Isotropic 3D nuclear morphometry of normal, fibrocystic and malignant breast epithelial cells reveals new structural alterations

Grading schemes for breast cancer diagnosis are predominantly based on pathologists' qualitative assessment of altered nuclear structure from 2D brightfield microscopy images. However, cells are three-dimensional (3D) objects with features that are inherently 3D and thus poorly characterized in...

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Bibliographic Details
Published in:	PloS one Vol. 7; no. 1; p. e29230
Main Authors:	Nandakumar, Vivek, Kelbauskas, Laimonas, Hernandez, Kathryn F, Lintecum, Kelly M, Senechal, Patti, Bussey, Kimberly J, Davies, Paul C W, Johnson, Roger H, Meldrum, Deirdre R
Format:	Journal Article
Language:	English
Published:	United States Public Library of Science 05-01-2012 Public Library of Science (PLoS)
Subjects:	Architectural engineering Automation Biology Biomarkers Breast - pathology Breast cancer Breast Neoplasms - pathology Cancer Cancer diagnosis Cancer metastasis Cell growth Cell Line Cell Nucleus - pathology Cells (Biology) Cellular structure Chromatin Computation Computed tomography Cytopathology Cytoplasm Diagnosis Engineering Enzymes Epithelial cells Epithelial Cells - pathology Epithelium Evaluation Female Fibrocystic Breast Disease - pathology Gene expression Humans Image contrast Imaging, Three-Dimensional - methods Kruskal-Wallis test Life sciences Lung cancer Malignancy Medical diagnosis Medical imaging Medical research Medicine Metastases Metastasis Microscopy Morphology Morphometry Neoplasm Metastasis Nuclear structure Nuclei Nuclei (cytology) Nucleoli Populations Quality Three dimensional imaging Tumors Variance analysis Variation United States > US Arizona
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Summary:	Grading schemes for breast cancer diagnosis are predominantly based on pathologists' qualitative assessment of altered nuclear structure from 2D brightfield microscopy images. However, cells are three-dimensional (3D) objects with features that are inherently 3D and thus poorly characterized in 2D. Our goal is to quantitatively characterize nuclear structure in 3D, assess its variation with malignancy, and investigate whether such variation correlates with standard nuclear grading criteria. We applied micro-optical computed tomographic imaging and automated 3D nuclear morphometry to quantify and compare morphological variations between human cell lines derived from normal, benign fibrocystic or malignant breast epithelium. To reproduce the appearance and contrast in clinical cytopathology images, we stained cells with hematoxylin and eosin and obtained 3D images of 150 individual stained cells of each cell type at sub-micron, isotropic resolution. Applying volumetric image analyses, we computed 42 3D morphological and textural descriptors of cellular and nuclear structure. We observed four distinct nuclear shape categories, the predominant being a mushroom cap shape. Cell and nuclear volumes increased from normal to fibrocystic to metastatic type, but there was little difference in the volume ratio of nucleus to cytoplasm (N/C ratio) between the lines. Abnormal cell nuclei had more nucleoli, markedly higher density and clumpier chromatin organization compared to normal. Nuclei of non-tumorigenic, fibrocystic cells exhibited larger textural variations than metastatic cell nuclei. At p<0.0025 by ANOVA and Kruskal-Wallis tests, 90% of our computed descriptors statistically differentiated control from abnormal cell populations, but only 69% of these features statistically differentiated the fibrocystic from the metastatic cell populations. Our results provide a new perspective on nuclear structure variations associated with malignancy and point to the value of automated quantitative 3D nuclear morphometry as an objective tool to enable development of sensitive and specific nuclear grade classification in breast cancer diagnosis.
Bibliography:	ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23 Conceived and designed the experiments: VN LK KJB RHJ. Performed the experiments: VN KFH KML PS. Analyzed the data: VN LK KJB RHJ. Contributed reagents/materials/analysis tools: VN PS. Wrote the paper: VN KJB RHJ. Critical manuscript review: PCWD DRM. Overall research guidance: DRM.
ISSN:	1932-6203 1932-6203
DOI:	10.1371/journal.pone.0029230