No abstract

Perturbation of the gut-associated microbial community may underlie many human illnesses, but the mechanisms that maintain homeostasis are poorly understood. We found depletion of butyrate-producing microbes by antibiotic treatment reduced epithelial signaling through the intracellular butyrate sensor PPAR-γ. Nitrate levels increased in the colonic lumen because epithelial expression of Nos2, the gene encoding inducible nitric oxide synthase (iNOS) was elevated in the absence of PPAR-γ-signaling. Microbiota-induced PPAR-γ-signaling also limits the luminal bioavailability of oxygen by driving the energy metabolism of colonic epithelial cells (colonocytes) towards β-oxidation. Therefore, microbiota-activated PPAR-γ-signaling is a homeostatic pathway that prevents a dysbiotic expansion of potentially pathogenic Escherichia and Salmonella by reducing the bioavailability of respiratory electron acceptors to Enterobacteriaceae in the lumen of the colon.

The production of NO ⅐ by mitochondria was investigated by electron paramagnetic resonance using the spin-trapping technique, and by the oxidation of oxymyoglobin. Percoll-purified rat liver mitochondria exhibited a negligible contamination with other subcellular fractions (1-4%) and high degree of functionality (respiratory control ratio ‫؍‬ 5-6). Toluene-permeabilized mitochondria, mitochondrial homogenates, and a crude preparation of nitric oxide synthase (NOS) incubated with the spin trap N-methyl-D-glucamine-dithiocarbamate-Fe II produced a signal ascribed to the NO ⅐ spin adduct (g ‫؍‬ 2.04; a N ‫؍‬ 12.5 G). The intensity of the signal increased with time, protein concentration, and L-Arg, and decreased with the addition of the NOS inhibitor N G -monomethyl-L-arginine. Intact mitochondria, mitochondrial homogenates, and submitochondrial particles produced NO ⅐ (followed by the oxidation of oxymyoglobin) at rates of 1.4, 4.9, and 7.1 nmol NO ⅐ ؋ (min⅐mg protein)؊1 , respectively, with a K m for L-Arg of 5-7 M. Comparison of the rates of NO ⅐ production obtained with homogenates and submitochondrial particles indicated that most of the enzymatic activity was localized in the mitochondrial inner membrane. This study demonstrates that mitochondria are a source of NO ⅐ , the production of which may effect energy metabolism, O 2 consumption, and O 2 free radical formation. Nitric oxide (NO ⅐ )1 is a free radical generated in biological systems by nitric oxide synthases (NOS). Because of its effect on neurotransmission, vasodilation, and immune response (1-3), NO ⅐ plays an important role in physiology, pathology, and pharmacology.Studies with brain tissue and macrophage lysates have shown that NOS is localized exclusively in the soluble fraction (3-6), and recent studies have indicated that the majority (Ͼ80%) of bovine endothelial NOS activity is bound to the particulate fraction of cell homogenates (7,8). Because the particulate fraction used in the studies was expected to contain plasma membranes, as well as microsomes, and, possibly, intracellular organelles, the actual subcellular location of the activity remained to be determined. Other lines of evidence have indicated the presence of NOS in the perinuclear region, in discrete regions of the plasma membrane of cultured endothelial cells, and in intact blood vessels (9, 10); immunocytochemical studies have revealed the presence of a NOS, or an antigenically related protein, in mitochondria isolated from different tissues (11-13). The predominant association of this mtNOS with the mitochondrial membrane (11, 12), and its co-localization with succinate dehydrogenase, a mitochondrial marker of the inner membrane (13), suggested that this enzyme has a particulate localization.These studies as well as the presence of substrates and cofactors in mitochondria required for NOS activity such as L-arginine (L-Arg), L-Arg transporters, Ca 2ϩ , calmodulin, NADPH, and the availability of O 2 , led us to postulate mitochondria as a potential source of NO ⅐ production.Fol...

No abstract

No abstract