Medical Sciences Commons^™

Identification Of Apoptosis Pathway In Treacher Collins Syndrome, Khaled Alsayegh

Theses and Dissertations

Treacher Collins Syndrome (TCS) is a rare autosomal dominant disorder characterized by severe craniofacial defects. The syndrome is associated with mutations in the TCOF1 gene, which encodes a nucleolar phosphoprotein called treacle. Model organisms have been generated to model the disease and have revealed knowledge about the etiology and pathogenesis of the disorder. The craniofacial abnormality observed in TCS patients is found to be caused by an increased level of apoptosis in the neuroepithelium and from this it has been suggested that treacle is important for proper formation and proliferation of neural crest cells that will ultimately contribute to the …

Transcriptional, Epigenetic, And Signal Events In Antifolate Therapeutics, Alexandra Racanelli

Theses and Dissertations

A targeted approach to the development of antifolate therapies has been sought for many years. Central to the success of such development is an understanding of the molecular mechanisms dictating the sensitivity of cells to antifolates and the fundamental differences of these processes between normal and neoplastic phenotypes. This dissertation addressed transcriptional mechanisms and cell-signaling events responsible for the efficacy of antifolate therapies. Transcriptional processes and cell signaling pathways are often aberrant in neoplastic tissues, providing a potential point of distinction between a normal and neoplastic cellular state. Folylpolyglutamate synthetase (FPGS) catalyzes the formation of poly-γ-glutamate derivatives of folates and …

Roles Of Krüppel Like Factors Klf1, Klf2, And Klf4 In Embryonic Beta-Globin Gene Expression, Yousef Alhashem

Theses and Dissertations

Krüppel like factors (KLFs) are a family of 17 proteins whose main function is gene regulation by binding to DNA elements in the promoters of various genes. KLF transcription factors recognize CACCC-elements and act as activators or repressors of the gene expression. Among the 17 family members, KLF1, KLF2, and KLF4 share high homology to each other. KLF1 is the founding member of the family and is an erythroid-specific protein. KLF2 is expressed in erythroid, endothelial, and other cells. KLF4 is expressed in endothelial, smooth muscle, and other cells. In this thesis, the functions of these KLFs were reviewed in …

The Effect Of Chronic Constriction Injury On Cellular Systems Within Nociceptive Pathways In The Mouse, Michelle Hoot

Theses and Dissertations

Chronic neuropathic pain is often difficult to treat due to its resistance to therapeutic intervention. This is due in part to the poor understanding of the physiological mechanisms involved in the establishment and maintenance of neuropathic pain states. The neuropathic pain model, chronic constriction injury of the sciatic nerve, produced robust pain hypersensitivity in our mice. It also induced significant changes in the mitogen activated protein kinase family, and the cannabinoid and µ-opioid systems in three different brain areas involved in the modulation or regulation of pain states. CCI induced a 2.5 fold increase in mRNA of the kinase Raf-1 …

Obesity, Adiposity, And Satiety In Mouse Models Of Smith-Magenis Syndrome And Dup(17)(P11.2) Syndrome, Brooke Burns

Theses and Dissertations

Smith-Magenis syndrome (SMS) is a complex disorder caused by haploinsufficiency of RAI1 and characterized by sleep disturbances, behavioral abnormalities, mental retardation, and obesity in teens and adults. Rai1+/- mice are obese after 20 weeks. Dup(17)(p11.2) syndrome is a complex disorder associated with overexpression of RAI1. A transgenic mouse model of dup(17)(p11.2) syndrome overexpresses Rai1 and results in a mouse that is growth delayed. In order to characterize the obese phenotypes of mouse models of SMS and the role of RAI1 in obesity, daily food intake and serum levels of insulin, glucose, PPY, and leptin were measured; adiposity was studied by …

Therapeutic Drugs In Cancer, Teneille Walker

Theses and Dissertations

The first study examined the interaction between low doses of the multi-kinase inhibitor sorafenib and the histone deacetylase inhibitor vorinostat in colon cancer cells. Sorafenib and vorinostat synergized to kill HCT116 and SW480 cells. In SW480 cells, sorafenib+vorinostat toxicity correlated with CD95 activation and CD95-stimulated autophagy. Drug lethality in SW480 cells was blocked by knock down of CD95. In SW620 cells that are patient matched to SW480 cells, sorafenib+vorinostat toxicity was significantly reduced that correlated with a lack of CD95 activation and lower expression of ceramide synthase 6 (LASS6). Overexpression of LASS6 in SW620 cells enhanced drug-induced CD95 activation and …

Therapeutic Drugs In Cancer And Resistance., Aditi Pandya Martin

Theses and Dissertations

We investigated the mechanism of toxicity and resistance development of small molecule tyrosine kinase inhibitor lapatinib in HCT 116 colon cancer cells. Lapatinib mediated cell death in HCT 116 cells was caspase independent and involved cytosolic release of apoptosis inducing factor. Treatment of HCT 116 cells with 10µM Lapatinib lead to the outgrowth of lapatinib resistant HCT 116 cells. Our studies show that alterations in the expression and activation of Bcl-2 family proteins allow lapatinib resistant HCT 116 cells to resist cytotoxic effects of lapatinib as well as of other commonly used chemotherapeutic agents. In hepatoma and pancreatic cancer cells, …

Apoptotic Engulfment Pathway And Schizophrenia, Xiangning Chen, Cuie Sun, Qi Chen, F. Anthony O'Neill, Dermot Walsh, Ayman H. Fanous, Kodavali V. Chowdari, Vishwajit L. Nimgaonkar, Adrian Scott, Sibylle G. Schwab, Dieter B. Wildenauer, Ronglin Che, Wei Tang, Yongyong Shi, Lin He, Xiong-Jian Luo, Bing Su, Todd L. Edwards, Zhongming Zhao, Kenneth S. Kendler

Human and Molecular Genetics Publications

Background

Apoptosis has been speculated to be involved in schizophrenia. In a previously study, we reported the association of the MEGF10 gene with the disease. In this study, we followed the apoptotic engulfment pathway involving the MEGF10, GULP1, ABCA1 and ABCA7 genes and tested their association with the disease.

Methodology/Principal Findings

Ten, eleven and five SNPs were genotyped in the GULP1, ABCA1 and ABCA7 genes respectively for the ISHDSF and ICCSS samples. In all 3 genes, we observed nominally significant associations. Rs2004888 at GULP1 was significant in both ISHDSF and ICCSS samples (p = 0.0083 and …

Synchronization Of Pancreatic Islet Oscillations By Intrapancreatic Ganglia: A Modeling Study, B. Fendler, M. Zhang, L. Satin, R. Bertram

Pharmacology and Toxicology Publications

Abstract

Plasma insulin measurements from mice, rats, dogs, and humans indicate that insulin levels are oscillatory, reflecting pulsatile insulin secretion from individual islets. An unanswered question, however, is how the activity of a population of islets is coordinated to yield coherent oscillations in plasma insulin. Here, using mathematical modeling, we investigate the feasibility of a potential islet synchronization mechanism, cholinergic signaling. This hypothesis is based on well-established experimental evidence demonstrating intrapancreatic parasympathetic (cholinergic) ganglia and recent in vitro evidence that a brief application of a muscarinic agonist can transiently synchronize islets. We demonstrate using mathematical modeling that periodic pulses of …

Creation Of A Mouse With A Humanized Fpgs Gene Compatible With Normal Development, Linying Xie

Theses and Dissertations

Abstract: Folylpoly-γ-glutamate synthetase (FPGS) catalyzes the formation of polyglutamate forms of the reduced folates and antifolates such as methotrexate (MTX) and pemetrexed; this allows the retention of folates and antifolate cancer drugs inside the cell. The enzyme activity of FPGS is essential for cell proliferation and survival. The mouse fpgs gene contains two promoters spaced 10 kb apart which are activated in a tissue-specific manner. The upstream promoter (P1) and exons A1a and A1b are used in some differentiated tissues, mainly liver and kidney, whereas the downstream promoter (P2) and exon 1 are used in rapidly dividing cells. In contrast, …

Identification Of Pharmacological And Molecular Mechanisms Involved In Nicotine Withdrawal, Kia Jackson

Theses and Dissertations

Tobacco dependence is the leading cause of preventable death in the United States. Despite currently available smoking cessation therapies, there is a high rate of relapse in smoking among those attempting to quit. While the somatic signs of nicotine withdrawal (insomnia, increased appetite, weight gain) contribute to the continuation of smoking behavior, it has been hypothesized that the affective signs (depression, anxiety, craving, irritability) are greater motivators of relapse and continued tobacco use. There are few studies that assess the molecular and receptor-mediated mechanisms of nicotine withdrawal; therefore, our studies focus on identifying the nicotinic acetylcholine receptor (nAChR) subtypes and …

Promotion Of Tumor Cell Death Through The Induction Of, Tuyen Nguyen

Theses and Dissertations

Microtubule poisons have proven to be effective in the treatment of a variety of malignancies. Although taxol-based derivatives promote microtubule stabilization, there is continuing interest in compounds that, like colchicines, act as microtubule destabilizing agents. Previous work from this laboratory showed that the novel microtubule poison, JG- 03-14, was active against breast tumor cells, promoting autophagic cell death. In the current work, we studied the influence of JG-03-14 on p53 wild type HCT116 colon carcinoma cells. A crystal violet sensitivity assay indicated that JG-03-14 induced growth inhibition, with 75% suppression of growth evident at a concentration of 500 nM. Time …

Effects Of Altered Superoxide Dismutase Expression On Age-Related Functional Declines And Survival In Drosophila, Ian Martin

Theses and Dissertations

Most organisms experience progressive declines in physiological function as they age. A number of studies in a variety of species support a strong link between oxidative damage, age-related functional declines and life span determination. Here, manipulating the expression levels of superoxide dismutase (SOD) isoenzymes SOD1 and SOD2, resulted in altered functional senescence and survival characteristics in Drosophila. Overexpression of cytosolic Sod1 using the yeast GAL4/UAS system conferred a 30-34% increase in mean life span and resulted in an attenuated senescence of odor avoidance behavior in aging flies. Tissue-specific Sod1 overexpression selectively in the nervous system or muscle failed to reproduce …

Nucleoplasmic And Cytoplasmic Degradation Of Telomerase: Implications Toward Telomerase-Based Cancer Therapy, Binh Nguyen

Theses and Dissertations

Telomerase is a ribonucleoprotein that is reactivated in cancer cells to allow for continuous cellular division and indefinite growth. With telomerase being expressed in more than 85% of all cancer, it is imperative that we understand how to selectively inactivate and degrade this unique DNA polymerase. In doing so, we can specifically target tumor cells to erode their telomeres so that they will undergo apoptosis or senescence. Through this research, we have learned that telomerase can be degraded in the nucleoplasm by Hsp90 chaperone inhibition and in the cytoplasm by the dominant negative mutant, D712A V713I. These findings should guide …

Morphine Exacerbates Hiv-1 Tat-Induced Cytokine Production In Astrocytes Through Convergent Effects On [Ca2+]I, Nf-Κb Trafficking And Transcription, Nazira El-Hage, Annadora J. Bruce-Keller, Tatiana Yakovleva, Igor Bazov, Georgy Bakalkin, Pamela E. Knapp, Kurt F. Hauser

Pharmacology and Toxicology Publications

Astroglia are key cellular sites where opiate drug signals converge with the proinflammatory effects of HIV-1 Tat signals to exacerbate HIV encephalitis. Despite this understanding, the molecular sites of convergence driving opiate-accelerated neuropathogenesis have not been deciphered. We therefore explored potential points of interaction between the signaling pathways initiated by HIV-1 Tat and opioids in striatal astrocytes. Profiling studies screening 152 transcription factors indicated that the nuclear factor-kappa B (NF-κB) subunit, c-Rel, was a likely candidate for Tat or Tat plus opiate-induced increases in cytokine and chemokine production by astrocytes. Pretreatment with the NF-κB inhibitor parthenolide provided evidence that Tat±morphine-induced …

Long Lasting Synchronization Of Calcium Oscillations By Cholinergic Stimulation In Isolated Pancreatic Islets, Min Zhang, Bernard Fendler, Bradford Peercy, Pranay Goel, Richard Bertram, Arthur Sherman, Leslie Satin

Pharmacology and Toxicology Publications

Abstract

Individual mouse pancreatic islets exhibit oscillations in [Ca²⁺]_i and insulin secretion in response to glucose in vitro, but how the oscillations of a million islets are coordinated within the human pancreas in vivo is unclear. Islet to islet synchronization is necessary, however, for the pancreas to produce regular pulses of insulin. To determine whether neurohormone release within the pancreas might play a role in coordinating islet activity, [Ca²⁺]_i changes in 4–6 isolated mouse islets were simultaneously monitored before and after a transient pulse of a putative synchronizing agent. The degree of synchronicity was …

Gap Junction Coupling And Calcium Waves In The Pancreatic Islet, Richard K.P. Benninger, Min Zhang, W. Steven Head, Leslie S. Satin, David W. Piston

Pharmacology and Toxicology Publications

Abstract

The pancreatic islet is a highly coupled, multicellular system that exhibits complex spatiotemporal electrical activity in response to elevated glucose levels. The emergent properties of islets, which differ from those arising in isolated islet cells, are believed to arise in part by gap junctional coupling, but the mechanisms through which this coupling occurs are poorly understood. To uncover these mechanisms, we have used both high-speed imaging and theoretical modeling of the electrical activity in pancreatic islets under a reduction in the gap junction mediated electrical coupling. Utilizing islets from a gap junction protein connexin 36 knockout mouse model together …

Examining The Behavioral And Molecular Aspects Of Adolescent Nicotine Dependence: Implications For Vulnerability To Drugs Of Abuse, Dena Heath Kota

Theses and Dissertations

Approximately 200 million men and 100 million women smoke worldwide. In the United States, an estimated 25.9 million men (23.9 percent) and 20.7 million women (18.1 percent) are smokers. The commencement of smoking at a young age is thought to increase addiction liability, decrease the probability of successful cessation, and correlate with a higher number of cigarettes smoked per day. Studies from the World Health Organization indicate that between 80,000 and 100,000 children start smoking every day worldwide. These statistics suggest that adolescence is a critical phase for developing nicotine dependence. The work in this dissertation contributes to the further …

Endocannabinoid Modulation Of Spatial Memory In Aversively And Appetitively Motivated Barnes Maze Tasks, John Pinckney Harloe

Theses and Dissertations

Genetic deletion or pharmacological blockade of the CB1 receptor has been reported to impair extinction learning in aversive conditioning (i.e., conditioned fear and Morris water maze) paradigms, but not in operant procedures in which food reinforcement is earned. It is difficult to discern whether the differential effects caused by CB1 receptor disruption on extinction result from the hedonics (i.e., aversive vs. appetitive) or is related to the required responses associated with these disparate tasks. In order to evaluate whether the hedonics is the determining factor, we used either aversive (i.e., escape from bright lights and air turbulence) or appetitive (i.e., …

Interaction Of Glycolysis And Mitochondrial Respiration In Metabolic Oscillations Of Pancreatic Islets, Richard Bertram, Leslie S. Satin, Morten Gram Pedersen, Dan S. Luciani, Arthur Sherman

Pharmacology and Toxicology Publications

Abstract

Insulin secretion from pancreatic β-cells is oscillatory, with a typical period of 2–7 min, reflecting oscillations in membrane potential and the cytosolic Ca²⁺ concentration. Our central hypothesis is that the slow 2–7 min oscillations are due to glycolytic oscillations, whereas faster oscillations that are superimposed are due to Ca²⁺ feedback onto metabolism or ion channels. We extend a previous mathematical model based on this hypothesis to include a more detailed description of mitochondrial metabolism. We demonstrate that this model can account for typical oscillatory patterns of membrane potential and Ca²⁺ concentration in islets. It also …

Telomerase Inhibition And Sensitization Of Breast Tumor Cells, Kennon R. Poynter

Theses and Dissertations

Telomerase, a ribonucleoprotein enzyme minimally composed of an RNA template (hTR) and a catalytically active protein subunit (hTERT), synthesizes telomeric repeats onto chromosome ends and is obligatory for continuous tumor cell proliferation, as well as malignant progression of breast cancer cells. Telomerase is an attractive anticancer therapeutic target because its activity is present in over 90% of human cancers, including more than 95% of breast carcinomas, but undetectable in most somatic cells. Traditions chemo- and radio-therapies lack the ability to effectively control and cure breast cancer, in part because residual cells are or become resistant to DNA damaging modalities.While various …

A Comparision Of Delta-9-Tetrahydrocannabinol Dependence In C57bl/6j Mice And Fatty Acid Amide Hydrolase Knock Out Mice, Brittany Leigh Alice Carlson

Theses and Dissertations

The idea that humans and laboratory animals can become physically dependent on marijuana or its primary psychoactive constituent, delta-9-tetrahydrocannabinol (THC), is gaining acceptance. However, there are no currently approved pharmacotherapies to treat cannabinoid withdrawal. The objective of this thesis was to evaluate whether elevating endogenous anandamide levels using mice lacking fatty acid amide hydrolase (FAAH), the enzyme responsible for anandamide metabolism, would ameliorate THC dependence. Mice were treated subchronically with a low or high THC dosing regimen and challenged with the CB1 receptor antagonist, rimonabant, to precipitate withdrawal. Following subchronic THC treatment, rimonabant precipitated a significant increase in paw flutters …

The Substituted Pyrrole Jb-03-14 Induces Autophagic Cell Death And Growth Arrest In Breast Tumor Cells, Christopher Ryan Arthur

Theses and Dissertations

The use of chemotherapy in the treatment of cancer has stimulated the demand for better chemotherapeutic agents that are more potent at destroying tumor cell populations and more selective for the specific tumor versus normal host tissues. This project is directed at discovering new anti-tumor agents that are effective against breast cancer based on structures derived from marine organisms, specifically brominated pyrroles. We utilized an in vitro breast cancer model to study the effects of pyrroles on tumor proliferation and survival, as well as growth arrest and cell death. Our findings indicate that the substituted pyrrole JG-03-14 induces time dependent …

The Quantitative Genetics Of Neurodevelopment: A Magnetic Resonance Imaging Study Of Childhood And Adolescence, James Eric Schmitt

Theses and Dissertations

Understanding the causes of individual differences in brain structure may give clues about the etiology of cognition, personality, and psychopathology, and also may identify endophenotypes for molecular genetic studies on brain development. We performed a comprehensive statistical genetic study of anatomic neuroimaging data from a large pediatric sample (N=600+) of twins and family members from the Child Psychiatry Branch at the NIMH. These analyses included variance decomposition of structural volumetric endophenotypes at several levels of resolution, voxel-level analysis of cortical thickness, assessment of gene by age interaction, several multivariate genetic analyses, and a search for genetically-mediated brain-behavioral relationships. These analyses …

Dual Regulation Of Telomerase Activity By Hsf1 And Its Role In Prostate Cancer Progression, Keith Douglas Ostergaard Jensen

Theses and Dissertations

It has been shown that the key components of the hsp90 chaperone complex, including hsp90, p23, hsp70, hsp40, and HOP (p60), associate with telomerase; however, their specific roles in telomerase function and tumor progression have not yet been defined. HSF1, the primary mammalian heat shock protein transcription factor, may affect telomerase activity and transformation by regulating the expression of several hsp90 chaperone complex proteins in response to stress as well as regulating the transcription of hTERT, the protein subunit of telomerase.In our in vitro model of prostate cancer progression, as cells progress from immortal but non-tumorigenic (P69) to tumorigenic (M2182) …

Glutamate Excitotoxicty Activates A Novel Calcium Permeable Ion Channel In Cultured Hippocampal Neurons, Laxmikant Sudhir Deshpande

Theses and Dissertations

Glutamate excitotoxicity is the predominant mechanism implicated in neuronal cell death associated with neurological disorders such as stroke, epilepsy, traumatic brain injury and ALS. Excessive stimulation of NMDA subtypes of glutamate receptors leads to protracted intracellular calcium elevations triggering calcium mediated neurotoxic mechanisms culminating in delayed neuronal cell death. In addition, glutamate excitotoxicity induces a NMDA dependent extended neuronal depolarization mediated by continuous calcium influx that correlates with delayed neuronal death. Attempts to prevent neuronal death by blocking calcium entry into the neurons using calcium channel blockers or NMDA receptor antagonists have failed to provide any beneficial effects in clinical …

Immunotoxicological Evaluation Of Critical Windows Of Development Following Exposure To 1,2:5,6 Dibenzanthracene In B6c3f1 Mice, Denise Marie Hernandez

Theses and Dissertations

Numerous findings have suggested that the increased prevalence of childhood illnesses such as cancer, infection, and allergy may be due to environmental exposures. One such cause may be maternal smoking or passive smoke exposure. Known immunotoxicants in cigarette smoke and environmental pollution include polycyclic aromatic hydrocarbons such as 1,2:5,6 Dibenzanthracene (DBA). The objective of these studies was to evaluate the immunosuppressive effects of DBA on various stages of immune system development. Adult mice were administered DBA daily in corn oil at dose levels of 158, 500, 1580, and 5000 µglkg S.C. for 28 days. Immunosuppression was not observed at levels …

Influence Of The Vitamin D3 Analog Eb 1089 On Senescence And Cell Death Pathways In The Response Of Breast Tumor Cells To Ionizing Radiation, Gerald Alan Demasters

Theses and Dissertations

A senescence-like growth arrest succeeded by rapid recovery of proliferative capacity is observed in MCF-7 breast tumor cells exposed to fractionated radiation (5 x 2Gy) alone. Exposure to the vitamin D3 analog EB 1089 (100nM) prior to irradiation converts the initial growth arrest response to cell death in part through the inhibition of radiation-induced senescence and promotion of both apoptotic and autophagic cell death. More importantly, EB 1089 was shown to profoundly reduce the rate of recovery following fractionated irradiation. The effect of EB 1089 on the temporal response to radiation is also observed in MCF-7 cells expressing caspase 3, …

Action Of Tyrosyl Dna Phosphodiesterase On 3'-Phosphoglycolate Terminated Dna Strand Breaks, Haritha Tatavarthi

Theses and Dissertations

Free radical-mediated DNA double strand breaks (DSBs) are induced either directly by ionizing radiation or by certain chemicals like bleomycin. These breaks are terminated by 3'-PG (PO4CH2COOˉ) or 3'-phosphate groups formed as a result of fragmentation of deoxyribose. To study the nature of repair of these 3'-blocked breaks, we constructed substrates mimicking free-radical induced DSBs. Human and yeast tyrosyl DNA-phosphodiesterase (Tdpl) efficiently processed substrates with 3'-PGs, in either the presence or absence of magnesium, to give a 3'-phosphate. Gel filtration chromatography and western blotting codmed that the putative enzyme in human extracts that efficiently processed PG was indeed tyrosyl DNA-phosphodiesterase. …

Pathological Upregulation Of A Calcium-Stimulated Phosphatase, Calcineurin, In Two Models Of Neuronal Injury, Jonathan Elledge Kurz

Theses and Dissertations

Excitotoxic calcium influx and activation of calcium-regulated systems is a common event in several types of neuronal injury. This mechanism has been the focus of intense research, with the hope that a more complete understanding of how neuronal injury affects calcium-regulated systems will provide effective treatment options. This study examines one such calcium-stimulated enzyme, calcineurin, in the context of two common neurological pathologies, status epilepticus and traumatic brain injury.Status epilepticus was induced by pilocarpine injection. NMDA-dependent increases in calcineurin activity were observed in cortical and hippocampal homogenates. Upon closer examination, the most profound increases in activity were found to be …