No abstract

CYP2J2 epoxygenase is a membrane bound cytochrome P450 that converts omega-3 and omega-6 fatty acids into physiologically active epoxides. In this work, we present a comprehensive comparison of the effects of N-terminal modifications on the properties of CYP2J2 with respect to the activity of the protein in model lipid bilayers using Nanodiscs. We demonstrate that the complete truncation of the N-terminus changes the association of this protein with the E.coli membrane but does not disrupt incorporation in the lipid bilayers of Nanodiscs. Notably, the introduction of silent mutations at the N-terminus was used to express full length CYP2J2 in E. coli while maintaining wild-type functionality. We further show that lipid bilayers are essential for the productive use of NADPH for ebastine hydroxylation by CYP2J2. Taken together, it was determined that the presence of the N-terminus is not as critical as the presence of a membrane environment for efficient electron transfer from cytochrome P450 reductase to CYP2J2 for ebastine hydroxylation in Nanodiscs. This suggests that adopting the native-like conformation of CYP2J2 and cytochrome P450 reductase in lipid bilayers is essential for effective use of reducing equivalents from NADPH for ebastine hydroxylation.

While ingestion of synbiotic yogurts containing Bifidobacterium animalis subsp. lactis and inulin is increasing, their effect on certain microbial groups in the human intestine is unclear. To further investigate this, a large-scale, crossover-design, placebocontrolled study was utilized to evaluate the effect of a synbiotic yogurt containing B. animalis subsp. lactis Bb-12 and inulin on the human intestinal bifidobacteria, clostridia, and enterobacteria. Fecal samples were collected at 14 time points from 46 volunteers who completed the study, and changes in the intestinal bacterial levels were monitored using real-time PCR. Strain Bb-12 could not be detected in feces after 2 weeks of washout. A live/dead PCR procedure indicated that the Bb-12 strain detected in the fecal samples was alive. A significant increase (P < 0.001) in the total bifidobacterial numbers was seen in both groups of subjects during the final washout period compared to the prefeeding period. This increase in total bifidobacteria corresponded with a significant decrease (P < 0.05) in numbers of clostridia but not enterobacteria. No significant differences in numbers of bifidobacteria, clostridia, or enterobacteria were observed between the probiotic and placebo groups during any of the feeding periods. However, subgrouping subjects based on lower initial bifidobacterial numbers or higher initial clostridial numbers did show corresponding significant differences between the synbiotic yogurt and placebo groups. This was not observed for a subgroup with higher initial enterobacterial numbers. While this synbiotic yogurt can increase bifidobacterial numbers and decrease clostridial numbers (but not enterobacterial numbers) in some individuals, it cannot modulate these microbial groups in the majority of individuals.

Background The definitive endoderm arises as a naive epithelial sheet that produces the entire gut tube and associated organs including the liver, pancreas and lungs. Murine explant studies demonstrate that Fibroblast Growth Factor (FGF) signaling from adjacent tissues is required to induce hepatic gene expression from isolated foregut endoderm. The requirement of FGF signaling during liver development is examined via small molecule inhibition during whole embryo culture. Results Loss of FGF signaling prior to hepatic induction results in morphological defects and gene expression changes that are confined to the anterior liver bud. In contrast the posterior portion of the liver bud remains relatively unaffected. Because FGF is thought to act as a morphogen during endoderm organogenesis, the ventral pancreas was also examined after FGF inhibition. Although the size of the ventral pancreas is not affected, loss of FGF signaling results in a significantly higher density of ventral pancreas cells. Conclusions The requirement for FGF-mediated induction of hepatic gene expression differs across the anterior-posterior axis of the developing liver bud. These results underscore the importance of studying tissue differentiation in the context of the whole embryo.

BMP signaling is required to maintain the hepato-pancreatobiliary boundary, at least in part, by indirectly repressing SOX9 in the hepatic endoderm. BMP and FGF signals are each required for the induction of spatially complementary subsets of hepatoblasts. These results underscore the importance of studying early inductive processes in the whole embryo. (Hepatology 2018;68:274-288).