Ferritins, an ancient family of protein nanocages, concentrate iron in iron–oxy minerals for iron–protein biosynthesis and protection against oxy radical damage. Of the two genetic mechanisms that regulate rates of ferritin-L synthesis, DNA transcription and mRNA translation, more is known about mRNA regulation where iron targets complexes of an mRNA structure, the iron-responsive element (IRE) sequence, and ferritin IRE repressors (iron regulatory proteins 1 and 2). Neither the integration of mRNA and DNA regulation nor the ferritin-L DNA promoter are well studied. We now report the combined effects of DNA transcription and mRNA translation regulation of ferritin-L synthesis. First, the promoter of human ferritin-L, encoding the animal-specific subunit associated with human diseases, was identified, and contained an overlapping Maf recognition element (MARE) and antioxidant responsive element (ARE) that was positively regulated by tert -butylhydroquinone, sulforaphane, and hemin with responses comparable to thioredoxin reductase (ARE regulator) or quinone reductase (MARE/ARE regulator). Iron, a poor regulator of the ferritin-L promoter, was 800 times less effective than sulforaphane. Combining the ferritin-L MARE/ARE and IRE produced a response to hemin that was 3-fold greater than the sum of responses of the MARE/ARE or IRE alone. Regulation of ferritin-L by a MARE/ARE DNA sequence emphasizes the importance of ferritin-L in oxidative stress that complements the mRNA regulation in iron stress. Combining DNA and mRNA mechanisms of regulation, as for ferritin-L, illustrates the advantages of using two types of genetic targets to achieve sensitive responses to multiple signals.

Traditionally, mouse humanization studies have used human fecal transfer to germ-free animals. This practice requires gnotobiotic facilities and is restricted to gnotobiotic mouse lines, which limits humanized mouse research. We have developed a generalizable method to humanize non germ-free mice using antibiotic treatment and human fecal transfer. The method involves depleting resident intestinal microbiota with broad-spectrum antibiotics, introducing human microbiota from frozen fecal samples by weekly gavage, and maintaining mice in HEPA-filtered microisolator cages. Pyrosequencing cecal microbiota 16S rRNA genes showed that recipient mice adopt a humanized microbiota profile analogous to their human donors, and distinct from mice treated with only antibiotics (no fecal transfer) or untreated control mice. In the humanized mice, 75% of the sequence mass was observed in their respective human donor and conversely, 68% of the donor sequence mass was recovered in the recipient mice. Principal component analyses of GC- and HPLC-separated cecal metabolites were performed to determine effects of transplanted microbiota on the metabolome. Cecal metabolite profiles of mice treated with only antibiotics (no fecal transfer) and control mice were dissimilar from each other and from humanized mice. Metabolite profiles for mice humanized from different donor samples clustered near each other, yet were sufficiently distinct that separate clusters were apparent for each donor. Also, cecal concentrations of 57 metabolites were significantly different between humanization treatments. These data demonstrate that our protocol can be used to humanize non germ-free mice and is sufficiently robust to generate metabolomic differences between mice humanized from different human donors.

Rodent models have been invaluable for biomedical research. Preclinical investigations with rodents allow researchers to investigate diseases by using study designs that are not suitable for human subjects. The primary criticism of preclinical animal models is that results are not always translatable to humans. Some of this lack of translation is due to inherent differences between species. However, rodent models have been refined over time, and translatability to humans has improved. Transgenic animals have greatly aided our understanding of interactions between genes and disease and have narrowed the translation gap between humans and model animals. Despite the technological innovations of animal models through advances in genetics, relatively little attention has been given to animal diets. Namely, developing diets that replicate what humans eat will help make animal models more relevant to human populations. This review focuses on commonly used rodent diets that are used to emulate the Western dietary pattern in preclinical studies of obesity and type 2 diabetes, nonalcoholic liver disease, maternal nutrition, and colorectal cancer.