Connective tissue growth factor and integrin αvß6: a new pair of regulators critical for ductular reaction and biliary fibrosis in mice.

UNLABELLED: Connective tissue growth factor (CTGF) is a matricellular protein that mediates cell-matrix interaction through various subtypes of integrin receptors. This study investigated the role of CTGF and integrin αvß6 in hepatic progenitor/oval cell activation, which often occurs in the form of ductular reactions (DRs) when hepatocyte proliferation is inhibited during severe liver injury. CTGF and integrin αvß6 proteins were highly expressed in DRs of human cirrhotic livers and cholangiocarcinoma. Confocal microscopy analysis of livers from Ctgf promoter-driven green fluorescent protein reporter mice suggested that oval cells and cholangiocytes were the main sources of CTGF and integrin αvß6 during liver injury induced by 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC). Deletion of exon 4 of the Ctgf gene using tamoxifen-inducible Cre-loxP system down-regulated integrin αvß6 in DDC-damaged livers of knockout mice. Ctgf deficiency or inhibition of integrin αvß6, by administrating the neutralizing antibody, 6.3G9 (10 mg/kg body weight), caused low levels of epithelial cell adhesion molecule and cytokeratin 19 gene messenger RNAs. Also, there were smaller oval cell areas, fewer proliferating ductular epithelial cells, and lower cholestasis serum markers within 2 weeks after DDC treatment. Associated fibrosis was attenuated, as indicated by reduced expression of fibrosis-related genes, smaller areas of alpha-smooth muscle actin staining, and low collagen production based on hydroxyproline content and Sirius Red staining. Finally, integrin αvß6 could bind to CTGF mediating oval cell adhesion to CTGF and fibronection substrata and promoting transforming growth factor (TGF)-ß1 activation in vitro. CONCLUSIONS: CTGF and integrin αvß6 regulate oval cell activation and fibrosis, probably through interacting with their common matrix and signal partners, fibronectin and TGF-ß1. CTGF and integrin αvß6 are potential therapeutic targets to control DRs and fibrosis in related liver disease.

Modulation of taste responsiveness by the satiation hormone peptide YY.

It has been hypothesized that the peripheral taste system may be modulated in the context of an animal's metabolic state. One purported mechanism for this phenomenon is that circulating gastrointestinal peptides modulate the functioning of the peripheral gustatory system. Recent evidence suggests endocrine signaling in the oral cavity can influence food intake (FI) and satiety. We hypothesized that these hormones may be affecting FI by influencing taste perception. We used immunohistochemistry along with genetic knockout models and the specific reconstitution of peptide YY (PYY) in saliva using gene therapy protocols to identify a role for PYY signaling in taste. We show that PYY is expressed in subsets of taste cells in murine taste buds. We also show, using brief-access testing with PYY knockouts, that PYY signaling modulates responsiveness to bitter-tasting stimuli, as well as to lipid emulsions. We show that salivary PYY augmentation, via viral vector therapy, rescues behavioral responsiveness to a lipid emulsion but not to bitter stimuli and that this response is likely mediated via activation of Y2 receptors localized apically in taste cells. Our findings suggest distinct functions for PYY produced locally in taste cells vs. that circulating systemically.

Neuronal localization of indoleamine 2,3-dioxygenase in mice.

Indoleamine 2,3-dioxygenase (IDO) catabolizes tryptophan to kynurenine. In the immune system, the reduction in tryptophan and increase in kynurenine act to suppress T-cell function. In the nervous system, kynurenine can be further metabolized to quinolinic acid, which can be neurotoxic. IDO is known to be expressed by microglia and its levels are upregulated by interferon-gamma (IFNgamma). We report here that IDO immunoreactivity is also localized in neurons, and that IDO is upregulated by IFNgamma in neurons of the hippocampus. Thus, neuronal IDO could contribute to the vulnerability of neurons to inflammatory conditions.

Repurposed biological scaffolds: kidney to pancreas.

Advances in organ regeneration have been facilitated by gentle decellularization protocols that maintain distinct tissue compartments, and thereby allow seeding of blood vessels with endothelial lineages separate from populations of the parenchyma with tissue-specific cells. We hypothesized that a reconstituted vasculature could serve as a novel platform for perfusing cells derived from a different organ: thus discordance of origin between the vascular and functional cells, leading to a hybrid repurposed organ. The need for a highly vascular bed is highlighted by tissue engineering approaches that involve transplantation of just cells, as attempted for insulin production to treat human diabetes. Those pancreatic islet cells present unique challenges since large numbers are needed to allow the cell-to-cell signaling required for viability and proper function; however, increasing their number is limited by inadequate perfusion and hypoxia. As proof of principle of the repurposed organ methodology we harnessed the vasculature of a kidney scaffold while seeding the collecting system with insulin-producing cells. Pig kidneys were decellularized by sequential detergent, enzymatic and rinsing steps. Maintenance of distinct vascular and collecting system compartments was demonstrated by both fluorescent 10 micron polystyrene microspheres and cell distributions in tissue sections. Sterilized acellular scaffolds underwent seeding separately via the artery (fibroblasts or endothelioma cells) and retrograde (murine ßTC-tet cells) up the ureter. After three-day bioreactor incubation, histology confirmed separation of cells in the vasculature from those in the collecting system. ßTC-tet clusters survived in tubules, glomerular Bowman's space, demonstrated insulin immunolabeling, and thereby supported the feasibility of kidney-to-pancreas repurposing.

Distribution of Y-receptors in murine lingual epithelia.

Peptide hormones and their cognate receptors belonging to neuropeptide Y (NPY) family mediate diverse biological functions in a number of tissues. Recently, we discovered the presence of the gut satiation peptide YY (PYY) in saliva of mice and humans and defined its role in the regulation of food intake and body weight maintenance. Here we report the systematic analysis of expression patterns of all NPY receptors (Rs), Y1R, Y2R, Y4R, and Y5R in lingual epithelia in mice. Using four independent assays, immunohistochemistry, in situ hybridization, immunocytochemistry and RT PCR, we show that the morphologically different layers of the keratinized stratified epithelium of the dorsal layer of the tongue express Y receptors in a very distinctive yet overlapping pattern. In particular, the monolayer of basal progenitor cells expresses both Y1 and Y2 receptors. Y1Rs are present in the parabasal prickle cell layer and the granular layer, while differentiated keratinocytes display abundant Y5Rs. Y4Rs are expressed substantially in the neuronal fibers innervating the lamina propria and mechanoreceptors. Basal epithelial cells positive for Y2Rs respond robustly to PYY(3-36) by increasing intracellular Ca(2+) suggesting their possible functional interaction with salivary PYY. In taste buds of the circumvallate papillae, some taste receptor cells (TRCs) express YRs localized primarily at the apical domain, indicative of their potential role in taste perception. Some of the YR-positive TRCs are co-localized with neuronal cell adhesion molecule (NCAM), suggesting that these TRCs may have synaptic contacts with nerve terminals. In summary, we show that all YRs are abundantly expressed in multiple lingual cell types, including epithelial progenitors, keratinocytes, neuronal dendrites and TRCs. These results suggest that these receptors may be involved in the mediation of a wide variety of functions, including proliferation, differentiation, motility, taste perception and satiation.