Maternal Obesity in Pregnancy Developmentally Programs Adipose Tissue Inflammation in Young, Lean Male Mice Offspring

Abstract

Obesity during pregnancy has a long-term effect on the health of the offspring including risk of developing the metabolic syndrome. Using a mouse model of maternal diet-induced obesity, we employed a genome-wide approach to investigate the microRNA (miRNA) and miRNA transcription profile in adipose tissue to understand mechanisms through which this occurs. Male offspring of diet-induced obese mothers, fed a control diet from weaning, showed no differences in body weight or adiposity at 8 weeks of age. However, offspring from the obese dams had up-regulated cytokine (Tnfα; P < .05) and chemokine (Ccl2 and Ccl7; P < .05) signaling in their adipose tissue. This was accompanied by reduced expression of miR-706, which we showed can directly regulate translation of the inflammatory proteins IL-33 (41% up-regulated; P < .05) and calcium/calmodulin-dependent protein kinase 1D (30% up-regulated; P < .01). We conclude that exposure to obesity during development primes an inflammatory environment in adipose tissue that is independent of offspring adiposity. Programming of adipose tissue miRNAs that regulate expression of inflammatory signaling molecules may be a contributing mechanism.

Figure 1.

Offspring phenotype at 8 weeks of age. Postweaning body weight trajectory; *, P < .05, n = 11 per group.

Figure 2.

Adipose microarray analysis. qPCR validation of top up-regulated (A) and down-regulated (B) transcripts; gene categories significantly enriched with up-regulated (C) and down-regulated (D) transcripts. qPCR expression was normalized to the geometric mean of the housekeepers β-actin, gapdh, and ppia. *, P < .05; **, P < .01; ***, P < .001 determined by one-tailed Student's t test, n = 10 for control, n = 11 for Mat-Ob.

Figure 3.

Inflammation in 8-week-old male adipose tissue. A, Chemokine expression. B, Inflammatory transcription factor expression at mRNA level. qPCR expression was normalized to the geomean of the housekeepers β-actin, gapdh, and ppia. *, P < .05; **, P < .01; ***, P < .001 determined by 2-tailed Student's t test, n = 10 for control, n = 11 for Mat-Ob.

Figure 4.

Inflammatory cell markers in 8-week-old male epididymal adipose tissue. A, qPCR expression of classical inflammatory cell markers, normalized to the geomean of the housekeeping genes β-actin, gapdh, and ppia (n = 10 for control, n = 11 for Mat-Ob). B, Average macrophage cell count per field of view (n = 5 for control, n = 6 for Mat-Ob).

Figure 5.

Adipose tissue histology in 8-week-old males. A, Adipocyte cell size analysis. B, Adipocyte cell number. Representative images of hemotoxylin stained adipocytes from control (C) and Mat-Ob (D) groups. Scale bar, 75 μm.

Figure 6.

microRNA and putative target validation. A, qPCR validation of selected miRNA from the microarray, normalized to the geomean of the housekeepers rnu6b_2, rnu5a, rnu1a, snord25, and scarna17 (n = 10 and 11 for control and Mat-Ob, respectively). B, Protein levels of CAMK1D determined by Western blotting with representative blots (n = 8 per group). C, Protein levels of IL-33 determined by Western blotting with representative blots (n = 8 per group). D, mRNA levels of Wnk1 (n = 10 per group). E, Target sequence and mutated target sequence of miR-706 on IL-33 3′-UTR subcloned in a pGL3-enhancer vector (pGL3-IL33 3′-UTR-wt and pGL3-IL-33 3′-UTR-mut, respectively). F, Normalized luciferase activity of the wild-type (pGL3-IL-33 3′-UTR-wt) and mutant (pGL3-IL33 3′-UTR-mut) IL-33 3′-UTR/miR-706 binding site in HeLa cells after negative control (NC) or miR-706 mimic transfection (n = 6 per group); *, P < .05; **, P < .01; ***, P < .001.