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Nat Methods. 2015 Jun;12(6):577-85. doi: 10.1038/nmeth.3363. Epub 2015 Apr 13.

In vivo cell-cycle profiling in xenograft tumors by quantitative intravital microscopy.

Author information

1
Department of Cell Biology, Harvard Medical School, Boston, Massachusetts, USA.
2
Department of Systems Biology, Harvard Medical School, Boston, Massachusetts, USA.
3
Center for Systems Biology, Massachusetts General Hospital, Boston, Massachusetts, USA.
4
Molecular, Cellular and Developmental Biology, University of Colorado Boulder, Boulder, Colorado, USA.
5
1] Department of Systems Biology, Harvard Medical School, Boston, Massachusetts, USA. [2] Center for Systems Biology, Massachusetts General Hospital, Boston, Massachusetts, USA.

Abstract

Quantification of cell-cycle state at a single-cell level is essential to understand fundamental three-dimensional (3D) biological processes such as tissue development and cancer. Analysis of 3D in vivo images, however, is very challenging. Today's best practice, manual annotation of select image events, generates arbitrarily sampled data distributions, which are unsuitable for reliable mechanistic inferences. Here, we present an integrated workflow for quantitative in vivo cell-cycle profiling. It combines image analysis and machine learning methods for automated 3D segmentation and cell-cycle state identification of individual cell-nuclei with widely varying morphologies embedded in complex tumor environments. We applied our workflow to quantify cell-cycle effects of three antimitotic cancer drugs over 8 d in HT-1080 fibrosarcoma xenografts in living mice using a data set of 38,000 cells and compared the induced phenotypes. In contrast to results with 2D culture, observed mitotic arrest was relatively low, suggesting involvement of additional mechanisms in their antitumor effect in vivo.

PMID:
25867850
PMCID:
PMC4579269
DOI:
10.1038/nmeth.3363
[Indexed for MEDLINE]
Free PMC Article

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