Cell and Developmental Biology Commons^™

Hiv-1 Effects On Neuropsychological Performance In A Resource-Limited Country, Zambia, Gabriel Holguin, Mwanza Banda, Elizabeth J. Willen, Costantine Malama, Kaseya O. Chiyenu, Victor C. Mudenda, Charles Wood

Nebraska Center for Virology: Faculty Publications

Zambia has substantially been affected by the HIV/AIDS epidemic with prevalence rates at 14% in a population estimated at 12 million. Yet, the extent of HIV-associated neurocognitive disorders (HAND) in this population remains to be clearly understood. A series of culturally appropriate neuropsychological (NP) assessments [International HIV Dementia Scale (IHDS), Color Trails Test 1 and 2, Grooved pegboard Test, and Time Gait Test] were used to test the effects of HIV on NP performance of HIV seropositive and seronegative individuals. Twenty-two percent HIV positive individuals ARV naïve met the criteria for IHDS-defined NP impairment. Gender significantly influenced the performance on …

Characterization Of Species C Human Adenovirus Serotype 6 (Ad6), Eric A. Weaver, Reeti Khare, Mathew L. Hillestad, Donna Palmer, Philip Ng, Michael A. Barry

Nebraska Center for Virology: Faculty Publications

Adenovirus serotype (Ad5) is the most studied Ad. Ad1, 2, and 6 are also members of species C Ad and are presumed to have biologies similar to Ad5. In this work, we have compared the ability of Ad1, 2, 5, and 6 to infect liver and muscle after intravenous and intramuscular injection. We found that Ad6 was surprisingly the most potent at liver gene delivery and that Ad1 and Ad2 were markedly weaker than Ad5 and 6. To understand these differences, we sequenced the Ad6 genome. This revealed that the Ad6 fiber protein is surprisingly three shaft repeats shorter than …

Generation Of A Kupffer Cell-Evading Adenovirus For Systemic And Liver-Directed Gene Transfer, Reeti Khare, Shannon M. May, Francesco Vetrini, Eric A. Weaver, Donna Palmer, Amanda Rosewell, Nathan Grove, Philip Ng, Michael A. Barry

Nebraska Center for Virology: Faculty Publications

As much as 90% of an intravenously (i.v.) injected dose of adenovirus serotype 5 (Ad5) is absorbed and destroyed by liver Kupffer cells. Viruses that escape these cells can then transduce hepatocytes after binding factor X (FX). Given that interactions with FX and Kupffer cells are thought to occur on the Ad5 hexon protein, we replaced its exposed hypervariable regions (HVR) with those from Ad6. When tested in vivo in BALB/c mice and in hamsters, the Ad5/6 chimera mediated >10 times higher transduction in the liver. This effect was not due to changes in FX binding. Rather, Ad5/6 appeared to …

Dose-Dependent Thresholds Of 10-Ns Electric Pulse Induced Plasma Membrane Disruption And Cytotoxicity In Multiple Cell Lines, Bennett L. Ibey, Caleb C. Roth, Andrei G. Pakhomov, Joshua A. Bernhard, Gerald J. Wilmink, Olga N. Pakhomova

Bioelectrics Publications

In this study, we determined the LD50 (50% lethal dose) for cell death, and the ED50 (50% of cell population staining positive) for propidium (Pr) iodide uptake, and phosphatidylserine (PS) externalization for several commonly studied cell lines (HeLa, Jurkat, U937, CHO-K1, and GH3) exposed to 10-ns electric pulses (EP). We found that the LD50 varied substantially across the cell lines studied, increasing from 51 J/g for Jurkat to 1861 J/g for HeLa. PS externalized at doses equal or lower than that required for death in all cell lines ranging from 51 J/g in Jurkat, to 199 J/g in CHO-K1. Pr …

Nanosecond Pulsed Electric Field Induced Cytoskeleton, Nuclear Membrane And Telomere Damage Adversely Impact Cell Survival, Michael W. Stacey, P. Fox, S. Buescher, Juergen F. Kolb

Bioelectrics Publications

We investigated the effects of nanosecond pulsed electric fields (nsPEF) on three human cell lines and demonstrated cell shrinkage, breakdown of the cytoskeleton, nuclear membrane and chromosomal telomere damage. There was a differential response between cell types coinciding with cell survival. Jurkat cells showed cytoskeleton, nuclear membrane and telomere damage that severely impacted cell survival compared to two adherent cell lines. Interestingly, disruption of the actin cytoskeleton in adherent cells prior to nsPEF exposure significantly reduced cell survival. We conclude that nsPEF applications are able to induce damage to the cytoskeleton and nuclear membrane. Telomere sequences, regions that tether and …

An Apoptosis Targeted Stimulus With Nanosecond Pulsed Electric Fields (Nspefs) In E4 Squamous Cell Carcinoma, Wei Ren, Stephen J. Beebe

Bioelectrics Publications

Stimuli directed towards activation of apoptosis mechanisms are an attractive approach to eliminate evasion of apoptosis, a ubiquitous cancer hallmark. In these in vitro studies, kinetics and electric field thresholds for several apoptosis characteristics are defined in E4 squamous carcinoma cells (SCC) exposed to ten 300 ns pulses with increasing electric fields. Cell death was [95% at the highest electric field and coincident with phosphatidylserine externalization, caspase and calpain activation in the presence and absence of cytochrome c release, decreases in Bid and mitochondria membrane potential (Δψm) without apparent changes reactive oxygen species levels or in Bcl2 and Bclxl levels. …

Roles Of The 15-Kda Selenoprotein (Sep15) In Redox Homeostasis And Cataract Development Revealed By The Analysis Of Sep 15 Knockout Mice, Marina V. Kasaikina, Dmitri E. Fomenko, Vyacheslav M. Labunskyy, Salil A. Lachke, Wenya Qiu, Juliet A. Moncaster, Jie Zhang, Mark W. Wojnarowicz, Sathish Kumar Natarajan, Mikalai I. Malinouski, Ulrich Schweizer, Petra A. Tsuji, Bradley A. Carlson, Richard L. Maas, Marjorie F. Lou, Lee E. Goldstein, Dolph L. Hatfield, Vadim N. Gladyshev

School of Veterinary and Biomedical Sciences: Faculty Publications

The 15-kDa selenoprotein (Sep15) is a thioredoxin-like, endoplasmic reticulum-resident protein involved in the quality control of glycoprotein folding through its interaction with UDP-glucose:glycoprotein glucosyltransferase. Expression of Sep15 is regulated by dietary selenium and the unfolded protein response, but its specific function is not known. In this study, we developed and characterized Sep15 KO mice by targeted removal of exon 2 of the Sep15 gene coding for the cysteinerich UDP-glucose:glycoprotein glucosyltransferase-binding domain. These KO mice synthesized a mutant mRNA, but the shortened protein product could be detected neither in tissues nor in Sep15 KO embryonic fibroblasts. Sep15 KO mice were viable …

The Impact Of Tricaine Methanesulfonate, 2-Phenoxyethanol, And Carvone-Methyl Salicylate On The Innate Immune Response Of Zebrafish (Danio Rerio), Charles R. Wulff

Honors Theses

Anesthesia plays a vital role in the maintenance of aquaculture species, where it is used to minimize stress during complex handling tasks such as transport, assessment, and harvesting. However, anesthetics have been shown to suppress the innate immune response, which could impact immunity and increase risk of infection. Tricaine methanesulfonate (MS-222) and 2-Phenoxyethanol (2-PE) represent two of the most commonly used anesthetics in aquaculture, with R-(+)-carvone, in the form of carvone-methyl salicylate (CMS) has recently been proposed as an alternative anesthetic for food fish. These three anesthetics were used to assess the influence of anesthetics on the immune system of …

Rho Kinase Regulates Basement Membrane Dynamics To Coordinate Salivary Gland Branching Morphogenesis And Tissue Organization, William Patrick Daley

Legacy Theses & Dissertations (2009 - 2024)

Branching morphogenesis is a developmental mechanism utilized by many organs, including the salivary gland, lung, kidney, and mammary gland, to increase the epithelial surface area for secretion or absorption. The embryonic submandibular salivary gland (SMG) is a classic model for studying tissue morphogenesis in three dimensions ex vivo. Salivary gland development requires distinct but overlapping processes: morphogenesis, the actual physical rearrangement of cells into complex three dimensional structures, and cytodifferentiation, the process by which these cells begin to take on their own specialized function. Polarization, or the organization of cells into a cohesive tissue structure, also occurs concurrently during organ …

A Breast Cancer Stem Cell Model Created From Mmtv-Pymt Mice Applicable To Human Breast Cancer, Denise Grant Lanza

Legacy Theses & Dissertations (2009 - 2024)

Cancer stem cells are the seeds of tumor growth, but there are limited cell-based methods that exist to study the properties of these cells. To create a model of breast cancer stem cells, we isolated tumors from MMTV-PyMT mice. Two out of the four different cell types isolated survived in culture, CD44+CD24- and CD24+CD49f+CD44low. We found that we could initiate tumors with as few as 10 cells injected subcutaneously in the hind leg or orthotopically in the cleared fat pad with CD24+ cells. However, we could not initiate tumors with injection of CD24- cells. We found a requirement for TICs …

The Role Of Trm9 In Stress Responses, Ashish Ravindra Patil

Legacy Theses & Dissertations (2009 - 2024)

Cells need to respond appropriately to environmental changes in order to maintain homeostasis. The cellular response to an environmental stress is regulated at transcriptional, translational and post translational levels. The tRNA, which acts as an adaptor molecule between the mRNA and the protein, plays an important role in the translational regulation of cellular responses to stress and is one of the most heavily modified biomolecules. In Saccharomyces cerevisiae , the wobble uracil of the tRNA(3'-UCU-5') Arg, tRNA(3'-UUC-5') Glu and certain other specific tRNAs are modified to 5-methoxycarbonylmethyluridine (mcm5U) and 5-methoxycarbonylmethyl-2-thiouridine (mcm5s2U) residues by the tRNA methyltransferase 9 (Trm9). Modifications at …

Regulation Of The Aryl Hydrocarbon Receptor And The Signal Transducer And Activator Of Transcription 1 By Long-Term Estrogen Exposure In Breast Carcinoma Cells, Neal Englert

Legacy Theses & Dissertations (2009 - 2024)

The risk of developing breast cancer is known to be associated with a woman's lifetime exposure to estrogens, both endogenous and exogenous. Increased exposure to estrogens stimulates cellular proliferation, which is a widely accepted theory of estrogen receptor positive mammary carcinogenesis. The molecular mechanisms of gene expression regulation in response to long-term estrogen exposure (LTEE) of MCF-7 breast cancer cells were addressed in this study, with a focus on the aryl hydrocarbon receptor (AHR) and signal transducer and activator of transcription 1 (STAT1).

The P38 Alpha Stress-Activated Protein Kinase Pathway In Mammary Gland Development And Cancer, Huei-Chi Wen

Legacy Theses & Dissertations (2009 - 2024)

Breast tumors arise from preinvasive lesions consisting of abnormal epithelial cells that accumulate within the mammary duct. However, the initiating steps and underlying pathways leading to the formation of these early stages of breast carcinoma, such as ductal carcinoma in situ (DCIS), is not fully understood.

Expression And Regulation Of Survivin In Prostate Cancer, Maygen K. Cardona

McNair Scholars Research Journal

Survivin is an Inhibitor of Apoptosis Protein (IAP), which makes it responsible for cell proliferation and survival by inhibiting regulated cell death and promoting mitosis. The exact role that it plays in prostate cancer is not yet fully understood; expression and regulation of Survivin is the primary focus of my project. The way Survivin was studied was through fractionating various cancer cells lines (LaPC-4, PC-3 E+, VCAP, C4-2 and DU145-Spindle, Round and Tight) and isolating the nuclear and cytosolic components. These fractionated samples were then treated and analyzed using Western Blot analysis being probed for nuclear and cytoplasmic Survivin, respectively. …

Banf1 Is Required To Maintain The Self-Renewal Of Both Mouse And Human Embryonic Stem Cells, Jesse L. Cox, Sunil K. Mallanna, Briana D. Ormsbee, Michelle Desler, Matthew S. Wiebe, Angie Rizzino

Nebraska Center for Virology: Faculty Publications

Self-renewal is a complex biological process necessary for maintaining the pluripotency of embryonic stem cells (ESCs). Recent studies have used global proteomic techniques to identify proteins that associate with the master regulators Oct4, Nanog, and Sox2 in ESCs or in ESCs during the early stages of differentiation. Through an unbiased proteomic screen, Banf1 was identified as a Sox2- associated protein. Banf1 has been shown to be essential for worm and fly development but, until now, its role in mammalian development and ESCs has not been explored. In this study, we examined the effect of knocking down Banf1 on ESCs. We …

An Upper Limit For Macromolecular Crowding Effects, Andrew C. Miklos, Congang Li, Courtney D. Sorell, L. Andrew Lyon, Gary J. Pielak

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Background: Solutions containing high macromolecule concentrations are predicted to affect a number of protein properties compared to those properties in dilute solution. In cells, these macromolecular crowders have a large range of sizes and can occupy 30% or more of the available volume. We chose to study the stability and ps-ns internal dynamics of a globular protein whose radius is similar to 2 nm when crowded by a synthetic microgel composed of poly(N-isopropylacrylamide-co-acrylic acid) with particle radii of similar to 300 nm.

Results: Our studies revealed no change in protein rotational or ps-ns backbone dynamics and only mild …

Examination Of The Mitochondrial Health Of The Basigin Null Mouse Retina, Kristine Anne V. Pablo

UNF Graduate Theses and Dissertations

Basigin gene products are cell adhesion molecules that are expressed by photoreceptor cells, Müller cells and endothelial cells of the mammalian retina. Previous studies have suggested that a lactate shuttle exists between the photoreceptor cells and the Müller cells, with Basigin being an essential component in this shuttle. Deletion of the Basigin gene in mice results in blindness with an eventual retinal degeneration. It was hypothesized that the lactate shuttle between photoreceptors and Müller cells does not form in Basigin null mice and that the blindness is attributed to faulty photoreceptor metabolism. Therefore, the purpose of this study was to …

Escherichia Coli Fpg Glycosylase Is Nonrendundant And Required For The Rapid Global Repair Of Oxidized Purine And Pyrimidine Damage In Vivo, Brandy J. Schalow, Charmain T. Courcelle, Justin Courcelle

Biology Faculty Publications and Presentations

Endonuclease (Endo) III and formamidopyrimidine-N-glycosylase (Fpg) are two of the predominant DNA glycosylases in Escherichia coli that remove oxidative base damage. In cell extracts and purified form, Endo III is generally more active toward oxidized pyrimidines, while Fpg is more active towards oxidized purines. However, the substrate specificities of these enzymes partially overlap in vitro. Less is known about the relative contribution of these enzymes in restoring the genomic template following oxidative damage. In this study, we examined how efficiently Endo III and Fpg repair their oxidative substrates in vivo following treatment with hydrogen peroxide. We found …

Role Of Vascular Endothelial Growth Factor, Placental Growth Factor And Cytokines On Outer Blood Retinal Barrier Breakdown., Sangeetha Sukumaran

Masters

The aim of this thesis was to investigate the effect of the VEGF family of growth factors and
cytokines TNF- α and IL-1β on oBRB function. The human RPE line ARPE-19 was
employed as an in vitro model of the oBRB. Two-Dimensional gel electrophoresis was
employed to study the effect of growth factors on the RPE proteome. The barrier properties of
the oBRB were evaluated by expression of the TJ proteins occludin or Zo-1, or claudins-1, 2,
3 and 4, determined by western blotting analysis. Claudins 2, 3 and 4 were found to be
significantly up-regulated by TNF-α, while interleukin …

Modeling Measurement Error In Tumor Characterization Studies, Cyril Rakovski, Daniel J. Weisenberger, Paul Marjoram, Peter W. Laird, Kimberly D. Siegmund

Mathematics, Physics, and Computer Science Faculty Articles and Research

Background: Etiologic studies of cancer increasingly use molecular features such as gene expression, DNA methylation and sequence mutation to subclassify the cancer type. In large population-based studies, the tumor tissues available for study are archival specimens that provide variable amounts of amplifiable DNA for molecular analysis. As molecular features measured from small amounts of tumor DNA are inherently noisy, we propose a novel approach to improve statistical efficiency when comparing groups of samples. We illustrate the phenomenon using the MethyLight technology, applying our proposed analysis to compare MLH1 DNA methylation levels in males and females studied in the Colon …

Nanosecond Pulse Electrical Fields Used In Conjunction With Multi-Wall Carbon Nanotubes As A Potential Tumor Treatment, Michael W. Stacey, Christopher Osgood, Bhargava Subhash Kalluri, Wei Cao, Hani Elsayed-Ali, Tarek Abdel-Fattah

Bioelectrics Publications

The objectives of this communication were to fabricate pure samples of multi-walled carbon nanotubes (MWCNTs) and to determine their toxicity in tumor cell lines. MWCNTs were dispersed in a concentration of the surfactant T80 that was minimally toxic. Cell-type variation in toxicity to MWCNTs was observed but was not significantly different to unexposed controls. Additionally, we investigated the increased cell killing of the pancreatic cancer cell line PANC1 when exposed to ultrashort (nanosecond) pulsed electrical fields (nsPEF) in the presence of MWCNTs as a potential form of cancer therapy. We hypothesized that the unique electronic properties of MWCNTs disrupt cell …

Comparative Study Of Long-And Short-Pulsed Electric Fields For Treating Melanoma In An In Vivo Mouse Model, Xinhua Chen, Xinmei Chen, Karl H. Schoenbach, Shusen Zheng, R. James Swanson

Bioelectrics Publications

A mouse melanoma model was set up with green fluorescent protein (GFP) expression in vivo. With the same energy, long- (1 ms) and short- (300 ns) pulsed electric fields were delivered to two melanomas injected into the same mouse. The tumor growth and green fluorescence were followed up to compare the different treatment efficacy of long and short pulses. After two days post treatment, short pulse-treated tumors showed a significantly lower tumor volume compared with long pulse-treated tumors (n=8, p

Identification And Characterization Of Fkbp52-Specific Inhibitors For The Treatment Of Prostate Cancer, Johanny Tonos De Leon

Open Access Theses & Dissertations

Steroid hormone receptors require the ordered assembly of various chaperone and cochaperone proteins in order to reach a functional state. The final stage in the receptor maturation process requires the formation of a mutimeric complex consisting of Hsp90 dimer, p23, and one of several large immunophilins. Studies conducted previously demonstrated that the large immunophilin FKBP52 acts to potentiate glucocorticoid, androgen, and progesterone receptor signaling pathways. The aim of these studies was to identify and characterize FKBP52-specific inhibitors that would not only serve as tools for the pharmacological analysis of FKBP52-receptor interactions, but may also lead to novel drugs with significant …

The Characterization Of Genes Involved In Response To The Phenol Derivative And Xenoestrogen Bisphenol-A In Saccharomyces Cerevisiae, Sasha N. Farina

CMC Senior Theses

Bisphenol A is an estrogenic compound that is found in polycarbonate plastics and epoxy resins; humans are continuously exposed to the compound and it is believed to possess the same carcinogenic effects as estrogen (Iso, 2006). In this study, I used Saccharomyces cerevisiae as a model organism to identify mechanisms by which BPA acts based on the genomic profiling of kinase genes from a Mat-α haploid deletion library. Kinases regulate many other proteins, so the identification of a single mutant could identify an entire affected pathway of genes. I conducted a systematic screen of these mutants using the phenotype of …

Cooperative And Antagonistic Roles For Heterochromatin Proteins In Transcriptional Regulation Of The Drosophila Sex Determination Masterswitch Gene, Hui Li

University of Kentucky Doctoral Dissertations

HOAP was originally identified as a component of an ORC-containing multi-protein complex of Heterochromatin Protein 1 (HP1) from early Drosophila embryos. HOAP immunostaining showed prominent association of it with telomeres, and mutants for HOAP (cav¹) showed it functions along with HP1 in forming a telomere capping complex that prevents telomeric fusions.

Weaker HOAP immunostaining is also observed in regions of pericentric heterochromatin and euchromatin. To examine the role of HOAP at these non-telomeric sites, we applied Affymetric Drosophila Genome Arrays to undertake a microarray expression profiling study of genes that are mis-expressed in cav¹ mutant larvae. The …

Bcc Skin Cancer Diagnosis Based On Texture Analysis Techniques, Shao-Hui Chuang, Xiaoyan Sun, Wen-Yu Chang, Gwo-Shing Chen, Adam Huang, Jiang Li, Frederic D. Mckenzie

Electrical & Computer Engineering Faculty Publications

In this paper, we present a texture analysis based method for diagnosing the Basal Cell Carcinoma (BCC) skin cancer using optical images taken from the suspicious skin regions. We first extracted the Run Length Matrix and Haralick texture features from the images and used a feature selection algorithm to identify the most effective feature set for the diagnosis. We then utilized a Multi-Layer Perceptron (MLP) classifier to classify the images to BCC or normal cases. Experiments showed that detecting BCC cancer based on optical images is feasible. The best sensitivity and specificity we achieved on our data set were 94% …

Variation, Systematics, And Relationships Of The Leptodactylus Bolivianus Complex (Amphibia: Anura: Leptodactylidae), W. Ronald Heyer, Rafael O. De Sá

Biology Faculty Publications

Heyer, W. Ronald, and Rafael O. de Sá. Variation, Systematics, and Relationships of the Leptodactylus bolivianus Complex (Amphibia: Anura: Leptodactylidae). Smithsonian Contributions to Zoology, number 635, viii + 58 pages, 21 figures, 20 tables, 2011.—The Leptodactylus bolivianuscomplex has been considered to consist of one or two species, L. bolivianus alone or L. bolivianus and L. insularum. Detailed morphological analyses were undertaken to evaluate variation in the complex, which ranges from Costa Rica through Panama, across northern South America in the river valleys draining to the Caribbean, and throughout much of the Amazon basin with southern limits in Bolivia. Members of …

Loss Of Bloom Syndrome Protein Causes Destabilization Of Genomic Architecture And Is Complemented By Ectopic Expression Of Escherichia Coli Recg In Human Cells, Michael Wayne Killen

University of Kentucky Doctoral Dissertations

Genomic instability driven by non-allelic homologous recombination (NAHR) provides a realistic mechanism that could account for the numerous chromosomal abnormalities that are hallmarks of cancer. We recently demonstrated that this type of instability could be assayed by analyzing the copy number variation of the human ribosomal RNA gene clusters (rDNA). Further, we found that gene cluster instability (GCI) was present in greater than 50% of the human cancer samples that were tested. Here, data is presented that confirms this phenomenon in the human GAGE gene cluster of those cancer patients. This adds credence to the hypothesis that NAHR could be …

Cardiac Calsequestrin Phosphorylation And Trafficking In The Mammalian Cardiomyocyte, Timothy Mcfarland

Wayne State University Dissertations

Cardiac CSQ (CSQ2) is a multifaceted protein, capable of binding significant quantities of Ca2+ and altering ryanodine receptor activity at the junctional sarcoplasmic reticulum (SR). Little is known about the trafficking of CSQ2 from its unknown site of biosynthesis, which appears to be of importance as its structure changes in a trafficking-dependent manner in various types of heart failure. Through the use of multiple antibodies specific to classic rough ER markers, and with the creation of CSQ-DsRed tetramer fusion protein, we were able to establish a juxtanuclear localization of rough ER in cardiomyocytes. Using fluorescence confocal microscopy, the translocon complex …

Amphiregulin (Areg) And Epidermal Growth Factor (Egf): Disparate In Egfr Signaling And Trafficking, Andrea Jacqueline Baillo

Wayne State University Dissertations

We have previously shown that SUM-149 human breast cancer cells require an AREG/EGFR autocrine loop for cell proliferation. We also demonstrated that AREG can increase EGFR stability and promote EGFR localization to the plasma membrane. In the presented dissertation we successfully knocked-down AREG expression in SUM-149 cells by lenti-viral infection of AREG shRNA. In the absence of AREG expression, SUM-149 cell growth was slowed, but not completely inhibited. Furthermore, cells infected with AREG shRNA constructs showed an increase in EGFR protein expression by western blot. Immunofluorescence and confocal microscopy showed that following AREG knock-down, EGFR continued to localize to the …