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Perinatal smoke/nicotine exposure alters lung development and causes asthma in exposed offspring, transmitted transgenerationally. The mechanism underlying the transgenerational inheritance of perinatal smoke/nicotine-induced asthma remains unknown, but germline epigenetic modulations may play a role. Using a well-established rat model of perinatal nicotine-induced asthma, we determined the DNA methylation pattern of spermatozoa of F1 rats exposed perinatally to nicotine in F0 gestation. To identify differentially methylated regions (DMRs), reduced representation bisulfite sequencing was performed on spermatozoa of F1 litters. The top regulated gene body and promoter DMRs were tested for lung gene expression levels, and key proteins involved in lung development and repair were determined. The overall CpG methylation in F1 sperms across gene bodies, promoters, 5'-UTRs, exons, introns, and 3'-UTRs was not affected by nicotine exposure. However, the methylation levels were different between the different genomic regions. Eighty one CpG sites, 16 gene bodies, and 3 promoter regions were differentially methylated. Gene enrichment analysis of DMRs revealed pathways involved in oxidative stress, nicotine response, alveolar and brain development, and cellular signaling. Among the DMRs, Dio1 and Nmu were the most hypermethylated and hypomethylated genes, respectively. Gene expression analysis showed that the mRNA expression and DNA methylation were incongruous. Key proteins involved in lung development and repair were significantly different (FDR < 0.05) between the nicotine and placebo-treated groups. Our data show that DNA methylation is remodeled in offspring spermatozoa upon perinatal nicotine exposure. These epigenetic alterations may play a role in transgenerational inheritance of perinatal smoke/nicotine induced asthma.

Intrauterine growth restriction (IUGR) leads to offspring obesity. In a maternal food restriction (MFR) during pregnancy-related IUGR rat model, bone marrow stem cells showed enhanced adipogenic programming; however, the effect of IUGR on white adipose tissue (WAT) progenitors is unknown. Here, by mRNA and functional profiling, we determined sex-specific adipogenic programming of WAT progenitors isolated from pups on the postnatal day (PND) 1 and 21. On PND1, PPARγ and Pref-1 expression was significantly downregulated in preadipocytes of both MFR males and females; however, at PND21, preadipocytes of MFR males showed upregulation in these genes. Even following adipogenic induction, both male and female MFR adipocytes exhibited lower PPARγ, ADRP, and adiponectin levels at PND1; however, at PND21 MFR male adipocytes showed an upward trend in the expression of these genes. An adipogenesis-specific RT-PCR array showed that male MFR adipocytes were programmed to exhibit stronger adipogenic propensity than females. Last, serum sex hormone and adipocyte estrogen/testosterone receptor expression profiles provide preliminary insights into the possible mechanism underlying sex-specific adipogenic programming in the IUGR offspring. In summary, IUGR programs WAT preadipocytes to greater adipogenic potential in males. Although the altered adipogenic programming following MFR was detectable at PND1, the changes were more pronounced at PND21, suggesting a potential role of postnatal nutrition in facilitating the sex-specific adipogenic programming in the IUGR offspring.

In a rat model, perinatal nicotine exposure results in an epigenetically driven multi- and trans-generationally transmitted asthmatic phenotype that tends to wane over successive generations. However, the effect of repeat nicotine exposure during the F1 (Filial 1) gestational period on the transmitted phenotype is unknown. Using a well-established rat model, we compared lung function, mesenchymal markers of airway reactivity, and global gonadal DNA methylation changes in F2 offspring in a sex-specific manner following perinatal exposure to nicotine in only the F0 gestation, in both F0 and F1 (F0/F1) gestations, and in neither (control group). Both F0 only and F0/F1 exposure groups showed an asthmatic phenotype, an effect that was more pronounced in the F0/F1 exposure group, especially in males. Testicular global DNA methylation increased, while ovarian global DNA methylation decreased in the F0/F1 exposed group. Since the offspring of smokers are more likely to smoke than the offspring of nonsmokers, this sets the stage for more severe asthma if both mother and grandmother had smoked during their pregnancies. Increased gonadal DNA methylation changes following nicotine reexposure in the F1 generation suggests that epigenetic mechanisms might well underlie the transgenerational inheritance of acquired phenotypic traits in general and nicotine-induced asthma in particular.

Perinatal smoke and nicotine exposure predisposes to chronic lung disease and morbidity. As the PPARγ pathway is implicated in structural and functional airway deficits in a well‐established rat model of perinatal nicotine exposure, mitochondrial derangement may accompany these deficits. Therefore, we hypothesized that perinatal nicotine exposure would result in lung mitochondrial dysfunction. Sprague‐Dawley dams received either placebo (CON) or nicotine (NIC, 1 mg.kg‐1) daily from embryonic day 6 to postnatal day 21. Postpartum, pups were breast feed ad libitum. Parenchymal lung sections (~10mg) were taken from adult males for mitochondrial assessment in situ (O2K, Oroboros). Body mass was not different at 5 months between CON and NIC (637±65 vs. 624±49 g; p=0.71). ADP‐stimulated O2 consumption (JO2) was ~50% less in NIC (n=6) compared to CON (n=6) (4.5±0.8 vs.8.8±2.5 pmol.s.mg‐1, respectively; p0.05). These data suggest reduced mitochondrial density, rather than respiratory chain complex dysfunction, with perinatal nicotine exposure. Low maximal JO2 with nicotine exposure, possibly via suppressed mitochondrial biogenesis, may predispose towards increased oxidative stress and chronic lung disease. Grant Funding Source: Supported by NIH HD51857, HD71731