Matching Items (162)
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
Viral protein U (Vpu) is a type-III integral membrane protein encoded by Human Immunodeficiency Virus-1 (HIV- 1). It is expressed in infected host cells and plays several roles in viral progeny escape from infected cells, including down-regulation of CD4 receptors. But key structure/function questions remain regarding the mechanisms by which the Vpu protein contributes to HIV-1 pathogenesis. Here we describe expression of Vpu in bacteria, its purification and characterization. We report the successful expression of PelB-Vpu in Escherichia coli using the leader peptide pectate lyase B (PelB) from Erwinia carotovora. The protein was detergent extractable and could be isolated in a very pure form. We demonstrate that the PelB signal peptide successfully targets Vpu to the cell membranes and inserts it as a type I membrane protein. PelB-Vpu was biophysically characterized by circular dichroism and dynamic light scattering experiments and was shown to be an excellent candidate for elucidating structural models.
ContributorsDeb, Arpan (Author) / Johnson, William (Author) / Kline, Alexander (Author) / Scott, Boston (Author) / Meador, Lydia (Author) / Srinivas, Dustin (Author) / Martin Garcia, Jose Manuel (Author) / Dorner, Katerina (Author) / Borges, Chad (Author) / Misra, Rajeev (Author) / Hogue, Brenda (Author) / Fromme, Petra (Author) / Mor, Tsafrir (Author) / ASU Biodesign Center Immunotherapy, Vaccines and Virotherapy (Contributor) / College of Liberal Arts and Sciences (Contributor) / School of Life Sciences (Contributor) / Biodesign Institute (Contributor) / School of Molecular Sciences (Contributor) / Applied Structural Discovery (Contributor) / Personalized Diagnostics (Contributor)
Created2017-02-22
Description
Plant viral vectors have previously been used to produce high expression levels of antibodies and other proteins of interest. By utilizing a transformed Agrobacterium with the vector containing the protein of interest for infiltration, viral vectors can easily reach the plant cells making it an effective form of transient protein expression. For this project two different plant viral vectors were compared; the geminiviral vector derived from Bean yellow dwarf virus (BeYDV) and the MagnICON vector system derived from Tobacco Mosaic Virus(TMV) and Potato Virus X(PVX). E16, an antibody against West Nile virus, has previously been expressed using both systems but expression levels between the systems were not directly compared. Agrobacterium tumefaciens EHA105 cells were transformed with both systems and expression levels of E16 were quantified using ELISAs. Results showed very low expression levels of E16 using the geminiviral vector indicating a need for further investigation into the clone used as previous studies reported much higher expression levels with the system.
ContributorsMurphy, Skylar (Author) / Chen, Qiang (Thesis director) / Jugler, Collin (Committee member) / College of Health Solutions (Contributor) / Barrett, The Honors College (Contributor)
Created2020-05
Description
Mitochondrial methionyl-tRNA-formyltransferase (MTFMT) is essential for mitochondrial protein translation. The MTFMT gene encodes for an enzyme of the same name, which acts to formylate the methionine of mitochondrial Met-tRNA(Met). In Homo sapiens, MTFMT-formylated-tRNA is an initiator and elongator for the synthesis of 13 mitochondrially-encoded proteins in complexes I, III and IV of the ETC. To understand this mechanism, it is necessary to perform a comprehensive analysis of energy metabolism and oxidative phosphorylation (OXPHOS) among impacted patients. Alterations to this gene vary, with the most documented as a single-splice-site mutation (c.626C>T). Here, we discuss MTFMT involvement in mitochondrial protein translation and neurodegenerative disorders, such as Leigh Syndrome and combined OXPHOS deficiency, in two families. We aim to delineate the impact of OXPHOS dysfunction in patients presenting with MTFMT mutation.
ContributorsChain, Kelsey (Author) / Chen, Qiang (Thesis director) / Rangasamy, Sampathkumar (Committee member) / Narayanan, Vinodh (Committee member) / School of Life Sciences (Contributor) / Barrett, The Honors College (Contributor)
Created2020-05
Description
“Tell It to the Frogs: Fukushima’s nuclear disaster and its impact on the Japanese Tree Frog” is a representation of the work from Giraudeau et. al’s “Carotenoid distribution in wild Japanese tree frogs (Hyla japonica) exposed to ionizing radiation in Fukushima.” This paper looked to see if carotenoid levels in the tree frog’s vocal sac, liver, and blood were affected by radiation from Fukushima’s power plant explosion. Without carotenoids, the pigment that gives the frogs their orange color on their necks, their courtship practices would be impacted and would not be as able to show off their fitness to potential mates. The artwork inspired by this research displayed the tree frog’s degradation over time due to radiation, starting with normal life and ending with their death and open on the table. The sculptures also pinpoint where the carotenoids were being measured with a brilliant orange glaze. Through ceramic hand building, the artist created larger than life frogs in hopes to elicit curiosity about them and their plight. While the paper did not conclude any changes in the frog’s physiology after 18 months of exposure, there are still questions that are left unanswered. Why did these frogs not have any reaction? Could there be any effects after more time has passed? Is radiation leakage as big of a problem as previously thought? The only way to get the answers to these questions is to be aware of these amphibians, the circumstances that led them to be involved, and continued research on them and radiation.
ContributorsWesterfield, Savannah (Author) / Beiner, Susan (Thesis director) / McGraw, Kevin (Committee member) / School of Life Sciences (Contributor) / School of Art (Contributor) / Barrett, The Honors College (Contributor)
Created2019-05
Description
Among the most ornate animal traits in nature are the angle-dependent (i.e. iridescent) structural colors of many birds, beetles, and butterflies. Though we now have a solid understanding of the mechanisms, function, and evolution of these features in several groups, less attention has been paid to the potential for angle-dependent reflectance in otherwise matte-appearing (i.e. not thought to be structurally colored) tissues. Here for the first time we describe non-iridescent angle-dependent coloration from the tail and wing feathers of several parrot species (Psittaciformes). We employed a novel approach \u2014 by calculating chromatic and achromatic contrasts (in just noticeable differences, JNDs) of straight and angled measurements of the same feather patch \u2014 to test for perceptually relevant angle-dependent changes in coloration on dorsal and ventral feather surfaces. We found, among the 15 parrot species studied, significant angle dependence for nearly all parameters (except chromatic JNDs on the ventral side of wing feathers). We then measured microstructural features on each side of feathers, including size and color of barbs and barbules, to attempt to predict interspecific variation in degree of angle-dependent reflectance. We found that hue, saturation, and brightness of feather barbs, barbule saturation, and barb:barbule coverage ratio were the strongest predictors of angle-dependent coloration. Interestingly, there was significant phylogenetic signal in only one of the seven angle-dependence models tested. These findings deepen our views on the importance of microscopic feather features in the production of directional animal coloration, especially in tissues that appear to be statically colored.
ContributorsReed, Steven Andrew (Co-author) / McGraw, Kevin (Thesis director) / Pratt, Stephen (Committee member) / Simpson, Richard (Committee member) / School of Mathematical and Statistical Sciences (Contributor) / School of Life Sciences (Contributor) / Barrett, The Honors College (Contributor)
Created2018-05
Description
A chimeric, humanized monoclonal antibody that recognizes a highly conserved fusion loop found on flaviviruses was constructed with a geminiviral replicon and transiently expressed in Nicotiana benthamiana plants through Agrobacterium tumefaciens infiltration. Characterization and expression studies were then conducted to confirm correct assembly of the antibody. Once the antibody was purified, an ELISA was conducted to validate that the antibody was able to bind to the flavivirus fusion loop.
ContributorsPardhe, Mary (Author) / Mason, Hugh (Thesis director) / Chen, Qiang (Committee member) / Mor, Tsafrir (Committee member) / School of Life Sciences (Contributor) / Department of Information Systems (Contributor) / W.P. Carey School of Business (Contributor) / Barrett, The Honors College (Contributor)
Created2018-05
Description
Historically, studies of condition-dependent signals in animals have been male-centric, but recent work suggests that female ornaments can also communicate individual quality (e.g., disease state, fecundity). There has been a surge of interest in how urbanization alters signaling traits, but we know little about if and how cities affect signal expression in female animals. We measured carotenoid-based plumage coloration and coccidian (Isospora spp) parasite burden in desert and city populations of house finches to examine urban impacts on male and female health and attractiveness. In earlier work, we showed that male house finches are less colorful and more parasitized in the city, and we again detected that pattern in this study for males. However, though city females are also less colorful than their rural counterparts, we found that rural females were more parasitized. Also, regardless of sex and unlike rural birds, more colorful birds in the city were more heavily infected with coccidia. These results show that urban environments can disrupt signal honesty in female animals and highlight the need for more studies on how cities affect disease and condition-dependent traits in both male and female animals.
ContributorsSykes, Brooke Emma (Author) / McGraw, Kevin (Thesis director) / Sweazea, Karen (Committee member) / Hutton, Pierce (Committee member) / School of Life Sciences (Contributor) / Barrett, The Honors College (Contributor)
Created2016-05
Description
Virus-Like Particles (VLPs) are self-assembling structures that lack the viral genetic material. Therefore they are safer and more immunogenic than other forms of vaccines. The Hepatitis B core (HBc) VLPs are a novel mechanism through which delivery of DNA-based human vaccines are plausible. Production of VLPs require recombinant, rapidly replicating, plant-based systems such as the geminiviral replicon system. This project entails the cloning process of HBc-DIII fusion protein, a VLP that should form Domain III of the Envelope protein on West Nile Virus, into deconstructed geminiviral vector. The cloning process includes the HBc-DIII fusion protein DNA isolation, restriction enzyme digestion with NcoI and SacI, PCR changing the NcoI site on the HBc-DIII insert to XbaI, sequencing, ligation into geminiviral vector and transformation into an agrobacterium strain. The major impediment to the cloning process was the presence of multiple bands instead of the expected two bands while doing restriction enzyme digests. The troubleshooting process enabled speculating that due to the excess of restriction enzymes in the digestion volume, some of the DNA was not digested completely. Hence, multiple bands were observed. However, sequencing analysis and further cloning process ensured the presence of HBc-DIII insert band (approximately 800bp) in the Gemini vector. Lastly, the construct HBc-DIII in Gemini vector was ensured to be in agrobacterium for further experiments such as agro-infiltration.
ContributorsSuresh Kumar, Reshma (Author) / Chen, Qiang (Thesis director) / Zhang, Peiming (Committee member) / School of Molecular Sciences (Contributor) / School of Life Sciences (Contributor) / Barrett, The Honors College (Contributor)
Created2016-05
Description
There are two electrophysiological states of sleep in birds (rapid-eye-movement sleep [REM] and slow-wave sleep [SWS]), which have different functions and costs. REM improves memory consolidation, while SWS is neuro-restorative but also exposes the animal to more risk during this deep-sleep phase. Birds who sleep in more exposed microsites are known to invest proportionally less in SWS (presumably to ensure proper vigilance), but otherwise little else is known about the ecological or behavioral predictors of how much time birds devote to REM v. SWS sleep. In this comparative analysis, we examine how proportional time spent in SWS v. REM is related to brain mass and duration of the incubation period in adults. Brain mass and incubation period were chosen as predictors of sleep state investment because brain mass is positively correlated with body size (and may show a relationship between physical development and sleep) and incubation period can be a link used to show similarities and differences between birds and mammals (using mammalian gestation period). We hypothesized that (1) species with larger brains (relative to body size and also while controlling for phylogeny) would have higher demands for information processing, and possibly proportionally outweigh neuro-repair, and thus devote more time to REM and that (2) species with longer incubation periods would have proportionally more REM due to the extended time required for overnight predator vigilance (and not falling into deep sleep) while on the nest. We found, using neurophysiological data from literature on 27 bird species, that adults from species with longer incubation periods spent proportionally more time in REM sleep, but that relative brain size was not significantly associated with relative time spent in REM or SWS. We therefore provide evidence that mammalian and avian REM in response to incubation/gestation period have convergently evolved. Our results suggest that overnight environmental conditions (e.g. sleep site exposure) might have a greater effect on sleep parameters than gross morphological attributes.
ContributorsRaiffe, Joshua Sapell (Author) / McGraw, Kevin (Thesis director) / Deviche, Pierre (Committee member) / Hutton, Pierce (Committee member) / School of Life Sciences (Contributor) / Barrett, The Honors College (Contributor)
Created2017-05