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1.
Exp Dermatol ; 22(10): 677-9, 2013 Oct.
Article in English | MEDLINE | ID: mdl-24079743

ABSTRACT

Keratinocyte sphingolipids are structural elements of epidermal permeability barrier and potential regulators of epidermal functions. We tested the influence of sphingoid bases sphinganine, sphingosine and phytosphingosine on in vitro keratinocyte differentiation. Lipidomic and transcriptomic analysis after treatment emphasizes sphinganine and phytosphingosine as potent modulators of keratinocyte differentiation and lipid metabolism. Sphinganine treatment regulated differentiation and sphingolipid metabolism-related genes, and also increased all major ceramide species. Sphingosine treatment increased ceramide and phytoceramide pools without changes in dihydroceramides. Phytosphingosine treatment markedly increased phytoceramide pools without raising ceramide or dihydroceramide levels. Sphinganine treatment increased specifically very long chain ceramides essential for intact barrier function. In summary, sphingoid bases, especially sphinganine, promote differentiation and ceramide production in keratinocytes. Free sphinganine may serve as a dermatological and cosmetic agent by enhancing formation and maintenance of an intact epidermal lipid barrier, with beneficial effects for skin and hair care applications.


Subject(s)
Cell Differentiation/drug effects , Keratinocytes/cytology , Keratinocytes/drug effects , Skin/drug effects , Sphingolipids/chemistry , Cells, Cultured , Ceramides/chemistry , Cosmetics , Fatty Acids/chemistry , Gene Expression Profiling , Gene Expression Regulation/drug effects , Humans , Lipids/chemistry , Permeability , Skin/metabolism , Sphingosine/analogs & derivatives , Sphingosine/chemistry , Time Factors
2.
J Photochem Photobiol B ; 116: 79-88, 2012 Nov 05.
Article in English | MEDLINE | ID: mdl-22982209

ABSTRACT

Many ATP binding cassette (ABC) transporters are important regulators of lipid homeostasis and have been implicated in keratinocyte lipid transport. Ultraviolet (UV) light exposure is a known epidermal stressor, which amongst other effects causes lipid alterations and defective lamellar body biogenesis. To elucidate the background of these lipid changes we studied the effect of UVB light on ABC transporter expression. The effect of UVB treatment on the levels of 47 known human ABC transporter mRNAs was analyzed in normal human epidermal keratinocytes. Immunoblots and promoter assays were carried out for ABCA1 and ABCG1. The mRNA levels of cholesterol transport regulators ABCA1 and ABCG1 were markedly downregulated by UVB, parallel to the lamellar ichthyosis related glucosylceramide transporter ABCA12 and the suspected sphingosine-1-phosphate and cholesterol sulfate transporter ABCC1. The long but not the short alternative splice variant of the ABCF2 was found to be markedly upregulated rapidly after UVB irradiation. Immunoblot confirmed ABCA1 and ABCG1 protein downregulation, and luciferase assays showed suppression of their promoters by UVB. These proteins mostly transport lipids, which account for the integrity of the epidermal barrier; therefore our findings on the UVB regulation of ABC transporters may explain the appearance of barrier dysfunction after UVB exposure.


Subject(s)
ATP-Binding Cassette Transporters/genetics , Gene Expression Regulation/radiation effects , Keratinocytes/metabolism , Keratinocytes/radiation effects , Ultraviolet Rays/adverse effects , Biological Transport , Cell Differentiation/drug effects , Cell Differentiation/radiation effects , Down-Regulation/drug effects , Down-Regulation/radiation effects , Epidermal Cells , Epidermis/drug effects , Epidermis/metabolism , Epidermis/radiation effects , Gene Expression Regulation/drug effects , Humans , Keratinocytes/cytology , Keratinocytes/drug effects , Lipid Metabolism/drug effects , Lipid Metabolism/radiation effects , Promoter Regions, Genetic/drug effects , Promoter Regions, Genetic/genetics , Promoter Regions, Genetic/radiation effects , RNA, Messenger/genetics , RNA, Messenger/metabolism , Radiation-Protective Agents/pharmacology
3.
Am J Hum Genet ; 89(4): 543-50, 2011 Oct 07.
Article in English | MEDLINE | ID: mdl-21981780

ABSTRACT

The disease classification neurodegeneration with brain iron accumulation (NBIA) comprises a clinically and genetically heterogeneous group of progressive neurodegenerative disorders characterized by brain iron deposits in the basal ganglia. For about half of the cases, the molecular basis is currently unknown. We used homozygosity mapping followed by candidate gene sequencing to identify a homozygous 11 bp deletion in the orphan gene C19orf12. Mutation screening of 23 ideopathic NBIA index cases revealed two mutated alleles in 18 of them, and one loss-of-function mutation is the most prevalent. We also identified compound heterozygous missense mutations in a case initially diagnosed with Parkinson disease at age 49. Psychiatric signs, optic atrophy, and motor axonal neuropathy were common findings. Compared to the most prevalent NBIA subtype, pantothenate kinase associated neurodegeneration (PKAN), individuals with two C19orf12 mutations were older at age of onset and the disease progressed more slowly. A polyclonal antibody against the predicted membrane spanning protein showed a mitochondrial localization. A histopathological examination in a single autopsy case detected Lewy bodies, tangles, spheroids, and tau pathology. The mitochondrial localization together with the immunohistopathological findings suggests a pathomechanistic overlap with common forms of neurodegenerative disorders.


Subject(s)
Brain/metabolism , Iron/metabolism , Mitochondrial Proteins/genetics , Neurodegenerative Diseases/genetics , Adolescent , Adult , Amino Acid Sequence , Case-Control Studies , Child , Child, Preschool , Cloning, Molecular , Cohort Studies , Female , Heterozygote , Homozygote , Humans , Male , Mitochondria/metabolism , Molecular Sequence Data , Mutation , Mutation, Missense , Pedigree , Sequence Homology, Amino Acid
4.
Exp Dermatol ; 19(3): 297-301, 2010 Mar.
Article in English | MEDLINE | ID: mdl-19961536

ABSTRACT

Keratinocyte differentiation plays a pivotal role in the epidermal barrier. Single keratinocyte differentiation genes have already been studied, but many important constituents of this process may have been missed so far. Gene expression profiling by microarray was carried out in cultured normal human epidermal keratinocytes undergoing confluence-induced differentiation to find novel differentiation genes. Candidate gene lists were established and genes of potential dermatological interest were validated by quantitative reverse transcription polymerase chain reaction and immunohistochemical analysis. Some of these points lead to the identification of counter-regulation of heme oxygenase and biliverdin reductase as well as glutaredoxin and glutathione reductase indicative of potential novel redox signaling in differentiating human keratinocytes. Others indicate a strong concert down-regulation of interleukin-1 signaling at previously unidentified levels during keratinocyte differentiation. We believe that identified genes contribute to a more comprehensive understanding of the complicated epidermal differentiation process and lead to better understanding of dermatological diseases.


Subject(s)
Cell Differentiation/genetics , Gene Expression Profiling , Keratinocytes/cytology , Keratinocytes/metabolism , Gene Regulatory Networks , Genome, Human , Humans , In Vitro Techniques , Oligonucleotide Array Sequence Analysis
5.
J Cereb Blood Flow Metab ; 25(1): 30-40, 2005 Jan.
Article in English | MEDLINE | ID: mdl-15678110

ABSTRACT

The transcription factor NF-kappaB is a key regulator of inflammation and cell survival. NF-kappaB is activated by cerebral ischemia in neurons and glia, but its function is controversial. To inhibit NF-kappaB selectively in neurons and glial cells, we have generated transgenic mice that express the IkappaBalpha superrepressor (IkappaBalpha mutated at serine-32 and serine-36, IkappaBalpha-SR) under transcriptional control of the neuron-specific enolase (NSE) and the glial fibrillary acidic protein (GFAP) promoter, respectively. In primary cortical neurons of NSE-IkappaBalpha-SR mice, NF-kappaB activity was partially inhibited. To assess NF-kappaB activity in vivo after permanent middle cerebral artery occlusion (MCAO), we measured the expression of NF-kappaB target genes by real-time polymerase chain reaction (PCR). The induction of c-myc and transforming growth factor-beta2 by cerebral ischemia was inhibited by neuronal expression of IkappaBalpha-SR, whereas induction of GFAP by MCAO was reduced by astrocytic expression of IkappaBalpha-SR. Neuronal, but not astrocytic, expression of the NF-kappaB inhibitor reduced both infarct size and cell death 48 hours after permanent MCAO. In summary, the data show that NF-kappaB is activated in neurons and astrocytes during cerebral ischemia and that NF-kappaB activation in neurons contributes to the ischemic damage.


Subject(s)
Brain Ischemia/metabolism , I-kappa B Proteins/metabolism , NF-kappa B/metabolism , Amino Acid Substitution/genetics , Animals , Astrocytes/metabolism , Astrocytes/pathology , Brain Ischemia/genetics , Brain Ischemia/pathology , Cell Death/genetics , Gene Expression Regulation/genetics , Glial Fibrillary Acidic Protein/genetics , I-kappa B Proteins/genetics , Infarction, Middle Cerebral Artery/metabolism , Infarction, Middle Cerebral Artery/pathology , Mice , Mice, Transgenic , NF-KappaB Inhibitor alpha , Neurons/metabolism , Neurons/pathology , Phosphopyruvate Hydratase/genetics , Point Mutation/genetics , Promoter Regions, Genetic , Proto-Oncogene Proteins c-myc/metabolism , Transcription Factor RelA , Transforming Growth Factor beta/metabolism
6.
Neuropharmacology ; 47(4): 580-92, 2004 Sep.
Article in English | MEDLINE | ID: mdl-15380375

ABSTRACT

Exogenous and endogenous cannabinoids have been shown to have neuroprotective effects in vitro and in vivo. Although many of the pharmacological effects of cannabinoids have been identified, the mechanism of neuroprotection still represents a controversy. Here we demonstrate for the first time protective effects of the synthetic cannabinoid dexanabinol by inhibiting apoptosis in a neuron-like cell line using nuclear staining and FACS analysis and in primary neurons. We provide further evidence of inhibition of nuclear factor-kappakappa B (NF-kappaB) by dexanabinol: Dexanabinol inhibits (1) phosphorylation and degradation of the inhibitor of NF-kappaB IkappaBalpha and translocation of NF-kappaB to the nucleus; dexanabinol reduces (2) the transcriptional activity of NF-kappaB and (3) mRNA accumulation of the NF-kappaB target genes tumor necrosis factor-alpha and interleukin-6 (TNF-alpha and IL-6). Dexanabinol does not bind to cannabinoid (CB) receptors 1 and 2. To investigate the mechanism of action, we employed the non-antioxidant CB1 receptor agonist WIN 55,212-2 and the antioxidant cannabinol, which binds to CB1 receptors only weakly. Both cannabinoids mimicked the effect of dexanabinol on NF-kappaB and apoptosis. This suggests that neither the antioxidant properties of cannabinoids nor binding to CB1 or CB2 receptors are responsible for the inhibition of NF-kappaB activity and apoptosis. Our results clearly demonstrate that dexanabinol inhibits NF-kappaB. NF-kappaB has been shown to be involved in brain damage and to promote neuronal cell death in vitro and in in vivo models of ischemic and neurodegenerative neurological diseases.


Subject(s)
Cannabinoids/pharmacology , Dronabinol/analogs & derivatives , Dronabinol/pharmacology , NF-kappa B/antagonists & inhibitors , Animals , Apoptosis/drug effects , Apoptosis/physiology , Cell Line, Tumor , Dose-Response Relationship, Drug , Humans , Mice , NF-kappa B/metabolism , PC12 Cells , Rats
7.
Naunyn Schmiedebergs Arch Pharmacol ; 369(6): 563-9, 2004 Jun.
Article in English | MEDLINE | ID: mdl-15141332

ABSTRACT

6-Hydroxydopamine (6-OHDA) is widely used to study the death of catecholaminergic cells related to Parkinson's disease. Oxidative stress and gene transcription are known to mediate the pro-apoptotic effect of 6-OHDA. As redox mechanisms are involved in activation of the transcription factor NF-kappaB, we studied the role of NF-kappaB in 6-OHDA-induced death of PC12 cells. We stably transfected PC12 cells with a doxycycline-regulated expression vector for the NF-kappaB super-repressor (IkappaBalpha mutated at serine-32 and serine-36, IkappaBalpha-SR). NF-kappaB transcriptional activity was evaluated by transient transfection of an NF-kappaB-driven luciferase reporter gene. Expression of IkappaBalpha-SR inhibited NF-kappaB stimulated by tumor necrosis factor alpha (TNFalpha) and 6-OHDA. Apoptosis was quantified by counting cells with condensed nuclei. IkappaBalpha-SR inhibited apoptosis induced by 6-OHDA but enhanced apoptosis that was triggered by TNFalpha. The converse effects of NF-kappaB could be due to different target genes that are induced in the context of TNFalpha and 6-OHDA stimulation. Indeed, TNFalpha stimulated mRNA accumulation of the anti-apoptotic superoxide dismutase 2 through NF-kappaB whereas 6-OHDA induced mRNA accumulation of the pro-apoptotic c-myc. These data demonstrate that NF-kappaB regulates survival of the neuron-like PC12 cells in a stimulus-specific manner. In the context of 6-OHDA stimulation, NF-kappaB mediates pro-apoptotic effects, suggesting that NF-kappaB signaling could be a target for drug development in Parkinson-related neurodegeneration.


Subject(s)
Apoptosis/physiology , NF-kappa B/physiology , 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine/pharmacology , Animals , Apoptosis/drug effects , Doxycycline/pharmacology , I-kappa B Proteins/biosynthesis , I-kappa B Proteins/genetics , NF-KappaB Inhibitor alpha , NF-kappa B/antagonists & inhibitors , NF-kappa B/genetics , Oxidopamine , PC12 Cells , Rats , Transfection , Tumor Necrosis Factor-alpha
8.
J Cereb Blood Flow Metab ; 23(4): 406-15, 2003 Apr.
Article in English | MEDLINE | ID: mdl-12679717

ABSTRACT

Although the function of fever is still unclear, it is now beyond doubt that body temperature influences the outcome of brain damage. An elevated body temperature is often found in stroke patients and denotes a bad prognosis. However, the pathophysiologic basis and treatment options of elevated body temperature after stroke are still unknown. Cerebral ischemia rapidly induced neuronal interleukin-6 (IL-6) expression in mice. In IL-6-deficient mice, body temperature was markedly decreased after middle cerebral artery occlusion (MCAO), but infarct size was comparable to that in control mice. If body temperature was controlled by external warming after MCAO, IL-6-deficient mice had a reduced survival, worse neurologic status, and larger infarcts than control animals. In cell culture, IL-6 exerted an antiapoptotic and neuroprotective effect. These data suggest that IL-6 is a key regulator of body temperature and an endogenous neuroprotectant in cerebral ischemia. Neuroprotective properties apparently compensate for its pyretic action after MCAO and enhance the safety of this endogenous pyrogen.


Subject(s)
Body Temperature Regulation/physiology , Brain Ischemia/physiopathology , Interleukin-6/genetics , Interleukin-6/metabolism , Animals , Behavior, Animal , Female , Gene Expression/physiology , Infarction, Middle Cerebral Artery/physiopathology , Male , Mice , Mice, Inbred C57BL , Mice, Mutant Strains , Motor Activity , PC12 Cells , Poly(ADP-ribose) Polymerases/metabolism , RNA, Messenger/analysis , Rats
9.
Neuroscientist ; 8(3): 268-75, 2002 Jun.
Article in English | MEDLINE | ID: mdl-12061506

ABSTRACT

IL-6 and its receptor(s) are found in the CNS in health and disease. Cellular sources are glial cells and neurons. Glial production of IL-6 has intensively been studied, but comparatively little is known about the induction of IL-6 in neurons. Emerging evidence suggests that IL-6 possesses neurotrophic properties. Recent data show that neuronal IL-6 expression is induced by excitatory amino acids or membrane depolarization. This implicates that IL-6 is produced not only under pathological conditions but may play a critical role as a physiological neuromodulator that is induced by neuronal activity and regulates brain functions. In the following article, the authors review the current data on IL-6 expression in neurons, with special reference to the induction of IL-6 by neuronal activity. They discuss its direct and indirect effects as a neuromodulator and speculate about the possible function of IL-6 as a physiological regulatory molecule and as a neuroprotective agent in brain pathology.


Subject(s)
Interleukin-6/physiology , Neurons/physiology , Receptors, Interleukin-6/physiology , Animals , Brain Ischemia/metabolism , Depression/metabolism , Excitatory Amino Acids/physiology , Fever/metabolism , Interleukin-6/metabolism , Membrane Potentials , Nerve Growth Factors/physiology , Neuroglia/physiology , Neurons/metabolism , Neuroprotective Agents/metabolism , Receptors, Interleukin-6/metabolism
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