Increasing maternal body mass index is associated with systemic inflammation in the mother and the activation of distinct placental inflammatory pathways

Abstract

Obese pregnant women have increased levels of proinflammatory cytokines in maternal circulation and placental tissues. However, the pathways contributing to placental inflammation in obesity are largely unknown. We tested the hypothesis that maternal body mass index (BMI) was associated with elevated proinflammatory cytokines in maternal and fetal circulations and increased activation of placental inflammatory pathways. A total of 60 women of varying pre-/early pregnancy BMI, undergoing delivery by Cesarean section at term, were studied. Maternal and fetal (cord) plasma were collected for analysis of insulin, leptin, IL-1beta, IL-6, IL-8, monocyte chemoattractant protein (MCP) 1, and TNFalpha by multiplex ELISA. Activation of the inflammatory pathways in the placenta was investigated by measuring the phosphorylated and total protein expression of p38-mitogen-activated protein kinase (MAPK), c-Jun-N-terminal kinase (JNK)-MAPK, signal transducer-activated transcription factor (STAT) 3, caspase-1, IL-1beta, IkappaB-alpha protein, and p65 DNA-binding activity. To determine the link between activated placental inflammatory pathways and elevated maternal cytokines, cultured primary human trophoblast (PHT) cells were treated with physiological concentrations of insulin, MCP-1, and TNFalpha, and inflammatory signaling analyzed by Western blot. Maternal BMI was positively correlated with maternal insulin, leptin, MCP-1, and TNFalpha, whereas only fetal leptin was increased with BMI. Placental phosphorylation of p38-MAPK and STAT3, and the expression of IL-1beta protein, were increased with maternal BMI; phosphorylation of p38-MAPK was also correlated with birth weight. In contrast, placental NFkappaB, JNK and caspase-1 signaling, and fetal cytokine levels were unaffected by maternal BMI. In PHT cells, p38-MAPK was activated by MCP-1 and TNFalpha, whereas STAT3 phosphorylation was increased following TNFalpha treatment. Maternal BMI is associated with elevated maternal cytokines and activation of placental p38-MAPK and STAT3 inflammatory pathways, without changes in fetal systemic inflammatory profile. Activation of p38-MAPK by MCP-1 and TNFalpha, and STAT3 by TNFalpha, suggests a link between elevated proinflammatory cytokines in maternal plasma and activation of placental inflammatory pathways. We suggest that inflammatory processes associated with elevated maternal BMI may influence fetal growth by altering placental function.

Keywords: cytokines; innate immune response; obesity.

© 2014 by the Society for the Study of Reproduction, Inc.

Figures

FIG. 1

Placental p38-MAPK and JNK-MAPK signaling in relation to maternal BMI. A) Representative Western blots of phosphorylated p38-MAPK (T182/Y180) and JNK-MAPK (T183/Y185) and total p38-MAPK and JNK-MAPK protein expression in homogenates of placentas from pregnancies with varying maternal BMI (n = 32). Scatter plots demonstrate relationship between maternal BMI and placental p38-MAPK (T182/Y180) phosphorylation (B), p38-MAPK expression (C), p46/54 JNK-MAPK (T183/Y185) phosphorylation (D), and p46/54 JNK-MAPK expression (E). Line of best fit indicates significant correlation with maternal BMI.

FIG. 2

Placental STAT3 and NFκB activity in relation to maternal BMI. Representative Western blots of phosphorylated STAT3 (Y705) and STAT3 (A), and IκBα (D) protein expression in homogenates of placentas from pregnancies with varying maternal BMI (n = 32). Scatter plots demonstrate relationship between maternal BMI and placental STAT3 (Y705) phosphorylation (B), STAT3 expression (C), IκBα expression (E), and nuclear p65 DNA-binding activity (n = 21; F). Line of best fit indicates significant correlation with maternal BMI.

FIG. 3

Activation of placental inflammasome complex in relation to maternal BMI. A) Representative Western blots of pro-caspase-1, caspase-1, and IL-1β protein expression in homogenates of placentas from pregnancies with varying maternal BMI (n = 32). Scatter plots demonstrate relationship between maternal BMI and placental pro-caspase-1 (B), caspase-1 (C), and IL-1β (D) expression. Line of best fit indicates significant correlation with maternal BMI.

FIG. 4

Influence of insulin on p38-MAPK and STAT3 phosphorylation in cultured PHTs. Representative Western blots of phosphorylated p38-MAPK (T182/Y180) and p38 MAPK (A), and phosphorylated STAT3 (Y705) and STAT3 expression (D) in PHT cells treated with insulin (5.8 ng/ml) or vehicle control. Histograms illustrate relative protein expression of phosphorylated p38-MAPK (T182/Y180) (B), p38-MAPK (C), phosphorylated STAT3 (Y705) (E), and STAT3 (F). Mean + SEM, n = 4. Cnt, control; Ins, insulin.

FIG. 5

Influence of MCP-1 and TNFα on p38-MAPK and STAT3 phosphorylation in cultured PHTs. Representative Western blots of phosphorylated p38-MAPK (T182/Y180) and p38 MAPK (A), and phosphorylated STAT3 (Y705) and STAT3 (D) expression in PHT cells treated with MCP-1 (100 pg/ml), TNFα (10 pg/ml), or vehicle control (Cnt). Histograms illustrate relative protein expression of phosphorylated p38-MAPK (T182/Y180) (B), p38-MAPK (C), phosphorylated STAT3 (Y705) (E), and STAT3 (F). Mean + SEM, n = 4. ** P < 0.01, *** P < 0.001 versus control.