Publications by authors named "Ralph Feltens"

23 Publications

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mediates the impact of prenatal bisphenol A exposure on long-term body weight development.

Clin Epigenetics 2018 20;10:58. Epub 2018 Apr 20.

1Department of Environmental Immunology, Helmholtz Centre for Environmental Research (UFZ), Leipzig, Germany.

Background: Exposure to endocrine-disrupting chemicals can alter normal physiology and increase susceptibility to non-communicable diseases like obesity. Especially the prenatal and early postnatal period is highly vulnerable to adverse effects by environmental exposure, promoting developmental reprogramming by epigenetic alterations. To obtain a deeper insight into the role of prenatal bisphenol A (BPA) exposure in children's overweight development, we combine epidemiological data with experimental models and BPA-dependent DNA methylation changes.

Methods: BPA concentrations were measured in maternal urine samples of the LINA mother-child-study obtained during pregnancy ( = 552), and BPA-associated changes in cord blood DNA methylation were analyzed by Illumina Infinium HumanMethylation450 BeadChip arrays ( = 472). Methylation changes were verified by targeted MassARRAY analyses, assessed for their functional translation by qPCR and correlated with children's body mass index (BMI) scores at the age of 1 and 6 years. Further, female BALB/c mice were exposed to BPA from 1 week before mating until delivery, and weight development of their pups was monitored ( ≥ 8/group). Additionally, human adipose-derived mesenchymal stem cells were treated with BPA during the adipocyte differentiation period and assessed for exposure-related epigenetic, transcriptional and morphological changes ( = 4).

Results: In prenatally BPA-exposed children two CpG sites with deviating cord blood DNA-methylation profiles were identified, among them a hypo-methylated CpG in the promoter of the obesity-associated mesoderm-specific transcript (. A mediator analysis suggested that prenatal BPA exposure was connected to cord blood promoter methylation and expression as well as BMI scores in early infancy. This effect could be confirmed in mice in which prenatal BPA exposure altered promoter methylation and transcription with a concomitant increase in the body weight of the juvenile offspring. An experimental model of in vitro differentiated human mesenchymal stem cells also revealed an epigenetically induced expression and enhanced adipogenesis following BPA exposure.

Conclusions: Our study provides evidence that mediates the impact of prenatal BPA exposure on long-term body weight development in offspring by triggering adipocyte differentiation.
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http://dx.doi.org/10.1186/s13148-018-0478-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5910578PMC
May 2019

Maternal phthalate exposure promotes allergic airway inflammation over 2 generations through epigenetic modifications.

J Allergy Clin Immunol 2018 02 6;141(2):741-753. Epub 2017 Apr 6.

Department of Environmental Immunology, UFZ-Helmholtz Centre for Environmental Research Leipzig-Halle, Leipzig, Germany; Department of Dermatology, Venerology and Allergology, Leipzig University Medical Center, Leipzig, Germany. Electronic address:

Background: Prenatal and early postnatal exposures to environmental factors are considered responsible for the increasing prevalence of allergic diseases. Although there is some evidence for allergy-promoting effects in children because of exposure to plasticizers, such as phthalates, findings of previous studies are inconsistent and lack mechanistic information.

Objective: We investigated the effect of maternal phthalate exposure on asthma development in subsequent generations and their underlying mechanisms, including epigenetic alterations.

Methods: Phthalate metabolites were measured within the prospective mother-child cohort Lifestyle and Environmental Factors and Their Influence on Newborns Allergy Risk (LINA) and correlated with asthma development in the children. A murine transgenerational asthma model was used to identify involved pathways.

Results: In LINA maternal urinary concentrations of mono-n-butyl phthalate, a metabolite of butyl benzyl phthalate (BBP), were associated with an increased asthma risk in the children. Using a murine transgenerational asthma model, we demonstrate a direct effect of BBP on asthma severity in the offspring with a persistently increased airway inflammation up to the F2 generation. This disease-promoting effect was mediated by BBP-induced global DNA hypermethylation in CD4 T cells of the offspring because treatment with a DNA-demethylating agent alleviated exacerbation of allergic airway inflammation. Thirteen transcriptionally downregulated genes linked to promoter or enhancer hypermethylation were identified. Among these, the GATA-3 repressor zinc finger protein 1 (Zfpm1) emerged as a potential mediator of the enhanced susceptibility for T2-driven allergic asthma.

Conclusion: These data provide strong evidence that maternal BBP exposure increases the risk for allergic airway inflammation in the offspring by modulating the expression of genes involved in T2 differentiation through epigenetic alterations.
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http://dx.doi.org/10.1016/j.jaci.2017.03.017DOI Listing
February 2018

Prenatal phthalate exposure associates with low regulatory T-cell numbers and atopic dermatitis in early childhood: Results from the LINA mother-child study.

J Allergy Clin Immunol 2017 04 5;139(4):1376-1379.e8. Epub 2016 Nov 5.

Department of Environmental Immunology, UFZ - Helmholtz Centre for Environmental Research Leipzig, Leipzig, Germany. Electronic address:

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http://dx.doi.org/10.1016/j.jaci.2016.09.034DOI Listing
April 2017

Di-(2-Ethylhexyl)-Phthalate (DEHP) Causes Impaired Adipocyte Function and Alters Serum Metabolites.

PLoS One 2015 2;10(12):e0143190. Epub 2015 Dec 2.

IFB AdiposityDiseases, University of Leipzig, Leipzig, Germany.

Di-(2-ethylhexyl)-phthalate (DEHP), an ubiquitous environmental contaminant, has been shown to cause adverse effects on glucose homeostasis and insulin sensitivity in epidemiological studies, but the underlying mechanisms are still unknown. We therefore tested the hypothesis that chronic DEHP exposure causes impaired insulin sensitivity, affects body weight, adipose tissue (AT) function and circulating metabolic parameters of obesity resistant 129S6 mice in vivo. An obesity-resistant mouse model was chosen to reduce a potential obesity bias of DEHP effects on metabolic parameters and AT function. The metabolic effects of 10-weeks exposure to DEHP were tested by insulin tolerance tests and quantitative assessment of 183 metabolites in mice. Furthermore, 3T3-L1 cells were cultured with DEHP for two days, differentiated into mature adipocytes in which the effects on insulin stimulated glucose and palmitate uptake, lipid content as well as on mRNA/protein expression of key adipocyte genes were investigated. We observed in female mice that DEHP treatment causes enhanced weight gain, fat mass, impaired insulin tolerance, changes in circulating adiponectin and adipose tissue Pparg, adiponectin and estrogen expression. Serum metabolomics indicated a general increase in phospholipid and carnitine concentrations. In vitro, DEHP treatment increases the proliferation rate and alters glucose uptake in adipocytes. Taken together, DEHP has significant effects on adipose tissue (AT) function and alters specific serum metabolites. Although, DEHP treatment led to significantly impaired insulin tolerance, it did not affect glucose tolerance, HOMA-IR, fasting glucose, insulin or triglyceride serum concentrations. This may suggest that DEHP treatment does not cause impaired glucose metabolism at the whole body level.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0143190PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4668085PMC
June 2016

Species determination of Culicoides biting midges via peptide profiling using matrix-assisted laser desorption ionization mass spectrometry.

Parasit Vectors 2014 Aug 24;7:392. Epub 2014 Aug 24.

Department of Proteomics, Helmholtz-Centre for Environmental Research-UFZ, 04318 Leipzig, Germany.

Background: Culicoides biting midges are vectors of bluetongue and Schmallenberg viruses that inflict large-scale disease epidemics in ruminant livestock in Europe. Methods based on morphological characteristics and sequencing of genetic markers are most commonly employed to differentiate Culicoides to species level. Proteomic methods, however, are also increasingly being used as an alternative method of identification. These techniques have the potential to be rapid and may also offer advantages over DNA-based techniques. The aim of this proof-of-principle study was to develop a simple MALDI-MS based method to differentiate Culicoides from different species by peptide patterns with the additional option of identifying discriminating peptides.

Methods: Proteins extracted from 7 Culicoides species were digested and resulting peptides purified. Peptide mass fingerprint (PMF) spectra were recorded using matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF-MS) and peak patterns analysed in R using the MALDIquant R package. Additionally, offline liquid chromatography (LC) MALDI-TOF tandem mass spectrometry (MS/MS) was applied to determine the identity of peptide peaks in one exemplary MALDI spectrum obtained using an unfractionated extract.

Results: We showed that the majority of Culicoides species yielded reproducible mass spectra with peak patterns that were suitable for classification. The dendrogram obtained by MS showed tentative similarities to a dendrogram generated from cytochrome oxidase I (COX1) sequences. Using offline LC-MALDI-TOF-MS/MS we determined the identity of 28 peptide peaks observed in one MALDI spectrum in a mass range from 1.1 to 3.1 kDa. All identified peptides were identical to other dipteran species and derived from one of five highly abundant proteins due to an absence of available Culicoides data.

Conclusion: Shotgun mass mapping by MALDI-TOF-MS has been shown to be compatible with morphological and genetic identification of specimens. Furthermore, the method performs at least as well as an alternative approach based on MS spectra of intact proteins, thus establishing the procedure as a method in its own right, with the additional option of concurrently using the same samples in other MS-based applications for protein identifications. The future availability of genomic information for different Culicoides species may enable a more stringent peptide detection based on Culicoides-specific sequence information.
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http://dx.doi.org/10.1186/1756-3305-7-392DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4158057PMC
August 2014

Chlorinated benzenes cause concomitantly oxidative stress and induction of apoptotic markers in lung epithelial cells (A549) at nonacute toxic concentrations.

J Proteome Res 2011 Feb 21;10(2):363-78. Epub 2010 Dec 21.

Department of Proteomics, Helmholtz Centre for Environmental Research-UFZ, Leipzig, Germany.

In industrialized countries, people spend more time indoors and are therefore increasingly exposed to volatile organic compounds that are emitted at working places and from consumer products, paintings, and furniture, with chlorobenzene (CB) and 1,2-dichlorobenzene (DCB) being representatives of the halogenated arenes. To unravel the molecular effects of low concentrations typical for indoor and occupational exposure, we exposed human lung epithelial cells to CB and DCB and analyzed the effects on the proteome level by 2-D DIGE, where 860 protein spots were detected. A set of 25 and 30 proteins were found to be significantly altered due to exposure to environmentally relevant concentrations of 10(-2) g/m(3) of CB or 10(-3) g/m(3) of DCB (2.2 and 0.17 ppm), respectively. The most enriched pathways were cell death signaling, oxidative stress response, protein quality control, and metabolism. The involvement of oxidative stress was validated by ROS measurement. Among the regulated proteins, 28, for example, voltage-dependent anion-selective channel protein 2, PDCD6IP protein, heat shock protein beta-1, proliferating cell nuclear antigen, nucleophosmin, seryl-tRNA synthetase, prohibitin, and protein arginine N-methyltransferase 1, could be correlated with the molecular pathway of cell death signaling. Caspase 3 activation by cleavage was confirmed for both CB and DCB by immunoblotting. Treatment with CB or DCB also caused differential protein phosphorylation, for example, at the proteins HNRNP C1/C2, serine-threonine receptor associated protein, and transaldolase 1. Compared to previous results, where cells were exposed to styrene, for the chlorinated aromatic substances besides oxidative stress, apoptosis was found as the predominant cellular response mechanism.
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http://dx.doi.org/10.1021/pr1005718DOI Listing
February 2011

Analysis of differentially expressed proteins in oral squamous cell carcinoma by MALDI-TOF MS.

J Oral Pathol Med 2011 May 17;40(5):369-79. Epub 2010 Dec 17.

Department of Otorhinolaryngology, Head and Neck Surgery, University Medical Centre of the Johannes Gutenberg University Mainz, Germany.

Purpose: To explore the presence of differentially expressed proteins in OSCC for discrimination of tumour and normal mucosa to establish potential biomarkers and therapeutic targets.

Experimental Design: Paired protein samples of 12 individuals (tongue cancer and non-cancerous mucosa) were separated by two-dimensional polyacrylamid gel electrophoresis. The protein patterns were compared pairwise and protein spots were quantified. We identified about 70 regulated proteins which we subsequently identified by MALDI-TOF mass spectrometry.

Results: Cancerous and non-cancerous tissues could be most precisely distinguished by a panel of proteins. They include the heat shock proteins (hsp)70 and 90, keratins (ck) 5, 6, 13, 14, 16, 17 and 19, beta globin, alpha-2-actin, stratifin, tropomyosin, calreticulin precursor, beta-2-tubulin, galectin7, thioredoxin, involucrin, adenylyl-cyclase-associated protein, disulfide isomerase-associated protein, thyrosine 3-monooxygenase, MYL2 and the s100 calcium binding protein. MYL3, cardiac muscle alpha actin 1 proprotein and transferrin were under-represented in OSCC. Six biomarkers, ck6 und ck13, beta globin, alpha-2-actin, hsp70 and hsp90 discriminated best between cancerous and non-cancerous oral tissues. All over-expressed proteins were analysed by STRING-analysis to highlight experimentally determined and computationally predicted interactions between the proteins. Especially involucrin, hsp70, calreticulin precursor, stratifin, (ck) 5, 6, 14, 19, tyrosine 3-monooxygenase, beta-2-tubulin and disulfide isomerase associated protein showed multiple relations.

Conclusion: We identified six proteins which are differentially expressed in most OSCC compared to healthy tissues. Of those, by string analysis, multiple interaction partners are assumed for hsp70. This protein is supposed to be the most promising candidate as marker molecule and target for OSCC therapy.
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http://dx.doi.org/10.1111/j.1600-0714.2010.00982.xDOI Listing
May 2011

Discrimination of different species from the genus Drosophila by intact protein profiling using matrix-assisted laser desorption ionization mass spectrometry.

BMC Evol Biol 2010 Apr 7;10:95. Epub 2010 Apr 7.

Department of Dermatology, Venerology and Allergology, Medical Faculty of the Leipzig University, Leipzig, Germany.

Background: The use of molecular biology-based methods for species identification and establishing phylogenetic relationships has supplanted traditional methods relying on morphological characteristics. While PCR-based methods are now the commonly accepted gold standards for these types of analysis, relatively high costs, time-consuming assay development or the need for a priori information about species-specific sequences constitute major limitations. In the present study, we explored the possibility to differentiate between 13 different species from the genus Drosophila via a molecular proteomic approach.

Results: After establishing a simple protein extraction procedure and performing matrix-assisted laser desorption/ionization (MALDI) mass spectrometry (MS) with intact proteins and peptides, we could show that most of the species investigated reproducibly yielded mass spectra that were adequate for species classification. Furthermore, a dendrogram generated by cluster analysis of total protein patterns agrees reasonably well with established phylogenetic relationships.

Conclusion: Considering the intra- and interspecies similarities and differences between spectra obtained for specimens of closely related Drosophila species, we estimate that species typing of insects and possibly other multicellular organisms by intact protein profiling (IPP) can be established successfully for species that diverged from a common ancestor about 3 million years ago.
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http://dx.doi.org/10.1186/1471-2148-10-95DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2858148PMC
April 2010

Proteome changes in human bronchoalveolar cells following styrene exposure indicate involvement of oxidative stress in the molecular-response mechanism.

Proteomics 2009 Nov;9(21):4920-33

Department of Proteomics, UFZ, Helmholtz-Centre for Environmental Research, Leipzig, Germany.

Styrene is a volatile organic compound that is widely used as an intermediate in many industrial settings. There are known adverse health effects at environmentally significant concentrations, but little is known about the molecular effect of exposure to styrene at sub-acute toxic concentrations. We exposed human lung epithelial cells, at a wide range of concentrations (1 mg/m(3)-10 g/m(3)), to styrene and analyzed the effects on the proteome level by 2-DE, where 1380 proteins spots were detected and 266 were identified unambiguously by MS. A set of 16 protein spots were found to be significantly altered due to exposure to styrene at environmentally significant concentrations of 1-10 mg/m(3) (0.2-2.3 ppm). Among these, superoxide dismutase as well as biliverdin reductase A could be correlated with the molecular pathway of oxidative stress, while eukaryotic translation initiation factor 5A-1, ezrin, lamin B2 and voltage-dependent anion channel 2 have been reported to be involved in apoptosis. Treatment with styrene also caused the formation of styrene oxide-protein adducts, specifically for thioredoxin reductase 1. These results underline the relevance of oxidative stress as a primary molecular response mechanism of lung epithelial cells to styrene exposure at indoor-relevant concentrations.
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http://dx.doi.org/10.1002/pmic.200800836DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3463241PMC
November 2009

Chlorobenzene induces oxidative stress in human lung epithelial cells in vitro.

Toxicol Appl Pharmacol 2010 Jan 2;242(1):100-8. Epub 2009 Oct 2.

UFZ- Helmholtz Centre for Environmental Research, Department of Environmental Immunology, Permoserstrasse 15, D-04318 Leipzig, Germany.

Chlorobenzene is a volatile organic compound (VOC) that is widely used as a solvent, degreasing agent and chemical intermediate in many industrial settings. Occupational studies have shown that acute and chronic exposure to chlorobenzene can cause irritation of the mucosa of the upper respiratory tract and eyes. Using in vitro assays, we have shown in a previous study that human bronchial epithelial cells release inflammatory mediators such as the cytokine monocyte chemoattractant protein-1 (MCP-1) in response to chlorobenzene. This response is mediated through the NF-kappaB signaling pathway. Here, we investigated the effects of monochlorobenzene on human lung cells, with emphasis on potential alterations of the redox equilibrium to clarify whether the chlorobenzene-induced inflammatory response in lung epithelial cells is caused via an oxidative stress-dependent mechanism. We found that expression of cellular markers for oxidative stress, such as heme oxygenase 1 (HO-1), glutathione S-transferase pi1 (GSTP1), superoxide dismutase 1 (SOD1), prostaglandin-endoperoxide synthase 2 (PTGS2) and dual specificity phosphatase 1 (DUSP1), were elevated in the presence of monochlorobenzene. Likewise, intracellular reactive oxygen species (ROS) were increased in response to exposure. However, in the presence of the antioxidants N-(2-mercaptopropionyl)-glycine (MPG) or bucillamine, chlorobenzene-induced upregulation of marker proteins and release of the inflammatory mediator MCP-1 are suppressed. These results complement our previous findings and point to an oxidative stress-mediated inflammatory response following chlorobenzene exposure.
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http://dx.doi.org/10.1016/j.taap.2009.09.020DOI Listing
January 2010

Noise exposure alters cyclooxygenase 1 (COX-1) and 5-lipoxygenase (5-LO) expression in the guinea pig cochlea.

Acta Otolaryngol 2010 Mar;130(3):358-65

Department of Otolaryngology, Head and Neck Surgery, Johannes Gutenberg University of Mainz, Medical School, Langenbeckstrasse 1, 55131 Mainz, Germany.

Conclusion: Changes in the metabolism of arachidonic acid (AA) might be part of a noise-induced compensatory mechanism with regional specificity.

Objectives: The released imbalance of prostaglandins and leukotrienes, both AA metabolites, might result in altered blood flow regulation in the inner ear and probably contributes to noise-induced hearing loss. The aim of this study was to gain further information about noise-dependent changes in AA metabolism in the mammalian cochlea.

Methods: In this prospective animal study, 10 male guinea pigs were exposed to tone bursts for 1 h at 70 dB sound pressure level (SPL) (n = 5) or 90 dB SPL (n = 5). Five animals were used as controls. Alterations in cyclooxygenase 1 (COX-1) and 5-lipoxygenase (5-LO) expression were determined by quantitative immunohistochemical analysis in 11 cochlear regions.

Results: COX-1 expression was decreased after both 70 dB SPL and 90 dB SPL exposure in most cell types of the organ of Corti and increased in the nerve fibers of the osseous spiral lamina. 5-LO was lowered after 90 dB SPL exposure, preferentially in the third cochlear turn in the organ of Corti, in the first and second turn in spiral ganglion cells, and in all turns in the stria vascularis.
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http://dx.doi.org/10.1080/00016480903168066DOI Listing
March 2010

Styrene induces an inflammatory response in human lung epithelial cells via oxidative stress and NF-kappaB activation.

Toxicol Appl Pharmacol 2008 Sep 29;231(2):241-7. Epub 2008 Apr 29.

UFZ-Helmholtz Centre for Environmental Research, Department of Environmental Immunology, Permoserstrasse 15, D-04318 Leipzig, Germany.

Styrene is a volatile organic compound (VOC) that is widely used as a solvent in many industrial settings. Chronic exposure to styrene can result in irritation of the mucosa of the upper respiratory tract. Contact of styrene with epithelial cells stimulates the expression of a variety of inflammatory mediators, including the chemotactic cytokine monocyte chemoattractant protein-1 (MCP-1). To characterise the underlying mechanisms of the induction of inflammatory signals by styrene, we investigated the influence of this compound on the induction of oxidative stress and the activation of the nuclear factor-kappa B (NF-kappaB) signalling pathway in human lung epithelial cells (A549). The results demonstrate that styrene-induced MCP-1 expression, as well as the expression of the oxidative stress marker glutathione S-transferase (GST), is associated with a concentration dependent pattern of NF-kappaB activity. An inhibitor of NF-kappaB, IKK-NBD, and the anti-inflammatory antioxidant N-acetylcysteine (NAC) were both effective in suppressing styrene-induced MCP-1 secretion. In addition, NAC was capable of inhibiting the upregulation of GST expression. Our findings suggest that the activation of the NF-kappaB signalling pathway by styrene is mediated via a redox-sensitive mechanism.
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http://dx.doi.org/10.1016/j.taap.2008.04.010DOI Listing
September 2008

Fast alterations of vascular endothelial growth factor (VEGF) expression and that of its receptors (Flt-1, Flk-1 and Neuropilin) in the cochlea of guinea pigs after moderate noise exposure.

Eur Arch Otorhinolaryngol 2007 Feb 10;264(2):121-8. Epub 2006 Oct 10.

Department of Otorhinolaryngology, Mainz Medical School, Mainz, Germany.

Vascular endothelial growth factor (VEGF) is a vascular permeability regulating, proangiogenic factor with neuroprotective properties. Its expression in the inner ear has been demonstrated, but little is known concerning its subcellular distribution or potential involvement in sound perception and adaptation to noise. Therefore, we determined the expression patterns and levels of VEGF and the three VEGF-receptors FLK, FLT and Neuropilin in the cochlea of guinea pigs, and examined the alterations occurring after noise exposure. After 70 dB exposure, VEGF expression was found to be reduced in all cell types of the organ of Corti, in the stria vascularis and in spiral ganglion cells. Additional down-regulation was observed in the spiral ligament and in interdental cells after 90 dB. In contrast, VEGF showed an in tendency increased level after both intensities in nerve fibers of the osseous spiral lamina. Expression of FLT was affected similarly, showing down-regulation after 70 and 90 dB on spiral ganglion cells, the nerve fibers of the osseous spiral lamina and on Deiters cells. Additionally, down-regulation was observed in the remaining cell types of the organ of Corti, the stria vascularis, the spiral ligament and the interdental cells. The Neuropilin levels remained unchanged by our experiments; apart from the blood vessel endothelium, there was no detectable expression in any of the cell types investigated. The FLK expression pattern was likewise unaffected by exposure to 70 or 90 dB, with the notable exception of an increased level occurring in Schwann cells after 90 dB. We postulate that modulation of VEGF and its receptors may be part of a neuroprotective mechanism in response to noise.
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http://dx.doi.org/10.1007/s00405-006-0154-3DOI Listing
February 2007

COX-2 expression in the guinea pig cochlea is partly altered by moderate sound exposure.

Neurosci Lett 2006 Feb 10;394(2):121-6. Epub 2005 Nov 10.

Department of Otolaryngology - Head and Neck Surgery, Johannes Gutenberg University Medical School, Mainz, Germany.

The cyclooxygenase-2 isoform (COX-2) was found recently to be constitutively expressed in the guinea pig inner ear. To gain knowledge about its role in sound perception, alterations in the COX-2 level of moderate noise-stimulated cochleae were determined. Staining intensities were quantified in different regions using an immunohistochemical staining procedure and computer-assisted system. After 70 dB and 90 dB noise exposure for 1 h at 8000 Hz, COX-2 downregulation was observed in the organ of Corti, which was most prominent in Deiters' cells near Hensen cells and outer hair cells. In pillar cells, COX-2 levels were only slightly reduced after 70 dB but strongly diminished after 90 dB exposure. In Hensen cells, COX-2 was downregulated after 70 dB stimulation, revealing a decreasing COX-2 content from the third to the first turn of the cochlea and a homogeneously reduced enzyme expression in all three turns after 90 dB. The COX-2 content in inner hair cells was nearly identical to unexposed cochleae after 70 dB exposure but significantly reduced after 90 dB stimulation. In spiral ganglion cells, stria vascularis, spiral ligament and limbus, COX-2 expression was unchanged after 70 dB and 90 dB. We suggest that alterations in COX-2 expression might contribute to diminished sensitivity at the cochlea after noise exposure to reduce subsequent noise distress, termed sound conditioning.
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http://dx.doi.org/10.1016/j.neulet.2005.10.039DOI Listing
February 2006

Endothelial nitric oxide synthase upregulation in the guinea pig organ of Corti after acute noise trauma.

Brain Res 2005 Jun;1047(1):85-96

Department of Otolaryngology--Head and Neck Surgery, Johannes Gutenberg University Medical School, 55131 Mainz, Germany.

Endothelial nitric oxide synthase (eNOS) upregulation was identified 60 h after acute noise trauma in morphologically intact cells of the reticular lamina in the organ of Corti of the guinea pig in the second turn of the cochlea. Using gold-coupled anti-eNOS antibodies and electron microscopy, it was shown that eNOS expression was upregulated in all cell areas and cell types except inner hair cells. Furthermore, eNOS was found in the organelle-free cytoplasm and in mitochondria of various cell types. The density of eNOS in mitochondria was considerably higher compared with the surrounding cytoplasm. Since eNOS activity is regulated by calcium, the eNOS detection was combined with calcium precipitation, a method for visualizing intracellular Ca2+ distribution. After acute noise trauma, intracellular Ca2+ was increased in all cell types and cell areas except in outer hair cells. Comparing the distribution patterns of eNOS and calcium, significantly elevated levels (P < 0.0001) of eNOS were detected within a 100 nm radius near calcium precipitates in all cuticular structures as well as microtubule-rich regions and Deiters' cells near Hensen cells. The observed colocalization lends support to the postulated mechanism of eNOS activation by Ca2+. eNOS upregulation after acute noise trauma might therefore be part of an induced stress response. The eNOS upregulation in cell areas with numerous microtubule- and actin-rich structures is discussed with respect to possible cytoskeleton-dependent processes in eNOS regulation.
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http://dx.doi.org/10.1016/j.brainres.2005.04.023DOI Listing
June 2005

Genetic association of cytokines polymorphisms with autoimmune hepatitis and primary biliary cirrhosis in the Chinese.

World J Gastroenterol 2005 May;11(18):2768-72

The Department of Medicine Laboratory, East Hospital, Ji Mo Road 150, Shanghai 200120, China.

Aim: To characterize gene polymorphism of several cytokine gene in-patients with AIH and PBC and to analyze the difference of the polymorphism distribution between Chinese patients and healthy controls.

Methods: The study population consisted of 62 patients with AIH, and 77 patients with PBC. The genetic profile of four cytokines was analyzed by restriction fragment length polymorphism after specific PCR amplification (PCR-RFLP) or sequence-specific primers PCR (SSP-PCR). The analyzed gene polymorphism included interleukin-1 (IL-1) (at position +3 953 and IL-1RN intron 2), IL-6 (at position -174), IL-10 promoter (at position -1 082, -819, and -592). The control group consisted of 160 healthy blood donors.

Results: The majority of Chinese people including patients and healthy controls exhibited IL-1B 1,1 genotype, and there was no significant difference in AIH, PBC patients and controls. There were highly statistically significant differences in the distribution of the IL-1RN gene polymorphism between the patients with PBC compared with controls. The frequency of IL-1RN 1,1 was significantly higher (90.9% vs 79.4%, P = 0.03) and the frequency of IL-1RN 1,2 was significantly lower in PBC patients (6.5% vs 17.5%, P = 0.01). No statistical difference was observed between AIH patients and controls. All of the 160 healthy controls and 62 cases of AIH patients exhibited IL-6-174GG genotype, and there were four cases, which expressed IL-6-174GC genotype in 77 cases of PBC patients. The frequency of IL-6-174GC was markedly significantly higher in PBC patients compared with controls (5.2% vs 0%, P = 0.004). No statistically significant difference was found in the distribution of IL-10 promoter genotype in AIH and PBC patients compared with controls.

Conclusion: The polymorphisms of IL-1RN and IL-6 -174G/C appear to be associated with PBC in Chinese patients.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4305913PMC
http://dx.doi.org/10.3748/wjg.v11.i18.2768DOI Listing
May 2005

Genetic association of vitamin D receptor polymorphisms with autoimmune hepatitis and primary biliary cirrhosis in the Chinese.

J Gastroenterol Hepatol 2005 Feb;20(2):249-55

Center of Clinical Immunology, Changzheng Hospital, Second Military Medical University, 4215 Feng Yang Road, Shanghai 200-003, China.

Background: Autoimmune hepatitis (AIH) and primary biliary cirrhosis (PBC) are two autoimmune diseases of unknown etiology. Genetic factors appear to be involved in the pathogenesis of both diseases. Vitamin D has been shown to exert multiple immunomodulatory effects, which acts through its own receptor (VDR). Polymorphisms of VDR had been implicated in several autoimmune diseases. In the present study, the association between Chinese patients with AIH, PBC and the polymorphisms in exon 2, intron 8 and exon 9 of vitamin D receptor genes was studied.

Methods: Four candidate gene loci were investigated in 49 patients with AIH, 58 patients with PBC, and 160 healthy controls. The VDR polymorphisms were assessed by FokI, BsmI, ApaI, and TaqI endonuclease digestion after specific polymerase chain reaction (PCR) amplification.

Results: The result show a significant difference in FokI polymorphism between AIH patients and controls (chi(2) = 5.47, P = 0.019), and a significant association in BsmI polymorphisms between PBC patients and controls (chi(2) = 6.52, P = 0.01). Furthermore the distribution of FokI, BsmI, ApaI, and TaqI gene types differed between Chinese healthy controls and Caucasian healthy controls.

Conclusion: It is suggested that there is a genetic link of VDR polymorphisms to autoimmune liver diseases such as AIH and PBC in Chinese patients. Further studies are needed to elucidate the mechanisms by which VDR polymorphisms contribute to the lose of immune tolerance in autoimmune diseases.
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http://dx.doi.org/10.1111/j.1440-1746.2005.03532.xDOI Listing
February 2005

Cytotoxic T lymphocyte associated antigen-4 gene polymorphisms confer susceptibility to primary biliary cirrhosis and autoimmune hepatitis in Chinese population.

World J Gastroenterol 2004 Oct;10(20):3056-9

The Center of Clinical Immunology, Changzheng Hospital, Second Military Medical University, Shanghai 200003, China.

Aim: To investigate the association between Chinese patients with autoimmune hepatitis (AIH), primary biliary cirrhosis (PBC) and the polymorphisms of cytotoxic T lymphocyte-associated antigen-4 (CTLA-4) gene promoter (-318) and exon 1 (+49).

Methods: CTLA-4 promoter (-318 T/C) and exon1 (+49A/G) polymorphisms were genotyped via restriction fragment length polymorphism methods in 62 Chinese AIH patients, 77 Chinese PBC patients and 160 healthy controls.

Results: We found a significant association in CTLA-4 gene exon1 49 A/G polymorphism between PBC patients and controls (P = 0.006) and the frequency of G alleles was significantly increased in comparison with controls (P = 0.0046, OR = 1.8). We also found the frequency of C alleles in promoter -318 was significantly increased in AIH patients compared with controls (P = 0.02, OR = 0.41). Although the genotype distribution of the CTLA-4 exon 1-promoter gene was not significantly different between AIH and PBC patients and controls, the occurrence of GG-CC was increased in two groups of patients (AIH: 32.3%, PBC: 37.7%, control: 22.5%).

Conclusion: Polymorphisms of CTLA-4 gene probably confer susceptibility to AIH and PBC in Chinese population.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4576272PMC
http://dx.doi.org/10.3748/wjg.v10.i20.3056DOI Listing
October 2004

[Genetic association of tumor necrosis factor (TNF)-alpha polymorphisms with primary biliary cirrhosis and autoimmune liver diseases in a Chinese population].

Zhonghua Gan Zang Bing Za Zhi 2004 Mar;12(3):160-2

Department of Experimental Diagnosis, Changzheng Hospital, Second Military Medical University, Shanghai 200003, China.

Objective: Autoimmune hepatitis (AIH) and primary biliary cirrhosis (PBC) are two autoimmune diseases of unknown etiology. Genetic factors appear to be involved in the pathogenesis of both diseases. Tumor necrosis factor (TNF)-alpha is one of the proinflammatory cytokines and immunomodulators, and is implicated in the pathogenesis of AIH and PBC. In this study, we studied the association between Chinese patients with AIH, PBC and the polymorphisms in promoter-region polymorphisms of the TNF-alpha gene at position -308 and -238.

Methods: We have investigated four candidate gene loci in 49 patients with AIH, 58 patients with PBC, and 160 healthy controls. The polymorphisms were assessed by the PCR specifically for the single-nucleotide polymorphisms.

Results: We found the difference in the TNF-alpha gene at position -308 genotype distributions between Chinese health controls and Caucasian health controls. Although the percent of TNF-alpha*2 was decrease on PBC patient group (10.34% vs. 16.88%), there was no significant difference between PBC patients and health control in the Chinese. There were also no significant differences between AIH and health control on the TNF-alpha gene at position -308 and -238.

Conclusion: Our findings suggest that the TNF-alpha promoter-region polymorphisms distribution is different between differe of ethnic groups; there are no genetic links of the TNF-alpha promoter-region polymorphisms to AIH and PBC in Chinese.
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March 2004

[Genetic association of vitamin D receptor polymorphisms with primary biliary cirrhosis and autoimmune liver diseases on Chinese].

Zhonghua Yi Xue Za Zhi 2003 Nov;83(21):1852-5

Department of Experimental Diagnosis, Changzheng Hospital, Second Military Medical University, Shanghai 200003, China.

Objective: To study the association between the polymorphisms of vitamin D receptor (VDR) gene and autoimmune liver diseases and (AIH) and primary biliary cirrhosis (PBC) in Chinese.

Methods: PCR-RELP was used to investigate the polymorphisms in exon 2, and exon 7 to exon 9 of VDR among 49 AIH patients, 58 PBC patients, and 160 healthy controls, all Chinese. VDR polymorphisms were assessed by FokI, BsmI, ApaI, and TaqI endonucleases restriction fragment length polymorphism analysis after specific polymerase chain reaction (PCR) amplification.

Results: The distribution of VDR gene polymorphism in Chinese was similar to those in Koreans and Japanese, and different from those in the Germans and Spaniards. The percentage of ff phenotype carriers was significantly higher in the AIH patients than in the healthy controls (34.7% vs. 48.1%, chi(2) = 5.47, P = 0.019) and the percentage of Ff phenotype carriers was lower in the AIH patients than in the healthy controls (34.7% vs. 48.1%, P = 0.057). The percentage of Bb phenotype carriers was significantly lower in the PBC patients than in the healthy controls (5.2% vs. 17.5%, P = 0.021) and. the percentage of bb phenotype carriers was significantly higher in the PBC patients than in the healthy controls (94.8% vs. 80.6%, chi(2) = 6.52, P = 0.01). We also detected a significant association of the BsmI polymorphisms in PBC patients in comparison with controls (P = 0.01). Furthermore we found the difference in the FokI, BsmI, ApaI, and TaqI genotype distribution between Chinese health controls and Caucasian health controls.

Conclusion: There is a significant association between FokI polymorphism and AIH and a significant association between the BsmI polymorphisms and PBC in Chinese.
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November 2003

An unusual mechanism of bacterial gene expression revealed for the RNase P protein of Thermus strains.

Proc Natl Acad Sci U S A 2003 May 28;100(10):5724-9. Epub 2003 Apr 28.

Institute für Biochemie and Molekulare Medizin, Universität zu Lübeck, Ratzeburger Allee 160, D-23538 Lübeck, Germany.

The RNase P protein gene (rnpA) completely overlaps the rpmH gene (encoding ribosomal protein L34) out of frame in the thermophilic bacterium Thermus thermophilus. This results in the synthesis of an extended RNase P protein (C5) of 163 aa and, by inference, of 240 aa in the related strain Thermus filiformis. Start codons of rnpA and rpmH, apparently governed by the same ribosome binding site, are separated by only 4 nt, which suggests a regulatory linkage between L34 and C5 translation and, accordingly, between ribosome and RNase P biosynthesis. Within the sequence encoding the N-terminal extensions and downstream of rpmH, several Thermus species exhibit in-frame deletionsinsertions, suggesting relaxed constraints for sequence conservation in this region. Roughly the N-terminal third of T. thermophilus C5 was further shown to be dispensable for RNase P function in vitro by using a precursor tRNA(Gly) substrate from the same organism. Taken together, these data reveal a mode of gene expression that is to our knowledge unprecedented in bacteria.
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http://dx.doi.org/10.1073/pnas.0931462100DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC156268PMC
May 2003

tRNA maturation in Aquifex aeolicus.

Biochimie 2002 Aug;84(8):713-22

Institut für Biochemie, Universität zu Lübeck, Ratzeburger Allee 160, 23538, Lübeck, Germany.

Several tRNAs in the hyperthermophilic bacterium Aquifex aeolicus are encoded in clusters and as part of ribosomal RNA operons, implying the requirement for tRNA processing by ribonuclease P (RNase P). Intriguingly, neither a gene for the RNA nor the protein component of this ubiquitous ribonucleoprotein enzyme has been hitherto identified in the sequenced genome of A. aeolicus, despite extensive data mining. As a result of the present study, primer extension analysis revealed that tRNAs in A. aeolicus possess canonical mature 5' ends; yet we were unable to demonstrate RNase P holoenzyme or RNase P RNA alone activity in A. aeolicus extracts under a variety of reaction conditions utilizing mono- and dimeric ptRNA substrates. Processing of dimeric ptRNA transcripts in extracts of A. aeolicus disclosed at least one endoribonuclease which cleaves in the A/U-rich spacer of the tandem ptRNA, reminiscent of bacterial RNase E-like enzymes.
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http://dx.doi.org/10.1016/s0300-9084(02)01447-5DOI Listing
August 2002

Potential contact sites between the protein and RNA subunit in the Bacillus subtilis RNase P holoenzyme.

J Mol Biol 2002 Jan;315(4):551-60

Institut für Biochemie, Medizinische Universität zu Lübeck, Ratzeburger Allee 160, Lübeck, D-23538, Germany.

We have detected by nucleotide analog interference mapping (NAIM) AMPalphaS and IMPalphaS modifications in Bacillus subtilis RNase P RNA that interfere with binding of the homologous protein subunit. Interference as well as some enhancement effects were clustered in two main areas, in P10.1a/L10.1 and P12 of the specificity domain (cluster 1, domain I) and in P2, P3, P15.1, J18/2 and J19/4 of the catalytic domain (cluster 2a, domain II). Minor interferences in P1 and P19 and a strong and weak enhancement effect in P19 represent a third area located in domain II (cluster 2b). Our results suggest that P3, P2-J18/2 and J19/4 are key elements for anchoring of the protein to the catalytic domain close to the scissile phosphodiester in enzyme-substrate complexes. Sites of interference or enhancement in clusters 1 and 2a are located at distances between 65 and 130 A from each other in the current 3D model of a full-length RNase P RNA-substrate complex. Taking into account that the RNase P protein monomer can bridge a maximum distance of about 40 A, simultaneous direct contacts to the two aforementioned potential RNA-binding areas would be incompatible with our current understanding of bacterial RNase P RNA architecture. Our findings suggest that the current 3D model has to be rearranged in order to reduce the distance between clusters 1 and 2a. Alternatively, based on the recent finding that B. subtilis RNase P forms a tetramer consisting of two protein and two RNA subunits, cluster 1 may reflect one protein contact site in domain I, and cluster 2a a separate one in domain II.
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http://dx.doi.org/10.1006/jmbi.2001.5261DOI Listing
January 2002