Matching Items (4)
Description
Background
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.
Methodology
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.
Principal Findings
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.
Conclusions
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.
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.
Methodology
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.
Principal Findings
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.
Conclusions
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.
ContributorsNandakumar, Vivek (Author) / Kelbauskas, Laimonas (Author) / Hernandez, Kathryn (Author) / Lintecum, Kelly (Author) / Senechal, Patti (Author) / Bussey, Kimberly (Author) / Davies, Paul (Author) / Johnson, Roger (Author) / Meldrum, Deirdre (Author) / Ira A. Fulton Schools of Engineering (Contributor) / School of Electrical, Computer and Energy Engineering (Contributor) / Biodesign Institute (Contributor) / Center for Biosignatures Discovery Automation (Contributor) / College of Liberal Arts and Sciences (Contributor) / School of Life Sciences (Contributor) / Department of Physics (Contributor)
Created2012-01-05
Description
ABSTRACT Genomes are biologically complex entities where an alteration in structure can yield no effect, or have a devastating effect on many pathways. Most of the focus has been on translocations that generate fusion proteins. However, this is only one of many outcomes. Recent work suggests alterations in topologically associated domains (TADs) can lead to changes in gene expression. It is hypothesized that alterations in genome structure can disrupt TADs leading to an alteration in the variability of gene expression within the contained gene expression neighborhood defined by the TAD. To test this hypothesis, variability of gene expression for genes contained within TADs between 37 cancer cell lines from the NCI-60 cell line panel was compared with normal expression data for the corresponding tissues of origin. Those results were correlated with the data on structural events within the NCI-60 cell lines that would disrupt a TAD. It was observed that 2.4% of the TADs displayed altered variance in gene expression when comparing cancer to normal tissue. Using array CGH data from the cancer cell lines to map breakpoints within TADS, it was discovered that altered variance is always associated with a TAD disrupted by a breakpoint, but a breakpoint within a TAD does not always lead to altered variance. TADs with altered variance in gene expression were no different in size than those without altered variance. There is evidence of recurrent pan-cancer alteration in variance for eleven genes within two TADs on two chromosomes (Chromosome 10 & 19) for all 37 cell lines. The genes located within these TADs are enriched in pathways related to RNA processing. This study supports altered variance as a signal of a breakpoint with a functional consequence.
ContributorsDunham, Jocelen Michaela (Author) / Kanthaswamy, Sreethan (Thesis advisor) / Mancenido, Michelle (Thesis advisor) / Bussey, Kimberly J. (Committee member) / Arizona State University (Publisher)
Created2022
Description
Fusion genes, arising from chromosomal translocations through nonallelic homologous recombination (NAHR), are pivotal in oncogenesis, leading to the formation of fusion proteins that contribute to cancer’s aggressive nature. The atavism theory posits that cancer is a throwback to an ancient cellular state, with reactivated ancestral cellular mechanisms driving uncontrolled growth and other cancerous traits. By comparing the evolutionary ages of the structural homologs of fusion proteins with those of their parental gene pairs, this study aims to determine whether these fusion proteins recapitulate ancient protein structures, thereby supporting the atavism theory.Utilizing data from the COSMIC database, fusion genes were constructed according to their corresponding cDNA sequences from parent gene pairs, and the 3D structures of resultant fusion proteins were predicted by using AlphaFold. Subsequent VAST analysis identified structural homologies with ancient proteins. The ages of original and fusion proteins were inferred by mapping homologous groups from the Ensembl Compara database to identify common ancestors. The TimeTree database was then used to assign gene ages based on the divergence of the most distantly related species in these groups. Finally, comparing these ages identified ancestral resemblances. The findings of this project demonstrate homology between the structures of most fusion proteins and those of ancient proteins found in humans, yeast, and bacteria, suggesting the re-emergency of ancient protein structures in cancer cells due to recurrent translocations. (Permutation test, p=0.0201). Additionally, a large portion (68%) of the examined fusion genes comprises one gene predating the advent of multicellularity and another emerging concurrently with or after this evolutionary milestone (One-sample proportions test, X-squared=13.291, df=1, p=0.00027). These results support the atavism theory, suggesting that such fusion events might bridge evolutionary gaps between unicellular and multicellular life forms. This could potentially explain the mechanisms behind cancer’s tendency to forsake multicellular characteristics, thereby enhancing malignancy. By illustrating how chromosomal translocations in cancer might be tapping into primordial protein architectures, this study not only provides evidence for the atavism theory but also opens new avenues for understanding cancer’s evolutionary underpinnings. This could lead to novel therapeutic strategies by exploiting the ancient vulnerabilities revealed through chromosomal translocations.
ContributorsSun, Shuyu (Author) / Bussey, Kimberly J. (Thesis advisor) / Broatch, Jennifer (Thesis advisor) / Marshall, Pamela A. (Committee member) / Arizona State University (Publisher)
Created2024
Description
The histone deacetylase (HDAC) inhibitor vorinostat has received significant attention in recent years as an ‘epigenetic’ drug used to treat solid tumors. However, its mechanisms of action are not entirely understood, particularly with regard to its interaction with the aberrations in 3D nuclear structure that accompany neoplastic progression. We investigated the impact of vorinostat on human esophageal epithelial cell lines derived from normal, metaplastic (pre-cancerous), and malignant tissue. Using a combination of novel optical computed tomography (CT)-based quantitative 3D absorption microscopy and conventional confocal fluorescence microscopy, we show that subjecting malignant cells to vorinostat preferentially alters their 3D nuclear architecture relative to non-cancerous cells. Optical CT (cell CT) imaging of fixed single cells showed that drug-treated cancer cells exhibit significant alterations in nuclear morphometry. Confocal microscopy revealed that vorinostat caused changes in the distribution of H3K9ac-marked euchromatin and H3K9me3-marked constitutive heterochromatin. Additionally, 3D immuno-FISH showed that drug-induced expression of the DNA repair gene MGMT was accompanied by spatial relocation toward the center of the nucleus in the nuclei of metaplastic but not in non-neoplastic cells. Our data suggest that vorinostat’s differential modulation of 3D nuclear architecture in normal and abnormal cells could play a functional role in its anti-cancer action.
ContributorsNandakumar, Vivek (Author) / Hansen Katdare, Nanna (Author) / Glenn, Honor (Author) / Han, Jessica (Author) / Helland, Stephanie (Author) / Hernandez, Kathryn (Author) / Senechal, Patti (Author) / Johnson, Roger (Author) / Bussey, Kimberly J. (Author) / Meldrum, Deirdre (Author) / Ira A. Fulton Schools of Engineering (Contributor)
Created2016-08-09