ABSTRACT
BACKGROUND: 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is a potent activator of the aryl hydrocarbon receptor (AhR) and causes chloracne in humans. The pathogenesis and role of AhR in chloracne remains incompletely understood. OBJECTIVE: To elucidate the mechanisms contributing to the development of the chloracne-like phenotype in a human epidermal equivalent model and identify potential biomarkers. METHODS: Using primary normal human epidermal keratinocytes (NHEK), we studied AhR activation by XRE-luciferase, AhR degradation and CYP1A1 induction. We treated epidermal equivalents with high affinity TCDD or two non-chloracnegens: ß-naphthoflavone (ß-NF) and 2-(1'H-indole-3'-carbonyl)-thiazole-4-carboxylic acid methyl ester (ITE). Using Western blotting and immunochemistry for filaggrin (FLG), involucrin (INV) and transglutaminase-1 (TGM-1), we compared the effects of the ligands on keratinocyte differentiation and development of the chloracne-like phenotype by H&E. RESULTS: In NHEKs, activation of an XRE-luciferase and CYP1A1 protein induction correlated with ligand binding affinity: TCDD>ß-NF>ITE. AhR degradation was induced by all ligands. In epidermal equivalents, TCDD induced a chloracne-like phenotype, whereas ß-NF or ITE did not. All three ligands induced involucrin and TGM-1 protein expression in epidermal equivalents whereas FLG protein expression decreased following treatment with TCDD and ß-NF. Inhibition of AhR by α-NF blocked TCDD-induced AhR activation in NHEKs and blocked phenotypic changes in epidermal equivalents; however, AhR knock down did not reproduce the phenotype. CONCLUSION: Ligand-induced CYP1A1 and AhR degradation did not correlate with their chloracnegenic potential, indicating that neither CYP1A1 nor AhR are suitable biomarkers. Mechanistic studies showed that the TCDD-induced chloracne-like phenotype depends on AhR activation whereas AhR knock down did not appear sufficient to induce the phenotype.
Subject(s)
Basic Helix-Loop-Helix Transcription Factors/agonists , Chloracne/etiology , Epidermis/drug effects , Gene Knockdown Techniques , Keratinocytes/drug effects , Polychlorinated Dibenzodioxins/toxicity , Receptors, Aryl Hydrocarbon/agonists , Basic Helix-Loop-Helix Transcription Factors/genetics , Basic Helix-Loop-Helix Transcription Factors/metabolism , Cell Differentiation/drug effects , Cells, Cultured , Chloracne/genetics , Chloracne/metabolism , Chloracne/pathology , Cytochrome P-450 CYP1A1/biosynthesis , Dose-Response Relationship, Drug , Enzyme Induction , Epidermis/metabolism , Epidermis/pathology , Filaggrin Proteins , Humans , Indoles/toxicity , Intermediate Filament Proteins/metabolism , Keratinocytes/metabolism , Keratinocytes/pathology , Ligands , Phenotype , Protein Precursors/metabolism , RNA Interference , Receptors, Aryl Hydrocarbon/genetics , Receptors, Aryl Hydrocarbon/metabolism , Thiazoles/toxicity , Transfection , Transglutaminases/metabolism , beta-Naphthoflavone/toxicityABSTRACT
Oral administration of buprenorphine is becoming a popular method of providing analgesia for laboratory rodents. The mixing of buprenorphine with flavoured jello, which rodents find palatable, is becoming a commonly used method as it is thought to improve the efficacy of oral buprenorphine by increasing the time available for it to be absorbed via the oral mucosa. The aim of this study was to assess the effect of various methods of buprenorphine administration (subcutaneous saline, subcutaneous buprenorphine [0.05 mg/kg], buprenorphine gavage [0.5 mg/kg], buprenorphine in jello [0.5 mg/kg] and buprenorphine in golden syrup [0.5 mg/kg]) on thermal antinociceptive thresholds in laboratory rats. Buprenorphine administered subcutaneously, by gavage, in jello and in syrup induced significant increases in thermal antinociceptive thresholds compared with saline. This effect was observed up to 5 h postadministration for buprenorphine administered subcutaneously and by gavage, but only for one hour postadministration for buprenorphine administered in jello and in syrup.
