Acute, reproductive, and developmental toxicity of essential oils assessed with alternative in vitro and in vivo systems

Abstract Essential oils (EOs) have attracted increased interest for different applications such as food preservatives, feed additives and ingredients in cosmetics. Due to their reported variable composition of components, they might be acutely toxic to humans and animals in small amounts. Despite the necessity, rigorous toxicity testing in terms of safety evaluation has not been reported so far, especially using alternatives to animal models. Here, we provide a strategy by use of alternative in vitro (cell cultures) and in vivo (Caenorhabditis elegans, hen’s egg test) approaches for detailed investigation of the impact of commonly used rosemary, citrus and eucalyptus essential oil on acute, developmental and reproductive toxicity as well as on mucous membrane irritation. In general, all EOs under study exhibited a comparable impact on measured parameters, with a slightly increased toxic potential of rosemary oil. In vitro cell culture results indicated a concentration-dependent decrease of cell viability for all EOs, with mean IC50 values ranging from 0.08 to 0.17% [v/v]. Similar results were obtained for the C. elegans model when using a sensitized bus-5 mutant strain, with a mean LC50 value of 0.42% [v/v]. In wild-type nematodes, approximately tenfold higher LC50 values were detected. C. elegans development and reproduction was already significantly inhibited at concentrations of 0.5% (wild-type) and 0.1% (bus-5) [v/v] of EO, respectively. Gene expression analysis revealed a significant upregulation of xenobiotic and oxidative stress genes such as cyp-14a3, gst-4, gpx-6 and sod-3. Furthermore, all three EOs under study showed an increased short-time mucous membrane irritation potential, already at 0.5% [v/v] of EO. Finally, GC–MS analysis was performed to quantitate the relative concentration of the most prominent EO compounds. In conclusion, our results demonstrate that EOs can exhibit severe toxic properties, already at low concentrations. Therefore, a detailed toxicological assessment is highly recommended for each EO and single intended application.

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The double-stranded DNA-binding proteins TEBP-1 and TEBP-2 form a telomeric complex with POT-1

Nature Communications ◽

10.1038/s41467-021-22861-2 ◽

2021 ◽

Vol 12 (1) ◽

Author(s):

Sabrina Dietz ◽

Miguel Vasconcelos Almeida ◽

Emily Nischwitz ◽

Jan Schreier ◽

Nikenza Viceconte ◽

...

Keyword(s):

Quantitative Proteomics ◽

Wild Type ◽

C Elegans ◽

Double Stranded Dna ◽

Telomere Sequence ◽

Proteomics Approach ◽

Telomeric Protein ◽

Regulate Telomere Length

AbstractTelomeres are bound by dedicated proteins, which protect them from DNA damage and regulate telomere length homeostasis. In the nematode Caenorhabditis elegans, a comprehensive understanding of the proteins interacting with the telomere sequence is lacking. Here, we harnessed a quantitative proteomics approach to identify TEBP-1 and TEBP-2, two paralogs expressed in the germline and embryogenesis that associate to telomeres in vitro and in vivo. tebp-1 and tebp-2 mutants display strikingly distinct phenotypes: tebp-1 mutants have longer telomeres than wild-type animals, while tebp-2 mutants display shorter telomeres and a Mortal Germline. Notably, tebp-1;tebp-2 double mutant animals have synthetic sterility, with germlines showing signs of severe mitotic and meiotic arrest. Furthermore, we show that POT-1 forms a telomeric complex with TEBP-1 and TEBP-2, which bridges TEBP-1/-2 with POT-2/MRT-1. These results provide insights into the composition and organization of a telomeric protein complex in C. elegans.

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67 Current Status and Future Prospective of the Use of Plant Bioactive Compounds in Dairy and Beef Cattle

Journal of Animal Science ◽

10.1093/jas/skab054.222 ◽

2021 ◽

Vol 99 (Supplement_1) ◽

pp. 132-132

Author(s):

Sergio Calsamiglia ◽

Maria Rodriguez-Prado ◽

Gonzalo Fernandez-Turren ◽

Lorena Castillejos

Keyword(s):

Beef Cattle ◽

Essential Oils ◽

Average Daily Gain ◽

Rumen Fermentation ◽

Production Performance ◽

In Vivo Studies ◽

Current Status ◽

The Impact

Abstract In the last 20 years there has been extensive in vitro research on the effects of plant extracts and essential oils on rumen microbial fermentation. The main objectives have been to improve energy metabolism through a reduction in methane emissions and an increase in propionate production; and to improve protein metabolism by reducing proteolysis and deamination. While the positive results from in vitro studies has stimulated the release of commercial products based on blends of essential oils, there is limited in vivo evidence on the rumen fermentation and production performance effects. A literature search was conducted to select in vivo studies where information on rumen fermentation and animal performance was reported. For dairy cattle, we identified 37 studies of which 21 were adequate to test production performance. Ten studies reported increases and 3 decreases in milk yield. For beef cattle, we identified 20 studies with rumen fermentation profile and 22 with performance data. Average daily gain improved in 7 and decreased in 1 study. Only 1 out of 16 studies reported an improvement in feed efficiency. Data indicate that out of more than 500 products tested in vitro, only around 20 have been tested in vivo in different mixtures and doses. The use of statistical approaches will allow to describe the conditions, doses and responses in dairy and beef cattle performance. The search for postruminal effects offers another alternative use. Evidence for effects on the intestinal and systemic effects on the immune system and antioxidant status (i.e., capsicum, garlic, eugenol, cinnamaldehyde curcuma, catechins, anethol or pinene), and in the modulation of metabolic regulation (capsicum, cinnamaldehyde, curcuma or garlic) may open the opportunity for future applications. However, stability of the product in the GI tract, description of the mechanisms of action and the impact of these changes on performance needs to be further demonstrated.

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Production of Noncapped Genomic RNAs Is Critical to Sindbis Virus Disease and Pathogenicity

mBio ◽

10.1128/mbio.02675-20 ◽

2020 ◽

Vol 11 (6) ◽

Author(s):

Autumn T. LaPointe ◽

V Douglas Landers ◽

Claire E. Westcott ◽

Kevin J. Sokoloski

Keyword(s):

Sindbis Virus ◽

Virus Disease ◽

Severe Disease ◽

Wild Type Virus ◽

Wild Type ◽

Genomic Rnas ◽

Mortality And Morbidity ◽

The Impact

ABSTRACT Alphaviruses are positive-sense RNA viruses that utilize a 5′ cap structure to facilitate translation of viral proteins and to protect the viral RNA genome. Nonetheless, significant quantities of viral genomic RNAs that lack a canonical 5′ cap structure are produced during alphaviral replication and packaged into viral particles. However, the role/impact of the noncapped genomic RNA (ncgRNA) during alphaviral infection in vivo has yet to be characterized. To determine the importance of the ncgRNA in vivo, the previously described D355A and N376A nsP1 mutations, which increase or decrease nsP1 capping activity, respectively, were incorporated into the neurovirulent AR86 strain of Sindbis virus to enable characterization of the impact of altered capping efficiency in a murine model of infection. Mice infected with the N376A nsP1 mutant exhibited slightly decreased rates of mortality and delayed weight loss and neurological symptoms, although levels of inflammation in the brain were similar to those of wild-type infection. Although the D355A mutation resulted in decreased antiviral gene expression and increased resistance to interferon in vitro, mice infected with the D355A mutant showed significantly reduced mortality and morbidity compared to mice infected with wild-type virus. Interestingly, expression of proinflammatory cytokines was found to be significantly decreased in mice infected with the D355A mutant, suggesting that capping efficiency and the production of ncgRNA are vital to eliciting pathogenic levels of inflammation. Collectively, these data indicate that the ncgRNA have important roles during alphaviral infection and suggest a novel mechanism by which noncapped viral RNAs aid in viral pathogenesis. IMPORTANCE Mosquito-transmitted alphaviruses have been the cause of widespread outbreaks of disease that can range from mild illness to lethal encephalitis or severe polyarthritis. There are currently no safe and effective vaccines or therapeutics with which to prevent or treat alphaviral disease, highlighting the need to better understand alphaviral pathogenesis to develop novel antiviral strategies. This report reveals production of noncapped genomic RNAs (ncgRNAs) to be a novel determinant of alphaviral virulence and offers insight into the importance of inflammation to pathogenesis. Taken together, the findings reported here suggest that the ncgRNAs contribute to alphaviral pathogenesis through the sensing of the ncgRNAs during alphaviral infection and are necessary for the development of severe disease.

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The double-stranded DNA-binding proteins TEBP-1 and TEBP-2 form a telomeric complex with POT-1

10.1101/2020.08.19.256388 ◽

2020 ◽

Author(s):

Sabrina Dietz ◽

Miguel Vasconcelos Almeida ◽

Emily Nischwitz ◽

Jan Schreier ◽

Nikenza Viceconte ◽

...

Keyword(s):

Quantitative Proteomics ◽

Binding Proteins ◽

Protein Complexes ◽

Wild Type ◽

C Elegans ◽

Double Stranded Dna ◽

Telomere Sequence ◽

Proteomics Approach

AbstractTelomeres are bound by dedicated protein complexes, like shelterin in mammals, which protect telomeres from DNA damage. In the nematode Caenorhabditis elegans, a comprehensive understanding of the proteins interacting with the telomere sequence is lacking. Here, we harnessed a quantitative proteomics approach to screen for proteins binding to C. elegans telomeres, and identified TEBP-1 and TEBP-2, two paralogs that associate to telomeres in vitro and in vivo. TEBP-1 and TEBP-2 are expressed in the germline and during embryogenesis. tebp-1 and tebp-2 mutants display strikingly distinct phenotypes: tebp-1 mutants have longer telomeres than wild-type animals, while tebp-2 mutants display shorter telomeres and a mortal germline, a phenotype characterized by transgenerational germline deterioration. Notably, tebp-1; tebp-2 double mutant animals have synthetic sterility, with germlines showing signs of severe mitotic and meiotic arrest. TEBP-1 and TEBP-2 form a telomeric complex with the known single-stranded telomere-binding proteins POT-1, POT-2, and MRT-1. Furthermore, we find that POT-1 bridges the double- stranded binders TEBP-1 and TEBP-2, with the single-stranded binders POT-2 and MRT-1. These results describe the first telomere-binding complex in C. elegans, with TEBP-1 and TEBP-2, two double-stranded telomere binders required for fertility and that mediate opposite telomere dynamics.

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Essential Oils as a Feed Additives: Pharmacokinetics and Potential Toxicity in Monogastric Animals

Animals ◽

10.3390/ani9060352 ◽

2019 ◽

Vol 9 (6) ◽

pp. 352 ◽

Cited By ~ 6

Author(s):

Pavel Horky ◽

Sylvie Skalickova ◽

Kristyna Smerkova ◽

Jiri Skladanka

Keyword(s):

Essential Oils ◽

Reproductive Toxicity ◽

Antimicrobial Effect ◽

Feed Additives ◽

The Body ◽

Syzygium Aromaticum ◽

High Antibacterial Activity ◽

Active Substances

Essential oils (EOs) are now a hot topic in finding modern substitutes for antibiotics. Many studies have shown positive results and confirmed their high antibacterial activity both in vitro and in vivo. Deservedly, there is an attempt to use EOs as a substitute for antibiotics, which are currently limited by legislation in animal breeding. Given the potential of EOs, studies on their fate in the body need to be summarized. The content of EO’s active substances varies depending on growing conditions and consequently on processing and storage. Their content also changes dynamically during the passage through the gastrointestinal tract and their effective concentration can be noticeably diluted at their place of action (small intestine and colon). Based on the solubility of the individual EO’s active substances, they are eliminated from the body at different rates. Despite a strong antimicrobial effect, some oils can be toxic to the body and cause damage to the liver, kidneys, or gastrointestinal tissues. Reproductive toxicity has been reported for Origanum vulgare and Mentha arvensis. Several publications also address the effect on the genome. It has been observed that EOs can show both genoprotective effects (Syzygium aromaticum) and genotoxicity, as is the case of Cinnamomum camphor. This review shows that although oils are mainly studied as promising antimicrobials, it is also important to assess animal safety.

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The experimentally obtained functional impact assessments of GT>GC 5’ splice site variants differ markedly from those predicted

10.1101/864843 ◽

2019 ◽

Author(s):

Jian-Min Chen ◽

Jin-Huan Lin ◽

Emmanuelle Masson ◽

Zhuan Liao ◽

Claude Férec ◽

...

Keyword(s):

Splice Site ◽

Disease Genes ◽

Wild Type ◽

Human Genetic Disease ◽

Functional Impact ◽

Prediction Tools ◽

Human Disease Genes ◽

The Impact

ABSTRACTGT>GC 5’ splice site (or +2T>C) variants have been frequently reported to cause human genetic disease. However, although we have demonstrated that GT>GC variants in human disease genes may not invariably be pathogenic, none of the currently available splicing prediction tools appear to be capable of reliably distinguishing those GT>GC variants that generate wild-type transcripts from those that do not. Recently, SpliceAI, a novel deep residual neural network tool, has been developed for splicing prediction. Methodologically distinct from previous approaches that either rely on human-engineered features and/or which focus on short nucleotide windows adjoining exon-intron boundaries, SpliceAI assesses splicing determinants by evaluating 10,000 nucleotides of flanking contextual sequence to predict the functional role in splicing of each position in the pre-mRNA transcript. Herein, we evaluated the performance of SpliceAI in the context of three datasets of GT>GC variants, all of which had been characterized functionally in terms of their impact on mRNA splicing. The first two datasets refer to our recently described “in vivo” dataset of 45 disease-causing GT>GC variants and the “in vitro” dataset of 103 GT>GC substitutions. The third dataset comprised 12 BRCA1 GT>GC variants that were recently analyzed by saturation genome editing. We processed all GT>GC variants using the default settings of SpliceAI. Comparison of the SpliceAI-predicted and experimentally obtained functional impact assessments of the analyzed GT>GC variants revealed that although SpliceAI performed rather better than other prediction tools, it was still far from perfect. A key issue is that the impact of GT>GC (as well as GT>GA or +2T>A) variants that generated wild-type transcripts represents a quantitative change that can vary from barely detectable to almost full expression of wild-type transcripts, with wild-type transcripts often co-existing with aberrantly spliced transcripts. Our findings highlight the challenges that we still face in attempting to accurately identify splice-altering variants.

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Testing the antimicrobial activity of essential oils

Acta Agraria Debreceniensis ◽

10.34101/actaagrar/44/2609 ◽

2011 ◽

pp. 71-74

Author(s):

Adela Frankova ◽

Pavel Kloucek ◽

Jakub Smid ◽

Lenka Nedorostova

Keyword(s):

Antimicrobial Activity ◽

Essential Oils ◽

Vapor Phase ◽

Salmonella Enteritidis ◽

Food Preservatives ◽

Fast Screening ◽

Wide Range ◽

In Vivo Testing

The vapor phase of some essential oils proved to have antimicrobial activity. Utilization of the vapor phase of Eos is presently understood as one of the possible alternatives to synthetic food preservatives which could be used in the future. However, testing the vapor phase of EOs against microorganisms causing food-borne diseases (e.g. Salmonella enteritidis or Staphylococcus aureus) or food spoilage is relatively new. Consequently, due to the large number of known EOs, research on their antimicrobial activity is still largely in the phase of in vitro rather than in vivo testing. Moreover, no standard and reliable method for fast screening of a wide range of samples exists. Thus, the aim of this study is to show results concerning tests of the antimicrobial activity of EOs against S. enteritidis or S. aureus, which were conducted by two modifications of the disc volatilization method we developed. The lately developed method has the potential to become widely used for fast screening of EO antimicrobial activity in the vapor phase.

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Different SUMO paralogues determine the fate of wild-type and mutant CFTRs: biogenesis versus degradation

Molecular Biology of the Cell ◽

10.1091/mbc.e18-04-0252 ◽

2019 ◽

Vol 30 (1) ◽

pp. 4-16 ◽

Cited By ~ 3

Author(s):

Xiaoyan Gong ◽

Yong Liao ◽

Annette Ahner ◽

Mads Breum Larsen ◽

Xiaohui Wang ◽

...

Keyword(s):

Physiological Role ◽

Transmembrane Conductance Regulator ◽

Wild Type ◽

Transmembrane Conductance ◽

Specific Effects ◽

Ubiquitin Conjugation ◽

Protein Array Analysis ◽

The Impact

A pathway for cystic fibrosis transmembrane conductance regulator (CFTR) degradation is initiated by Hsp27, which cooperates with Ubc9 and binds to the common F508del mutant to modify it with SUMO-2/3. These SUMO paralogues form polychains, which are recognized by the ubiquitin ligase, RNF4, for proteosomal degradation. Here, protein array analysis identified the SUMO E3, protein inhibitor of activated STAT 4 (PIAS4), which increased wild-type (WT) and F508del CFTR biogenesis in CFBE airway cells. PIAS4 increased immature CFTR threefold and doubled expression of mature CFTR, detected by biochemical and functional assays. In cycloheximide chase assays, PIAS4 slowed immature F508del degradation threefold and stabilized mature WT CFTR at the plasma membrance. PIAS4 knockdown reduced WT and F508del CFTR expression by 40–50%, suggesting a physiological role in CFTR biogenesis. PIAS4 modified F508del CFTR with SUMO-1 in vivo and reduced its conjugation to SUMO-2/3. These SUMO paralogue-specific effects of PIAS4 were reproduced in vitro using purified F508del nucleotide-binding domain 1 and SUMOylation reaction components. PIAS4 reduced endogenous ubiquitin conjugation to F508del CFTR by ∼50% and blocked the impact of RNF4 on mutant CFTR disposal. These findings indicate that different SUMO paralogues determine the fates of WT and mutant CFTRs, and they suggest that a paralogue switch during biogenesis can direct these proteins to different outcomes: biogenesis versus degradation.

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Comparative phytochemical analysis of Phlogacanthus thyrsiflorus Nees: Implications on attenuation of pro-oxidants and pathogen virulence in Caenorhabditis elegans model system

Asian Journal of Pharmaceutical and Clinical Research ◽

10.22159/ajpcr.2017.v10i5.17319 ◽

2017 ◽

Vol 10 (5) ◽

pp. 361

Author(s):

Kitlangki Suchiang ◽

Nitasha H Kayde

Keyword(s):

Oxidative Stress ◽

Free Radicals ◽

Free Radical ◽

Caenorhabditis Elegans ◽

Model System ◽

Wild Type ◽

C Elegans ◽

Mutant Strains

Background: Phlogacanthus thyrsiflorus Nees (P. thyrsiflorus) of Acanthaceae family is endogenous to sub-tropical Himalayas. It has been reported to be used traditionally in Jaintia tribe of Meghalaya, India for treatment of many ailments.Objectives: The aim was to detect the active compounds present in the leaves for evaluation of in vitro free radicals scavenging potentials. Leaves protective actions in vivo will be investigated using Caenorhabditis elegans (C. elegans) model system utilizing wild type and mutant strains and the phenomena of host-pathogens interactions.Materials and methods: Gas chromatography/ Mass spectrometry (GC/MS) was used for detection of different compounds present. The versatility of leaf extracts to scavenge different free radicals generated in vitro was assessed with different in vitro methods. Survival analysis of wild type and mutant strains C. elegans under enhanced pro-oxidants exposure was investigated in vivo. Fast killing assay was also performed to study the extracts modulatory activity on host C. elegans survival under pathogen Pseudomonas aeruginosa infection.Results: Forty compounds were detected in methanolic fraction of the extract with variable percentages. Both aqueous and methanol extract possessed remarkable, versatile free radical scavenging activity irrespective of the types of free radical generated. The in vivo experiments are in compliance, with observable increased survival ability percentage of C. elegans under intense exogenous oxidative stress and pathogen infection.Conclusion: Our findings enlightened the different compounds present with versatility of P. thyrsiflorus in tackling different free radicals generated both in vitro and in vivo that highly support for its candidature as a good antioxidant source. Our findings may justify the historical relevance of this plant in herbal remedies that could form the basis for inquiry of new active principles.Keywords: Free radicals, Oxidative stress, Caenorhabditis elegans, Phlogacanthus thyrsiflorus, Phytochemicals

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FSMP-07. CYSTATHIONINE-Γ-LYASE DRIVES ANTIOXIDANT DEFENSE IN CYSTEINE-RESTRICTED IDH1 MUTANT ASTROCYTOMAS

Neuro-Oncology Advances ◽

10.1093/noajnl/vdab024.071 ◽

2020 ◽

Vol 3 (Supplement_1) ◽

pp. i17-i17

Author(s):

Andrés Cano-Galiano ◽

Anais Oudin ◽

Fred Fack ◽

Maria-Francesca Allega ◽

David Sumpton ◽

...

Keyword(s):

Antioxidant Defense ◽

De Novo ◽

Idh1 Mutation ◽

Wild Type ◽

Isocitrate Dehydrogenase 1 ◽

Astrocytic Gliomas ◽

The Impact ◽

Reducing Potential

Abstract Mutations in isocitrate dehydrogenase 1 or 2 (IDH1/2) define glioma subtypes and are considered primary events in gliomagenesis, impacting tumor epigenetics and metabolism. IDH enzymes are crucial for the generation of reducing potential, yet the impact of the mutation on the cellular antioxidant system is not understood. Here, we investigate how glutathione (GSH) levels are maintained in IDH1 mutant gliomas, despite an altered NADPH/NADP balance. We find that IDH1 mutant astrocytomas specifically upregulate cystathionine γ-lyase (CSE), the enzyme responsible for cysteine production upstream of GSH biosynthesis. Genetic and chemical interference with CSE in patient-derived glioma cells carrying the endogenous IDH1 mutation, sensitized tumor cells to cysteine depletion, an effect not observed in IDH1 wild-type gliomas. This correlated with reduced GSH synthesis as shown by in vitro and in vivo serine tracing and led to delayed tumor growth in mice. Thus we show that IDH1 mutant astrocytic gliomas critically rely on NADPH-independent de novo GSH synthesis to maintain the antioxidant defense, which uncovers a novel metabolic vulnerability in this dismal disease.

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