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Lucie Hloušková Supervisor : Ale ks andra Kapuscik

28 . 07. 2011. Lucie Hloušková Supervisor : Ale ks andra Kapuscik. Application of cytotoxicity assays on HeLa and HepG2 cells for estimation of health risk for humans as response to cyanobacterial metabolites. Introduction. Places of finding Unicellular and colonial species

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Lucie Hloušková Supervisor : Ale ks andra Kapuscik

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  1. 28. 07. 2011 Lucie Hloušková Supervisor : Aleksandra Kapuscik Application of cytotoxicity assays on HeLa and HepG2 cells for estimation of health risk for humans as response to cyanobacterial metabolites.

  2. Introduction Places of finding Unicellular and colonial species Secondary metabolites of cyanobacteria Biological activity - antifungal, anticancer, immunosupresants, ion channel blockers, enzyme inhibitors, cytotoxic agents…

  3. Aim of the project Characterization of toxic effects ofcyanobacterial metabolites tovarious human and murine cancer cell lines

  4. Material 15 strains(Nostoc, Phormidium, Leptolyngbia,Oscillatoria and Wiollea) The cyanobacterial biomass was lyophilized and used for experiments

  5. Methods • Sample preparation (70% MeOH extraction) • Cytotoxicity methyl-tetrazolium test (MTT) performed on cell lines: Hep G2(human liver carcinoma), HeLa (human cervix carcinoma), Sp2 (murine myeloma), Yac-1 (murine lymphoma) • Caspase 3/7 assay – detection of apoptosis (Hep G2 cell line)

  6. Our samples for testing…

  7. Cytotoxicity screening – MTT test + 10mg/mlextracts MTT (methyl-tetrazolium salt) • centrifugation • crystals dissolved in DMSO • absorbance measurement in 590nm yellow purplecrystals

  8. MTT results Cytotoxicity groups: non cytotoxic: >75% viability modetate cytotox: 50-75% strong cytotox: <50%

  9. Caspase assay results

  10. Photos Yac – 1 control Sp 2 control

  11. HeLa 77 5h HepG2 control

  12. Our work

  13. Conclusions • HeLa cells are the most resistant cell line to tested extracts, whereas Yac-1 shows the highest sensitivity • For some cyanobacterial strains a good cytotoxicity correlation to all tested cell lines has been proved • Both 77 - Phormidium sp. CALU 52 and 183 - Nostoc sp. CALU 1533 strains, in concentration 2 mg/ml, activated caspase 3 and 7 what correlates also with observed morfology

  14. Acknowledgements • to my supervisor Aleksandra Kapuscik for her help and advice, and to PavelHrouzek • my colleague Sofia Chysta • to all organizers of the Summer School Thankyouforyourattention !

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