The potentially important role of environmentally responsive epigenetic changes has been recognized with respect to the fetal origins of adult disease (10
). However, there are limited experimental data investigating the impact of prenatal and early developmental environmental exposures on consequent intestinal inflammatory responses. Early postnatal maternal deprivation resulted in increased offspring colitis susceptibility in young adult rats (27
) and influenced colonic mast cell–nerve interactions (28
). Additionally, the lipid composition of the maternal diet has been shown to modify colonic phospholipid constitution and young adult colitis susceptibility in rat offspring (29
). However, the epigenetic consequences of early developmental environmental exposures have not been addressed in regards to mammalian intestinal inflammation. Here we studied the effects of an epigenetically active diet (11
) on murine colitis susceptibility.
MD supplementation during childhood did not influence colitis susceptibility following 10 days of reversal in young adult animals. Consequently, we studied the effect of maternal MD supplementation on offspring intestinal inflammation and found a profound and prolonged sensitization to DSS in the pups resulting from the dietary intervention. The increased susceptibility to chemically induced colitis associated with low volume, but persistent DNA methylation changes at a functionally select, small number of genomic loci. Although we expected mostly increased levels of DNA methylation to result from the supplementation of MDs, both increases and decreases took place. Therefore, developmental reprogramming of DNA methylation appeared to occur in the offspring colonic mucosa as a result of the maternal dietary modification. Interestingly, the X chromosome was particularly sensitive to the MD supplementation with decreased levels of methylation independent from gender.
The MD maternal diet also persistently modified gene expression in the offspring, affecting ~3% of all murine transcripts. Importantly, genes involved in immune responses (Table ) were enriched in the MD responsive group. This observation supports the developmental reprogramming of colonic mucosal immune functions resulting from fetal MD exposure. In addition to the ontologically identified immune-related genes (Table ), we found decreased expression of Cpn2
(Fig. ). CPN2
encodes the large subunit of carboxypeptidase N (CPN) and stabilizes the smaller and catalytically active subunit, CPN1. CPN participates in the degradation of biologically active peptides. The targeted disruption of Cpn1
causes increased susceptibility to anaphylatoxin-mediated shock in mice (30
). Consequently, the decreased expression of Cpn2
may lead to decreased amounts of active Cpn1 during the experimentally induced inflammatory cascade of DSS colitis and may increase MD offspring vulnerability. In the meantime, CPN1 and 2 were increased in the colonic mucosa of patients suffering from IBD compared with healthy controls (20
). It is not known, however, whether the CPN response in IBD patients is dampened compared with acute forms of colitis, or what the baseline levels of CPN1 and 2 are from non-inflamed areas of the mucosa in IBD. Nevertheless, our Cpn2
results further support the persistent reprogramming of colonic mucosal immune responses.
We were able to delineate overlaps between DNA methylation and gene expression changes at a limited number of genomic intervals (18
). The identified inverse correlation between methylation and expression of Ppara
highlights the functionally important DNA methylation changes induced by the fetal exposure to MD supplementation. The observed increased expression of Ppara
in the MD offspring would indicate a relative protection against chemically induced colitis based on previous studies (31
). Therefore, the induced epigenetic change at Ppara
may serve as a protective mechanism to lessen pro-inflammatory alterations (such as decreased Cpn2
expression for instance) and contribute to the grossly normal phenotype of the MD offspring.
Persistent microbiomic alterations in the colonic mucosa of MD offspring were not identified at the genus level. Only one species, Staphylococcus saprophyticus
, had similar changes in composition at P30 and P90, but not at the same volume. Staphylococcus saprophyticus
has been associated with sporadic diarrhea in children (32
), but its relevance in murine colitis models has not been determined. Nevertheless, our study implicates that there is a significant, maternal diet driven prolonged mucosal microbial composition difference in mammalian offspring, regardless of the same dietary and housing environment following weaning. This maternal diet-induced variation is most pronounced in infancy, but persists into young adulthood. While none of the genera differences was preserved from P30 to P90, there were still seven genera that were significantly influenced by MD supplementation of the maternal diet in the offspring colonic mucosa at P90. These particular genera may be important with respect to colitis susceptibility. For example, Roseburia
was decreased in the MD offspring at P90 (Supplementary Material, Fig. S2
) which has been previously observed in patients with predominantly ileal Crohn's disease (33
). The longstanding influence of maternal diet on the offspring microbiota is surprising considering that enteral microbial composition can shift significantly within a day following dietary changes in gnotobiotic murine models (22
). Therefore, it is plausible that the prenatal reprogramming of mucosal immunology upon maternal MD supplementation creates a persistent effect on the enteral microbiome by inducing a longstanding modification of its physiologic pediatric development. Consequently, an overall inflammation prone interaction is created within the key elements (mucosal microbiome, mucosa, host immunity) of IBD pathology.
This study includes the first epigenetically focused developmental dietary intervention attempting to unravel key environmental factors for the rising incidence of IBD. Because the utilized MDs include many common components of prenatal dietary supplements (folate, methionine, betaine, vitamin B12), the results of this experiment could be of great significance. Conclusive results from subsequent studies may support the development of guidelines for the judicious use of MD supplements in future public health policies.