TY - JOUR
T1 - An automated fluorescence videomicroscopy assay for the detection of mitotic catastrophe
AU - Rello-Varona, S.
AU - Kepp, O.
AU - Vitale, I.
AU - Michaud, M.
AU - Senovilla, L.
AU - Jemaà, M.
AU - Joza, N.
AU - Galluzzi, L.
AU - Castedo, M.
AU - Kroemer, G.
N1 - Funding Information:
Acknowledgements. We thank Verónica Labrador Catarero from the Optical and Confocal Microscopy Facility of Centro de Biología Molecular Severo Ochoa (Madrid, Spain) for helpful discussion. GK is supported by the Ligue Nationale contre le Cancer (Equipe labellisée), Agence Nationale pour la Recherche, European Commission (Apo-Sys, ChemoRes, ApopTrain), Fondation pour la Recherche Médicale, Institut National du Cancer, Cancéropôle Ile-de-France. SR-V, MM and LS are supported by FRM. OK is supported by EMBO. LG is supported by the Apo-Sys consortium of the European Union. MC is supported by the Association pour la Recherche sur le Cancer (ARC).
PY - 2010/2/1
Y1 - 2010/2/1
N2 - Mitotic catastrophe can be defined as a cell death mode that occurs during or shortly after a prolonged/aberrant mitosis, and can show apoptotic or necrotic features. However, conventional procedures for the detection of apoptosis or necrosis, including biochemical bulk assays and cytofluorometric techniques, cannot discriminate among pre-mitotic, mitotic and post-mitotic death, and hence are inappropriate to monitor mitotic catastrophe. To address this issue, we generated isogenic human colon carcinoma cell lines that differ in ploidy and p53 status, yet express similar amounts of fluorescent biosensors that allow for the visualization of chromatin (histone H2B coupled to green fluorescent protein (GFP)) and centrosomes (centrin coupled to the Discosoma striata red fluorescent protein (DsRed)). By combining high-resolution fluorescence videomicroscopy and automated image analysis, we established protocols and settings for the simultaneous assessment of ploidy, mitosis, centrosome number and cell death (which in our model system occurs mainly by apoptosis). Time-lapse videomicroscopy showed that this approach can be used for the high-throughput detection of mitotic catastrophe induced by three mechanistically distinct anti-mitotic agents (dimethylenastron (DIMEN), nocodazole (NDZ) and paclitaxel (PTX)), and - in this context - revealed an important role of p53 in the control of centrosome number.
AB - Mitotic catastrophe can be defined as a cell death mode that occurs during or shortly after a prolonged/aberrant mitosis, and can show apoptotic or necrotic features. However, conventional procedures for the detection of apoptosis or necrosis, including biochemical bulk assays and cytofluorometric techniques, cannot discriminate among pre-mitotic, mitotic and post-mitotic death, and hence are inappropriate to monitor mitotic catastrophe. To address this issue, we generated isogenic human colon carcinoma cell lines that differ in ploidy and p53 status, yet express similar amounts of fluorescent biosensors that allow for the visualization of chromatin (histone H2B coupled to green fluorescent protein (GFP)) and centrosomes (centrin coupled to the Discosoma striata red fluorescent protein (DsRed)). By combining high-resolution fluorescence videomicroscopy and automated image analysis, we established protocols and settings for the simultaneous assessment of ploidy, mitosis, centrosome number and cell death (which in our model system occurs mainly by apoptosis). Time-lapse videomicroscopy showed that this approach can be used for the high-throughput detection of mitotic catastrophe induced by three mechanistically distinct anti-mitotic agents (dimethylenastron (DIMEN), nocodazole (NDZ) and paclitaxel (PTX)), and - in this context - revealed an important role of p53 in the control of centrosome number.
KW - Apoptosis
KW - Cell death detection
KW - Centrosome
KW - Mitotic catastrophe
KW - Multipolar mitosis
KW - p53
UR - http://www.scopus.com/inward/record.url?scp=79952164297&partnerID=8YFLogxK
U2 - 10.1038/cddis.2010.6
DO - 10.1038/cddis.2010.6
M3 - Article
C2 - 21364633
AN - SCOPUS:79952164297
SN - 2041-4889
VL - 1
JO - Cell Death and Disease
JF - Cell Death and Disease
IS - 2
M1 - e25
ER -