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Iskalni niz: "ključne besede" (flow cytometry) .

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1.
HepG2 spheroids as a biosensor-like cell-based system for (geno)toxicity assessment
Martina Štampar, Sonja Žabkar, Metka Filipič, Bojana Žegura, 2022, izvirni znanstveni članek

Povzetek: 3D spheroids developed from HepG2 cells were used as a biosensor-like system for the detection of (geno)toxic effects induced by chemicals. Benzo(a)pyrene (B(a)P) and amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) with well-known mechanisms of action were used for system validation. HepG2 spheroids grown for 3 days were exposed to BaP and PhIP for 24 and 72 h. The growth and viability of spheroids were monitored by planimetry and Live/Dead staining of cells. Multi-parametric flow cytometric analysis was applied for simultaneous detection of specific end-effects including cell cycle analysis (Hoechst staining), cell proliferation (KI67 marker), and DNA double-strand breaks (ℽH2AX) induced by genotoxic compounds. Depending on the exposure concentration/time, BaP reduced spheroid growth, affected cell proliferation by arresting cells in S and G2 phase and induced DNA double-strand breaks (DSB). Simultaneous staining of ℽH2AX formation and cell cycle analysis revealed that after BaP (10 μM; 24 h) exposure 60% of cells in G0/G1 phase had DNA DSB, while after 72 h only 20% of cells contained DSB indicating efficient repair of DNA lesions. PhIP did not influence the spheroid size whereas accumulation of cells in the G2 phase occurred after both treatment times. The evaluation of DNA damage revealed that at 200 μM PhIP 50% of cells in G0/G1 phase had DNA DSB, which after 72-h exposure dropped to 40%, showing lower repair capacity of PhIP-induced DSB compared to BaP-induced. The developed approach using simultaneous detection of several parameters provides mechanistic data and thus contributes to more reliable genotoxicity assessment of chemicals as a high-content screening tool.
Ključne besede: in vitro 3D cell model, HepG2, flow cytometry, cell cycle, proliferation, DNA strand, breaks
Objavljeno v DiRROS: 16.07.2024; Ogledov: 110; Prenosov: 84
.pdf Celotno besedilo (6,21 MB)
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2.
Impact of deoxynivalenol and zearalenone as single and combined treatment on DNA, cell cycle and cell proliferation in HepG2 cells
Ana-Marija Domijan, Klara Hercog, Martina Štampar, Goran Gajski, Marko Gerić, Marijana Sokolović, Bojana Žegura, 2023, izvirni znanstveni članek

Povzetek: The study aimed to investigate toxicity and the mechanism of toxicity of two Fusarium mycotoxins, deoxynivalenol (DON) and zearalenone (ZEA). DON and ZEA were applied to HepG2 cells as single compounds and in combination at low environmentally relevant concentrations. HepG2 cells were exposed to DON (0.5, 1, and 2 µM), ZEA (5, 10, and 20 µM) or their combinations (1 µM DON + 5 µM ZEA, 1 µM DON + 10 µM ZEA and 1 µM DON + 20 µM ZEA) for 24 h and cell viability, DNA damage, cell cycle and proliferation were assessed. Both mycotoxins reduced cell viability, however, combined treatment with DON and ZEA resulted in higher reduction of cell viability. DON (1 µM) induced primary DNA damage, while DON (1 µM) in combination with higher ZEA concentrations showed antagonistic effects compared to DON alone at 1 µM. DON arrested HepG2 cells in G2 phase and significantly inhibited cell proliferation, while ZEA had no significant effect on cell cycle. The combined treatment with DON and ZEA arrested cells in G2 phase to a higher extend compared to treatment with single mycotoxins. Potentiating effect observed after DON and ZEA co-exposure at environmentally relevant concentrations indicates that in risk assessment and setting governments’ regulations, mixtures of mycotoxins should be considered.
Ključne besede: mycotoxins, comet assay, flow cytometry, co-exposure, food monitoring
Objavljeno v DiRROS: 12.07.2024; Ogledov: 248; Prenosov: 121
.pdf Celotno besedilo (1,60 MB)
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