Maternal cigarette smoke exposure contributes to glucose intolerance and decreased brain insulin action in mice offspring independent of maternal diet

Abstract

Background: Maternal smoking leads to intrauterine undernutrition and is associated with low birthweight and higher risk of offspring obesity. Intrauterine smoke exposure (SE) may alter neuroendocrine mediators regulating energy homeostasis as chemicals in cigarette smoke can reach the fetus. Maternal high-fat diet (HFD) consumption causes fetal overnutrition; however, combined effects of HFD and SE are unknown. Thus we investigated the impact of combined maternal HFD and SE on adiposity and energy metabolism in offspring.

Method: Female Balb/c mice had SE (2 cigarettes/day, 5 days/week) or were sham exposed for 5 weeks before mating. Half of each group was fed HFD (33% fat) versus chow as control. The same treatment continued throughout gestation and lactation. Female offspring were fed chow after weaning and sacrificed at 12 weeks.

Results: Birthweights were similar across maternal groups. Faster growth was evident in pups from SE and/or HFD dams before weaning. At 12 weeks, offspring from HFD-fed dams were significantly heavier than those from chow-fed dams (chow-sham 17.6±0.3 g; chow-SE 17.8±0.2 g; HFD-sham 18.7±0.3 g; HFD-SE 18.8±0.4 g, P<0.05 maternal diet effect); fat mass was significantly greater in offspring from chow+SE, HFD+SE and HFD+sham dams. Both maternal HFD and SE affected brain lactate transport. Glucose intolerance and impaired brain response to insulin were observed in SE offspring, and this was aggravated by maternal HFD consumption.

Conclusion: While maternal HFD led to increased body weight in offspring, maternal SE independently programmed adverse health outcomes in offspring. A smoke free environment and healthy diet during pregnancy is desirable to optimize offspring health.

Figure 1. Body weight changes of the offspring mice.

Body weight changes during the suckling and post-weaning periods in chow-sham (open square n = 24), chow-SE (solid square n = 21), HFD-sham (open circle n = 17), and HFD-SE (solid circle n = 15) groups. Results are expressed as mean ± SEM. Data were analysed by ANOVA with repeated measures followed by a post hoc LSD test. *, P<0.05, maternal HFD effect. Ψ, P<0.05, chow-sham significantly different from the other 3 groups. Chow-sham: offspring from dam fed chow & sham exposed; chow-SE: offspring from dam fed chow & cigarette smoke exposed; HFD-sham: offspring from dam fed HFD & sham exposed; HFD-SE: offspring from dams fed HFD & cigarette smoke exposed.

Figure 2. Plasma level and fat mRNA expression of leptin.

Plasma leptin (n = 15–24) and leptin mRNA expression in Rp fat (n = 10–12). Results are expressed as mean ± SEM. Data were analysed by two-way ANOVA followed by a post hoc LSD test. #, P<0.05, significantly different from chow-sham group. Chow-sham: offspring from dam fed chow & sham exposed; chow-SE: offspring from dam fed chow & cigarette smoke exposed; HFD-sham: offspring from dam fed HFD & sham exposed; HFD-SE: offspring from dams fed HFD & cigarette smoke exposed.

Figure 3. Glucose tolerance and markers related to insulin sensitivity.

(A) Change in blood glucose levels above the baseline values during an IPGTT at 11 weeks (glucose 2 g/kg, n = 6) in chow-sham (open square), chow-SE (solid square), HFD-sham (open circle), and HFD-SE (solid circle) pups. Results are expressed as mean ± S.E.M. Data were analysed by ANOVA with repeated measures followed by a post hoc LSD test. Area under the curve for (A) is shown in (B). (C) TNFα mRNA expression in the Rp fat at 12 weeks (n = 12). (D) Liver phosphorylated-PKB/total PKB protein 10 min after exogenous insulin injection (1 U/kg, ip) at 12 weeks (n = 5–6) and representative blots. Results are expressed as mean ± SEM. Data in (B), (C), and (D) were analysed by two-way ANOVA followed by post hoc LSD tests. (A) #, P<0.05, maternal SE effect, HFD-SE & chow-SE different from HFD-sham & chow-sham, respectively. θ, P<0.05, HFD-SE different from chow-SE. (B, C) #, P<0.05, maternal SE effect; ψ, P<0.05, chow-sham different from the other 3 groups. Chow-sham: offspring from dam fed chow & sham exposed; chow-SE: offspring from dam fed chow & cigarette smoke exposed; HFD-sham: offspring from dam fed HFD & sham exposed; HFD-SE: offspring from dams fed HFD & cigarette smoke exposed.

Figure 4. mRNA and protein expression in the hypothalamus.

mRNA expression of NPY (A), Y1 receptor (B), POMC (C), and Sim1 (D), and phosphorylated/total GSK protein (E) and phosphorylated-PKB protein (F) before and 10 min after exogenous insulin injection (1 U/kg, ip) in the offspring hypothalamus at 12 weeks (n = 8–9). Results are expressed as mean ± SEM. Data in (A) and (C) were analysed by multiple-factor ANOVA followed by post hoc LSD tests. Data in (B) and (D) were analysed by two-way ANOVA followed by post hoc LSD tests. *, P<0.05, maternal HFD effect; **, P<0.05, significantly different from chow-SE group; η, P<0.05, insulin injection effect. (A) saline injection (open bars), insulin injection (1 U/kg, closed bars). Chow-sham: offspring from dam fed chow & sham exposed; chow-SE: offspring from dam fed chow & cigarette smoke exposed; HFD-sham: offspring from dam fed HFD & sham exposed; HFD-SE: offspring from dams fed HFD & cigarette smoke exposed. NPY: neuropeptide Y; POMC: proopiomelanocortin; Sim1: single-minded gene 1.

Figure 5. mRNA expression of MCTs in the hypothalamus.

mRNA expression of MCT2 (A), and MCT4 (B) in the offspring hypothalamus at 12 weeks (n = 8–9). Results are expressed as mean ± SEM. Data were analysed by two-way ANOVA followed by a post hoc LSD tests. *, P<0.05, maternal HFD effect; ψ, P<0.05, significantly different from all the other 3 groups. Chow-sham: offspring from dam fed chow & sham exposed; chow-SE: offspring from dam fed chow & cigarette smoke exposed; HFD-sham: offspring from dam fed HFD & sham exposed; HFD-SE: offspring from dams fed HFD & cigarette smoke exposed.

Figure 6. mRNA expression of CPT-1 α and ATGL in the fat.

mRNA expression of CPT-1α (A), and ATGL (B) in the Rp fat at 12 weeks (n = 8–9). Results are expressed as mean ± SEM. Data were analysed by two-way ANOVA followed by post hoc LSD tests. *, P<0.05, maternal HFD effect; #, P<0.05, maternal SE effect; ψ, P<0.05, significantly different from the other 3 groups. Chow-sham: offspring from dam fed chow & sham exposed; chow-SE: offspring from dam fed chow & cigarette smoke exposed; HFD-sham: offspring from dam fed HFD & sham exposed; HFD-SE: offspring from dams fed HFD & cigarette smoke exposed.