Showing papers by "Kettering University published in 2004"
TL;DR: Conditions to direct hES cells into neurons of midbrain dopaminergic identity are described and high-yield DA neuron derivation was confirmed from three independent hES and two monkey embryonic stem cell lines.
Abstract: Human embryonic stem (hES) cells are defined by their extensive self-renewal capacity and their potential to differentiate into any cell type of the human body. The challenge in using hES cells for developmental biology and regenerative medicine has been to direct the wide differentiation potential toward the derivation of a specific cell fate. Within the nervous system, hES cells have been shown to differentiate in vitro into neural progenitor cells, neurons, and astrocytes. However, to our knowledge, the selective derivation of any given neuron subtype has not yet been demonstrated. Here, we describe conditions to direct hES cells into neurons of midbrain dopaminergic identity. Neuroectodermal differentiation was triggered on stromal feeder cells followed by regional specification by means of the sequential application of defined patterning molecules that direct in vivo midbrain development. Progression toward a midbrain dopamine (DA) neuron fate was monitored by the sequential expression of key transcription factors, including Pax2, Pax5, and engrailed-1 (En1), measurements of DA release, the presence of tetrodotoxin-sensitive action potentials, and the electron-microscopic visualization of tyrosinehydroxylase-positive synaptic terminals. High-yield DA neuron derivation was confirmed from three independent hES and two monkey embryonic stem cell lines. The availability of unlimited numbers of midbrain DA neurons is a first step toward exploring the potential of hES cells in preclinical models of Parkinson's disease. This experimental system also provides a powerful tool to probe the molecular mechanisms that control the development and function of human midbrain DA neurons.
1,072 citations
TL;DR: This work has mainly targeted the extraction of blood vessels, neurosvascular structure in particular, but has also reviewed some of the segmentation methods for the tubular objects that show similar characteristics to vessels.
Abstract: Vessel segmentation algorithms are the critical components of circulatory blood vessel analysis systems. We present a survey of vessel extraction techniques and algorithms. We put the various vessel extraction approaches and techniques in perspective by means of a classification of the existing research. While we have mainly targeted the extraction of blood vessels, neurosvascular structure in particular, we have also reviewed some of the segmentation methods for the tubular objects that show similar characteristics to vessels. We have divided vessel segmentation algorithms and techniques into six main categories: (1) pattern recognition techniques, (2) model-based approaches, (3) tracking-based approaches, (4) artificial intelligence-based approaches, (5) neural network-based approaches, and (6) tube-like object detection approaches. Some of these categories are further divided into subcategories. We have also created tables to compare the papers in each category against such criteria as dimensionality, input type, preprocessing, user interaction, and result type.
1,020 citations
TL;DR: Responses to open-ended questions on an exploratory survey indicate that students identify common themes in describing both temptations to cheat or to violate workplace policies and factors which caused them to hesitate in acting unethically, supporting the first hypothesis and laying the foundation for future surveys having forced-choice responses.
Abstract: Previous research indicates that students in engineering self-report cheating in college at higher rates than those in most other disciplines. Prior work also suggests that participation in one deviant behavior is a reasonable predictor of future deviant behavior. This combination of factors leads to a situation where engineering students who frequently participate in academic dishonesty are more likely to make unethical decisions in professional practice. To investigate this scenario, we propose the hypotheses that (1) there are similarities in the decision-making processes used by engineering students when considering whether or not to participate in academic and professional dishonesty, and (2) prior academic dishonesty by engineering students is an indicator of future decisions to act dishonestly. Our sample consisted of undergraduate engineering students from two technically-oriented private universities. As a group, the sample reported working full-time an average of six months per year as professionals in addition to attending classes during the remaining six months. This combination of both academic and professional experience provides a sample of students who are experienced in both settings. Responses to open-ended questions on an exploratory survey indicate that students identify common themes in describing both temptations to cheat or to violate workplace policies and factors which caused them to hesitate in acting unethically, thus supporting our first hypothesis and laying the foundation for future surveys having forced-choice responses. As indicated by the responses to forced-choice questions for the engineering students surveyed, there is a relationship between self-reported rates of cheating in high school and decisions to cheat in college and to violate workplace policies; supporting our second hypothesis. Thus, this exploratory study demonstrates connections between decision-making about both academic and professional dishonesty. If better understood, these connections could lead to practical approaches for encouraging ethical behavior in the academic setting, which might then influence future ethical decision-making in workplace settings.
278 citations
TL;DR: It is demonstrated that this histone fusion reporter allows the direct visualization of active chromatin in situ and permits the visualization of individual cells within a population, and so facilitates tracking cell position over time and is attractive for use in multidimensional studies of in vivo cell behavior and cell fate.
Abstract: Advances in optical imaging modalities and the continued evolution of genetically-encoded fluorescent proteins are coming together to facilitate the study of cell behavior at high resolution in living organisms. As a result, imaging using autofluorescent protein reporters is gaining popularity in mouse transgenic and targeted mutagenesis applications. We have used embryonic stem cell-mediated transgenesis to label cells at sub-cellular resolution in vivo, and to evaluate fusion of a human histone protein to green fluorescent protein for ubiquitous fluorescent labeling of nucleosomes in mice. To this end we have generated embryonic stem cells and a corresponding strain of mice that is viable and fertile and exhibits widespread chromatin-localized reporter expression. High levels of transgene expression are maintained in a constitutive manner. Viability and fertility of homozygous transgenic animals demonstrates that this reporter is developmentally neutral and does not interfere with mitosis or meiosis. Using various optical imaging modalities including wide-field, spinning disc confocal, and laser scanning confocal and multiphoton excitation microscopy, we can identify cells in various stages of the cell cycle. We can identify cells in interphase, cells undergoing mitosis or cell death. We demonstrate that this histone fusion reporter allows the direct visualization of active chromatin in situ. Since this reporter segments three-dimensional space, it permits the visualization of individual cells within a population, and so facilitates tracking cell position over time. It is therefore attractive for use in multidimensional studies of in vivo cell behavior and cell fate.
251 citations
TL;DR: The x-ray structure of the unphosphorylated form of the kinase catalytic domain of Syk is determined, suggesting the existence of two distinct Gleevec binding modes: an extended, trans-conformation characteristic of tight binding to the inactive conformation of a protein kinase and a second compact, cis-conform characteristic of weakerbinding to the active conformation.
196 citations
TL;DR: Biochemical characterization of the three SUMO isoforms into which an additional Gly-Gly di-peptide was inserted, or whereby the respective SUMO tails from the three isoforms were swapped, suggests a strict dependence for SUMO isopeptidase activity on residues C-terminal to the conserved Gly-gly motif and preferred cleavage site forsumO proteases.
183 citations
TL;DR: The first crystal structure of an RNA ligase suggests that contemporary DNA ligases, RNA ligases and RNA capping enzymes evolved by fusion of ancillary effector domains to an ancestral catalytic module involved in RNA repair.
167 citations
TL;DR: Mature DCs from 13 of 13 chronic HCV patients expressed typical maturation markers and were capable of priming allogeneic T lymphocytes, as well as stimulating influenza-specific memory T cells, consistent with clinical and immunologic data that the deficit in the patient's immune repertoire is HCV-specific.
156 citations
TL;DR: The enzyme-ligand structures, together with new mutational data, fully account for the biochemical specificity of the cap guanine-N7 methylation reaction, an essential and defining step of eukaryotic mRNA synthesis.
148 citations
TL;DR: The demographics and clinical features, the radiologic and pathologic features as well as the immunohistochemistry are discussed and this series of 17 breast GCT cases represent the largest series published to date.
Abstract: Seventeen cases of granular cell tumor (GCT) of the breast are reviewed. The demographics and clinical features are reviewed and the radiologic and pathologic features as well as the immunohistochemistry are discussed. To our knowledge, our series of 17 breast GCT cases represent the largest series published to date.
146 citations
TL;DR: A new approach, which is termed the "ynolide method", is directed to the synthesis of resorcinylic macrolides, including cycloproparadicicol and aigialomycin D, which were evaluated for their inhibitory activity against the growth of breast cancer cell line, MCF-7.
Abstract: A program currently ongoing in our laboratory envisions natural macrolide radicicol-based inhibitors targeting the molecular chaperone Hsp90. Such inhibitors can be potential anticancer agents due to their ability to induce the breakdown of a variety of oncogenic proteins. In this account, we first concern ourselves with a vastly important total synthesis of such an inhibitor. We accomplished this via a new approach, which we term the “ynolide method”, directed to the synthesis of resorcinylic macrolides, including cycloproparadicicol and aigialomycin D. The key features of the syntheses involve cobalt-complexation-promoted ring-closing metathesis (RCM) to generate ynolides, followed by Diels−Alder reaction with dimedone-derived bis-siloxy dienes to elaborate the benzo system. A number of interesting analogues were synthesized using this protocol. They were evaluated for their inhibitory activity against the growth of breast cancer cell line, MCF-7. The potency of their cytotoxicity was found to be consis...
TL;DR: The 1.9 A crystal structure of the ligase domain with AMP bound at the active site is reported, which reveals a shared fold, catalytic mechanism, and evolutionary history for RNA ligases, DNA ligase, and mRNA capping enzymes.
TL;DR: It was found that simplified analogues can retain the full potency of proteasome inhibition and some preliminary results from SAR studies are provided.
Abstract: A full account of the total syntheses of proteasome inhibitors TMC-95A and -B is provided. A key feature of the syntheses involved installation of a cis-propenylamide moiety by a thermal rearrangement of an α-silylallyl amide. The scope and mechanism of the enamide-forming reaction are discussed. Also provided are some preliminary results from SAR studies. It was found that simplified analogues can retain the full potency of proteasome inhibition.
TL;DR: It is reported that mice lacking functional ADAM19 exhibit severe defects in cardiac morphogenesis, including a ventricular septal defect (VSD), abnormal formation of the aortic and pulmonic valves, leading to valvular stenosis, and abnormalities of the cardiac vasculature.
Abstract: Congenital heart disease is the most common form of human birth defects, yet much remains to be learned about its underlying causes. Here we report that mice lacking functional ADAM19 (mnemonic for a disintegrin and metalloprotease 19) exhibit severe defects in cardiac morphogenesis, including a ventricular septal defect (VSD), abnormal formation of the aortic and pulmonic valves, leading to valvular stenosis, and abnormalities of the cardiac vasculature. During mouse development, ADAM19 is highly expressed in the conotruncus and the endocardial cushion, structures that give rise to the affected heart valves and the membranous ventricular septum. ADAM19 is also highly expressed in osteoblast-like cells in the bone, yet it does not appear to be essential for bone growth and skeletal development. Most adam19 / animals die perinatally, likely as a result of their cardiac defects. These findings raise the possibility that mutations in ADAM19 may contribute to human congenital heart valve and septal defects.
TL;DR: In this paper, the UEP approach is used for the construction of wavelet tight frames with two (anti-) symmetric wavelets, and some results and examples that complement recent results by Q. Jiang are provided.
TL;DR: Structural and biochemical analysis suggests an autoregulatory mechanism whereby premature decapping mRNA is prevented by blocking the conformational changes that are required to form a closed productive active site capable of cap hydrolysis.
TL;DR: It is shown that a member of the FK506-binding protein family, FKBP8, is an essential antagonist of SHH signaling in CNS development, suggesting that hedgehog signal transduction is subject to cell-type specific modulation during mammalian development.
Abstract: Sonic hedgehog (SHH) is a secreted morphogen that regulates the patterning and growth of many tissues in the developing mouse embryo, including the central nervous system (CNS). We show that a member of the FK506-binding protein family, FKBP8, is an essential antagonist of SHH signaling in CNS development. Loss of FKBP8 causes ectopic and ligand-independent activation of the Shh pathway, leading to expansion of ventral cell fates in the posterior neural tube and suppression of eye development. Although it is expressed broadly, FKBP8 is required to antagonize SHH signaling primarily in neural tissues, suggesting that hedgehog signal transduction is subject to cell-type specific modulation during mammalian development.
TL;DR: This work reports that bacteriophage T4 RNA ligase 2 (Rnl2) is an efficient catalyst of RNA ligation at a 3′-OH/5′-PO4 nick in a double-stranded RNA or an RNA·DNA hybrid and identifies 3 amino acids as essential for ligase activity.
Patent•
12 Aug 2004TL;DR: In this article, a method of treating cancer in a subject in need thereof, by administering to the patient a first amount of a histone deacetylase (HDAC) inhibitor or a pharmaceutically acceptable salt or hydrate thereof, in a first treatment procedure, and a second amount of an anti-cancer agent in a second treatment procedure.
Abstract: not available for EP1667680Abstract of corresponding document: WO2005023179The present invention relates to a method of treating cancer in a subject in need thereof, by administering to a subject in need thereof a first amount of a histone deacetylase (HDAC) inhibitor or a pharmaceutically acceptable salt or hydrate thereof, in a first treatment procedure, and a second amount of an anti-cancer agent in a second treatment procedure. The first and second amounts together comprise a therapeutically effective amount. The effect of the HDAC inhibitor and the anti-cancer agent may be additive or synergistic.
TL;DR: In obese patients, use of 31 G x 6 mm and 29 G x 12.7 mm needles produced comparable HbA(1c) values, double-blind pain and leakage scores, convenience, and ease of use, however, patients preferred the shorter needle.
TL;DR: The Kit receptor functions in hematopoiesis, lymphocyte development, gastrointestinal tract motility, melanogenesis, and gametogenesis and Kit-mediated PI 3-kinase signaling and Src kinase family signaling is highly specific for different cellular contexts in vivo.
Abstract: The Kit receptor functions in hematopoiesis, lymphocyte development, gastrointestinal tract motility, melanogenesis, and gametogenesis. To investigate the roles of different Kit signaling pathways in vivo, we have generated knock-in mice in which docking sites for PI 3-kinase (KitY719) or Src kinase (KitY567) have been mutated. Whereas steady-state hematopoiesis is normal in KitY719F/Y719F and KitY567F/Y567F mice, lymphopoiesis is affected differentially. The KitY567F mutation, but not the KitY719F mutation, blocks pro T cell and pro B cell development in an age-dependent manner. Thus, the Src family kinase, but not the PI 3-kinase docking site in Kit, mediates a critical signal for lymphocyte development. In agreement with these results, treatment of normal mice with the Kit tyrosine kinase inhibitor imatinib (Gleevec®) leads to deficits in pro T and pro B cell development, similar to those seen in KitY567F/Y567F and KitW/W mice. The two mutations do not affect embryonic gametogenesis but the KitY719F mutation blocks spermatogenesis at the spermatogonial stages and in contrast the KitY567F mutation does not affect this process. Therefore, Kit-mediated PI 3-kinase signaling and Src kinase family signaling is highly specific for different cellular contexts in vivo.
TL;DR: Rnl2 efficiently seals 3'-OH/5'-PO4 RNA nicks in either a duplex RNA or an RNA:DNA hybrid but cannot seal DNA nicks, providing insights to the evolution of nucleic acid repair systems.
TL;DR: This study demonstrates that AMN is an essential component of the Cubn receptor complex in vivo and suggests that Amn/Cubn is required for endocytosis/transcytosis of one or more ligands in the VE during gastrulation to coordinate growth and patterning of the embryo.
Abstract: Impaired primitive streak assembly in the mouse amnionless (amn) mutant results in the absence of non-axial trunk mesoderm, a derivative of the middle region of the primitive streak. In addition, the epiblast of amn mutants fails to increase significantly in size after E7.0, indicating that middle primitive streak assembly is mechanistically tied to the growth of the embryo during gastrulation. Amn, a novel transmembrane protein, is expressed exclusively in an extra-embryonic tissue, visceral endoderm (VE), during the early post-implantation stages. We show that Amn is also expressed in kidney proximal tubules (KPT) and intestinal epithelium, which, like the VE, are polarized epithelia specialized for resorption and secretion. To explore whether Amn participates in the development or function of KPT and intestinal epithelia and to gain insight into the function of Amn during gastrulation, we constructed Amn(-/-) ES cell +/+ blastocyst chimeras. While chimeras form anatomically normal kidneys and intestine, they exhibit variable, selective proteinuria, a sign of KPT malfunction. In humans, AMN has been genetically connected to Cubilin (CUBN), a multi-ligand scavenger receptor expressed by KPT, intestine and yolk sac. Loss of CUBN, the intestinal intrinsic factor (IF)-vitamin B12 receptor, results in hereditary megaloblastic anemia (MGA1), owing to vitamin B12 malabsorption. The recent report of MGA1 families with mutations in AMN suggests that AMN functions in the same pathway as CUBN. We demonstrate that Cubn is not properly localized to the cell surface in Amn(-/-) tissues in the embryo and adult mouse, and that adult chimeras exhibit selective proteinuria of Cubn ligands. This study demonstrates that Amn is an essential component of the Cubn receptor complex in vivo and suggests that Amn/Cubn is required for endocytosis/transcytosis of one or more ligands in the VE during gastrulation to coordinate growth and patterning of the embryo. Furthermore, as AMN is apparently not required for gastrulation in humans, the developmental requirements for Amn/Cubn function may not be evolutionarily conserved, possibly reflecting differences between species in the role and organization of extra-embryonic tissues.
TL;DR: Chemical syntheses are reported for prostate specific antigen (PSA) N-linked glycopeptide fragments consisting of an uneicosapeptide with di-, tri-, and tetrabranched N-acetyllactosamine-type glycans that will be used for the generation of antibodies that may form the basis for a new prostate cancer diagnostic assay.
Abstract: Chemical syntheses are reported for prostate specific antigen (PSA) N-linked glycopeptide fragments consisting of an uneicosapeptide (residues 27-47 of PSA) with di-, tri-, and tetrabranched N-acetyllactosamine-type glycans. The syntheses involve simultaneous, multiple glycosylations of the corresponding pentasaccharide acceptors prepared from a common trisaccharide precursor. Globally deprotected glycans are aminated and then aspartylated with a hexapeptide, which is then extended using native chemical ligation (NCL). The glycopeptides will be used for the generation of antibodies that may form the basis for a new prostate cancer diagnostic assay.
TL;DR: The first crystal structure of a member of this novel GTPase family, IIGP1, in its nucleotide-free, GDP-, and GppNHp-bound form is presented and it is shown that the dimer is required for cooperative GTP hydrolysis and GTP-dependent oligomerization of II GP1.
TL;DR: PapA5 represents the first structure solved for a protein involved in polyketide synthesis in Mycobacteria and shares unexpected similarity to structures of chloramphenicol acetyl transferase, dihydrolipoyl transacetylase, carnitine acetyltransferase, and VibH, a non-ribosomal peptide synthesis condensation enzyme.
TL;DR: Systemic AL amyloidosis with NHL has distinctive clinical features but employs the same Ig VL gene repertoire as AL with clonal plasma cell dyscrasias, reflecting previously observed correlations between germline gene use and organ tropism.
Abstract: Systemic AL amyloidosis (AL) is a disorder in which light chains form fibrillar deposits, leading to organ dysfunction and death. Rarely, AL has been associated with non-Hodgkin's lymphoma (NHL), although this association has not been well characterized. We report a series of six patients with AL associated with NHL, primarily lymphoplasmacytic lymphoma. Organ involvement was variable, with frequent bulky lymphadenopathy and visceral cavity deposits, but no cardiac involvement. Positron emission tomography scans were negative. Bone marrow and lymph node biopsies showed a mixed population of CD20+ lymphoid and CD138+ plasma cells. Serum free light chains were elevated, and correlated with response to therapy. Immunoglobulin light chain variable region (Ig VL) germline gene use was typical for AL, reflecting previously observed correlations between germline gene use and organ tropism. Five patients received rituximab-based therapies with two responses. Two patients underwent autologous stem cell transplantation with one complete haematological response. Four patients survive at 10-132 months from diagnosis. AL with NHL has distinctive clinical features but employs the same Ig VL gene repertoire as AL with clonal plasma cell dyscrasias. Serial serum free light chain levels are useful for tracking response to therapy. Treatments aimed at both lymphoid and plasma cell components appear warranted.