The global rise in obesity, particularly among women of reproductive age, is a significant concern. Maternal obesity during pregnancy has been linked to long-term, programmed effects on offspring, increasing their susceptibility to obesity and related diseases. These effects may be mediated by epigenetic processes, including changes in gene expression and microRNA (miRNA) profiles. MiRNAs are small molecules that regulate gene expression by binding to target mRNAs. Disturbances in miRNA expression are associated with metabolic conditions like type 2 diabetes, cardiovascular disease, and liver steatosis. Maternal obesity and high-fat diets can program miRNA expression in offspring, affecting their health. However, most studies focus on young adults, neglecting the impact of aging on these programmed phenotypes. The aging process itself begins prenatally, and individuals with suboptimal maternal environments are more vulnerable to age-associated diseases. This study aimed to investigate the liver phenotype of 12-month-old mice born to obese dams and explore the relationship with age-related changes in miRNA expression, specifically miR-582, previously implicated in liver cirrhosis and metabolic disorders.

Numerous studies have established the link between maternal obesity and the increased risk of metabolic disorders in offspring. These studies highlight epigenetic mechanisms, particularly focusing on DNA methylation, histone modification, and the role of non-coding RNAs, such as microRNAs. The involvement of specific miRNAs in the development of metabolic diseases, including type 2 diabetes, cardiovascular disease, and liver steatosis, has been extensively documented. Several studies show that maternal obesity or high-fat diets during pregnancy and lactation can alter miRNA expression in various tissues of the offspring. However, most of these studies focused on younger offspring, limiting our understanding of the long-term consequences and the interplay with the aging process. Research examining the impact of maternal obesity on the aged offspring is relatively scarce, particularly concerning sex-specific effects and the progression of age-related diseases like non-alcoholic fatty liver disease (NAFLD). This gap in the literature underscores the importance of longitudinal studies to assess the long-term effects of maternal obesity on offspring health.

This study utilized a mouse model (C57BL/6 J) of maternal diet-induced obesity. Female mice were fed either a standard chow diet or an obesogenic high-fat diet from weaning until mating. The obesogenic diet consisted of a high-fat diet supplemented with sweetened condensed milk to induce obesity. After mating, females remained on their respective diets throughout pregnancy and lactation. Litter sizes were standardized to six pups per litter on postnatal day 2, and male offspring were weaned onto a control diet on day 21. Body weight and composition were assessed weekly/monthly until 52 weeks of age. At 12 months of age, serum metabolites (insulin, leptin, cholesterol, triglycerides, HDL, LDL, free fatty acids), glucose tolerance (GTT), hepatic lipid content, liver histology (steatosis, fibrosis), and apoptosis (TUNEL assay) were measured. Hepatic expression of miR-582 (3p and 5p) was measured in both 8-week-old and 12-month-old offspring using qPCR, along with mRNA levels of genes involved in fibrosis, apoptosis, and mitochondrial function. Western blotting was used to assess protein expression of key components in these pathways. Statistical analyses included t-tests, Mann-Whitney U tests, ANOVA, and correlation analysis.

At 12 months of age, offspring from obese mothers showed significantly increased body weight (48.8 ± 1.5 g vs 39.1 ± 1.4 g, p<0.0001) and fat mass (19.5 ± 0.8 g vs 10.4 ± 0.9 g, p<0.0001) compared to controls. Serum levels of LDL and leptin were elevated in the offspring of obese mothers (p<0.01 and p<0.001, respectively). Hepatic lipid content was significantly higher in the OC group (p<0.01). Histopathological analysis revealed that a significantly greater percentage of offspring from obese mothers had severe steatosis (56% vs 25%). The offspring of obese dams displayed reduced phosphorylation and total protein levels of AMPKa (p<0.01), along with reduced protein expression of several mitochondrial OXPHOS complexes (p<0.01), despite increased mRNA levels for some complexes, indicating potential post-transcriptional regulation. Although no severe fibrosis was observed, collagen deposition was increased, and relative mRNA levels of *Col1a1* were significantly increased (p<0.05) in the OC group, reflecting an increased risk of fibrosis. The Bax/Bcl-2 ratio, indicating apoptosis, was higher in offspring from obese dams (p<0.05). Notably, expression of miR-582 (both 3p and 5p) was significantly increased in 12-month-old offspring from obese dams (p<0.001 and p<0.05, respectively). This increase was more pronounced in the 12-month-old animals compared to the 8-week-old animals, indicating an age-dependent effect, more exaggerated in the offspring of obese dams. A positive correlation was observed between hepatic miR-582 levels and lipid accumulation.

This study demonstrates that maternal diet-induced obesity programs adverse metabolic changes in male offspring, with effects becoming more evident with age. The increase in body weight, adiposity, and hepatic steatosis in 12-month-old offspring suggests the development of NAFLD. The observed reduction in AMPKa activity and mitochondrial complex protein levels, along with increased expression of pro-fibrotic and pro-apoptotic markers, support the notion of mitochondrial dysfunction and increased susceptibility to liver injury. The age-dependent increase in hepatic miR-582 expression in offspring of obese dams further suggests a potential mechanistic link between this miRNA and the observed accelerated metabolic aging. The study's findings highlight the importance of considering the long-term consequences of maternal obesity, which extend beyond early adulthood and may contribute to the accelerated progression of age-related metabolic diseases. The marked increase in miR-582 levels with age in offspring exposed to maternal obesity suggests that this miRNA may play a crucial role in mediating the long-term metabolic consequences of developmental programming.

This study demonstrates that maternal diet-induced obesity negatively impacts offspring metabolic health, leading to increased adiposity and hepatic steatosis, which are exacerbated with age. The findings highlight the role of miR-582 in this process and support the hypothesis that accelerated metabolic aging contributes to the development of programmed phenotypes. Future research should investigate the mechanisms underlying miR-582's role in mediating the long-term effects of maternal obesity and explore potential therapeutic interventions targeting this miRNA pathway. Studies including female offspring are also warranted to understand sex-specific responses.

The study focused solely on male offspring, limiting the generalizability of the findings to both sexes. While 12 months of age represents a relatively advanced age in mice, it's still only about half their lifespan. Therefore, further research is needed to assess the progression of these effects into later stages of life. The study does not distinguish between effects of maternal obesity itself versus the effects of the specific high-fat/high-sugar diet used. While the latter mimics human dietary excesses, disentangling the effects will be essential in future research.