Cell and Developmental Biology Commons^™

The Effect Of Acp1-Ada1 Genetic Interaction On Human Life Span, Nazzareno Lucarini, Valerio Napolioni, Andrea Magrini, Fulvia Gloria

Human Biology Open Access Pre-Prints

Acid phosphatase (ACP1) is a polymorphic enzyme which catalyzes the conversion of flavinmononucleotide (FMN) to riboflavin and regulates the cellular concentration of flavin-adeninedinucleotide (FAD) and, consequently, energy metabolism. Its activity is modulated by adenosine deaminase (ADA1) genotype. Aim of our work is to verify whether individuals with a high proportion of ACP1 f isozyme and carrying ADA*2 allele, displaying the highest phosphatase activity, may have a higher life expectancy. Genomic DNA was extracted from peripheral blood of 569 females and 509 males (18-106 years) randomly recruited from Central Italy. These samples were subdivided into three sexspecific age groups …

The Methuselah Family Of G Protein Coupled Receptors, Meghna V. Patel

Wayne State University Dissertations

G protein-coupled receptors (GPCRs) are one of the largest class of transmembrane signaling proteins that regulate essential developmental and physiological processes in a cell. GPCR success is illustrated by their abundance across both invertebrate and vertebrate genomes. Phylogenetic analyses show that GPCR families have undergone a lot of gene gain and loss during insect evolution. In Drosophila melanogaster, the fifteen Methuselah/Methuselah-like (Mth/Mthl) genes are in fact an insect specific family of GPCRs. In our study, we conducted a phylogenetic analysis using receptor sequences from five Drosophila species and two related insects, including Tribolium and Anopheles to examine the evolution of …

The Role Of Cad,Flash And Fam129b In Cancer Cell Survial And Apoptosis, Song Chen

Wayne State University Dissertations

Apoptosis is a normal process in the human body. However, apoptosis is desregulated in cancer cells. Most cancer cells gain resistance to apoptosis, leading to uncontrolled proliferation. In this dissertation, we identified three proteins, associated with apoptosis pathway. 1) CAD, a large multifunctional complex that is invariably elevated in tumor cells, 2) FLASH, a large protein with multiple growth related functions and 3) FAM129B. We demonstrate that CAD could interact with FLASH by using yeast two hybrid, co-immunopreciptation and fluorescence microscopy. In addition, functional analysis using siRNA technology further indicated that CAD could co-operate with FLASH and play roles in …

A Role For Reactive Oxygen Species In Photodynamic Therapy, Michael Price

Wayne State University Dissertations

Photodynamic therapy (PDT) is based on the ability of certain photosensitizing agents to selectively localize in neoplastic cells and their vasculature. Subsequent irradiation at a wavelength corresponding to a photosensitizer absorbance band excites the photosensitizer molecules, leading to energy transfer reactions and fluorescence. It was initially concluded that the phototoxic effect occurred when energy from the excited state of the photosensitizer was transferred to dissolved oxygen to form singlet oxygen. This product has a very brief half-life and will cause cellular damage only in the immediate vicinity of its formation. But an excited-state photosensitizer can also interact with oxygen to …

Molecular Mechanisms Of Strain-Stimulated Intestinal Epithelial Cell Differentiation And Migration, Lisi Yuan

Wayne State University Dissertations

The intestinal epithelium is subjected to repetitive deformation during normal gut function by peristalsis and villous motility. In vitro, cyclic strain promotes intestinal epithelial proliferation and induces an absorptive phenotype characterized by increased dipeptidyl dipeptidase (DPPIV) expression. Schlafen 3 is a novel gene recently associated with cellular differentiation. We sought to evaluate whether Schlafen 3 mediates the effects of strain on the differentiation of IEC-6 intestinal epithelial cells in the absence or presence of cyclic strain. Strain increased Schlafen 3 mRNA and protein. In cells transfected with a control non-targeting siRNA, strain increased DPPIV specific activity. However, Schlafen

3 reduction …

Hyperosmotic Stress Enzyme Signaling Modulates Oct4, Nanog, And Rex1 Expression And Induces Prioritized Differentiation Of Murine Embryonic Stem Cells, Jill A. Slater

Wayne State University Dissertations

HYPEROSMOTIC STRESS ENZYME SIGNALING MODULATES OCT4, NANOG,

AND REX1 EXPRESSION AND INDUCES PRIORITIZED DIFFERENTIATION OF

MURINE EMBRYONIC STEM CELLS

by

JILL SLATER

MAY 2013

Advisor: Daniel Rappolee, Ph.D.

Major: Physiology

Degree: Doctor of Philosophy

Transcription factor expression and therefore lineage identity in the periimplantation

embryo and its stem cells may be influenced by extracellular stresses,

potentially affecting pregnancy outcome. Cellular stress forces cells to suppress some

normal activities (such as protein synthesis and cell proliferation) in order to repair

stress-damaged macromolecules and restore homeostasis. Therefore, any new

activities that embryonic cells initiate while concurrently funding the demands of the

stress …

Pax6 Is Required For The Development Of The Lateral Procephalon In Tribolium, Qing Luan

Wayne State University Theses

In Tribolium, combinatorial knockdown of the Pax6 orthologs eyeless (ey) and twin of eyeless (toy) affects the peripheral visual system but also other areas of the dorsal larval head capsule. To elucidate the role of Pax6 genes during Tribolium embryonic head development in detail, we performed an extensive analysis of cuticle

elements, brain anatomy, embryonic head morphogenesis and developmental marker gene expression. Our results reveal that Pax6 is required for the development of a large contiguous area of the lateral anterior head, morphologically addressed as the embryonic head lobes, which encompass the neuroectodermal precursor tissues of the visual system, parts …

Cellular Plasticity In White Adipose Tissue: In Vivo Identification Of Bipotent Adipocyte Progenitors In Adult White Adipose Tissue, Yun-Hee Lee

Wayne State University Dissertations

Nutritional and pharmacological stimuli can dramatically alter the cellular composition and phenotype of white adipose tissue (WAT). Nonetheless, the identity of progenitors that contribute to this cellular plasticity in vivo remains poorly understood. Utilizing genetic lineage tracing techniques in combination with in situ imunohistochemical analysis, we demonstrate that brown adipocytes (BA) that are induced by beta3-adrenergic receptor activation (ADRB3) in WAT arise from the proliferation and differentiation of cells that express platelet-derived growth factor receptor alpha (PDGFRA), CD34 and Sca1 (PDGFRA+ cells). PDGFRA+ cells have a unique morphology in which extended processes contact multiple cells in the tissue microenvironment. Surprisingly, …

Identifying Sm22 As A Key Player In Arterial Diseases, Jianbin Shen

Wayne State University Dissertations

Background : Expression of vascular smooth muscle cell (VSMC) cytoskeleton markers including SM22 is down-regulated in arterial diseases including atherosclerosis where inflammation and osteochondrogenesis are present. However, the role of this downregulation in arterial pathogenesis is unknown. Hypothesis : Downregulation of SM22 may actively contribute to arterial pathogenesis. Methods : Five Sm22 knockout (Sm22^-/-) mice and their wild type littermates were subjected to carotid artery denudation, an artery injury model. Analyses were conducted on carotid arteries 2 weeks after injury. Primary VSMCs were isolated from mouse aortas and investigated individually at passage 2 to 4. Sm22 knockdown was …

Identification Of Cellular Functions Of Cardiolipin As Physiological Modifiers Of Barth Syndrome, Amit Shridhar Joshi

Wayne State University Dissertations

Cardiolipin (CL) is an anionic phospholipid synthesized in the mitochondrial inner membrane. Perturbation of CL metabolism leads to Barth syndrome (BTHS), a life threatening genetic disorder. I utilized genetic, biochemical and cell biological approaches in yeast to elucidate the cellular functions of CL. Understanding the functions of CL is expected to shed light on the pathology and possible treatments for BTHS.

BTHS is caused by mutations in TAZ1, which encodes a CL remodeling enzyme called tafazzin. BTHS patients exhibit a wide range of clinical presentations, indicating that physiological modifiers influence the BTHS phenotype. A targeted synthetic lethality screen was performed …

Biophysical And Biochemical Factors In The Cellular Microenvironment; Effects On Cell Migration And Invasion, Shalini Menon

Wayne State University Dissertations

Cellular migration is a vital process central to many physiological events including development, immune surveillance and wound healing. However, migration and invasion are not unique to normal physiology, they are also key determinants in the progression of disease states such as cancer. Given the significance of migration it is important that we understand how the process is regulated intracellularly and the various stimuli that can promote it. Even though the role of biochemical factors in mediating migration has been studied extensively, the role of biophysical factors in modulating migration and invasion is less appreciation. The biochemical and biophysical components of …

Hdm2 Small-Molecule Inhibitors For Therapeutic Intervention In B-Cell Lymphoma, Angela Sosin

Wayne State University Dissertations

Lymphomas frequently retain wild-type (wt) p53 function but overexpress HDM2, compromising p53 activity. Therefore, lymphoma is a suitable model for studying therapeutic value of disrupting HDM2-p53 association by small-molecule inhibitors (SMIs). HDM2 SMIs have been developed and are currently under various stages of preclinical and clinical investigation. This study examined various molecular mechanisms associated and biological effects of two different classes of HDM2 SMIs: the spiro-oxindoles (MI-219) and cis-imidazoline (Nutlin-3) in lymphoma cell lines and patient-derived B-lymphoma cells. Surprisingly, results revealed significant quantitative and qualitative differences between these two agents. At the molecular level, effect of Nutlin-3 was generally more …

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 …

Axogial Communication Mediated By Soluble Neuregulin-1 And Bdnf, Zhenzhong Ma

Wayne State University Dissertations

During peripheral nervous system development, successful communication between axons and glial cells including Schwann cells in peripheral nervous system and oligodendrocytes in central nervous system, is required for the proper functions of both neurons and glia. Three types of alternatively-spliced proteins belonging to the neuregulin1 (NRG1) gene family of growth and differentiation factors are essential for Schwann cell survival and peripheral nerve development. While membrane-bound NRG1 forms (type III) has been strongly implicated in the regulation of myelination process at late stage of Schwann cell development, little is known about the role of soluble, heparin-binding forms of NRG1 (type I/II) …

The Process Of Autophagy In An In Vitro Model Of Myocardial Ischemia-Reperfusion Injury, Kadija Abounit

Wayne State University Dissertations

Autophagy has been implicated in the pathogenesis of myocardial ischemia-reperfusion (I/R) injury; however, it is not clear whether autophagy is beneficial or detrimental to cell survival. We hypothesized that autophagy is beneficial to cardiomyocytes during I/R and that the paucity of ATP may limit its occurrence. To test this hypothesis, we developed a model of ischemia (hypoxia) involving exposure of HL-1 murine atrial cardiomyocytes to two concentrations of oxygen (0.5% and 2.0%) and 2-deoxyglucose.

Hypoxia, irrespective of the oxygen concentration, caused a dramatic drop in cellular ATP and an increase in the phosphorylation of AMPK (an indication of energy starvation),. …

Mechanical Forces And Tumor Cells: Insight Into The Biophysical Aspects Of Cancer Progression, Indrajyoti Indra

Wayne State University Dissertations

Mechanical forces play an important role in the regulation of cellular behavior and physiological processes including adhesion, migration, proliferation, tissue repair, embryogenesis and development. In addition, a number of diseases including cancer, have been linked to changes in cellular and extracellular mechanical properties. However, whether a correlation exists between the progression of cancer towards metastasis and mechanical factors has not been clearly defined. Additionally, how a cell responds to changes in extracellular mechanical cues as it gains metastatic abilities is poorly understood. To address these questions, we have utilized a panel of murine breast cancer cell lines with progressive metastatic. …

Functional In Vitro Analyses Of Lipid Raft-Associated Cathepsin B: Implication For The Invasive Phenotype Of Inflammatory Breast Cancer, Bernadette Caroline Victor

Wayne State University Dissertations

FUNCTIONAL IN VITRO ANALYSES OF LIPID RAFT-ASSOCIATED CATHEPSIN B: IMPLICATION FOR THE INVASIVE PHENOTYPE

OF INFLAMMATORY BREAST CANCER

by

BERNADETTE C. VICTOR

December 2011

Advisor: Dr. Bonnie F. Sloane

Major: Cancer Biology

Degree: Doctor of Philosophy

Inflammatory breast cancer (IBC) is an aggressive, metastatic and

highly angiogenic form of locally advanced breast cancer. Breast

cancer invasion has been linked to proteolytic activity at the tumor

cell surface. We observed that uPA, uPAR and enzymatically active

cathepsin B were all present in caveolae fractions isolated from

SUM149 cells. Using a live-cell proteolysis assay, we demonstrated

that both IBC cell lines degrade …

Human Trophoblast Survival And Invasion In The Developing Placenta: Autocrine Regulation By Hbegf, Philip Jessmon

Wayne State University Dissertations

HBEGF is a multifunctional protein in early pregnancy that induces cytotrophoblast (CTB) cell differentiation to an invasive phenotype, protects against apoptosis, and is involved in an autocrine signaling mechanism that leads to its own protein synthesis. CTBs exist in a low O₂ environment during the first 10 weeks of implantation, during which they invade the decidualized uterine stroma. Inhibitors of intracellular signaling pathways demonstrated that at 20% O₂ HBEGF induces an increase in cell migration through the ERK, MAPK14, JNK, or PIK3 pathways downstream of signaling through its ERBB receptors. Also downstream of these four pathways, HBEGF induces …

Palmitoylation And The Yeast Casein Kinase Yck2, Irene Papanayotou

Wayne State University Dissertations

Palmitoylation is a post-translational lipid modification that allows proteins to interact with membranes. In the yeast Saccharomyces cerevisiae, the casein kinase Yck2 is palmitoylated twice at its two C-terminal palmitoyl-accepting cysteine residues, by the palmitoyl-transferring enzyme Akr1. Once palmitoylated, Yck2 traffics through the well characterized secretory pathway to the plasma membrane where it participates in many cellular functions, including bud morphogenesis, cytokinesis, nutrient sensing, and receptor internalization. While the hydrophilic Yck2 is presumably synthesized on cytosolic ribosomes, it gains access to the membrane system by interaction with the six transmembrane-spanning Golgi-localized Akr1. Since palmitoylation occurs at membranes and the palmitoyl-transferases …

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 …

The Role Of The Sparc Acidic Domain And Egf-Like Module In Glioma Migration, Invasion, And Signaling, Heather M. Mcclung

Wayne State University Dissertations

THE ROLE OF THE SPARC ACIDIC DOMAIN AND EGF-LIKE MODULE IN GLIOMA MIGRATION, INVASION, AND SIGNALING

HEATHER M. MCCLUNG

Advisor: Sandra A. Rempel, Ph.D.

Major: Pharmacology

Degree: Doctor of Philosophy

We have previously shown that Secreted Protein Acidic and Rich in Cysteine (SPARC) is upregulated in all astrocytoma grades and increases tumor cell migration and invasion. It is thought that different domains within the protein may regulate SPARC functions, suggesting domain-specific targeting to inhibit invasion. To enhance our understanding of SPARC-mediated invasion, we first confirm, at the protein level, our previous cDNA array results, that SPARC increases expression of the …

Frazzled And Abelson Interact To Regulate The Actin Cytoskeleton In Drosophila, Bridget Elsa Varughese

Wayne State University Dissertations

Guidance receptors such as Frazzled affect cell shape and motility by directly or indirectly modulating the cytoskeleton. Fra is particularly needed for the formation of the posterior commissures in a developing Drosophila embryo. The cytoplasmic tyrosine kinase, Abelson Kinase (Abl) enhances the loss of commissures observed in fra mutant. Abl physically interacts with Frazzled to help guide commissural axons across the midline. Furthermore, the loss of commissural axons is only seen when the actin dynamics are perturbed. Abl is also known to regulate actin-dependent processes underlying formation of filopodia, microspikes and membrane ruffles. So, we established a Drosophila S2 cell …

The Cellular And Genetic Mechanisms Underlying Morphological Diversity Of Insects, Nataliya Turchyn

Wayne State University Dissertations

The vast diversity of insect appendages provides an excellent model for examining the cellular and molecular basis of phenotypic changes in nature. One of the hallmarks of evolution of insect appendages is the differential enlargement of hind (T3) legs compared to their fore (T1) and mid (T2) counterparts. While basic information regarding the genetic mechanisms that regulate the size of the T3 legs became available in the past several years, virtually nothing is known about cellular mechanisms involved. To address this issue, we measured the relative mitotic activity (RMA) in Acheta domesticus (house cricket) and Oncopeltus fasciatus (milkweed bug), two …

Regulatory And Functional Aspects Of Foxo3a Transcription Factor And Their Implications In Prostate Cancer, Melissa Elise Dobson

Wayne State University Dissertations

The P13K/Akt pathway is a critical mediator of growth factor signaling involving many cellular functions. The deregulation of this pathway has been shown to be involved in the development of various cancers. One of the main targets of this pathway is FoxO3a, a transcription factor whose target genes are involved in important cellular processes such as apoptosis, cell cycle control, and glucose metabolism. FoxO3a is regulated by various post translational modifications including acetylation, ubiquitination and phosphorylation. The transcription factor is directly phosphorylated by Akt on 3 residues: Threonine 32, Serine 253 and Serine 315. Phosphorylation by Akt generates binding sites …

Matriptase/Pdgf D/Beta-Pdgfr Signaling Axis In Human Prostate Cancer: The Role Of Pten In The Regulation Of Pdgf D Expression, M. Katie Conley-Lacomb

Wayne State University Dissertations

Platelet Derived Growth Factor (PDGF) is a family of mesenchymal growth factors that regulate cell proliferation, migration, and differentiation. Unlike the classic PDGF ligands A and B, which are secreted as active dimers, PDGF D must undergo extracellular proteolytic processing to remove its N-terminal CUB domain from the C-terminal PDGF growth domain before the ligand is able to stimulate its receptor, PDGF receptor beta (?-PDGFR). Importantly, recent clinical studies have shown that ?-PDGFR is upregulated in primary prostate cancer and bone metastases. However, PDGF B, formerly thought to be the sole ligand for ?-PDGFR, is not expressed in clinical prostate …

Induction And Regulation Of Autophagy By Novel Prenylation Inhibitors In Sts-26t Malignant Peripheral Nerve Sheath Tumor (Mpnst) Cells, Komal Madhukar Sane

Wayne State University Dissertations

Prenylation pathways have been targeted via several different compounds that inhibit farnesyl transferase (FTase) and/or geranylgeranyl transferase (GGTase) enzymes in many cellular and animal models of cancer. Some of these have also been evaluated in clinical trials with limited success. Multiple mechanisms of action have been elucidated for such compounds, including cell cycle arrest, proteasome inhibition, apoptosis and most recently, autophagy. However, there is still an urgent need of effective agents of this class of anti-tumor therapeutics. In this dissertation, I sought to delve into this issue by characterizing our novel prenylation inhibitors and their potential as anti-tumor agents. Novel …

Fine Structure Of The Substantia Nigra Pars Lateralis In The Opossum, Terrence Paul Ma

Wayne State University Dissertations

The cytoarchitecture of the substantia nigra pars lateralis (SNl) of the opossum was examined by light and electron microscopic procedures. Nissl-stained, Golgi-impregnated, HRP-reacted and electron microscopic preparations were utilized in this study. SNl neurons were divided into three groups on the basis of cell size; small, medium-sized, and large neurons. Small cells had a nucleus with some clumped heterochromatin, a relatively organelle-free cytoplasm, and long, thin, sparcely branching dendrites. Electron microscopic examination of small cells confirmed the light microscopic observations. Medium-sized neurons possessed a relatively clear nucleus, clumps of Nissl substance, especially at a juxtanuclear position near the emergence of …