Cell and Developmental Biology Commons^™

Oral Immunization Of Rhesus Macaques With Adenoviral Hiv Vaccines Using Enteric-Coated Capsules, George T. Mercier, Pramod N. Nehete, Marco F. Passeri, Bharti N. Nehete, Eric A. Weaver, Nancy Smyth Templeton, Kimberly Schluns, Stephanie S. Buchl, K. Buchl, Michael A. Barry

Nebraska Center for Virology: Faculty Publications

Targeted delivery of vaccine candidates to the gastrointestinal (GI) tract holds potential for mucosal immunization, particularly against mucosal pathogens like the human immunodeficiency virus (HIV). Among the different strategies for achieving targeted release in the GI tract, namely the small intestine, pH sensitive enteric coating polymers have been shown to protect solid oral dosage forms from the harsh digestive environment of the stomach and dissolve relatively rapidly in the small intestine by taking advantage of the luminal pH gradient. We developed an enteric polymethacrylate formulation for coating hydroxy-propyl-methyl-cellulose (HPMC) capsules containing lyophilized Adenoviral type 5 (Ad5) vectors expressing HIV-1 gag …

Thermodynamic Evidence That Ganglioside-Mediated Insertion Of Botulinum A Into The Cholinergic Nerve Ending May Precede Endocytosis And Acidification: A Langmuir Film Study, Bradley Adam Strongin

Theses and Dissertations

Botulinum Neurotoxin (BoNT) is one of the most potent toxins known to human kind. The Atomic Force Microscope (AFM) was employed to investigate the conditions under which BoNT type A heavy chain would bind and/or insert into mica supported dipalmitoylphosphatidylcholine (DPPC) lipid bilayers. As an alternate technique, DPPC/GT1b or total ganglioside extract (80:20) monolayers of a Langmuir Blodgett (LB) Trough were adapted to be artificial membrane models for toxin insertion studies. We conclude that LB monolayer studies are a promising candidate for BoNT/A membrane insertion investigation. Botulinum neurotoxin serotype A insertions into the LB monolayers in the presence of BoNT/A …

Effects Of Endurance Training On The Ampk Response To Exercise., David Gerald Chesser

Theses and Dissertations

Activation of AMP-activated protein kinase (AMPK) results in the upregulation of several intracellular systems which help to prepare a cell for a high energy challenge. The magnitude of the AMPK response to a 10 min bout of exercise has been found to decrease in red quadriceps (RQ) following training, while putative AMPK roles seem to be maintained; specifically, the biogenesis of mitochondria and higher levels of hexokinase II and glucose transporter 4 (GLUT4). If the AMPK response to exercise is responsible in part for these adaptations, how can they be maintained if the AMPK response is attenuated? The purpose of …

Light-Evoked Calcium Responses Of Isolated Melanopsin- Expressing Retinal Ganglion Cells, Andrew T. E. Hartwick, Jayne R. Bramley, Jianing Yu, Kelly T. Stevens, Charles N. Allen, William H. Baldridge, Patricia J. Sollars, Gary E. Pickard

School of Veterinary and Biomedical Sciences: Faculty Publications

A small number (<2%) of mammalian retinal ganglion cells express the photopigment melanopsin and are intrinsically photosensitive (ipRGCs). Light depolarizes ipRGCs and increases intracellular calcium levels ( [Ca²⁺]_i ) but the signaling cascades underlying these responses have yet to be elucidated. To facilitate physiological studies on these rare photoreceptors, highly enriched ipRGC cultures from neonatal rats were generated using anti-melanopsin-mediated plate adhesion (immunopanning). This novel approach enabled experiments on isolated ipRGCs, eliminating the potential confounding influence of rod/cone-driven input. Light induced a rise in [Ca²⁺]_i (monitored using fura-2 imaging) in the immunopanned ipRGCs and the source of this Ca²⁺ signal was investigated. The Ca²⁺ responses were inhibited by 2-aminoethoxydiphenyl borate, SKF-96365 (1–2-(4-methoxyphenyl)-2-[3-(4-methoxyphenyl)propoxy]ethyl-1H-imidazole), flufenamic acid, …

Genome-Wide Transcriptional Profiling Of The Cyclic Amp-Dependent Signaling Pathway During Morphogenic Transitions Of Candida Albicans, Yong-Sun Bahn, Matthew Molenda, Janet F. Staab, Courtney A. Lyman, Laura J. Gordon, Paula Sundstrom

Dartmouth Scholarship

Candida albicans is an opportunistic human fungal pathogen that causes systemic candidiasis as well as superficial mucosal candidiasis. In response to the host environment, C. albicans transitions between yeast and hyphal forms. In particular, hyphal growth is important in facilitating adhesion and invasion of host tissues, concomitant with the expression of various hypha-specific virulence factors. In previous work, we showed that the cyclic AMP (cAMP) signaling pathway plays a crucial role in morphogenic transitions and virulence of C. albicans by studying genes encoding adenylate cyclase-associated protein (CAP1) and high-affinity phosphodiesterase (PDE2) (Y. S. Bahn, J. Staab, and P. Sundstrom, Mol. …

Mapk Survival Signaling In Melanoma, Matthew W. Vanbrocklin

Dissertations

Extracellular signals activate mitogen-activated protein kinase (MAPK) cascades potentiating biological activities such as cell proliferation, differentiation, and survival. Constitutive activation of MAPK signaling pathways is implicated in the development and progression of many human cancers, including melanoma. Mutually exclusive activating mutations in NRAS or BRAF are found in ∼85% of all melanomas resulting in constitutive activation of the MAPK pathway (RAS-BRAF-MEK-ERK-RSK). We have previously demonstrated that inhibition of this pathway with small molecule MEK inhibitors selectively induces apoptosis in human melanoma cells both in vitro and in vivo, but not in normal melanocytes. These results support the notion that the …

A Conserved Cam- And Radial Spoke–Associated Complex Mediates Regulation Of Flagellar Dynein Activity, Erin E. Dymek, Elizabeth F. Smith

Dartmouth Scholarship

For virtually all cilia and eukaryotic flagella, the second messengers calcium and cyclic adenosine monophosphate are implicated in modulating dynein- driven microtubule sliding to regulate beating. Calmodulin (CaM) localizes to the axoneme and is a key calcium sensor involved in regulating motility. Using immunoprecipitation and mass spectrometry, we identify members of a CaM-containing complex that are involved in regulating dynein activity. This complex includes flagellar-associated protein 91 (FAP91), which shares considerable sequence similarity to AAT-1, a protein originally identified in testis as an A-kinase anchor protein (AKAP)- binding protein. FAP91 directly interacts with radial spoke protein 3 (an AKAP), which …

Cell Death In Health And Disease, Richard A. Lockshin, Zahra Zakeri

Publications and Research

Cell death is clearly an important factor in development, homeostasis, pathology and in aging, but medical efforts based on controlling cell death have not become major aspects of medicine. There are several reasons why hopes have been slow to be fulfilled, and they present indications for new directions in research. Most effort has focused on the machinery of cell death, or the proximate effectors of apoptosis and their closely associated and interacting proteins. But cells have many options other than apoptosis. These include autophagy, necrosis, atrophy and stepwise or other alternate means of self-disassembly. The response of a cell to …

Enhancement Of Efficacy Of Prrsv Vaccines By Altering The Glycosylation Pattern Of Viral Glycoproteins, Asit K. Pattnaik

School of Veterinary and Biomedical Sciences: Faculty Publications

PRRSV, the causative agent of PRRS is of major economic significance to the pork industry in the USA and around the world. Current commercial vaccine does not provide adequate protection against PRRSV outbreaks. Therefore, there is an urgent need for development of more efficacious vaccine to combat PRRS. Our previous studies have suggested that (i) induction of neutralizing antibody response is an important correlate of evaluating the efficacy of a vaccine; (ii) neutralizing antibodies can be enhanced by hypoglycosylation of the major surface glycoprotein (GP5). While it is known that GP5 plays a prominent role in neutralizing antibody induction, it …

Role Of Actin Cytoskeletal Dynamics In Activation Of The Cyclic Amp Pathway And Hwp1 Gene Expression In Candida Albicans, Michael J. Wolyniak, Paula Sundstrom

Dartmouth Scholarship

Changes in gene expression during reversible bud-hypha transitions of the opportunistic fungal pathogen Candida albicans permit adaptation to environmental conditions that are critical for proliferation in host tissues. Our previous work has shown that the hypha-specific adhesin gene HWP1 is up-regulated by the cyclic AMP (cAMP) signaling pathway. However, little is known about the potential influences of determinants of cell morphology on HWP1 gene expression. We found that blocking hypha formation with cytochalasin A, which destabilizes actin filaments, and with latrunculin A, which sequesters actin monomers, led to a loss of HWP1 gene expression. In contrast, high levels of HWP1 …

Mutations Affecting A Putative Mutla Endonuclease Motif Impact Multiple Dna Mismatch Repair Functions, Naz Erdeniz, Megan Nguyen, Suzanne M. Deschênes, R. Michael Liskay

Biology Faculty Publications

Mutations in DNA mismatch repair (MMR) lead to increased mutation rates and higher recombination between similar, but not identical sequences, as well as resistance to certain DNA methylating agents. Recently, a component of human MMR machinery, MutLα, has been shown to display a latent endonuclease activity. The endonuclease active site appears to include a conserved motif, DQHA(X)₂E(X)₄E, within the COOH-terminus of human PMS2. Substitution of the glutamic acid residue (E705) abolished the endonuclease activity and mismatch-dependent excision in vitro. Previously, we showed that the PMS2-E705K mutation and the corresponding mutation in Saccharomyces cerevisiae were …

A Group M Consensus Envelope Glycoprotein Induces Antibodies That Neutralize Subsets Of Subtype B And C Hiv-1 Primary Viruses, Hua-Xin Liao, Laura L. Sutherland, Shi-Mao Xia, Mary E. Brock, Richard M. Scearce, Stacie Vanleeuwen, S. Munir Alam, Mildred Mcadams, Eric A. Weaver, Zenaido T. Camacho, Ben-Jiang Ma, Yingying Li, Julie M. Decker, Gary J. Nabel, David C. Montefiori, Beatrice H. Hahn, Bette T. Korber, Feng Gao, Barton F. Haynes

Nebraska Center for Virology: Faculty Publications

HIV-1 subtype C is the most common HIV-1 group M subtype in Africa and many parts of Asia. However, to date HIV-1 vaccine candidate immunogens have not induced potent and broadly neutralizing antibodies against subtype C primary isolates. We have used a centralized gene strategy to address HIV-1 diversity, and generated a group M consensus envelope gene with shortened consensus variable loops (CON-S) for comparative studies with wildtype (WT) Env immunogens. Our results indicate that the consensus HIV-1 group M CON-S Env elicited cross-subtype neutralizing antibodies of similar or greater breadth and titer than the WT Envs tested, indicating the …

Effects Of A Series Of Triorganotins On Atp Levels In Human Natural Killer Cells, Laurin N. Holloway, Keith H. Pannell, Margaret M. Whalen

Chemistry Faculty Research

Natural killer (NK) cells are our initial immune defense against viral infections and cancer development. Thus, agents that are able to interfere with their function increase the risk of cancer and/or infection. A series of triorganotins (trimethyltin (TMT), dimethylphenyltin (DMPT), methyldiphenyltin (MDPT), and triphenyltin (TPT)) have been shown to decrease the lytic function of human NK cells. TPT and MDPT were much more effective than DMPT or TMT at reducing lytic function. This study investigates the role that decreased ATP levels may play in decreases in the lytic function of NK cells induced by these organotins (OTs). A 24h exposure …

The Yeast Integral Membrane Protein Apq12 Potentially Links Membrane Dynamics To Assembly Of Nuclear Pore Complexes, John J. Scarcelli, Christin A. Hodge, Charles N. Cole

Dartmouth Scholarship

Although the structure and function of components of the nuclear pore complex (NPC) have been the focus of many studies, relatively little is known about NPC biogenesis. In this study, we report that Apq12 is required for efficient NPC biogenesis in Saccharomyces cerevisiae. Apq12 is an integral membrane protein of the nuclear envelope (NE) and endoplasmic reticulum. Cells lacking Apq12 are cold sensitive for growth, and a subset of their nucleoporins (Nups), those that are primarily components of the cytoplasmic fibrils of the NPC, mislocalize to the cytoplasm. APQ12 deletion also causes defects in NE morphology. In the absence of …

Vitamin E (Tocotrienols) And Prostate Cancer: A Proteomics Approach., Christian Mbangha Muenyi

Electronic Theses and Dissertations

Proteomics is the large scale study of proteins in cells or organisms. The purpose of this study was to characterize the proteomic alterations occurring in a prostate cancer (LNCaP) cell line after treatment with delta-tocotrienol (a form of vitamin E not very prevalent from most dietary sources). We found that both gamma- and delta-tocotrienols induced time and concentration dependent growth inhibition and programmed cell dead (apoptosis) in LNCaP cells. Secondly, we used two-dimensional gel electrophoresis (2-DE) to characterize changes in protein expression levels associated with this treatment. Our results show that a specific set of proteins are regulated at both …

Low Molecular Weight Protein Tyrosine Phosphatase (Lmw-Ptp) And Its Possible Physiological Functions Of Redox Signaling In The Eye Lens, Kuiyi Xing, Ashraf Raza, Stefan Lofgren, M. Rohan Fernando, Ye-Shih Ho, Marjorie F. Lou

School of Veterinary and Biomedical Sciences: Faculty Publications

Low molecular weight protein tyrosine phosphatase (LMW-PTP) was cloned from human lens epithelial B3 cells (HLE B3) and the recombinant enzyme was purified to homogeneity. The pure enzyme reacted positively with anti-LMW-PTP antibody, displayed tyrosine-specific phosphatase activity and was extremely sensitive to H2O2. The inactivated LMW-PTP could be regenerated by thioltransferase (TTase)/GSH system as demonstrated by both activity assay and by mass spectrometry (MS). The MS study also showed that an intramolecular disulfide bond was formed between C13 and C18 at the active site, and was reduced by the TTase/GSH system. The putative role of LMW-PTP in regulating platelet derived …

Maternal High-Salt Diet During Pregnancy In Sprague Dawley Rats Programs Exaggerated Stress-Induced Blood Pressure And Heart Rate Responses In Adult Female Offspring, Summer Hayes King

Theses and Dissertations

The prenatal environment has been shown to have lasting effects on cardiovascular health. In the present study, pregnant rats were fed a 0.7% NaCl normal salt (NS) diet or an 8% NaCl high salt (HS) diet throughout pregnancy. Adult offspring were fitted with radiotelemetry probes to continuously measure blood pressure and heart rate. Rats were placed in restraining cages to test for a programmed acute stress hyperresponsiveness. Offspring were challenged with HS diet for one week to determine if blood pressure salt sensitivity had been programmed by the prenatal HS diet. Animals were killed following resting and acute stress conditions, …

Effects Of Calcium Availability On The Reproductive Output Of Insectivorous Bats, Christina M. Booher

All-Inclusive List of Electronic Theses and Dissertations

Reproduction is a period of high calcium demand in vertebrates, and calcium deficiency can limit reproductive output in mammals. Insects are a poor calcium source, suggesting that insectivorous species are more likely to be calcium deficient. Pregnant big brown bats (Eptesicus fuscus) consume 10 times less calcium than they are estimated to require. To accommodate calcium demand during pregnancy, many mammals mobilize more calcium from the skeleton, and extensive bone loss has been observed in some bats during pregnancy. A conflict may arise between the female and developing embryo over allocation of limited calcium supplies, which could limit reproductive output …

The Effects Of Aging And Activity On Glial Cell Line-Derived Neurotrophic Factor Expression In Skeletal Muscle, Nathan Peplinski

Dissertations

Glial cell line-derived neurotrophic factor (GDNF) is an extracellular signaling protein that is produced by skeletal muscle and is important for the motor neurons that control muscle movement. GDNF has been shown to keep neurons alive under conditions that they would otherwise not persist. In skeletal muscle, GDNF has been shown to be one of the most potent neurotrophic factors that influence motor neuron survival. While the role of GDNF has been well studied during early development, not much is known about what happens to GDNF expression in the adult and with advanced aging. Previous results from our lab have …

Amp-Activated Protein Kinase Kinase Activity And Phosphorylation Of Amp-Activated Protein Kinase In Contracting Muscle Of Sedentary And Endurance Trained Rats, Denise Hurst

Theses and Dissertations

This study was designed to examine activity of AMP-activated protein kinase kinase (AMPKK) and AMP-activated protein kinase (AMPK) in muscles from control (C) and endurance trained (T) rats. Rats were trained 5 days/wk, 2 hr/d for 8 wks at a final intensity of 32 m/min up a 15% grade with 30 second sprints at 52 m/min every 10 min. Gastrocnemius muscles were stimulated in situ in T and C rats for 5 min at frequencies of 0.4/sec and 1/sec. Gastrocnemius LKB1 protein, a putative component of the AMPKK complex (LKB1, STRAD, and MO25), increased approximately 2-fold in response to training. …

Regulation Of Adrenal Steroidogenesis By Interleukin-6, Stephen A. Mcilmoil

Theses and Dissertations

Cortisol and dehydroepiandrosterone (DHEA) are steroids produced by the zona fasciculata (ZF) and reticularis (ZR), respectively, of the adrenal cortex. Both steroids are upregulated in response to adrenocorticotropic hormone (ACTH). Cortisol is a glucorticoid that is important in the regulation of inflammation and metabolism. DHEA is an adrenal androgen important in fetal growth and puberty but tends to decrease gradually after puberty in both men and women. DHEA has various effects on metabolism and immune function including inhibiting the effects of cortisol on some tissues. During the acute phase of stress, cortisol and DHEA rise due to an increase in …

Regulation Of Lkb1-Strad-Mo25 Complex Expression And Activation Of Ampk In Skeletal Muscle By Thyroid Hormone, Devon Jack Branvold

Theses and Dissertations

AMP-activated protein kinase (AMPK), a heterotrimeric protein which serves as a metabolic master switch in skeletal muscle, is a research target for the pharmaceutical treatment and prevention of type 2 diabetes. The expression of all of the isoforms of the subunits of AMPK and AMPK activity are increased in skeletal muscle tissue of hyperthyroid rats. Activity of AMPK is regulated by an upstream kinase (AMPKK). The LKB1-STRAD-MO25 complex is a major AMPKK in skeletal muscle. This experiment was designed to determine whether the increase in AMPK activity is accompanied by a thyroid hormone-induced increase in the expression of the LKB1-STRAD-MO25 …

Regulation Of Meiotic Cohesion And Chromosome Core Morphogenesis During Pachytene In Drosophila Oocytes, Radhika S. Khetani, Sharon E. Bickel

Dartmouth Scholarship

During meiosis, cohesion between sister chromatids is required for normal levels of homologous recombination, maintenance of chiasmata and accurate chromosome segregation during both divisions. In Drosophila, null mutations in the ord gene abolish meiotic cohesion, although how ORD protein promotes cohesion has remained elusive. We show that SMC subunits of the cohesin complex colocalize with ORD at centromeres of ovarian germ-line cells. In addition, cohesin SMCs and ORD are visible along the length of meiotic chromosomes during pachytene and remain associated with chromosome cores following DNase I digestion. In flies lacking ORD activity, cohesin SMCs fail to accumulate at oocyte …

Physiological Notch Signaling Promotes Gliogenesis In The Developing Peripheral And Central Nervous Systems, Merritt Taylor, Kelly Yeager, Sean J. Morrison

Peer Reviewed Articles

Constitutive activation of the Notch pathway can promote gliogenesis by peripheral (PNS) and central (CNS) nervous system progenitors. This raises the question of whether physiological Notch signaling regulates gliogenesis in vivo. To test this, we conditionally deleted Rbpsuh (Rbpj) from mouse PNS or CNS progenitors using Wnt1-Cre or Nestin-Cre. Rbpsuh encodes a DNAbinding protein (RBP/J) that is required for canonical signaling by all Notch receptors. In most regions of the developing PNS and spinal cord, Rbpsuh deletion caused only mild defects in neurogenesis, but severe defects in gliogenesis. These resulted from defects in glial specification or differentiation, …

Comparative Functional Analysis Of Aquaporins/Glyceroporins In Mammals And Anurans, Carissa M. Krane, David L. Goldstein

Biology Faculty Publications

Maintenance of fluid homeostasis is critical to establishing and maintaining normal physiology. The landmark discovery of membrane water channels (aquaporins; AQPs) ushered in a new area in osmoregulatory biology that has drawn from and contributed to diverse branches of biology, from molecular biology and genomics to systems biology and evolution, and from microbial and plant biology to animal and translational physiology. As a result, the study of AQPs provides a unique and integrated backdrop for exploring the relationships between genes and genome systems, the regulation of gene expression, and the physiologic consequences of genetic variation. The wide species distribution of …

Water. New Waters And New Life, Juan Enriquez

New England Journal of Public Policy

An excerpt from an article about life sciences taken from the proceedings of the Education for Public Inquiry and International Citizenship (EPIIC) Symposium held at Tufts University in Massachusetts in February 2005 is presented.

Somatostatin Receptor Subtype 2 Is Functionally Expressed In Raw264.7 Cells, Matthew Sung

Seton Hall University Dissertations and Theses (ETDs)

Chronic inflammation left unchecked can be quite harmful to the tissue with the pro-inflammatory stimulus. It is marked by the recruitment and activation of leukocytes, including lymphocytes and macrophages with their subsequent proliferation and reactive oxygen species release. Macrophages are also one of the primary players in propagating the inflammatory response as they secrete pro-inflammatory cytokines to sustain local tissue responses. Current therapies for chronic inflammation include non-steroidal anti­ inflammatory drugs and glucocorticoids; however, both have various side effects and setbacks. Somatostatin is an endogenous hormone which inhibits cellular secretion and proliferation throughout the body. Somatostatin receptor activation is mediated …

Characterization Of Molecular Events Following Heavy Metal Cadmium Exposure: A Cellular Regulatory Symphony In Response To Early Cadmium Insult, Chin-Ju Jean Hsiao

Dissertations

Cadmium, a non-essential toxic heavy metal, has been classified as a human carcinogen and one of the most hazardous substances in our environment. Since it can circulate within the food chain, has a long biological half-life and can bio-accumulate, it has the potential to induce detrimental health effects linked to toxicity, cancer and chronic diseases.

In recent years progress has been made to dissect and understand the molecular mechanisms underlying the effects of cadmium exposure. However, few studies have focused on investigating potential protective and/or defense mechanisms naturally applied by cells upon initial exposure, prior to any damage or toxic …

The E705k Mutation In Hpms2 Exerts Recessive, Not Dominant, Effects On Mismatch Repair, Suzanne M. Deschênes, Guy Tomer, Megan Nguyen, Naz Erdeniz, Nicole C. Juba, Natalia Sepúlveda, Jenna E. Pisani, R. Michael Liskay

Biology Faculty Publications

The hPMS2 mutation E705K is associated with Turcot syndrome. To elucidate the pathogenesis of hPMS2-E705K, we modeled this mutation in yeast and characterized its expression and effects on mutation avoidance in mammalian cells. We found that while hPMS2-E705K (pms1-E738K in yeast) did not significantly affect hPMS2 (Pms1p in yeast) stability or interaction with MLH1, it could not complement the mutator phenotype in MMR-deficient mouse or yeast cells. Furthermore, hPMS2-E705K/pms1-E738K inhibited MMR in wild-type (WT) mammalian cell extracts or yeast cells only when present in excess amounts relative to WT PMS2. Our results strongly suggest that hPMS2-E705K is a …

Stoichiometric Controls Of Mercury Dilution By Growth, Roxanne Karimi, Celia Y. Chen, Paul C. Pickhardt, Nicholas S. Fisher, Carol L. Folt

Dartmouth Scholarship

Rapid growth could significantly reduce methylmercury (MeHg) concentrations in aquatic organisms by causing a greater than proportional gain in biomass relative to MeHg (somatic growth dilution). We hypothesized that rapid growth from the consumption of high-quality algae, defined by algal nutrient stoichiometry, reduces MeHg concentrations in zooplankton, a major source of MeHg for lake fish. Using a MeHg radiotracer, we measured changes in MeHg concentrations, growth and ingestion rates in juvenile Daphnia pulex fed either high (C:P = 139) or low-quality (C:P = 1317) algae (Ankistrodesmus falcatus) for 5 d. We estimated Daphnia steady-state MeHg concentrations, using a …