The effect of dietary prebiotics and probiotics on body weight, large intestine indices, and fecal bile acid profile in wild type and IL10-/- mice.

Previous studies have suggested roles of probiotics and prebiotics on body weight management and intestinal function. Here, the effects of a dietary prebiotic, inulin (50 mg/g diet), and probiotic, Bfidobacterium animalis subsp. lactis (Bb12) (final dose verified at 10(5) colony forming unit (cfu)/g diet, comparable to human consumption), were determined separately and in combination in mice using cellulose-based AIN-93G diets under conditions allowed for the growth of commensal bacteria. Continuous consumption of Bb12 and/or inulin did not affect food intake or body, liver, and spleen weights of young and adult mice. Fecal bile acid profiles were determined by nanoESI-MS/MS tandem mass spectrometry. In the presence of inulin, more bacterial deconjugation of taurine from primary bile acids was observed along with an increased cecal weight. Consumption of inulin in the absence or presence of Bb12 also increased the villus cell height in the proximal colon along with a trend of higher bile acid sulfation by intestinal cells. Feeding Bb12 alone at the physiological dose did not affect bile acid deconjugation and had little effect on other intestinal indices. Although interleukin (IL)10-null mice are susceptible to enterocolitis, they maintained the same body weight as the wild type mice under our specific pathogen-free housing condition and showed no signs of inflammation. Nevertheless, they had smaller cecum suggesting a mildly compromised intestinal development even before the disease manifestation. Our results are consistent with the notion that dietary factors such as prebiotics play important roles in the growth of intestinal microbiota and may impact on the intestinal health. In addition, fecal bile acid profiling could potentially be a non-invasive tool in monitoring the intestinal environment.

Loss of IL-10 decreases mouse postpubertal mammary gland development in the absence of inflammation.

IL-10 is a pleiotrophic anti-inflammatory cytokine. Decreased IL-10 expression is associated with an increased breast cancer risk but the mechanism is not clear. This study was designed to test the hypothesis that the loss of IL-10 alters mammary development, even in the absence of inflammation. Wild-type and IL-10-/- mouse littermates were similar in growth, development, and breeding success. Using whole-mounts and paraffin sections, mammary glands from pre-pubertal mice (d21) were found to not be affected by the IL-10 null genotype. However, after the onset of estrous cycling, ductal structure, but not lymph nodes or adipocytes, of IL-10 knockout mice were found to moderately decrease at day 55, 80, and 150 of age. This phenotype was not rescued by lactogenesis. At day 2 of lactation, IL-10 null mice had reduced lobular complexity and glandular area with the retention of adipocytes. These results support the hypothesis that absence of IL-10 reduces glandular development during postnatal development, at maturity, and during the early stages of lactation. Although our study cannot distinguish between a direct IL-10 effect on the epithelial cells and an indirect systemic effect, epithelial cell responses to IL-10 should be considered in the therapeutic applications of cytokines or cytokine ablation.

RNA-protein complexes that form in the spinach chloroplast atpI 5' untranslated region can be divided into two subcomplexes, each comprised of unique cis-elements and trans-factors.

Control of gene expression in chloroplasts is critically dependent upon post-transcriptional mechanisms, most of which require formation of RNA-protein complexes. The 5' untranslated regions (5'UTRs) of chloroplast mRNAs have been shown to affect stability and/or translation of the message. These effects are mediated by the binding of specific protein(s) to the 5'UTR. We can detect such 5'UTR-protein complexes in vitro and have previously shown that the same polypeptide(s) bind many spinach chloroplast 5'UTRs (Robida et al. 2002). Here we report investigations on the RNA elements and protein factors involved in formation of these complexes. Comparison of the atpI 5'UTR, which serves as the representative 5'UTR for these experiments, among 12 angiosperms revealed two phylogenetically conserved regions upstream of a putative ribosome binding site. To determine whether the two conserved regions interact to form a single polypeptide-binding site, binding assays were performed with RNAs containing only one of the two. Those experiments revealed that the entire 5'UTR could be separated into two binding sites for chloroplast polypeptides, each containing one of the two conserved regions. Competition binding assays using the individual binding sites established that each was bound by different polypeptide(s). These data support the hypothesis that there are at least two unique polypeptides involved in these 5'UTR-protein complexes, each binding specifically to a different site within the 5'UTR.

Proteins are shared among RNA-protein complexes that form in the 5' untranslated regions of spinach chloroplast mRNAs.

Gene expression in chloroplasts is strongly regulated at the post-transcriptional level. Most post-transcriptional mechanisms require RNA-protein complexes. Here we report an analysis of RNA-protein complexes that form in the 5' untranslated regions (5'UTRs) of spinach chloroplast mRNAs. Previous studies from our laboratory showed that four ATP synthase 5'UTRs were able to compete with each other for binding by proteins in a chloroplast extract. This implied that at least some of the binding proteins recognized all four of those ATP synthase 5'UTRs. Here, we examine whether the binding proteins are ATP synthase-specific by performing competition-binding assays between an ATP synthase 5'UTR and 5'UTRs from other chloroplast genes. Competition substrates were chosen to represent a wide range of chloroplast mRNAs, including those encoding the photosystems, NADH dehydrogenase, cytochromes and ribosomal subunits, and two previously unexamined ATP synthase subunits. Results from these experiments revealed that, although the ATP synthase-binding proteins do not bind universally to every chloroplast 5'UTR, they do bind to the majority (12/14) of those examined. Thus, these RNA-binding proteins are candidates for factors that link the post-transcriptional expression of many chloroplast genes of disparate function.