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Video time lapse imaging of mitotic catastrophe events in response to thiopurines
Author(s) -
Patel Shikha,
Pittman Douglas
Publication year - 2013
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.27.1_supplement.1027.2
Subject(s) - multinucleate , mitosis , mitotic catastrophe , programmed cell death , interphase , cell division , biology , cell , dna damage , cell cycle , dna repair , cancer research , microbiology and biotechnology , genetics , dna , apoptosis
Thiopurines are a class of chemotherapy drugs used as immunosuppressants in organ transplant patients and for the treatment of childhood cancers. However, approximately 40% of patients develop life‐threatening problems later in life as a direct result of treatment. 6‐thioguanine (6TG) is a thiopurine directly incorporated into the DNA. Processing of the damage by the mismatch DNA repair pathway leads to the formation of double‐stranded breaks repaired by the homologous recombination (HR) machinery. RAD51D is an HR protein, and absence of RAD51D leads to the formation of large multinucleated cells in response to 6TG treatment. Using live cell imaging, I am attempting to determine how these multinucleated cells form. Treated and untreated mouse embryo fibroblasts (MEFs) were cultured in a 37°C chamber containing 5% CO 2 and visualized using a Leica ASMDW microscope. Images were captured every three minutes and translated into individual videos. Cell division times and outcomes were determined by manually following each cell and daughter cells. The data suggest multinucleated cells form after aberrant mitotic events and subsequent fusion of the daughter cells during the second cell cycle following treatment. Current data also indicate that multinucleation is more prevalent at lower doses while interphase arrest is more common at a higher dose of 6TG. These studies have the potential to determine the mechanisms of thiopurine‐induced secondary cancers and may possibly be used to identify individuals who may benefit from alternative treatments.