Mitochondrial enzyme activity is reduced in skeletal muscle of OGDM women.

<p>Quantitative bar graphs of mitochondrial DNA copy number (<i>A</i>) and enzyme activity of citrate synthase (<i>B</i>) complex I (<i>C</i>), complex II (<i>D</i>), complex III (<i>E</i>), and complex IV (<i>F</i>) of the respiratory chain in pyramidalis muscle collected during scheduled cesarean section. Data are mean ± SEM. *<i>P</i><0.05 <i>vs</i>. NGT.</p

Work flow of statistical analysis of single proteins.

<p>Work flow of statistical analysis of single proteins.</p

Mitochondrial respiratory chain complex proteins are not different between ONGT and OGDM women.

<p>Complex I (<i>A</i>), complex II (<i>B</i>), complex III (<i>C</i>), complex IV (<i>D</i>). Representative Western blots where calnexin is used as loading control (<i>E</i>).</p

AMPK phosphorylation is reduced in OGDM women, though regulators of mitochondrial biogenesis are not.

<p>Quantitative bar graphs of PPARα (<i>A</i>) and PGC-1α (<i>B</i>), total AMPK (<i>C</i>), and phospho-AMPK (<i>D</i>) protein content in pyramidalis muscle collected during scheduled cesarean section. Representative Western blots where calnexin is used as loading control (<i>E</i>). Data are mean ± SEM. *<i>P</i><0.05 <i>vs</i>. NGT.</p

Clinical characteristics of the subjects for Study 2.

<p>Data are mean ± SEM. OGTT, oral glucose tolerance test. Independent <i>t</i>-test *<i>P</i><0.05.</p><p>Clinical characteristics of the subjects for Study 2.</p

Nicotinamide Promotes Adipogenesis in Umbilical Cord-Derived Mesenchymal Stem Cells and Is Associated with Neonatal Adiposity: The Healthy Start BabyBUMP Project

<div><p>The cellular mechanisms whereby excess maternal nutrition during pregnancy increases adiposity of the offspring are not well understood. However, nicotinamide (NAM), a fundamental micronutrient that is important in energy metabolism, has been shown to regulate adipogenesis through inhibition of SIRT1. Here we tested three novel hypotheses: 1) NAM increases the adipogenic response of human umbilical cord tissue-derived mesenchymal stem cells (MSCs) through a SIRT1 and PPARγ pathway; 2) lipid potentiates the NAM-enhanced adipogenic response; and 3) the adipogenic response to NAM is associated with increased percent fat mass (%FM) among neonates. MSCs were derived from the umbilical cord of 46 neonates born to non-obese mothers enrolled in the Healthy Start study. Neonatal %FM was measured using air displacement plethysmography (Pea Pod) shortly after birth. Adipogenic differentiation was induced for 21 days in the 46 MSC sets under four conditions, +NAM (3mM)/–lipid (200 μM oleate/palmitate mix), +NAM/+lipid, –NAM/+lipid, and vehicle-control (–NAM/–lipid). Cells incubated in the presence of NAM had significantly higher PPARγ protein (+24%, p <0.01), FABP4 protein (+57%, p <0.01), and intracellular lipid content (+51%, p <0.01). Lipid did not significantly increase either PPARγ protein (p = 0.98) or FABP4 protein content (p = 0.82). There was no evidence of an interaction between NAM and lipid on adipogenic response of PPARγ or FABP4 protein (p = 0.99 and p = 0.09). In a subset of 9 MSC, SIRT1 activity was measured in the +NAM/-lipid and vehicle control conditions. SIRT1 enzymatic activity was significantly lower (-70%, p <0.05) in the +NAM/-lipid condition than in vehicle-control. In a linear model with neonatal %FM as the outcome, the percent increase in PPARγ protein in the +NAM/-lipid condition compared to vehicle-control was a significant predictor (β = 0.04, 95% CI 0.01–0.06, p <0.001). These are the first data to support that chronic NAM exposure potentiates adipogenesis in human MSCs <i>in-vitro</i>, and that this process involves PPARγ and SIRT1.</p></div

Effects of NAM (3mM) and lipid (200uM) during adipose differentiation on SIRT1 protein and enzyme activity.

<p>SIRT1 protein content in MSCs measured by ICE assay at day 21 in all four treatment conditions in the full experimental cohort of 46 cell sets (A). A representative sample of 9 sets of cells from the 46 experimental sets was used for SIRT1 enzyme activity at day 21 in vehicle-control and NAM conditions only (B). *p<0.05 for t-test of NAM vs. vehicle-control; #p<0.05 for main effect of NAM regardless of lipid treatment using MANOVA.</p

Scatter plots depicting the correlation between neonatal adiposity and the percent change of PPARγ (A) and FABP4 (B) protein content at day 21 between vehicle-control and NAM only conditions (N = 46).

<p>Scatter plots depicting the correlation between neonatal adiposity and the percent change of PPARγ (A) and FABP4 (B) protein content at day 21 between vehicle-control and NAM only conditions (N = 46).</p

Effects of NAM (3mM) incubation during adipocyte cell differentiation in human MSC.

<p>Cells were harvested at day 9 of differentiation and mRNA expression of PPARγ (A), SIRT1 enzyme activity (B), and acetylation of SIRT1 protein targets, PPARγ (C) and β-catenin (D), and mRNA expression of NAMPT (E) in the vehicle-control and NAM only conditions as described in Methods. N = 9 per group. *p<0.05 vs. vehicle-control.</p