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1.
Article in German | MEDLINE | ID: mdl-34670308

ABSTRACT

OBJECTIVES: Survey concerning the situation of small animal anesthesia in Europe and assessment of the compliance with the guidelines of the AVA (American Society of Anesthesiologists) and the DVG specialty group VAINS (Veterinary Anesthesia, Intensive Care, Emergency Medicine, and Pain Management). MATERIAL AND METHODS: A link to an online survey with questions concerning anesthesia management (topics include demographics, equipment, monitoring, thermal management, pre-anesthetic examination, anesthetic protocol, and others) was sent to small animal practitioners in several countries. RESULTS: A total of 767 evaluable questionnaires came from Germany (n = 343), Austria (n = 216), Switzerland (n = 83), the United Kingdom (n = 38), France (n = 25), Hungary (n = 25), Scandinavia (n = 23), and "other countries" (n = 11). On average, 91 % of respondents complied with the AVA guideline and 58 % complied with the VAINS specialty group guideline even before its publication. Practices/clinics with higher staff possessed superior equipment, and practices/clinics performing higher numbers of anesthesias per week were more likely to implement "good preliminary examination." CONCLUSION AND CLINICAL RELEVANCE: Although the guidelines were found to be implemented to a certain degree, the presented study reveals a potential for optimization of the anesthesia regime in many practices/clinics, e. g. by improving the equipment or allowing for better use of present apparatus.


Subject(s)
Anesthesia , Anesthetics , Anesthesia/veterinary , Animals , Europe , Germany , Surveys and Questionnaires
2.
Sci Rep ; 9(1): 5656, 2019 04 04.
Article in English | MEDLINE | ID: mdl-30948772

ABSTRACT

Human immunodeficiency virus (HIV-1) entry is initiated by the binding between the viral envelope glycoprotein gp120 and the host receptor CD4, and followed by reduction of structural disulfides of gp120 and CD4. The host thioredoxin-1 (Trx1) efficiently reduces disulfides of gp120 and CD4 in vitro, and recently CD4-dependent HIV-1 entry was shown to be inhibited by anti-Trx1-antibodies, indicating a central role for Trx1. 1-methylpropyl-2-imidazolyl disulfide (PX-12) is a reversible inhibitor of the Trx1 system that may also cause a slow irreversible thioalkylation of Trx1. It was developed as an antitumor agent, however, the current study aimed to determine if it also has an anti-HIV-1 effect. We show that PX-12 has anti-HIV-1(IIIB) activity in TZM-bl cells, in fact, no virus was detected inside the cells in the presence of 10 µM PX-12. Moreover, PX-12 inhibited the enzymatic activity of Trx1 and the Trx1-dependent disulfide reduction of gp120. Microtubule polymerization and formation of acetylated microtubules were also inhibited, activities shown to be required for HIV-1 life cycle propagation. In conclusion, our data strengthens the notion that the early steps of the HIV-1 life cycle depends on the Trx1 system and indicate that the Trx1 system may be a rational drug target for HIV-1 treatment.


Subject(s)
Disulfides/pharmacology , HIV Infections/drug therapy , Imidazoles/pharmacology , Thioredoxins/metabolism , CD4 Antigens/metabolism , Cell Line , Disulfides/metabolism , HIV Envelope Protein gp120/metabolism , HIV Infections/metabolism , HIV Infections/virology , HIV-1/physiology , Humans , Imidazoles/metabolism , Oxidation-Reduction , Protein Binding , Thioredoxins/drug effects , Virus Internalization/drug effects
3.
PLoS One ; 11(1): e0147773, 2016.
Article in English | MEDLINE | ID: mdl-26816344

ABSTRACT

BACKGROUND: The entry of HIV into its host cell is an interesting target for chemotherapeutic intervention in the life-cycle of the virus. During entry, reduction of disulfide bridges in the viral envelope glycoprotein gp120 by cellular oxidoreductases is crucial. The cellular thioredoxin reductase-1 plays an important role in this oxidoreduction process by recycling electrons to thioredoxin-1. Therefore, thioredoxin reductase-1 inhibitors may inhibit gp120 reduction during HIV-1 entry. In this present study, tellurium-based thioredoxin reductase-1 inhibitors were investigated as potential inhibitors of HIV entry. RESULTS: The organotellurium compounds inhibited HIV-1 and HIV-2 replication in cell culture at low micromolar concentrations by targeting an early event in the viral infection cycle. Time-of-drug-addition studies pointed to virus entry as the drug target, more specifically: the organotellurium compound TE-2 showed a profile similar or close to that of the fusion inhibitor enfuvirtide (T-20). Surface plasmon resonance-based interaction studies revealed that the compounds do not directly interact with the HIV envelope glycoproteins gp120 and gp41, nor with soluble CD4, but instead, dose-dependently bind to thioredoxin reductase-1. By inhibiting the thioredoxin-1/thioredoxin reductase-1-directed oxidoreduction of gp120, the organotellurium compounds prevent conformational changes in the viral glycoprotein which are necessary during viral entry. CONCLUSION: Our findings revealed that thioredoxin-1/thioredoxin reductase-1 acts as a cellular target for the inhibition of HIV entry.


Subject(s)
Antiviral Agents/pharmacology , HIV Infections/drug therapy , HIV-1/drug effects , Tellurium/pharmacology , Thioredoxin Reductase 1/antagonists & inhibitors , Thioredoxins/metabolism , Virus Internalization/drug effects , Antiviral Agents/chemistry , Cell Line , HIV Envelope Protein gp120/metabolism , HIV Infections/metabolism , HIV-1/physiology , Humans , Oxidation-Reduction/drug effects , Tellurium/chemistry , Thioredoxin Reductase 1/metabolism
4.
PLoS One ; 9(7): e103554, 2014.
Article in English | MEDLINE | ID: mdl-25075746

ABSTRACT

Thioredoxin-1 (Trx1) is a protein antioxidant involved in major cellular processes. Increased plasma levels of Trx1 have been associated with human diseases suggesting that Trx1 is a marker for oxidative stress with putative clinical use. However, the reported mean levels of Trx1 in the control cohorts vary a hundred-fold between studies (0.8-87 ng/ml), possibly due to methodological differences between the capture ELISA used in the different studies. The aim of this study was to investigate methodological aspects related to the ELISA measurement of Trx1. ELISAs utilizing different capture and detection combinations of antibodies to Trx1 and as well as recombinant human (rh) Trx1 standards from two sources were characterized. The different ELISAs were subsequently used to measure Trx1 in human plasma and cerebrospinal fluid samples (CSF) from healthy donors and from patients with various neurological diagnoses. The Trx1 standards differed in their content of monomeric and oligomeric Trx1, which affected the ELISAs composed of different antibody combinations. Thus, the levels of Trx1 determined in human plasma and CSF samples varied depending on the antibody used in the ELISAs and on the rhTrx1 standard. Furthermore, the relevance of preventing interference by heterophilic antibodies (HA) in human plasma and CSF was investigated. The addition of a HA blocking buffer to human samples drastically reduced the ELISA signals in many samples showing that HA are likely to cause false positive results unless they are blocked. In conclusion, the study shows that the design of a Trx1 ELISA in regards to antibodies and standards used has an impact on the measured Trx1 levels. Importantly, analyses of human plasma and CSF without preventing HA interference may obscure the obtained data. Overall, the results of this study are crucial for the improvement of future studies on the association of Trx1 levels with various diseases.


Subject(s)
Enzyme-Linked Immunosorbent Assay , Thioredoxins/analysis , Adult , Antibodies/immunology , Blotting, Western , Female , Humans , Male , Middle Aged , Nervous System Diseases/cerebrospinal fluid , Nervous System Diseases/diagnosis , Oxidative Stress , Recombinant Proteins/biosynthesis , Recombinant Proteins/genetics , Recombinant Proteins/immunology , Thioredoxins/blood , Thioredoxins/cerebrospinal fluid
5.
Antioxid Redox Signal ; 18(7): 795-9, 2013 Mar 01.
Article in English | MEDLINE | ID: mdl-23043315

ABSTRACT

The global protein thiol pool has been reported to play a major role in the defense against oxidative stress as a redox buffer similar to glutathione. The present study uses a novel method to visualize cellular changes of the global protein thiol pool in response to induced oxidative stress. Unexpectedly, the results showed an uneven distribution of protein thiols in resting cells with no apparent change in their level or distribution in response to diamide as has been reported previously. Further analysis revealed that thiol pool oxidation is artificially high due to insufficient activity of the widely used sample quencher trichloroacetic acid (TCA). This suggests that previously published articles based on TCA as a quencher should be interpreted with caution as TCA could have caused similar artifacts. Overall, the results presented here question the major role for the global thiol pool in the defense against oxidative stress. Instead our hypothesis is that the fraction of proteins involved in response to oxidative stress is much smaller than previously anticipated in support of a fine-tuned cell signaling by redox regulation.


Subject(s)
Actins/metabolism , Trichloroacetic Acid/metabolism , Actins/chemistry , Artifacts , Cell Line, Tumor , Diamide/pharmacology , HeLa Cells , Humans , Oxidation-Reduction/drug effects , Oxidative Stress/drug effects , Sulfhydryl Compounds/metabolism , Sulfhydryl Compounds/pharmacology
6.
Int J Biochem Cell Biol ; 44(3): 556-62, 2012 Mar.
Article in English | MEDLINE | ID: mdl-22230366

ABSTRACT

HIV-1 enters cells via interaction of the viral glycoprotein gp120, the host cell surface receptor CD4 and the co-receptors CCR5 or CXCR4. For entry, gp120 undergoes conformational changes that depend on the reduction of one or more disulfides. Previous studies indicate that protein disulfide isomerase (PDI), thioredoxin-1 (Trx1), and glutaredoxin-1 (Grx1) catalyze gp120 reduction, but their specific disulfide targets are not known. Here, it was demonstrated that PDI and Trx1 have similar gp120 disulfide targets as determined by labeling after reduction, but with some pattern differences, including overall stronger labeling with Trx1 than with PDI. Furthermore, uneven labeling of the residues of a disulfide may reflect altered accessibility by conformational changes upon the reduction process. Since both PDI and Trx1 may be involved in viral entry, compounds that target the host redox system or the viral gp120 were tested in vitro to investigate whether redox regulation is a target for anti-HIV therapy. Carbohydrate binding agents (CBAs), previously shown to bind gp120 and inhibit HIV entry, were now demonstrated to inhibit gp120 disulfide reduction. Auranofin, an inhibitor of thioredoxin reductase 1 (TrxR1), also showed inhibitory activity towards HIV infection, although close to its cytotoxic concentration. Our results demonstrate that both the host redox system and the viral surface glycoproteins are of interest for the development of new generations of anti-HIV therapeutics.


Subject(s)
HIV Envelope Protein gp120/metabolism , HIV Infections/metabolism , HIV-1/physiology , Protein Disulfide-Isomerases/metabolism , Thioredoxins/metabolism , Allosteric Regulation , Animals , Antiviral Agents/pharmacology , Auranofin/pharmacology , Cattle , Disulfides/chemistry , Disulfides/metabolism , HIV Envelope Protein gp120/chemistry , HIV Infections/physiopathology , HIV Infections/virology , HIV-1/pathogenicity , Humans , Mass Spectrometry , Oxidation-Reduction/drug effects , Protein Conformation , Rats , Virus Internalization , Virus Replication
7.
Arch Microbiol ; 192(5): 373-82, 2010 May.
Article in English | MEDLINE | ID: mdl-20354844

ABSTRACT

Aspergillus nidulans can use a variety of fatty acids as sole carbon and energy sources via its peroxisomal and mitochondrial beta-oxidation pathways. Prior to channelling the fatty acids into beta-oxidation, they need to be activated to their acyl-CoA derivates. Analysis of the genome sequence identified a number of possible fatty acyl-CoA synthetases (FatA, FatB, FatC, FatD, FaaA and FaaB). FaaB was found to be the major long-chain synthetase for fatty acid degradation. FaaB was shown to localise to the peroxisomes, and the corresponding gene was induced in the presence of short and long chain fatty acids. Deletion of the faaB gene leads to a reduced/abolished growth on a variety of fatty acids. However, at least one additional fatty acyl-CoA synthetase with a preference for short chain fatty acids and a potential mitochondrial candidate (AN4659.3) has been identified via genome analysis.


Subject(s)
Acyl Coenzyme A/metabolism , Aspergillus nidulans/enzymology , Coenzyme A Ligases/genetics , Coenzyme A Ligases/metabolism , Fungal Proteins/genetics , Fungal Proteins/metabolism , Aspergillus nidulans/genetics , Aspergillus nidulans/growth & development , Gene Deletion , Genome, Fungal , Genomics
8.
Curr Genet ; 56(2): 139-50, 2010 Apr.
Article in English | MEDLINE | ID: mdl-20043225

ABSTRACT

Filamentous fungi can use a variety of fatty acids (FA) as sole carbon and energy sources. Aspergillus nidulans has been shown to possess both peroxisomal and mitochondrial beta-oxidation pathways. In these studies, the major peroxisomal long chain fatty acyl coenzyme A oxidase AoxA was identified. AoxA was shown to be localised to peroxisomes and deletion of the aoxA gene leads to reduced growth on long chain FA, but not on short chain FA. AoxA is predicted to be part of the same peroxisomal beta-oxidation pathway as the bifunctional protein FoxA. In addition, an aoxA(p)lacZ reporter gene construct is induced by short and long chain FA and the induction is dependent on the transcriptional regulators FarA, FarB and ScfA with FarA being required for the induction by short chain as well as long chain FA and FarB and ScfA being required for induction of aoxA by short chain FA. It is proposed that there are additional peroxisomal beta-oxidation pathways in A. nidulans, which include fatty acyl-CoA dehydrogenases with a partially overlapping substrate range and include a pathway for short chain FA.


Subject(s)
Acyl-CoA Oxidase/metabolism , Aspergillus nidulans/genetics , Aspergillus nidulans/metabolism , Fatty Acids/metabolism , Oxidoreductases/metabolism , Acyl-CoA Dehydrogenase/genetics , Acyl-CoA Dehydrogenase/metabolism , Acyl-CoA Dehydrogenases/genetics , Acyl-CoA Dehydrogenases/metabolism , Acyl-CoA Oxidase/genetics , Aspergillus nidulans/growth & development , Fatty Acids/genetics , Genes, Reporter , Mitochondria/genetics , Mitochondria/metabolism , Oxidation-Reduction , Peroxisomes/genetics , Peroxisomes/metabolism
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