Objective The inner mucus layer in mouse colon normally separates bacteria from the epithelium. Do humans have a similar inner mucus layer and are defects in this mucus layer a common denominator for spontaneous colitis in mice models and ulcerative colitis? Methods and Results The colon mucus layer of mice deficient in the Muc2 mucin, Core 1 O-glycans, Tlr5, IL10 and Slc9a3 (Nhe3) together with dextran sulfate (DSS) treated mice was immunostained for Muc2 and the bacterial localization in the mucus was analyzed. All murine colitis models revealed bacteria in contact with the epithelium. Additional analysis of the less inflamed IL10−/− mice revealed a thicker mucus layer than WT, but the properties were different as the inner mucus layer could be penetrated both by bacteria in vivo and by fluorescent beads the size of bacteria ex vivo. Clear separation between bacteria or fluorescent beads and the epithelium mediated by the inner mucus layer was also evident in normal human sigmoid colon biopsies. In contrast, mucus on colon biopsies of ulcerative colitis (UC) patients with acute inflammation had a highly penetrable mucus. Most UC patients in remission had similar to controls an impenetrable mucus layer. Conclusions Normal human sigmoid colon has an inner mucus layer impenetrable to bacteria. The colon mucus in animal models that spontaneously develop colitis and in UC patients with active disease allows bacteria to penetrate and reach the epithelium. Thus colon mucus properties can be modulated and suggest a novel model of UC pathophysiology.

Virtually all nutrients from the diet are absorbed into blood across the highly polarized epithelial cell layer forming the small and large intestinal mucosa. Anatomical, histological, and functional specializations along the gastrointestinal tract are responsible for the effective and regulated nutrient transport via both passive and active mechanisms. In this chapter, we summarize the current state of knowledge regarding the mechanism of intestinal absorption of key nutrients such as sodium, anions (chloride, sulfate, oxalate), carbohydrates, amino acids and peptides, lipids, lipidand water-soluble vitamins, as well as the major minerals and micronutrients. This outline, including the molecular identity, specificity, and coordinated activities of key transport proteins and genes involved, serves as the background for the following chapters focused on the pathophysiology of acquired and congenital intestinal malabsorption, as well as clinical tools to test and treat malabsorptive symptoms.

Fibroblast growth factor (FGF)23 is a phosphaturic hormone that decreases circulating 1alpha,25-dihydroxyvitamin D(3) [1,25(OH)(2)D(3)] and elicits hypophosphatemia, both of which contribute to rickets/osteomalacia. It has been shown recently that serum FGF23 increases after treatment with renal 1,25(OH)(2)D(3) hormone, suggesting that 1,25(OH)(2)D(3) negatively feedback controls its levels by inducing FGF23. To establish the tissue of origin and the molecular mechanism by which 1,25(OH)(2)D(3) increases circulating FGF23, we administered 1,25(OH)(2)D(3) to C57BL/6 mice. Within 24 h, these mice displayed a dramatic elevation in serum immunoreactive FGF23, and the expression of FGF23 mRNA in bone was significantly upregulated by 1,25(OH)(2)D(3), but there was no effect in several other tissues. Furthermore, we treated rat UMR-106 osteoblast-like cells with 1,25(OH)(2)D(3), and real-time PCR analysis revealed a dose- and time-dependent stimulation of FGF23 mRNA concentrations. The maximum increase in FGF23 mRNA was 1,024-fold at 10(-7) M 1,25(OH)(2)D(3) after 24-h treatment, but statistically significant differences were observed as early as 4 h after 1,25(OH)(2)D(3) treatment. In addition, using cotreatment with actinomycin D or cycloheximide, we observed that 1,25(OH)(2)D(3) regulation of FGF23 gene expression occurs at the transcriptional level, likely via the nuclear vitamin D receptor, and is dependent on synthesis of an intermediary transfactor. These results indicate that bone is a major site of FGF23 expression and source of circulating FGF23 after 1,25(OH)(2)D(3) administration or physiological upregulation. Our data also establish FGF23 induction by 1,25(OH)(2)D(3) in osteoblasts as a feedback loop between these two hormones that completes a kidney-intestine-bone axis that mediates phosphate homeostasis.

Glutamate is the major excitatory neurotransmitter in the mammalian central nervous system. Packaging and storage of glutamate into glutamatergic neuronal vesicles requires ATP-dependent vesicular glutamate uptake systems, which utilize the electrochemical proton gradient as a driving force. VGLUT1, the first identified vesicular glutamate transporter, is only expressed in a subset of glutamatergic neurons. We report here the molecular cloning and functional characterization of a novel glutamate transporter, VGLUT2, from mouse brain. VGLUT2 has all major functional characteristics of a synaptic vesicle glutamate transporter, including ATP dependence, chloride stimulation, substrate specificity, and substrate affinity. It has 75 and 79% amino acid identity with human and rat VGLUT1, respectively. However, expression patterns of VGLUT2 in brain are different from that of VGLUT1. In addition, VGLUT2 activity is dependent on both membrane potential and pH gradient of the electrochemical proton gradient, whereas VGLUT1 is primarily dependent on only membrane potential. The presence of VGLUT2 in brain regions lacking VGLUT1 suggests that the two isoforms together play an important role in vesicular glutamate transport in glutamatergic neurons.Neurotransmission depends on the regulated exocytotic release of vesicular transmitter molecules to the synaptic cleft, where they interact with postsynaptic receptors that subsequently transduce the information. Two types of neurotransmitter transporters have been identified based on membrane localization on plasma membrane or vesicular membrane. Removal of the transmitter from the synaptic cleft results in termination of the signal, and this requires destruction of transmitter or reuptake of transmitter back to the presynaptic terminal or glial cells via a sodium-dependent uptake system on the plasma membrane (1). Packaging and storage of neurotransmitters into specialized secretory vesicles in neurons ensures their regulated release. This storage is also crucial for protecting the neurotransmmitter molecules from leakage or intraneuronal metabolism and for protecting the neuron from possible toxic effects. This process is mediated by specific transporters on the vesicular membranes. At least four different types of vesicular transporters have been functionally identified that are specific for transport of classic neurotransmitters: monoamines, acetylcholine, ␥-aminobutyric acid (GABA), and glutamate (2, 3). Unlike the plasma membrane transporters, which rely on a sodium gradient across the plasma membrane, all of these vesicular transport processes depend on the proton electrochemical gradient (⌬ Hϩ ) 1 generated by a Mg 2ϩ -activated vacuolar H ϩ -ATPase (V-ATPase) on the vesicular membrane (4). When protons are pumped into the vesicular lumen, a proton gradient (⌬pH) and a membrane potential (⌬) occur across the membrane to form ⌬ Hϩ, which favors the exchange of luminal protons for cytoplasmic transmitter. The transport of monoamines and acetylcholine rely predominantly on ...

We sought to identify novel genes involved in intestinal iron absorption by inducing iron deficiency in rats during postnatal development from the suckling period through adulthood. We then performed comparative gene chip analyses (RAE230A and RAE230B chips; Affymetrix) with cRNA derived from duodenal mucosa. Real-time PCR was used to confirm changes in gene expression. Genes encoding the apical iron transport-related proteins [divalent metal transporter 1 (DMT1) and duodenal cytochrome b] were strongly induced at all ages studied, whereas increases in mRNA encoding the basolateral proteins iron-regulated gene 1 and hephaestin were observed only by real-time PCR. In addition, transferrin receptor 1 and heme oxygenase 1 were induced. We also identified induction of novel genes not previously associated with intestinal iron transport. The Menkes copper ATPase (ATP7a) and metallothionein were strongly induced at all ages studied, suggesting increased copper absorption by enterocytes during iron deficiency. We also found significantly increased liver copper levels in 7- to 12-wk-old iron-deficient rats. Also upregulated at most ages examined were the sodium-dependent vitamin C transporter, tripartite motif protein 27, aquaporin 4, lipocalin-interacting membrane receptor, and the breast cancer-resistance protein (ABCG2). Some genes also showed decreased expression with iron deprivation, including several membrane transporters, metabolic enzymes, and genes involved in the oxidative stress response. We speculate that dietary iron deprivation leads to increased intestinal copper absorption via DMT1 on the brush-border membrane and the Menkes copper ATPase on the basolateral membrane. These findings may thus explain copper loading in the iron-deficient state. We also demonstrate that many other novel genes may be differentially regulated in the setting of iron deprivation.