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Epigenomic and metabolic responses of hypothalamic POMC neurons to gestational nicotine exposure in adult offspring

View Article: PubMed Central - PubMed

ABSTRACT

Background: Epidemiological and animal studies have reported that prenatal nicotine exposure (PNE) leads to obesity and type-2 diabetes in offspring. Central leptin-melanocortin signaling via hypothalamic arcuate proopiomelanocortin (POMC) neurons is crucial for the regulation of energy and glucose balance. Furthermore, hypothalamic POMC neurons were recently found to mediate the anorectic effects of nicotine through activation of acetylcholine receptors. Here, we hypothesized that PNE impairs leptin-melanocortinergic regulation of energy balance in first-generation offspring by altering expression of long non-coding RNAs (lncRNAs) putatively regulating development and/or function of hypothalamic POMC neurons.

Methods: C57BL/6J females were exposed ad libitum to nicotine through drinking water and crossed with C57BL/6J males. Nicotine exposure was sustained during pregnancy and discontinued at parturition. Offspring development was monitored from birth into adulthood. From the age of 8 weeks, central leptin-melanocortin signaling, diabetes, and obesity susceptibility were assessed in male offspring fed a low-fat or high-fat diet for 16 weeks. Nicotine-exposed and non-exposed C57BL/6J females were also crossed with C57BL/6J males expressing the enhanced green fluorescent protein specifically in POMC neurons. Transgenic male offspring were subjected to laser microdissections and RNA sequencing (RNA-seq) analysis of POMC neurons for determination of nicotine-induced gene expression changes and regulatory lncRNA/protein-coding gene interactions.

Results: Contrary to expectation based on previous studies, PNE did not impair but rather enhanced leptin-melanocortinergic regulation of energy and glucose balance via POMC neurons in offspring. RNA-seq of laser microdissected POMC neurons revealed only one consistent change, upregulation of Gm15851, a lncRNA of yet unidentified function, in nicotine-exposed offspring. RNA-seq further suggested 82 cis-regulatory lncRNA/protein-coding gene interactions, 19 of which involved coding genes regulating neural development and/or function, and revealed expression of several previously unidentified metabolic, neuroendocrine, and neurodevelopment pathways in POMC neurons.

Conclusions: PNE does not result in obesity and type 2 diabetes but instead enhances leptin-melanocortinergic feeding and body weight regulation via POMC neurons in adult offspring. PNE leads to selective upregulation of Gm15851, a lncRNA, in adult offspring POMC neurons. POMC neurons express several lncRNAs and pathways possibly regulating POMC neuronal development and/or function.

Electronic supplementary material: The online version of this article (doi:10.1186/s13073-016-0348-2) contains supplementary material, which is available to authorized users.

No MeSH data available.


Related in: MedlinePlus

PNE decreased body weight gain and moderately increased sensitivity to leptin-induced and melanotan II (MTII)-induced body weight loss of offspring in refeeding experiments. Male offspring were fasted for 12 h, injected i.p. leptin (5 mg/kg), MTII (5 mg/kg), or vehicle (PBS), and immediately offered food. Body weight gain (% of pre-injection body weight) and food intake were determined at defined time points post injection. a–d Leptin experiments: body weight change of STD mice (a) and HFD mice (b), cumulative food intake of STD mice (c) and HFD mice (d). e–h MTII experiments: body weight change of STD mice (e) and HFD mice (f), cumulative food intake of STD mice (g) and HFD mice (h). Data were analyzed by two-way ANOVA followed by Bonferroni post-tests (*, p < 0.05; **, p < 0.01; ***, p < 0.001) or unpaired two-tailed t test: (#, p < 0.05 and numerical p values in b and d). The number of male offspring in each group was n = 7–9 originating from at least three different dams. All data are expressed as mean ± SEM
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Fig3: PNE decreased body weight gain and moderately increased sensitivity to leptin-induced and melanotan II (MTII)-induced body weight loss of offspring in refeeding experiments. Male offspring were fasted for 12 h, injected i.p. leptin (5 mg/kg), MTII (5 mg/kg), or vehicle (PBS), and immediately offered food. Body weight gain (% of pre-injection body weight) and food intake were determined at defined time points post injection. a–d Leptin experiments: body weight change of STD mice (a) and HFD mice (b), cumulative food intake of STD mice (c) and HFD mice (d). e–h MTII experiments: body weight change of STD mice (e) and HFD mice (f), cumulative food intake of STD mice (g) and HFD mice (h). Data were analyzed by two-way ANOVA followed by Bonferroni post-tests (*, p < 0.05; **, p < 0.01; ***, p < 0.001) or unpaired two-tailed t test: (#, p < 0.05 and numerical p values in b and d). The number of male offspring in each group was n = 7–9 originating from at least three different dams. All data are expressed as mean ± SEM

Mentions: Leptin administration examined the response of first order signaling neurons controlling food intake and body weight, including hypothalamic POMC neurons. Vehicle-injected and leptin-injected PNE offspring tended to gain less body weight than vehicle-injected and leptin-injected control offspring both under STD conditions (t = 1 h: F1,27 = 4.71, p = 0.04; t = 2 h: F1,27 = 14.5, p = 0.0007; t = 8 h: F1,27 = 9.6, p = 0.005; t = 24 h: F1,27 = 9.9, p = 0.004; two-way ANOVA) (Fig. 3a) and HFD conditions (t = 1 h: F1,28 = 7.9, p = 0.009; t = 2 h: F1,28 = 3.3, p = 0.08; two-way ANOVA) (Fig. 3b). PNE offspring also tended to be more sensitive to leptin-mediated inhibition of body weight gain under STD conditions (t = 2 h: t14 = 1.89, p = 0.048; t test) (Fig. 3a) and HFD conditions (t = 1 h: t14 = 1.89, p = 0.079; t = 2 h: t14 = 2.21, p = 0.045; t test) (Fig. 3b). Moreover, vehicle-injected PNE offspring gained less body weight than vehicle-injected control offspring both on the STD (t = 2 h: p < 0.05; t = 8 h: p < 0.01; Bonferroni post-test) (Fig. 3a) and HFD (t = 1 h: t14 = 2.08, p = 0.057; t test) (Fig. 3b). PNE did not impact food intake of vehicle-treated and leptin-treated offspring on the STD (Fig. 3c) but moderately lowered food consumption of vehicle-treated and leptin-treated offspring on the HFD (t = 1 h: F1,28 = 4.6, p = 0.04; two-way ANOVA). Furthermore, PNE modestly increased sensitivity of HFD offspring to feeding inhibition by leptin 1 h after refeeding (t14 = 2.11, p = 0.054; t test) (Fig. 3d).Fig. 3


Epigenomic and metabolic responses of hypothalamic POMC neurons to gestational nicotine exposure in adult offspring
PNE decreased body weight gain and moderately increased sensitivity to leptin-induced and melanotan II (MTII)-induced body weight loss of offspring in refeeding experiments. Male offspring were fasted for 12 h, injected i.p. leptin (5 mg/kg), MTII (5 mg/kg), or vehicle (PBS), and immediately offered food. Body weight gain (% of pre-injection body weight) and food intake were determined at defined time points post injection. a–d Leptin experiments: body weight change of STD mice (a) and HFD mice (b), cumulative food intake of STD mice (c) and HFD mice (d). e–h MTII experiments: body weight change of STD mice (e) and HFD mice (f), cumulative food intake of STD mice (g) and HFD mice (h). Data were analyzed by two-way ANOVA followed by Bonferroni post-tests (*, p < 0.05; **, p < 0.01; ***, p < 0.001) or unpaired two-tailed t test: (#, p < 0.05 and numerical p values in b and d). The number of male offspring in each group was n = 7–9 originating from at least three different dams. All data are expressed as mean ± SEM
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Fig3: PNE decreased body weight gain and moderately increased sensitivity to leptin-induced and melanotan II (MTII)-induced body weight loss of offspring in refeeding experiments. Male offspring were fasted for 12 h, injected i.p. leptin (5 mg/kg), MTII (5 mg/kg), or vehicle (PBS), and immediately offered food. Body weight gain (% of pre-injection body weight) and food intake were determined at defined time points post injection. a–d Leptin experiments: body weight change of STD mice (a) and HFD mice (b), cumulative food intake of STD mice (c) and HFD mice (d). e–h MTII experiments: body weight change of STD mice (e) and HFD mice (f), cumulative food intake of STD mice (g) and HFD mice (h). Data were analyzed by two-way ANOVA followed by Bonferroni post-tests (*, p < 0.05; **, p < 0.01; ***, p < 0.001) or unpaired two-tailed t test: (#, p < 0.05 and numerical p values in b and d). The number of male offspring in each group was n = 7–9 originating from at least three different dams. All data are expressed as mean ± SEM
Mentions: Leptin administration examined the response of first order signaling neurons controlling food intake and body weight, including hypothalamic POMC neurons. Vehicle-injected and leptin-injected PNE offspring tended to gain less body weight than vehicle-injected and leptin-injected control offspring both under STD conditions (t = 1 h: F1,27 = 4.71, p = 0.04; t = 2 h: F1,27 = 14.5, p = 0.0007; t = 8 h: F1,27 = 9.6, p = 0.005; t = 24 h: F1,27 = 9.9, p = 0.004; two-way ANOVA) (Fig. 3a) and HFD conditions (t = 1 h: F1,28 = 7.9, p = 0.009; t = 2 h: F1,28 = 3.3, p = 0.08; two-way ANOVA) (Fig. 3b). PNE offspring also tended to be more sensitive to leptin-mediated inhibition of body weight gain under STD conditions (t = 2 h: t14 = 1.89, p = 0.048; t test) (Fig. 3a) and HFD conditions (t = 1 h: t14 = 1.89, p = 0.079; t = 2 h: t14 = 2.21, p = 0.045; t test) (Fig. 3b). Moreover, vehicle-injected PNE offspring gained less body weight than vehicle-injected control offspring both on the STD (t = 2 h: p < 0.05; t = 8 h: p < 0.01; Bonferroni post-test) (Fig. 3a) and HFD (t = 1 h: t14 = 2.08, p = 0.057; t test) (Fig. 3b). PNE did not impact food intake of vehicle-treated and leptin-treated offspring on the STD (Fig. 3c) but moderately lowered food consumption of vehicle-treated and leptin-treated offspring on the HFD (t = 1 h: F1,28 = 4.6, p = 0.04; two-way ANOVA). Furthermore, PNE modestly increased sensitivity of HFD offspring to feeding inhibition by leptin 1 h after refeeding (t14 = 2.11, p = 0.054; t test) (Fig. 3d).Fig. 3

View Article: PubMed Central - PubMed

ABSTRACT

Background: Epidemiological and animal studies have reported that prenatal nicotine exposure (PNE) leads to obesity and type-2 diabetes in offspring. Central leptin-melanocortin signaling via hypothalamic arcuate proopiomelanocortin (POMC) neurons is crucial for the regulation of energy and glucose balance. Furthermore, hypothalamic POMC neurons were recently found to mediate the anorectic effects of nicotine through activation of acetylcholine receptors. Here, we hypothesized that PNE impairs leptin-melanocortinergic regulation of energy balance in first-generation offspring by altering expression of long non-coding RNAs (lncRNAs) putatively regulating development and/or function of hypothalamic POMC neurons.

Methods: C57BL/6J females were exposed ad libitum to nicotine through drinking water and crossed with C57BL/6J males. Nicotine exposure was sustained during pregnancy and discontinued at parturition. Offspring development was monitored from birth into adulthood. From the age of 8&nbsp;weeks, central leptin-melanocortin signaling, diabetes, and obesity susceptibility were assessed in male offspring fed a low-fat or high-fat diet for 16&nbsp;weeks. Nicotine-exposed and non-exposed C57BL/6J females were also crossed with C57BL/6J males expressing the enhanced green fluorescent protein specifically in POMC neurons. Transgenic male offspring were subjected to laser microdissections and RNA sequencing (RNA-seq) analysis of POMC neurons for determination of nicotine-induced gene expression changes and regulatory lncRNA/protein-coding gene interactions.

Results: Contrary to expectation based on previous studies, PNE did not impair but rather enhanced leptin-melanocortinergic regulation of energy and glucose balance via POMC neurons in offspring. RNA-seq of laser microdissected POMC neurons revealed only one consistent change, upregulation of Gm15851, a lncRNA of yet unidentified function, in nicotine-exposed offspring. RNA-seq further suggested 82 cis-regulatory lncRNA/protein-coding gene interactions, 19 of which involved coding genes regulating neural development and/or function, and revealed expression of several previously unidentified metabolic, neuroendocrine, and neurodevelopment pathways in POMC neurons.

Conclusions: PNE does not result in obesity and type 2 diabetes but instead enhances leptin-melanocortinergic feeding and body weight regulation via POMC neurons in adult offspring. PNE leads to selective upregulation of Gm15851, a lncRNA, in adult offspring POMC neurons. POMC neurons express several lncRNAs and pathways possibly regulating POMC neuronal development and/or function.

Electronic supplementary material: The online version of this article (doi:10.1186/s13073-016-0348-2) contains supplementary material, which is available to authorized users.

No MeSH data available.


Related in: MedlinePlus