Cell and Developmental Biology Commons^™

Never Let Me Clone? Countering An Ethical Argument Against The Reproductive Cloning Of Humans, Yvette Pearson

Philosophy Faculty Publications

In the March 2006 issue of EMBO reports, Christof Tannert, a bioethicist at the Max Delbrück Research Centre in Berlin, Germany, presented a moral argument against human reproductive cloning on the basis of Immanuel Kant’s categorical imperative (Tannert, 2006). In this article, I address some problems with Tannert’s views and show that our concerns about this prospective procedure should prompt us to scrutinize carefully the conventional procreative practices and attitudes. Indeed, if we set aside objections that are grounded in genetic determinism, many of the offensive features of human cloning are identical to problems with procreation by more conventional means, …

Chromosomal Localization Of The Islet Neogenesis Associated Protein (Ingap) Gene In Syrian Hamster By Tyramide Signal Amplification-Fluorescence In Situ Hybridization (Tsa-Fish), Sallie A. Smith

Biological Sciences Theses & Dissertations

Diabetes mellitus is a group of conditions characterized by hyperglycemia due to an inability to produce or properly utilize insulin. The majority of cases fall into two categories, Type I and Type 2. Type I results from the autoimmune destruction of pancreatic β-cells of the islets. The beta cells are the exclusive source of insulin and the patient becomes entirely dependent on exogenous insulin to survive. Patients with Type 2 are distinguished by insulin resistance, a condition that develops due to the inability of the body to effectively use the insulin being produced. The β-cells gradually lose their ability to …

Morphological And Molecular Defects In Zebrafish Embryos (Danio Rerio) After Chromated Copper Arsenate Exposure, Kerry Jean Lee

Biological Sciences Theses & Dissertations

This investigation identified the morphological defects caused by different concentrations of chromium, copper, and arsenic (CCA) on the developing zebrafish embryo; in addition to defects caused by the three combined components (complete CCA). A change in the DNA of the developing embryos in response to the three components of CCA and complete CCA was also determined. Early Dania rerio embryos were cultured for five days with various concentrations of CCA components and complete CCA. Morphological defects were assessed with light microscopy and DNA fragmentation was determined by agarose gel electrophoresis. Embryonic exposure of D. rerio to sub lethal concentrations of …

The Antitumor Agent, Arglabin-Dma, Preferentially Induces Apoptosis In Human Colon Tumor Cells, Sung Wook Kwon

Theses and Dissertations in Biomedical Sciences

Arglabin-DMA, an analog of farnesyl pyrophosphate (FPP), reportedly inhibits farnesyltransferase (FTase) directly by competitively blocking the binding of Ras protein and its posttranslational modification, as suggested in previous studies. But, the mechanisms by which Arglabin-DMA inhibits tumor growth in vivo and in vitro are still relatively poorly characterized. To determine the mechanism by which this drug inhibits tumor growth, the effects of Arglabin-DMA in two human colon tumor cell lines (mutant K-ras HCT 116 and wild-type ras HT-29) were explored on cell proliferation, apoptosis, and cell cycle kinetics in vitro. In cell viability studies, we showed that Arglabin-DMA …

Developing And Optimizing Conditions For Single Cell Genetic Analysis, Susan Adele Gitlin

Theses and Dissertations in Biomedical Sciences

Preimplantation Genetic Diagnosis (PGD) allows for couples to obtain genetic information at the embryo stage, therefore preventing lethal genetic diseases in their offspring. A recent multi-center study determined a 17% amplification failure rate overall and a 3.4% misdiagnosis rate (ESHRE PGD consortium steering committee, 2002). Experiments presented here were conducted to decrease those rates by optimizing single cell sensitivity and specificity of PGD techniques. Primer extension preamplification (PEP) was used after initial amplification failure, followed by gene specific PCR. Reamplification experiments yielded 54% amplification rates using blastomeres that previously failed amplification. In the clinical trial, four of six blastomeres were …

Nanosecond Pulsed Electric Fields: A New Stimulus To Activate Intracellular Signaling, Stephen J. Beebe, Karl H. Schoenbach

Bioelectrics Publications

When new technologies are introduced into the scientific community, controversy is expected and both excitement and disappointment enrich the lives of those who initiate the new ideas. It becomes the mission of the “inventors” to embrace the burden of proof to establish their ideas and convince the skeptics and disbelievers who will undoubtedly temper their enthusiasm and test their patience. While open mindedness is generally a scientific motto, those who review patents, manuscripts, and grants do not always readily practice it, even when the evidence is convincingly presented; old ideas and concepts often die hard. So it has been and …

Selective Field Effects On Intracellular Vacuoles And Vesicle Membranes With Nanosecond Electric Pulses, Ephrem Tekle, Hammou Oubrahim, Sergey M. Dzekunov, Juergen F. Kolb, Karl H. Schoenbach

Bioelectrics Publications

Electric pulses across intact vesicles and cells can lead to transient increase in permeability of their membranes. We studied the integrity of these membranes in response to external electric pulses of high amplitude and submicrosecond duration with a primary aim of achieving selective permeabilization. These effects were examined in two separate model systems comprising of 1), a mixed population of 1,2-di-oleoyl-sn-glycero-3-phosphocholine phospholipid vesicles and in 2), single COS-7 cells, in which large endosomal membrane vacuoles were induced by stimulated endocytosis. It has been shown that large and rapidly varying external electric fields, with pulses shorter than the charging time of …

Simulations Of Nanopore Formation And Phosphatidylserine Externalization In Lipid Membranes Subjected To A High-Intensity, Ultrashort Electric Pulse, Q. Hu, R. P. Joshi, K. H. Schoenbach

Bioelectrics Publications

A combined MD simulator and time dependent Laplace solver are used to analyze the electrically driven phosphatidylserine externalization process in cells. Time dependent details of nanopore formation at cell membranes in response to a high-intensity (100kV∕cm), ultrashort (10ns) electric pulse are also probed. Our results show that nanosized pores could typically be formed within about 5ns. These predictions are in very good agreement with recent experimental data. It is also demonstrated that defect formation and PS externalization in membranes should begin on the anode side. Finally, the simulations confirm that PS externalization is a nanopore facilitated event, rather than the …

Regulation Of Sparc Gene Expression By The Activator Protein 1 Transcription Factor, Joseph William Briggs

Theses and Dissertations in Biomedical Sciences

Overexpression of the c-Jun proto-oncogene in MCF7 breast cancer cells results in a variety of phenotypic changes related to malignant progression including a shift to estrogen independent growth, increased cell motility and invasion. Concurrent with these phenotypic changes are alterations to cellular gene expression patterns. One gene that becomes highly upregulated is SPARC (secreted protein acidic and rich in cysteine). Increased SPARC expression is associated with malignant progression in a variety of different cancers, although little is known regarding the mechanisms of SPARC gene regulation. Therefore, the objectives of this study were: (1) to determine the mechanisms by which c-Jun …

The Ultrastructure Of The Olfactory System In Two Species Of Short-Tailed Shrews, Blarina Brevicauda And Blarina Carolinensis, Lisa Johnson Byrum

Theses and Dissertations in Biomedical Sciences

Several studies of the fine structure of the olfactory system of rodents have been conducted, but very little research has been done on members of the Insectivora. The olfactory systems of the northern short-tailed shrew, Blarina brevicauda, and the southern short-tailed shrew, Blarina carolinensis, were examined by light and electron microscopy. These shrews were live trapped in the vicinity of Norfolk, Virginia throughout all months of the year. Olfactory tissues were processed following standard transmission and scanning electron microscopy protocols. The olfactory system structures investigated included the olfactory epitheliumlmucosa (OEM), main olfactory bulb (MOB), accessory olfactory bulb (AOB), …

Does Antarctic Krill Employ Body Shrinkage As An Overwintering Strategy?, Se-Jong Ju, H. Rodger Harvey, H. C. Shinn, Y. Kim, S. H. Kang

OES Faculty Publications

To determine if Antarctic krill employ body shrinkage as one of its overwintering mechanisms in the field, Euphausia superba and Euphausia crystallorophias were collected during fall and winter in and around Marguerite Bay through US Southern Ocean GLOBEC field programs during fall and winter 2001 and 2002. The relationships between the body length and weight of both krill species were exponentially correlated with no significant differences between the two species (p>0.05). The ratio between eye diameter and body length of individual krill was examined in an expectation that it could be used as an indicator of the body shrinkage …

Expression Of Glutathione-S-Transferase Mu 1 (Gstm1) And Glutathione-S-Transferase Theta 1 (Gstt1) In Patients With Aplastic Anemia And Myelodysplastic Syndromes, Aaron Conrad Spivey

Theses and Dissertations in Biomedical Sciences

Aplastic Anemia is a disorder of the hematopoietic bone marrow stem cells that is frequently associated with drug or chemical exposures. Reduced ability to detoxify drugs or chemicals may result in an increased risk of disease. Glutathione S-transferases are Phase II enzymes, which help detoxify xenobiotics. Many patients who have developed aplastic anemia and myelodysplastic syndrome have the null genotype for GSTMI and/or GSTTI. For patients with GSTMI and GSTTI positive genotypes, non-expression of these enzymes may cause an increased risk in developing aplastic anemia or myelodysplastic syndrome. To test whether genotype positive patients have GSTMI or GSTTI null …

The Quantitative Assessment Of Osteoinductivity Of Human Demineralized Bone Matrix And Cdna Array Analysis Of Osteogenic Differentiation In Human Periosteal Cells, Sittisak Honsawek

Theses and Dissertations in Biomedical Sciences

Demineralized bone matrix (DBM) has been extensively utilized in orthopeadic, periodontal, and maxillofacial applications. DBM comprises bone morphogenetic proteins (BMPs) that are essential regulators for endochondral bone formation. The current study was undertaken to establish the quantitative in vitro assays for determining the osteoinductive potential of human DBM and the use of cDNA gene expression array technology for examining gene expression profiles during osteoblast differentiation in human periosteal cells.

BMP-4 is one of the most osteoinductive factors and has been investigated for clinical applications. For this reason, BMP-4 was designated as the osteoinductive protein for use in the quantitative in …

Demographic Assessment Of The Blue Crab (Callinectes Sapidus) In Chesapeake Bay Using Extractable Lipofuscins As Age Markers, Se-Jong Ju, David H. Secor, H. Rodger Harvey

OES Faculty Publications

The blue crab (Callinectes sapidus) plays an important economic and ecological role in estuaries and coastal habitats from the Gulf of Mexico to the east coast of North America, but demographic assessments are limited by length-based methods. We applied an alternative aging method using biochemical measures of metabolic byproducts (lipofuscins) sequestered in the neural tissue of eyestalks to examine population age structure. From Chesapeake Bay, subsamples of animals collected from the 1998-99 (n-769) and 1999-2000 (n=367) winter dredge surveys were collected and lipofuscin was measured. Modal analysis of the lipofuscin index provided separation into three modes, whereas carapace-width …

Embryo And Gamete Development Upon Exposure To Cca Components: Cro3, Cuo, And As2o5, Nerbana Talaat-Elsebaei Mahmoud

Theses and Dissertations in Biomedical Sciences

Production of functional gametes and healthy embryos is essential for proliferation of all vertebrates, especially humans. Many compounds have toxic effects on developing gametes and embryos among which are chromium trioxide (CrO₃), cupric oxide (CuO) and arsenic pentaoxide (As₂O₅) as a mixture (CCA) or individually. Controversy surrounding the safety of CCA-treated wood centers primarily on the toxicity of its components and the potential for these metals to be released from the wood.

The aim of this research is to test the hypothesis that CCA components have deleterious effects on embryo development, oocyte maturation and …

Involvement Of Platelet-Derived Growth Factor, Pdgf Receptor Signaling, Focal Adhesion Kinase And Src In Pressure-Induced Vascular Smooth Muscle Hypertrophy, Darian Clark Rice

Theses and Dissertations in Biomedical Sciences

Elevated blood pressure is associated with varying degrees of arterial remodeling. The mechanisms by which extracellular mechanical stress is converted into intracellular alterations in signal transduction and gene expression have yet to be fully elucidated. Our goal was to investigate the early events in the vascular smooth muscle response to acute hypertension and to identify mediators involved in long-term hypertensive remodeling.

In the acute phase of hypertension we targeted cell surface integrin and growth factor receptors thought to be mechanically sensitive. The signaling molecules FAK, Src and ERK-MAPK are known to be triggered by integrin engagement and growth factor receptor …

Determination Of Nuclear Factor-Kappa B Activation In Cultured Renal Epithelial Cells And Cardiac Myocytes Exposed To Cocaine And Morphine, David Angeles Tiangco

Biological Sciences Theses & Dissertations

Nuclear factor-kappa B (NF-kB) is an important transcription factor that participates in the genetic regulation of inflammation in many tissues. Cocaine and heroin are drugs of abuse associated with renal and cardiac pathology. The purpose of this study was to test the hypothesis that exposure to cocaine or morphine (a metabolite of heroin) would activate NF-kB in renal epithelial cells (COS-7) and cardiac myocytes (H9c2) grown in culture. COS-7 and H9c2 cells were co-transfected with an experimental reporter specific for NF-kB activation and a control reporter with constitutive activity. A dual-luciferase assay was used to determine levels of NF-kB activation …

Prostate Specific Membrane Antigen (Psma): Immunoassay Development And Characterization Of Transcriptional Regulation, Zhen Xiao

Theses and Dissertations in Biomedical Sciences

Prostate cancer (PCA) is the most common cancer and the second leading cause of death among American men. The high mortality is greatly attributed to the lack of early detection tools and effective treatment for metastasis and relapses. Biomarkers that can discriminate benign from malignant tumor and signal the development of androgen independent and metastatic tumor are needed. A biomarker designated prostate specific membrane antigen (PSMA) has the potential to fulfill this need. The objective of this study is to develop a clinically useful immunoassay for quantitation of serum PSMA and to study the molecular mechanism underlying the upregulation of …

Improved Energy Model For Membrane Electroporation In Biological Cells Subjected To Electrical Pulses, R. P. Joshi, Q. Hu, K. H. Schoenbach, H. P. Hjalmarson

Bioelectrics Publications

A self-consistent model analysis of electroporation in biological cells has been carried out based on an improved energy model. The simple energy model used in the literature is somewhat incorrect and unphysical for a variety of reasons. Our model for the pore formation energy E(r) includes a dependence on pore population and density. It also allows for variable surface tension, incorporates the effects of finite conductivity on the electrostatic correction term, and is dynamic in nature. Self-consistent calculations, based on a coupled scheme involving the Smoluchowski equation and the improved energy model, are presented. It is shown that E(r) becomes …

Theoretical Predictions Of Electromechanical Deformation Of Cells Subjected To High Voltages For Membrane Electroporation, R. P. Joshi, Q. Hu, K. H. Schoenbach, H. P. Hjalmarson

Bioelectrics Publications

An electromechanical analysis based on thin-shell theory is presented to analyze cell shape changes in response to external electric fields. This approach can be extended to include osmotic-pressure changes. Our calculations demonstrate that at large fields, the spherical cell geometry can be significantly modified, and even ellipsoidal forms would be inappropriate to account for the deformation. Values of the surface forces obtained from our calculations are in very good agreement with the 1–10 mN/m range for membrane rupture reported in the literature. The results, in keeping with reports in the literature, demonstrate that the final shape depends on membrane thickness. …

Apoptosis Pathways: Presence And Significance In Ejaculated Human Spermatozoa, Steven Lewis Taylor

Theses and Dissertations in Biomedical Sciences

Ejaculated sperm display markers that are indicative of apoptosis in somatic cells. The question remains as to whether sperm have operative apoptosis mechanisms. The aim of this research was to test the hypothesis that apoptosis markers in sperm and somatic cells are different.

Ejaculated human sperm from patients and donors were separated into high and low motility fractions using Percoll™ gradients. Contaminating cells were removed using anti-CD45 conjugated paramagnetic beads. Fractions were divided into groups: staurosporine, anti-Fas antibody, and hydrogen peroxide treated and control. Direct enzymatic measurement of caspase activity, flow cytometric evaluation of phosphatidylserine translocation, immunoblots, and immunocytochemistry were …

Mechanism For Membrane Electroporation Irreversibility Under High-Intensity, Ultrashort Electrical Pulse Conditions, R. P. Joshi, K. H. Schoenbach

Bioelectrics Publications

An improved electroporation model is used to address membrane irreversibility under ultrashort electric pulse conditions. It is shown that membranes can survive a strong electric pulse and recover provided the pore distribution has a relatively large spread. If, however, the population consists predominantly of larger radii pores, then irreversibility can result. Physically, such a distribution could arise if pores at adjacent sites coalesce. The requirement of close proximity among the pore sites is more easily satisfied in smaller organelles than in outer cell membranes. Model predictions are in keeping with recent observations of cell damage to intracellular organelles (e.g., mitochondria), …

In Vivo Effect Of Leukemia Inhibitory Factor (Lif) And An Anti-Lif Polyclonal Antibody On Murine Embryo And Fetal Development Following Exposure At The Time Of Transcervical Blastocyst Transfer, Michael H. Mitchell, R. James Swanson, Sergio Oehninger

Biological Sciences Faculty Publications

Leukemia inhibitory factor (LIF) enhances in vitro murine preimplantation development in a time- and dose-dependent fashion. Knockout experiments have demonstrated that endometrial LIF is essential for in vivo murine implantation. We assessed the impact of LIF and an anti-LIF polyclonal antibody (pab) on in vivo development and developed a novel and successful nonsurgical method of embryo transfer for this species, a transcervical blastocyst transfer technique. The objectives of this study were to evaluate the effects of LIF and the anti-LIF pab on 1) implantation, resorption, pregnancy, and viability rates and 2) the overall structural and skeletal development. Two-cell embryos were …

Positive Regulation Of Pka On Human Gonadotropin-Releasing Hormone (Hgnrh) Gene Expression In Human Placental Jeg-3 Cells, Zhaoyang Wen

Chemistry & Biochemistry Theses & Dissertations

Using the human placental choriocarcinoma JEG-3 cell line as an in vitro human placental model, we studied the mechanisms of the PKA positive regulation of the hGnRH gene expression in the human placenta. Studies in JEG3 cells showed that through the PKA catalytic subunit a, human GnRH upstream promoter activity was stimulated by PKA signaling pathway in a cAMP dependent mechanism. The sequence between —202 (Afl II) and —554 (BamH I) base pair in the human GnRH upstream promoter region appeared to be responsible for the PKA positive regulation of the gene expression. Furthermore, Western blot analysis demonstrated the involvement …

Self-Consistent Simulations Of Electroporation Dynamics In Biological Cells Subjected To Ultrashort Electrical Pulses, R. P. Joshi, Q. Hu, R. Aly, K. H. Schoenbach, H. P. Hjalmarson

Bioelectrics Publications

The temporal dynamics of electroporation of cells subjected to ultrashort voltage pulses are studied based on a coupled scheme involving the Laplace, Nernst-Plank, and Smoluchowski equations. A pore radius dependent energy barrier for ionic transport, accounts for cellular variations. It is shown that a finite time delay exists in pore formation, and leads to a transient overshoot of the transmembrane potential V_mem beyond 1.0 V. Pore resealing is shown to consist of an initial fast process, a 10⁻⁴s delay, followed by a much slower closing at a time constant of about 10 ⁻¹s. This establishes a …

Growth Rate Variability And Lipofuscin Accumulation Rates In The Blue Crab Callinectes Sapidus, Se-Jong Ju, David H. Secor, H. Rodger Harvey

OES Faculty Publications

To better understand growth and age-pigment (lipofuscin) accumulation rates of the blue crab Callinectes sapidus under natural conditions, juveniles (33 to 94 mm carapace width) were reared in outdoor ponds for over 1 yr. Growth rates, measured by carapace width, during summer and fall exceeded all those reported in the literature; the initial carapace width of 59 ± 14 mm (mean ± SD) increased to 164 ± 15 mm within a 3 mo period. No growth occurred during winter months (November to April) at low water temperatures. Growth rates of crabs in ponds were substantially higher (von Bertalanffy growth parameter …

Characterization Of The Biological Activities Of Recombinant Fusion Protein Green Fluorescent Protein/Human Zona Pellucida Protein 3 (Gfp/Hzp3), Zhiyong Lin

Theses and Dissertations in Biomedical Sciences

Despite numerous reports indicating the successful production of bioactive recombinant ZP3, no report has shown the rhZP3 having direct binding activity with human sperm. Recombinant ZP3 generated from our previous study displayed binding activity with human sperm through indirect evidence from hemizona assay (HZA).

This present study focused on the production of recombinant ZP3 with direct binding activity with human sperm. Through the application of a pEGFP expression vector, fusion protein GFP/ZP3 was successfully generated and expressed. The expression of GFP/ZP3 was evidenced by RT-PCR and western blot. The fusion protein was partially purified by Ni-NTA affinity column from cell …

Induction Of Apoptosis In Human Prostate Cancer Cells By Resveratrol, Gary Zulfikar Morris

Chemistry & Biochemistry Theses & Dissertations

Recently attention has been brought to trans-resveratrol's {TR) anticancer activity, as determined through a number of cultured cancer cell models. This activity was attributed to TR behaving as an estrogen, and the orientation of TR' s hydroxyl groups. Based on this work it was of interest to determine whether TR would also be toxic in prostate cancer cells; if toxic, did TR induce necrosis or apoptosis in the cells; was it toxic through hormone mediated pathways; and were TR's hydroxyl groups responsible for its biological activity. To this end, cellular viability was assessed in two different prostate cancer cell …

An Ultrastructural Survey Of The Nuchal Glands Of Rhabdophis Tigrinus (Serpentes: Colubridae), Kathleen A. Roberts

Biological Sciences Theses & Dissertations

Rhabdophis tigrinus, an Asian natricine snake, possesses a series of integumentary glands located in the nuchal skin. Previous studies indicated that the nuchal glands are of mesodermal origin, contain cardiac steroids analogous to bufotoxins found in the parotoid glands of toads, which are included in the snake's diet. The nuchal glands are defensive in function. Preliminary histological studies revealed differences in quality of the gland tissue compared to previous studies, and difference in fixation techniques were suspected. Seven specimens of R. tigrinus were fixed by perfusion using a conservative fixation process standard for electron microscopy. The nuchal gland series …

Electroporation Dynamics In Biological Cells Subjected To Ultrafast Electrical Pulses: A Numerical Simulation Study, R. P. Joshi, K. H. Schoenbach

Bioelectrics Publications

A model analysis of electroporation dynamics in biological cells has been carried out based on the Smoluchowski equation. Results of the cellular response to short, electric pulses are presented, taking account of the growth and resealing dynamics of transient aqueous pores. It is shown that the application of large voltages alone may not be sufficient to cause irreversible breakdown, if the time duration is too short. Failure to cause irreversible damage at small pulse widths could be attributed to the time inadequacy for pores to grow and expand beyond a critical threshold radius. In agreement with earlier studies, it is …