obese women (12-lean and 12-obese). TAC was normalized to total protein and expressed as nM TAC per mg protein. B and D: Western blot (D) and densitometric analysis (B) of FoxO4 levels in the nuclear and cytoplasmic fractions isolated from 8-lean and 8-obese placenta. (#) Represents statistical outliers that were removed from the analysis. Values are expressed as means ± SE. C: Correlations between the densitometry values for placental activated JNK (obtained from our previous publication [13]) and nuclear FoxO4. r = Pearson’s correlation coefficient, where a positive value indicates a positive relationship. Statistical significance was determined when P < 0.05 (*).

Figure 4. The effects of maternal obesity on placental VEGF-A and HIF-1α and vessel density

A and B: Western blot (A) and densitometric analysis (B) of VEGF-A and HIF-1α protein in term placenta from lean (n =12) and obese women (n = 12). C: Vessel density was determined as the number of vessels (stained with CD31) per field. Mean values were determined from 4 fields per section for 6-lean and 6-obese subjects. Values are expressed as means ± SE where * indicates statistical significance (P < 0.05).

Figure 5. Proposed model of the effects of maternal obesity on placental lipotoxicity, oxidative stress, and inflammation

Figure 5 summarizes the effects of maternal obesity on the placental environment. Boxes colored grey indicated decreased levels in obese placenta, whereas boxes colored black with white writing indicate increased levels. Arrows indicate a positive regulation and bar-headed lines show negative regulation. Interactions depicted are based on studies performed in various tissues (in some cases placenta) and have been previously published.