ABSTRACT
The dwarf planet (1) Ceres, the largest object in the main asteroid belt with a mean diameter of about 950 kilometres, is located at a mean distance from the Sun of about 2.8 astronomical units (one astronomical unit is the Earth-Sun distance). Thermal evolution models suggest that it is a differentiated body with potential geological activity. Unlike on the icy satellites of Jupiter and Saturn, where tidal forces are responsible for spewing briny water into space, no tidal forces are acting on Ceres. In the absence of such forces, most objects in the main asteroid belt are expected to be geologically inert. The recent discovery of water vapour absorption near Ceres and previous detection of bound water and OH near and on Ceres (refs 5-7) have raised interest in the possible presence of surface ice. Here we report the presence of localized bright areas on Ceres from an orbiting imager. These unusual areas are consistent with hydrated magnesium sulfates mixed with dark background material, although other compositions are possible. Of particular interest is a bright pit on the floor of crater Occator that exhibits probable sublimation of water ice, producing haze clouds inside the crater that appear and disappear with a diurnal rhythm. Slow-moving condensed-ice or dust particles may explain this haze. We conclude that Ceres must have accreted material from beyond the 'snow line', which is the distance from the Sun at which water molecules condense.
ABSTRACT
BACKGROUND AND OBJECTIVE: Oral submucous fibrosis, a disease of collagen disorder, has been attributed to arecoline present in the saliva of betel quid chewers. However, the molecular basis of the action of arecoline in the pathogenesis of oral submucous fibrosis is poorly understood. The basic aim of our study was to elucidate the mechanism underlying the action of arecoline on the expression of genes in oral fibroblasts. MATERIAL AND METHODS: Human keratinocytes (HaCaT cells) and primary human gingival fibroblasts were treated with arecoline in combination with various pathway inhibitors, and the expression of transforming growth factor-beta isoform genes and of collagen isoforms was assessed using reverse transcription-polymerase chain reaction analysis. RESULTS: We observed the induction of transforming growth factor-beta2 by arecoline in HaCaT cells and this induction was found to be caused by activation of the M-3 muscarinic acid receptor via the induction of calcium and the protein kinase C pathway. Most importantly, we showed that transforming growth factor-beta2 was significantly overexpressed in oral submucous fibrosis tissues (p = 0.008), with a median of 2.13 (n = 21) compared with 0.75 (n = 18) in normal buccal mucosal tissues. Furthermore, arecoline down-regulated the expression of collagens 1A1 and 3A1 in human primary gingival fibroblasts; however these collagens were induced by arecoline in the presence of spent medium of cultured human keratinocytes. Treatment with a transforming growth factor-beta blocker, transforming growth factor-beta1 latency-associated peptide, reversed this up-regulation of collagen, suggesting a role for profibrotic cytokines, such as transforming growth factor-beta, in the induction of collagens. CONCLUSION: Taken together, our data highlight the importance of arecolineinduced epithelial changes in the pathogenesis of oral submucous fibrosis.
Subject(s)
Arecoline/pharmacology , Cholinergic Agonists/pharmacology , Extracellular Matrix/drug effects , Fibroblasts/drug effects , Gingiva/drug effects , Keratinocytes/drug effects , Cell Line , Chelating Agents/pharmacology , Collagen/drug effects , Collagen/genetics , Collagen Type I/drug effects , Collagen Type I/genetics , Collagen Type I, alpha 1 Chain , Collagen Type III/drug effects , Collagen Type III/genetics , Collagen Type IV/drug effects , Collagen Type IV/genetics , Collagen Type VI/drug effects , Collagen Type VI/genetics , Collagen Type VII/drug effects , Collagen Type VII/genetics , Down-Regulation , Egtazic Acid/analogs & derivatives , Egtazic Acid/pharmacology , Extracellular Matrix/genetics , Gingiva/cytology , Humans , Mouth Mucosa/pathology , Oral Submucous Fibrosis/genetics , Oral Submucous Fibrosis/pathology , Protein Isoforms/drug effects , Protein Isoforms/genetics , Protein Kinase C/antagonists & inhibitors , Protein Kinase C/drug effects , Protein Kinase C/genetics , Receptor, Muscarinic M3/drug effects , Receptor, Muscarinic M3/genetics , Staurosporine/pharmacology , Transforming Growth Factor beta1/pharmacology , Transforming Growth Factor beta2/drug effects , Transforming Growth Factor beta2/geneticsABSTRACT
BACKGROUND AND OBJECTIVE: Arecoline, an arecanut alkaloid present in the saliva of betel quid chewers, has been implicated in the pathogenesis of a variety of inflammatory oral diseases, including oral submucous fibrosis and periodontitis. To understand the molecular basis of arecoline action in epithelial changes associated with these diseases, we investigated the effects of arecoline on human keratinocytes with respect to cell growth regulation and the expression of stress-responsive genes. MATERIAL AND METHODS: Human keratinocyte cells (of the HaCaT cell line) were treated with arecoline, following which cell viability was assessed using the Trypan Blue dye-exclusion assay, cell growth and proliferation were analyzed using the MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide) and 5-bromo-2-deoxyuridine incorporation assays, cell cycle arrest and generation of reactive oxygen species were examined using flow cytometry, and gene expression changes were investigated using the reverse transcription-polymerase chain reaction technique. The role of oxidative stress, muscarinic acetylcholine receptor and mitogen-activated protein kinase (MAPK) pathways were studied using specific inhibitors. Western blot analysis was performed to study p38 MAPK activation. RESULTS: Arecoline induced the generation of reactive oxygen species and cell cycle arrest at the G1/G0 phase in HaCaT cells without affecting the expression of p21/Cip1. Arecoline-induced epithelial cell death at higher concentrations was caused by oxidative trauma without eliciting apoptosis. Sublethal concentrations of arecoline upregulated the expression of the following stress-responsive genes: heme oxygenase-1; ferritin light chain; glucose-6-phosphate dehydrogenase; glutamate-cysteine ligase catalytic subunit; and glutathione reductase. Additionally, there was a dose-dependent induction of interleukin-1alfa mRNA by arecoline via oxidative stress and p38 MAPK activation. CONCLUSION: Our data highlight the role of oxidative stress in arecoline-mediated cell death, gene regulation and inflammatory processes in human keratinocytes.
Subject(s)
Arecoline/pharmacology , Cholinergic Agonists/pharmacology , Keratinocytes/drug effects , Oxidative Stress/drug effects , Apoferritins , Cell Cycle/drug effects , Cell Death/drug effects , Cell Line , Cell Proliferation/drug effects , Cell Survival/drug effects , Coloring Agents , Cyclin-Dependent Kinase Inhibitor p21/drug effects , Dose-Response Relationship, Drug , Enzyme Activation/drug effects , Gene Expression Regulation/drug effects , Glucosephosphate Dehydrogenase/drug effects , Glutamate-Cysteine Ligase/drug effects , Glutathione Reductase/drug effects , Heme Oxygenase-1/drug effects , Humans , Interleukin-1alpha/biosynthesis , Mitogen-Activated Protein Kinases/drug effects , Oxidative Stress/genetics , Protein Kinase Inhibitors/metabolism , Reactive Oxygen Species/analysis , Receptors, Muscarinic/drug effects , p38 Mitogen-Activated Protein Kinases/drug effectsABSTRACT
Transglutaminase-2 (TGM-2) stabilizes extracellular matrix (ECM) proteins by cross-linking and has been implicated in several fibrotic disorders. Arecoline present in betel quid has been proposed as one of the causative factors for oral submucous fibrosis (OSMF). Hence, we hypothesize that arecoline may regulate TGM-2 and may have a role in the pathogenesis of OSMF. The expression of TGM-2 was studied in OSMF tissues by real-time RT-PCR analysis, and significant overexpression was observed in most OSMF tissues (P=0.0112) compared with normal tissues. Arecoline induced TGM-2 mRNA and protein expression as well as TGM-2 activity in human gingival fibroblast cells. The addition of methocramine hemihydrate (M-2 muscarinic acetylcholine receptor selective antagonist) or 8'-bromo-cAMP abolished arecoline-mediated TGM-2 induction, suggesting a role for M-2 muscarinic acid receptor and a repressor role for cAMP. Our study provides evidence for TGM-2 overexpression in OSMF and its regulation by arecoline in oral fibroblasts.
