Angelina Huseinovic  Annelieke Jaspers  Annina P. van Splunter  Hanne Srgrd  Saskia M. Wilting  Dorian R. A. Swarts  Ida H. van der Meulen  Victor W. van Beusechem  Rene X. de Menezes  Renske D. M. Steenbergen 《International journal of molecular sciences》2022,23(9)

The progression of anchorage-dependent epithelial cells to anchorage-independent growth represents a critical hallmark of malignant transformation. Using an in vitro model of human papillomavirus (HPV)-induced transformation, we previously showed that acquisition of anchorage-independent growth is associated with marked (epi)genetic changes, including altered expression of microRNAs. However, the laborious nature of the conventional growth method in soft agar to measure this phenotype hampers a high-throughput analysis. We developed alternative functional screening methods using 96- and 384-well ultra-low attachment plates to systematically investigate microRNAs regulating anchorage-independent growth. SiHa cervical cancer cells were transfected with a microRNA mimic library (n = 2019) and evaluated for cell viability. We identified 84 microRNAs that consistently suppressed growth in three independent experiments. Further validation in three cell lines and comparison of growth in adherent and ultra-low attachment plates yielded 40 microRNAs that specifically reduced anchorage-independent growth. In conclusion, ultra-low attachment plates are a promising alternative for soft-agar assays to study anchorage-independent growth and are suitable for high-throughput functional screening. Anchorage independence suppressing microRNAs identified through our screen were successfully validated in three cell lines. These microRNAs may provide specific biomarkers for detecting and treating HPV-induced precancerous lesions progressing to invasive cancer, the most critical stage during cervical cancer development.  相似文献   

    

Manouk K. Bos  Teoman Deger  Stefan Sleijfer  John W. M. Martens  Saskia M. Wilting 《International journal of molecular sciences》2022,23(10)

ESR1 methylation was proposed as mechanism for endocrine resistance in metastatic breast cancer patients. To evaluate its potential as a minimally invasive biomarker, we investigated the feasibility of measuring ESR1 methylation in cell-free DNA (cfDNA) and its association with endocrine resistance. First, we provided evidence that demethylation in vitro restores ER expression. Subsequently, we found that ESR1 methylation in cfDNA was not enriched in endocrine-resistant versus endocrine-sensitive patients. Interestingly, we found a correlation between ESR1 methylation and age. Publicly available data confirm an age-related increase in ESR1 methylation in leukocytes, confounding the determination of the ESR1 methylation status of tumors using cfDNA.  相似文献   

    

Maëlle Deshoux  Baptiste Monsion  Elodie Pichon  Jaime Jimnez  Arnzazu Moreno  Bastien Cayrol  Gaël Thbaud  Sam T. Mugford  Saskia A. Hogenhout  Stphane Blanc  Alberto Fereres  Marilyne Uzest 《International journal of molecular sciences》2022,23(23)

To avoid the activation of plant defenses and ensure sustained feeding, aphids are assumed to use their mouthparts to deliver effectors into plant cells. A recent study has shown that effectors detected near feeding sites are differentially distributed in plant tissues. However, the precise process of effector delivery into specific plant compartments is unknown. The acrostyle, a cuticular organ located at the tip of maxillary stylets that transiently binds plant viruses via its stylin proteins, may participate in this specific delivery process. Here, we demonstrate that Mp10, a saliva effector released into the plant cytoplasm during aphid probing, binds to the acrostyles of Acyrthosiphon pisum and Myzus persicae. The effector probably interacts with Stylin-03 as a lowered Mp10-binding to the acrostyle was observed upon RNAi-mediated reduction in Stylin-03 production. In addition, Stylin-03 and Stylin-01 RNAi aphids exhibited changes in their feeding behavior as evidenced by electrical penetration graph experiments showing longer aphid probing behaviors associated with watery saliva release into the cytoplasm of plant cells. Taken together, these data demonstrate that the acrostyle also has effector binding capacity and supports its role in the delivery of aphid effectors into plant cells.  相似文献   

    

Annett Kuehne  Saskia Floerl  Yohannes Hagos 《International journal of molecular sciences》2022,23(24)

Multiple drugs are used to treat various indications as well as pesticides that are ingested unintentionally and enter the bloodstream. The residence time or bioavailability of these substances in circulation depends on several mechanisms, such as drug–drug interaction (DDI), drug–pesticide interaction, metabolizing enzymes and the hepatic and renal transport systems, involved in the elimination of the compounds from the body. One of these transporters is the Organic Cation Transporter 2 (OCT2) member of the solute carrier (SLC22) transporter family. OCT2 is highly expressed in the proximal tubule epithelial cells in human and mouse kidney, where it mediates the uptake of endogenous organic cations as well as numerous drugs and xenobiotics, and contributes to the first step of renal clearance. In this study, we examined OCT2 on two subjects: First, the transferability of data from mouse to human, since mice are initially examined in the development of new drugs to assess the renal excretion of organic cations. Second, to what extent the choice of substrate affects the properties of an inhibitor. For this purpose, the functional properties of hOCT2 and mOct2 were validated under the same experimental conditions with the known substrates metformin and 1-Methyl-4-phenylpyridinium iodide (MPP). While hOCT2 and mOct2 showed very low affinities for metformin with K_m values of 3.9 mM and 3.5 mM, the affinity of hOCT2 and mOct2 for MPP (62 and 40 µM) was 64- and 89-fold higher, respectively. For our positive control inhibitor decynium22, we determined the following IC₅₀ values for hOCT2 and mOct2: 2.2 and 2.6 µM for metformin uptake, and 16 and 6.9 µM for MPP uptake. A correlation analysis of the inhibitory effects of 13 drugs and 9 pesticides on hOCT2- and mOct2-mediated transport of metformin showed a correlation coefficient R² of 0.88, indicating good interspecies correlation. Nevertheless, the bioenhancer elacridar and the fungicide imazalil showed species-dependent inhibitory potentials. Concentration-dependent inhibition of hOCT2- and mOct2-mediated metformin uptake by elacridar showed IC₅₀ values of 20 µM and 1.9 µM and by imazalil 4.7 µM and 0.58 µM, respectively. In conclusion, although our data show comparable species-independent interactions for most compounds, there can be large species–specific differences in the interactions of individual compounds, which should be considered when extrapolating data from mice to humans. Furthermore, a comparison of the inhibitory potential of elacridar and imazalil on metformin uptake with that on MPP uptake reveals substrate-dependent differences in hOCT2 and mOct2 for both inhibitors. Therefore, it might be useful to test two different substrates in inhibition studies.  相似文献   

    

Saskia Schmidt  Catherine Jane Messner  Carine Gaiser  Carina Hmmerli  Laura Suter-Dick 《International journal of molecular sciences》2022,23(23)

Low-dose methotrexate (MTX) is a standard therapy for rheumatoid arthritis due to its low cost and efficacy. Despite these benefits, MTX has been reported to cause chronic drug-induced liver injury, namely liver fibrosis. The hallmark of liver fibrosis is excessive scarring of liver tissue, triggered by hepatocellular injury and subsequent activation of hepatic stellate cells (HSCs). However, little is known about the precise mechanisms through which MTX causes hepatocellular damage and activates HSCs. Here, we investigated the mechanisms leading to hepatocyte injury in HepaRG and used immortalized stellate cells (hTERT-HSC) to elucidate the mechanisms leading to HSC activation by exposing mono- and co-cultures of HepaRG and hTERT-HSC to MTX. The results showed that at least two mechanisms are involved in MTX-induced toxicity in HepaRG: (i) oxidative stress through depletion of glutathione (GSH) and (ii) impairment of cellular respiration in a GSH-independent manner. Furthermore, we measured increased levels of endoplasmic reticulum (ER) stress in activated HSC following MTX treatment. In conclusion, we established a human-relevant in vitro model to gain mechanistical insights into MTX-induced hepatotoxicity, linked oxidative stress in HepaRG to a GSH-dependent and -independent pathway, and hypothesize that not only oxidative stress in hepatocytes but also ER stress in HSCs contribute to MTX-induced activation of HSCs.  相似文献   

    

Christina B. Bielmeier  Saskia Roth  Sabrina I. Schmitt  Stefaniya K. Boneva  Anja Schlecht  Mario Vallon  Ernst R. Tamm  Süleyman Ergün  Andreas Neueder  Barbara M. Braunger 《International journal of molecular sciences》2021,22(12)

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Saskia Schwarzer  Michael Radzieta  Slade O. Jensen  Matthew Malone 《International journal of molecular sciences》2021,22(17)

A topical desiccating wound agent containing methanesulfonic acid, dimethylsulfoxide and amorphous silica was evaluated in three in vitro models for its efficacy against biofilms produced by Pseudomonas aeruginosa (ATCC-15442) and Staphylococcus aureus (ATCC-6538). The in vitro biofilm models used were; the MBEC Assay^®, Centre for Disease Control (CDC) Biofilm Reactor^® and a Semi-solid biofilm model. A 30-s exposure of a topical wound desiccating agent was used in each model. A complete eradication of viable cells was demonstrated in all models for both strains (p < 0.0001). Imaging with scanning electron microscopy (SEM) was performed where possible. All three models demonstrated complete eradication of viable cells with a 30 s application of a topical wound desiccating agent.  相似文献   

    

Saskia Franke  Kati Fröhlich  Susanne Werner  Volker Böhm  Friedrich Schöne 《European Journal of Lipid Science and Technology》2010,112(10):1122-1129

Carotenoids and vitamin E in oils from the market – 6 rapeseed and 6 sunflower oils, half of each cold pressed and refined – and in the oils of rape, sunflower, flax and safflower as well as the respective seeds and press cakes from a local oil mill were quantified by HPLC. Furthermore, a photometric determination of carotenoid content was tested and checked against the chromatographic method. In the cold pressed oils minor amounts of xanthophylls (all‐E)‐lutein and (all‐E)‐zeaxanthin were determined. With exception of traces of (all‐E)‐β‐carotene in cold‐pressed rapeseed oil this provitamin A active compound did not occur. Cold pressed rapeseed oils contained 0.5–1.5 mg total carotenoids/100 g which was manifold the content of the further oils. Vitamin E was found in all vegetable oils at plant‐typic tocopherol patterns. The photometric determination of carotenoids resulted in significantly higher concentrations compared to the HPLC. This overestimation bases on the carotenoid pattern which was validated by comparison with known high‐carotenoid materials, i.e. maize flour with an abundant amount of xanthophylls and carrots with an abundant amount of carotenes.  相似文献   

    

Alba Tres  Rory O'Neill  Saskia M. van Ruth 《Lipid Technology》2011,23(2):40-42

Organic products (such as organic eggs) usually have a higher price than the corresponding conventional products. This makes organic products susceptible to fraud. Administrative controls are conducted to detect this type of frauds. However, an analytical verification of the organic identity of food products would be very useful in this respect. It is unlikely that there is a single compound, or a few compounds, that differ sufficiently between organic and conventional eggs to be used as a marker of organic identity. Instead, fingerprinting of the organic products (that is to say, analyzing a wide range of compounds instead of only a few) might be used as a tool for their verification. Here, we have used the fatty acid composition of egg yolks as a fingerprint to verify the organic identity of eggs. From the fingerprints, chemometric models were built which predict the identity of eggs (organic or conventional) with high success rates.  相似文献   

    

Sven Käselau  Saskia Scheel  Linnea Petersson  Chau‐Hon Ho  Gerrit A Luinstra 《Polymer International》2018,67(10):1359-1367

Composites of linear low‐density polyethylene were obtained in toluene slurry by in situ copolymerization of ethylene and 1‐octene in the presence of untreated magnesium oxide–hydroxide nanoparticles (MgO@Mg(OH)₂) of ±50 nm and such treated with dibutylmagnesium (DBM) as support for a bis(n‐butylcyclopentadienyl)zirconium dichloride–methylaluminoxane (MAO) catalyst system. Treatment of the nanoparticles with DBM (0.5–6 mmol g^?1 MgO@Mg(OH)₂) allows one to decrease the amount of MAO by 1.2 mmol Al g^?1 MgO@Mg(OH)₂, while reaching the same average catalyst activity and a finer distribution of the particles. Energy‐dispersive X‐ray mapping shows that the MAO is mainly associated with the filler. The crystallinity of the matrix polymer decreases with filler content. © 2018 Society of Chemical Industry  相似文献