Gap Junctions

PubMed Central

Nielsen, Morten Schak; Axelsen, Lene Nygaard; Sorgen, Paul L.; Verma, Vandana; Delmar, Mario; Holstein-Rathlou, Niels-Henrik

2013-01-01

Gap junctions are essential to the function of multicellular animals, which require a high degree of coordination between cells. In vertebrates, gap junctions comprise connexins and currently 21 connexins are known in humans. The functions of gap junctions are highly diverse and include exchange of metabolites and electrical signals between cells, as well as functions, which are apparently unrelated to intercellular communication. Given the diversity of gap junction physiology, regulation of gap junction activity is complex. The structure of the various connexins is known to some extent; and structural rearrangements and intramolecular interactions are important for regulation of channel function. Intercellular coupling is further regulated by the number and activity of channels present in gap junctional plaques. The number of connexins in cell-cell channels is regulated by controlling transcription, translation, trafficking, and degradation; and all of these processes are under strict control. Once in the membrane, channel activity is determined by the conductive properties of the connexin involved, which can be regulated by voltage and chemical gating, as well as a large number of posttranslational modifications. The aim of the present article is to review our current knowledge on the structure, regulation, function, and pharmacology of gap junctions. This will be supported by examples of how different connexins and their regulation act in concert to achieve appropriate physiological control, and how disturbances of connexin function can lead to disease. © 2012 American Physiological Society. Compr Physiol 2:1981-2035, 2012. PMID:23723031

The enteric microbiota regulates jejunal Paneth cell number and function without impacting intestinal stem cells.

PubMed

Schoenborn, Alexi A; von Furstenberg, Richard J; Valsaraj, Smrithi; Hussain, Farah S; Stein, Molly; Shanahan, Michael T; Henning, Susan J; Gulati, Ajay S

2018-06-08

Paneth cells (PCs) are epithelial cells found in the small intestine, next to intestinal stem cells (ISCs) at the base of the crypts. PCs secrete antimicrobial peptides (AMPs) that regulate the commensal gut microbiota. In contrast, little is known regarding how the enteric microbiota reciprocally influences PC function. In this study, we sought to characterize the impact of the enteric microbiota on PC biology in the mouse small intestine. This was done by first enumerating jejunal PCs in germ-free (GF) versus conventionally-raised (CR) mice. We next evaluated the possible functional consequences of altered PC biology in these experimental groups by assessing epithelial proliferation, ISC numbers, and the production of AMPs. We found that PC numbers were significantly increased in CR versus GF mice; however, there were no differences in ISC numbers or cycling activity between groups. Of the AMPs assessed, only Reg3γ transcript expression was significantly increased in CR mice. Intriguingly, this increase was abrogated in cultured CR versus GF enteroids, and could not be re-induced with various bacterial ligands. Our findings demonstrate the enteric microbiota regulates PC function by increasing PC numbers and inducing Reg3γ expression, though the latter effect may not involve direct interactions between bacteria and the intestinal epithelium. In contrast, the enteric microbiota does not appear to regulate jejunal ISC census and proliferation. These are critical findings for investigators using GF mice and the enteroid system to study PC and ISC biology.

Integrating physiological regulation with stem cell and tissue homeostasis

PubMed Central

Nakada, Daisuke; Levi, Boaz P.; Morrison, Sean J.

2015-01-01

Summary Stem cells are uniquely able to self-renew, to undergo multilineage differentiation, and to persist throughout life in a number of tissues. Stem cells are regulated by a combination of shared and tissue-specific mechanisms and are distinguished from restricted progenitors by differences in transcriptional and epigenetic regulation. Emerging evidence suggests that other aspects of cellular physiology, including mitosis, signal transduction, and metabolic regulation also differ between stem cells and their progeny. These differences may allow stem cells to be regulated independently of differentiated cells in response to circadian rhythms, changes in metabolism, diet, exercise, mating, aging, infection, and disease. This allows stem cells to sustain homeostasis or to remodel relevant tissues in response to physiological change. Stem cells are therefore not only regulated by short-range signals that maintain homeostasis within their tissue of origin, but also by long-range signals that integrate stem cell function with systemic physiology. PMID:21609826

Hormonal Regulation of Dendritic Cell Differentiation in the Thymus.

PubMed

Shirshev, S V; Orlova, E G; Loginova, O A; Nekrasova, I V; Gorbunova, O L; Maslennikova, I L

2018-06-19

We studied the effect of hormones estriol, ghrelin, kisspeptin, and chorionic gonadotropin in concentrations corresponding to their content in the peripheral blood in each trimester of pregnancy on the expression of membrane molecules on myeloid and plasmacytoid dendritic cells of the thymus. It was found that thymic myeloid dendritic cells are sensitive to the action of estriol and kisspeptin. Estriol in a concentration of the first trimester of pregnancy reduces the number of myeloid dendritic cells expressing receptor for thymic stromal lymphopoietin (CD11c+TSLP-R + ) and inhibitory molecule B7-H3 (CD11c + CD276 + ). In contrast to estriol, kisspeptin regulates the processes of differentiation of thymic myeloid dendritic cells in concentrations typical of the second-third trimesters and reduced their total number (CD11c + ) and the number of cells expressing TSLP-R (CD11c + TSLP-R + ). Estriol and kisspeptin do not affect the total number of plasmacytoid dendritic cells (CD303 + ) and expression of TSLP-R and CD276 by these cells. Ghrelin and chorionic gonadotropin in the studied concentrations had no significant effect on the total number of thymic myeloid and plasmacytoid dendritic cells and on the expression of membrane molecules of TSLP-R and CD276.

Regulation of naturally occurring autoantibody secretion by a radiosensitive lymphocyte: initial characterization and ontogeny

DOE Office of Scientific and Technical Information (OSTI.GOV)

Moticka, E.J.

1983-10-01

Low level irradiation (400-500 R) of normal mice or of murine spleen cells resulted in the detection of an enhanced number of plaque-forming cells against bromelain-treated autologous red cells (Br MRBC) 1 day later. The mechanism responsible for the increased numbers of plaques is apparently the elimination of a suppressor T cell since the addition of thymocytes or of Lyt 1+2+ splenic cells to cultures of irradiated cells reversed the radiation-induced increase. Studies on the ontogeny of the phenomenon indicate that anti-Br MRBC plaques can be formed by spleen cells taken from mice shortly after birth although adult levels aremore » not reached until after 3 weeks of age. Radiation-induced increases in the number of plaques were not seen until 3 weeks of age, thus, suggesting a temporal developmental sequence of the ability to produce autoantibodies and to regulate such production.« less

Msi2 Regulates the Aggressiveness of Non Small Cell Lung Cancer (NSCLC)

DTIC Science & Technology

2016-10-01

AWARD NUMBER: W81XWH-15-1-0192 TITLE: Msi2 Regulates the Aggressiveness of Non -Small Cell Lung Cancer (NSCLC) PRINCIPAL INVESTIGATOR: Yanis...Annual 3. DATES COVERED (From - To) 15 Sep 2015 - 14 Sep 2016 4. TITLE AND SUBTITLE Msi2 Regulates the Aggressiveness of Non -Small Cell Lung Cancer...in vitro and in vivo are ongoing, while immunohistochemistry studies are starting Fall 2016. 15. SUBJECT TERMS Non -small cell lung cancer

APOPTOSIS DURING DEVELOPMENT AND AGING AND IN RESPONSE TO MERCURY EXPOSURE.

EPA Science Inventory

In the central nervous system from embryogenesis through senescence, cell number is regulated, in part, by apoptosis. Each region of the nervous system has a characteristic temporal pattern of programmed cell death, which includes far greater numbers of cells undergoing apop...

Regulation of DNA replication during development

PubMed Central

Nordman, Jared; Orr-Weaver, Terry L.

2012-01-01

As development unfolds, DNA replication is not only coordinated with cell proliferation, but is regulated uniquely in specific cell types and organs. This differential regulation of DNA synthesis requires crosstalk between DNA replication and differentiation. This dynamic aspect of DNA replication is highlighted by the finding that the distribution of replication origins varies between differentiated cell types and changes with differentiation. Moreover, differential DNA replication in some cell types can lead to increases or decreases in gene copy number along chromosomes. This review highlights the recent advances and technologies that have provided us with new insights into the developmental regulation of DNA replication. PMID:22223677

Oncogenic KRAS Regulates Tumor Cell Signaling via Stromal Reciprocation

PubMed Central

Tape, Christopher J.; Ling, Stephanie; Dimitriadi, Maria; McMahon, Kelly M.; Worboys, Jonathan D.; Leong, Hui Sun; Norrie, Ida C.; Miller, Crispin J.; Poulogiannis, George; Lauffenburger, Douglas A.; Jørgensen, Claus

2016-01-01

Summary Oncogenic mutations regulate signaling within both tumor cells and adjacent stromal cells. Here, we show that oncogenic KRAS (KRASG12D) also regulates tumor cell signaling via stromal cells. By combining cell-specific proteome labeling with multivariate phosphoproteomics, we analyzed heterocellular KRASG12D signaling in pancreatic ductal adenocarcinoma (PDA) cells. Tumor cell KRASG12D engages heterotypic fibroblasts, which subsequently instigate reciprocal signaling in the tumor cells. Reciprocal signaling employs additional kinases and doubles the number of regulated signaling nodes from cell-autonomous KRASG12D. Consequently, reciprocal KRASG12D produces a tumor cell phosphoproteome and total proteome that is distinct from cell-autonomous KRASG12D alone. Reciprocal signaling regulates tumor cell proliferation and apoptosis and increases mitochondrial capacity via an IGF1R/AXL-AKT axis. These results demonstrate that oncogene signaling should be viewed as a heterocellular process and that our existing cell-autonomous perspective underrepresents the extent of oncogene signaling in cancer. Video Abstract PMID:27087446

Identifying microRNAs that Regulate Neuroblastoma Cell Differentiation

DTIC Science & Technology

2015-10-01

Award Number: W81XWH-13-1-0241 TITLE: Identifying that Regulate Neuroblastoma Cell Differentiation PRINCIPAL INVESTIGATOR: Dr. Liqin Du...inducing miRNA, miR- 449a. We examined the differentiation-inducing function of miR-449a in multiple neuroblastoma cell lines. We have demonstrated that...miR-449a functions as an inducer of cell differentiation in neuroblastoma cell lines with distinct genetic backgrounds, including the MYCN

Notch pathway signaling in the skin antagonizes Merkel cell development.

PubMed

Logan, Gregory J; Wright, Margaret C; Kubicki, Adam C; Maricich, Stephen M

2018-02-15

Merkel cells are mechanosensitive skin cells derived from the epidermal lineage whose development requires expression of the basic helix-loop-helix transcription factor Atoh1. The genes and pathways involved in regulating Merkel cell development during embryogenesis are poorly understood. Notch pathway signaling antagonizes Atoh1 expression in many developing body regions, so we hypothesized that Notch signaling might inhibit Merkel cell development. We found that conditional, constitutive overexpression of the Notch intracellular domain (NICD) in mouse epidermis significantly decreased Merkel cell numbers in whisker follicles and touch domes of hairy skin. Conversely, conditional deletion of the obligate NICD binding partner RBPj in the epidermis significantly increased Merkel cell numbers in whisker follicles, led to the development of ectopic Merkel cells outside of touch domes in hairy skin epidermis, and altered the distribution of Merkel cells in touch domes. Deletion of the downstream Notch effector gene Hes1 also significantly increased Merkel cell numbers in whisker follicles. Together, these data demonstrate that Notch signaling regulates Merkel cell production and patterning. Copyright © 2018 Elsevier Inc. All rights reserved.

Successful In Vitro Expansion and Differentiation of Cord Blood Derived CD34+ Cells into Early Endothelial Progenitor Cells Reveals Highly Differential Gene Expression

PubMed Central

Topcic, Denijal; Haviv, Izhak; Merivirta, Ruusu-Maaria; Agrotis, Alexander; Leitner, Ephraem; Jowett, Jeremy B.; Bode, Christoph; Lappas, Martha; Peter, Karlheinz

2011-01-01

Endothelial progenitor cells (EPCs) can be purified from peripheral blood, bone marrow or cord blood and are typically defined by a limited number of cell surface markers and a few functional tests. A detailed in vitro characterization is often restricted by the low cell numbers of circulating EPCs. Therefore in vitro culturing and expansion methods are applied, which allow at least distinguishing two different types of EPCs, early and late EPCs. Herein, we describe an in vitro culture technique with the aim to generate high numbers of phenotypically, functionally and genetically defined early EPCs from human cord blood. Characterization of EPCs was done by flow cytometry, immunofluorescence microscopy, colony forming unit (CFU) assay and endothelial tube formation assay. There was an average 48-fold increase in EPC numbers. EPCs expressed VEGFR-2, CD144, CD18, and CD61, and were positive for acetylated LDL uptake and ulex lectin binding. The cells stimulated endothelial tube formation only in co-cultures with mature endothelial cells and formed CFUs. Microarray analysis revealed highly up-regulated genes, including LL-37 (CAMP), PDK4, and alpha-2-macroglobulin. In addition, genes known to be associated with cardioprotective (GDF15) or pro-angiogenic (galectin-3) properties were also significantly up-regulated after a 72 h differentiation period on fibronectin. We present a novel method that allows to generate high numbers of phenotypically, functionally and genetically characterized early EPCs. Furthermore, we identified several genes newly linked to EPC differentiation, among them LL-37 (CAMP) was the most up-regulated gene. PMID:21858032

Hyaluronan as an Immune Regulator in Human Diseases

PubMed Central

NOBLE, PAUL W.; LIANG, JIURONG; JIANG, DIANHUA

2010-01-01

Accumulation and turnover of extracellular matrix components are the hallmarks of tissue injury. Fragmented hyaluronan stimulates the expression of inflammatory genes by a variety of immune cells at the injury site. Hyaluronan binds to a number of cell surface proteins on a variety of cell types. Hyaluronan fragments signal through both Toll-like receptor (TLR) 4 and TLR2 as well as CD44 to stimulate inflammatory genes in inflammatory cells. Hyaluronan is also present on the cell surface of epithelial cells and provides protection against tissue damage by interacting with TLR2 and TLR4 on these parenchymal cells. Hyaluronan and hyaluronan-binding proteins regulate inflammation, tissue injury and repair through regulating inflammatory cell recruitment, release of inflammatory cytokines, and stem cell migration. This review focuses on the role of hyaluronan as an immune regulator in human diseases. PMID:21248167

The regulation of tooth morphogenesis is associated with epithelial cell proliferation and the expression of Sonic hedgehog through epithelial-mesenchymal interactions

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ishida, Kentaro; Murofushi, Mayumi; Nakao, Kazuhisa

2011-02-18

Research highlights: {yields} Bioengineered teeth regulated the contact area of epithelium and mesenchyme. {yields} The crown width is regulated by the contact area of the epithelium and mesenchyme. {yields} This regulation is associated with cell proliferation and Sonic hedgehog expression. {yields} The cusp number is correlated with the crown width of the bioengineered tooth. {yields} Cell proliferation and Shh expression areas regulate the tooth morphogenesis. -- Abstract: Ectodermal organs, such as the tooth, salivary gland, hair, and mammary gland, develop through reciprocal epithelial-mesenchymal interactions. Tooth morphologies are defined by the crown width and tooth length (macro-morphologies), and by the numbermore » and locations of the cusp and roots (micro-morphologies). In our current study, we report that the crown width of a bioengineered molar tooth, which was reconstructed using dissociated epithelial and mesenchymal cells via an organ germ method, can be regulated by the contact area between epithelial and mesenchymal cell layers. We further show that this is associated with cell proliferation and Sonic hedgehog (Shh) expression in the inner enamel epithelium after the germ stage has formed a secondary enamel knot. We also demonstrate that the cusp number is significantly correlated with the crown width of the bioengineered tooth. These findings suggest that the tooth micro-morphology, i.e. the cusp formation, is regulated after the tooth width, or macro-morphology, is determined. These findings also suggest that the spatiotemporal patterning of cell proliferation and the Shh expression areas in the epithelium regulate the crown width and cusp formation of the developing tooth.« less

A novel IL-10-independent regulatory role for B cells in suppressing autoimmunity by maintenance of regulatory T cells via GITR ligand.

PubMed

Ray, Avijit; Basu, Sreemanti; Williams, Calvin B; Salzman, Nita H; Dittel, Bonnie N

2012-04-01

B cells are important for the regulation of autoimmune responses. In experimental autoimmune encephalomyelitis (EAE), B cells are required for spontaneous recovery in acute models. Production of IL-10 by regulatory B cells has been shown to modulate the severity EAE and other autoimmune diseases. Previously, we suggested that B cells regulated the number of CD4(+)Foxp3(+) T regulatory cells (Treg) in the CNS during EAE. Because Treg suppress autoimmune responses, we asked whether B cells control autoimmunity by maintenance of Treg numbers. B cell deficiency achieved either genetically (μMT) or by depletion with anti-CD20 resulted in a significant reduction in the number of peripheral but not thymic Treg. Adoptive transfer of WT B cells into μMT mice restored both Treg numbers and recovery from EAE. When we investigated the mechanism whereby B cells induce the proliferation of Treg and EAE recovery, we found that glucocorticoid-induced TNF ligand, but not IL-10, expression by B cells was required. Of clinical significance is the finding that anti-CD20 depletion of B cells accelerated spontaneous EAE and colitis. Our results demonstrate that B cells play a major role in immune tolerance required for the prevention of autoimmunity by maintenance of Treg via their expression of glucocorticoid-induced TNFR ligand.

Testosterone regulates the autophagic clearance of androgen binding protein in rat Sertoli cells

PubMed Central

Ma, Yi; Yang, Hao-Zheng; Xu, Long-Mei; Huang, Yi-Ran; Dai, Hui-Li; Kang, Xiao-Nan

2015-01-01

Dysregulation of androgen-binding protein (ABP) is associated with a number of endocrine and andrology diseases. However, the ABP metabolism in Sertoli cells is largely unknown. We report that autophagy degrades ABP in rat Sertoli cells, and the autophagic clearance of ABP is regulated by testosterone, which prolongs the ABP biological half-life by inhibiting autophagy. Further studies identified that the autophagic clearance of ABP might be selectively regulated by testosterone, independent of stress (hypoxia)-induced autophagic degradation. These data demonstrate that testosterone up-regulates ABP expression at least partially by suppressing the autophagic degradation. We report a novel finding with respect to the mechanisms by which ABP is cleared, and by which the process is regulated in Sertoli cells. PMID:25745956

Sertoli cell androgen receptor expression regulates temporal fetal and adult Leydig cell differentiation, function, and population size.

PubMed

Hazra, Rasmani; Jimenez, Mark; Desai, Reena; Handelsman, David J; Allan, Charles M

2013-09-01

We recently created a mouse model displaying precocious Sertoli cell (SC) and spermatogenic development induced by SC-specific transgenic androgen receptor expression (TgSCAR). Here we reveal that TgSCAR regulates the development, function, and absolute number of Leydig cells (LCs). Total fetal and adult type LC numbers were reduced in postnatal and adult TgSCAR vs control testes, despite normal circulating LH levels. Normal LC to SC ratios found in TgSCAR testes indicate that SC androgen receptor (SCAR)-mediated activity confers a quorum-dependent relationship between total SC and LC numbers. TgSCAR enhanced LC differentiation, shown by elevated ratios of advanced to immature LC types, and reduced LC proliferation in postnatal TgSCAR vs control testes. Postnatal TgSCAR testes displayed up-regulated expression of coupled ligand-receptor transcripts (Amh-Amhr2, Dhh-Ptch1, Pdgfa-Pdgfra) for potential SCAR-stimulated paracrine pathways, which may coordinate LC differentiation. Neonatal TgSCAR testes displayed normal T and dihydrotestosterone levels despite differential changes to steroidogenic gene expression, with down-regulated Star, Cyp11a1, and Cyp17a1 expression contrasting with up-regulated Hsd3b1, Hsd17b3, and Srd5a1 expression. TgSCAR males also displayed elevated postnatal and normal adult serum testosterone levels, despite reduced LC numbers. Enhanced adult-type LC steroidogenic output was revealed by increased pubertal testicular T, dihydrotestosterone, 3α-diol and 3β-diol levels per LC and up-regulated steroidogenic gene (Nr5a1, Lhr, Cyp11a1, Cyp17a1, Hsd3b6, Srd5a1) expression in pubertal or adult TgSCAR vs control males, suggesting regulatory mechanisms maintain androgen levels independently of absolute LC numbers. Our unique gain-of-function TgSCAR model has revealed that SCAR activity controls temporal LC differentiation, steroidogenic function, and population size.

β-Catenin–regulated myeloid cell adhesion and migration determine wound healing

PubMed Central

Amini-Nik, Saeid; Cambridge, Elizabeth; Yu, Winston; Guo, Anne; Whetstone, Heather; Nadesan, Puviindran; Poon, Raymond; Hinz, Boris; Alman, Benjamin A.

2014-01-01

A β-catenin/T cell factor–dependent transcriptional program is critical during cutaneous wound repair for the regulation of scar size; however, the relative contribution of β-catenin activity and function in specific cell types in the granulation tissue during the healing process is unknown. Here, cell lineage tracing revealed that cells in which β-catenin is transcriptionally active express a gene profile that is characteristic of the myeloid lineage. Mice harboring a macrophage-specific deletion of the gene encoding β-catenin exhibited insufficient skin wound healing due to macrophage-specific defects in migration, adhesion to fibroblasts, and ability to produce TGF-β1. In irradiated mice, only macrophages expressing β-catenin were able to rescue wound-healing deficiency. Evaluation of scar tissue collected from patients with hypertrophic and normal scars revealed a correlation between the number of macrophages within the wound, β-catenin levels, and cellularity. Our data indicate that β-catenin regulates myeloid cell motility and adhesion and that β-catenin–mediated macrophage motility contributes to the number of mesenchymal cells and ultimate scar size following cutaneous injury. PMID:24837430

Bach2 Controls Homeostasis of Eosinophils by Restricting the Type-2 Helper Function of T Cells.

PubMed

Sato, Yuki; Kato, Hiroki; Ebina-Shibuya, Risa; Itoh-Nakadai, Ari; Okuyama, Ryuhei; Igarashi, Kazuhiko

2017-03-01

Bach2 is a transcription factor which represses its target genes and plays important roles in the differentiation of B and T lymphoid cells. Bach2-deficient (KO) mice develop severe pulmonary alveolar proteinosis, which is associated with increased numbers of granulocytes and T cells. Bach2 is essential for the regulation of T cells, but its role in the regulation of granulocytes is not clear. Here, we observed increased numbers of eosinophils but not neutrophils in the bone marrow, spleen, peripheral blood, and bronchoalveolar lavage fluids of Bach2 KO mice compared with those of wild-type (WT) mice. Upon co-transplantation of the bone marrow cells from CD45.2 Bach2 KO and CD45.1/CD45.2 double-positive WT mice to irradiated WT CD45.1/CD45.2 mice, the reconstituted numbers of eosinophils were similar between Bach2 KO and WT cells. These results showed that the deficiency of Bach2 in eosinophils did not directly drive the differentiation of eosinophils. To investigate the effect of Bach2 KO CD4 + T cells upon eosinophils, we analyzed Rag2/Bach2-double deficient (dKO) mice which lack lymphocytes including CD4 + T cells. Rag2/Bach2 dKO mice did not show any increase in the numbers of eosinophils. Importantly, Bach2 KO mice showed an increase of interleukin-5 (Il-5) in the sera compared with WT mice. These results suggest that up-regulated functions of CD4 + T cells including secretion of Il-5 resulted in proliferation and/or migration to peripheral tissues of eosinophils in Bach2 KO mice. We propose that Bach2 controls homeostasis of eosinophils via restricting the production of Il-5 in CD4 + T cells.

Substrate-specific regulation of ubiquitination by the anaphase-promoting complex

PubMed Central

Song, Ling

2011-01-01

By orchestrating the sequential degradation of a large number of cell cycle regulators, the ubiquitin ligase anaphase-promoting complex (APC/C) is essential for proliferation in all eukaryotes. The correct timing of APC/C-dependent substrate degradation, a critical feature of progression through mitosis, was long known to be controlled by mechanisms targeting the core APC/C-machinery. Recent experiments, however have revealed an important contribution of substrate-specific regulation of the APC/C to achieve accurate cell division. In this perspective, we describe different mechanisms of substrate-specific APC/C-regulation and discuss their importance for cell division. PMID:21191176

Interaction of Proteins Identified in Human Thyroid Cells

PubMed Central

Pietsch, Jessica; Riwaldt, Stefan; Bauer, Johann; Sickmann, Albert; Weber, Gerhard; Grosse, Jirka; Infanger, Manfred; Eilles, Christoph; Grimm, Daniela

2013-01-01

Influence of gravity forces on the regulation of protein expression by healthy and malignant thyroid cells was studied with the aim to identify protein interactions. Western blot analyses of a limited number of proteins suggested a time-dependent regulation of protein expression by simulated microgravity. After applying free flow isoelectric focusing and mass spectrometry to search for differently expressed proteins by thyroid cells exposed to simulated microgravity for three days, a considerable number of candidates for gravi-sensitive proteins were detected. In order to show how proteins sensitive to microgravity could directly influence other proteins, we investigated all polypeptide chains identified with Mascot scores above 100, looking for groups of interacting proteins. Hence, UniProtKB entry numbers of all detected proteins were entered into the Search Tool for the Retrieval of Interacting Genes/Proteins (STRING) and processed. The program indicated that we had detected various groups of interacting proteins in each of the three cell lines studied. The major groups of interacting proteins play a role in pathways of carbohydrate and protein metabolism, regulation of cell growth and cell membrane structuring. Analyzing these groups, networks of interaction could be established which show how a punctual influence of simulated microgravity may propagate via various members of interaction chains. PMID:23303277

The Endocytic Adaptor Eps15 Controls Marginal Zone B Cell Numbers

PubMed Central

Pozzi, Benedetta; Amodio, Stefania; Lucano, Caterina; Sciullo, Anna; Ronzoni, Simona; Castelletti, Daniela; Adler, Thure; Treise, Irina; Betsholtz, Ingrid Holmberg; Rathkolb, Birgit; Busch, Dirk H.; Wolf, Eckhard; Fuchs, Helmut; Gailus-Durner, Valérie; de Angelis, Martin Hrabě; Betsholtz, Christer; Casola, Stefano; Di Fiore, Pier Paolo; Offenhäuser, Nina

2012-01-01

Eps15 is an endocytic adaptor protein involved in clathrin and non-clathrin mediated endocytosis. In Caenorhabditis elegans and Drosophila melanogaster lack of Eps15 leads to defects in synaptic vesicle recycling and synapse formation. We generated Eps15-KO mice to investigate its function in mammals. Eps15-KO mice are born at the expected Mendelian ratio and are fertile. Using a large-scale phenotype screen covering more than 300 parameters correlated to human disease, we found that Eps15-KO mice did not show any sign of disease or neural deficits. Instead, altered blood parameters pointed to an immunological defect. By competitive bone marrow transplantation we demonstrated that Eps15-KO hematopoietic precursor cells were more efficient than the WT counterparts in repopulating B220+ bone marrow cells, CD19− thymocytes and splenic marginal zone (MZ) B cells. Eps15-KO mice showed a 2-fold increase in MZ B cell numbers when compared with controls. Using reverse bone marrow transplantation, we found that Eps15 regulates MZ B cell numbers in a cell autonomous manner. FACS analysis showed that although MZ B cells were increased in Eps15-KO mice, transitional and pre-MZ B cell numbers were unaffected. The increase in MZ B cell numbers in Eps15 KO mice was not dependent on altered BCR signaling or Notch activity. In conclusion, in mammals, the endocytic adaptor protein Eps15 is a regulator of B-cell lymphopoiesis. PMID:23226392

microRNAs as regulators of adipogenic differentiation of mesenchymal stem cells.

PubMed

Hamam, Dana; Ali, Dalia; Kassem, Moustapha; Aldahmash, Abdullah; Alajez, Nehad M

2015-02-15

microRNAs (miRNAs) constitute complex regulatory network, fine tuning the expression of a myriad of genes involved in different biological and physiological processes, including stem cell differentiation. Mesenchymal stem cells (MSCs) are multipotent stem cells present in the bone marrow stroma, and the stroma of many other tissues, and can give rise to a number of mesoderm-type cells including adipocytes and osteoblasts, which form medullary fat and bone tissues, respectively. The role of bone marrow fat in bone mass homeostasis is an area of intensive investigation with the aim of developing novel approaches for enhancing osteoblastic bone formation through inhibition of bone marrow fat formation. A number of recent studies have reported several miRNAs that enhance or inhibit adipogenic differentiation of MSCs and with potential use in microRNA-based therapy to regulate adipogenesis in the context of treating bone diseases and metabolic disorders. The current review focuses on miRNAs and their role in regulating adipogenic differentiation of MSCs.

Intra-islet endothelial cell and β-cell crosstalk: Implication for islet cell transplantation

PubMed Central

Narayanan, Siddharth; Loganathan, Gopalakrishnan; Dhanasekaran, Maheswaran; Tucker, William; Patel, Ankit; Subhashree, Venugopal; Mokshagundam, SriPrakash; Hughes, Michael G; Williams, Stuart K; Balamurugan, Appakalai N

2017-01-01

The intra-islet microvasculature is a critical interface between the blood and islet endocrine cells governing a number of cellular and pathophysiological processes associated with the pancreatic tissue. A growing body of evidence indicates a strong functional and physical interdependency of β-cells with endothelial cells (ECs), the building blocks of islet microvasculature. Intra-islet ECs, actively regulate vascular permeability and appear to play a role in fine-tuning blood glucose sensing and regulation. These cells also tend to behave as “guardians”, controlling the expression and movement of a number of important immune mediators, thereby strongly contributing to the physiology of islets. This review will focus on the molecular signalling and crosstalk between the intra-islet ECs and β-cells and how their relationship can be a potential target for intervention strategies in islet pathology and islet transplantation. PMID:28507914

[The implementation of computer model in research of dynamics of proliferation of cells of thyroid gland follicle].

PubMed

Abduvaliev, A A; Gil'dieva, M S; Khidirov, B N; Saĭdalieva, M; Khasanov, A A; Musaeva, Sh N; Saatov, T S

2012-04-01

The article deals with the results of computational experiments in research of dynamics of proliferation of cells of thyroid gland follicle in normal condition and in the case of malignant neoplasm. The model studies demonstrated that the chronic increase of parameter of proliferation of cells of thyroid gland follicle results in abnormal behavior of numbers of cell cenosis of thyroid gland follicle. The stationary state interrupts, the auto-oscillations occur with transition to irregular oscillations with unpredictable cell proliferation and further to the "black hole" effect. It is demonstrated that the present medical biologic experimental data and theory propositions concerning the structural functional organization of thyroid gland on cell level permit to develop mathematical models for quantitative analysis of numbers of cell cenosis of thyroid gland follicle in normal conditions. The technique of modeling of regulative mechanisms of living systems and equations of cell cenosis regulations was used

FADD and the NF-κB family member Bcl-3 regulate complementary pathways to control T-cell survival and proliferation

PubMed Central

Rangelova, Svetla; Kirschnek, Susanne; Strasser, Andreas; Häcker, Georg

2008-01-01

Fas-associated protein with death domain/mediator of receptor induced toxicity (FADD/MORT1) was first described as a transducer of death receptor signalling but was later recognized also to be important for proliferation of T cells. B-cell lymphoma 3 (Bcl-3) is a relatively little understood member of the nuclear factor (NF)-κB family of transcription factors. We recently found that Bcl-3 is up-regulated in T cells from mice where FADD function is blocked by a dominant negative transgene (FADD-DN). To understand the importance of this, we generated FADD-DN/bcl-3−/− mice. Here, we report that T cells from these mice show massive cell death and severely reduced proliferation in response to T-cell receptor (TCR) stimulation in vitro. Transgenic co-expression of Bcl-2 (FADD-DN/bcl-3−/−/vav-bcl-2 mice) rescued the survival but not the proliferation of T cells. FADD-DN/bcl-3−/− mice had normal thymocyte numbers but reduced numbers of peripheral T cells despite an increase in cycling T cells in vivo. However, activation of the classical NF-κB and extracellular regulated kinase (ERK) pathways and expression of interleukin (IL)-2 mRNA upon stimulation were normal in T cells from FADD-DN/bcl-3−/− mice. These data suggest that FADD and Bcl-3 regulate separate pathways that both contribute to survival and proliferation in mouse T cells. PMID:18557791

Effect of promoter architecture on the cell-to-cell variability in gene expression.

PubMed

Sanchez, Alvaro; Garcia, Hernan G; Jones, Daniel; Phillips, Rob; Kondev, Jané

2011-03-01

According to recent experimental evidence, promoter architecture, defined by the number, strength and regulatory role of the operators that control transcription, plays a major role in determining the level of cell-to-cell variability in gene expression. These quantitative experiments call for a corresponding modeling effort that addresses the question of how changes in promoter architecture affect variability in gene expression in a systematic rather than case-by-case fashion. In this article we make such a systematic investigation, based on a microscopic model of gene regulation that incorporates stochastic effects. In particular, we show how operator strength and operator multiplicity affect this variability. We examine different modes of transcription factor binding to complex promoters (cooperative, independent, simultaneous) and how each of these affects the level of variability in transcriptional output from cell-to-cell. We propose that direct comparison between in vivo single-cell experiments and theoretical predictions for the moments of the probability distribution of mRNA number per cell can be used to test kinetic models of gene regulation. The emphasis of the discussion is on prokaryotic gene regulation, but our analysis can be extended to eukaryotic cells as well.

Effect of Promoter Architecture on the Cell-to-Cell Variability in Gene Expression

PubMed Central

Sanchez, Alvaro; Garcia, Hernan G.; Jones, Daniel; Phillips, Rob; Kondev, Jané

2011-01-01

According to recent experimental evidence, promoter architecture, defined by the number, strength and regulatory role of the operators that control transcription, plays a major role in determining the level of cell-to-cell variability in gene expression. These quantitative experiments call for a corresponding modeling effort that addresses the question of how changes in promoter architecture affect variability in gene expression in a systematic rather than case-by-case fashion. In this article we make such a systematic investigation, based on a microscopic model of gene regulation that incorporates stochastic effects. In particular, we show how operator strength and operator multiplicity affect this variability. We examine different modes of transcription factor binding to complex promoters (cooperative, independent, simultaneous) and how each of these affects the level of variability in transcriptional output from cell-to-cell. We propose that direct comparison between in vivo single-cell experiments and theoretical predictions for the moments of the probability distribution of mRNA number per cell can be used to test kinetic models of gene regulation. The emphasis of the discussion is on prokaryotic gene regulation, but our analysis can be extended to eukaryotic cells as well. PMID:21390269

Over-expression of Stat5b confers protection against diabetes in the non-obese diabetic (NOD) mice via up-regulation of CD4{sup +}CD25{sup +} regulatory T cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jin, Yulan; Purohit, Sharad; Department of Pathology, Medical College of Georgia, Georgia Health Sciences University, GA

Highlights: Black-Right-Pointing-Pointer This is the first study to provide direct evidence of the role of Stat5b in NOD mice. Black-Right-Pointing-Pointer Over-expression of wild type Stat5b transgene protects NOD mice against diabetes. Black-Right-Pointing-Pointer This protection may be mediated by the up-regulation of CD4{sup +}CD25{sup +} Tregs. -- Abstract: The signal transducers and activators of transcription (STAT) family of proteins play a critical role in cytokine signaling required for fine tuning of immune regulation. Previous reports showed that a mutation (L327M) in the Stat5b protein leads to aberrant cytokine signaling in the NOD mice. To further elaborate the role of Stat5b inmore » diabetes, we established a NOD transgenic mouse that over-expresses the wild type Stat5b gene. The incidences of spontaneous diabetes as well as cyclophosphamide-induced diabetes were significantly reduced and delayed in the Stat5b transgenic NOD mice compared to their littermate controls. The total cell numbers of CD4{sup +} T cells and especially CD8{sup +} T cells in the spleen and pancreatic lymph node were increased in the Stat5b transgenic NOD mice. Consistent with these findings, CD4{sup +} and CD8{sup +} T cells from the Stat5b transgenic NOD mice showed a higher proliferation capacity and up-regulation of multiple cytokines including IL-2, IFN-{gamma}, TNF-{alpha} and IL-10 as well as anti-apoptotic gene Bcl-xl. Furthermore, the number and proportion of CD4{sup +}CD25{sup +} regulatory T cells were significantly increased in transgenic mice although in vitro suppression ability of the regulatory T-cells was not affected by the transgene. Our results suggest that Stat5b confers protection against diabetes in the NOD mice by regulating the numbers and function of multiple immune cell types, especially by up-regulating CD4{sup +}CD25{sup +} regulatory T cells.« less

Leaf area and photosynthesis of newly emerged trifoliolate leaves are regulated by mature leaves in soybean.

PubMed

Wu, Yushan; Gong, Wanzhuo; Wang, Yangmei; Yong, Taiwen; Yang, Feng; Liu, Weigui; Wu, Xiaoling; Du, Junbo; Shu, Kai; Liu, Jiang; Liu, Chunyan; Yang, Wenyu

2018-03-29

Leaf anatomy and the stomatal development of developing leaves of plants have been shown to be regulated by the same light environment as that of mature leaves, but no report has yet been written on whether such a long-distance signal from mature leaves regulates the total leaf area of newly emerged leaves. To explore this question, we created an investigation in which we collected data on the leaf area, leaf mass per area (LMA), leaf anatomy, cell size, cell number, gas exchange and soluble sugar content of leaves from three soybean varieties grown under full sunlight (NS), shaded mature leaves (MS) or whole plants grown in shade (WS). Our results show that MS or WS cause a marked decline both in leaf area and LMA in newly developing leaves. Leaf anatomy also showed characteristics of shade leaves with decreased leaf thickness, palisade tissue thickness, sponge tissue thickness, cell size and cell numbers. In addition, in the MS and WS treatments, newly developed leaves exhibited lower net photosynthetic rate (Pn), stomatal conductance (Gs) and transpiration rate (E), but higher carbon dioxide (CO 2 ) concentration in the intercellular space (Ci) than plants grown in full sunlight. Moreover, soluble sugar content was significantly decreased in newly developed leaves in MS and WS treatments. These results clearly indicate that (1) leaf area, leaf anatomical structure, and photosynthetic function of newly developing leaves are regulated by a systemic irradiance signal from mature leaves; (2) decreased cell size and cell number are the major cause of smaller and thinner leaves in shade; and (3) sugars could possibly act as candidate signal substances to regulate leaf area systemically.

Identification of host transcriptional networks showing concentration-dependent regulation by HPV16 E6 and E7 proteins in basal cervical squamous epithelial cells

PubMed Central

Smith, Stephen P.; Scarpini, Cinzia G.; Groves, Ian J.; Odle, Richard I.; Coleman, Nicholas

2016-01-01

Development of cervical squamous cell carcinoma requires increased expression of the major high-risk human-papillomavirus (HPV) oncogenes E6 and E7 in basal cervical epithelial cells. We used a systems biology approach to identify host transcriptional networks in such cells and study the concentration-dependent changes produced by HPV16-E6 and -E7 oncoproteins. We investigated sample sets derived from the W12 model of cervical neoplastic progression, for which high quality phenotype/genotype data were available. We defined a gene co-expression matrix containing a small number of highly-connected hub nodes that controlled large numbers of downstream genes (regulons), indicating the scale-free nature of host gene co-expression in W12. We identified a small number of ‘master regulators’ for which downstream effector genes were significantly associated with protein levels of HPV16 E6 (n = 7) or HPV16 E7 (n = 5). We validated our data by depleting E6/E7 in relevant cells and by functional analysis of selected genes in vitro. We conclude that the network of transcriptional interactions in HPV16-infected basal-type cervical epithelium is regulated in a concentration-dependent manner by E6/E7, via a limited number of central master-regulators. These effects are likely to be significant in cervical carcinogenesis, where there is competitive selection of cells with elevated expression of virus oncoproteins. PMID:27457222

NtKRP, a kinesin-12 protein, regulates embryo/seed size and seed germination via involving in cell cycle progression at the G2/M transition.

PubMed

Tian, Shujuan; Wu, Jingjing; Li, Fen; Zou, Jianwei; Liu, Yuwen; Zhou, Bing; Bai, Yang; Sun, Meng-Xiang

2016-10-25

Kinesins comprise a superfamily of microtubule-based motor proteins involved in essential processes in plant development, but few kinesins have been functionally identified during seed development. Especially, few kinesins that regulate cell division during embryogenesis have been identified. Here we report the functional characterization of NtKRP, a motor protein of the kinesin-12 family. NtKRP is predominantly expressed in embryos and embryonic roots. NtKRP RNAi lines displayed reductions in cell numbers in the meristematic zone, in embryonic root length, and in mature embryo and seed sizes. Furthermore, we also show that CDKA;1 binds to NtKRP at the consensus phosphorylation sites and that the decreased cell numbers in NtKRP-silenced embryos are due to a delay in cell division cycle at the G2/M transition. In addition, binding between the cargo-binding tail domain of NtKRP and CDKA; 1 was also determined. Our results reveal a novel molecular pathway that regulates embryo/seed development and critical role of kinesin in temporal and spatial regulation of a specific issue of embryo developmental.

Megakaryocytes regulate hematopoietic stem cell quiescence through CXCL4 secretion.

PubMed

Bruns, Ingmar; Lucas, Daniel; Pinho, Sandra; Ahmed, Jalal; Lambert, Michele P; Kunisaki, Yuya; Scheiermann, Christoph; Schiff, Lauren; Poncz, Mortimer; Bergman, Aviv; Frenette, Paul S

2014-11-01

In the bone marrow, hematopoietic stem cells (HSCs) lodge in specialized microenvironments that tightly control the proliferative state of HSCs to adapt to the varying needs for replenishment of blood cells while also preventing HSC exhaustion. All putative niche cells suggested thus far have a nonhematopoietic origin. Thus, it remains unclear how feedback from mature cells is conveyed to HSCs to adjust their proliferation. Here we show that megakaryocytes (MKs) can directly regulate HSC pool size in mice. Three-dimensional whole-mount imaging revealed that endogenous HSCs are frequently located adjacent to MKs in a nonrandom fashion. Selective in vivo depletion of MKs resulted in specific loss of HSC quiescence and led to a marked expansion of functional HSCs. Gene expression analyses revealed that MKs are the source of chemokine C-X-C motif ligand 4 (CXCL4, also named platelet factor 4 or PF4) in the bone marrow, and we found that CXCL4 regulates HSC cell cycle activity. CXCL4 injection into mice resulted in a reduced number of HSCs because of their increased quiescence. By contrast, Cxcl4(-/-) mice exhibited an increased number of HSCs and increased HSC proliferation. Combined use of whole-mount imaging and computational modeling was highly suggestive of a megakaryocytic niche capable of independently influencing HSC maintenance by regulating quiescence. These results indicate that a terminally differentiated cell type derived from HSCs contributes to the HSC niche, directly regulating HSC behavior.

LNK mutations and myeloproliferative disorders.

PubMed

McMullin, Mary Frances; Cario, Holger

2016-02-01

The lymphocyte adaptor protein (LNK) is one of a family of adaptor proteins involved cell signaling and control of B cell populations. It has a critical role in regulation of signaling in hematopoiesis. Lnk negatively regulates cytokine initiated cell signaling and it functions as a negative regulator of the mutant protein in myeloproliferative neoplasms JAK2V617F. A number of mutations in LNK have been described in a variety of myeloproliferative neoplasms some of which have been demonstrated to cause increased cellular proliferation. The majority of mutations occur in exon 2. In a small number of cases idiopathic erythrocytosis with subnormal erythropoietin levels LNK mutations have been found which may account for the clinical phenotype. Thus investigation for LNK mutations should be considered in the investigation of idiopathic erythrocytosis and perhaps other myeloproliferative neoplasms. © 2015 Wiley Periodicals, Inc.

The contribution of hypothalamic macroglia to the regulation of energy homeostasis

PubMed Central

Buckman, Laura B.; Ellacott, Kate L. J.

2014-01-01

The hypothalamus is critical for the regulation of energy homeostasis. Genetic and pharmacologic studies have identified a number of key hypothalamic neuronal circuits that integrate signals controlling food intake and energy expenditure. Recently, studies have begun to emerge demonstrating a role for non-neuronal cell types in the regulation of energy homeostasis. In particular the potential importance of different glial cell types is increasingly being recognized. A number of studies have described changes in the activity of hypothalamic macroglia (principally astrocytes and tanycytes) in response to states of positive and negative energy balance, such as obesity and fasting. This article will review these studies and discuss how these findings are changing our understanding of the cellular mechanisms by which energy homeostasis is regulated. PMID:25374514

Manipulating memory CD8 T cell numbers by timed enhancement of IL-2 signals1

PubMed Central

Kim, Marie T.; Kurup, Samarchith P.; Starbeck-Miller, Gabriel R.; Harty, John T.

2016-01-01

Due to the growing burden of tumors and chronic infections, manipulating CD8 T cell responses for clinical use has become an important goal for immunologists. Here, we show that dendritic cell (DC) immunization coupled with relatively early (days 1–3) or late (days 4–6) administration of enhanced IL-2-signals both increase peak effector CD8 T cell numbers, but only early IL-2 signals enhance memory numbers. IL-2 signals delivered at relatively late time points drive terminal differentiation, marked Bim mediated contraction and do not increase memory T cell numbers. In contrast, early IL-2 signals induce effector cell metabolic profiles more conducive to memory formation. Of note, down-regulation of CD80 and CD86 was observed on DCs in vivo following early IL-2 treatment. Mechanistically, early IL-2 treatment enhanced CTLA-4 expression on regulatory T (Treg) cells, and CTLA-4 blockade alongside IL-2 treatment in vivo prevented the decrease in CD80 and CD86, supporting a cell-extrinsic role of CTLA-4 in down-regulating B7-ligand expression on DCs. Finally, DC immunization followed by early IL-2 treatment and αCTLA-4 blockade resulted in lower memory CD8 T cell numbers compared to the DC + early IL-2 treatment group. These data suggest that curtailed signaling through the B7-CD28 co-stimulatory axis during CD8 T cell activation limits terminal differentiation and preserves memory CD8 T cell formation and thus, should be considered in future T cell vaccination strategies. PMID:27439516

Professional Regulation: A Potentially Valuable Tool in Responding to “Stem Cell Tourism”

PubMed Central

Zarzeczny, Amy; Caulfield, Timothy; Ogbogu, Ubaka; Bell, Peter; Crooks, Valorie A.; Kamenova, Kalina; Master, Zubin; Rachul, Christen; Snyder, Jeremy; Toews, Maeghan; Zoeller, Sonja

2014-01-01

The growing international market for unproven stem cell-based interventions advertised on a direct-to-consumer basis over the internet (“stem cell tourism”) is a source of concern because of the risks it presents to patients as well as their supporters, domestic health care systems, and the stem cell research field. Emerging responses such as public and health provider-focused education and national regulatory efforts are encouraging, but the market continues to grow. Physicians play a number of roles in the stem cell tourism market and, in many jurisdictions, are members of a regulated profession. In this article, we consider the use of professional regulation to address physician involvement in stem cell tourism. Although it is not without its limitations, professional regulation is a potentially valuable tool that can be employed in response to problematic types of physician involvement in the stem cell tourism market. PMID:25241736

The lupus susceptibility gene Pbx1 regulates the balance between follicular helper T cell and regulatory T cell differentiation

PubMed Central

Choi, Seung-Chul; Hutchinson, Tarun E.; Titov, Anton A.; Seay, Howard R.; Li, Shiwu; Brusko, Todd M.; Croker, Byron P.; Salek-Ardakani, Shahram; Morel, Laurence

2016-01-01

Pbx1 controls chromatin accessibility to a large number of genes and is entirely conserved between mice and humans. The Pbx1-d dominant negative isoform is more frequent in the CD4+ T cells from lupus patients than from healthy controls. Pbx1-d is associated with the production of autoreactive T cells in mice carrying the Sle1a1 lupus susceptibility locus. Transgenic expression of Pbx1-d in CD4+ T cells reproduced the phenotypes of Sle1a1 mice, with increased inflammatory functions of CD4+ T cells and impaired regulatory T cell homeostasis. Pbx1-d Tg also expanded the number of follicular helper T cells in a cell-intrinsic and antigen-specific manner that was enhanced in recall responses, and resulted in TH1-biased antibodies. Moreover, Pbx1-d Tg CD4+ T cells upregulated the expression of miR-10a, miR-21 and miR-155, which have been implicated in Treg and TFH cell homeostasis. Our results suggest that Pbx1-d impacts lupus development by regulating effector T cell differentiation and promoting TFH cells at the expense of Treg cells. In addition, our results identify Pbx1 as a novel regulator of CD4+ T cell effector function. PMID:27296664

Regulation of CD4 T cells and their effects on immunopathological inflammation following viral infection.

PubMed

Bhattacharyya, Mitra; Madden, Patrick; Henning, Nathan; Gregory, Shana; Aid, Malika; Martinot, Amanda J; Barouch, Dan H; Penaloza-MacMaster, Pablo

2017-10-01

CD4 T cells help immune responses, but knowledge of how memory CD4 T cells are regulated and how they regulate adaptive immune responses and induce immunopathology is limited. Using adoptive transfer of virus-specific CD4 T cells, we show that naive CD4 T cells undergo substantial expansion following infection, but can induce lethal T helper type 1-driven inflammation. In contrast, memory CD4 T cells exhibit a biased proliferation of T follicular helper cell subsets and were able to improve adaptive immune responses in the context of minimal tissue damage. Our analyses revealed that type I interferon regulates the expansion of primary CD4 T cells, but does not seem to play a critical role in regulating the expansion of secondary CD4 T cells. Strikingly, blockade of type I interferon abrogated lethal inflammation by primary CD4 T cells following viral infection, despite that this treatment increased the numbers of primary CD4 T-cell responses. Altogether, these data demonstrate important aspects of how primary and secondary CD4 T cells are regulated in vivo, and how they contribute to immune protection and immunopathology. These findings are important for rational vaccine design and for improving adoptive T-cell therapies against persistent antigens. © 2017 John Wiley & Sons Ltd.

Proliferating cell nuclear antigen (PCNA)-associated KIAA0101/PAF15 protein is a cell cycle-regulated anaphase-promoting complex/cyclosome substrate.

PubMed

Emanuele, Michael J; Ciccia, Alberto; Elia, Andrew E H; Elledge, Stephen J

2011-06-14

The anaphase-promoting complex/cyclosome (APC/C) is a cell cycle-regulated E3 ubiquitin ligase that controls the degradation of substrate proteins at mitotic exit and throughout the G1 phase. We have identified an APC/C substrate and cell cycle-regulated protein, KIAA0101/PAF15. PAF15 protein levels peak in the G2/M phase of the cell cycle and drop rapidly at mitotic exit in an APC/C- and KEN-box-dependent fashion. PAF15 associates with proliferating cell nuclear antigen (PCNA), and depletion of PAF15 decreases the number of cells in S phase, suggesting a role for it in cell cycle regulation. Following irradiation, PAF15 colocalized with γH2AX foci at sites of DNA damage through its interaction with PCNA. Finally, PAF15 depletion led to an increase in homologous recombination-mediated DNA repair, and overexpression caused sensitivity to UV-induced DNA damage. We conclude that PAF15 is an APC/C-regulated protein involved in both cell cycle progression and the DNA damage response.

Regulation of the actin cytoskeleton-plasma membrane interplay by phosphoinositides.

PubMed

Saarikangas, Juha; Zhao, Hongxia; Lappalainen, Pekka

2010-01-01

The plasma membrane and the underlying cortical actin cytoskeleton undergo continuous dynamic interplay that is responsible for many essential aspects of cell physiology. Polymerization of actin filaments against cellular membranes provides the force for a number of cellular processes such as migration, morphogenesis, and endocytosis. Plasma membrane phosphoinositides (especially phosphatidylinositol bis- and trisphosphates) play a central role in regulating the organization and dynamics of the actin cytoskeleton by acting as platforms for protein recruitment, by triggering signaling cascades, and by directly regulating the activities of actin-binding proteins. Furthermore, a number of actin-associated proteins, such as BAR domain proteins, are capable of directly deforming phosphoinositide-rich membranes to induce plasma membrane protrusions or invaginations. Recent studies have also provided evidence that the actin cytoskeleton-plasma membrane interactions are misregulated in a number of pathological conditions such as cancer and during pathogen invasion. Here, we summarize the wealth of knowledge on how the cortical actin cytoskeleton is regulated by phosphoinositides during various cell biological processes. We also discuss the mechanisms by which interplay between actin dynamics and certain membrane deforming proteins regulate the morphology of the plasma membrane.

p53 Mutation suppresses adult neurogenesis in medaka fish (Oryzias latipes)

DOE Office of Scientific and Technical Information (OSTI.GOV)

Isoe, Yasuko; Okuyama, Teruhiro; Taniguchi, Yoshihito

2012-07-13

Highlights: Black-Right-Pointing-Pointer Progenitor migration is accompanied by an increase in their numbers in the adult brain. Black-Right-Pointing-Pointer p53 Mutation suppressed an increase in the number of the migrated progenitors. Black-Right-Pointing-Pointer The decreased progenitor number is not due to enhanced cell death. Black-Right-Pointing-Pointer p53 Mutation did not affect proliferation of stem cells. -- Abstract: Tumor suppressor p53 negatively regulates self-renewal of neural stem cells in the adult murine brain. Here, we report that the p53 null mutation in medaka fish (Oryzias latipes) suppressed neurogenesis in the telencephalon, independent of cell death. By using 5-bromo-29-deoxyuridine (BrdU) immunohistochemistry, we identified 18 proliferation zonesmore » in the brains of young medaka fish; in situ hybridization showed that p53 was expressed selectively in at least 12 proliferation zones. We also compared the number of BrdU-positive cells present in the whole telencephalon of wild-type (WT) and p53 mutant fish. Immediately after BrdU exposure, the number of BrdU-positive cells did not differ significantly between them. One week after BrdU-exposure, the BrdU-positive cells migrated from the proliferation zone, which was accompanied by an increased number in the WT brain. In contrast, no significant increase was observed in the p53 mutant brain. Terminal deoxynucleotidyl transferase (dUTP) nick end-labeling revealed that there was no significant difference in the number of apoptotic cells in the telencephalon of p53 mutant and WT medaka, suggesting that the decreased number of BrdU-positive cells in the mutant may be due to the suppression of proliferation rather than the enhancement of neural cell death. These results suggest that p53 positively regulates neurogenesis via cell proliferation.« less

Co-regulation of pluripotency and genetic integrity at the genomic level.

PubMed

Cooper, Daniel J; Walter, Christi A; McCarrey, John R

2014-11-01

The Disposable Soma Theory holds that genetic integrity will be maintained at more pristine levels in germ cells than in somatic cells because of the unique role germ cells play in perpetuating the species. We tested the hypothesis that the same concept applies to pluripotent cells compared to differentiated cells. Analyses of transcriptome and cistrome databases, along with canonical pathway analysis and chromatin immunoprecipitation confirmed differential expression of DNA repair and cell death genes in embryonic stem cells and induced pluripotent stem cells relative to fibroblasts, and predicted extensive direct and indirect interactions between the pluripotency and genetic integrity gene networks in pluripotent cells. These data suggest that enhanced maintenance of genetic integrity is fundamentally linked to the epigenetic state of pluripotency at the genomic level. In addition, these findings demonstrate how a small number of key pluripotency factors can regulate large numbers of downstream genes in a pathway-specific manner. Copyright © 2014. Published by Elsevier B.V.

Distinct subpopulations of FOXD1 stroma-derived cells regulate renal erythropoietin

PubMed Central

Liu, Qingdu; Binns, Thomas C.; Davidoff, Olena; Kapitsinou, Pinelopi P.; Pfaff, Andrew S.; Olauson, Hannes; Fogo, Agnes B.; Fong, Guo-Hua; Gross, Kenneth W.

2016-01-01

Renal peritubular interstitial fibroblast-like cells are critical for adult erythropoiesis, as they are the main source of erythropoietin (EPO). Hypoxia-inducible factor 2 (HIF-2) controls EPO synthesis in the kidney and liver and is regulated by prolyl-4-hydroxylase domain (PHD) dioxygenases PHD1, PHD2, and PHD3, which function as cellular oxygen sensors. Renal interstitial cells with EPO-producing capacity are poorly characterized, and the role of the PHD/HIF-2 axis in renal EPO-producing cell (REPC) plasticity is unclear. Here we targeted the PHD/HIF-2/EPO axis in FOXD1 stroma-derived renal interstitial cells and examined the role of individual PHDs in REPC pool size regulation and renal EPO output. Renal interstitial cells with EPO-producing capacity were entirely derived from FOXD1-expressing stroma, and Phd2 inactivation alone induced renal Epo in a limited number of renal interstitial cells. EPO induction was submaximal, as hypoxia or pharmacologic PHD inhibition further increased the REPC fraction among Phd2–/– renal interstitial cells. Moreover, Phd1 and Phd3 were differentially expressed in renal interstitium, and heterozygous deficiency for Phd1 and Phd3 increased REPC numbers in Phd2–/– mice. We propose that FOXD1 lineage renal interstitial cells consist of distinct subpopulations that differ in their responsiveness to Phd2 inactivation and thus regulation of HIF-2 activity and EPO production under hypoxia or conditions of pharmacologic or genetic PHD inactivation. PMID:27088801

Tropomodulins are negative regulators of neurite outgrowth

PubMed Central

Fath, Thomas; Fischer, Robert S.; Dehmelt, Leif; Halpain, Shelley; Fowler, Velia M.

2010-01-01

Regulation of the actin cytoskeleton is critical for neurite formation. Tropomodulins (Tmods) regulate polymerization at actin filament pointed ends. Previous experiments using a mouse model deficient for the neuron specific isoform Tmod2 suggested a role for Tmods in neuronal function by impacting processes underlying learning and memory. However, the role of Tmods in neuronal function on the cellular level remains unknown. Immunofluorescence localization of the neuronal isoforms Tmod1 and Tmod2 in cultured rat primary hippocampal neurons revealed that Tmod1 is enriched along the proximal part of F-actin bundles in lamellipodia of spreading cells and in growth cones of extending neurites, while Tmod2 appears largely cytoplasmic. Functional analysis of these Tmod isoforms in a mouse neuroblastoma N2a cell line showed that knockdown of Tmod2 resulted in a significant increase in number of neurite-forming cells and in neurite length. While N2a cells compensated for Tmod2 knockdown by increasing Tmod1 levels, over-expression of exogenous Tmod1 had no effect on neurite outgrowth. Moreover, knockdown of Tmod1 increased the number of neurites formed per cell, without effect on number of neurite-forming cells or neurite length. Taken together, these results indicate that Tmod1 and Tmod2 have mechanistically distinct inhibitory roles in neurite formation, likely mediated via different effects on F-actin dynamics and via differential localizations during early neuritogenesis. PMID:21146252

Suppression of human fibrosarcoma cell growth by transcription factor, Egr-1, involves down-regulation of Bcl-2.

PubMed

Huang, R P; Fan, Y; Peng, A; Zeng, Z L; Reed, J C; Adamson, E D; Boynton, A L

1998-09-11

Previously, we showed that the transcription factor Egr-1 suppressed the proliferation of v-sis transformed NIH3T3 cells and also a number of human tumor cells. Here, we investigate the possible mechanisms responsible for this function. We show that transfected Egr-1 in human fibrosarcoma cells HT1080 leads to down-regulation of Bcl-2. Transient CAT transfection assays reveal that expression of Egr-1 suppresses Bcl-2 promoter activity in a dose-dependent manner. Furthermore, overexpression of Bcl-2 in Egr-1-expressing HT1080 cells enhanced cell proliferation in monolayer culture and increased anchorage-independent growth. Our results suggest that suppression of tumor cell proliferation by Egr-1 may be at least partially mediated through the down-regulation of Bcl-2.

Regulated internalization of NMDA receptors drives PKD1-mediated suppression of the activity of residual cell-surface NMDA receptors.

PubMed

Fang, Xiao-Qian; Qiao, Haifa; Groveman, Bradley R; Feng, Shuang; Pflueger, Melissa; Xin, Wen-Kuan; Ali, Mohammad K; Lin, Shuang-Xiu; Xu, Jindong; Duclot, Florian; Kabbaj, Mohamed; Wang, Wei; Ding, Xin-Sheng; Santiago-Sim, Teresa; Jiang, Xing-Hong; Salter, Michael W; Yu, Xian-Min

2015-11-19

Constitutive and regulated internalization of cell surface proteins has been extensively investigated. The regulated internalization has been characterized as a principal mechanism for removing cell-surface receptors from the plasma membrane, and signaling to downstream targets of receptors. However, so far it is still not known whether the functional properties of remaining (non-internalized) receptor/channels may be regulated by internalization of the same class of receptor/channels. The N-methyl-D-aspartate receptor (NMDAR) is a principal subtype of glutamate-gated ion channel and plays key roles in neuronal plasticity and memory functions. NMDARs are well-known to undergo two types of regulated internalization - homologous and heterologous, which can be induced by high NMDA/glycine and DHPG, respectively. In the present work, we investigated effects of regulated NMDAR internalization on the activity of residual cell-surface NMDARs and neuronal functions. In electrophysiological experiments we discovered that the regulated internalization of NMDARs not only reduced the number of cell surface NMDARs but also caused an inhibition of the activity of remaining (non-internalized) surface NMDARs. In biochemical experiments we identified that this functional inhibition of remaining surface NMDARs was mediated by increased serine phosphorylation of surface NMDARs, resulting from the activation of protein kinase D1 (PKD1). Knockdown of PKD1 did not affect NMDAR internalization but prevented the phosphorylation and inhibition of remaining surface NMDARs and NMDAR-mediated synaptic functions. These data demonstrate a novel concept that regulated internalization of cell surface NMDARs not only reduces the number of NMDARs on the cell surface but also causes an inhibition of the activity of remaining surface NMDARs through intracellular signaling pathway(s). Furthermore, modulating the activity of remaining surface receptors may be an effective approach for treating receptor internalization-induced changes in neuronal functions of the CNS.

What determines organ size differences between species? A meta-analysis of the cellular basis.

PubMed

Gázquez, Ayelén; Beemster, Gerrit T S

2017-07-01

Little is known about how the characteristic differences in organ size between species are regulated. At the cellular level, the size of an organ is strictly regulated by cell division and expansion during its development. We performed a meta-analysis of the growth parameters of roots, and Graminae and eudicotyledonous leaves, to address the question of how quantitative variation in these two processes contributes to size differences across a range of species. We extracted or derived cellular parameters from published kinematic growth analyses. These data were subjected to linear regression analyses to identify the parameters that determine differences in organ growth. Our results demonstrate that, across all species and organs, similar conclusions can be made: cell number rather than cell size determines the final size of plant organs; cell number is determined by meristem size rather than the rate at which cells divide; cells that are small when leaving the meristem compensate by expanding for longer; mature cell size is primarily determined by the duration of cell expansion. These results identify the regulation of the transition from cell division to expansion as the key cellular mechanism targeted by the evolution of organ size. © 2017 The Authors. New Phytologist © 2017 New Phytologist Trust.

High Concentration of Melatonin Regulates Leaf Development by Suppressing Cell Proliferation and Endoreduplication in Arabidopsis.

PubMed

Wang, Qiannan; An, Bang; Shi, Haitao; Luo, Hongli; He, Chaozu

2017-05-05

N -acetyl-5-methoxytryptamine (Melatonin), as a crucial messenger in plants, functions in adjusting biological rhythms, stress tolerance, plant growth and development. Several studies have shown the retardation effect of exogenous melatonin treatment on plant growth and development. However, the in vivo role of melatonin in regulating plant leaf growth and the underlying mechanism are still unclear. In this study, we found that high concentration of melatonin suppressed leaf growth in Arabidopsis by reducing both cell size and cell number. Further kinetic analysis of the fifth leaves showed that melatonin remarkably inhibited cell division rate. Additionally, flow cytometic analysis indicated that melatonin negatively regulated endoreduplication during leaf development. Consistently, the expression analysis revealed that melatonin regulated the transcriptional levels of key genes of cell cycle and ribosome. Taken together, this study suggests that high concentration of melatonin negatively regulated the leaf growth and development in Arabidopsis , through modulation of endoreduplication and the transcripts of cell cycle and ribosomal key genes.

High Concentration of Melatonin Regulates Leaf Development by Suppressing Cell Proliferation and Endoreduplication in Arabidopsis

PubMed Central

Wang, Qiannan; An, Bang; Shi, Haitao; Luo, Hongli; He, Chaozu

2017-01-01

N-acetyl-5-methoxytryptamine (Melatonin), as a crucial messenger in plants, functions in adjusting biological rhythms, stress tolerance, plant growth and development. Several studies have shown the retardation effect of exogenous melatonin treatment on plant growth and development. However, the in vivo role of melatonin in regulating plant leaf growth and the underlying mechanism are still unclear. In this study, we found that high concentration of melatonin suppressed leaf growth in Arabidopsis by reducing both cell size and cell number. Further kinetic analysis of the fifth leaves showed that melatonin remarkably inhibited cell division rate. Additionally, flow cytometic analysis indicated that melatonin negatively regulated endoreduplication during leaf development. Consistently, the expression analysis revealed that melatonin regulated the transcriptional levels of key genes of cell cycle and ribosome. Taken together, this study suggests that high concentration of melatonin negatively regulated the leaf growth and development in Arabidopsis, through modulation of endoreduplication and the transcripts of cell cycle and ribosomal key genes. PMID:28475148

Identification of Cell Cycle-regulated Genes in Fission YeastD⃞

PubMed Central

Peng, Xu; Karuturi, R. Krishna Murthy; Miller, Lance D.; Lin, Kui; Jia, Yonghui; Kondu, Pinar; Wang, Long; Wong, Lim-Soon; Liu, Edison T.; Balasubramanian, Mohan K.; Liu, Jianhua

2005-01-01

Cell cycle progression is both regulated and accompanied by periodic changes in the expression levels of a large number of genes. To investigate cell cycle-regulated transcriptional programs in the fission yeast Schizosaccharomyces pombe, we developed a whole-genome oligonucleotide-based DNA microarray. Microarray analysis of both wild-type and cdc25 mutant cell cultures was performed to identify transcripts whose levels oscillated during the cell cycle. Using an unsupervised algorithm, we identified 747 genes that met the criteria for cell cycle-regulated expression. Peaks of gene expression were found to be distributed throughout the entire cell cycle. Furthermore, we found that four promoter motifs exhibited strong association with cell cycle phase-specific expression. Examination of the regulation of MCB motif-containing genes through the perturbation of DNA synthesis control/MCB-binding factor (DSC/MBF)-mediated transcription in arrested synchronous cdc10 mutant cell cultures revealed a subset of functional targets of the DSC/MBF transcription factor complex, as well as certain gene promoter requirements. Finally, we compared our data with those for the budding yeast Saccharomyces cerevisiae and found ∼140 genes that are cell cycle regulated in both yeasts, suggesting that these genes may play an evolutionarily conserved role in regulation of cell cycle-specific processes. Our complete data sets are available at http://giscompute.gis.a-star.edu.sg/~gisljh/CDC. PMID:15616197

Antagonistic Regulation of Cystic Fibrosis Transmembrane Conductance Regulator Cell Surface Expression by Protein Kinases WNK4 and Spleen Tyrosine Kinase ▿

PubMed Central

Mendes, Ana Isabel; Matos, Paulo; Moniz, Sónia; Luz, Simão; Amaral, Margarida D.; Farinha, Carlos M.; Jordan, Peter

2011-01-01

Members of the WNK (with-no-lysine [K]) subfamily of protein kinases regulate various ion channels involved in sodium, potassium, and chloride homeostasis by either inducing their phosphorylation or regulating the number of channel proteins expressed at the cell surface. Here, we describe findings demonstrating that the cell surface expression of the cystic fibrosis transmembrane conductance regulator (CFTR) is also regulated by WNK4 in mammalian cells. This effect of WNK4 is independent of the presence of kinase and involves interaction with and inhibition of spleen tyrosine kinase (Syk), which phosphorylates Tyr512 in the first nucleotide-binding domain 1 (NBD1) of CFTR. Transfection of catalytically active Syk into CFTR-expressing baby hamster kidney cells reduces the cell surface expression of CFTR, whereas that of WNK4 promotes it. This is shown by biotinylation of cell surface proteins, immunofluorescence microscopy, and functional efflux assays. Mutation of Tyr512 to either glutamic acid or phenylalanine is sufficient to alter CFTR surface levels. In human airway epithelial cells, downregulation of endogenous Syk and WNK4 confirms their roles as physiologic regulators of CFTR surface expression. Together, our results show that Tyr512 phosphorylation is a novel signal regulating the prevalence of CFTR at the cell surface and that WNK4 and Syk perform an antagonistic role in this process. PMID:21807898

Modulation of Mitochondrial DNA Copy Number to Induce Hepatocytic Differentiation of Human Amniotic Epithelial Cells.

PubMed

Vaghjiani, Vijesh; Cain, Jason E; Lee, William; Vaithilingam, Vijayaganapathy; Tuch, Bernard E; St John, Justin C

2017-10-15

Mitochondrial deoxyribonucleic acid (mtDNA) copy number is tightly regulated during pluripotency and differentiation. There is increased demand of cellular adenosine triphosphate (ATP) during differentiation for energy-intensive cell types such as hepatocytes and neurons to meet the cell's functional requirements. During hepatocyte differentiation, mtDNA copy number should be synchronously increased to generate sufficient ATP through oxidative phosphorylation. Unlike bone marrow mesenchymal cells, mtDNA copy number failed to increase by 28 days of differentiation of human amniotic epithelial cells (hAEC) into hepatocyte-like cells (HLC) despite their expression of some end-stage hepatic markers. This was due to higher levels of DNA methylation at exon 2 of POLGA, the mtDNA-specific replication factor. Treatment with a DNA demethylation agent, 5-azacytidine, resulted in increased mtDNA copy number, reduced DNA methylation at exon 2 of POLGA, and reduced hepatic gene expression. Depletion of mtDNA followed by subsequent differentiation did not increase mtDNA copy number, but reduced DNA methylation at exon 2 of POLGA and increased expression of hepatic and pluripotency genes. We encapsulated hAEC in barium alginate microcapsules and subsequently differentiated them into HLC. Encapsulation resulted in no net increase of mtDNA copy number but a significant reduction in DNA methylation of POLGA. RNAseq analysis showed that differentiated HLC express hepatocyte-specific genes but also increased expression of inflammatory interferon genes. Differentiation in encapsulated cells showed suppression of inflammatory genes as well as increased expression of genes associated with hepatocyte function pathways and networks. This study demonstrates that an increase in classical hepatic gene expression can be achieved in HLC through encapsulation, although they fail to effectively regulate mtDNA copy number.

DEC1/STRA13 is a key negative regulator of activation-induced proliferation of human B cells highly expressed in anergic cells.

PubMed

Camponeschi, Alessandro; Todi, Laura; Cristofoletti, Cristina; Lazzeri, Cristina; Carbonari, Maurizio; Mitrevski, Milica; Marrapodi, Ramona; Del Padre, Martina; Fiorilli, Massimo; Casato, Milvia; Visentini, Marcella

2018-06-01

The transcription factor DEC1/STRA13 (also known as BHLHE40 and SHARP2) is involved in a number of processes including inhibition of cell proliferation and delay of cell cycle, and is a negative regulator of B cell activation and development in mice. We show here that, unlike in mice, DEC1/STRA13 expression is induced in human naïve and memory resting B cells by activation through the B-cell receptor (BCR) or Toll-like receptor 9 (TLR9). siRNA silencing of DEC1/STRA13 increases the capacity of activated B cells to perform a high number of divisions after TLR9 ligation. This identifies DEC1/STRA13 as a critical negative regulator of clonal expansion of activated human B cells. We also show that DEC1/STRA13 is upregulated in human anergic CD21 low B cells clonally expanded in patients with HCV-associated mixed cryoglobulinemia, which fail to proliferate in response to BCR or TLR9 ligation. siRNA knockdown of DEC1/STRA13, however, fails to restore responsiveness to stimuli in these cells, although it might improve the proliferative capacity in a subset of anergic cells with less pronounced proliferative defect. Copyright © 2018 European Federation of Immunological Societies. Published by Elsevier B.V. All rights reserved.

Chloroquine synergizes with FTS to enhance cell growth inhibition and cell death

PubMed Central

Schmukler, Eran; Wolfson, Eya; Haklai, Roni; Elad-Sfadia, Galit; Kloog, Yoel; Pinkas-Kramarski, Ronit

2014-01-01

The Ras family of small GTPases transmits extracellular signals that regulate cell growth, differentiation, motility and death. Ras signaling is constitutively active in a large number of human cancers. Ras can also regulate autophagy by affecting several signaling pathways including the mTOR pathway. Autophagy is a process that regulates the balance between protein synthesis and protein degradation. It is important for normal growth control, but may be defective in diseases. Previously, we have shown that Ras inhibition by FTS induces autophagy, which partially protects cancer cells and may limit the use of FTS as an anti-cancer drug. Since FTS is a non toxic drug we hypothesized that FTS and chloroquine (an autophagy inhibitor) will synergize in cell growth inhibition and cell death. Thus, in the present study, we explored the mechanism of each individual drug and their combined action. Our results demonstrate that in HCT-116 and in Panc-1 cells, FTS induces autophagy, which can be inhibited by chloroquine. Furthermore, the combined treatment synergistically decreased the number of viable cells. Interestingly, the combined treatment enhanced apoptotic cell death as indicated by increased sub-G1 cell population, increased Hoechst staining, activation of caspase 3, decrease in survivin expression and release of cytochrome c. Thus, chloroquine treatment may promote FTS-mediated inhibition of tumor cell growth and may stimulate apoptotic cell death. PMID:24368422

Regulation of diet-induced adipose tissue and systemic inflammation by salicylates and pioglitazone.

PubMed

Kim, Myung-Sunny; Yamamoto, Yasuhiko; Kim, Kyungjin; Kamei, Nozomu; Shimada, Takeshi; Liu, Libin; Moore, Kristin; Woo, Ju Rang; Shoelson, Steven E; Lee, Jongsoon

2013-01-01

It is increasingly accepted that chronic inflammation participates in obesity-induced insulin resistance and type 2 diabetes (T2D). Salicylates and thiazolidinediones (TZDs) both have anti-inflammatory and anti-hyperglycemic properties. The present study compared the effects of these drugs on obesity-induced inflammation in adipose tissue (AT) and AT macrophages (ATMs), as well as the metabolic and immunological phenotypes of the animal models. Both drugs improved high fat diet (HFD)-induced insulin resistance. However, salicylates did not affect AT and ATM inflammation, whereas Pioglitazone improved these parameters. Interestingly, HFD and the drug treatments all modulated systemic inflammation as assessed by changes in circulating immune cell numbers and activation states. HFD increased the numbers of circulating white blood cells, neutrophils, and a pro-inflammatory monocyte subpopulation (Ly6C(hi)), whereas salicylates and Pioglitazone normalized these cell numbers. The drug treatments also decreased circulating lymphocyte numbers. These data suggest that obesity induces systemic inflammation by regulating circulating immune cell phenotypes and that anti-diabetic interventions suppress systemic inflammation by normalizing circulating immune phenotypes.

Monocyte recruitment and expression of monocyte chemoattractant protein-1 are developmentally regulated in remodeling bone in the mouse.

PubMed Central

Volejnikova, S.; Laskari, M.; Marks, S. C.; Graves, D. T.

1997-01-01

Tooth eruption is defined as the movement of a tooth from its site of development within the alveolar bone to its position of function in the oral cavity. It represents an excellent model to examine osseous metabolism as bone resorption and bone formation occur simultaneously and are spatially separated. Bone resorption occurs in the coronal (occlusal) area, whereas bone formation occurs in the basal area. Monocytes are thought to have a significant role in the regulation of osseous metabolism. The goal of this study was to examine the recruitment of monocytes to bone in C57BL/6J mice that are undergoing developmentally regulated bone remodeling. Monocytes were detected by immunohistochemistry and osteoclasts were counted as bone-associated multi-nucleated, tartrate-resistant acid phosphatase (TRAP)-positive cells. Cell numbers were obtained from histological sections of animals sacrificed daily for 14 days after birth; an image analysis system was used for quantification. The results demonstrated that, immediately after birth, there were relatively few monocytic cells. In the area of bone resorption, the number of monocytes increased with time, reaching peaks at 5 and 9 days, and decreased thereafter. A similar pattern was observed for osteoclasts. In the area of bone formation, there was a time-dependent increase in the number of monocytes. In contrast, the number of osteoclasts in this area was highest at the earliest time points and decreased after day 3. To investigate potential mechanisms for the recruitment of monocytes, expression of monocyte chemoattractant protein (MCP)-1 was assessed. The number of MCP-1-positive cells increased with time and was generally proportional to the recruitment of mononuclear phagocytes. Osteoblasts were the principal bone cell type expressing MCP-1. The results demonstrate that the recruitment of mononuclear cells in the occlusal area is associated with bone resorption. In contrast, recruitment of monocytes in the basal area is associated with bone formation and a decrease in the number of osteoclasts. These results suggest that monocytes have different functional roles in areas of bone formation compared with bone resorption. Furthermore, the expression of MCP-1 is developmentally regulated and may provide a mechanistic basis to explain the recruitment of monocytic cells. Images Figure 1 Figure 2 Figure 3 Figure 4 PMID:9137095

Csk Homologous Kinase, a Potential Regulator of CXCR4-mediated Breast Cancer Cell Metastasis

DTIC Science & Technology

2010-08-31

SH2 ) and SH3 domains and lacks the consensus tyrosine phosphorylation and myristylation sites found in Src family kinases . CHK has been shown to...0350 TITLE: Csk Homologous Kinase , a Potential Regulator of CXCR4-mediated Breast Cancer Cell Metastasis PRINCIPAL INVESTIGATOR: Byeong-Chel...1 AUG 2009 - 31 JUL 2010 4. TITLE AND SUBTITLE 5a. CONTRACT NUMBER W81XWH-09-1-0350 Csk Homologous Kinase , a Potential Regulator

Cell cycle nucleic acids, polypeptides and uses thereof

DOEpatents

Gordon-Kamm, William J [Urbandale, IA; Lowe, Keith S [Johnston, IA; Larkins, Brian A [Tucson, AZ; Dilkes, Brian R [Tucson, AZ; Sun, Yuejin [Westfield, IN

2007-08-14

The invention provides isolated nucleic acids and their encoded proteins that are involved in cell cycle regulation. The invention further provides recombinant expression cassettes, host cells, transgenic plants, and antibody compositions. The present invention provides methods and compositions relating to altering cell cycle protein content, cell cycle progression, cell number and/or composition of plants.

Three-tier regulation of cell number plasticity by neurotrophins and Tolls in Drosophila

PubMed Central

Phizacklea, Mark; Gay, Nicholas J.

2017-01-01

Cell number plasticity is coupled to circuitry in the nervous system, adjusting cell mass to functional requirements. In mammals, this is achieved by neurotrophin (NT) ligands, which promote cell survival via their Trk and p75NTR receptors and cell death via p75NTR and Sortilin. Drosophila NTs (DNTs) bind Toll receptors instead to promote neuronal survival, but whether they can also regulate cell death is unknown. In this study, we show that DNTs and Tolls can switch from promoting cell survival to death in the central nervous system (CNS) via a three-tier mechanism. First, DNT cleavage patterns result in alternative signaling outcomes. Second, different Tolls can preferentially promote cell survival or death. Third, distinct adaptors downstream of Tolls can drive either apoptosis or cell survival. Toll-6 promotes cell survival via MyD88–NF-κB and cell death via Wek-Sarm-JNK. The distribution of adaptors changes in space and time and may segregate to distinct neural circuits. This novel mechanism for CNS cell plasticity may operate in wider contexts. PMID:28373203

Sonic hedgehog controls growth of external genitalia by regulating cell cycle kinetics

PubMed Central

Seifert, Ashley W.; Zheng, Zhengui; Ormerod, Brandi K.; Cohn, Martin J.

2010-01-01

During embryonic development, cells are instructed which position to occupy, they interpret these cues as differentiation programmes, and expand these patterns by growth. Sonic hedgehog (Shh) specifies positional identity in many organs; however, its role in growth is not well understood. In this study, we show that inactivation of Shh in external genitalia extends the cell cycle from 8.5 to 14.4 h, and genital growth is reduced by ∼75%. Transient Shh signalling establishes pattern in the genital tubercle; however, transcriptional levels of G1 cell cycle regulators are reduced. Consequently, G1 length is extended, leading to fewer progenitor cells entering S-phase. Cell cycle genes responded similarly to Shh inactivation in genitalia and limbs, suggesting that Shh may regulate growth by similar mechanisms in different organ systems. The finding that Shh regulates cell number by controlling the length of specific cell cycle phases identifies a novel mechanism by which Shh elaborates pattern during appendage development. PMID:20975695

Differentiation induction of mouse embryonic stem cells into sinus node-like cells by suramin

PubMed Central

Wiese, Cornelia; Nikolova, Teodora; Zahanich, Ihor; Sulzbacher, Sabine; Fuchs, Joerg; Yamanaka, Satoshi; Graf, Eva; Ravens, Ursula; Boheler, Kenneth R.; Wobus, Anna M.

2015-01-01

Background Embryonic stem (ES) cells differentiate into cardiac phenotypes representing early pacemaker-, atrial-, ventricular-, and sinus node-like cells, however, ES-derived specification into sinus nodal cells is not yet known. By using the naphthylamine derivative of urea, suramin, we were able to follow the process of cardiac specialization into sinus node-like cells. Methods Differentiating mouse ES cells were treated with suramin (500 μM) from day 5 to 7 of embryoid body formation, and cells were analysed for their differentiation potential via morphological analysis, flow cytometry, RT-PCR, immunohistochemistry and patch clamp analysis. Results Application of suramin resulted in an increased number of cardiac cells, but inhibition of neuronal, skeletal muscle and definitive endoderm differentiation. Immediately after suramin treatment, a decreased mesendoderm differentiation was found. Brachyury, FGF10, Wnt8 and Wnt3a transcript levels were significantly down-regulated, followed by a decrease in mesoderm- and cardiac progenitor-specific markers BMP2, GATA4/5, Wnt11, Isl1, Nkx2.5 and Tbx5 immediately after removal of the substance. With continued differentiation, a significant up-regulation of Brachyury, FGF10 and GATA5 transcript levels was observed, whereas Nkx2.5, Isl1, Tbx5, BMP2 and Wnt11 levels were normalized to control levels. At advanced differentiation stages, sinus node-specific HCN4, Tbx2 and Tbx3 transcript levels were significantly up-regulated. Immunofluorescence and patch-clamp analysis confirmed the increased number of sinus node-like cells, and electrophysiological analysis revealed a lower number of atrial- and ventricular-like cardiomyocytes following suramin treatment. Conclusion We conclude that the interference of suramin with the cardiac differentiation process modified mesoderm- and cardiac-specific gene expression resulting in enhanced formation of sinus node-like cells. PMID:19775764

Purinergic Receptors in Quiescence and Localization of Leukemic Stem Cells

DTIC Science & Technology

2011-05-01

2011. Ewing’s Sarcoma Gene EWS regulates Hematopoietic Stem Cell Senescence. Blood, 117:1156-66. Conclusion How leukemia stem cells gained...findings contained in this report are those of the author( s ) and should not be construed as an official Department of the Army position, policy or...Receptors in Quiescence and Localization of Leukemic Stem Cells 5b. GRANT NUMBER W81XWH-09-1-0364 5c. PROGRAM ELEMENT NUMBER 6. AUTHOR( S ) 5d

Integrative analysis of copy number and gene expression data suggests novel pathogenetic mechanisms in primary myelofibrosis.

PubMed

Salati, Simona; Zini, Roberta; Nuzzo, Simona; Guglielmelli, Paola; Pennucci, Valentina; Prudente, Zelia; Ruberti, Samantha; Rontauroli, Sebastiano; Norfo, Ruggiero; Bianchi, Elisa; Bogani, Costanza; Rotunno, Giada; Fanelli, Tiziana; Mannarelli, Carmela; Rosti, Vittorio; Salmoiraghi, Silvia; Pietra, Daniela; Ferrari, Sergio; Barosi, Giovanni; Rambaldi, Alessandro; Cazzola, Mario; Bicciato, Silvio; Tagliafico, Enrico; Vannucchi, Alessandro M; Manfredini, Rossella

2016-04-01

Primary myelofibrosis (PMF) is a Myeloproliferative Neoplasm (MPN) characterized by megakaryocyte hyperplasia, progressive bone marrow fibrosis, extramedullary hematopoiesis and transformation to Acute Myeloid Leukemia (AML). A number of phenotypic driver (JAK2, CALR, MPL) and additional subclonal mutations have been described in PMF, pointing to a complex genomic landscape. To discover novel genomic lesions that can contribute to disease phenotype and/or development, gene expression and copy number signals were integrated and several genomic abnormalities leading to a concordant alteration in gene expression levels were identified. In particular, copy number gain in the polyamine oxidase (PAOX) gene locus was accompanied by a coordinated transcriptional up-regulation in PMF patients. PAOX inhibition resulted in rapid cell death of PMF progenitor cells, while sparing normal cells, suggesting that PAOX inhibition could represent a therapeutic strategy to selectively target PMF cells without affecting normal hematopoietic cells' survival. Moreover, copy number loss in the chromatin modifier HMGXB4 gene correlates with a concomitant transcriptional down-regulation in PMF patients. Interestingly, silencing of HMGXB4 induces megakaryocyte differentiation, while inhibiting erythroid development, in human hematopoietic stem/progenitor cells. These results highlight a previously un-reported, yet potentially interesting role of HMGXB4 in the hematopoietic system and suggest that genomic and transcriptional imbalances of HMGXB4 could contribute to the aberrant expansion of the megakaryocytic lineage that characterizes PMF patients. © 2015 UICC.

EGY1 plays a role in regulation of endodermal plastid size and number that are involved in ethylene-dependent gravitropism of light-grown Arabidopsis hypocotyls.

PubMed

Guo, Di; Gao, Xiaorong; Li, Hao; Zhang, Tao; Chen, Gu; Huang, Pingbo; An, Lijia; Li, Ning

2008-03-01

Egy1 was isolated as an ethylene-dependent gravitropism-deficient Arabidopsis mutant. Molecular studies reveal that EGY1 gene encodes a 59-kDa plastid-targeted metalloprotease. It is actively expressed in hypocotyl tissue and targets to endodermal and cortex plastid. Its protein level is up-regulated by both ethylene and light. CAB protein accumulation and chlorophyll level is severely reduced in hypocotyls and endodermal cells, respectively. Sucrose is able to restore the severely reduced starch and lipid contents as well as the deficient endodermal plastid size found in light-grown egy1 hypocotyls yet it fails to rescue the reduced plastid number and chlorophyll level in egy1 endodermal cells. The loss-of-function egy1 mutation results in a smaller size (1.9 +/- 0.3 microm in diameter) and less number (5 +/- 1) of plastids in endodermal cells, which are nearly 50% of the wild-type. EGY1 is specially required for the development of full-size endodermal plastid in seedlings that are grown on sucrose-free media under light. It plays a direct role in controlling the light-induced chlorophyll production, grana formation and plastid replication in endodermal cell. However, it plays an indirect role in regulation of endodermal plastid size. It is likely that the ethylene-dependent gravitropism-deficient phenotype of egy1 hypocotyls may result from the smaller size and less number of endodermal plastids. Gravicurvature assays performed on ethylene-insensitive mutants, etr1-1, etr2-1, ers2-1, ein4-1 and ein2-5, have clearly demonstrated the necessary role for ethylene in vigorous gravitropism of light-grown hypocotyls. The degree of ethylene-dependent gravicurvature is positively correlated with the combined state of endodermal plastid mass and number. Neither ethylene nor EGY1-regulated full-size endodermal plastid is sufficient for promotion of vigorous hypocotyl gravitropism. Presence of 4 full-size plastids per endodermal cell together with ethylene pretreatment of hypocotyls becomes sufficient to trigger vigorous gravicurvature in light-grown seedlings. A model is therefore proposed to address the role of EGY1 in regulation of endodermal plastid size and number as well as the stimulatory effect of ethylene on hypocotyl gravitropism.

Megakaryocytes regulate hematopoietic stem cell quiescence via Cxcl4 secretion

PubMed Central

Bruns, Ingmar; Lucas, Daniel; Pinho, Sandra; Ahmed, Jalal; Lambert, Michele P.; Kunisaki, Yuya; Scheiermann, Christoph; Schiff, Lauren; Poncz, Mortimer; Bergman, Aviv; Frenette, Paul S.

2014-01-01

In the bone marrow (BM), hematopoietic stem cells (HSCs) lodge in specialized microenvironments that tightly control their proliferative state to adapt to the varying needs for replenishment of blood cells while also preventing exhaustion1. All putative niche cells suggested thus far have a non-hematopoietic origin2-8. Thus, it remains unclear how feedback from mature cells is conveyed to HSCs to adjust proliferation. Here we show that megakaryocytes (Mk) can directly regulate HSC pool size. Three-dimensional whole-mount imaging revealed that endogenous HSCs are frequently located adjacent to Mk in a non-random fashion. Selective in vivo depletion of Mk resulted in specific loss of HSC quiescence and led to a marked expansion of functional HSCs. Gene expression analyses revealed that Mk were the source of chemokine C-X-C motif ligand 4 (Cxcl4, also named platelet factor 4, Pf4) in the BM and Cxcl4 injection reduced HSC numbers via increased quiescence. By contrast, Cxcl4−/− mice exhibited increased HSC numbers and proliferation. Combined use of whole-mount imaging and computational modelling was highly suggestive of a megakaryocytic niche capable of influencing independently HSC maintenance by regulating quiescence. Thus, these results indicate that a terminally differentiated HSC progeny contributes to niche activity by directly regulating HSC behavior. PMID:25326802

The regulation of tooth morphogenesis is associated with epithelial cell proliferation and the expression of Sonic hedgehog through epithelial-mesenchymal interactions.

PubMed

Ishida, Kentaro; Murofushi, Mayumi; Nakao, Kazuhisa; Morita, Ritsuko; Ogawa, Miho; Tsuji, Takashi

2011-02-18

Ectodermal organs, such as the tooth, salivary gland, hair, and mammary gland, develop through reciprocal epithelial-mesenchymal interactions. Tooth morphologies are defined by the crown width and tooth length (macro-morphologies), and by the number and locations of the cusp and roots (micro-morphologies). In our current study, we report that the crown width of a bioengineered molar tooth, which was reconstructed using dissociated epithelial and mesenchymal cells via an organ germ method, can be regulated by the contact area between epithelial and mesenchymal cell layers. We further show that this is associated with cell proliferation and Sonic hedgehog (Shh) expression in the inner enamel epithelium after the germ stage has formed a secondary enamel knot. We also demonstrate that the cusp number is significantly correlated with the crown width of the bioengineered tooth. These findings suggest that the tooth micro-morphology, i.e. the cusp formation, is regulated after the tooth width, or macro-morphology, is determined. These findings also suggest that the spatiotemporal patterning of cell proliferation and the Shh expression areas in the epithelium regulate the crown width and cusp formation of the developing tooth. Copyright © 2011 Elsevier Inc. All rights reserved.

A new class of cyclin dependent kinase in Chlamydomonas is required for coupling cell size to cell division

PubMed Central

Li, Yubing; Liu, Dianyi; López-Paz, Cristina; Olson, Bradley JSC; Umen, James G

2016-01-01

Proliferating cells actively control their size by mechanisms that are poorly understood. The unicellular green alga Chlamydomonas reinhardtii divides by multiple fission, wherein a ‘counting’ mechanism couples mother cell-size to cell division number allowing production of uniform-sized daughters. We identified a sizer protein, CDKG1, that acts through the retinoblastoma (RB) tumor suppressor pathway as a D-cyclin-dependent RB kinase to regulate mitotic counting. Loss of CDKG1 leads to fewer mitotic divisions and large daughters, while mis-expression of CDKG1 causes supernumerous mitotic divisions and small daughters. The concentration of nuclear-localized CDKG1 in pre-mitotic cells is set by mother cell size, and its progressive dilution and degradation with each round of cell division may provide a link between mother cell-size and mitotic division number. Cell-size-dependent accumulation of limiting cell cycle regulators such as CDKG1 is a potentially general mechanism for size control. DOI: http://dx.doi.org/10.7554/eLife.10767.001 PMID:27015111

STAT3 is a critical cell-intrinsic regulator of human unconventional T cell numbers and function

PubMed Central

Wilson, Robert P.; Ives, Megan L.; Rao, Geetha; Lau, Anthony; Payne, Kathryn; Kobayashi, Masao; Arkwright, Peter D.; Peake, Jane; Wong, Melanie; Adelstein, Stephen; Smart, Joanne M.; French, Martyn A.; Fulcher, David A.; Picard, Capucine; Bustamante, Jacinta; Boisson-Dupuis, Stephanie; Gray, Paul; Stepensky, Polina; Warnatz, Klaus; Freeman, Alexandra F.; Rossjohn, Jamie; McCluskey, James; Holland, Steven M.; Casanova, Jean-Laurent; Uzel, Gulbu; Ma, Cindy S.

2015-01-01

Unconventional T cells such as γδ T cells, natural killer T cells (NKT cells) and mucosal-associated invariant T cells (MAIT cells) are a major component of the immune system; however, the cytokine signaling pathways that control their development and function in humans are unknown. Primary immunodeficiencies caused by single gene mutations provide a unique opportunity to investigate the role of specific molecules in regulating human lymphocyte development and function. We found that individuals with loss-of-function mutations in STAT3 had reduced numbers of peripheral blood MAIT and NKT but not γδ T cells. Analysis of STAT3 mosaic individuals revealed that this effect was cell intrinsic. Surprisingly, the residual STAT3-deficient MAIT cells expressed normal levels of the transcription factor RORγt. Despite this, they displayed a deficiency in secretion of IL-17A and IL-17F, but were able to secrete normal levels of cytokines such as IFNγ and TNF. The deficiency in MAIT and NKT cells in STAT3-deficient patients was mirrored by loss-of-function mutations in IL12RB1 and IL21R, respectively. Thus, these results reveal for the first time the essential role of STAT3 signaling downstream of IL-23R and IL-21R in controlling human MAIT and NKT cell numbers. PMID:25941256

Leaf shape: genetic controls and environmental factors.

PubMed

Tsukaya, Hirokazu

2005-01-01

In recent years, many genes have been identified that are involved in the developmental processes of leaf morphogenesis. Here, I review the mechanisms of leaf shape control in a model plant, Arabidopsis thaliana, focusing on genes that fulfill special roles in leaf development. The lateral, two-dimensional expansion of leaf blades is highly dependent on the determination of the dorsoventrality of the primordia, a defining characteristic of leaves. Having a determinate fate is also a characteristic feature of leaves and is controlled by many factors. Lateral expansion is not only controlled by general regulators of cell cycling, but also by the multi-level regulation of meristematic activities, e.g., specific control of cell proliferation in the leaf-length direction, in leaf margins and in parenchymatous cells. In collaboration with the polarized control of leaf cell elongation, these redundant and specialized regulating systems for cell cycling in leaf lamina may realize the elegantly smooth, flat structure of leaves. The unified, flat shape of leaves is also dependent on the fine integration of cell proliferation and cell enlargement. Interestingly, while a decrease in the number of cells in leaf primordia can trigger a cell volume increase, an increase in the number of cells does not trigger a cell volume decrease. This phenomenon is termed compensation and suggests the existence of some systems for integration between cell cycling and cell enlargement in leaf primordia via cell-cell communication. The environmental adjustment of leaf expansion to light conditions and gravity is also summarized.

Hypoxia promotes noncanonical autophagy in nucleus pulposus cells independent of MTOR and HIF1A signaling.

PubMed

Choi, Hyowon; Merceron, Christophe; Mangiavini, Laura; Seifert, Erin L; Schipani, Ernestina; Shapiro, Irving M; Risbud, Makarand V

2016-09-01

Nucleus pulposus (NP) cells reside in the avascular and hypoxic microenvironment of intervertebral discs. Importantly, many activities related to survival and function of NP cells are controlled by the HIF-family of transcription factors. We hypothesize that NP cells adapt to their hypoxic niche through modulation of macroautophagy/autophagy. In various cell types, hypoxia induces autophagy in a HIF1A-dependent fashion; however, little is known about hypoxic regulation of autophagy in NP cells. Hypoxia increases the number of autophagosomes as seen by TEM analysis and LC3-positive puncta in NP cells. Hypoxic induction of autophagy was also demonstrated by a significantly higher number of autophagosomes and smaller change in autolysosomes in NP cells expressing tandem-mCherry-EGFP-LC3B. Increased LC3-II levels were not accompanied by a concomitant increase in BECN1 or the ATG12-ATG5 complex. In addition, ULK1 phosphorylation at Ser757 and Ser777 responsive to MTOR and AMPK, respectively, was not affected in hypoxia. Interestingly, when MTOR activity was inhibited by rapamycin or Torin1, LC3-II levels did not change, suggesting a novel MTOR-independent regulation. Noteworthy, while silencing of HIF1A affected hypoxic induction of BNIP3, it did not affect LC3-II levels, indicating hypoxia-induced autophagy is HIF1-independent. Importantly, there was no change in the number of LC3-positive autophagosomes in NP-specific Hif1a null mice. Finally, inhibition of autophagic flux did not affect the glycolytic metabolism of NP cells, suggesting a possible nonmetabolic role of autophagy. Taken together, our study for the first time shows that NP cells regulate autophagy in a noncanonical fashion independent of MTOR and HIF1A signaling.

Influence and timing of arrival of murine neural crest on pancreatic beta cell development and maturation.

PubMed

Plank, Jennifer L; Mundell, Nathan A; Frist, Audrey Y; LeGrone, Alison W; Kim, Thomas; Musser, Melissa A; Walter, Teagan J; Labosky, Patricia A

2011-01-15

Interactions between cells from the ectoderm and mesoderm influence development of the endodermally-derived pancreas. While much is known about how mesoderm regulates pancreatic development, relatively little is understood about how and when the ectodermally-derived neural crest regulates pancreatic development and specifically, beta cell maturation. A previous study demonstrated that signals from the neural crest regulate beta cell proliferation and ultimately, beta cell mass. Here, we expand on that work to describe timing of neural crest arrival at the developing pancreatic bud and extend our knowledge of the non-cell autonomous role for neural crest derivatives in the process of beta cell maturation. We demonstrated that murine neural crest entered the pancreatic mesenchyme between the 26 and 27 somite stages (approximately 10.0 dpc) and became intermingled with pancreatic progenitors as the epithelium branched into the surrounding mesenchyme. Using a neural crest-specific deletion of the Forkhead transcription factor Foxd3, we ablated neural crest cells that migrate to the pancreatic primordium. Consistent with previous data, in the absence of Foxd3, and therefore the absence of neural crest cells, proliferation of insulin-expressing cells and insulin-positive area are increased. Analysis of endocrine cell gene expression in the absence of neural crest demonstrated that, although the number of insulin-expressing cells was increased, beta cell maturation was significantly impaired. Decreased MafA and Pdx1 expression illustrated the defect in beta cell maturation; we discovered that without neural crest, there was a reduction in the percentage of insulin-positive cells that co-expressed Glut2 and Pdx1 compared to controls. In addition, transmission electron microscopy analyses revealed decreased numbers of characteristic insulin granules and the presence of abnormal granules in insulin-expressing cells from mutant embryos. Together, these data demonstrate that the neural crest is a critical regulator of beta cell development on two levels: by negatively regulating beta cell proliferation and by promoting beta cell maturation. Copyright © 2010 Elsevier Inc. All rights reserved.

Professional regulation: a potentially valuable tool in responding to "stem cell tourism".

PubMed

Zarzeczny, Amy; Caulfield, Timothy; Ogbogu, Ubaka; Bell, Peter; Crooks, Valorie A; Kamenova, Kalina; Master, Zubin; Rachul, Christen; Snyder, Jeremy; Toews, Maeghan; Zoeller, Sonja

2014-09-09

The growing international market for unproven stem cell-based interventions advertised on a direct-to-consumer basis over the internet ("stem cell tourism") is a source of concern because of the risks it presents to patients as well as their supporters, domestic health care systems, and the stem cell research field. Emerging responses such as public and health provider-focused education and national regulatory efforts are encouraging, but the market continues to grow. Physicians play a number of roles in the stem cell tourism market and, in many jurisdictions, are members of a regulated profession. In this article, we consider the use of professional regulation to address physician involvement in stem cell tourism. Although it is not without its limitations, professional regulation is a potentially valuable tool that can be employed in response to problematic types of physician involvement in the stem cell tourism market. Copyright © 2014 The Authors. Published by Elsevier Inc. All rights reserved.

Hippocampal cell proliferation regulation by repeated stress and antidepressants.

PubMed

Chen, Hu; Pandey, Ghanshyam N; Dwivedi, Yogesh

2006-06-26

A recent hypothesis suggests reduced hippocampal neurogenesis in depression. Here, we examined cell proliferation in the dentate gyrus and the subventricular zone of rats given repeated stress, a paradigm that prolongs learned helplessness behavior, and whether antidepressants modulate the learned helplessness-associated altered cell proliferation. Decreased cell proliferation, number of clusters, and cells/cluster were noted in the dentate gyrus, but not in the subventricular zone, of learned helplessness rats. Both fluoxetine and desipramine reversed the learned helplessness behavior and increased the cell proliferation and the number of clusters in learned helplessness rats; only fluoxetine did so significantly. Both fluoxetine and desipramine significantly increased the number of cells/cluster. Our results suggest modified hippocampal neurogenesis in prolonged depression and in the mechanism of antidepressant action.

Compensatory regulation of Na+ absorption by Na+/H+ exchanger and Na+-Cl- cotransporter in zebrafish (Danio rerio)

PubMed Central

2013-01-01

Introduction In mammals, internal Na+ homeostasis is maintained through Na+ reabsorption via a variety of Na+ transport proteins with mutually compensating functions, which are expressed in different segments of the nephrons. In zebrafish, Na+ homeostasis is achieved mainly through the skin/gill ionocytes, namely Na+/H+ exchanger (NHE3b)-expressing H+-ATPase rich (HR) cells and Na+-Cl- cotransporter (NCC)-expressing NCC cells, which are functionally homologous to mammalian proximal and distal convoluted tubular cells, respectively. The present study aimed to investigate whether or not the functions of HR and NCC ionocytes are differentially regulated to compensate for disruptions of internal Na+ homeostasis and if the cell differentiation of the ionocytes is involved in this regulation pathway. Results Translational knockdown of ncc caused an increase in HR cell number and a resulting augmentation of Na+ uptake in zebrafish larvae, while NHE3b loss-of-function caused an increase in NCC cell number with a concomitant recovery of Na+ absorption. Environmental acid stress suppressed nhe3b expression in HR cells and decreased Na+ content, which was followed by up-regulation of NCC cells accompanied by recovery of Na+ content. Moreover, knockdown of ncc resulted in a significant decrease of Na+ content in acid-acclimated zebrafish. Conclusions These results provide evidence that HR and NCC cells exhibit functional redundancy in Na+ absorption, similar to the regulatory mechanisms in mammalian kidney, and suggest this functional redundancy is a critical strategy used by zebrafish to survive in a harsh environment that disturbs body fluid Na+ homeostasis. PMID:23924428

Regulatory considerations for pluripotent stem cell therapies.

PubMed

Carpenter, Melissa K

2017-01-01

The development of pluripotent stem cell (PSC) therapies is rapidly advancing, and a number of PSC-derived cell products are currently being tested in clinical trials. The biological complexity of these therapies results in specific challenges in complying with regulatory guidelines. This includes the choice of starting material, reproducible and consistent manufacturing, and preclinical safety and efficacy assessment of the PSC-derived product. This review discusses current US cell therapy regulations and strategies for compliance with these regulations when developing PSC-derived products. © 2017 Elsevier B.V. All rights reserved.

Bach2 Promotes Regulatory T-cell Development by Limiting Effector Differentiation | Center for Cancer Research

Cancer.gov

A number of allergic and autoimmune disorders, such as Crohn’s disease, asthma, type I diabetes and multiple sclerosis, are associated with polymorphisms in a gene encoding the transcription factor, BACH2. Despite this, the mechanism Bach2 uses to prevent immune-mediated diseases was not known. To function appropriately, the immune system relies on a delicate balance between immune-stimulating and immune-regulating cells. When immune-stimulating cells become too active, or immune-regulating cells become ineffective, autoimmune and allergic diseases can ensue.

Coherent organization in gene regulation: a study on six networks

NASA Astrophysics Data System (ADS)

Aral, Neşe; Kabakçıoğlu, Alkan

2016-04-01

Structural and dynamical fingerprints of evolutionary optimization in biological networks are still unclear. Here we analyze the dynamics of genetic regulatory networks responsible for the regulation of cell cycle and cell differentiation in three organisms or cell types each, and show that they follow a version of Hebb's rule which we have termed coherence. More precisely, we find that simultaneously expressed genes with a common target are less likely to act antagonistically at the attractors of the regulatory dynamics. We then investigate the dependence of coherence on structural parameters, such as the mean number of inputs per node and the activatory/repressory interaction ratio, as well as on dynamically determined quantities, such as the basin size and the number of expressed genes.

Hydrogen monitoring requirements in the global technical regulation on hydrogen and fuel cell vehicles

DOE PAGES

Buttner, William; Rivkin, C.; Burgess, R.; ...

2017-02-04

Here, the United Nations Economic Commission for Europe Global Technical Regulation (GTR) Number 13 ( Global Technical Regulation on Hydrogen and Fuel Cell Vehicles) is the defining document regulating safety requirements in hydrogen vehicles, and in particular, fuel cell electric vehicles (FCEVs). GTR Number 13 has been formally adopted and will serve as the basis for the national regulatory standards for FCEV safety in North America (led by the United States), Japan, Korea, and the European Union. The GTR defines safety requirements for these vehicles, including specifications on the allowable hydrogen levels in vehicle enclosures during in-use and post-crash conditionsmore » and on the allowable hydrogen emissions levels in vehicle exhaust during certain modes of normal operation. However, in order to be incorporated into national regulations, that is, to be legally binding, methods to verify compliance with the specific requirements must exist. In a collaborative program, the Sensor Laboratories at the National Renewable Energy Laboratory in the United States and the Joint Research Centre, Institute for Energy and Transport in the Netherlands have been evaluating and developing analytical methods that can be used to verify compliance with the hydrogen release requirements as specified in the GTR.« less

Purpose and regulation of stem cells: a systems-biology view from the Caenorhabditis elegans germ line.

PubMed

Cinquin, Olivier

2009-01-01

Stem cells are expected to play a key role in the development and maintenance of organisms, and hold great therapeutic promises. However, a number of questions must be answered to achieve an understanding of stem cells and put them to use. Here I review some of these questions, and how they relate to the model system provided by the Caenorhabditis elegans germ line, which is exceptional in its thorough genetic characterization and experimental accessibility under in vivo conditions. A fundamental question is how to define a stem cell; different definitions can be adopted that capture different features of interest. In the C. elegans germ line, stem cells can be defined by cell lineage or by cell commitment ('commitment' must itself be carefully defined). These definitions are associated with two other important questions about stem cells: their functions (which must be addressed following a systems approach, based on an evolutionary perspective) and their regulation. I review possible functions and their evolutionary groundings, including genome maintenance and powerful regulation of cell proliferation and differentiation, and possible regulatory mechanisms, including asymmetrical division and control of transit amplification by a developmental timer. I draw parallels between Drosophila and C. elegans germline stem cells; such parallels raise intriguing questions about Drosophila stem cells. I conclude by showing that the C. elegans germ line bears similarities with a number of other stem cell systems, which underscores its relevance to the understanding of stem cells.

Interferon-Gamma and Fas Are Involved in Porphyromonas gingivalis-Induced Apoptosis of Human Extravillous Trophoblast-Derived HTR8/SVneo Cells via Extracellular Signal-Regulated Kinase 1/2 Pathway.

PubMed

Ren, Hongyu; Li, Yuhong; Jiang, Han; Du, Minquan

2016-11-01

A number of studies recently revealed a link between periodontal disease and preterm birth (PTB). PTB can be induced by dental infection with Porphyromonas gingivalis (Pg), a periodontopathic bacterium. This study aims to investigate responses of human extravillous trophoblast-derived HTR8/SVneo cells to Pg infection. Cell apoptosis, cell viability, protein expression, and cytokine production in HTR8 cells were measured via: 1) flow cytometry, 2) CCK-8 assay, 3) western blot, and 4) enzyme-linked immunosorbent assay methods, respectively. Pg decreased cell viability and increased cell apoptosis, active caspase-3 and Fas expression, and interferon-gamma (IFN-γ) secretion in HTR8 cells. Extracellular signal-regulated kinase (ERK) 1/2 inhibitor U0126 and FasL neutralizing antibody NOK1 that blocks FasL/Fas interaction both significantly suppressed Pg-induced apoptosis. U0126 also inhibited IFN-γ secretion and Fas expression close to control levels. Moreover, treatment with recombinant IFN-γ also significantly decreased number of viable HTR8 cells and increased Fas expression, suggesting IFN-γ may play an important role in Pg-induced apoptosis of HTR8 cells, at least partially through regulation of Fas expression. To the best of the authors' knowledge, this is the first study to demonstrate Pg induces IFN-γ secretion, Fas expression, and apoptosis in human extravillous trophoblast-derived HTR8/SVneo cells in an ERK1/2-dependent manner, and IFN-γ (explored by recombinant IFN-γ) and Fas are involved in Pg-induced apoptosis. The finding that Pg infection abnormally regulates inflammation and apoptosis of human trophoblasts may give new insights into the possible link of PTB with maternal periodontal disease and periodontal pathogens.

Apoptosis in activated T cells: what are the triggers, and what the signal transducers?

PubMed

Häcker, Georg; Bauer, Anette; Villunger, Andreas

2006-11-01

At the end of an immune response, apoptosis drastically reduces the numbers of activated T cells. It has been a matter of intense research how this form of apoptosis is regulated and initiated, and a number of proteins have been identified that contribute to this process. The present, widely accepted model assumes that the interplay of pro- and anti -apoptotic Bcl-2 family members determines the onset of activated T cell death, with the BH3-only protein Bim activating pro-apoptotic Bax/Bak. In the search for up-stream signals, factors from other immune cells have been shown to play a role, and the NFkappaB family member Bcl-3 has been implicated as a signalling-intermediate in T cells. Recent work has tested the interrelation of these factors and has suggested that Bcl-3 acts as a regulator of Bim activation, that the induction of apoptosis through Bim can be complemented by its relative Puma, and that the presence of certain cytokines during T cell activation delays the activation of Bim and Puma. Here we discuss these recent insights and provide a view on how the regulation of activated T cell death is achieved and how extrinsic signals may translate into the activation of the apoptotic pathway.

Redox-dependent transcriptional regulation.

PubMed

Liu, Hongjun; Colavitti, Renata; Rovira, Ilsa I; Finkel, Toren

2005-11-11

Reactive oxygen species contribute to the pathogenesis of a number of disparate disorders including tissue inflammation, heart failure, hypertension, and atherosclerosis. In response to oxidative stress, cells activate expression of a number of genes, including those required for the detoxification of reactive molecules as well as for the repair and maintenance of cellular homeostasis. In many cases, these induced genes are regulated by transcription factors whose structure, subcellular localization, or affinity for DNA is directly or indirectly regulated by the level of oxidative stress. This review summarizes the recent progress on how cellular redox status can regulate transcription-factor activity and the implications of this regulation for cardiovascular disease.

Transcriptional control of stem cell fate by E2Fs and pocket proteins

PubMed Central

Julian, Lisa M.; Blais, Alexandre

2015-01-01

E2F transcription factors and their regulatory partners, the pocket proteins (PPs), have emerged as essential regulators of stem cell fate control in a number of lineages. In mammals, this role extends from both pluripotent stem cells to those encompassing all embryonic germ layers, as well as extra-embryonic lineages. E2F/PP-mediated regulation of stem cell decisions is highly evolutionarily conserved, and is likely a pivotal biological mechanism underlying stem cell homeostasis. This has immense implications for organismal development, tissue maintenance, and regeneration. In this article, we discuss the roles of E2F factors and PPs in stem cell populations, focusing on mammalian systems. We discuss emerging findings that position the E2F and PP families as widespread and dynamic epigenetic regulators of cell fate decisions. Additionally, we focus on the ever expanding landscape of E2F/PP target genes, and explore the possibility that E2Fs are not simply regulators of general ‘multi-purpose’ cell fate genes but can execute tissue- and cell type-specific gene regulatory programs. PMID:25972892

Regulation of cell wall biosynthesis.

PubMed

Zhong, Ruiqin; Ye, Zheng-Hua

2007-12-01

Plant cell walls differ in their amount and composition among various cell types and even in different microdomains of the wall of a given cell. Plants must have evolved regulatory mechanisms controlling biosynthesis, targeted secretion, and assembly of wall components to achieve the heterogeneity in cell walls. A number of factors, including hormones, the cytoskeleton, glycosylphosphatidylinositol-anchored proteins, phosphoinositides, and sugar nucleotide supply, have been implicated in the regulation of cell wall biosynthesis or deposition. In the past two years, there have been important discoveries in transcriptional regulation of secondary wall biosynthesis. Several transcription factors in the NAC and MYB families have been shown to be the key switches for activation of secondary wall biosynthesis. These studies suggest a transcriptional network comprised of a hierarchy of transcription factors is involved in regulating secondary wall biosynthesis. Further investigation and integration of the regulatory players participating in the making of cell walls will certainly lead to our understanding of how wall amounts and composition are controlled in a given cell type. This may eventually allow custom design of plant cell walls on the basis of our needs.

Drosophila Activin- and the Activin-like product Dawdle function redundantly to regulate proliferation in the larval brain.

PubMed

Zhu, Changqi C; Boone, Jason Q; Jensen, Philip A; Hanna, Scott; Podemski, Lynn; Locke, John; Doe, Chris Q; O'Connor, Michael B

2008-02-01

The Drosophila Activin-like ligands Activin-beta and Dawdle control several aspects of neuronal morphogenesis, including mushroom body remodeling, dorsal neuron morphogenesis and motoneuron axon guidance. Here we show that the same two ligands act redundantly through the Activin receptor Babo and its transcriptional mediator Smad2 (Smox), to regulate neuroblast numbers and proliferation rates in the developing larval brain. Blocking this pathway results in the development of larvae with small brains and aberrant photoreceptor axon targeting, and restoring babo function in neuroblasts rescued these mutant phenotypes. These results suggest that the Activin signaling pathway is required for producing the proper number of neurons to enable normal connection of incoming photoreceptor axons to their targets. Furthermore, as the Activin pathway plays a key role in regulating propagation of mouse and human embryonic stem cells, our observation that it also regulates neuroblast numbers and proliferation in Drosophila suggests that involvement of Activins in controlling stem cell propagation may be a common regulatory feature of this family of TGF-beta-type ligands.

Reverse-engineering of gene networks for regulating early blood development from single-cell measurements.

PubMed

Wei, Jiangyong; Hu, Xiaohua; Zou, Xiufen; Tian, Tianhai

2017-12-28

Recent advances in omics technologies have raised great opportunities to study large-scale regulatory networks inside the cell. In addition, single-cell experiments have measured the gene and protein activities in a large number of cells under the same experimental conditions. However, a significant challenge in computational biology and bioinformatics is how to derive quantitative information from the single-cell observations and how to develop sophisticated mathematical models to describe the dynamic properties of regulatory networks using the derived quantitative information. This work designs an integrated approach to reverse-engineer gene networks for regulating early blood development based on singel-cell experimental observations. The wanderlust algorithm is initially used to develop the pseudo-trajectory for the activities of a number of genes. Since the gene expression data in the developed pseudo-trajectory show large fluctuations, we then use Gaussian process regression methods to smooth the gene express data in order to obtain pseudo-trajectories with much less fluctuations. The proposed integrated framework consists of both bioinformatics algorithms to reconstruct the regulatory network and mathematical models using differential equations to describe the dynamics of gene expression. The developed approach is applied to study the network regulating early blood cell development. A graphic model is constructed for a regulatory network with forty genes and a dynamic model using differential equations is developed for a network of nine genes. Numerical results suggests that the proposed model is able to match experimental data very well. We also examine the networks with more regulatory relations and numerical results show that more regulations may exist. We test the possibility of auto-regulation but numerical simulations do not support the positive auto-regulation. In addition, robustness is used as an importantly additional criterion to select candidate networks. The research results in this work shows that the developed approach is an efficient and effective method to reverse-engineer gene networks using single-cell experimental observations.

Regulation of NK92-MI cell cytotoxicity by substance P.

PubMed

Fu, W X; Qin, B; Zhou, A P; Yu, Q Y; Huang, Q J; Liang, Z F

2011-08-01

The neuropeptide substance P (SP) can regulate a number of immunological functions in vitro and in vivo and may regulate natural killer (NK) cell activity. Here, we investigated whether SP has a role in regulating NK92-MI cell function in vitro, and how it influences NK cell activity. We found that SP dose dependently increased the cytotoxicity of NK92-MI cells and had a maximal effect at a concentration of 10(-12) and 10(-10) m. Furthermore, the expression of cytotoxic-associated molecules (perforin, granzyme) and activating receptor NKp46 [a member of natural cytotoxicity receptors (NCRs)] was observed to be upregulated by SP at optimal concentration, at which SP enhanced the cytotoxicity of NK92-MI cells. Neurokinin-1 receptor (NK-1R), a functional receptor of SP, was found on NK92-MI cells, and the observed effects of SP on NK92-MI cells could be more partially blocked by an NK-1R antagonist. Our data suggest that SP induces NK92-MI cell cytotoxicity by directly increasing the expression of cytotoxic granules and upregulates NK92-MI cell receptor-mediated functions indirectly. Thus, SP may regulate NK cell function mainly through NK-1R. © 2011 The Authors. Scandinavian Journal of Immunology © 2011 Blackwell Publishing Ltd.

Building strong bones: molecular regulation of the osteoblast lineage.

PubMed

Long, Fanxin

2011-12-22

The past 15 years have witnessed tremendous progress in the molecular understanding of osteoblasts, the main bone-forming cells in the vertebrate skeleton. In particular, all of the major developmental signals (including WNT and Notch signalling), along with an increasing number of transcription factors (such as RUNX2 and osterix), have been shown to regulate the differentiation and/or function of osteoblasts. As evidence indicates that osteoblasts may also regulate the behaviour of other cell types, a clear understanding of the molecular identity and regulation of osteoblasts is important beyond the field of bone biology.

Synaptotagmin 4 Regulates Pancreatic β Cell Maturation by Modulating the Ca2+ Sensitivity of Insulin Secretion Vesicles.

PubMed

Huang, Chen; Walker, Emily M; Dadi, Prasanna K; Hu, Ruiying; Xu, Yanwen; Zhang, Wenjian; Sanavia, Tiziana; Mun, Jisoo; Liu, Jennifer; Nair, Gopika G; Tan, Hwee Yim Angeline; Wang, Sui; Magnuson, Mark A; Stoeckert, Christian J; Hebrok, Matthias; Gannon, Maureen; Han, Weiping; Stein, Roland; Jacobson, David A; Gu, Guoqiang

2018-05-07

Islet β cells from newborn mammals exhibit high basal insulin secretion and poor glucose-stimulated insulin secretion (GSIS). Here we show that β cells of newborns secrete more insulin than adults in response to similar intracellular Ca 2+ concentrations, suggesting differences in the Ca 2+ sensitivity of insulin secretion. Synaptotagmin 4 (Syt4), a non-Ca 2+ binding paralog of the β cell Ca 2+ sensor Syt7, increased by ∼8-fold during β cell maturation. Syt4 ablation increased basal insulin secretion and compromised GSIS. Precocious Syt4 expression repressed basal insulin secretion but also impaired islet morphogenesis and GSIS. Syt4 was localized on insulin granules and Syt4 levels inversely related to the number of readily releasable vesicles. Thus, transcriptional regulation of Syt4 affects insulin secretion; Syt4 expression is regulated in part by Myt transcription factors, which repress Syt4 transcription. Finally, human SYT4 regulated GSIS in EndoC-βH1 cells, a human β cell line. These findings reveal the role that altered Ca 2+ sensing plays in regulating β cell maturation. Copyright © 2018 Elsevier Inc. All rights reserved.

β1 integrin is a crucial regulator of pancreatic β-cell expansion

PubMed Central

Diaferia, Giuseppe R.; Jimenez-Caliani, Antonio J.; Ranjitkar, Prerana; Yang, Wendy; Hardiman, Gary; Rhodes, Christopher J.; Crisa, Laura; Cirulli, Vincenzo

2013-01-01

Development of the endocrine compartment of the pancreas, as represented by the islets of Langerhans, occurs through a series of highly regulated events encompassing branching of the pancreatic epithelium, delamination and differentiation of islet progenitors from ductal domains, followed by expansion and three-dimensional organization into islet clusters. Cellular interactions with the extracellular matrix (ECM) mediated by receptors of the integrin family are postulated to regulate key functions in these processes. Yet, specific events regulated by these receptors in the developing pancreas remain unknown. Here, we show that ablation of the β1 integrin gene in developing pancreatic β-cells reduces their ability to expand during embryonic life, during the first week of postnatal life, and thereafter. Mice lacking β1 integrin in insulin-producing cells exhibit a dramatic reduction of the number of β-cells to only ∼18% of wild-type levels. Despite the significant reduction in β-cell mass, these mutant mice are not diabetic. A thorough phenotypic analysis of β-cells lacking β1 integrin revealed a normal expression repertoire of β-cell markers, normal architectural organization within islet clusters, and a normal ultrastructure. Global gene expression analysis revealed that ablation of this ECM receptor in β-cells inhibits the expression of genes regulating cell cycle progression. Collectively, our results demonstrate that β1 integrin receptors function as crucial positive regulators of β-cell expansion. PMID:23863477

Role of T Cells in Malnutrition and Obesity

PubMed Central

Gerriets, Valerie A.; MacIver, Nancie J.

2014-01-01

Nutritional status is critically important for immune cell function. While obesity is characterized by inflammation that promotes metabolic syndrome including cardiovascular disease and insulin resistance, malnutrition can result in immune cell defects and increased risk of mortality from infectious diseases. T cells play an important role in the immune adaptation to both obesity and malnutrition. T cells in obesity have been shown to have an early and critical role in inducing inflammation, accompanying the accumulation of inflammatory macrophages in obese adipose tissue, which are known to promote insulin resistance. How T cells are recruited to adipose tissue and activated in obesity is a topic of considerable interest. Conversely, T cell number is decreased in malnourished individuals, and T cells in the setting of malnutrition have decreased effector function and proliferative capacity. The adipokine leptin, which is secreted in proportion to adipocyte mass, may have a key role in mediating adipocyte-T cell interactions in both obesity and malnutrition, and has been shown to promote effector T cell function and metabolism while inhibiting regulatory T cell proliferation. Additionally, key molecular signals are involved in T cell metabolic adaptation during nutrient stress; among them, the metabolic regulator AMP kinase and the mammalian target of rapamycin have critical roles in regulating T cell number, function, and metabolism. In summary, understanding how T cell number and function are altered in obesity and malnutrition will lead to better understanding of and treatment for diseases where nutritional status determines clinical outcome. PMID:25157251

Bone marrow adipocytes: a neglected target tissue for growth hormone.

PubMed

Gevers, Evelien F; Loveridge, Nigel; Robinson, Iain C A F

2002-10-01

Bone marrow (BM) contains numerous adipocytes. These share a common precursor with osteoblasts and chondrocytes, but their function is unknown. It is unclear what regulates the differentiation of these three different cell types, though their subsequent metabolic activity is under hormonal regulation. GH and estrogen stimulate bone growth and mineralization, by direct effects on chondrocytes and osteoblasts. GH also stimulates lipolysis in subcutaneous and visceral adipocytes. However, adipocytes in BM have largely been ignored as potential targets for GH or estrogen action. We have addressed this by measuring BM adipocyte number, perimeter and area as well as bone area and osteoblast activity in GH-deficient dwarf (dw/dw), normal, or ovariectomized (Ovx) rats, with or without GH, IGF-1, PTH, or estrogen treatment or high fat feeding. Marrow adipocyte numbers were increased 5-fold (P < 0.001) in dw/dw rats, and cell size was also increased by 20%. These values returned toward normal in dw/dw rats given GH but not when given IGF-1. Cancellous bone area and osteoblast number were significantly (P < 0.005) lower in dw/dw rats, though alkaline phosphatase (ALP) activity in individual osteoblasts was unchanged. GH treatment increased % osteoblast covered bone surface without affecting individual cell ALP activity. Ovariectomy in normal or dw/dw rats had no affect on marrow adipocyte number nor size, although estrogen treatment in ovariectomized (Ovx) normal rats did increase adipocyte number. Ovx decreased tibial cancellous bone area in normal rats (64%; P < 0.05) and decreased osteoblast ALP-activity (P < 0.01) but did not affect the percentage of osteoblast-covered bone surface. Estrogen replacement reversed these changes. While treatment with PTH by continuous sc infusion decreased cancellous bone (P < 0.05) and high fat feeding increased the size of BM adipocytes (P < 0.01), they did not affect BM adipocyte number. These results suggest that GH has a specific action on BM adipocytes that is not simply due to altered bone or fat metabolism. We conclude that the marrow adipocyte lineage is an important and specific target for GH action. The inverse relationship between adipocyte number and osteoblast covered bone surface, together with the well-known effects of GH on epiphysial chondrocytes leads us to propose that GH plays two important roles on cells of all three lineages. During differentiation, it regulates the numbers of each cell type that are maintained from the common precursor lineage. Subsequently it has cell-specific effects on the metabolic activities of the differentiated cells. In the case of marrow adipocytes, GH-dependent lipolysis could provide an important hormonally regulated local high energy source in bone.

mTORC2 regulates multiple aspects of NKT-cell development and function

PubMed Central

Sklarz, Tammarah; Guan, Peng; Gohil, Mercy; Cotton, Renee M.; Ge, Moyar Q.; Haczku, Angela; Das, Rupali; Jordan, Martha S.

2017-01-01

Invariant NKT (iNKT) cells bridge innate and adaptive immunity by rapidly secreting cytokines and lysing targets following TCR recognition of lipid antigens. Based on their ability to secrete IFN-γ, IL-4 and IL-17A, iNKT-cells are classified as NKT-1, NKT-2 and NKT-17 subsets, respectively. The molecular pathways regulating iNKT-cell fate are not fully defined. Recent studies implicate Rictor, a required component of mTORC2, in the development of select iNKT-cell subsets, however these reports are conflicting. To resolve these questions, we used Rictorfl/fl CD4cre+ mice and found that Rictor is required for NKT-17 cell development and normal iNKT-cell cytolytic function. Conversely, Rictor is not absolutely required for IL-4 and IFN-γ production as peripheral iNKT-cells make copious amounts of these cytokines. Overall iNKT-cell numbers are dramatically reduced in the absence of Rictor. We provide data indicating Rictor regulates cell survival as well as proliferation of developing and mature iNKT-cells. Thus, mTORC2 regulates multiple aspects of iNKT-cell development and function. PMID:28078715

A serine residue in ClC-3 links phosphorylation-dephosphorylation to chloride channel regulation by cell volume.

PubMed

Duan, D; Cowley, S; Horowitz, B; Hume, J R

1999-01-01

In many mammalian cells, ClC-3 volume-regulated chloride channels maintain a variety of normal cellular functions during osmotic perturbation. The molecular mechanisms of channel regulation by cell volume, however, are unknown. Since a number of recent studies point to the involvement of protein phosphorylation/dephosphorylation in the control of volume-regulated ionic transport systems, we studied the relationship between channel phosphorylation and volume regulation of ClC-3 channels using site-directed mutagenesis and patch-clamp techniques. In native cardiac cells and when overexpressed in NIH/3T3 cells, ClC-3 channels were opened by cell swelling or inhibition of endogenous PKC, but closed by PKC activation, phosphatase inhibition, or elevation of intracellular Ca2+. Site-specific mutational studies indicate that a serine residue (serine51) within a consensus PKC-phosphorylation site in the intracellular amino terminus of the ClC-3 channel protein represents an important volume sensor of the channel. These results provide direct molecular and pharmacological evidence indicating that channel phosphorylation/dephosphorylation plays a crucial role in the regulation of volume sensitivity of recombinant ClC-3 channels and their native counterpart, ICl.vol.

Phosphoinositides: Tiny Lipids With Giant Impact on Cell Regulation

PubMed Central

2013-01-01

Phosphoinositides (PIs) make up only a small fraction of cellular phospholipids, yet they control almost all aspects of a cell's life and death. These lipids gained tremendous research interest as plasma membrane signaling molecules when discovered in the 1970s and 1980s. Research in the last 15 years has added a wide range of biological processes regulated by PIs, turning these lipids into one of the most universal signaling entities in eukaryotic cells. PIs control organelle biology by regulating vesicular trafficking, but they also modulate lipid distribution and metabolism via their close relationship with lipid transfer proteins. PIs regulate ion channels, pumps, and transporters and control both endocytic and exocytic processes. The nuclear phosphoinositides have grown from being an epiphenomenon to a research area of its own. As expected from such pleiotropic regulators, derangements of phosphoinositide metabolism are responsible for a number of human diseases ranging from rare genetic disorders to the most common ones such as cancer, obesity, and diabetes. Moreover, it is increasingly evident that a number of infectious agents hijack the PI regulatory systems of host cells for their intracellular movements, replication, and assembly. As a result, PI converting enzymes began to be noticed by pharmaceutical companies as potential therapeutic targets. This review is an attempt to give an overview of this enormous research field focusing on major developments in diverse areas of basic science linked to cellular physiology and disease. PMID:23899561

Plasma cell output from germinal centers is regulated by signals from Tfh and stromal cells

PubMed Central

George, Laura A.; Acs, Andreas; Durrett, Russell E.

2018-01-01

Germinal centers (GCs) are the sites where B cells undergo affinity maturation. The regulation of cellular output from the GC is not well understood. Here, we show that from the earliest stages of the GC response, plasmablasts emerge at the GC–T zone interface (GTI). We define two main factors that regulate this process: Tfh-derived IL-21, which supports production of plasmablasts from the GC, and TNFSF13 (APRIL), which is produced by a population of podoplanin+ CD157high fibroblastic reticular cells located in the GTI that are also rich in message for IL-6 and chemokines CXCL12, CCL19, and CCL21. Plasmablasts in the GTI express the APRIL receptor TNFRSF13B (TACI), and blocking TACI interactions specifically reduces the numbers of plasmablasts appearing in the GTI. Plasma cells generated in the GTI may provide an early source of affinity-matured antibodies that may neutralize pathogens or provide feedback regulating GC B cell selection. PMID:29549115

Centriole maturation requires regulated Plk1 activity during two consecutive cell cycles.

PubMed

Kong, Dong; Farmer, Veronica; Shukla, Anil; James, Jana; Gruskin, Richard; Kiriyama, Shigeo; Loncarek, Jadranka

2014-09-29

Newly formed centrioles in cycling cells undergo a maturation process that is almost two cell cycles long before they become competent to function as microtubule-organizing centers and basal bodies. As a result, each cell contains three generations of centrioles, only one of which is able to form cilia. It is not known how this long and complex process is regulated. We show that controlled Plk1 activity is required for gradual biochemical and structural maturation of the centrioles and timely appendage assembly. Inhibition of Plk1 impeded accumulation of appendage proteins and appendage formation. Unscheduled Plk1 activity, either in cycling or interphase-arrested cells, accelerated centriole maturation and appendage and cilia formation on the nascent centrioles, erasing the age difference between centrioles in one cell. These findings provide a new understanding of how the centriole cycle is regulated and how proper cilia and centrosome numbers are maintained in the cells.

Centriole maturation requires regulated Plk1 activity during two consecutive cell cycles

PubMed Central

Kong, Dong; Farmer, Veronica; Shukla, Anil; James, Jana; Gruskin, Richard; Kiriyama, Shigeo

2014-01-01

Newly formed centrioles in cycling cells undergo a maturation process that is almost two cell cycles long before they become competent to function as microtubule-organizing centers and basal bodies. As a result, each cell contains three generations of centrioles, only one of which is able to form cilia. It is not known how this long and complex process is regulated. We show that controlled Plk1 activity is required for gradual biochemical and structural maturation of the centrioles and timely appendage assembly. Inhibition of Plk1 impeded accumulation of appendage proteins and appendage formation. Unscheduled Plk1 activity, either in cycling or interphase-arrested cells, accelerated centriole maturation and appendage and cilia formation on the nascent centrioles, erasing the age difference between centrioles in one cell. These findings provide a new understanding of how the centriole cycle is regulated and how proper cilia and centrosome numbers are maintained in the cells. PMID:25246616

Transcriptional and posttranscriptional regulation of class I major histocompatibility complex genes following transformation with human adenoviruses.

PubMed Central

Shemesh, J; Rotem-Yehudar, R; Ehrlich, R

1991-01-01

Transformation of rodent cells by human adenoviruses is a well-established model system for studying the expression, regulation, and function of class I antigens. In this report, we demonstrate that the highly oncogenic adenovirus type 12 operates at the transcriptional and posttranscriptional levels in regulating the activity of major histocompatibility complex class I genes and products in transformed cells. Adenovirus type 12 suppresses the cell surface expression of class I antigens in most cell lines. Nevertheless, in a number of cell lines suppression is the result of reduction in the amount of stable specific mRNA, while in another group of cell lines suppression involves interference with processing of a posttranscriptional product. The two mechanisms operate both for the endogenous H-2 genes and for a miniature swine class I transgene that is expressed in the cells. Images PMID:1895404

Multi-gene fluorescence in situ hybridization to detect cell cycle gene copy number aberrations in young breast cancer patients

PubMed Central

Li, Chunyan; Bai, Jingchao; Hao, Xiaomeng; Zhang, Sheng; Hu, Yunhui; Zhang, Xiaobei; Yuan, Weiping; Hu, Linping; Cheng, Tao; Zetterberg, Anders; Lee, Mong-Hong; Zhang, J

2014-01-01

Breast cancer is a disease of cell cycle, and the dysfunction of cell cycle checkpoints plays a vital role in the occurrence and development of breast cancer. We employed multi-gene fluorescence in situ hybridization (M-FISH) to investigate gene copy number aberrations (CNAs) of 4 genes (Rb1, CHEK2, c-Myc, CCND1) that are involved in the regulation of cell cycle, in order to analyze the impact of gene aberrations on prognosis in the young breast cancer patients. Gene copy number aberrations of these 4 genes were more frequently observed in young breast cancer patients when compared with the older group. Further, these CNAs were more frequently seen in Luminal B type, Her2 overexpression, and tiple-negative breast cancer (TNBC) type in young breast cancer patients. The variations of CCND1, Rb1, and CHEK2 were significantly correlated with poor survival in the young breast cancer patient group, while the amplification of c-Myc was not obviously correlated with poor survival in young breast cancer patients. Thus, gene copy number aberrations (CNAs) of cell cycle-regulated genes can serve as an important tool for prognosis in young breast cancer patients. PMID:24621502

Dynein light chain regulates adaptive and innate B cell development by distinctive genetic mechanisms

PubMed Central

King, Ashleigh; Li, Lingli; Wong, David M.; Liu, Rui; Bamford, Rebecca; Strasser, Andreas

2017-01-01

Mechanistic differences in the development and function of adaptive, high-affinity antibody-producing B-2 cells and innate-like, “natural” antibody-producing B-1a cells remain poorly understood. Here we show that the multi-functional dynein light chain (DYNLL1/LC8) plays important roles in the establishment of B-1a cells in the peritoneal cavity and in the ongoing development of B-2 lymphoid cells in the bone marrow of mice. Epistasis analyses indicate that Dynll1 regulates B-1a and early B-2 cell development in a single, linear pathway with its direct transcriptional activator ASCIZ (ATMIN/ZNF822), and that the two genes also have complementary functions during late B-2 cell development. The B-2 cell defects caused by loss of DYNLL1 were associated with lower levels of the anti-apoptotic protein BCL-2, and could be supressed by deletion of pro-apoptotic BIM which is negatively regulated by both DYNLL1 and BCL-2. Defects in B cell development caused by loss of DYNLL1 could also be partially suppressed by a pre-arranged SWHEL Igm-B cell receptor transgene. In contrast to the rescue of B-2 cell numbers, the B-1a cell deficiency in Dynll1-deleted mice could not be suppressed by the loss of Bim, and was further compounded by the SWHEL transgene. Conversely, oncogenic MYC expression, which is synthetic lethal with Dynll1 deletion in B-2 cells, did not further reduce B-1a cell numbers in Dynll1-defcient mice. Finally, we found that the ASCIZ-DYNLL1 axis was also required for the early-juvenile development of aggressive MYC-driven and p53-deficient B cell lymphomas. These results identify ASCIZ and DYNLL1 as the core of a transcriptional circuit that differentially regulates the development of the B-1a and B-2 B lymphoid cell lineages and plays a critical role in lymphomagenesis. PMID:28922373

SRC-like adaptor protein regulates B cell development and function.

PubMed

Dragone, Leonard L; Myers, Margaret D; White, Carmen; Sosinowski, Tomasz; Weiss, Arthur

2006-01-01

The avidity of BCRs and TCRs influences signal strength during processes of lymphocyte development. Avidity is determined by both the intrinsic affinity for Ag and surface levels of the Ag receptor. The Src-like adaptor protein (SLAP) is a regulator of TCR levels on thymocytes, and its deficiency alters thymocyte development. We hypothesized that SLAP, which is expressed in B cells, also is important in regulating BCR levels, signal strength, and B cell development. To test this hypothesis, we analyzed the B cell compartment in SLAP-deficient mice. We found increased splenic B cell numbers and decreased surface IgM levels on mature, splenic B cells deficient in SLAP. Immature bone marrow and splenic B cells from BCR-transgenic, SLAP-deficient mice were found to express higher surface levels of IgM. In contrast, mature splenic B cells from BCR-transgenic mice expressed decreased levels of surface BCR associated with decreased calcium flux and activation-induced markers, compared with controls. These data suggest that SLAP regulates BCR levels and signal strength during lymphocyte development.

AIRE is a critical spindle-associated protein in embryonic stem cells

PubMed Central

Gu, Bin; Lambert, Jean-Philippe; Cockburn, Katie; Gingras, Anne-Claude; Rossant, Janet

2017-01-01

Embryonic stem (ES) cells go though embryo-like cell cycles regulated by specialized molecular mechanisms. However, it is not known whether there are ES cell-specific mechanisms regulating mitotic fidelity. Here we showed that Autoimmune Regulator (Aire), a transcription coordinator involved in immune tolerance processes, is a critical spindle-associated protein in mouse ES(mES) cells. BioID analysis showed that AIRE associates with spindle-associated proteins in mES cells. Loss of function analysis revealed that Aire was important for centrosome number regulation and spindle pole integrity specifically in mES cells. We also identified the c-terminal LESLL motif as a critical motif for AIRE’s mitotic function. Combined maternal and zygotic knockout further revealed Aire’s critical functions for spindle assembly in preimplantation embryos. These results uncovered a previously unappreciated function for Aire and provide new insights into the biology of stem cell proliferation and potential new angles to understand fertility defects in humans carrying Aire mutations. DOI: http://dx.doi.org/10.7554/eLife.28131.001 PMID:28742026

PGE2 maintains self-renewal of human adult stem cells via EP2-mediated autocrine signaling and its production is regulated by cell-to-cell contact.

PubMed

Lee, Byung-Chul; Kim, Hyung-Sik; Shin, Tae-Hoon; Kang, Insung; Lee, Jin Young; Kim, Jae-Jun; Kang, Hyun Kyoung; Seo, Yoojin; Lee, Seunghee; Yu, Kyung-Rok; Choi, Soon Won; Kang, Kyung-Sun

2016-05-27

Mesenchymal stem cells (MSCs) possess unique immunomodulatory abilities. Many studies have elucidated the clinical efficacy and underlying mechanisms of MSCs in immune disorders. Although immunoregulatory factors, such as Prostaglandin E2 (PGE2), and their mechanisms of action on immune cells have been revealed, their effects on MSCs and regulation of their production by the culture environment are less clear. Therefore, we investigated the autocrine effect of PGE2 on human adult stem cells from cord blood or adipose tissue, and the regulation of its production by cell-to-cell contact, followed by the determination of its immunomodulatory properties. MSCs were treated with specific inhibitors to suppress PGE2 secretion, and proliferation was assessed. PGE2 exerted an autocrine regulatory function in MSCs by triggering E-Prostanoid (EP) 2 receptor. Inhibiting PGE2 production led to growth arrest, whereas addition of MSC-derived PGE2 restored proliferation. The level of PGE2 production from an equivalent number of MSCs was down-regulated via gap junctional intercellular communication. This cell contact-mediated decrease in PGE2 secretion down-regulated the suppressive effect of MSCs on immune cells. In conclusion, PGE2 produced by MSCs contributes to maintenance of self-renewal capacity through EP2 in an autocrine manner, and PGE2 secretion is down-regulated by cell-to-cell contact, attenuating its immunomodulatory potency.

Microarray analysis on gene regulation by estrogen, progesterone and tamoxifen in human endometrial stromal cells.

PubMed

Ren, Chun-E; Zhu, Xueqiong; Li, Jinping; Lyle, Christian; Dowdy, Sean; Podratz, Karl C; Byck, David; Chen, Hai-Bin; Jiang, Shi-Wen

2015-03-13

Epithelial stromal cells represent a major cellular component of human uterine endometrium that is subject to tight hormonal regulation. Through cell-cell contacts and/or paracrine mechanisms, stromal cells play a significant role in the malignant transformation of epithelial cells. We isolated stromal cells from normal human endometrium and investigated the morphological and transcriptional changes induced by estrogen, progesterone and tamoxifen. We demonstrated that stromal cells express appreciable levels of estrogen and progesterone receptors and undergo different morphological changes upon hormonal stimulation. Microarray analysis indicated that both estrogen and progesterone induced dramatic alterations in a variety of genes associated with cell structure, transcription, cell cycle, and signaling. However, divergent patterns of changes, and in some genes opposite effects, were observed for the two hormones. A large number of genes are identified as novel targets for hormonal regulation. These hormone-responsive genes may be involved in normal uterine function and the development of endometrial malignancies.

Distribution of cocaine and amphetamine regulated transcript in ureters and urinary bladder of hypertensive rats.

PubMed

Janiuk, I; Kasacka, I

2013-01-01

Cocaine and amphetamine regulated transcript (CART), a neuropeptide of the central and peripheral nervous system plays an essential role in maintaining body homeostasis by regulating body temperature, orexia, digestive motility and blood pressure. Very few studies describe the relationship of hyperten¬sion with CART. Therefore, the present research was undertaken to identify, locate and determine the number of CART-immunopositive neuroendocrine cells (NE) and structures in the urinary bladder and ureter of rats with experimentally induced nephrogenic hypertension. The experiments were conducted on 20 Wistar rats in which hypertension was experimentally induced by applying a clamp on the left renal artery based on the two kidney, one clip experimental model (2K1C). After 6 weeks, fragments of the ureters and urinary bladder were sampled from rats with permanent hypertension. Immunohisto¬chemical analyses revealed a salient effect of renovascular hypertension on the neuroendocrine system of rat ureters and urinary bladder. Differences in the number of neuroendocrine cells and in the density of CART-positive structures were identified between the hypertensive and normotensive (control) rats. Hypertension greatly increased the number of NE cells and the density of CART- immunoreactive (IR) structures in the analysed urinary system organs.

Molecular control of brain size: Regulators of neural stem cell life, death and beyond

DOE Office of Scientific and Technical Information (OSTI.GOV)

Joseph, Bertrand; Hermanson, Ola, E-mail: ola.hermanson@ki.se

2010-05-01

The proper development of the brain and other organs depends on multiple parameters, including strictly controlled expansion of specific progenitor pools. The regulation of such expansion events includes enzymatic activities that govern the correct number of specific cells to be generated via an orchestrated control of cell proliferation, cell cycle exit, differentiation, cell death etc. Certain proteins in turn exert direct control of these enzymatic activities and thus progenitor pool expansion and organ size. The members of the Cip/Kip family (p21Cip1/p27Kip1/p57Kip2) are well-known regulators of cell cycle exit that interact with and inhibit the activity of cyclin-CDK complexes, whereas membersmore » of the p53/p63/p73 family are traditionally associated with regulation of cell death. It has however become clear that the roles for these proteins are not as clear-cut as initially thought. In this review, we discuss the roles for proteins of the Cip/Kip and p53/p63/p73 families in the regulation of cell cycle control, differentiation, and death of neural stem cells. We suggest that these proteins act as molecular interfaces, or 'pilots', to assure the correct assembly of protein complexes with enzymatic activities at the right place at the right time, thereby regulating essential decisions in multiple cellular events.« less

Regulation of tomato fruit pericarp development by an interplay between CDKB and CDKA1 cell cycle genes

PubMed Central

Czerednik, Anna; Busscher, Marco; Bielen, Bram A.M.; Wolters-Arts, Mieke; de Maagd, Ruud A.; Angenent, Gerco C.

2012-01-01

Growth of tomato fruits is determined by cell division and cell expansion, which are tightly controlled by factors that drive the core cell cycle. The cyclin-dependent kinases (CDKs) and their interacting partners, the cyclins, play a key role in the progression of the cell cycle. In this study the role of CDKA1, CDKB1, and CDKB2 in fruit development was characterized by fruit-specific overexpression and down-regulation. CDKA1 is expressed in the pericarp throughout development, but is strongly up-regulated in the outer pericarp cell layers at the end of the growth period, when CDKB gene expression has ceased. Overexpression of the CDKB genes at later stages of development and the down-regulation of CDKA1 result in a very similar fruit phenotype, showing a reduction in the number of cell layers in the pericarp and alterations in the desiccation of the fruits. Expression studies revealed that CDKA1 is down-regulated by the expression of CDKB1/2 in CDKB1 and CDKB2 overexpression mutants, suggesting opposite roles for these types of CDK proteins in tomato pericarp development. PMID:22282536

Coculture with endothelial cells reduces the population of cycling LeX neural precursors but increases that of quiescent cells with a side population phenotype

DOE Office of Scientific and Technical Information (OSTI.GOV)

Mathieu, Celine; Fouchet, Pierre; Gauthier, Laurent R.

2006-04-01

Neural stem cell proliferation and differentiation are regulated by external cues from their microenvironment. As endothelial cells are closely associated with neural stem cell in brain germinal zones, we investigated whether endothelial cells may interfere with neurogenesis. Neural precursor cells (NPC) from telencephalon of EGFP mouse embryos were cocultured in direct contact with endothelial cells. Endothelial cells did not modify the overall proliferation and apoptosis of neural cells, albeit they transiently delayed spontaneous apoptosis. These effects appeared to be specific to endothelial cells since a decrease in proliferation and a raise in apoptosis were observed in cocultures with fibroblasts. Endothelialmore » cells stimulated the differentiation of NPC into astrocytes and into neurons, whereas they reduced differentiation into oligodendrocytes in comparison to adherent cultures on polyornithine. Determination of NPC clonogenicity and quantification of LeX expression, a marker for NPC, showed that endothelial cells decreased the number of cycling NPC. On the other hand, the presence of endothelial cells increased the number of neural cells having 'side population' phenotype, another marker reported on NPC, which we have shown to contain quiescent cells. Thus, we show that endothelial cells may regulate neurogenesis by acting at different level of NPC differentiation, proliferation and quiescence.« less

Biophysical force regulation in 3D tumor cell invasion

NASA Astrophysics Data System (ADS)

Wu, Mingming

When embedded within 3D extracellular matrices (ECM), animal cells constantly probe and adapt to the ECM locally (at cell length scale) and exert forces and communicate with other cells globally (up to 10 times of cell length). It is now well accepted that mechanical crosstalk between animal cells and their microenvironment critically regulate cell function such as migration, proliferation and differentiation. Disruption of the cell-ECM crosstalk is implicated in a number of pathologic processes including tumor progression and fibrosis. Central to the problem of cell-ECM crosstalk is the physical force that cells generate. By measuring single cell generated force within 3D collagen matrices, we revealed a mechanical crosstalk mechanism between the tumor cells and the ECM. Cells generate sufficient force to stiffen collagen fiber network, and stiffer matrix, in return promotes larger cell force generation. Our work highlights the importance of fibrous nonlinear elasticity in regulating tumor cell-ECM interaction, and results may have implications in the rapid tissue stiffening commonly found in tumor progression and fibrosis. This work is partially supported by NIH Grants R21RR025801 and R21GM103388.

Telomere dynamics and homeostasis in a transmissible cancer.

PubMed

Ujvari, Beata; Pearse, Anne-Maree; Taylor, Robyn; Pyecroft, Stephen; Flanagan, Cassandra; Gombert, Sara; Papenfuss, Anthony T; Madsen, Thomas; Belov, Katherine

2012-01-01

Devil Facial Tumour Disease (DFTD) is a unique clonal cancer that threatens the world's largest carnivorous marsupial, the Tasmanian devil (Sarcophilus harrisii) with extinction. This transmissible cancer is passed between individual devils by cell implantation during social interactions. The tumour arose in a Schwann cell of a single devil over 15 years ago and since then has expanded clonally, without showing signs of replicative senescence; in stark contrast to a somatic cell that displays a finite capacity for replication, known as the "Hayflick limit". In the present study we investigate the role of telomere length, measured as Telomere Copy Number (TCN), and telomerase and shelterin gene expression, as well as telomerase activity in maintaining hyperproliferation of Devil Facial Tumour (DFT) cells. Our results show that DFT cells have short telomeres. DFTD TCN does not differ between geographic regions or between strains. However, TCN has increased over time. Unlimited cell proliferation is likely to have been achieved through the observed up-regulation of the catalytic subunit of telomerase (TERT) and concomitant activation of telomerase. Up-regulation of the central component of shelterin, the TRF1-intercating nuclear factor 2 (TINF2) provides DFT a mechanism for telomere length homeostasis. The higher expression of both TERT and TINF2 may also protect DFT cells from genomic instability and enhance tumour proliferation. DFT cells appear to monitor and regulate the length of individual telomeres: i.e. shorter telomeres are elongated by up-regulation of telomerase-related genes; longer telomeres are protected from further elongation by members of the shelterin complex, which may explain the lack of spatial and strain variation in DFT telomere copy number. The observed longitudinal increase in gene expression in DFT tissue samples and telomerase activity in DFT cell lines might indicate a selection for more stable tumours with higher proliferative potential.

Spdef null mice lack conjunctival goblet cells and provide a model of dry eye.

PubMed

Marko, Christina K; Menon, Balaraj B; Chen, Gang; Whitsett, Jeffrey A; Clevers, Hans; Gipson, Ilene K

2013-07-01

Goblet cell numbers decrease within the conjunctival epithelium in drying and cicatrizing ocular surface diseases. Factors regulating goblet cell differentiation in conjunctival epithelium are unknown. Recent data indicate that the transcription factor SAM-pointed domain epithelial-specific transcription factor (Spdef) is essential for goblet cell differentiation in tracheobronchial and gastrointestinal epithelium of mice. Using Spdef(-/-) mice, we determined that Spdef is required for conjunctival goblet cell differentiation and that Spdef(-/-) mice, which lack conjunctival goblet cells, have significantly increased corneal surface fluorescein staining and tear volume, a phenotype consistent with dry eye. Microarray analysis of conjunctival epithelium in Spdef(-/-) mice revealed down-regulation of goblet cell-specific genes (Muc5ac, Tff1, Gcnt3). Up-regulated genes included epithelial cell differentiation/keratinization genes (Sprr2h, Tgm1) and proinflammatory genes (Il1-α, Il-1β, Tnf-α), all of which are up-regulated in dry eye. Interestingly, four Wnt pathway genes were down-regulated. SPDEF expression was significantly decreased in the conjunctival epithelium of Sjögren syndrome patients with dry eye and decreased goblet cell mucin expression. These data demonstrate that Spdef is required for conjunctival goblet cell differentiation and down-regulation of SPDEF may play a role in human dry eye with goblet cell loss. Spdef(-/-) mice have an ocular surface phenotype similar to that in moderate dry eye, providing a new, more convenient model for the disease. Copyright © 2013 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

Role of Tat-interacting protein of 110 kDa and microRNAs in the regulation of hematopoiesis.

PubMed

Liu, Ying; He, Johnny J

2016-07-01

Hematopoiesis is regulated by cellular factors including transcription factors, microRNAs, and epigenetic modifiers. Understanding how these factors regulate hematopoiesis is pivotal for manipulating them to achieve their desired potential. In this review, we will focus on HIV-1 Tat-interacting protein of 110 kDa (Tip110) and its regulation of hematopoiesis. There are several pathways in hematopoiesis that involve Tip110 regulation. Tip110 is expressed in human cord blood CD34 cells; its expression decreases when CD34 cells begin to differentiate. Tip110 is also expressed in mouse marrow hematopoietic stem cells (HSC) and hematopoietic progenitor cells (HPC). Tip110 expression increases the number, survival, and cell cycling of HPC. Tip110-mediated regulation of hematopoiesis has been linked to its reciprocal control of proto-oncogene expression. Small noncoding microRNAs (miRs) have been shown to play important roles in regulation of hematopoiesis. miR-124 specifically targets 3'-untranslated region of Tip110 and subsequently regulates Tip110 expression in HSC. Our recent findings for manipulating expression levels of Tip110 in HSC and HPC could be useful for expanding HSC and HPC and for improving engraftment of cord blood HSC/HPC.

FoxO is a critical regulator of stem cell maintenance in immortal Hydra.

PubMed

Boehm, Anna-Marei; Khalturin, Konstantin; Anton-Erxleben, Friederike; Hemmrich, Georg; Klostermeier, Ulrich C; Lopez-Quintero, Javier A; Oberg, Hans-Heinrich; Puchert, Malte; Rosenstiel, Philip; Wittlieb, Jörg; Bosch, Thomas C G

2012-11-27

Hydra's unlimited life span has long attracted attention from natural scientists. The reason for that phenomenon is the indefinite self-renewal capacity of its stem cells. The underlying molecular mechanisms have yet to be explored. Here, by comparing the transcriptomes of Hydra's stem cells followed by functional analysis using transgenic polyps, we identified the transcription factor forkhead box O (FoxO) as one of the critical drivers of this continuous self-renewal. foxO overexpression increased interstitial stem cell and progenitor cell proliferation and activated stem cell genes in terminally differentiated somatic cells. foxO down-regulation led to an increase in the number of terminally differentiated cells, resulting in a drastically reduced population growth rate. In addition, it caused down-regulation of stem cell genes and antimicrobial peptide (AMP) expression. These findings contribute to a molecular understanding of Hydra's immortality, indicate an evolutionarily conserved role of FoxO in controlling longevity from Hydra to humans, and have implications for understanding cellular aging.

Stability of Control Networks in Autonomous Homeostatic Regulation of Stem Cell Lineages.

PubMed

Komarova, Natalia L; van den Driessche, P

2018-05-01

Design principles of biological networks have been studied extensively in the context of protein-protein interaction networks, metabolic networks, and regulatory (transcriptional) networks. Here we consider regulation networks that occur on larger scales, namely the cell-to-cell signaling networks that connect groups of cells in multicellular organisms. These are the feedback loops that orchestrate the complex dynamics of cell fate decisions and are necessary for the maintenance of homeostasis in stem cell lineages. We focus on "minimal" networks that are those that have the smallest possible numbers of controls. For such minimal networks, the number of controls must be equal to the number of compartments, and the reducibility/irreducibility of the network (whether or not it can be split into smaller independent sub-networks) is defined by a matrix comprised of the cell number increments induced by each of the controlled processes in each of the compartments. Using the formalism of digraphs, we show that in two-compartment lineages, reducible systems must contain two 1-cycles, and irreducible systems one 1-cycle and one 2-cycle; stability follows from the signs of the controls and does not require magnitude restrictions. In three-compartment systems, irreducible digraphs have a tree structure or have one 3-cycle and at least two more shorter cycles, at least one of which is a 1-cycle. With further work and proper biological validation, our results may serve as a first step toward an understanding of ways in which these networks become dysregulated in cancer.

Kid cleaves specific mRNAs at UUACU sites to rescue the copy number of plasmid R1

PubMed Central

Pimentel, Belén; Madine, Mark A; de la Cueva-Méndez, Guillermo

2005-01-01

Stability and copy number of extra-chromosomal elements are tightly regulated in prokaryotes and eukaryotes. Toxin Kid and antitoxin Kis are the components of the parD stability system of prokaryotic plasmid R1 and they can also function in eukaryotes. In bacteria, Kid was thought to become active only in cells that lose plasmid R1 and to cleave exclusively host mRNAs at UA(A/C/U) trinucleotide sites to eliminate plasmid-free cells. Instead, we demonstrate here that Kid becomes active in plasmid-containing cells when plasmid copy number decreases, cleaving not only host- but also a specific plasmid-encoded mRNA at the longer and more specific target sequence UUACU. This specific cleavage by Kid inhibits bacterial growth and, at the same time, helps to restore the plasmid copy number. Kid targets a plasmid RNA that encodes a repressor of the synthesis of an R1 replication protein, resulting in increased plasmid DNA replication. This mechanism resembles that employed by some human herpesviruses to regulate viral amplification during infection. PMID:16163387

Identification of Akt Interaction Protein PHF20/TZP That Transcriptionally Regulates p53*

PubMed Central

Park, Sungman; Kim, Donghwa; Dan, Han C.; Chen, Huihua; Testa, Joseph R.; Cheng, Jin Q.

2012-01-01

Akt regulates a diverse array of cellular functions, including cell survival, proliferation, differentiation, and metabolism. Although a number of molecules have been identified as upstream regulators and downstream targets of Akt, the mechanisms by which Akt regulates these cellular processes remain elusive. Here, we demonstrate that a novel transcription factor, PHF20/TZP (referring to Tudor and zinc finger domain containing protein), binds to Akt and induces p53 expression at the transcription level. Knockdown of PHF20 significantly reduces p53. PHF20 inhibits cell growth, DNA synthesis, and cell survival. Akt phosphorylates PHF20 at Ser291 in vitro and in vivo, which results in its translocation from the nucleus to the cytoplasm and attenuation of PHF20 function. These data indicate that PHF20 is a substrate of Akt and plays a role in Akt cell survival/growth signaling. PMID:22334668

Sequoia establishes tip-cell number in Drosophila trachea by regulating FGF levels.

PubMed

Araújo, Sofia J; Casanova, Jordi

2011-07-15

Competition and determination of leading and trailing cells during collective cell migration is a widespread phenomenon in development, wound healing and tumour invasion. Here, we analyse this issue during in vivo ganglionic branch cell migration in the Drosophila tracheal system. We identify Sequoia (Seq) as a negative transcriptional regulator of Branchless (Bnl), a Drosophila FGF homologue, and observe that modulation of Bnl levels determines how many cells will lead this migrating cluster, regardless of Notch lateral inhibition. Our results show that becoming a tip cell does not prevent others in the branch taking the same position, suggesting that leader choice does not depend only on sensing relative amounts of FGF receptor activity.

Tim-3 and PD-1 regulate CD8+ T cell function to maintain early pregnancy in mice

PubMed Central

XU, Yuan-Yuan; WANG, Song-Cun; LIN, Yi-Kong; LI, Da-Jin; DU, Mei-Rong

2017-01-01

During pregnancy, CD8+ T cells are important regulators in the balance of fetal tolerance and antiviral immunity. T-cell immunoglobulin mucin-3 (Tim-3) and programmed cell death-1 (PD-1) are well-recognized negative co-stimulatory molecules involved in viral persistence and tumor metastasis. Here, we demonstrate that CD8+ T cells co-expressing Tim-3 and PD-1 were down-regulated in the deciduae of female mice in abortion-prone matings compared with normal pregnant mice. In addition to their reduced numbers, the Tim-3+PD-1+CD8+ T cells produced lower levels of the anti-inflammatory cytokines interleukin (IL)-4 and IL-10, as well as a higher level of the pro-inflammatory cytokine interferon (IFN)-γ, relative to those from normal pregnancy. Furthermore, normal pregnant CBA/J females challenged with Tim-3- and/or PD-1-blocking antibodies were more susceptible to fetal resorption. These findings indicate that Tim-3 and PD-1 pathways play critical roles in regulating CD8+ T cell function and maintaining normal pregnancy. PMID:28331165

Budesonide suppresses pulmonary antibacterial host defense by down-regulating cathelicidin-related antimicrobial peptide in allergic inflammation mice and in lung epithelial cells

PubMed Central

2013-01-01

Background Glucocorticoids are widely regarded as the most effective treatment for asthma. However, the direct impact of glucocorticoids on the innate immune system and antibacterial host defense during asthma remain unclear. Understanding the mechanisms underlying this process is critical to the clinical application of glucocorticoids for asthma therapy. After sensitization and challenge with ovalbumin (OVA), BALB/c mice were treated with inhaled budesonide and infected with Pseudomonas aeruginosa (P. aeruginosa). The number of viable bacteria in enflamed lungs was evaluated, and levels of interleukin-4 (IL-4) and interferon-γ (IFN-γ) in serum were measured. A lung epithelial cell line was pretreated with budesonide. Levels of cathelicidin-related antimicrobial peptide (CRAMP) were measured by immunohistochemistry and western blot analysis. Intracellular bacteria were observed in lung epithelial cells. Results Inhaled budesonide enhanced lung infection in allergic mice exposed to P. aeruginosa and increased the number of viable bacteria in lung tissue. Higher levels of IL-4 and lower levels of IFN-γ were observed in the serum. Budesonide decreased the expression of CRAMP, increased the number of internalized P. aeruginosa in OVA-challenged mice and in lung epithelial cell lines. These data indicate that inhaled budesonide can suppress pulmonary antibacterial host defense by down-regulating CRAMP in allergic inflammation mice and in cells in vitro. Conclusions Inhaled budesonide suppressed pulmonary antibacterial host defense in an asthmatic mouse model and in lung epithelium cells in vitro. This effect was dependent on the down-regulation of CRAMP. PMID:23387852

Intermediate Filaments Play a Pivotal Role in Regulating Cell Architecture and Function*

PubMed Central

Lowery, Jason; Kuczmarski, Edward R.; Herrmann, Harald; Goldman, Robert D.

2015-01-01

Intermediate filaments (IFs) are composed of one or more members of a large family of cytoskeletal proteins, whose expression is cell- and tissue type-specific. Their importance in regulating the physiological properties of cells is becoming widely recognized in functions ranging from cell motility to signal transduction. IF proteins assemble into nanoscale biopolymers with unique strain-hardening properties that are related to their roles in regulating the mechanical integrity of cells. Furthermore, mutations in the genes encoding IF proteins cause a wide range of human diseases. Due to the number of different types of IF proteins, we have limited this short review to cover structure and function topics mainly related to the simpler homopolymeric IF networks composed of vimentin, and specifically for diseases, the related muscle-specific desmin IF networks. PMID:25957409

Characterization of the Th Profile of the Bovine Endometrium during the Oestrous Cycle and Early Pregnancy

PubMed Central

Oliveira, Lilian J.; Mansourri-Attia, Nadéra; Fahey, Alan G.; Browne, John; Forde, Niamh; Roche, James F.; Lonergan, Patrick; Fair, Trudee

2013-01-01

Despite extensive research in the area of cow fertility, the extent to which the maternal immune system is modulated during pregnancy in cattle remains unclear. Therefore, the objective of the current study was to characterize the presence and response profile of B, T-helper (LTh), T- cytotoxic (LTc), gamma delta-T (γδT) and natural killer (NK) lymphocytes in terms of cell number, distribution and cytokine expression in bovine endometrial tissue to pregnancy. Endometrial tissue samples were collected from beef heifers on Days 5, 7, 13 and 16 of the estrous cycle or pregnancy. Samples were analysed by immunofluorescence to identify the presence and abundance of B-B7 (B-cells), CD4 (LTh), CD8 (LTc), γδT cell receptor (TCR) and CD335/NKp46 (NK cells) -positive immune cells. Quantitative real time PCR (QPCR) was carried out to analyse mRNA relative abundance of FOXP3 (a marker of regulatory T (Treg) cells) and a panel of immune factors, including MHC-I, LIF, Interleukins 1, 2, 6, 8, 10, 11,12A, IFNa and IFNG. Results indicate that B-B7+ cells are quite populous in bovine endometrial tissue, CD4+ and CD8+ -cells are present in moderate numbers and γδTCR+ and CD335+ cells are present in low numbers. Pregnancy affected the total number and distribution pattern of the NK cell population, with the most significant variation observed on Day 16 of pregnancy. Neither B lymphocytes nor T lymphocyte subsets were regulated temporally during the oestrous cycle or by pregnancy prior to implantation. mRNA transcript abundance of the immune factors LIF, IL1b, IL8 and IL12A, IFNa and IFNG, expression was regulated temporally during the estrous cycle and LIF, IL1b, IL-10, IL11, IL12A were also temporally regulated during pregnancy. In conclusion, the endometrial immune profile of the oestrous cycle favours a Th2 environment in anticipation of pregnancy and the presence of an embryo acts to fine tune this environment. PMID:24204576

Somatic mutations and affinity maturation are impaired by excessive numbers of T follicular helper cells and restored by Treg cells or memory T cells.

PubMed

Preite, Silvia; Baumjohann, Dirk; Foglierini, Mathilde; Basso, Camilla; Ronchi, Francesca; Fernandez Rodriguez, Blanca M; Corti, Davide; Lanzavecchia, Antonio; Sallusto, Federica

2015-11-01

We previously reported that Cd3e-deficient mice adoptively transferred with CD4(+) T cells generate high numbers of T follicular helper (Tfh) cells, which go on to induce a strong B-cell and germinal center (GC) reaction. Here, we show that in this system, GC B cells display an altered distribution between the dark and light zones, and express low levels of activation-induced cytidine deaminase. Furthermore, GC B cells from Cd3e(-/-) mice accumulate fewer somatic mutations as compared with GC B cells from wild-type mice, and exhibit impaired affinity maturation and reduced differentiation into long-lived plasma cells. Reconstitution of Cd3e(-/-) mice with regulatory T (Treg) cells restored Tfh-cell numbers, GC B-cell numbers and B-cell distribution within dark and light zones, and the rate of antibody somatic mutations. Tfh-cell numbers and GC B-cell numbers and dynamics were also restored by pre-reconstitution of Cd3e(-/-) mice with Cxcr5(-/-) Treg cells or non-regulatory, memory CD4(+) T cells. Taken together, these findings underline the importance of a quantitatively regulated Tfh-cell response for an efficient and long-lasting serological response. © 2015 The Authors. European Journal of Immunology published by WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Regulation of insulin-like growth factor I receptors on vascular smooth muscle cells by growth factors and phorbol esters.

PubMed

Ververis, J J; Ku, L; Delafontaine, P

1993-06-01

Insulin-like growth factor I (IGF I) is an important mitogen for vascular smooth muscle cells. To characterize regulation of vascular IGF I receptors, we performed radioligand displacement experiments using rat aortic smooth muscle cells (RASMs). Serum deprivation for 48 hours caused a 40% decrease in IGF I receptor number. Exposure of quiescent RASMs to platelet-derived growth factor (PDGF), fibroblast growth factor (FGF), or angiotensin II (Ang II) caused a 1.5-2.0-fold increase in IGF I receptors per cell. After FGF exposure, there was a marked increase in the mitogenic response to IGF I. IGF I downregulated its receptors in the presence of platelet-poor plasma. Stimulation of protein kinase C (PKC) by exposure of quiescent RASMs to phorbol 12-myristate 13-acetate caused a biphasic response in IGF I binding; there was a 42% decrease in receptor number at 45 minutes and a 238% increase at 24 hours. To determine the role of PKC in growth factor-induced regulation of IGF I receptors, we downregulated PKC by exposing RASMs to phorbol 12,13-dibutyrate (PDBu) for 48 hours. PDGF- and FGF- but not Ang II-mediated upregulation of IGF I receptors was completely inhibited in PDBu-treated cells. Thus, acute PKC activation by phorbol esters inhibits IGF I binding, whereas chronic PKC activation increases IGF I binding. PDGF and FGF but not Ang II regulate vascular IGF I receptors through a PKC-dependent pathway. These data provide new insights into the regulation of vascular smooth muscle cell IGF I receptors in vitro and are of potential importance in characterizing vascular proliferative responses in vivo.

The Akt signaling pathway is required for tissue maintenance and regeneration in planarians.

PubMed

Peiris, T Harshani; Ramirez, Daniel; Barghouth, Paul G; Oviedo, Néstor J

2016-04-11

Akt (PKB) is a serine threonine protein kinase downstream of the phosphoinositide 3-kinase (PI3K) pathway. In mammals, Akt is ubiquitously expressed and is associated with regulation of cellular proliferation, metabolism, cell growth and cell death. Akt has been widely studied for its central role in physiology and disease, in particular cancer where it has become an attractive pharmacological target. However, the mechanisms by which Akt signaling regulates stem cell behavior in the complexity of the whole body are poorly understood. Planarians are flatworms with large populations of stem cells capable of dividing to support adult tissue renewal and regeneration. The planarian ortholog Smed-Akt is molecularly conserved providing unique opportunities to analyze the function of Akt during cellular turnover and repair of adult tissues. Our findings abrogating Smed-Akt with RNA-interference in the planarian Schmidtea mediterranea led to a gradual decrease in stem cell (neoblasts) numbers. The reduced neoblast numbers largely affected the maintenance of adult tissues including the nervous and excretory systems and ciliated structures in the ventral epithelia, which impaired planarian locomotion. Downregulation of Smed-Akt function also resulted in an increase of cell death throughout the animal. However, in response to amputation, levels of cell death were decreased and failed to localize near the injury site. Interestingly, the neoblast mitotic response was increased around the amputation area but the regenerative blastema failed to form. We demonstrate Akt signaling is essential for organismal physiology and in late stages of the Akt phenotype the reduction in neoblast numbers may impair regeneration in planarians. Functional disruption of Smed-Akt alters the balance between cell proliferation and cell death leading to systemic impairment of adult tissue renewal. Our results also reveal novel roles for Akt signaling during regeneration, specifically for the timely localization of cell death near the injury site. Thus, Akt signaling regulates neoblast biology and mediates in the distribution of injury-mediated cell death during tissue repair in planarians.

Pro-hormone Secretogranin II Regulates Dense Core Secretory Granule Biogenesis in Catecholaminergic Cells*

PubMed Central

Courel, Maïté; Soler-Jover, Alex; Rodriguez-Flores, Juan L.; Mahata, Sushil K.; Elias, Salah; Montero-Hadjadje, Maïté; Anouar, Youssef; Giuly, Richard J.; O'Connor, Daniel T.; Taupenot, Laurent

2010-01-01

Processes underlying the formation of dense core secretory granules (DCGs) of neuroendocrine cells are poorly understood. Here, we present evidence that DCG biogenesis is dependent on the secretory protein secretogranin (Sg) II, a member of the granin family of pro-hormone cargo of DCGs in neuroendocrine cells. Depletion of SgII expression in PC12 cells leads to a decrease in both the number and size of DCGs and impairs DCG trafficking of other regulated hormones. Expression of SgII fusion proteins in a secretory-deficient PC12 variant rescues a regulated secretory pathway. SgII-containing dense core vesicles share morphological and physical properties with bona fide DCGs, are competent for regulated exocytosis, and maintain an acidic luminal pH through the V-type H+-translocating ATPase. The granulogenic activity of SgII requires a pH gradient along this secretory pathway. We conclude that SgII is a critical factor for the regulation of DCG biogenesis in neuroendocrine cells, mediating the formation of functional DCGs via its pH-dependent aggregation at the trans-Golgi network. PMID:20061385

Myeloid Wnt ligands are required for normal development of dermal lymphatic vasculature.

PubMed

Muley, Ajit; Odaka, Yoshi; Lewkowich, Ian P; Vemaraju, Shruti; Yamaguchi, Terry P; Shawber, Carrie; Dickie, Belinda H; Lang, Richard A

2017-01-01

Resident tissue myeloid cells play a role in many aspects of physiology including development of the vascular systems. In the blood vasculature, myeloid cells use VEGFC to promote angiogenesis and can use Wnt ligands to control vascular branching and to promote vascular regression. Here we show that myeloid cells also regulate development of the dermal lymphatic vasculature using Wnt ligands. Using myeloid-specific deletion of the WNT transporter Wntless we show that myeloid Wnt ligands are active at two distinct stages of development of the dermal lymphatics. As lymphatic progenitors are emigrating from the cardinal vein and intersomitic vessels, myeloid Wnt ligands regulate both their numbers and migration distance. Later in lymphatic development, myeloid Wnt ligands regulate proliferation of lymphatic endothelial cells (LEC) and thus control lymphatic vessel caliber. Myeloid-specific deletion of WNT co-receptor Lrp5 or Wnt5a gain-of-function also produce elevated caliber in dermal lymphatic capillaries. These data thus suggest that myeloid cells produce Wnt ligands to regulate lymphatic development and use Wnt pathway co-receptors to regulate the balance of Wnt ligand activity during the macrophage-LEC interaction.

Pro-hormone secretogranin II regulates dense core secretory granule biogenesis in catecholaminergic cells.

PubMed

Courel, Maïté; Soler-Jover, Alex; Rodriguez-Flores, Juan L; Mahata, Sushil K; Elias, Salah; Montero-Hadjadje, Maïté; Anouar, Youssef; Giuly, Richard J; O'Connor, Daniel T; Taupenot, Laurent

2010-03-26

Processes underlying the formation of dense core secretory granules (DCGs) of neuroendocrine cells are poorly understood. Here, we present evidence that DCG biogenesis is dependent on the secretory protein secretogranin (Sg) II, a member of the granin family of pro-hormone cargo of DCGs in neuroendocrine cells. Depletion of SgII expression in PC12 cells leads to a decrease in both the number and size of DCGs and impairs DCG trafficking of other regulated hormones. Expression of SgII fusion proteins in a secretory-deficient PC12 variant rescues a regulated secretory pathway. SgII-containing dense core vesicles share morphological and physical properties with bona fide DCGs, are competent for regulated exocytosis, and maintain an acidic luminal pH through the V-type H(+)-translocating ATPase. The granulogenic activity of SgII requires a pH gradient along this secretory pathway. We conclude that SgII is a critical factor for the regulation of DCG biogenesis in neuroendocrine cells, mediating the formation of functional DCGs via its pH-dependent aggregation at the trans-Golgi network.

Ursolic Acid Attenuates Diabetic Mesangial Cell Injury through the Up-Regulation of Autophagy via miRNA-21/PTEN/Akt/mTOR Suppression

PubMed Central

Lu, Xinxing; Fan, Qiuling; Xu, Li; Li, Lin; Yue, Yuan; Xu, Yanyan; Su, Yan; Zhang, Dongcheng; Wang, Lining

2015-01-01

Objective To investigate the effect of ursolic acid on autophagy mediated through the miRNA-21-targeted phosphoinositide 3 kinase (PI3K)/protein kinase B (Akt)/mammalian target of rapamycin (mTOR) pathway in rat mesangial cells cultured under high glucose (HG) conditions. Methods Rat glomerular mesangial cells were cultured under normal glucose, HG, HG with the PI3K inhibitor LY294002 or HG with ursolic acid conditions. Cell proliferation and hypertrophy were assayed using an MTT assay and the ratio of total protein to cell number, respectively. The miRNA-21 expression was detected using RT-qPCR. The expression of phosphatase and tensin homolog (PTEN)/AKT/mTOR signaling signatures, autophagy-associated protein and collagen I was detected by western blotting and RT-qPCR. Autophagosomes were observed using electron microscopy. Results Compared with mesangial cells cultured under normal glucose conditions, the cells exposed to HG showed up-regulated miRNA-21 expression, down-regulated PTEN protein and mRNA expression, up-regulated p85PI3K, pAkt, pmTOR, p62/SQSTMI, and collagen I expression and down-regulated LC3II expression. Ursolic acid and LY294002 inhibited HG-induced mesangial cell hypertrophy and proliferation, down-regulated p85PI3K, pAkt, pmTOR, p62/SQSTMI, and collagen I expression and up-regulated LC3II expression. However, LY294002 did not affect the expression of miRNA-21 and PTEN. Ursolic acid down-regulated miRNA-21 expression and up-regulated PTEN protein and mRNA expression. Conclusions Ursolic acid inhibits the glucose-induced up-regulation of mesangial cell miRNA-21 expression, up-regulates PTEN expression, inhibits the activation of PI3K/Akt/mTOR signaling pathway, and enhances autophagy to reduce the accumulation of the extracellular matrix and ameliorate cell hypertrophy and proliferation. PMID:25689721

Translating Research into Clinical Scale Manufacturing of Mesenchymal Stromal Cells

PubMed Central

Bieback, Karen; Kinzebach, Sven; Karagianni, Marianna

2010-01-01

It sounds simple to obtain sufficient numbers of cells derived from fetal or adult human tissues, isolate and/or expand the stem cells, and then transplant an appropriate number of these cells into the patient at the correct location. However, translating basic research into routine therapies is a complex multistep process which necessitates product regulation. The challenge relates to managing the expected therapeutic benefits with the potential risks and to balance the fast move to clinical trials with time-consuming cautious risk assessment. This paper will focus on the definition of mesenchymal stromal cells (MSCs), and challenges and achievements in the manufacturing process enabling their use in clinical studies. It will allude to different cellular sources, special capacities of MSCs, but also to current regulations, with a special focus on accessory material of human or animal origin, like media supplements. As cellular integrity and purity, formulation and lot release testing of the final product, validation of all procedures, and quality assurance are of utmost necessity, these topics will be addressed. PMID:21318154

Regulation of Memory T Cells by Interleukin-23.

PubMed

Li, Yanchun; Wang, Hongbo; Lu, Honghua; Hua, Shucheng

2016-01-01

Interleukin-23 (IL-23), a member of the IL-12 family of cytokines, is a heterodimeric cytokine. It is composed of subunits p40 (shared with IL-12) and p19 (an IL-12 p35-related subunit) and is secreted by several types of immune cells, such as natural killer cells and dendritic cells. The IL-23 receptor is composed of the subunit IL-12Rβ1 and the IL-23-specific subunit IL-23R. The binding of IL-23 to its specific cell surface receptor regulates a number of functions, including proliferation and differentiation of cells and secretion of cell factors. Memory T cells are a subset of T cells that secrete numerous important cell factors, and they function in the immune response to infection and diseases like cancer, autoimmune disease and bronchial asthma. IL-23R is expressed on the surface of memory T cells, which suggests that it can specifically regulate memory T cell function. IL-23 has been widely used as a clinical indicator in immune-related diseases and shows potential for use in disease treatment. Here we review the current progress in the study of the role of IL-23 in the regulation of memory T cells. © 2016 S. Karger AG, Basel.

Yes-associated protein and WW-containing transcription regulator 1 regulate the expression of sex-determining genes in Sertoli cells, but their inactivation does not cause sex reversal.

PubMed

Levasseur, Adrien; Paquet, Marilène; Boerboom, Derek; Boyer, Alexandre

2017-07-01

Yes-associated protein (YAP) and WW-containing transcription regulator 1 (WWTR1) are two functionally redundant transcriptional regulators that are downstream effectors of the Hippo signaling pathway, and that act as major regulators of cell growth and differentiation. To elucidate their role in Sertoli cells, primary Sertoli cell culture from Yapflox/flox; Wwtr1flox/flox animals were infected with a Cre recombinase-expressing adenovirus. Concomitant inactivation of Yap and Wwtr1 resulted in a decrease in the mRNA levels of the male sex differentiation genes Dhh, Dmrt1, Sox9, and Wt1, whereas those of genes involved in female differentiation (Wnt4, Rspo1, and Foxl2) were induced. SOX9, FOXL2, and WNT4 proteins were regulated in the same manner as their mRNAs in response to loss of YAP and WWTR1. To further characterize the role of YAP and WWTR1 in Sertoli cells, we generated a mouse model (Yapflox/flox; Wwtr1flox/flox; Amhcre/+) in which Yap and Wwtr1 were conditionally deleted in Sertoli cells. An increase in the number of apoptotic cells was observed in the seminiferous tubules of 4 dpp mutant mice, leading to a reduction in testis weights and a decrease in the number of Sertoli cells in adult animals. Gene expression analyses of testes from 4 dpp Yapflox/flox; Wwtr1flox/flox; Amhcre/+ mice showed that Sertoli cell differentiation is initially altered, as Dhh, Dmrt1, and Sox9 mRNA levels were downregulated, whereas Wnt4 mRNA levels were increased. However, expression of these genes was not changed in older animals. Together, these results suggest a novel role of the Hippo signaling pathway in the mechanisms of sex differentiation. © The Authors 2017. Published by Oxford University Press on behalf of Society for the Study of Reproduction. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

A role for autophagic protein beclin 1 early in lymphocyte development.

PubMed

Arsov, Ivica; Adebayo, Adeola; Kucerova-Levisohn, Martina; Haye, Joanna; MacNeil, Margaret; Papavasiliou, F Nina; Yue, Zhenyu; Ortiz, Benjamin D

2011-02-15

Autophagy is a highly regulated and evolutionarily conserved process of cellular self-digestion. Recent evidence suggests that this process plays an important role in regulating T cell homeostasis. In this study, we used Rag1(-/-) (recombination activating gene 1(-/-)) blastocyst complementation and in vitro embryonic stem cell differentiation to address the role of Beclin 1, one of the key autophagic proteins, in lymphocyte development. Beclin 1-deficient Rag1(-/-) chimeras displayed a dramatic reduction in thymic cellularity compared with control mice. Using embryonic stem cell differentiation in vitro, we found that the inability to maintain normal thymic cellularity is likely caused by impaired maintenance of thymocyte progenitors. Interestingly, despite drastically reduced thymocyte numbers, the peripheral T cell compartment of Beclin 1-deficient Rag1(-/-) chimeras is largely normal. Peripheral T cells displayed normal in vitro proliferation despite significantly reduced numbers of autophagosomes. In addition, these chimeras had greatly reduced numbers of early B cells in the bone marrow compared with controls. However, the peripheral B cell compartment was not dramatically impacted by Beclin 1 deficiency. Collectively, our results suggest that Beclin 1 is required for maintenance of undifferentiated/early lymphocyte progenitor populations. In contrast, Beclin 1 is largely dispensable for the initial generation and function of the peripheral T and B cell compartments. This indicates that normal lymphocyte development involves Beclin 1-dependent, early-stage and distinct, Beclin 1-independent, late-stage processes.

Spdef Null Mice Lack Conjunctival Goblet Cells and Provide a Model of Dry Eye

PubMed Central

Marko, Christina K.; Menon, Balaraj B.; Chen, Gang; Whitsett, Jeffrey A.; Clevers, Hans; Gipson, Ilene K.

2014-01-01

Goblet cell numbers decrease within the conjunctival epithelium in drying and cicatrizing ocular surface diseases. Factors regulating goblet cell differentiation in conjunctival epithelium are unknown. Recent data indicate that the transcription factor SAM-pointed domain epithelial-specific transcription factor (Spdef) is essential for goblet cell differentiation in tracheobronchial and gastrointestinal epithelium of mice. Using Spdef−/− mice, we determined that Spdef is required for conjunctival goblet cell differentiation and that Spdef−/− mice, which lack conjunctival goblet cells, have significantly increased corneal surface fluorescein staining and tear volume, a phenotype consistent with dry eye. Microarray analysis of conjunctival epithelium in Spdef−/− mice revealed down-regulation of goblet cell–specific genes (Muc5ac, Tff1, Gcnt3). Up-regulated genes included epithelial cell differentiation/keratinization genes (Sprr2h, Tgm1) and proinflammatory genes (Il1-α, Il-1β, Tnf-α), all of which are up-regulated in dry eye. Interestingly, four Wnt pathway genes were down-regulated. SPDEF expression was significantly decreased in the conjunctival epithelium of Sjögren syndrome patients with dry eye and decreased goblet cell mucin expression. These data demonstrate that Spdef is required for conjunctival goblet cell differentiation and down-regulation of SPDEF may play a role in human dry eye with goblet cell loss. Spdef−/− mice have an ocular surface phenotype similar to that in moderate dry eye, providing a new, more convenient model for the disease. PMID:23665202

Density-dependent regulation of growth of BSC-1 cells in cell culture: control of growth by serum factors.

PubMed Central

Holley, R W; Armour, R; Baldwin, J H; Brown, K D; Yeh, Y C

1977-01-01

BSC-1 cells grow slowly, to high cell density, in medium with 0.1% calf serum. An increase in the serum concentration increases both the growth rate of the cells and the final cell density. The serum can be replaced to some extent by epidermal growth factor (EGF). Initiation of DNA synthesis in BSC-1 cells that have spread into a "wound" in a crowded cell layer requires the addition of a trace of serum or EGF, if the cells have previously been deprived of serum. The binding of 125I-labeled EGF to low-density and high-density BSC-1 cells has been studied. Binding is faster to low-density cells. Cells at low cell density also bind much more EGF per cell than cells at high cell density. The fraction of bound 125I-labeled EGF that is present on the cell surface as intact EGF is larger at low than at high cell density. The results indicate that the number of available EGF receptors per cell decreases drastically as the cell density increases. It is suggested that a decrease in the number of available EGF receptor sites per cell, and the accompanying decrease in sensitivity of the cells to EGF, contributes to density-dependent regulation of growth of these cells. Images PMID:303774

Differential signaling and regulation of apical vs. basolateral EGFR in polarized epithelial cells.

PubMed

Kuwada, S K; Lund, K A; Li, X F; Cliften, P; Amsler, K; Opresko, L K; Wiley, H S

1998-12-01

Overexpression of the epidermal growth factor receptors (EGFR) in polarized kidney epithelial cells caused them to appear in high numbers at both the basolateral and apical cell surfaces. We utilized these cells to look for differences in the regulation and signaling of apical vs. basolateral EGFR. Apical and basolateral EGFR were biologically active and mediated EGF-induced cell proliferation to similar degrees. Receptor downregulation and endocytosis were less efficient at the apical surface, resulting in prolonged EGF-induced tyrosine kinase activity at the apical cell membrane. Tyrosine phosphorylation of EGFR substrates known to mediate cell proliferation, Src-homologous and collagen protein (SHC), extracellularly regulated kinase 1 (ERK1), and ERK2 could be induced similarly by activation of apical or basolateral EGFR. Focal adhesion kinase was tyrosine phosphorylated more by basolateral than by apical EGFR; however, beta-catenin was tyrosine phosphorylated to a much greater degree following the activation of mislocalized apical EGFR. Thus EGFR regulation and EGFR-mediated phosphorylation of certain substrates differ at the apical and basolateral cell membrane domains. This suggests that EGFR mislocalization could result in abnormal signal transduction and aberrant cell behavior.

Hematopoietic Stem Cells: Transcriptional Regulation, Ex Vivo Expansion and Clinical Application

PubMed Central

Aggarwal, R.; Lu, J.; Pompili, V.J.; Das, H.

2012-01-01

Maintenance of ex vivo hematopoietic stem cells (HSC) pool and its differentiated progeny is regulated by complex network of transcriptional factors, cell cycle proteins, extracellular matrix, and their microenvironment through an orchestrated fashion. Strides have been made to understand the mechanisms regulating in vivo quiescence and proliferation of HSCs to develop strategies for ex vivo expansion. Ex vivo expansion of HSCs is important to procure sufficient number of stem cells and as easily available source for HSC transplants for patients suffering from hematological disorders and malignancies. Our lab has established a nanofiber-based ex vivo expansion strategy for HSCs, while preserving their stem cell characteristics. Ex vivo expanded cells were also found biologically functional in various disease models. However, the therapeutic potential of expanded stem cells at clinical level still needs to be verified. This review outlines transcriptional factors that regulate development of HSCs and their commitment, genes that regulate cell cycle status, studies that attempt to develop an effective and efficient protocol for ex vivo expansion of HSCs and application of HSC in various non-malignant and malignant disorders. Overall the goal of the current review is to deliver an understanding of factors that are critical in resolving the challenges that limit the expansion of HSCs in vivo and ex vivo. PMID:22082480

Neurotrophin-4 regulates the survival of gustatory neurons earlier in development using a different mechanism than brain-derived neurotrophic factor.

PubMed

Patel, Ami V; Krimm, Robin F

2012-05-01

The number of neurons in the geniculate ganglion that are available to innervate taste buds is regulated by neurotrophin-4 (NT-4) and brain-derived neurotrophic factor (BDNF). Our goal for the current study was to examine the timing and mechanism of NT-4-mediated regulation of geniculate neuron number during development. We discovered that NT-4 mutant mice lose 33% of their geniculate neuronal cells between E10.5 and E11.5. By E11.5, geniculate axons have just reached the tongue and do not yet innervate their gustatory targets; thus, NT-4 does not function as a target-derived growth factor. At E11.5, no difference was observed in proliferating cells or the rate at which cells exit the cell cycle between NT-4 mutant and wild type ganglia. Instead, there was an increase in TUNEL-labeling, indicating an increase in cell death in Ntf4(-/-) mice compared with wild types. However, activated caspase-3, which is up-regulated in the absence of BDNF, was not increased. This finding indicates that cell death initiated by NT-4-removal occurs through a different cell death pathway than BDNF-removal. We observed no additional postnatal loss of taste buds or neurons in Ntf4(-/-) mice. Thus, during early embryonic development, NT-4 produced in the ganglion and along the projection pathway inhibits cell death through an activated caspase-3 independent mechanism. Therefore, compared to BDNF, NT-4 plays distinct roles in gustatory development; differences include timing, source of neurotrophin, and mechanism of action. Published by Elsevier Inc.

CUDC-907 Promotes Bone Marrow Adipocytic Differentiation Through Inhibition of Histone Deacetylase and Regulation of Cell Cycle.

PubMed

Ali, Dalia; Alshammari, Hassan; Vishnubalaji, Radhakrishnan; Chalisserry, Elna Paul; Hamam, Rimi; Alfayez, Musaad; Kassem, Moustapha; Aldahmash, Abdullah; Alajez, Nehad M

2017-03-01

The role of bone marrow adipocytes (BMAs) in overall energy metabolism and their effects on bone mass are currently areas of intensive investigation. BMAs differentiate from bone marrow stromal cells (BMSCs); however, the molecular mechanisms regulating BMA differentiation are not fully understood. In this study, we investigated the effect of CUDC-907, identified by screening an epigenetic small-molecule library, on adipocytic differentiation of human BMSCs (hBMSCs) and determined its molecular mechanism of action. Human bone marrow stromal cells exposed to CUDC-907 (500 nM) exhibited enhanced adipocytic differentiation (∼2.9-fold increase, P < 0.005) compared with that of control cells. Global gene expression and signaling pathway analyses of differentially expressed genes revealed a strong enrichment of genes involved in adipogenesis, cell cycle, and DNA replication. Chromatin immune precipitation combined with quantitative polymerase chain reaction showed significant increase in H3K9ac epigenetic marker in the promoter regions of AdipoQ, FABP4, PPARγ, KLF15, and CEBPA in CUDC-907-treated hBMSCs. Follow-up experiments corroborated that the inhibition of histone deacetylase (HDAC) activity enhanced adipocytic differentiation, while the inhibition of PI3K decreased adipocytic differentiation. In addition, CUDC-907 arrested hBMSCs in the G0-G1 phase of the cell cycle and reduced the number of S-phase cells. Our data reveal that HDAC, PI3K, and cell cycle genes are important regulators of BMA formation and demonstrate that adipocyte differentiation of hBMSCs is associated with complex changes in a number of epigenetic and genetic pathways, which can be targeted to regulate BMA formation.

Brassinosteroid insensitive 1-associated kinase 1 (OsI-BAK1) is associated with grain filling and leaf development in rice.

PubMed

Khew, Choy-Yuen; Teo, Chin-Jit; Chan, Wai-Sun; Wong, Hann-Ling; Namasivayam, Parameswari; Ho, Chai-Ling

2015-06-15

Brassinosteroid Insensitive 1 (BRI1)-Associated Kinase I (BAK1) has been reported to interact with BRI1 for brassinosteroid (BR) perception and signal transduction that regulate plant growth and development. The aim of this study is to investigate the functions of a rice OsBAK1 homologue, designated as OsI-BAK1, which is highly expressed after heading. Silencing of OsI-BAK1 in rice plants produced a high number of undeveloped green and unfilled grains compared to the untransformed plants. Histological analyses demonstrated that embryos were either absent or retarded in their development in these unfilled rice grains of OsI-BAK1 RNAi plants. Down regulation of OsI-BAK1 caused a reduction in cell number and enlargement in leaf bulliform cells. Furthermore, transgenic rice plants overexpressing OsI-BAK1 were demonstrated to have corrugated and twisted leaves probably due to increased cell number that caused abnormal bulliform cell structure which were enlarged and plugged deep into leaf epidermis. The current findings suggest that OsI-BAK1 may play an important role in the developmental processes of rice grain filling and leaf cell including the bulliform cells. Copyright © 2015 Elsevier GmbH. All rights reserved.

MicroRNA inhibition fine-tunes and provides robustness to the restriction point switch of the cell cycle.

PubMed

Del Rosario, Ricardo C H; Damasco, Joseph Ray Clarence G; Aguda, Baltazar D

2016-09-09

The restriction point marks a switch in G1 from growth factor-dependent to growth factor-independent progression of the cell cycle. The proper regulation of this switch is important for normal cell processes; aberrations could result in a number of diseases such as cancer, neurodegenerative disorders, stroke and myocardial infarction. To further understand the regulation of the restriction point, we extended a mathematical model of the Rb-E2F pathway to include members of the microRNA cluster miR-17-92. Our mathematical analysis shows that microRNAs play an essential role in fine-tuning and providing robustness to the switch. We also demonstrate how microRNA regulation can steer cells in or out of cancer states.

MicroRNA inhibition fine-tunes and provides robustness to the restriction point switch of the cell cycle

PubMed Central

del Rosario, Ricardo C. H.; Damasco, Joseph Ray Clarence G.; Aguda, Baltazar D.

2016-01-01

The restriction point marks a switch in G1 from growth factor-dependent to growth factor-independent progression of the cell cycle. The proper regulation of this switch is important for normal cell processes; aberrations could result in a number of diseases such as cancer, neurodegenerative disorders, stroke and myocardial infarction. To further understand the regulation of the restriction point, we extended a mathematical model of the Rb-E2F pathway to include members of the microRNA cluster miR-17-92. Our mathematical analysis shows that microRNAs play an essential role in fine-tuning and providing robustness to the switch. We also demonstrate how microRNA regulation can steer cells in or out of cancer states. PMID:27610602

Anti-B-Cell Therapies in Autoimmune Neurological Diseases: Rationale and Efficacy Trials.

PubMed

Alexopoulos, Harry; Biba, Angie; Dalakas, Marinos C

2016-01-01

B cells have an ever-increasing role in the etiopathology of a number of autoimmune neurological disorders, acting as antibody-producing cells and, most importantly, as sensors, coordinators, and regulators of the immune response. B cells, among other functions, regulate the T-cell activation process through their participation in antigen presentation and production of cytokines. The availability of monoclonal antibodies or fusion proteins against B-cell surface molecules or B-cell trophic factors bestows a rational approach for treating autoimmune neurological disorders, even when T cells are the main effector cells. This review summarizes basic aspects of B-cell biology, discusses the role(s) of B cells in neurological autoimmunity, and presents anti-B-cell drugs that are either currently on the market or are expected to be available in the near future for treating neurological autoimmune disorders.

The Reactivity, Distribution and Abundance of Non-Astrocytic Inner Retinal Glial (NIRG) Cells Are Regulated by Microglia, Acute Damage, and IGF1

PubMed Central

Zelinka, Christopher P.; Scott, Melissa A.; Volkov, Leo; Fischer, Andy J.

2012-01-01

Recent studies have described a novel type of glial cell that is scattered across the inner layers of the avian retina and possibly the retinas of primates. These cells have been termed Non-astrocytic Inner Retinal Glial (NIRG) cells. These cells are stimulated by insulin-like growth factor 1 (IGF1) to proliferate, migrate distally into the retina, and become reactive. These changes in glial activity correlate with increased susceptibility of retinal neurons and Müller glia to excitotoxic damage. The purpose of this study was to further study the NIRG cells in retinas treated with IGF1 or acute damage. In response to IGF1, the reactivity, proliferation and migration of NIRG cells persists through 3 days after treatment. At 7 days after treatment, the numbers and distribution of NIRG cells returns to normal, suggesting that homeostatic mechanisms are in place within the retina to maintain the numbers and distribution of these glial cells. By comparison, IGF1-induced microglial reactivity persists for at least 7 days after treatment. In damaged retinas, we find a transient accumulation of NIRG cells, which parallels the accumulation of reactive microglia, suggesting that the reactivity of NIRG cells and microglia are linked. When the microglia are selectively ablated by the combination of interleukin 6 and clodronate-liposomes, the NIRG cells down-regulate transitin and perish within the following week, suggesting that the survival and phenotype of NIRG cells are somehow linked to the microglia. We conclude that the abundance, reactivity and retinal distribution of NIRG cells can be dynamic, are regulated by homoestatic mechanisms and are tethered to the microglia. PMID:22973454

CP-25, a Novel Anti-inflammatory and Immunomodulatory Drug, Inhibits the Functions of Activated Human B Cells through Regulating BAFF and TNF-alpha Signaling and Comparative Efficacy with Biological Agents.

PubMed

Zhang, Feng; Shu, Jin-Ling; Li, Ying; Wu, Yu-Jing; Zhang, Xian-Zheng; Han, Le; Tang, Xiao-Yu; Wang, Chen; Wang, Qing-Tong; Chen, Jing-Yu; Chang, Yan; Wu, Hua-Xun; Zhang, Ling-Ling; Wei, Wei

2017-01-01

Paeoniflorin-6'- O -benzene sulfonate (code: CP-25) was the chemistry structural modifications of Paeoniflorin (Pae). CP-25 inhibited B cells proliferation stimulated by B cell activating factor belonging to the TNF family (BAFF) or Tumor necrosis factor alpha (TNF-alpha). CP-25, Rituximab and Etanercept reduced the percentage and numbers of CD19 + B cells, CD19 + CD20 + B cells, CD19 + CD27 + B cells and CD19 + CD20 + CD27 + B cells induced by BAFF or TNF-alpha. There was significant difference between CP-25 and Rituximab or CP-25 and Etanercept. CP-25 down-regulated the high expression of BAFFR, BCMA, and TACI stimulated by BAFF or TNF-alpha. The effects of Rituximab and Etanercept on BAFFR or BCMA were stronger than that of CP-25. CP-25, Rituximab and Etanercept down-regulated significantly the expression of TNFR1 and TNFR2 on B cell stimulated by BAFF or TNF-alpha. CP-25, Rituximab and Etanercept down-regulated the expression of MKK3, P-p38, P-p65, TRAF2, and p52 in B cells stimulated by BAFF and the expression of TRAF2 and P-p65 in B cells stimulated by TNF-alpha. These results suggest that CP-25 regulated moderately activated B cells function by regulating the classical and alternative NF-κB signaling pathway mediated by BAFF and TNF-alpha-TRAF2-NF-κB signaling pathway. This study suggests that CP-25 may be a promising anti-inflammatory immune and soft regulation drug.

Mechanisms of epigenetic and cell-type specific regulation of Hey target genes in ES cells and cardiomyocytes.

PubMed

Weber, David; Heisig, Julia; Kneitz, Susanne; Wolf, Elmar; Eilers, Martin; Gessler, Manfred

2015-02-01

Hey bHLH transcription factors are critical effectors of Notch signaling. During mammalian heart development they are expressed in atrial and ventricular cardiomyocytes and in the developing endocardium. Hey knockout mice suffer from lethal cardiac defects, such as ventricular septum defects, valve defects and cardiomyopathy. Despite this functional relevance, little is known about the regulation of downstream targets in relevant cell types. The objective of this study was to elucidate the regulatory mechanisms by which Hey proteins affect gene expression in a cell type specific manner. We used an in vitro cardiomyocyte differentiation system with inducible Hey1 or Hey2 expression to study target gene regulation in cardiomyocytes (CM) generated from murine embryonic stem cells (ESC). The effects of Hey1 and Hey2 are largely redundant, but cell type specific. The number of regulated genes is comparable between ESC and CM, but the total number of binding sites is much higher, especially in ESC, targeting mainly genes involved in transcriptional regulation and developmental processes. Repression by Hey proteins generally correlates with the extent of Hey-binding to target promoters, Hdac recruitment and lower histone acetylation. Functionally, treatment with the Hdac inhibitor TSA abolished Hey target gene regulation. However, in CM the repressive effect of Hey-binding is lost for a subset of genes. These also lack Hey-dependent histone deacetylation in CM and are enriched for binding sites of cardiac specific activators like Srf, Nkx2-5, and Gata4. Ectopic Nkx2-5 overexpression in ESC blocks Hey-mediated repression of these genes. Thus, Hey proteins mechanistically repress target genes via Hdac recruitment and histone deacetylation. In CM Hey-repression is counteracted by cardiac activators, which recruit histone acetylases and prevent Hey mediated deacetylation and subsequent repression for a subset of genes. Copyright © 2014 Elsevier Ltd. All rights reserved.

Calcineurin/NFAT signalling inhibits myeloid haematopoiesis.

PubMed

Fric, Jan; Lim, Clarice X F; Koh, Esther G L; Hofmann, Benjamin; Chen, Jinmiao; Tay, Hock Soon; Mohammad Isa, Siti Aminah Bte; Mortellaro, Alessandra; Ruedl, Christiane; Ricciardi-Castagnoli, Paola

2012-04-01

Nuclear factor of activated T cells (NFAT) comprises a family of transcription factors that regulate T cell development, activation and differentiation. NFAT signalling can also mediate granulocyte and dendritic cell (DC) activation, but it is unknown whether NFAT influences their development from progenitors. Here, we report a novel role for calcineurin/NFAT signalling as a negative regulator of myeloid haematopoiesis. Reconstituting lethally irradiated mice with haematopoietic stem cells expressing an NFAT-inhibitory peptide resulted in enhanced development of the myeloid compartment. Culturing bone marrow cells in media supplemented with Flt3-L in the presence of the calcineurin/NFAT inhibitor Cyclosporin A increased numbers of differentiated DC. Global gene expression analysis of untreated DC and NFAT-inhibited DC revealed differential expression of transcripts that regulate cell cycle and apoptosis. In conclusion, these results provide evidence that calcineurin/NFAT signalling negatively regulates myeloid lineage development. The finding that inhibition of NFAT enhances myeloid development provides a novel insight into understanding how the treatment with drugs targeting calcineurin/NFAT signalling influence the homeostasis of the innate immune system. Copyright © 2012 EMBO Molecular Medicine.

Homozygously deleted gene DACH1 regulates tumor-initiating activity of glioma cells

PubMed Central

Watanabe, Akira; Ogiwara, Hideki; Ehata, Shogo; Mukasa, Akitake; Ishikawa, Shumpei; Maeda, Daichi; Ueki, Keisuke; Ino, Yasushi; Todo, Tomoki; Yamada, Yasuhiro; Fukayama, Masashi; Saito, Nobuhito; Miyazono, Kohei; Aburatani, Hiroyuki

2011-01-01

Loss or reduction in function of tumor suppressor genes contributes to tumorigenesis. Here, by allelic DNA copy number analysis using single-nucleotide polymorphism genotyping array and mass spectrometry, we report homozygous deletion in glioblastoma multiformes at chromosome 13q21, where DACH1 gene is located. We found decreased cell proliferation of a series of glioma cell lines by forced expression of DACH1. We then generated U87TR-Da glioma cells, where DACH1 expression could be activated by exposure of the cells to doxycycline. Both ex vivo cellular proliferation and in vivo growth of s.c. transplanted tumors in mice are reduced in U87TR-Da cells with DACH1 expression (U87-DACH1-high), compared with DACH1-nonexpressing U87TR-Da cells (U87-DACH1-low). U87-DACH1-low cells form spheroids with CD133 and Nestin expression in serum-free medium but U87-DACH1-high cells do not. Compared with spheroid-forming U87-DACH1-low cells, adherent U87-DACH1-high cells display lower tumorigenicity, indicating DACH1 decreases the number of tumor-initiating cells. Gene expression analysis and chromatin immunoprecipitation assay reveal that fibroblast growth factor 2 (FGF2/bFGF) is transcriptionally repressed by DACH1, especially in cells cultured in serum-free medium. Exogenous bFGF rescues spheroid-forming activity and tumorigenicity of the U87-DACH1-high cells, suggesting that loss of DACH1 increases the number of tumor-initiating cells through transcriptional activation of bFGF. These results illustrate that DACH1 is a distinctive tumor suppressor, which does not only suppress growth of tumor cells but also regulates bFGF-mediated tumor-initiating activity of glioma cells. PMID:21750150

FAP positive fibroblasts induce immune checkpoint blockade resistance in colorectal cancer via promoting immunosuppression.

PubMed

Chen, Lingling; Qiu, Xiangting; Wang, Xinhua; He, Jian

2017-05-20

Immune checkpoint blockades that significantly prolonged survival of melanoma patients have been less effective on colorectal cancer (CRC) patients. Growing evidence suggested that fibroblast activation protein-alpha (FAP) on cancer associate fibroblasts (CAFs) has critical roles in regulating antitumor immune response by inducing tumor-promoting inflammation. In this study, we explored the roles of FAP in regulating the tumor immunity and immune checkpoint blockades resistance in CRC experimental systems. We found that CAFs with high FAP expression could induce immune checkpoint blockade resistance in CRC mouse model. Mechanistically, CAFs with high FAP expression promoted immunosuppression in the CRC tumor immune microenvironment by up-regulating CCL2 secretion, recruiting myeloid cells, and decreasing T-cell activity. In human CRC samples, FAP expression was proportional to myeloid cells number, but inversely related to T-cell number. High FAP expression also predicted poor survival of CRC patients. Taken together, our study suggested that high FAP expression in CAFs is one reason leading to immune checkpoint blockades resistance in CRC patients and FAP is an optional target for reversing immune checkpoint blockades resistance. Copyright © 2017 Elsevier Inc. All rights reserved.

The Maize MID-COMPLEMENTING ACTIVITY homolog CELL NUMBER REGULATOR13/NARROW ODD DWARF, coordinates organ growth and tissue patterning

USDA-ARS?s Scientific Manuscript database

Organogenesis occurs from cell division, expansion and differentiation. How these cellular processes are coordinated remains elusive. The maize leaf provides an excellent system to study cellular differentiation because it has several different tissues and cell types. The narrow odd dwarf (nod) mut...

SHOEBOX Modulates Root Meristem Size in Rice through Dose-Dependent Effects of Gibberellins on Cell Elongation and Proliferation

PubMed Central

Li, Jintao; Zhao, Yu; Chu, Huangwei; Wang, Likai; Fu, Yanru; Liu, Ping; Upadhyaya, Narayana; Chen, Chunli; Mou, Tongmin; Feng, Yuqi; Kumar, Prakash; Xu, Jian

2015-01-01

Little is known about how the size of meristem cells is regulated and whether it participates in the control of meristem size in plants. Here, we report our findings on shoebox (shb), a mild gibberellin (GA) deficient rice mutant that has a short root meristem size. Quantitative analysis of cortical cell length and number indicates that shb has shorter, rather than fewer, cells in the root meristem until around the fifth day after sowing, from which the number of cortical cells is also reduced. These defects can be either corrected by exogenous application of bioactive GA or induced in wild-type roots by a dose-dependent inhibitory effect of paclobutrazol on GA biosynthesis, suggesting that GA deficiency is the primary cause of shb mutant phenotypes. SHB encodes an AP2/ERF transcription factor that directly activates transcription of the GA biosynthesis gene KS1. Thus, root meristem size in rice is modulated by SHB-mediated GA biosynthesis that regulates the elongation and proliferation of meristem cells in a developmental stage-specific manner. PMID:26275148

SHOEBOX Modulates Root Meristem Size in Rice through Dose-Dependent Effects of Gibberellins on Cell Elongation and Proliferation.

PubMed

Li, Jintao; Zhao, Yu; Chu, Huangwei; Wang, Likai; Fu, Yanru; Liu, Ping; Upadhyaya, Narayana; Chen, Chunli; Mou, Tongmin; Feng, Yuqi; Kumar, Prakash; Xu, Jian

2015-08-01

Little is known about how the size of meristem cells is regulated and whether it participates in the control of meristem size in plants. Here, we report our findings on shoebox (shb), a mild gibberellin (GA) deficient rice mutant that has a short root meristem size. Quantitative analysis of cortical cell length and number indicates that shb has shorter, rather than fewer, cells in the root meristem until around the fifth day after sowing, from which the number of cortical cells is also reduced. These defects can be either corrected by exogenous application of bioactive GA or induced in wild-type roots by a dose-dependent inhibitory effect of paclobutrazol on GA biosynthesis, suggesting that GA deficiency is the primary cause of shb mutant phenotypes. SHB encodes an AP2/ERF transcription factor that directly activates transcription of the GA biosynthesis gene KS1. Thus, root meristem size in rice is modulated by SHB-mediated GA biosynthesis that regulates the elongation and proliferation of meristem cells in a developmental stage-specific manner.

The biochemistry of hematopoietic stem cell development.

PubMed

Kaimakis, P; Crisan, M; Dzierzak, E

2013-02-01

The cornerstone of the adult hematopoietic system and clinical treatments for blood-related disease is the cohort of hematopoietic stem cells (HSC) that is harbored in the adult bone marrow microenvironment. Interestingly, this cohort of HSCs is generated only during a short window of developmental time. In mammalian embryos, hematopoietic progenitor and HSC generation occurs within several extra- and intraembryonic microenvironments, most notably from 'hemogenic' endothelial cells lining the major vasculature. HSCs are made through a remarkable transdifferentiation of endothelial cells to a hematopoietic fate that is long-lived and self-renewable. Recent studies are beginning to provide an understanding of the biochemical signaling pathways and transcription factors/complexes that promote their generation. The focus of this review is on the biochemistry behind the generation of these potent long-lived self-renewing stem cells of the blood system. Both the intrinsic (master transcription factors) and extrinsic regulators (morphogens and growth factors) that affect the generation, maintenance and expansion of HSCs in the embryo will be discussed. The generation of HSCs is a stepwise process involving many developmental signaling pathways, morphogens and cytokines. Pivotal hematopoietic transcription factors are required for their generation. Interestingly, whereas these factors are necessary for HSC generation, their expression in adult bone marrow HSCs is oftentimes not required. Thus, the biochemistry and molecular regulation of HSC development in the embryo are overlapping, but differ significantly from the regulation of HSCs in the adult. HSC numbers for clinical use are limiting, and despite much research into the molecular basis of HSC regulation in the adult bone marrow, no panel of growth factors, interleukins and/or morphogens has been found to sufficiently increase the number of these important stem cells. An understanding of the biochemistry of HSC generation in the developing embryo provides important new knowledge on how these complex stem cells are made, sustained and expanded in the embryo to give rise to the complete adult hematopoietic system, thus stimulating novel strategies for producing increased numbers of clinically useful HSCs. This article is part of a Special Issue entitled Biochemistry of Stem Cells. Copyright © 2012 Elsevier B.V. All rights reserved.

Regulation of epidermal cell fate in Arabidopsis roots: the importance of multiple feedback loops

PubMed Central

Schiefelbein, John; Huang, Ling; Zheng, Xiaohua

2014-01-01

The specification of distinct cell types in multicellular organisms is accomplished via establishment of differential gene expression. A major question is the nature of the mechanisms that establish this differential expression in time and space. In plants, the formation of the hair and non-hair cell types in the root epidermis has been used as a model to understand regulation of cell specification. Recent findings show surprising complexity in the number and the types of regulatory interactions between the multiple transcription factor genes/proteins influencing root epidermis cell fate. Here, we describe this regulatory network and the importance of the multiple feedback loops for its establishment and maintenance. PMID:24596575

Concise Review: Geminin-A Tale of Two Tails: DNA Replication and Transcriptional/Epigenetic Regulation in Stem Cells.

PubMed

Patmanidi, Alexandra L; Champeris Tsaniras, Spyridon; Karamitros, Dimitris; Kyrousi, Christina; Lygerou, Zoi; Taraviras, Stavros

2017-02-01

Molecular mechanisms governing maintenance, commitment, and differentiation of stem cells are largely unexploited. Molecules involved in the regulation of multiple cellular processes are of particular importance for stem cell physiology, as they integrate different signals and coordinate cellular decisions related with self-renewal and fate determination. Geminin has emerged as a critical factor in DNA replication and stem cell differentiation in different stem cell populations. Its inhibitory interaction with Cdt1, a member of the prereplicative complex, ensures the controlled timing of DNA replication and, consequently, genomic stability in actively proliferating cells. In embryonic as well as somatic stem cells, Geminin has been shown to interact with transcription factors and epigenetic regulators to drive gene expression programs and ultimately guide cell fate decisions. An ever-growing number of studies suggests that these interactions of Geminin and proteins regulating transcription are conserved among metazoans. Interactions between Geminin and proteins modifying the epigenome, such as members of the repressive Polycomb group and the SWI/SNF proteins of the permissive Trithorax, have long been established. The complexity of these interactions, however, is only just beginning to unravel, revealing key roles on maintaining stem cell self-renewal and fate specification. In this review, we summarize current knowledge and give new perspectives for the role of Geminin on transcriptional and epigenetic regulation, alongside with its regulatory activity in DNA replication and their implication in the regulation of stem and progenitor cell biology. Stem Cells 2017;35:299-310. © 2016 AlphaMed Press.

MicroRNA-9 regulates non-small cell lung cancer cell invasion and migration by targeting eukaryotic translation initiation factor 5A2.

PubMed

Xu, Guodong; Shao, Guofeng; Pan, Qiaoling; Sun, Lebo; Zheng, Dawei; Li, Minghui; Li, Ni; Shi, Huoshun; Ni, Yiming

2017-01-01

MicroRNAs (miRNAs) play a critical role in cancer development and progression. Bioinformatics analyses has identified eukaryotic translation initiation factor 5A2 (eIF5A2) as a target of miR-9. In this study, we attempted to determine whether miR-9 regulates non-small cell lung cancer (NSCLC) cell invasion and migration by targeting eIF5A2 We examined eIF5A2 expression using reverse transcription-quantitative PCR (RT-qPCR) and subsequently transfected A549 and NCI-H1299 NSCLC cells with a miR-9 mimic or miR-9 inhibitor to determine the migration and invasive capability of the cells via wound healing assay and Transwell invasion assay, respectively. E-cadherin and vimentin expression was detected with western blotting. The miR-9 mimic significantly reduced NSCLC cell invasive and metastatic ability, and the miR-9 inhibitor enhanced NSCLC cell migration activity, increasing the number of migrated cells. There was no significant difference between the negative control siRNA and miR-9 mimic groups after knockdown of eIF5A2; western blotting showed that miR-9 regulated E-cadherin and vimentin expression. These data show that miR-9 regulates NSCLC cell invasion and migration through regulating eIF5A2 expression. Taken together, our findings suggest that the mechanism of miR-9-regulated NSCLC cell invasion and migration may be related to epithelial-mesenchymal transition.

Indole-3-Carbinol Promotes Goblet-Cell Differentiation Regulating Wnt and Notch Signaling Pathways AhR-Dependently.

PubMed

Park, Joo-Hung; Lee, Jeong-Min; Lee, Eun-Jin; Hwang, Won-Bhin; Kim, Da-Jeong

2018-04-30

Using an in vitro model of intestinal organoids derived from intestinal crypts, we examined effects of indole-3-carbinol (I3C), a phytochemical that has anticancer and aryl hydrocarbon receptor (AhR)-activating abilities and thus is sold as a dietary supplement, on the development of intestinal organoids and investigated the underlying mechanisms. I3C inhibited the in vitro development of mouse intestinal organoids. Addition of α-naphthoflavone, an AhR antagonist or AhR siRNA transfection, suppressed I3C function, suggesting that I3C-mediated interference with organoid development is AhR-dependent. I3C increased the expression of Muc2 and lysozyme, lineage-specific genes for goblet cells and Paneth cells, respectively, but inhibits the expression of IAP, a marker gene for enterocytes. In the intestines of mice treated with I3C, the number of goblet cells was reduced, but the number of Paneth cells and the depth and length of crypts and villi were not changed. I3C increased the level of active nonphosphorylated β-catenin, but suppressed the Notch signal. As a result, expression of Hes1, a Notch target gene and a transcriptional repressor that plays a key role in enterocyte differentiation, was reduced, whereas expression of Math1, involved in the differentiation of secretory lineages, was increased. These results provide direct evidence for the role of AhR in the regulation of the development of intestinal stem cells and indicate that such regulation is likely mediated by regulation of Wnt and Notch signals.

Increased Differentiation of Dermal Mast Cells in Mice Lacking the Mpl Gene

PubMed Central

Ghinassi, Barbara; Zingariello, Maria; Martelli, Fabrizio; Lorenzini, Rodolfo; Vannucchi, Alessandro M.; Rana, Rosa Alba; Nishikawa, Mitsuo; Migliaccio, Giovanni; Mascarenhas, John

2009-01-01

Thrombopoietin interactions with its receptor, Mpl, play an important role in the regulation of hematopoietic stem/progenitor cell proliferation and differentiation. In this study, we report that the mast cell restricted progenitor cells (MCP) and the mast cell precursors in the bone marrow of wild-type mice express Mpl on their surface. Furthermore, targeted deletion of the Mpl gene in mice decreases the number of MCP while increasing the number of mast cell precursors present in the marrow and spleen. It also increases the number of mast cells present in the dermis, in the peritoneal cavity, and in the gut of the mice. In addition, serosal mast cells from Mplnull mice have a distinctive differentiation profile similar to that expressed by wild-type dermal mast cells. These results suggest that not only does ligation of thrombopoietin with the Mpl receptor exert an effect at the mast cell restricted progenitor cell level, but also plays an unexpected yet important role in mast cell maturation. PMID:19025339

The negative regulation of red cell mass by neocytolysis: physiologic and pathophysiologic manifestations.

PubMed

Rice, Lawrence; Alfrey, Clarence P

2005-01-01

We have uncovered a physiologic process which negatively regulates the red cell mass by selectively hemolyzing young circulating red blood cells. This allows fine control of the number of circulating red blood cells under steady-state conditions and relatively rapid adaptation to new environments. Neocytolysis is initiated by a fall in erythropoietin levels, so this hormone remains the major regulator of red cell mass both with anemia and with red cell excess. Physiologic situations in which there is increased neocytolysis include the emergence of newborns from the hypoxic uterine environment and the descent of polycythemic high-altitude dwellers to sea level. The process first became apparent while investigating the mechanism of the anemia that invariably occurs after spaceflight. Astronauts experience acute central plethora on entering microgravity resulting in erythropoietin suppression and neocytolysis, but the reduced blood volume and red cell mass become suddenly maladaptive on re-entry to earth's gravity. The pathologic erythropoietin deficiency of renal disease precipitates neocytolysis, which explains the prolongation of red cell survival consistently resulting from erythropoietin therapy and points to optimally efficient erythropoietin dosing schedules. Implications should extend to a number of other physiologic and pathologic situations including polycythemias, hemolytic anemias, 'blood-doping' by elite athletes, and oxygen therapy. It is likely that erythropoietin influences endothelial cells which in turn signal reticuloendothelial phagocytes to destroy or permit the survival of young red cells marked by surface molecules. Ongoing studies to identify the molecular targets and cytokine intermediaries should facilitate detection, dissection and eventual therapeutic manipulation of the process. Copyright (c) 2005 S. Karger AG, Basel.

Intermediate Filaments Play a Pivotal Role in Regulating Cell Architecture and Function.

PubMed

Lowery, Jason; Kuczmarski, Edward R; Herrmann, Harald; Goldman, Robert D

2015-07-10

Intermediate filaments (IFs) are composed of one or more members of a large family of cytoskeletal proteins, whose expression is cell- and tissue type-specific. Their importance in regulating the physiological properties of cells is becoming widely recognized in functions ranging from cell motility to signal transduction. IF proteins assemble into nanoscale biopolymers with unique strain-hardening properties that are related to their roles in regulating the mechanical integrity of cells. Furthermore, mutations in the genes encoding IF proteins cause a wide range of human diseases. Due to the number of different types of IF proteins, we have limited this short review to cover structure and function topics mainly related to the simpler homopolymeric IF networks composed of vimentin, and specifically for diseases, the related muscle-specific desmin IF networks. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

NFIB-mediated repression of the epigenetic factor Ezh2 regulates cortical development.

PubMed

Piper, Michael; Barry, Guy; Harvey, Tracey J; McLeay, Robert; Smith, Aaron G; Harris, Lachlan; Mason, Sharon; Stringer, Brett W; Day, Bryan W; Wray, Naomi R; Gronostajski, Richard M; Bailey, Timothy L; Boyd, Andrew W; Richards, Linda J

2014-02-19

Epigenetic mechanisms are essential in regulating neural progenitor cell self-renewal, with the chromatin-modifying protein Enhancer of zeste homolog 2 (EZH2) emerging as a central player in promoting progenitor cell self-renewal during cortical development. Despite this, how Ezh2 is itself regulated remains unclear. Here, we demonstrate that the transcription factor nuclear factor IB (NFIB) plays a key role in this process. Nfib(-/-) mice exhibit an increased number of proliferative ventricular zone cells that express progenitor cell markers and upregulation of EZH2 expression within the neocortex and hippocampus. NFIB binds to the Ezh2 promoter and overexpression of NFIB represses Ezh2 transcription. Finally, key downstream targets of EZH2-mediated epigenetic repression are misregulated in Nfib(-/-) mice. Collectively, these results suggest that the downregulation of Ezh2 transcription by NFIB is an important component of the process of neural progenitor cell differentiation during cortical development.

Active hexose correlated compound enhances tumor surveillance through regulating both innate and adaptive immune responses.

PubMed

Gao, Yunfei; Zhang, Dongqing; Sun, Buxiang; Fujii, Hajime; Kosuna, Ken-Ichi; Yin, Zhinan

2006-10-01

Active hexose correlated compound (AHCC) is a mixture of polysaccharides, amino acids, lipids and minerals derived from cocultured mycelia of several species of Basidiomycete mushrooms. AHCC has been implicated to modulate immune functions and plays a protective role against infection. However, the potential role of AHCC in tumor immune surveillance is unknown. In this study, C57BL/6 mice were orally administered AHCC or water, followed by tumor cell inoculation. We showed that compared to pure water-treated mice, AHCC treatment significantly delayed tumor development after inoculation of either melanoma cell line B16F0 or lymphoma cell line EL4. Treatment with AHCC enhanced both Ag-specific activation and proliferation of CD4(+) and CD8(+) T cells, increased the number of tumor Ag-specific CD8(+) T cells, and more importantly, increased the frequency of tumor Ag-specific IFN-gamma producing CD8(+) T cells. Interestingly, AHCC treatment also showed increased cell number of NK and gammadelta T cells, indicating the role of AHCC in activating these innate-like lymphocytes. In summary, our results demonstrate that AHCC can enhance tumor immune surveillance through regulating both innate and adaptive immune responses.

Aggregatibacter actinomycetemcomitans regulates the expression of integrins and reduces cell adhesion via integrin α5 in human gingival epithelial cells.

PubMed

Kochi, Shinsuke; Yamashiro, Keisuke; Hongo, Shoichi; Yamamoto, Tadashi; Ugawa, Yuki; Shimoe, Masayuki; Kawamura, Mari; Hirata-Yoshihara, Chiaki; Ideguchi, Hidetaka; Maeda, Hiroshi; Takashiba, Shogo

2017-12-01

Gingival epithelial cells form a physiological barrier against bacterial invasion. Excessive bacterial invasion destroys the attachment between the tooth surface and the epithelium, resulting in periodontitis. Integrins play a significant role in cell attachment; therefore, we hypothesized that bacterial infection might decrease the expressions of these integrins in gingival epithelial cells, resulting in reduced cell adhesion. Immortalized human gingival epithelial cells were co-cultured with Aggregatibacter actinomycetemcomitans Y4 (Aa Y4), and the gene expression levels of IL-8, proliferating cell nuclear antigen (PCNA), and integrins (α2, α3, α5, β4, and β6) were measured using quantitative reverse transcription polymerase chain reaction. Expression of PCNA and integrins, except integrin α5, was significantly downregulated, while expression of IL-8 and integrin α5 was significantly upregulated in the cells co-cultured with Aa Y4. The number of adherent cells significantly decreased when co-cultured with Aa Y4, as determined using cell adhesion assays. In the cells co-cultured with Aa Y4 and an integrin α5 neutralizing antibody, there was no effect on the expression of IL-8 and PCNA, while the expressions of integrins α2, α3, β4, and β6, and the number of adherent cells did not decrease. The number of invading bacteria in the cells was reduced in the presence of the antibody and increased in the presence of TLR2/4 inhibitor. Therefore, integrin α5 might be involved in Aa Y4 invasion into gingival epithelial cells, and the resulting signal transduction cascade reduces cell adhesion by decreasing the expression of integrins, while the TLR2/4 signaling cascade regulates IL-8 expression.

PD-1 regulates T cell proliferation in a tissue and subset specific manner during normal mouse pregnancy

PubMed Central

Shepard, Michelle T.; Bonney, Elizabeth A.

2014-01-01

The regulation of T cell homeostasis during pregnancy has important implications for maternal tolerance and immunity. Evidence suggests that Programmed Death-1 (PD-1) participates in regulation of T cell homeostasis and peripheral tolerance. To examine the contribution of PD-1 signaling on T cell homeostasis during normal mouse pregnancy, we examined T cell number or proportion, PD-1 expression, proliferation, and apoptosis by flow cytometry, BrdU incorporation, and TUNEL assay in pregnant mice given anti-PD-1 blocking antibody or control on days 10, 12, and 14 of gestation. We observed tissue, treatment, and T cell-specific differences in PD-1 expression. Both pregnancy and PD-1 blockade increased T cell proliferation in the spleen while this effect was limited to CD4 T cells in the uterine- draining nodes. In the uterus, PD-1 blockade markedly altered the composition of the T cell pool. These studies support the idea that pregnancy is a state of dynamic T cell homeostasis and suggest that this state is partially supported by PD-1 signaling. PMID:23782245

Mitochondrial DNA copy number is regulated in a tissue specific manner by DNA methylation of the nuclear-encoded DNA polymerase gamma A

PubMed Central

Kelly, Richard D. W.; Mahmud, Arsalan; McKenzie, Matthew; Trounce, Ian A.; St John, Justin C.

2012-01-01

DNA methylation is an essential mechanism controlling gene expression during differentiation and development. We investigated the epigenetic regulation of the nuclear-encoded, mitochondrial DNA (mtDNA) polymerase γ catalytic subunit (PolgA) by examining the methylation status of a CpG island within exon 2 of PolgA. Bisulphite sequencing identified low methylation levels (<10%) within exon 2 of mouse oocytes, blastocysts and embryonic stem cells (ESCs), while somatic tissues contained significantly higher levels (>40%). In contrast, induced pluripotent stem (iPS) cells and somatic nuclear transfer ESCs were hypermethylated (>20%), indicating abnormal epigenetic reprogramming. Real time PCR analysis of 5-methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC) immunoprecipitated DNA suggests active DNA methylation and demethylation within exon 2 of PolgA. Moreover, neural differentiation of ESCs promoted de novo methylation and demethylation at the exon 2 locus. Regression analysis demonstrates that cell-specific PolgA expression levels were negatively correlated with DNA methylation within exon 2 and mtDNA copy number. Finally, using chromatin immunoprecipitation (ChIP) against RNA polymerase II (RNApII) phosphorylated on serine 2, we show increased DNA methylation levels are associated with reduced RNApII transcriptional elongation. This is the first study linking nuclear DNA epigenetic regulation with mtDNA regulation during differentiation and cell specialization. PMID:22941637

[Harringtonine induces apoptosis in NB4 cells through down-regulation of Mcl-1].

PubMed

Wu, Chunxiao; Shen, Hongqiang; Xia, Dajing

2013-07-01

To investigate the growth inhibition effect, cytotoxicity and apoptotic induction of harringtonine (HT) in human acute promyelocytic leukemia (APL) NB4 cells,and the related mechanism. NB4 cells were treated with HT. Total cell numbers were counted by hemocytometer, and cell viabilities were determined by trypan blue exclusion. Apoptotic cells were determined by fluorescence microscopy and FACS after staining with AO and EB or PI, respectively. The cleavage of PARP and the activation of Bax and the expression of anti-apoptotic proteins were determined by Western Blot. siRNA was used to silence the expression of target genes. Primary cells were isolated following Ficoll-Hypaque density gradient centrifugation method. HT inhibited cell growth and induced apoptosis of NB4 cells in a dose- and time-dependent manner. Apoptosis induced by HT was correlated with the down-regulation of Mcl-1 and the cleavage of PARP, while HT did not affect the protein level of Bax and Bak or change the protein level of Bcl-2. The silence of Bcl-XL sensitized HT-induced apoptosis in NB4 cells.Apoptosis induced by HT in primarily cultured APL cells was also correlated with the down-regulation of Mcl-1. HT inhibits cell growth and induces apoptosis in NB4 cells and primarily cultured APL cells, which may be associated with down-regulation of Mcl-1.

PbrmiR397a regulates lignification during stone cell development in pear fruit.

PubMed

Xue, Cheng; Yao, Jia-Long; Qin, Meng-Fan; Zhang, Ming-Yue; Allan, Andrew C; Wang, De-Fu; Wu, Jun

2018-05-13

Lignified stone cells substantially reduce fruit quality. Therefore, it is desirable to inhibit stone cell development by using genetic technologies. However, the molecular mechanisms regulating lignification are poorly understood in fruit stone cells. In this study, we have shown that microRNA (miR) miR397a regulates fruit cell lignification by inhibiting laccase (LAC) genes that encode key lignin biosynthesis enzymes. Transient overexpression of PbrmiR397a, which is the miR397a of Chinese pear (Pyrus bretschneideri), and simultaneous silencing of three LAC genes reduced the lignin content and stone cell number in pear fruit. A single nucleotide polymorphism (SNP) identified in the promoter of the PbrmiR397a gene was found to associate with low levels of fruit lignin, after analysis of the genome sequences of sixty pear varieties. This SNP created a TCA-element that responded to salicylic acid (SA) to induce gene expression as confirmed using a cell-based assay system. Furthermore, stable overexpression of PbrmiR397a in transgenic tobacco plants reduced the expression of target LAC genes and decreased the content of lignin but did not change the ratio of syringyl and guaiacyl lignin monomers. Consistent with reduction of lignin content, the transgenic plants showed fewer numbers of vessel elements and thinner secondary walls in the remaining elements compared to wild-type control plants. This study has advanced our understanding of the regulation of lignin biosynthesis and provided useful molecular genetic information for improving pear fruit quality. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

Nitric oxide regulates tumor cell cross-talk with stromal cells in the tumor microenvironment of the liver.

PubMed

Decker, Ningling Kang; Abdelmoneim, Soha S; Yaqoob, Usman; Hendrickson, Helen; Hormes, Joe; Bentley, Mike; Pitot, Henry; Urrutia, Raul; Gores, Greg J; Shah, Vijay H

2008-10-01

Tumor progression is regulated through paracrine interactions between tumor cells and stromal cells in the microenvironment, including endothelial cells and myofibroblasts. Nitric oxide (NO) is a key molecule in the regulation of tumor-microenvironment interactions, although its precise role is incompletely defined. By using complementary in vitro and in vivo approaches, we studied the effect of endothelial NO synthase (eNOS)-derived NO on liver tumor growth and metastasis in relation to adjacent stromal myofibroblasts and matrix because liver tumors maintain a rich, vascular stromal network enriched with phenotypically heterogeneous myofibroblasts. Mice with an eNOS deficiency developed liver tumors more frequently in response to carcinogens compared with control animals. In a surgical model of pancreatic cancer liver metastasis, eNOS overexpression in the tumor microenvironment attenuated both the number and size of tumor implants. NO promoted anoikis of tumor cells in vitro and limited their invasive capacity. Because tumor cell anoikis and invasion are both regulated by myofibroblast-derived matrix, we explored the effect of NO on tumor cell protease expression. Both microarray and Western blot analysis revealed eNOS-dependent down-regulation of the matrix protease cathepsin B within tumor cells, and silencing of cathepsin B attenuated tumor cell invasive capacity in a similar manner to that observed with eNOS overexpression. Thus, a NO gradient within the tumor microenvironment influences tumor progression through orchestrated molecular interactions between tumor cells and stroma.

Asymmetric cell division during T cell development controls downstream fate

PubMed Central

Pham, Kim; Shimoni, Raz; Charnley, Mirren; Ludford-Menting, Mandy J.; Hawkins, Edwin D.; Ramsbottom, Kelly; Oliaro, Jane; Izon, David; Ting, Stephen B.; Reynolds, Joseph; Lythe, Grant; Molina-Paris, Carmen; Melichar, Heather; Robey, Ellen; Humbert, Patrick O.; Gu, Min

2015-01-01

During mammalian T cell development, the requirement for expansion of many individual T cell clones, rather than merely expansion of the entire T cell population, suggests a possible role for asymmetric cell division (ACD). We show that ACD of developing T cells controls cell fate through differential inheritance of cell fate determinants Numb and α-Adaptin. ACD occurs specifically during the β-selection stage of T cell development, and subsequent divisions are predominantly symmetric. ACD is controlled by interaction with stromal cells and chemokine receptor signaling and uses a conserved network of polarity regulators. The disruption of polarity by deletion of the polarity regulator, Scribble, or the altered inheritance of fate determinants impacts subsequent fate decisions to influence the numbers of DN4 cells arising after the β-selection checkpoint. These findings indicate that ACD enables the thymic microenvironment to orchestrate fate decisions related to differentiation and self-renewal. PMID:26370500

Red blood cells as modulators of T cell growth and survival.

PubMed

Arosa, Fernando A; Pereira, Carlos F; Fonseca, Ana M

2004-01-01

T cell homeostasis is largely controlled by a balance between cell death and survival and anomalies in either process account for a number of diseases linked to excessive or faulty T cell growth. Yet, the influence that cells outside the immunological system have on these processes has only recently received attention. Accumulated evidence indicate that homeostasis of the CD4+ and CD8+ T cell pools is highly dynamic and regulated by signals delivered by cells and molecules present in the different internal microenvironments. The major function of red blood cells (RBC) is generally considered to be oxygen and carbon dioxide transport. In recent years, however, RBC have been implicated in the regulation of basic physiological processes, from vascular contraction and platelet aggregation to T cell growth and survival. Regulation of T cell survival by RBC may influence the response of selected subsets of T cells to internal or external stimuli and may help explaining the immunomodulatory activities of red blood cells. By interfering in the balance between death and survival RBC become potential tools that can be manipulated to improve or reverse pathological situations characterized by anomalies in the control of T cell growth.

Seed Embryo Development Is Regulated via an AN3-MINI3 Gene Cascade

PubMed Central

Meng, Lai-Sheng; Wang, Yi-Bo; Loake, Gary J.; Jiang, Ji-Hong

2016-01-01

In agriculture, seed mass is one of the most important components related to seed yield. MINISEED3 (MINI3) which encodes the transcriptional activator WRKY10, is thought to be a pivotal regulator of seed mass. In Arabidopsis SHORT HYPOCOTYL UNDER BLUE1 (SHB1) associates with the promoter of MINI3, regulating embryo cell proliferation (both cell division and elongation), which, in turn, modulates seed mass. Furthermore, the recruitment of SHB1 via MINI3 to both its cognate promoter and that of IKU2 implies a two-step amplification for countering the low expression level of IKU2, which is thought to function as a molecular switch for seed cavity enlargement. However, it is largely unknown how embryo cell proliferation, which encompasses both cell division and elongation, is regulated by SHB1 and MINI3 function. Here, we show that a loss of function mutation within the transcriptional coactivator ANGUSTIFOLIA3 (AN3), increases seed mass. Further, AN3 associates with the MINI3 promoter in vivo. Genetic evidence indicates that the absence of MINI3 function suppresses the decrease of cell number observed in an3-4 mutants by regulating cell division and in turn inhibits increased cell size of the an3-4 line by controlling cell elongation. Thus, seed embryo development is modulated via an AN3-MINI3 gene cascade. This regulatory model provides a deeper understanding of seed mass regulation, which may in turn lead to increased crop yields. PMID:27857719

Organic Cation Transporter-Mediated Ergothioneine Uptake in Mouse Neural Progenitor Cells Suppresses Proliferation and Promotes Differentiation into Neurons

PubMed Central

Ishimoto, Takahiro; Nakamichi, Noritaka; Hosotani, Hiroshi; Masuo, Yusuke; Sugiura, Tomoko; Kato, Yukio

2014-01-01

The aim of the present study is to clarify the functional expression and physiological role in neural progenitor cells (NPCs) of carnitine/organic cation transporter OCTN1/SLC22A4, which accepts the naturally occurring food-derived antioxidant ergothioneine (ERGO) as a substrate in vivo. Real-time PCR analysis revealed that mRNA expression of OCTN1 was much higher than that of other organic cation transporters in mouse cultured cortical NPCs. Immunocytochemical analysis showed colocalization of OCTN1 with the NPC marker nestin in cultured NPCs and mouse embryonic carcinoma P19 cells differentiated into neural progenitor-like cells (P19-NPCs). These cells exhibited time-dependent [3H]ERGO uptake. These results demonstrate that OCTN1 is functionally expressed in murine NPCs. Cultured NPCs and P19-NPCs formed neurospheres from clusters of proliferating cells in a culture time-dependent manner. Exposure of cultured NPCs to ERGO or other antioxidants (edaravone and ascorbic acid) led to a significant decrease in the area of neurospheres with concomitant elimination of intracellular reactive oxygen species. Transfection of P19-NPCs with small interfering RNA for OCTN1 markedly promoted formation of neurospheres with a concomitant decrease of [3H]ERGO uptake. On the other hand, exposure of cultured NPCs to ERGO markedly increased the number of cells immunoreactive for the neuronal marker βIII-tubulin, but decreased the number immunoreactive for the astroglial marker glial fibrillary acidic protein (GFAP), with concomitant up-regulation of neuronal differentiation activator gene Math1. Interestingly, edaravone and ascorbic acid did not affect such differentiation of NPCs, in contrast to the case of proliferation. Knockdown of OCTN1 increased the number of cells immunoreactive for GFAP, but decreased the number immunoreactive for βIII-tubulin, with concomitant down-regulation of Math1 in P19-NPCs. Thus, OCTN1-mediated uptake of ERGO in NPCs inhibits cellular proliferation via regulation of oxidative stress, and also promotes cellular differentiation by modulating the expression of basic helix-loop-helix transcription factors via an unidentified mechanism different from antioxidant action. PMID:24586778

Fluoroorotic acid-selected Nicotiana plumbaginifolia cell lines with a stable thymine starvation phenotype have lost the thymine-regulated transcriptional program.

PubMed

Santoso, D; Thornburg, R

2000-08-01

We have selected 143 independent Nicotiana plumbaginifolia cell lines that survive in the presence of 5-fluoroorotic acid. These lines show several diverse phenotypes. The majority of these cell lines showed reduced levels of UMP synthase. However, one particular phenotype, which represents 14% of the total independent lines (20 cell lines), showed an unexpected, high level of UMP synthase and was therefore analyzed in detail. The selected cell lines showed no differences with wild-type cells with respect to uptake of orotic acid, affinity of UMP synthase for its substrates, or UMP synthase gene-copy number. Alternative detoxification mechanisms were also excluded. The elevated enzyme activity was correlated with elevated UMP synthase protein levels as well as elevated UMP synthase mRNA levels. In contrast to wild-type cell lines, the fluoroorotic acid-selected cell lines did not respond to thymine or to other biochemicals that affect thymine levels. In addition, there was also a concomitant up-regulation of aspartate transcarbamoylase, however, dihydroorotase and dihydroorotate dehydrogenase are not up-regulated in these cell lines.

Fluoroorotic Acid-Selected Nicotiana plumbaginifolia Cell Lines with a Stable Thymine Starvation Phenotype Have Lost the Thymine-Regulated Transcriptional Program1

PubMed Central

Santoso, Djoko; Thornburg, Robert

2000-01-01

We have selected 143 independent Nicotiana plumbaginifolia cell lines that survive in the presence of 5-fluoroorotic acid. These lines show several diverse phenotypes. The majority of these cell lines showed reduced levels of UMP synthase. However, one particular phenotype, which represents 14% of the total independent lines (20 cell lines), showed an unexpected, high level of UMP synthase and was therefore analyzed in detail. The selected cell lines showed no differences with wild-type cells with respect to uptake of orotic acid, affinity of UMP synthase for its substrates, or UMP synthase gene-copy number. Alternative detoxification mechanisms were also excluded. The elevated enzyme activity was correlated with elevated UMP synthase protein levels as well as elevated UMP synthase mRNA levels. In contrast to wild-type cell lines, the fluoroorotic acid-selected cell lines did not respond to thymine or to other biochemicals that affect thymine levels. In addition, there was also a concomitant up-regulation of aspartate transcarbamoylase, however, dihydroorotase and dihydroorotate dehydrogenase are not up-regulated in these cell lines. PMID:10938367

[Apoptosis: cellular and clinical aspects].

PubMed

Løvschall, H; Mosekilde, L

1997-04-01

Removal of damaged cells is essential for the maintenance of life in multicellular organisms. The process of self destruction, apoptosis, eliminates surplus or damaged cells as part of the pathophysiological defence system. Apoptosis is essential in structural and functional organogenesis during embryological development. The physiological regulation of tissue kinetics is a product of both cell proliferation and cell death. Internal and external regulatory stimuli regulate the balance between apoptosis and mitosis by genetic interaction. Apoptosis is characterized by condensation of chromatine as a result of DNA degradation, formation of blebs in the plasma and nuclear membranes, condensation of cytoplasma, formation of vesicular apoptotic bodies, and phagocytosis by neighbouring cells without inflammatory response. A number of observations indicate that programmed cell death plays an important role in the regulation of cytofunctional homeostasis and defense against accumulation of damaged cells, eg with DNA alterations. Dysregulation of the apoptotic gene program, eg by mutations, may not only lead to loss or degeneration of tissue, but also to hyperproliferative and tumorigenic disorders. New evidence indicates that apoptosis regulation is important both in aging processes and diseases such as: neuropathies, immunopathies, viral infections, cancer, etc. Pharmacological intervention designed to modulate apoptosis seems to raise new possibilities in the treatment of disease.

Regulation of Sodium Transport in the Inner Ear

PubMed Central

Kim, Sung Huhn; Marcus, Daniel C.

2011-01-01

Na+ concentrations in endolymph must be controlled to maintain hair cell function since the transduction channels of hair cells are cation-permeable, but not K+-selective. Flooding or fluctuations of the hair cell cytosol with Na+ would be expected to lead to cellular dysfunction, hearing loss and vertigo. This review briefly describes cellular mechanisms known to be responsible for Na+homeostasis in each compartment of the inner ear, including the cochlea, saccule, semicircular canals and endolymphatic sac. The influx of Na+into endolymph of each of the organs is likely via passive diffusion, but these pathways have not yet been identified or characterized. Na+ absorption is controlled by gate -keeper channels in the apical (endolymphatic) membrane of the transporting cells. Highly Na+-selective epithelial sodium channels (ENaC) control absorption by Reissner’s membrane, saccular extramacular epithelium, semicircular canal duct epithelium and endolymphatic sac. ENaC activity is controlled by a number of signal pathways, but most notably by genomic regulation of channel numbers in the membrane via glucocorticoid signaling. Nonselective cation channels in the apical membrane of outer sulcus epithelial cells and vestibular transitional cells mediate Na+ and parasensory K+ absorption. The K+-mediated transduction current in hair cells is also accompanied by a Na+ flux since the transduction channels are nonselective cation channels. Cation absorption by all of these cells is regulated by extracellular ATP via apical nonselective cation channels (P2X receptors). The heterogeneous population of epithelial cells in the endolymphatic sac is thought to have multiple absorptive pathways for Na+ with regulatory pathways that include glucocorticoids and purinergic agonists. PMID:21620939

Mucosal tolerance to experimental autoimmune myasthenia gravis is associated with down-regulation of AChR-specific IFN-gamma-expressing Th1-like cells and up-regulation of TGF-beta mRNA in mononuclear cells.

PubMed

Ma, C G; Zhang, G X; Xiao, B G; Wang, Z Y; Link, J; Olsson, T; Link, H

1996-02-13

Oral and nasal administration of nicotinic acetylcholine receptor (AChR) to Lewis rats prior to myasthenogenic immunization with AChR and complete Freund's adjuvant (CFA) resulted in prevention or marked decrease of the severity of experimental autoimmune myasthenia gravis (EAMG) and suppression of AChR-specific B-cell responses and of AChR-reactive T-cell function. To examine the involvement of immunoregulatory cytokines and the underlying mechanisms involved in tolerance induction, in situ hybridization with radiolabeled cDNA oligonucleotide proves was adopted to enumerate mononuclear cells (MNC) expressing mRNA for the proinflammatory cytokine interferon-gamma (IFN-gamma), the B cell-stimulating interleukin-4 (IL-4), and the immunosuppressive transforming growth factor-beta (TGF-beta). Popliteal and inguinal lymph nodes from EAMG rats contained elevated numbers of AChR-reactive IFN-gamma, IL-4, and TGF-beta mRNA-expressing cells, compared to control rats receiving PBS orally or nasally and injected with CFA only. Oral and nasal tolerance was accompanied by decreased numbers of AChR-reactive IFN-gamma and IL-4 mRNA-expressing cells and strong up-regulation of TGF-beta mRNA-positive cells in lymphoid organs when compared to nontolerized EAMG control rats. The results suggest that IFN-gamma and IL-4 are central effector molecules in the development of EAMG and that TGF-beta plays an important role in tolerance induction to EAMG.

The intestinal B-cell response in celiac disease

PubMed Central

Mesin, Luka; Sollid, Ludvig M.; Niro, Roberto Di

2012-01-01

The function of intestinal immunity is to provide protection toward pathogens while preserving the composition of the microflora and tolerance to orally fed nutrients. This is achieved via a number of tightly regulated mechanisms including production of IgA antibodies by intestinal plasma cells. Celiac disease is a common gut disorder caused by a dysfunctional immune regulation as signified, among other features, by a massive intestinal IgA autoantibody response. Here we review the current knowledge of this B-cell response and how it is induced, and we discuss key questions to be addressed in future research. PMID:23060888

Protein phosphatase 2A: a highly regulated family of serine/threonine phosphatases implicated in cell growth and signalling.

PubMed Central

Janssens, V; Goris, J

2001-01-01

Protein phosphatase 2A (PP2A) comprises a family of serine/threonine phosphatases, minimally containing a well conserved catalytic subunit, the activity of which is highly regulated. Regulation is accomplished mainly by members of a family of regulatory subunits, which determine the substrate specificity, (sub)cellular localization and catalytic activity of the PP2A holoenzymes. Moreover, the catalytic subunit is subject to two types of post-translational modification, phosphorylation and methylation, which are also thought to be important regulatory devices. The regulatory ability of PTPA (PTPase activator), originally identified as a protein stimulating the phosphotyrosine phosphatase activity of PP2A, will also be discussed, alongside the other regulatory inputs. The use of specific PP2A inhibitors and molecular genetics in yeast, Drosophila and mice has revealed roles for PP2A in cell cycle regulation, cell morphology and development. PP2A also plays a prominent role in the regulation of specific signal transduction cascades, as witnessed by its presence in a number of macromolecular signalling modules, where it is often found in association with other phosphatases and kinases. Additionally, PP2A interacts with a substantial number of other cellular and viral proteins, which are PP2A substrates, target PP2A to different subcellular compartments or affect enzyme activity. Finally, the de-regulation of PP2A in some specific pathologies will be touched upon. PMID:11171037

The Cdk4-E2f1 pathway regulates early pancreas development by targeting Pdx1+ progenitors and Ngn3+ endocrine precursors

PubMed Central

Kim, So Yoon; Rane, Sushil G.

2011-01-01

Cell division and cell differentiation are intricately regulated processes vital to organ development. Cyclin-dependent kinases (Cdks) are master regulators of the cell cycle that orchestrate the cell division and differentiation programs. Cdk1 is essential to drive cell division and is required for the first embryonic divisions, whereas Cdks 2, 4 and 6 are dispensable for organogenesis but vital for tissue-specific cell development. Here, we illustrate an important role for Cdk4 in regulating early pancreas development. Pancreatic development involves extensive morphogenesis, proliferation and differentiation of the epithelium to give rise to the distinct cell lineages of the adult pancreas. The cell cycle molecules that specify lineage commitment within the early pancreas are unknown. We show that Cdk4 and its downstream transcription factor E2f1 regulate mouse pancreas development prior to and during the secondary transition. Cdk4 deficiency reduces embryonic pancreas size owing to impaired mesenchyme development and fewer Pdx1+ pancreatic progenitor cells. Expression of activated Cdk4R24C kinase leads to increased Nkx2.2+ and Nkx6.1+ cells and a rise in the number and proliferation of Ngn3+ endocrine precursors, resulting in expansion of the β cell lineage. We show that E2f1 binds and activates the Ngn3 promoter to modulate Ngn3 expression levels in the embryonic pancreas in a Cdk4-dependent manner. These results suggest that Cdk4 promotes β cell development by directing E2f1-mediated activation of Ngn3 and increasing the pool of endocrine precursors, and identify Cdk4 as an important regulator of early pancreas development that modulates the proliferation potential of pancreatic progenitors and endocrine precursors. PMID:21490060

Cell life and death in the anterior pituitary gland: role of oestrogens.

PubMed

Seilicovich, A

2010-07-01

Apoptotic processes play an important role in the maintenance of cell numbers in the anterior pituitary gland during physiological endocrine events. In this review, we summarise the regulation of apoptosis of anterior pituitary cells, particularly lactotrophs, somatotrophs and gonadotrophs, and analyse the possible mechanisms involved in oestrogen-induced apoptosis in anterior pituitary cells. Oestrogens exert apoptotic actions in several cell types and act as modulators of pituitary cell renewal, sensitising cells to both mitogenic and apoptotic signals. Local synthesis of growth factors and cytokines induced by oestradiol as well as changes in phenotypic features that enhance the responsiveness of anterior pituitary cells to pro-apoptotic factors may account for cyclical apoptotic activity in anterior pituitary cells during the oestrous cycle. Considering that tissue homeostasis results from a balance between cell proliferation and death and that mechanisms involved in apoptosis are tightly regulated, defects in cell death processes could have a considerable physiopathological impact.

Integrated analysis of chromosome copy number variation and gene expression in cervical carcinoma

PubMed Central

Yan, Deng; Yi, Song; Chiu, Wang Chi; Qin, Liu Gui; Kin, Wong Hoi; Kwok Hung, Chung Tony; Linxiao, Han; Wai, Choy Kwong; Yi, Sui; Tao, Yang; Tao, Tang

2017-01-01

Objective This study was conducted to explore chromosomal copy number variations (CNV) and transcript expression and to examine pathways in cervical pathogenesis using genome-wide high resolution microarrays. Methods Genome-wide chromosomal CNVs were investigated in 6 cervical cancer cell lines by Human Genome CGH Microarray Kit (4x44K). Gene expression profiles in cervical cancer cell lines, primary cervical carcinoma and normal cervical epithelium tissues were also studied using the Whole Human Genome Microarray Kit (4x44K). Results Fifty common chromosomal CNVs were identified in the cervical cancer cell lines. Correlation analysis revealed that gene up-regulation or down-regulation is significantly correlated with genomic amplification (P=0.009) or deletion (P=0.006) events. Expression profiles were identified through cluster analysis. Gene annotation analysis pinpointed cell cycle pathways was significantly (P=1.15E-08) affected in cervical cancer. Common CNVs were associated with cervical cancer. Conclusion Chromosomal CNVs may contribute to their transcript expression in cervical cancer. PMID:29312578

Biological pathways involved in the development of inflammatory bowel disease.

PubMed

Zemljic, Mateja; Pejkovic, Bozena; Krajnc, Ivan; Lipovsek, Saska

2014-10-01

Apoptosis, autophagy and necrosis are three distinct functional types of the mammalian cell death network. All of them are characterized by a number of cell's morphological changes. The inappropriate induction of cell death is involved in the pathogenesis of a number of diseases.Pathogenesis of inflammatory bowel diseases (ulcerative colitis, Crohn's disease) includes an abnormal immunological response to disturbed intestinal microflora. One of the most important reason in pathogenesis of chronic inflammatory disease and subsequent multiple organ pathology is a barrier function of the gut, regulating cellular viability. Recent findings have begun to explain the mechanisms by which intestinal epithelial cells are able to survive in such an environment and how loss of normal regulatory processes may lead to inflammatory bowel disease (IBD).This review focuses on the regulation of biological pathways in development and homeostasis in IBD. Better understanding of the physiological functions of biological pathways and their influence on inflammation, immunity, and barrier function will simplify our expertice of homeostasis in the gastrointestinal tract and in upgrading diagnosis and treatment.

Regulation of EGFR signal transduction by analogue-to-digital conversion in endosomes

PubMed Central

Villaseñor, Roberto; Nonaka, Hidenori; Del Conte-Zerial, Perla; Kalaidzidis, Yannis; Zerial, Marino

2015-01-01

An outstanding question is how receptor tyrosine kinases (RTKs) determine different cell-fate decisions despite sharing the same signalling cascades. Here, we uncovered an unexpected mechanism of RTK trafficking in this process. By quantitative high-resolution FRET microscopy, we found that phosphorylated epidermal growth factor receptor (p-EGFR) is not randomly distributed but packaged at constant mean amounts in endosomes. Cells respond to higher EGF concentrations by increasing the number of endosomes but keeping the mean p-EGFR content per endosome almost constant. By mathematical modelling, we found that this mechanism confers both robustness and regulation to signalling output. Different growth factors caused specific changes in endosome number and size in various cell systems and changing the distribution of p-EGFR between endosomes was sufficient to reprogram cell-fate decision upon EGF stimulation. We propose that the packaging of p-RTKs in endosomes is a general mechanism to ensure the fidelity and specificity of the signalling response. DOI: http://dx.doi.org/10.7554/eLife.06156.001 PMID:25650738

DNA Replication Control During Drosophila Development: Insights into the Onset of S Phase, Replication Initiation, and Fork Progression

PubMed Central

Hua, Brian L.; Orr-Weaver, Terry L.

2017-01-01

Proper control of DNA replication is critical to ensure genomic integrity during cell proliferation. In addition, differential regulation of the DNA replication program during development can change gene copy number to influence cell size and gene expression. Drosophila melanogaster serves as a powerful organism to study the developmental control of DNA replication in various cell cycle contexts in a variety of differentiated cell and tissue types. Additionally, Drosophila has provided several developmentally regulated replication models to dissect the molecular mechanisms that underlie replication-based copy number changes in the genome, which include differential underreplication and gene amplification. Here, we review key findings and our current understanding of the developmental control of DNA replication in the contexts of the archetypal replication program as well as of underreplication and differential gene amplification. We focus on the use of these latter two replication systems to delineate many of the molecular mechanisms that underlie the developmental control of replication initiation and fork elongation. PMID:28874453

Integrated analysis of chromosome copy number variation and gene expression in cervical carcinoma.

PubMed

Yan, Deng; Yi, Song; Chiu, Wang Chi; Qin, Liu Gui; Kin, Wong Hoi; Kwok Hung, Chung Tony; Linxiao, Han; Wai, Choy Kwong; Yi, Sui; Tao, Yang; Tao, Tang

2017-12-12

This study was conducted to explore chromosomal copy number variations (CNV) and transcript expression and to examine pathways in cervical pathogenesis using genome-wide high resolution microarrays. Genome-wide chromosomal CNVs were investigated in 6 cervical cancer cell lines by Human Genome CGH Microarray Kit (4x44K). Gene expression profiles in cervical cancer cell lines, primary cervical carcinoma and normal cervical epithelium tissues were also studied using the Whole Human Genome Microarray Kit (4x44K). Fifty common chromosomal CNVs were identified in the cervical cancer cell lines. Correlation analysis revealed that gene up-regulation or down-regulation is significantly correlated with genomic amplification ( P =0.009) or deletion ( P =0.006) events. Expression profiles were identified through cluster analysis. Gene annotation analysis pinpointed cell cycle pathways was significantly ( P =1.15E-08) affected in cervical cancer. Common CNVs were associated with cervical cancer. Chromosomal CNVs may contribute to their transcript expression in cervical cancer.

Cullin-4 regulates Wingless and JNK signaling-mediated cell death in the Drosophila eye

PubMed Central

Tare, Meghana; Sarkar, Ankita; Bedi, Shimpi; Kango-Singh, Madhuri; Singh, Amit

2016-01-01

In all multicellular organisms, the fundamental processes of cell proliferation and cell death are crucial for growth regulation during organogenesis. Strict regulation of cell death is important to maintain tissue homeostasis by affecting processes like regulation of cell number, and elimination of unwanted/unfit cells. The developing Drosophila eye is a versatile model to study patterning and growth, where complex signaling pathways regulate growth and cell survival. However, the molecular mechanisms underlying regulation of these processes is not fully understood. In a gain-of-function screen, we found that misexpression of cullin-4 (cul-4), an ubiquitin ligase, can rescue reduced eye mutant phenotypes. Previously, cul-4 has been shown to regulate chromatin remodeling, cell cycle and cell division. Genetic characterization of cul-4 in the developing eye revealed that loss-of-function of cul-4 exhibits a reduced eye phenotype. Analysis of twin-spots showed that in comparison with their wild-type counterparts, the cul-4 loss-of-function clones fail to survive. Here we show that cul-4 clones are eliminated by induction of cell death due to activation of caspases. Aberrant activation of signaling pathways is known to trigger cell death in the developing eye. We found that Wingless (Wg) and c-Jun-amino-terminal-(NH2)-Kinase (JNK) signaling are ectopically induced in cul-4 mutant clones, and these signals co-localize with the dying cells. Modulating levels of Wg and JNK signaling by using agonists and antagonists of these pathways demonstrated that activation of Wg and JNK signaling enhances cul-4 mutant phenotype, whereas downregulation of Wg and JNK signaling rescues the cul-4 mutant phenotypes of reduced eye. Here we present evidences to demonstrate that cul-4 is involved in restricting Wg signaling and downregulation of JNK signaling-mediated cell death during early eye development. Overall, our studies provide insights into a novel role of cul-4 in promoting cell survival in the developing Drosophila eye. PMID:28032862

Quantitative Phosphoproteomics Reveals SLP-76 Dependent Regulation of PAG and Src Family Kinases in T Cells

PubMed Central

Cao, Lulu; Ding, Yiyuan; Hung, Norris; Yu, Kebing; Ritz, Anna; Raphael, Benjamin J.; Salomon, Arthur R.

2012-01-01

The SH2-domain-containing leukocyte protein of 76 kDa (SLP-76) plays a critical scaffolding role in T cell receptor (TCR) signaling. As an adaptor protein that contains multiple protein-binding domains, SLP-76 interacts with many signaling molecules and links proximal receptor stimulation to downstream effectors. The function of SLP-76 in TCR signaling has been widely studied using the Jurkat human leukaemic T cell line through protein disruption or site-directed mutagenesis. However, a wide-scale characterization of SLP-76-dependant phosphorylation events is still lacking. Quantitative profiling of over a hundred tyrosine phosphorylation sites revealed new modes of regulation of phosphorylation of PAG, PI3K, and WASP while reconfirming previously established regulation of Itk, PLCγ, and Erk phosphorylation by SLP-76. The absence of SLP-76 also perturbed the phosphorylation of Src family kinases (SFKs) Lck and Fyn, and subsequently a large number of SFK-regulated signaling molecules. Altogether our data suggests unique modes of regulation of positive and negative feedback pathways in T cells by SLP-76, reconfirming its central role in the pathway. PMID:23071622

Quantitative phosphoproteomics reveals SLP-76 dependent regulation of PAG and Src family kinases in T cells.

PubMed

Cao, Lulu; Ding, Yiyuan; Hung, Norris; Yu, Kebing; Ritz, Anna; Raphael, Benjamin J; Salomon, Arthur R

2012-01-01

The SH2-domain-containing leukocyte protein of 76 kDa (SLP-76) plays a critical scaffolding role in T cell receptor (TCR) signaling. As an adaptor protein that contains multiple protein-binding domains, SLP-76 interacts with many signaling molecules and links proximal receptor stimulation to downstream effectors. The function of SLP-76 in TCR signaling has been widely studied using the Jurkat human leukaemic T cell line through protein disruption or site-directed mutagenesis. However, a wide-scale characterization of SLP-76-dependant phosphorylation events is still lacking. Quantitative profiling of over a hundred tyrosine phosphorylation sites revealed new modes of regulation of phosphorylation of PAG, PI3K, and WASP while reconfirming previously established regulation of Itk, PLCγ, and Erk phosphorylation by SLP-76. The absence of SLP-76 also perturbed the phosphorylation of Src family kinases (SFKs) Lck and Fyn, and subsequently a large number of SFK-regulated signaling molecules. Altogether our data suggests unique modes of regulation of positive and negative feedback pathways in T cells by SLP-76, reconfirming its central role in the pathway.

Mechanical influence of tissue culture plates and extracellular matrix on mesenchymal stem cell behavior: A topical review.

PubMed

Tatullo, Marco; Marrelli, Massimo; Falisi, Giovanni; Rastelli, Claudio; Palmieri, Francesca; Gargari, Marco; Zavan, Barbara; Paduano, Francesco; Benagiano, Vincenzo

2016-03-01

Tissue engineering applications need a continuous development of new biomaterials able to generate an ideal cell-extracellular matrix interaction. The stem cell fate is regulated by several factors, such as growth factors or transcription factors. The most recent literature has reported several publications able to demonstrate that environmental factors also contribute to the regulation of stem cell behavior, leading to the opinion that the environment plays the major role in the cell differentiation.The interaction between mesenchymal stem cells (MSCs) and extracellular environment has been widely described, and it has a crucial role in regulating the cell phenotype. In our laboratory (Tecnologica Research Institute, Crotone, Italy), we have recently studied how several physical factors influence the distribution and the morphology of MSCs isolated from dental pulp, and how they are able to regulate stem cell differentiation. Mechanical and geometrical factors are only a small part of the environmental factors able to influence stem cell behavior, however, this influence should be properly known: in fact, this assumption must be clearly considered during those studies involving MSCs; furthermore, these interactions should be considered as an important bias that involves an high number of studies on the MSCs, since in worldwide laboratories the scientists mostly use tissue culture plates for their experiments. © The Author(s) 2015.

Metabolic Plasticity in Cancer Cells: Reconnecting Mitochondrial Function to Cancer Control

PubMed Central

Ramanujan, V. Krishnan

2015-01-01

Anomalous increase in glycolytic activity defines one of the key metabolic alterations in cancer cells. A realization of this feature has led to critical advancements in cancer detection techniques such as positron emission tomography (PET) as well as a number of therapeutic avenues targeting the key glycolytic steps within a cancer cell. A normal healthy cell’s survival relies on a sensitive balance between the primordial glycolysis and a more regulated mitochondrial bioenergetics. The salient difference between these two bioenergetics pathways is that oxygen availability is an obligatory requirement for mitochondrial pathway while glycolysis can function without oxygen. Early observations that some cancer cells up-regulate glycolytic activity even in the presence of oxygen (aerobic glycolysis) led to a hypothesis that such an altered cancer cell metabolism stems from inherent mitochondrial dysfunction. While a general validity of this hypothesis is still being debated, a number of recent research efforts have yielded clarity on the physiological origins of this aerobic glycolysis phenotype in cancer cells. Building on these recent studies, we present a generalized scheme of cancer cell metabolism and propose a novel hypothesis that might rationalize new avenues of cancer intervention. PMID:26457230

Neuroimmune modulation of gut function

USDA-ARS?s Scientific Manuscript database

There is considerable interest in the mechanisms and pathways involved in the neuro-immune regulation of gut function. The number of cell types and possible interactions is staggering and there are a number of recent reviews detailing various aspects of these interactions, many of which focus on ...

Activating receptors promote NK cell expansion for maintenance, IL-10 production, and CD8 T cell regulation during viral infection.

PubMed

Lee, Seung-Hwan; Kim, Kwang-Sin; Fodil-Cornu, Nassima; Vidal, Silvia M; Biron, Christine A

2009-09-28

Natural killer (NK) cells have the potential to deliver both direct antimicrobial effects and regulate adaptive immune responses, but NK cell yields have been reported to vary greatly during different viral infections. Activating receptors, including the Ly49H molecule recognizing mouse cytomegalovirus (MCMV), can stimulate NK cell expansion. To define Ly49H's role in supporting NK cell proliferation and maintenance under conditions of uncontrolled viral infection, experiments were performed in Ly49h(-/-), perforin 1 (Prf1)(-/-), and wild-type (wt) B6 mice. NK cell numbers were similar in uninfected mice, but relative to responses in MCMV-infected wt mice, NK cell yields declined in the absence of Ly49h and increased in the absence of Prf1, with high rates of proliferation and Ly49H expression on nearly all cells. The expansion was abolished in mice deficient for both Ly49h and Prf1 (Ly49h(-/-)Prf1(-/-)), and negative consequences for survival were revealed. The Ly49H-dependent protection mechanism delivered in the absence of Prf1 was a result of interleukin 10 production, by the sustained NK cells, to regulate the magnitude of CD8 T cell responses. Thus, the studies demonstrate a previously unappreciated critical role for activating receptors in keeping NK cells present during viral infection to regulate adaptive immune responses.

Rho/ROCK signaling in regulation of corneal epithelial cell cycle progression.

PubMed

Chen, Jian; Guerriero, Emily; Lathrop, Kira; SundarRaj, Nirmala

2008-01-01

The authors' previous study showed that the expression of a Rho-associated serine/threonine kinase (ROCK) is regulated during cell cycle progression in corneal epithelial cells. The present study was conducted to determine whether and how Rho/ROCK signaling regulates cell cycle progression. Rabbit corneal epithelial cells (RCECs) in culture were arrested in the G(0) phase of the cell cycle by serum deprivation and then allowed to re-enter the cell cycle in the presence or absence of the ROCK inhibitor (Y27632) in serum-supplemented medium. The number of cells in the S phase, the relative levels of specific cyclins and CDKs and their intracellular distribution, and the relative levels of mRNAs were determined by BrdU labeling, Western blot and immunocytochemical analyses, and real-time RT-PCR, respectively. ROCK inhibition delayed the progression of G(1) to S phase and led to a decrease in the number of RCECs entering the S phase between 12 and 24 hours from 31.5% +/- 4.5% to 8.1% +/- 2.6%. During the cell cycle progression, protein and mRNA levels of cyclin-D1 and -D3 and cyclin-dependent kinases CDK4 and CDK6 were significantly lower, whereas the protein levels of the CDK inhibitor p27(Kip1) were higher in ROCK-inhibited cells. Intracellular mRNA or protein levels of cyclin-E and protein levels of CDK2 were not significantly affected, but their nuclear translocation was delayed by ROCK inhibition. ROCK signaling is involved in cell cycle progression in RCECs, possibly by upregulation of cyclin-D1 and -D3 and CDK4, -6, and -2; nuclear translocation of CDK2 and cyclin-E; and downregulation of p27(Kip1).

Generation of mice deficient in RNA-binding motif protein 3 (RBM3) and characterization of its role in innate immune responses and cell growth

DOE Office of Scientific and Technical Information (OSTI.GOV)

Matsuda, Atsushi; Core Research for Evolution Science and Technology, Japan Science and Technology Agency, Chiyoda-ku, Tokyo 102-0075; Ogawa, Masahiro

Highlights: {yields} We identified RNA-binding motif protein 3 (RBM3) as CpG-B DNA-binding protein. {yields} RBM3 translocates from the nucleus to the cytoplasm and co-localized with CpG-B DNA. {yields} We newly generated Rbm3-deficient (Rbm3{sup -/-}) mice. {yields} DNA-mediated cytokine gene induction was normally occured in Rbm3{sup -/-} cells. {yields}Rbm3{sup -/-} MEFs showed poorer proliferation rate and increased number of G2-phase cells. -- Abstract: The activation of innate immune responses is critical to host defense against microbial infections, wherein nucleic acid-sensing pattern recognition receptors recognize DNA or RNA from viruses or bacteria and activate downstream signaling pathways. In a search for newmore » DNA-sensing molecules that regulate innate immune responses, we identified RNA-binding motif protein 3 (RBM3), whose role has been implicated in the regulation of cell growth. In this study, we generated Rbm3-deficient (Rbm3{sup -/-}) mice to study the role of RBM3 in immune responses and cell growth. Despite evidence for its interaction with immunogenic DNA in a cell, no overt phenotypic abnormalities were found in cells from Rbm3{sup -/-} mice for the DNA-mediated induction of cytokine genes. Interestingly, however, Rbm3{sup -/-} mouse embryonic fibroblasts (MEFs) showed poorer proliferation rates as compared to control MEFs. Further cell cycle analysis revealed that Rbm3{sup -/-} MEFs have markedly increased number of G2-phase cells, suggesting a hitherto unknown role of RBM3 in the G2-phase control. Thus, these mutant mice and cells may provide new tools with which to study the mechanisms underlying the regulation of cell cycle and oncogenesis.« less

Zinc enhances the number of regulatory T cells in allergen-stimulated cells from atopic subjects.

PubMed

Rosenkranz, Eva; Hilgers, Ralf-Dieter; Uciechowski, Peter; Petersen, Arnd; Plümäkers, Birgit; Rink, Lothar

2017-03-01

The trace element zinc is essential for immune function and its regulation. Since zinc deficiency and allergic hyperresponsive reactions are often accompanied, the influence of zinc on allergen-induced cell growth, CD4+ regulatory T (Treg) cell numbers and cytokine expression during allergic immune reactions was investigated. Peripheral blood mononuclear cells (PBMCs) from non-atopic and atopic subjects were treated with timothy grass allergen pre-incubated with or without zinc. Proliferation was determined by analyzing the incorporation of 3 H-thymidine. Intracellular zinc and Foxp3 levels and cell surface antigens were measured by FACS, cytokine expression by ELISA and real-time PCR. Incubation with 50 μM zinc sulfate (Zn50) enhances cytosolic zinc concentrations in CD3+ T cells. The data also reveal that the combination of Zn50 plus allergen significantly reduces PBMC proliferation of atopic subjects. Additionally, Zn50 plus allergen enhances Th1 cytokine responses shown by increased interferon (IFN)-γ/interleukin (IL)-10 ratios as well as enhanced tumor necrosis factor-α release. In response to allergen, zinc increases Treg cells and upregulates the mRNA expression of cytotoxic T-lymphocyte antigen-4 in atopic subjects. Interestingly, Zn50 alone leads to an increase of CD4+CD25high(hi)+ cells in atopic and non-atopic subjects. Zinc may regulate unwanted hyperresponsive immune reactions by suppressing proliferation through a significant shift from IL-10 to the Th1 cytokine IFN-γ, and enhanced regulatory T cell numbers. Therefore, zinc supplementation may be a promising tool for the therapy of allergies, without negatively affecting the immune system.

Genitourinary mast cells and survival

PubMed Central

Stewart, Julia M.

2015-01-01

Mast cells (MCs) are ubiquitous in the body, but they have historically been associated with allergies, and most recently with regulation of immunity and inflammation. However, it remains a puzzle why so many MCs are located in the diencephalon, which regulates emotions and in the genitourinary tract, including the bladder, prostate, penis, vagina and uterus that hardly ever get allergic reactions. A number of papers have reported that MCs have estrogen, gonadotropin and corticotropin-releasing hormone (CRH) receptors. Moreover, animal experiments have shown that diencephalic MCs increase in number during courting in doves. We had reported that allergic stimulation of nasal MCs leads to hypothalamic-pituitary adrenal (HPA) activation. Interestingly, anecdotal information indicates that female patients with mastocytosis or mast cell activation syndrome may have increased libido. Preliminary evidence also suggests that MCs may have olfactory receptors. MCs may, therefore, have been retained phylogenetically not only to “smell danger”, but to promote survival and procreation. PMID:26813805

Genitourinary mast cells and survival.

PubMed

Theoharides, Theoharis C; Stewart, Julia M

2015-10-01

Mast cells (MCs) are ubiquitous in the body, but they have historically been associated with allergies, and most recently with regulation of immunity and inflammation. However, it remains a puzzle why so many MCs are located in the diencephalon, which regulates emotions and in the genitourinary tract, including the bladder, prostate, penis, vagina and uterus that hardly ever get allergic reactions. A number of papers have reported that MCs have estrogen, gonadotropin and corticotropin-releasing hormone (CRH) receptors. Moreover, animal experiments have shown that diencephalic MCs increase in number during courting in doves. We had reported that allergic stimulation of nasal MCs leads to hypothalamic-pituitary adrenal (HPA) activation. Interestingly, anecdotal information indicates that female patients with mastocytosis or mast cell activation syndrome may have increased libido. Preliminary evidence also suggests that MCs may have olfactory receptors. MCs may, therefore, have been retained phylogenetically not only to "smell danger", but to promote survival and procreation.

Dynamics of cell proliferation in the adult dentate gyrus of two inbred strains of mice

NASA Technical Reports Server (NTRS)

Hayes, N. L.; Nowakowski, R. S.

2002-01-01

The output potential of proliferating populations in either the developing or the adult nervous system is critically dependent on the length of the cell cycle (T(c)) and the size of the proliferating population. We developed a new approach for analyzing the cell cycle, the 'Saturate and Survive Method' (SSM), that also reveals the dynamic behaviors in the proliferative population and estimates of the size of the proliferating population. We used this method to analyze the proliferating population of the adult dentate gyrus in 60 day old mice of two inbred strains, C57BL/6J and BALB/cByJ. The results show that the number of cells labeled by exposure to BUdR changes dramatically with time as a function of the number of proliferating cells in the population, the length of the S-phase, cell division, the length of the cell cycle, dilution of the S-phase label, and cell death. The major difference between C57BL/6J and BALB/cByJ mice is the size of the proliferating population, which differs by a factor of two; the lengths of the cell cycle and the S-phase and the probability that a newly produced cell will die within the first 10 days do not differ in these two strains. This indicates that genetic regulation of the size of the proliferating population is independent of the genetic regulation of cell death among those newly produced cells. The dynamic changes in the number of labeled cells as revealed by the SSM protocol also indicate that neither single nor repeated daily injections of BUdR accurately measure 'proliferation.'.

Determination of the Absolute Number of Cytokine mRNA Molecules within Individual Activated Human T Cells

NASA Technical Reports Server (NTRS)

Karr, Laurel J.; Marshall, Gwen; Hockett, Richard D.; Bucy, R. Pat; Curreri, Peter A. (Technical Monitor)

2002-01-01

A primary function of activated T cells is the expression and subsequent secretion of cytokines, which orchestrate the differentiation of other lymphocytes, modulate antigen presenting cell activity, and alter vascular endothelium to mediate an immune response. Since many features of immune regulation probably result from modest alterations of endogenous rates of multiple interacting processes, quantitative analysis of the frequency and specific activity of individual T cells is critically important. Using a coordinated set of quantitative methods, the absolute number of molecules of several key cytokine mRNA species in individual T cells has been determined. The frequency of human blood T cells activated in vitro by mitogens and recall protein antigens was determined by intracellular cytokine protein staining, in situ hybridization for cytokine mRNA, and by limiting dilution analysis for cytokine mRNA+ cells. The absolute number of mRNA molecules was simultaneously determined in both homogenates of the entire population of cells and in individual cells obtained by limiting dilution, using a quantitative, competitive RT-PCR assay. The absolute numbers of mRNA molecules in a population of cells divided by the frequency of individual positive cells, yielded essentially the same number of mRNA molecules per cell as direct analysis of individual cells by limiting dilution analysis. Mean numbers of mRNA per positive cell from both mitogen and antigen activated T cells, using these stimulation conditions, were 6000 for IL-2, 6300 for IFN-gamma, and 1600 for IL-4.

Coexistence of proguanylin (1-15) and somatostatin in the gastrointestinal tract.

PubMed

Ieda, H; Naruse, S; Furuya, S; Ozaki, T; Ando, E; Nokihara, K; Hori, S; Kitagawa, M; Hayakawa, T

1998-12-01

In order to identify proguanylin-secreting cells, we have raised an antiserum against the synthetic fragment of human proguanylin (1-15) and have examined the proguanylin-positive cells in the human and rat gastrointestinal tract by immunohistochemical methods. Numerous proguanylin (1-15)-immunoreactive cells were found in the gastrointestinal tract. They were either pyramidal or spindle shaped in the stomach. Spindle-shaped cells, frequently possessing long slender processes, were located at the base of the pyloric epithelium and did not extend to the lumen. In the duodenum and jejunum, these cells were mostly pyramidal in shape and often had a slender process towards the lumen. The immunostaining was completely blocked by the human proguanylin (1-15) fragment. Paneth and goblet cells were negative against this antiserum. The number of serotonin-positive cells was much larger than that of proguanylin-positive cells in all the segments tested. The number of proguanylin-positive cells decreased from the jejunum to the ileum and very few cells were observed in the colon. In contrast to serotonin-positive cells, most somatostatin-positive cells were also positive for proguanylin. Thus, proguanylin (1-15) or its related protein appears to coexist with somatostatin in intestinal endocrine D cells which may be a source of circulating proguanylin. Proguanylin, like somatostatin, may also regulate intestinal function as a local regulator.

Regulation of gonadotropin receptors on cultured porcine Leydig and Sertoli cells: effect of potassium depletion.

PubMed

Bernier, M; Laferrere, B; Jaillard, C; Clerget, M; Saez, J M

1986-06-01

We have examined the role of the NaK-ATPase pump activity on the ligand-induced down-regulation of gonadotropin receptors in cultured porcine Leydig and Sertoli cells. In both cells, inhibition of the NaK pump by ouabain produced a depletion of intracellular K+ levels (ID50, 10(-7) M) after a lag period of about 8 h. In the absence of ligand, the number of FSH receptors in ouabain-treated Sertoli cells was unaffected or slightly reduced, whereas a 2-fold increase in the number of human CG (hCG)/LH receptors with small changes in the binding affinity was observed in Leydig cells treated by ouabain. The effect of ouabain was dose dependent. Differences were also observed in the down-regulation process of gonadotropin receptors in ouabain-treated cells. The hCG-induced receptor loss in Leydig cells was completely reversed by ouabain whereas the drug had no effect on ligand-induced loss of FSH receptors in Sertoli cells. Similar results were observed when the cells were incubated in K+-free medium. Kinetics studies with labeled hCG have shown that ouabain treatment slows down significantly the rate of [125I]iodo-hCG internalization (t 1/2, 18 h; control cells, t 1/2, 6 h), but had no effect on the degradation of internalized hormone. The internalization of receptor-bound [125I]iodo-hCG was also reduced when Leydig cells were incubated in K+-free medium, but was restored when this medium was supplemented with rubidium. The influence of the NaK pump on the receptor regulation of a ligand common to both types of cells, such as epidermal growth factor, was studied under the same experimental conditions. Neither ouabain nor K+-free medium were able to prevent the epidermal growth factor-induced reduction of receptor levels in Leydig and Sertoli cells. Thus, it appears that modulation of ligand-induced receptor loss by depletion of cellular K+ levels is not dependent on the cell type, but on the ligand-receptor complex. The data also show a striking difference in the dynamics of gonadotropin-receptor interaction of two structurally related hormones.

BARHL2 differentially regulates the development of retinal amacrine and ganglion neurons

PubMed Central

Ding, Qian; Chen, Hui; Xie, Xiaoling; Libby, Richard T.; Tian, Ning; Gan, Lin

2009-01-01

Summary Through transcriptional regulations the BarH family of homeodomain proteins play essential roles in cell fate specification, cell differentiation, migration and survival. Barhl2, a member of the Barh gene family, is expressed in retinal ganglion cells (RGCs), amacrine cells (ACs) and horizontal cells. Here, to investigate the role of Barhl2 in retinal development, Barhl2 deficient mice were generated. Analysis of AC subtypes in Barhl2 deficient retinas suggests that Barhl2 plays a critical role in AC subtype determination. A significant reduction of glycinergic and GABAergic ACs with a substantial increase in the number of cholinergic ACs was observed in Barhl2-null retinas. Barhl2 is also critical for the development of a normal complement of RGCs. Barhl2 deficiency resulted in a 35% increase in RGCs undergoing apoptosis during development. Genetic analysis revealed that Barhl2 functions downstream of the Atoh7-Pou4f3 regulatory pathway and regulates the maturation and/or survival of RGCs. Thus, BARHL2 appears to have numerous roles in retinal development, including regulating neuronal subtype specification, differentiation, and survival. PMID:19339595

Yeast as a model to study apoptosis?

PubMed

Fleury, Christophe; Pampin, Mathieu; Tarze, Agathe; Mignotte, Bernard

2002-02-01

Programmed cell death (PCD) serves as a major mechanism for the precise regulation of cell numbers, and as a defense mechanism to remove unwanted and potentially dangerous cells. Despite the striking heterogeneity of cell death induction pathways, the execution of the death program is often associated with characteristic morphological and biochemical changes termed apoptosis. Although for a long time the absence of mitochondrial changes was considered as a hallmark of apoptosis, mitochondria appear today as the central executioner of programmed cell death. This crucial position of mitochondria in programmed cell death control is not due to a simple loss of function (deficit in energy supplying), but rather to an active process in the regulation of effector mechanisms. The large diversity of regulators of apoptosis in mammals and their numerous interactions complicate the analysis of their individual functions. Yeast, eukaryotic but unicellular organism, lack the main regulators of apoptosis (caspases, Bcl-2 family members, ...) found in mammals. This absence render them a powerful tool for heterologous expression, functional studies, and even cloning of new regulators of apoptosis. Great advances have thus been made in our understanding of the molecular mechanisms of Bcl-2 family members interactions with themselves and other cellular proteins, specially thanks to the two hybrid system and the easy manipulation of yeast (molecular biology and genetics). This review will focus on the use of yeast as a tool to identify new regulators and study function of mammalian apoptosis regulators.

Ectopic expression of Capsicum-specific cell wall protein Capsicum annuum senescence-delaying 1 (CaSD1) delays senescence and induces trichome formation in Nicotiana benthamiana.

PubMed

Seo, Eunyoung; Yeom, Seon-In; Jo, Sunghwan; Jeong, Heejin; Kang, Byoung-Cheorl; Choi, Doil

2012-04-01

Secreted proteins are known to have multiple roles in plant development, metabolism, and stress response. In a previous study to understand the roles of secreted proteins, Capsicum annuum secreted proteins (CaS) were isolated by yeast secretion trap. Among the secreted proteins, we further characterized Capsicum annuum senescence-delaying 1 (CaSD1), a gene encoding a novel secreted protein that is present only in the genus Capsicum. The deduced CaSD1 contains multiple repeats of the amino acid sequence KPPIHNHKPTDYDRS. Interestingly, the number of repeats varied among cultivars and species in the Capsicum genus. CaSD1 is constitutively expressed in roots, and Agrobacterium-mediated transient overexpression of CaSD1 in Nicotiana benthamiana leaves resulted in delayed senescence with a dramatically increased number of trichomes and enlarged epidermal cells. Furthermore, senescence- and cell division-related genes were differentially regulated by CaSD1-overexpressing plants. These observations imply that the pepper-specific cell wall protein CaSD1 plays roles in plant growth and development by regulating cell division and differentiation.

Germ cell control of testin production is inverse to that of other Sertoli cell products.

PubMed

Jégou, B; Pineau, C; Velez de la Calle, J F; Touzalin, A M; Bardin, C W; Cheng, C Y

1993-06-01

Recent studies have shown that germ cells can regulate testins, two newly identified Sertoli cell proteins that are associated with junctional complexes. To investigate this possibility, several parameters of Sertoli cell function were investigated over 2-120 days post exposure of the rat testes to x-rays (3 Grays). The irradiation-induced loss of spermatogonia resulted in a maturation-depletion process progressively affecting all germ cell classes. Testis weight began to decrease when the most numerous germ cell type (spermatids) began to decline. A complete or near complete recovery of spermatogenesis and of the testis weight had occurred by day 120 post irradiation. There was no significant change in FSH, epididymal androgen-binding protein, and tubule fluid levels during the first weeks after irradiation, when the seminiferious epithelium was depleted of spermatogonia and germ cells up to early spermatids. In contrast, when the number of the more mature forms of spermatids declined (between day 21 and 54), FSH rose and androgen-binding protein as well as fluid production declined. The subsequent recovery of these parameters was also highly correlated with the number of late spermatids. By contrast, testicular testin contents reacted to the depletion of germ cells with a biphasic increase; a doubling occurred when spermatogonia, spermatocytes, and early spermatids were absent (days 4-28), and a 7-fold rise occurred by day 37 when the number of late spermatids had decreased by 50%. By day 54, when the sperm counts had reached a nadir, testin contents had returned to levels corresponding to about four times the control levels; they progressively recovered thereafter. These observations support the postulate that germ cells negatively regulate testins. This possibility was investigated with in vitro experiments showing that addition of germ cell-conditioned medium to Sertoli cell monolayers inhibited testin secretion in a dose-dependent manner. In conclusion this study; 1) highlights the complex interplay between the various germ cell classes in the control of the Sertoli cell function in the adult testis; 2) establishes that germ cell effects may be opposite on different Sertoli cell products; 3) demonstrates that several classes of germ cells negatively control testicular testin contents; and 4) emphasizes the particular role of late spermatids in Sertoli cell regulation.

Nicotine affects rat Leydig cell function in vivo and vitro via down-regulating some key steroidogenic enzyme expressions.

PubMed

Guo, Xiaoling; Wang, Huang; Wu, Xiaolong; Chen, Xianwu; Chen, Yong; Guo, Jingjing; Li, Xiaoheng; Lian, Qingquan; Ge, Ren-Shan

2017-12-01

Nicotine is consumed largely as a component of cigarettes and has a potential effect on pubertal development of Leydig cells in males. To investigate its effects, 49-day-old male Sprague Dawley rats received intraperitoneal injections of nicotine (0.5 or 1 mg/kg/day) for 2 weeks and immature Leydig cells were isolated from the testes of 35-day-old rats and treated with nicotine (0.05-50 μM). Serum hormones, Leydig cell number and related gene expression levels after in vivo treatment were determined and medium androgen levels were measured and cell cycle, apoptosis, mitochondrial membrane potential (△Ψm), and reactive oxygen species (ROS) of Leydig cells after in vitro treatment were measured. In vivo exposure to nicotine lowered serum luteinizing hormone, follicle stimulating hormone, and testosterone levels and reduced Leydig cell number and gene expression levels. Nicotine in vitro inhibited androgen production in Leydig cells by downregulating the expression levels of P450 cholesterol side cleavage enzyme, 3β-hydroxysteroid dehydrogenase 1, and steroidogenic factor 1 at different concentration ranges. In conclusion, nicotine disrupts Leydig cell steroidogenesis during puberty possibly via down-regulating some key steroidogenic enzyme expressions. Copyright © 2017. Published by Elsevier Ltd.

Integrin traffic - the update.

PubMed

De Franceschi, Nicola; Hamidi, Hellyeh; Alanko, Jonna; Sahgal, Pranshu; Ivaska, Johanna

2015-03-01

Integrins are a family of transmembrane cell surface molecules that constitute the principal adhesion receptors for the extracellular matrix (ECM) and are indispensable for the existence of multicellular organisms. In vertebrates, 24 different integrin heterodimers exist with differing substrate specificity and tissue expression. Integrin-extracellular-ligand interaction provides a physical anchor for the cell and triggers a vast array of intracellular signalling events that determine cell fate. Dynamic remodelling of adhesions, through rapid endocytic and exocytic trafficking of integrin receptors, is an important mechanism employed by cells to regulate integrin-ECM interactions, and thus cellular signalling, during processes such as cell migration, invasion and cytokinesis. The initial concept of integrin traffic as a means to translocate adhesion receptors within the cell has now been expanded with the growing appreciation that traffic is intimately linked to the cell signalling apparatus. Furthermore, endosomal pathways are emerging as crucial regulators of integrin stability and expression in cells. Thus, integrin traffic is relevant in a number of pathological conditions, especially in cancer. Nearly a decade ago we wrote a Commentary in Journal of Cell Science entitled 'Integrin traffic'. With the advances in the field, we felt it would be appropriate to provide the growing number of researchers interested in integrin traffic with an update. © 2015. Published by The Company of Biologists Ltd.

An increase or a decrease in myosin II phosphorylation inhibits macrophage motility

PubMed Central

1991-01-01

Myosin II purified from mammalian non-muscle cells is phosphorylated on the 20-kD light chain subunit (MLC20) by the Ca2+/calmodulin-dependent enzyme myosin light chain kinase (MLCK). The importance of MLC20 phosphorylation in regulating cell motility was investigated by introducing either antibodies to MLCK (MK-Ab) or a Ca2+/calmodulin- independent, constitutively active form of MLCK (MK-) into macrophages. The effects of these proteins on cell motility were then determined using a quantitative chemotaxis assay. Chemotaxis is significantly diminished in macrophages containing MK-Ab compared to macrophages containing control antibodies. Moreover, there is an inverse relationship between the number of cells that migrate and the amount of MK-Ab introduced into cells. Interestingly, there is also an inverse relationship between the number of cells that migrate and the amount of MK- introduced into cells. Other experiments demonstrated that MK-Ab decreased intracellular MLC20 phosphorylation while MK- increased MLC20 phosphorylation. MK- also increased the amount of myosin associated with the cytoskeleton. These data demonstrate that the regulation of MLCK is an important aspect of cell motility and suggest that MLC20 phosphorylation must be maintained within narrow limits during translational motility by mammalian cells. PMID:2071674

A practical guide for the identification of membrane and plasma membrane proteins in human embryonic stem cells and human embryonal carcinoma cells.

PubMed

Dormeyer, Wilma; van Hoof, Dennis; Mummery, Christine L; Krijgsveld, Jeroen; Heck, Albert J R

2008-10-01

The identification of (plasma) membrane proteins in cells can provide valuable insights into the regulation of their biological processes. Pluripotent cells such as human embryonic stem cells and embryonal carcinoma cells are capable of unlimited self-renewal and share many of the biological mechanisms that regulate proliferation and differentiation. The comparison of their membrane proteomes will help unravel the biological principles of pluripotency, and the identification of biomarker proteins in their plasma membranes is considered a crucial step to fully exploit pluripotent cells for therapeutic purposes. For these tasks, membrane proteomics is the method of choice, but as indicated by the scarce identification of membrane and plasma membrane proteins in global proteomic surveys it is not an easy task. In this minireview, we first describe the general challenges of membrane proteomics. We then review current sample preparation steps and discuss protocols that we found particularly beneficial for the identification of large numbers of (plasma) membrane proteins in human tumour- and embryo-derived stem cells. Our optimized assembled protocol led to the identification of a large number of membrane proteins. However, as the composition of cells and membranes is highly variable we still recommend adapting the sample preparation protocol for each individual system.

SEMI-ROLLED LEAF1 Encodes a Putative Glycosylphosphatidylinositol-Anchored Protein and Modulates Rice Leaf Rolling by Regulating the Formation of Bulliform Cells1[W][OA

PubMed Central

Xiang, Jing-Jing; Zhang, Guang-Heng; Qian, Qian; Xue, Hong-Wei

2012-01-01

Leaf rolling is an important agronomic trait in rice (Oryza sativa) breeding and moderate leaf rolling maintains the erectness of leaves and minimizes shadowing between leaves, leading to improved photosynthetic efficiency and grain yields. Although a few rolled-leaf mutants have been identified and some genes controlling leaf rolling have been isolated, the molecular mechanisms of leaf rolling still need to be elucidated. Here we report the isolation and characterization of SEMI-ROLLED LEAF1 (SRL1), a gene involved in the regulation of leaf rolling. Mutants srl1-1 (point mutation) and srl1-2 (transferred DNA insertion) exhibit adaxially rolled leaves due to the increased numbers of bulliform cells at the adaxial cell layers, which could be rescued by complementary expression of SRL1. SRL1 is expressed in various tissues and is expressed at low levels in bulliform cells. SRL1 protein is located at the plasma membrane and predicted to be a putative glycosylphosphatidylinositol-anchored protein. Moreover, analysis of the gene expression profile of cells that will become epidermal cells in wild type but probably bulliform cells in srl1-1 by laser-captured microdissection revealed that the expression of genes encoding vacuolar H+-ATPase (subunits A, B, C, and D) and H+-pyrophosphatase, which are increased during the formation of bulliform cells, were up-regulated in srl1-1. These results provide the transcript profile of rice leaf cells that will become bulliform cells and demonstrate that SRL1 regulates leaf rolling through inhibiting the formation of bulliform cells by negatively regulating the expression of genes encoding vacuolar H+-ATPase subunits and H+-pyrophosphatase, which will help to understand the mechanism regulating leaf rolling. PMID:22715111

Temporal Control of Plant Organ Growth by TCP Transcription Factors.

PubMed

Huang, Tengbo; Irish, Vivian F

2015-06-29

The Arabidopsis petal is a simple laminar organ whose development is largely impervious to environmental effects, making it an excellent model for dissecting the regulation of cell-cycle progression and post-mitotic cell expansion that together sculpt organ form. Arabidopsis petals grow via basipetal waves of cell division, followed by a phase of cell expansion. RABBIT EARS (RBE) encodes a C2H2 zinc finger transcriptional repressor and is required for petal development. During the early phase of petal initiation, RBE regulates a microRNA164-dependent pathway that controls cell proliferation at the petal primordium boundaries. The effects of rbe mutations on petal lamina growth suggest that RBE is also required to regulate later developmental events during petal organogenesis. Here, we demonstrate that, early in petal development, RBE represses the transcription of a suite of CIN-TCP genes that in turn act to inhibit the number and duration of cell divisions; the temporal alleviation of that repression results in the transition from cell division to post-mitotic cell expansion and concomitant petal maturation. Copyright © 2015 Elsevier Ltd. All rights reserved.

Telomere Dynamics and Homeostasis in a Transmissible Cancer

PubMed Central

Ujvari, Beata; Pearse, Anne-Maree; Taylor, Robyn; Pyecroft, Stephen; Flanagan, Cassandra; Gombert, Sara; Papenfuss, Anthony T.; Madsen, Thomas; Belov, Katherine

2012-01-01

Background Devil Facial Tumour Disease (DFTD) is a unique clonal cancer that threatens the world's largest carnivorous marsupial, the Tasmanian devil (Sarcophilus harrisii) with extinction. This transmissible cancer is passed between individual devils by cell implantation during social interactions. The tumour arose in a Schwann cell of a single devil over 15 years ago and since then has expanded clonally, without showing signs of replicative senescence; in stark contrast to a somatic cell that displays a finite capacity for replication, known as the “Hayflick limit”. Methodology/Principal Findings In the present study we investigate the role of telomere length, measured as Telomere Copy Number (TCN), and telomerase and shelterin gene expression, as well as telomerase activity in maintaining hyperproliferation of Devil Facial Tumour (DFT) cells. Our results show that DFT cells have short telomeres. DFTD TCN does not differ between geographic regions or between strains. However, TCN has increased over time. Unlimited cell proliferation is likely to have been achieved through the observed up-regulation of the catalytic subunit of telomerase (TERT) and concomitant activation of telomerase. Up-regulation of the central component of shelterin, the TRF1-intercating nuclear factor 2 (TINF2) provides DFT a mechanism for telomere length homeostasis. The higher expression of both TERT and TINF2 may also protect DFT cells from genomic instability and enhance tumour proliferation. Conclusions/Significance DFT cells appear to monitor and regulate the length of individual telomeres: i.e. shorter telomeres are elongated by up-regulation of telomerase-related genes; longer telomeres are protected from further elongation by members of the shelterin complex, which may explain the lack of spatial and strain variation in DFT telomere copy number. The observed longitudinal increase in gene expression in DFT tissue samples and telomerase activity in DFT cell lines might indicate a selection for more stable tumours with higher proliferative potential. PMID:22952882

microRNA-150 Regulates Mobilization and Migration of Bone Marrow-Derived Mononuclear Cells by Targeting Cxcr4

PubMed Central

Tano, Nobuko; Kim, Ha Won; Ashraf, Muhammad

2011-01-01

The interaction between chemokine receptor type 4 (CXCR4) and its ligand, stromal cell-derived factor (SDF)-1, plays an important role in stem cell mobilization and migration in ischemic tissues. MicroRNAs (miRs) are key regulators of stem cell function and are involved in regulation of stem cell survival and differentiation to adopt different cell lineages. In this study, we show that ischemia inhibits the expression of miR-150 in BM-derived mononuclear cells (MNC) and activates its target Cxcr4 gene. Our results show that miR-150/CXCR4 cascade enhances MNC mobilization and migration. By using mouse acute myocardial infarction (MI) model, we found that MNCs in peripheral blood (PB) were increased significantly at day 5 after AMI as compared to control group and the number of CXCR4 positive MNCs both in bone marrow (BM) and PB was also markedly increased after MI. Analysis by microarray-based miRNA profiling and real-time PCR revealed that the expression of miR-150 which targets Cxcr4 gene as predicted was significantly downregulated in BM-MNCs after MI. Abrogation of miR-150 markedly increased CXCR4 protein expression suggesting its target gene. To show that miR-150 regulates MNC mobilization, knockdown of miR-150 in BM-MNCs by specific antisense inhibitor resulted in their higher migration ability in vitro as compared to scramble-transfected MNCs. Furthermore, in vivo BM transplantation of MNCs lacking miR-150 expression by lentiviral vector into the irradiated wild type mice resulted in the increased number of MNCs in PB after AMI as compared to control. In conclusion, this study demonstrates that ischemia mobilizes BM stem cells via miR-150/CXCR4 dependent mechanism and miR-150 may be a novel therapeutic target for stem cell migration to the ischemic tissue for neovascularization and repair. PMID:22039399

Managing the complexity of communication: regulation of gap junctions by post-translational modification

PubMed Central

Axelsen, Lene N.; Calloe, Kirstine; Holstein-Rathlou, Niels-Henrik; Nielsen, Morten S.

2013-01-01

Gap junctions are comprised of connexins that form cell-to-cell channels which couple neighboring cells to accommodate the exchange of information. The need for communication does, however, change over time and therefore must be tightly controlled. Although the regulation of connexin protein expression by transcription and translation is of great importance, the trafficking, channel activity and degradation are also under tight control. The function of connexins can be regulated by several post translational modifications, which affect numerous parameters; including number of channels, open probability, single channel conductance or selectivity. The most extensively investigated post translational modifications are phosphorylations, which have been documented in all mammalian connexins. Besides phosphorylations, some connexins are known to be ubiquitinated, SUMOylated, nitrosylated, hydroxylated, acetylated, methylated, and γ-carboxyglutamated. The aim of the present review is to summarize our current knowledge of post translational regulation of the connexin family of proteins. PMID:24155720

Functional Analysis of the Drosophila Embryonic Germ Cell Transcriptome by RNA Interference

PubMed Central

Bujna, Ágnes; Vilmos, Péter; Spirohn, Kerstin; Boutros, Michael; Erdélyi, Miklós

2014-01-01

In Drosophila melanogaster, primordial germ cells are specified at the posterior pole of the very early embryo. This process is regulated by the posterior localized germ plasm that contains a large number of RNAs of maternal origin. Transcription in the primordial germ cells is actively down-regulated until germ cell fate is established. Bulk expression of the zygotic genes commences concomitantly with the degradation of the maternal transcripts. Thus, during embryogenesis, maternally provided and zygotically transcribed mRNAs determine germ cell development collectively. In an effort to identify novel genes involved in the regulation of germ cell behavior, we carried out a large-scale RNAi screen targeting both maternal and zygotic components of the embryonic germ line transcriptome. We identified 48 genes necessary for distinct stages in germ cell development. We found pebble and fascetto to be essential for germ cell migration and germ cell division, respectively. Our data uncover a previously unanticipated role of mei-P26 in maintenance of embryonic germ cell fate. We also performed systematic co-RNAi experiments, through which we found a low rate of functional redundancy among homologous gene pairs. As our data indicate a high degree of evolutionary conservation in genetic regulation of germ cell development, they are likely to provide valuable insights into the biology of the germ line in general. PMID:24896584

Adrenocortical Gap Junctions and Their Functions

PubMed Central

Bell, Cheryl L.; Murray, Sandra A.

2016-01-01

Adrenal cortical steroidogenesis and proliferation are thought to be modulated by gap junction-mediated direct cell–cell communication of regulatory molecules between cells. Such communication is regulated by the number of gap junction channels between contacting cells, the rate at which information flows between these channels, and the rate of channel turnover. Knowledge of the factors regulating gap junction-mediated communication and the turnover process are critical to an understanding of adrenal cortical cell functions, including development, hormonal response to adrenocorticotropin, and neoplastic dedifferentiation. Here, we review what is known about gap junctions in the adrenal gland, with particular attention to their role in adrenocortical cell steroidogenesis and proliferation. Information and insight gained from electrophysiological, molecular biological, and imaging (immunocytochemical, freeze fracture, transmission electron microscopic, and live cell) techniques will be provided. PMID:27445985

The interrelationship between bone and fat: from cellular see-saw to endocrine reciprocity.

PubMed

Sadie-Van Gijsen, H; Crowther, N J; Hough, F S; Ferris, W F

2013-07-01

The number of mature osteoblasts and marrow adipocytes in bone is influenced by the differentiation of the common mesenchymal progenitor cell towards one phenotype and away from the other. Consequently, factors which promote adipogenesis not only lead to fatty marrow but also inhibit osteoblastogenesis, resulting in decreased osteoblast numbers, diminished bone formation and, potentially, inadequate bone mass and osteoporosis. In addition to osteoblast and bone adipocyte numbers being influenced by this skewing of progenitor cell differentiation towards one phenotype, mature osteoblasts and adipocytes secrete factors which may evoke changes in the cell fate and function of each other. This review examines the endogenous factors, such as PPAR-γ2, Wnt, IGF-1, GH, FGF-2, oestrogen, the GP130 signalling cytokines, vitamin D and glucocorticoids, which regulate the selection between osteoblastogenesis and adipogenesis and the interrelationship between fat and bone. The role of adipokines on bone, such as adiponectin and leptin, as well as adipose-derived oestrogen, is reviewed and the role of bone as an energy regulating endocrine organ is discussed.

Depletion of nuclear import protein karyopherin alpha 7 (KPNA7) induces mitotic defects and deformation of nuclei in cancer cells.

PubMed

Vuorinen, Elisa M; Rajala, Nina K; Ihalainen, Teemu O; Kallioniemi, Anne

2018-03-27

Nucleocytoplasmic transport is a tightly regulated process carried out by specific transport machinery, the defects of which may lead to a number of diseases including cancer. Karyopherin alpha 7 (KPNA7), the newest member of the karyopherin alpha nuclear importer family, is expressed at a high level during embryogenesis, reduced to very low or absent levels in most adult tissues but re-expressed in cancer cells. We used siRNA-based knock-down of KPNA7 in cancer cell lines, followed by functional assays (proliferation and cell cycle) and immunofluorescent stainings to determine the role of KPNA7 in regulation of cancer cell growth, proper mitosis and nuclear morphology. In the present study, we show that the silencing of KPNA7 results in a dramatic reduction in pancreatic and breast cancer cell growth, irrespective of the endogenous KPNA7 expression level. This growth inhibition is accompanied by a decrease in the fraction of S-phase cells as well as aberrant number of centrosomes and severe distortion of the mitotic spindles. In addition, KPNA7 depletion leads to reorganization of lamin A/C and B1, the main nuclear lamina proteins, and drastic alterations in nuclear morphology with lobulated and elongated nuclei. Taken together, our data provide new important evidence on the contribution of KPNA7 to the regulation of cancer cell growth and the maintenance of nuclear envelope environment, and thus deepens our understanding on the impact of nuclear transfer proteins in cancer pathogenesis.

Dehydrated human amnion/chorion membrane regulates stem cell activity in vitro

PubMed Central

Massee, Michelle; Chinn, Kathryn; Lei, Jennifer; Lim, Jeremy J.; Young, Conan S.

2015-01-01

Abstract Human‐derived placental tissues have been shown in randomized clinical trials to be effective for healing chronic wounds, and have also demonstrated the ability to recruit stem cells to the wound site in vitro and in vivo. In this study, PURION® Processed dehydrated human amnion/chorion membrane allografts (dHACM, EpiFix®, MiMedx Group, Marietta, GA) were evaluated for their ability to alter stem cell activity in vitro. Human bone marrow mesenchymal stem cells (BM‐MSCs), adipose derived stem cells (ADSCs), and hematopoietic stem cells (HSCs) were treated with soluble extracts of dHACM tissue, and were evaluated for cellular proliferation, migration, and cytokine secretion. Stem cells were analyzed for cell number by DNA assay after 24 h, closure of an acellular zone using microscopy over 3 days, and soluble cytokine production in the medium of treated stem cells was analyzed after 3 days using a multiplex ELISA array. Treatment with soluble extracts of dHACM tissue stimulated BM‐MSCs, ADSCs, and HSCs to proliferate with a significant increase in cell number after 24 h. dHACM treatment accelerated closure of an acellular zone by ADSCs and BM‐MSCs after 3 days, compared to basal medium. BM‐MSCs, ADSCs, and HSCs also modulated endogenous production of a number of various soluble signals, including regulators of inflammation, mitogenesis, and wound healing. dHACM treatment promoted increased proliferation and migration of ADSCs, BM‐MSCs, and HSCs, along with modulation of secreted proteins from those cells. Therefore, dHACM may impact wound healing by amplifying host stem cell populations and modulating their responses in treated wound tissues. © 2015 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 104B: 1495–1503, 2016. PMID:26175122

Benzo(a)pyrene quinones increase cell proliferation, generate reactive oxygen species, and transactivate the epidermal growth factor receptor in breast epithelial cells.

PubMed

Burdick, Andrew D; Davis, John W; Liu, Ke Jian; Hudson, Laurie G; Shi, Honglian; Monske, Michael L; Burchiel, Scott W

2003-11-15

Polycyclic aromatic hydrocarbons, such as benzo(a)pyrene (BaP), are known mammary carcinogens in rodents and may be involved in human breast cancer. We have reported previously that BaP can mimic growth factor signaling and increase cell proliferation in primary human mammary epithelial cells and the human mammary epithelial cell line MCF-10A. BaP-quinones (BPQs) are important metabolites of BaP that have been associated with the production of reactive oxygen species. Using a model of epidermal growth factor (EGF) withdrawal in MCF-10A, we hypothesized that production of reactive oxygen species by BPQs could lead to the activation of the EGF receptor (EGFR). Here, we demonstrate through electron paramagnetic resonance spectroscopy and flow cytometry that 1,6-BPQ and 3,6-BPQ produce superoxide anion and hydrogen peroxide in MCF-10A cells. Furthermore, we show that BPQs increase EGFR, Akt, and extracellular signal-regulated kinase activity, leading to increased cell number in the absence of EGF. The BPQ-induced EGFR activity and associated cell proliferation were attenuated by the EGFR inhibitor AG1478, as well as by the antioxidant N-acetyl cysteine. Overexpression of catalase, but not Cu/Zn superoxide dismutase, reduced the extent of BPQ-dependent increased cell number and EGFR pathway activation. Moreover, the direct treatment of MCF-10A cells with hydrogen peroxide enhanced EGFR, Akt, and extracellular-regulated kinase phosphorylation that could be similarly inhibited by AG1478, N-acetyl cysteine, and catalase. Taken together, these data indicate that BPQs, through the generation of hydrogen peroxide, activate the EGFR in MCF-10A cells, leading to increased cell number under EGF-deficient conditions.

Sarcoptes scabiei Mites Modulate Gene Expression in Human Skin Equivalents

PubMed Central

Morgan, Marjorie S.; Arlian, Larry G.; Markey, Michael P.

2013-01-01

The ectoparasitic mite, Sarcoptes scabiei that burrows in the epidermis of mammalian skin has a long co-evolution with its hosts. Phenotypic studies show that the mites have the ability to modulate cytokine secretion and expression of cell adhesion molecules in cells of the skin and other cells of the innate and adaptive immune systems that may assist the mites to survive in the skin. The purpose of this study was to identify genes in keratinocytes and fibroblasts in human skin equivalents (HSEs) that changed expression in response to the burrowing of live scabies mites. Overall, of the more than 25,800 genes measured, 189 genes were up-regulated >2-fold in response to scabies mite burrowing while 152 genes were down-regulated to the same degree. HSEs differentially expressed large numbers of genes that were related to host protective responses including those involved in immune response, defense response, cytokine activity, taxis, response to other organisms, and cell adhesion. Genes for the expression of interleukin-1α (IL-1α) precursor, IL-1β, granulocyte/macrophage-colony stimulating factor (GM-CSF) precursor, and G-CSF precursor were up-regulated 2.8- to 7.4-fold, paralleling cytokine secretion profiles. A large number of genes involved in epithelium development and keratinization were also differentially expressed in response to live scabies mites. Thus, these skin cells are directly responding as expected in an inflammatory response to products of the mites and the disruption of the skin’s protective barrier caused by burrowing. This suggests that in vivo the interplay among these skin cells and other cell types, including Langerhans cells, dendritic cells, lymphocytes and endothelial cells, is responsible for depressing the host’s protective response allowing these mites to survive in the skin. PMID:23940705

Pelle Modulates dFoxO-Mediated Cell Death in Drosophila.

PubMed

Wu, Chenxi; Chen, Yujun; Wang, Feng; Chen, Changyan; Zhang, Shiping; Li, Chaojie; Li, Wenzhe; Wu, Shian; Xue, Lei

2015-10-01

Interleukin-1 receptor-associated kinases (IRAKs) are crucial mediators of the IL-1R/TLR signaling pathways that regulate the immune and inflammation response in mammals. Recent studies also suggest a critical role of IRAKs in tumor development, though the underlying mechanism remains elusive. Pelle is the sole Drosophila IRAK homolog implicated in the conserved Toll pathway that regulates Dorsal/Ventral patterning, innate immune response, muscle development and axon guidance. Here we report a novel function of pll in modulating apoptotic cell death, which is independent of the Toll pathway. We found that loss of pll results in reduced size in wing tissue, which is caused by a reduction in cell number but not cell size. Depletion of pll up-regulates the transcription of pro-apoptotic genes, and triggers caspase activation and cell death. The transcription factor dFoxO is required for loss-of-pll induced cell death. Furthermore, loss of pll activates dFoxO, promotes its translocation from cytoplasm to nucleus, and up-regulates the transcription of its target gene Thor/4E-BP. Finally, Pll physically interacts with dFoxO and phosphorylates dFoxO directly. This study not only identifies a previously unknown physiological function of pll in cell death, but also shed light on the mechanism of IRAKs in cell survival/death during tumorigenesis.

Repression of cell proliferation by miR319-regulated TCP4.

PubMed

Schommer, Carla; Debernardi, Juan M; Bresso, Edgardo G; Rodriguez, Ramiro E; Palatnik, Javier F

2014-10-01

Leaf development has been extensively studied on a genetic level. However, little is known about the interplay between the developmental regulators and the cell cycle machinery--a link that ultimately affects leaf form and size. miR319 is a conserved microRNA that regulates TCP transcription factors involved in multiple developmental pathways, including leaf development and senescence, organ curvature, and hormone biosynthesis and signaling. Here, we analyze the participation of TCP4 in the control of cell proliferation. A small increase in TCP4 activity has an immediate impact on leaf cell number, by significantly reducing cell proliferation. Plants with high TCP4 levels have a strong reduction in the expression of genes known to be active in G2-M phase of the cell cycle. Part of these effects is mediated by induction of miR396, which represses Growth-Regulating Factor (GRF) transcription factors. Detailed analysis revealed TCP4 to be a direct regulator of MIR396b. However, we found that TCP4 can control cell proliferation through additional pathways, and we identified a direct connection between TCP4 and ICK1/KRP1, a gene involved in the progression of the cell cycle. Our results show that TCP4 can activate different pathways that repress cell proliferation. © The Author 2014. Published by the Molecular Plant Shanghai Editorial Office in association with Oxford University Press on behalf of CSPB and IPPE, SIBS, CAS.

Temporal regulation of Stat5 activity in determination of cell differentiation program

PubMed Central

Hoshino, Akemi; Fujii, Hodaka

2007-01-01

Although Stat5 is activated by various cytokines, only ethrytopoietin (Epo) and a small number of cytokines induce Stat5-dependent erythroid differentiation. Here, by using a reporter gene system to monitor transcriptional activity of Stat5, we showed that Epo but not interleukin (IL)-3 supports sustained activation of Stat5, which induces globin gene expression. IL-3 or IL-2 stimulation inhibits Epo-induced globin gene expression. The acidic region of the IL-2 receptor β chain was essential for this inhibition. These results underscore the importance of temporal regulation of Stat activity for regulation of cytokine-specific cell differentiation. PMID:17511959

Fascin regulates nuclear actin during Drosophila oogenesis

PubMed Central

Kelpsch, Daniel J.; Groen, Christopher M.; Fagan, Tiffany N.; Sudhir, Sweta; Tootle, Tina L.

2016-01-01

Drosophila oogenesis provides a developmental system with which to study nuclear actin. During Stages 5–9, nuclear actin levels are high in the oocyte and exhibit variation within the nurse cells. Cofilin and Profilin, which regulate the nuclear import and export of actin, also localize to the nuclei. Expression of GFP-tagged Actin results in nuclear actin rod formation. These findings indicate that nuclear actin must be tightly regulated during oogenesis. One factor mediating this regulation is Fascin. Overexpression of Fascin enhances nuclear GFP-Actin rod formation, and Fascin colocalizes with the rods. Loss of Fascin reduces, whereas overexpression of Fascin increases, the frequency of nurse cells with high levels of nuclear actin, but neither alters the overall nuclear level of actin within the ovary. These data suggest that Fascin regulates the ability of specific cells to accumulate nuclear actin. Evidence indicates that Fascin positively regulates nuclear actin through Cofilin. Loss of Fascin results in decreased nuclear Cofilin. In addition, Fascin and Cofilin genetically interact, as double heterozygotes exhibit a reduction in the number of nurse cells with high nuclear actin levels. These findings are likely applicable beyond Drosophila follicle development, as the localization and functions of Fascin and the mechanisms regulating nuclear actin are widely conserved. PMID:27535426

Interferon regulatory factor 4 (IRF4) controls myeloid-derived suppressor cell (MDSC) differentiation and function.

PubMed

Nam, Sorim; Kang, Kyeongah; Cha, Jae Seon; Kim, Jung Woo; Lee, Hee Gu; Kim, Yonghwan; Yang, Young; Lee, Myeong-Sok; Lim, Jong-Seok

2016-12-01

Myeloid-derived suppressor cells (MDSCs) are immature cells that do not differentiate into mature myeloid cells. Two major populations of PMN-MDSCs (Ly6G high Ly6C low Gr1 high CD11b + ) and MO-MDSCs (Ly6G - Ly6C high Gr-1 int CD11b + ) have an immune suppressive function. Interferon regulatory factor 4 (IRF4) has a role in the negative regulation of TLR signaling and is associated with lymphoid cell development. However, the roles of IRF4 in myeloid cell differentiation are unclear. In this study, we found that IRF4 expression was remarkably suppressed during the development of MDSCs in the tumor microenvironment. Both the mRNA and protein levels of IRF4 in MDSCs were gradually reduced, depending on the development of tumors in the 4T1 model. siRNA-mediated knockdown of IRF4 in bone marrow cells promoted the differentiation of PMN-MDSCs. Similarly, IRF4 inhibition in bone marrow cells using simvastatin, which has been known to inhibit IRF4 expression, increased PMN-MDSC numbers. In contrast, IRF4 overexpression in bone marrow cells inhibited the total numbers of MDSCs, especially PMN-MDSCs. Notably, treatment with IL-4, an upstream regulator of IRF4, induced IRF4 expression in the bone marrow cells, and consequently, IL-4-induced IRF4 expression resulted in a decrease in PMN-MDSC numbers. Finally, we confirmed that IRF4 expression in MDSCs can modulate their activity to inhibit T cell proliferation through IL-10 production and ROS generation, and myeloid-specific deletion of IRF4 leads to the increase of MDSC differentiation. Our present findings indicate that IRF4 reduction induced by tumor formation can increase the number of MDSCs, and increases in the IRF4 expression in MDSCs may infringe on the immune-suppressive function of MDSCs. © Society for Leukocyte Biology.

Transcriptional and Functional Plasticity Induced by Chronic Insulin Exposure in a Mast Cell-Like Basophilic Leukemia Cell Model

PubMed Central

Jansen, Chad; Speck, Mark; Greineisen, William E; Maaetoft-Udsen, Kristina; Cordasco, Edward; Shimoda, Lori MN; Stokes, Alexander J; Turner, Helen

2018-01-01

Objective Secretory granules (SG) and lipid bodies (LB) are the primary organelles that mediate functional responses in mast cells. SG contains histamine and matrix-active proteases, while LB are reservoirs of arachidonic acid and its metabolites, precursors for rapid synthesis of eicosanoids such as LTC4. Both of these compartments can be dynamically or ontologically regulated, with metabolic and immunological stimuli altering lipid body content and granule numbers responding to contextual signals from tissue. We previously described that chronic in vitro or in vivo hyperinsulinemia expands the LB compartment with a concomitant loss of SG capacity, suggesting that this ratio is dynamically regulated. The objective of the current study is to determine if chronic insulin exposure initiates a transcriptional program that biases model mast cells towards a lipogenic state with accompanying loss of secretory granule biogenesis. Methods We used a basophilic leukemic cell line with mucosal mast cell-like features as a model system. We tested the hypothesis that chronic insulin exposure initiates a transcriptional program that biases these model mast cells towards a lipogenic state with accompanying loss of secretory granule biogenesis. Transcriptional arrays were used to map gene expression patterns. Biochemical, immunocytochemical and mediator release assays were used to evaluate organelle numbers and functional responses. Results In a mucosal mast cell model, the rat basophilic leukemia line RBL2H3, mast cell granularity and SG numbers are inversely correlated with LB numbers. Chronic insulin exposure appears to modulate gene networks involved in both lipid body biogenesis and secretory granule formation. Western blot analysis confirms upregulation of protein levels for LB proteins, and decreases in proteins that are markers for SG cargo. Conclusions The levels of insulin in the extracellular milieu may modify the phenotype of mast cell-like cells in vitro. PMID:29430572

Increasing RpoS expression causes cell death in Borrelia burgdorferi.

PubMed

Chen, Linxu; Xu, Qilong; Tu, Jiagang; Ge, Yihe; Liu, Jun; Liang, Fang Ting

2013-01-01

RpoS, one of the two alternative σ factors in Borrelia burgdorferi, is tightly controlled by multiple regulators and, in turn, determines expression of many critical virulence factors. Here we show that increasing RpoS expression causes cell death. The immediate effect of increasing RpoS expression was to promote bacterial division and as a consequence result in a rapid increase in cell number before causing bacterial death. No DNA fragmentation or degradation was observed during this induced cell death. Cryo-electron microscopy showed induced cells first formed blebs, which were eventually released from dying cells. Apparently blebbing initiated cell disintegration leading to cell death. These findings led us to hypothesize that increasing RpoS expression triggers intracellular programs and/or pathways that cause spirochete death. The potential biological significance of induced cell death may help B. burgdorferi regulate its population to maintain its life cycle in nature.

Functions of TAM RTKs in regulating spermatogenesis and male fertility in mice.

PubMed

Chen, Yongmei; Wang, Huizhen; Qi, Nan; Wu, Hui; Xiong, Weipeng; Ma, Jing; Lu, Qingxian; Han, Daishu

2009-10-01

Mice lacking TYRO3, AXL and MER (TAM) receptor tyrosine kinases (RTKs) are male sterile. The mechanism of TAM RTKs in regulating male fertility remains unknown. In this study, we analyzed in more detail the testicular phenotype of TAM triple mutant (TAM(-/-)) mice with an effort to understand the mechanism. We demonstrate that the three TAM RTKs cooperatively regulate male fertility, and MER appears to be more important than AXL and TYRO3. TAM(-/-) testes showed a progressive loss of germ cells from elongated spermatids to spermatogonia. Young adult TAM(-/-) mice exhibited oligo-astheno-teratozoospermia and various morphological malformations of sperm cells. As the mice aged, the germ cells were eventually depleted from the seminiferous tubules. Furthermore, we found that TAM(-/-) Sertoli cells have an impaired phagocytic activity and a large number of differentially expressed genes compared to wild-type controls. By contrast, the function of Leydig cells was not apparently affected by the mutation of TAM RTKs. Therefore, we conclude that the suboptimal function of Sertoli cells leads to the impaired spermatogenesis in TAM(-/-) mice. The results provide novel insight into the mechanism of TAM RTKs in regulating male fertility.

IL-33 activates eosinophils of visceral adipose tissue both directly and via innate lymphoid cells.

PubMed

Hashiguchi, Masaaki; Kashiwakura, Yuji; Kojima, Hidefumi; Kobayashi, Ayano; Kanno, Yumiko; Kobata, Tetsuji

2015-03-01

Eosinophils are multifunctional leukocytes involved in allergic reactions as well as adipose tissue regulation. IL-5 is required for eosinophil survival; however, the in vivo mechanisms of eosinophil regulation are not fully understood. A tg mouse model with il5 promoter-driven EGFP expression was established for detecting the IL-5-producing cells in vivo. Il5-egfp tg mice expressed high levels of EGFP in gonadal adipose tissue (GAT) cells. EGFP(+) cells in GAT were mainly group 2 innate lymphoid cells (ILCs). IL-33 preferentially expanded EGFP(+) cells and eosinophils in GAT in vivo. EGFP(+) ILCs were found to upregulate prg2 mRNA expression in GAT eosinophils. These results demonstrate that ILCs activate eosinophils in GAT. The blockage of IL-33Rα, on the other hand, did not impair EGFP(+) ILC numbers but did impair eosinophil numbers in vivo. GAT eosinophils expressed IL-33Rα and IL-33 expanded eosinophil numbers in CD90(+) cell-depleted mice. IL-33 was further observed to induce the expression of retnla and epx mRNA in eosinophils. These findings demonstrate that IL-33 directly activates eosinophils in GAT, and together with our other findings described above, our findings show that IL-33 has dual pathways via which it activates eosinophils in vivo: a direct activation pathway and a group 2 ILC-mediated pathway. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

HMGB proteins involved in TOR signaling as general regulators of cell growth by controlling ribosome biogenesis.

PubMed

Vizoso-Vázquez, A; Barreiro-Alonso, A; González-Siso, M I; Rodríguez-Belmonte, E; Lamas-Maceiras, M; Cerdán, M E

2018-04-30

The number of ribosomes and their activity need to be highly regulated because their function is crucial for the cell. Ribosome biogenesis is necessary for cell growth and proliferation in accordance with nutrient availability and other external and intracellular signals. High-mobility group B (HMGB) proteins are conserved from yeasts to human and are decisive in cellular fate. These proteins play critical functions, from the maintenance of chromatin structure, DNA repair, or transcriptional regulation, to facilitation of ribosome biogenesis. They are also involved in cancer and other pathologies. In this review, we summarize evidence of how HMGB proteins contribute to ribosome-biogenesis control, with special emphasis on a common nexus to the target of rapamycin (TOR) pathway, a signaling cascade essential for cell growth and proliferation from yeast to human. Perspectives in this field are also discussed.

AlphaB-crystallin regulates remyelination after peripheral nerve injury

PubMed Central

Lim, Erin-Mai F.; Nakanishi, Stan T.; Hoghooghi, Vahid; Eaton, Shane E. A.; Palmer, Alexandra L.; Frederick, Ariana; Stratton, Jo A.; Stykel, Morgan G.; Zochodne, Douglas W.; Biernaskie, Jeffrey; Ousman, Shalina S.

2017-01-01

AlphaB-crystallin (αBC) is a small heat shock protein that is constitutively expressed by peripheral nervous system (PNS) axons and Schwann cells. To determine what role this crystallin plays after peripheral nerve damage, we found that loss of αBC impaired remyelination, which correlated with a reduced presence of myelinating Schwann cells and increased numbers of nonmyelinating Schwann cells. The heat shock protein also seems to regulate the cross-talk between Schwann cells and axons, because expected changes in neuregulin levels and ErbB2 receptor expression after PNS injury were disrupted in the absence of αBC. Such dysregulations led to defects in conduction velocity and motor and sensory functions that could be rescued with therapeutic application of the heat shock protein in vivo. Altogether, these findings show that αBC plays an important role in regulating Wallerian degeneration and remyelination after PNS injury. PMID:28137843

Cancer stem cells and differentiation therapy.

PubMed

Jin, Xiong; Jin, Xun; Kim, Hyunggee

2017-10-01

Cancer stem cells can generate tumors from only a small number of cells, whereas differentiated cancer cells cannot. The prominent feature of cancer stem cells is its ability to self-renew and differentiate into multiple types of cancer cells. Cancer stem cells have several distinct tumorigenic abilities, including stem cell signal transduction, tumorigenicity, metastasis, and resistance to anticancer drugs, which are regulated by genetic or epigenetic changes. Like normal adult stem cells involved in various developmental processes and tissue homeostasis, cancer stem cells maintain their self-renewal capacity by activating multiple stem cell signaling pathways and inhibiting differentiation signaling pathways during cancer initiation and progression. Recently, many studies have focused on targeting cancer stem cells to eradicate malignancies by regulating stem cell signaling pathways, and products of some of these strategies are in preclinical and clinical trials. In this review, we describe the crucial features of cancer stem cells related to tumor relapse and drug resistance, as well as the new therapeutic strategy to target cancer stem cells named "differentiation therapy."

Activation of GLP-1 Receptor Promotes Bone Marrow Stromal Cell Osteogenic Differentiation through β-Catenin

PubMed Central

Meng, Jingru; Ma, Xue; Wang, Ning; Jia, Min; Bi, Long; Wang, Yunying; Li, Mingkai; Zhang, Huinan; Xue, Xiaoyan; Hou, Zheng; Zhou, Ying; Yu, Zhibin; He, Gonghao; Luo, Xiaoxing

2016-01-01

Summary Glucagon-like peptide 1 (GLP-1) plays an important role in regulating bone remodeling, and GLP-1 receptor agonist shows a positive relationship with osteoblast activity. However, GLP-1 receptor is not found in osteoblast, and the mechanism of GLP-1 receptor agonist on regulating bone remodeling is unclear. Here, we show that the GLP-1 receptor agonist exendin-4 (Ex-4) promoted bone formation and increased bone mass and quality in a rat unloading-induced bone loss model. These functions were accompanied by an increase in osteoblast number and serum bone formation markers, while the adipocyte number was decreased. Furthermore, GLP-1 receptor was detected in bone marrow stromal cells (BMSCs), but not in osteoblast. Activation of GLP-1 receptor by Ex-4 promoted the osteogenic differentiation and inhibited BMSC adipogenic differentiation through regulating PKA/β-catenin and PKA/PI3K/AKT/GSK3β signaling. These findings reveal that GLP-1 receptor regulates BMSC osteogenic differentiation and provide a molecular basis for therapeutic potential of GLP-1 against osteoporosis. PMID:26947974

DOE Office of Scientific and Technical Information (OSTI.GOV)

Buttner, William; Rivkin, Carl; Burgess, Robert

The United Nations Global Technical Regulation (GTR) Number 13 (Global Technical Regulation on Hydrogen and Fuel Cell Vehicles) is the defining document regulating safety requirements in hydrogen vehicles, and in particular fuel cell electric vehicles (FCEV). GTR Number 13 has been formally implemented and will serve as the basis for the national regulatory standards for FCEV safety in North America (Canada, United States), Japan, Korea, and the European Union. The GTR defines safety requirement for these vehicles, including specifications on the allowable hydrogen levels in vehicle enclosures during in-use and post-crash conditions and on the allowable hydrogen emissions levels inmore » vehicle exhaust during certain modes of normal operation. However, in order to be incorporated into national regulations, that is, in order to be binding, methods to verify compliance to the specific requirements must exist. In a collaborative program, the Sensor Laboratories at the National Renewable Energy Laboratory in the United States and the Joint Research Centre, Institute for Energy and Transport in the Netherlands have been evaluating and developing analytical methods that can be used to verify compliance to the hydrogen release requirement as specified in the GTR.« less

Pyruvate dehydrogenase has a major role in mast cell function, and its activity is regulated by mitochondrial microphthalmia transcription factor.

PubMed

Sharkia, Israa; Hadad Erlich, Tal; Landolina, Nadine; Assayag, Miri; Motzik, Alex; Rachmin, Inbal; Kay, Gillian; Porat, Ziv; Tshori, Sagi; Berkman, Neville; Levi-Schaffer, Francesca; Razin, Ehud

2017-07-01

We have recently observed that oxidative phosphorylation-mediated ATP production is essential for mast cell function. Pyruvate dehydrogenase (PDH) is the main regulator of the Krebs cycle and is located upstream of the electron transport chain. However, the role of PDH in mast cell function has not been described. Microphthalmia transcription factor (MITF) regulates the development, number, and function of mast cells. Localization of MITF to the mitochondria and its interaction with mitochondrial proteins has not been explored. We sought to explore the role played by PDH in mast cell exocytosis and to determine whether MITF is localized in the mitochondria and involved in regulation of PDH activity. Experiments were performed in vitro by using human and mouse mast cells, as well as rat basophil leukemia cells, and in vivo in mice. The effect of PDH inhibition on mast cell function was examined. PDH interaction with MITF was measured before and after immunologic activation. Furthermore, mitochondrial localization of MITF and its effect on PDH activity were determined. PDH is essential for immunologically mediated degranulation of mast cells. After activation, PDH is serine dephosphorylated. In addition, for the first time, we show that MITF is partially located in the mitochondria and interacts with PDH. This interaction is dependent on the phosphorylation state of PDH. Furthermore, mitochondrial MITF regulates PDH activity. The association of mitochondrial MITF with PDH emerges as an important regulator of mast cell function. Our findings indicate that PDH could arise as a new target for the manipulation of allergic diseases. Copyright © 2016 American Academy of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.

Dynamic genome wide expression profiling of Drosophila head development reveals a novel role of Hunchback in retinal glia cell development and blood-brain barrier integrity

PubMed Central

Torres-Oliva, Montserrat; Schneider, Julia; Wiegleb, Gordon

2018-01-01

Drosophila melanogaster head development represents a valuable process to study the developmental control of various organs, such as the antennae, the dorsal ocelli and the compound eyes from a common precursor, the eye-antennal imaginal disc. While the gene regulatory network underlying compound eye development has been extensively studied, the key transcription factors regulating the formation of other head structures from the same imaginal disc are largely unknown. We obtained the developmental transcriptome of the eye-antennal discs covering late patterning processes at the late 2nd larval instar stage to the onset and progression of differentiation at the end of larval development. We revealed the expression profiles of all genes expressed during eye-antennal disc development and we determined temporally co-expressed genes by hierarchical clustering. Since co-expressed genes may be regulated by common transcriptional regulators, we combined our transcriptome dataset with publicly available ChIP-seq data to identify central transcription factors that co-regulate genes during head development. Besides the identification of already known and well-described transcription factors, we show that the transcription factor Hunchback (Hb) regulates a significant number of genes that are expressed during late differentiation stages. We confirm that hb is expressed in two polyploid subperineurial glia cells (carpet cells) and a thorough functional analysis shows that loss of Hb function results in a loss of carpet cells in the eye-antennal disc. Additionally, we provide for the first time functional data indicating that carpet cells are an integral part of the blood-brain barrier. Eventually, we combined our expression data with a de novo Hb motif search to reveal stage specific putative target genes of which we find a significant number indeed expressed in carpet cells. PMID:29360820

PTTG1 Attenuates Drug-Induced Cellular Senescence

PubMed Central

Tong, Yunguang; Zhao, Weijiang; Zhou, Cuiqi; Wawrowsky, Kolja; Melmed, Shlomo

2011-01-01

As PTTG1 (pituitary tumor transforming gene) abundance correlates with adverse outcomes in cancer treatment, we determined mechanisms underlying this observation by assessing the role of PTTG1 in regulating cell response to anti-neoplastic drugs. HCT116 cells devoid of PTTG1 (PTTG1−/−) exhibited enhanced drug sensitivity as assessed by measuring BrdU incorporation in vitro. Apoptosis, mitosis catastrophe or DNA damage were not detected, but features of senescence were observed using low doses of doxorubicin and TSA. The number of drug-induced PTTG1−/− senescent cells increased ∼4 fold as compared to WT PTTG1-replete cells (p<0.001). p21, an important regulator of cell senescence, was induced ∼3 fold in HCT116 PTTG1−/− cells upon doxorubicin or Trichostatin A treatment. Binding of Sp1, p53 and p300 to the p21 promoter was enhanced in PTTG1−/− cells after treatment, suggesting transcriptional regulation of p21. p21 knock down abrogated the observed senescent effects of these drugs, indicating that PTTG1 likely suppresses p21 to regulate drug-induced senescence. PTTG1 also regulated SW620 colon cancer cells response to doxorubicin and TSA mediated by p21. Subcutaneously xenografted PTTG1−/− HCT116 cells developed smaller tumors and exhibited enhanced responses to doxorubicin. PTTG1−/− tumor tissue derived from excised tumors exhibited increased doxorubicin-induced senescence. As senescence is a determinant of cell responses to anti-neoplastic treatments, these findings suggest PTTG1 as a tumor cell marker to predict anti-neoplastic treatment outcomes. PMID:21858218

Counting molecules in single organelles with superresolution microscopy allows tracking of the endosome maturation trajectory

PubMed Central

Puchner, Elias M.; Walter, Jessica M.; Kasper, Robert; Huang, Bo; Lim, Wendell A.

2013-01-01

Cells tightly regulate trafficking of intracellular organelles, but a deeper understanding of this process is technically limited by our inability to track the molecular composition of individual organelles below the diffraction limit in size. Here we develop a technique for intracellularly calibrated superresolution microscopy that can measure the size of individual organelles as well as accurately count absolute numbers of molecules, by correcting for undercounting owing to immature fluorescent proteins and overcounting owing to fluorophore blinking. Using this technique, we characterized the size of individual vesicles in the yeast endocytic pathway and the number of accessible phosphatidylinositol 3-phosphate binding sites they contain. This analysis reveals a characteristic vesicle maturation trajectory of composition and size with both stochastic and regulated components. The trajectory displays some cell-to-cell variability, with smaller variation between organelles within the same cell. This approach also reveals mechanistic information on the order of events in this trajectory: Colocalization analysis with known markers of different vesicle maturation stages shows that phosphatidylinositol 3-phosphate production precedes fusion into larger endosomes. This single-organelle analysis can potentially be applied to a range of small organelles to shed light on their precise composition/structure relationships, the dynamics of their regulation, and the noise in these processes. PMID:24043832

FoxO1 regulates apoptosis induced by asbestos in the MT-2 human T-cell line.

PubMed

Matsuzaki, Hidenori; Lee, Suni; Maeda, Megumi; Kumagai-Takei, Naoko; Nishimura, Yasumitsu; Otsuki, Takemi

2016-09-01

Asbestos is known to cause malignant mesothelioma and lung cancer. Recent studies implicate tumor immunity in the development of various tumors, including malignant mesothelioma. In order to establish an in vitro T-cell model to clarify the effects of long-term exposure of asbestos on tumor immunity, in this study, human T-cell line MT-2 cells were cultured with asbestos for longer than 8 months and the resultant cells (MT-2Rst) were assessed for the expression of forkhead transcription factor FoxO1. Gene expression analysis revealed that the amount of FoxO1 mRNA decreased after long-term exposure of the MT-2 cells to asbestos. In accordance with this reduction in FoxO1, pro-apoptotic Foxo1 target genes Puma, Fas ligand and Bim were also seen to be down-regulated in MT-2Rst cells. Furthermore, shRNA-mediated knock-down of FoxO1 reduced the number of apoptotic parental MT-2 cells after treatment with asbestos. On the other hand, over-expression of FoxO1 did not affect asbestos-induced apoptosis in MT-2Rst cells. These results suggested that FoxO1 played an important role in regulating asbestos-induced apoptosis and confirmed the presence of multiple pathways regulating resistance to asbestos in MT-2Rst cells.

Regulation of the Arabidopsis root vascular initial population by LONESOME HIGHWAY

PubMed Central

Ohashi-Ito, Kyoko; Bergmann, Dominique C.

2011-01-01

Complex organisms consist of a multitude of cell types arranged in precise spatial relation to each other. Arabidopsis roots generally exhibit radial tissue organization; however, within a tissue layer, cells are not identical. Specific vascular cell types are arranged in diametrically opposed longitudinal files that maximize the distance between them and create a bilaterally symmetric (diarch) root. Mutations in the LONESOME HIGHWAY (LHW) gene eliminate bilateral symmetry and reduce the number of cells in the center of the root, resulting in roots with only single and xylem and phloem poles. LHW does not appear to be required for the creation of any specific cell type, but coordinately controls the number of all vascular cell types by regulating the size of the pool of cells from which they arise. We cloned LHW and found that it encodes a protein with weak sequence similarity to basic helix-loop-helix (bHLH) domain proteins. LHW is a transcriptional activator in vitro. In plants, LHW is nuclear localized and is expressed in the root meristems where we hypothesize it acts independently of other known root patterning genes to promote the production of stele cells, but may also indirectly feed into established regulatory networks for the maintenance of the root meristem. PMID:17626058

Polyamine spermidine is an upstream negator of ethylene-regulated pathogenesis of botrytis cinerea in tomato leaf

USDA-ARS?s Scientific Manuscript database

Polyamines are biogenic polycationic compounds implicated in a number of processes including plant cell division, cell elongation, flowering, fruit set and development, fruit ripening, senescence and responses to abiotic stresses. Comparatively, little is known about their role in plant-microbe int...

SH2 domain containing leukocyte phosphoprotein of 76-kDa (SLP-76) feedback regulation of ZAP-70 microclustering.

PubMed

Liu, Hebin; Purbhoo, Marco A; Davis, Daniel M; Rudd, Christopher E

2010-06-01

T cell receptor (TCR) signaling involves CD4/CD8-p56lck recruitment of ZAP-70 to the TCR receptor, ZAP-70 phosphorylation of LAT that is followed by LAT recruitment of the GADS-SLP-76 complex. Back regulation of ZAP-70 by SLP-76 has not been documented. In this paper, we show that anti-CD3 induced ZAP-70 cluster formation is significantly reduced in the absence of SLP-76 (i.e., J14 cells) and in the presence of a mutant of SLP-76 (4KE) in Jurkat and primary T cells. Both the number of cells with clusters and the number of clusters per cell were reduced. This effect was not mediated by SLP-76 SH2 domain binding to ZAP-70 because SLP-76 failed to precipitate ZAP-70 and an inactivating SH2 domain mutation (i.e., R448L) on SLP-76 4KE did not reverse the inhibition of ZAP-70 clustering. Mutation of R448 on WT SLP-76 still supported ZAP-70 clustering. Intriguingly, by contrast, LAT clustering occurred normally in the absence of SLP-76, or the presence of 4KE SLP-76 indicating that this transmembrane adaptor can operate independently of ZAP-70-GADS-SLP-76. Our findings reconfigure the TCR signaling pathway by showing SLP-76 back-regulation of ZAP-70, an event that could ensure that signaling components are in balance for optimal T cell activation.

Zic deficiency in the cortical marginal zone and meninges results in cortical lamination defects resembling those in type II lissencephaly.

PubMed

Inoue, Takashi; Ogawa, Masaharu; Mikoshiba, Katsuhiko; Aruga, Jun

2008-04-30

The formation of the highly organized cortical structure depends on the production and correct placement of the appropriate number and types of neurons. The Zic family of zinc-finger transcription factors plays essential roles in regulating the proliferation and differentiation of neuronal progenitors in the medial forebrain and the cerebellum. Examination of the expression of Zic genes demonstrated that Zic1, Zic2, and Zic3 were expressed by the progenitor cells in the septum and cortical hem, the sites of generation of the Cajal-Retzius (CR) cells. Immunohistochemical studies have revealed that Zic proteins were abundantly expressed in the meningeal cells and that the majority of the CR cells distributed in the medial and dorsal cortex also expressed Zic proteins in the mid-late embryonic and postnatal cortical marginal zones. During embryonic cortical development, Zic1/Zic3 double-mutant and hypomorphic Zic2 mutant mice showed a reduction in the number of CR cells in the rostral cortex, whereas the cell number remained unaffected in the caudal cortex. These mutants also showed mislocalization of the CR cells and cortical lamination defects, resembling the changes noted in type II (cobblestone) lissencephaly, throughout the brain. In the Zic1/3 mutant, reduced proliferation of the meningeal cells was observed before the thinner and disrupted organization of the pial basement membrane (BM) with reduced expression of the BM components and the meningeal cell-derived secretory factor. These defects correlated with the changes in the end feet morphology of the radial glial cells. These findings indicate that the Zic genes play critical roles in cortical development through regulating the proliferation of meningeal cells and the pial BM assembly.

MLH1 function is context dependent in colorectal cancers.

PubMed

Jackson, Thomas; Ahmed, Mohamed A H; Seth, Rashmi; Jackson, Darryl; Ilyas, Mohammad

2011-02-01

Loss of mismatch repair (MMR) function in sporadic colorectal cancer occurs most commonly because of inactivation of MLH1, and it causes an increase in mutation rate. However, it is uncertain whether loss of MMR alters any other cellular function. The aim of this study was to investigate the role of MMR in regulating cell numbers and apoptosis. MLH1 protein levels were manipulated by (a) cloning and forcibly expressing MLH1 in HCT116 (a cell line with MLH1 mutation) and RKO (a cell line with MLH1 silencing), and (b) knockdown of MLH1 in SW480 (a cell line with normal MMR function). Cell number and apoptotic bodies were measured in standard and 'high stress' (ie, after staurosporine exposure) conditions. Restoration of MLH1 function in HCT116 and RKO resulted in increased cell number (p<0.001 for both cell lines) and decreased numbers of floating apoptotic bodies (p<0.01 in HCT116) in standard culture conditions. However, on induction of apoptotic stress, restoration of MLH1 resulted in reduced cell numbers (p<0.005). Knockdown of MLH1 in SW480 had no effect on cell numbers or apoptosis. MLH1 function may be context dependent: in 'low stress' conditions it may act to inhibit apoptosis, while in 'high stress' conditions it may induce apoptosis. However, within the context of chromosomal instability, the effect of MLH1 on cell numbers is limited.

Inducible and Conditional Deletion of Extracellular Signal-regulated Kinase 5 Disrupts Adult Hippocampal Neurogenesis*

PubMed Central

Pan, Yung-Wei; Zou, Junhui; Wang, Wenbin; Sakagami, Hiroyuki; Garelick, Michael G.; Abel, Glen; Kuo, Chay T.; Storm, Daniel R.; Xia, Zhengui

2012-01-01

Recent studies have led to the exciting idea that adult-born neurons in the dentate gyrus of the hippocampus may play a role in hippocampus-dependent memory formation. However, signaling mechanisms that regulate adult hippocampal neurogenesis are not well defined. Here we report that extracellular signal-regulated kinase 5 (ERK5), a member of the mitogen-activated protein kinase family, is selectively expressed in the neurogenic regions of the adult mouse brain. We present evidence that shRNA suppression of ERK5 in adult hippocampal neural stem/progenitor cells (aNPCs) reduces the number of neurons while increasing the number of cells expressing markers for stem/progenitor cells or proliferation. Furthermore, shERK5 attenuates both transcription and neuronal differentiation mediated by Neurogenin 2, a transcription factor expressed in adult hippocampal neural progenitor cells. By contrast, ectopic activation of endogenous ERK5 signaling via expression of constitutive active MEK5, an upstream activating kinase for ERK5, promotes neurogenesis in cultured aNPCs and in the dentate gyrus of the mouse brain. Moreover, neurotrophins including NT3 activate ERK5 and stimulate neuronal differentiation in aNPCs in an ERK5-dependent manner. Finally, inducible and conditional deletion of ERK5 specifically in the neurogenic regions of the adult mouse brain delays the normal progression of neuronal differentiation and attenuates adult neurogenesis in vivo. These data suggest ERK5 signaling as a critical regulator of adult hippocampal neurogenesis. PMID:22645146

A Novel Role for C5a in B-1 Cell Homeostasis

PubMed Central

Bröker, Katharina; Figge, Julia; Magnusen, Albert F.; Manz, Rudolf A.; Köhl, Jörg; Karsten, Christian M.

2018-01-01

B-1 cells constitute a unique subpopulation of lymphocytes residing mainly in body cavities like the peritoneal cavity (PerC) but are also found in spleen and bone marrow (BM). As innate-like B cells, they mediate first line immune defense through low-affinity natural IgM (nIgM) antibodies. PerC B-1 cells can egress to the spleen and differentiate into nIgM antibody-secreting plasma cells that recognize conserved exogenous and endogenous cellular structures. Homing to and homeostasis within the PerC are regulated by the chemokine CXCL13 released by PerC macrophages and stroma cells. However, the exact mechanisms underlying the regulation of CXCL13 and B-1 homeostasis are not fully explored. B-1 cells play important roles in the inflammatory response to infection, autoimmunity, ischemia/reperfusion injury, obesity, and atherosclerosis. Remarkably, this list of inflammatory entities has a strong overlap with diseases that are regulated by complement suggesting a link between B-1 cells and the complement system. Interestingly, up to now, no data exist regarding the role of complement in B-1 cell biology. Here, we demonstrate for the first time that C5a regulates B-1 cell steady-state dynamics within the peritoneum, the spleen, and the BM. We found decreased B-1a cell numbers in the peritoneum and the spleen of C5aR1−/− mice associated with increased B1-a and B1-b numbers in the spleen and high serum titers of nIgM antibodies directed against phosphorylcholine and several pneumococcal polysaccharides. Similarly, peritoneal B-1a cells were decreased in the peritoneum and splenic B-1a and B-1b cells were increased in C5aR2−/− mice. The decrease in peritoneal B-1 cell numbers was associated with decreased peritoneal CXCL13 levels in C5aR1−/− and C5aR2−/− mice. In search for mechanisms, we found that combined TLR2 and IL-10 receptor activation in PerC macrophages induced strong CXCL13 production, which was significantly reduced in cells from C5aR1- and C5aR2-deficient mice and after combined C5aR-targeting. Such stimulation also induced marked local C5 production by PerC macrophages and C5a generation. Importantly, peritoneal in vivo administration of C5a increased CXCL13 production. Taken together, our findings suggest that local non-canonical C5 activation in PerC macrophages fuels CXCL13 production as a novel mechanism to control B-1 cell homeostasis. PMID:29520270

The NOTCH Ligand JAG1 Regulates GDNF Expression in Sertoli Cells

PubMed Central

Garcia, Thomas X.; Parekh, Parag; Gandhi, Pooja; Sinha, Krishna

2017-01-01

In the seminiferous epithelium of the testis, Sertoli cells are key niche cells directing proliferation and differentiation of spermatogonial stem cells (SSCs) into spermatozoa. Sertoli cells produce glial cell line-derived neurotrophic factor (GDNF), which is essential for SSC self-renewal and progenitor expansion. While the role of GDNF in the testis stem cell niche is established, little is known about how this factor is regulated. Our previous studies on NOTCH activity in Sertoli cells demonstrated a role of this pathway in limiting stem/progenitor cell numbers, thus ultimately downregulating sperm cell output. In this study we demonstrate through a double-mutant mouse model that NOTCH signaling in Sertoli cells functions solely through the canonical pathway. Further, we demonstrate through Dual luciferase assay and chromatin immunoprecipitation quantitative polymerase chain reaction (ChIP-qPCR) analysis that the NOTCH targets HES1 and HEY1, which are transcriptional repressors, directly downregulate GDNF expression by binding to the Gdnf promoter, thus antagonizing the effects of FSH/cAMP. Finally, we demonstrate that testicular stem/progenitors cells are activating NOTCH signaling in Sertoli cells in vivo and in vitro through the NOTCH ligand JAG1 at their surface, indicating that these cells may ensure their own homeostasis through negative feedback regulation. PMID:28051360

Transcriptional and post-transcriptional regulation of NK cell development and function

PubMed Central

Leong, Jeffrey W.; Wagner, Julia A.; Ireland, Aaron R.; Fehniger, Todd A.

2016-01-01

Natural killer (NK) cells are specialized innate lymphoid cells that survey against viral infections and malignancy. Numerous advances have improved our understanding of the molecular mechanisms that control NK cell development and function over the past decade. These include both studies on the regulatory effects of transcription factors and translational repression via microRNAs. In this review, we summarize our current knowledge of DNA-binding transcription factors that regulate gene expression and thereby orchestrate NK cell development and activation, with an emphasis on recent discoveries. Additionally, we highlight our understanding of how RNA-bindings microRNAs fine tune the NK cell molecular program. We also underscore the large number of open questions in field that are now being addressed using new technological approaches and genetically engineered model organisms. Ultimately, a deeper understanding of the basic molecular biology of NK cells will facilitate new strategies to manipulate NK cells for the treatment of human disease. PMID:26948928

TSA suppresses miR-106b-93-25 cluster expression through downregulation of MYC and inhibits proliferation and induces apoptosis in human EMC.

PubMed

Zhao, Zhi-Ning; Bai, Jiu-Xu; Zhou, Qiang; Yan, Bo; Qin, Wei-Wei; Jia, Lin-Tao; Meng, Yan-Ling; Jin, Bo-Quan; Yao, Li-Bo; Wang, Tao; Yang, An-Gang

2012-01-01

Histone deacetylase (HDAC) inhibitors are emerging as a novel class of anti-tumor agents and have manifested the ability to decrease proliferation and increase apoptosis in different cancer cells. A significant number of genes have been identified as potential effectors responsible for the anti-tumor function of HDAC inhibitor. However, the molecular mechanisms of these HDAC inhibitors in this process remain largely undefined. In the current study, we searched for microRNAs (miRs) that were affected by HDAC inhibitor trichostatin (TSA) and investigated their effects in endometrial cancer (EMC) cells. Our data showed that TSA significantly inhibited the growth of EMC cells and induced their apoptosis. Among the miRNAs that altered in the presence of TSA, the miR-106b-93-25 cluster, together with its host gene MCM7, were obviously down-regulated in EMC cells. p21 and BIM, which were identified as target genes of miR-106b-93-25 cluster, increased in TSA treated tumor cells and were responsible for cell cycle arrest and apoptosis. We further identified MYC as a regulator of miR-106b-93-25 cluster and demonstrated its down-regulation in the presence of TSA resulted in the reduction of miR-106b-93-25 cluster and up-regulation of p21 and BIM. More important, we found miR-106b-93-25 cluster was up-regulated in clinical EMC samples in association with the overexpression of MCM7 and MYC and the down-regulation of p21 and BIM. Thus our studies strongly indicated TSA inhibited EMC cell growth and induced cell apoptosis and cell cycle arrest at least partially through the down-regulation of the miR-106b-93-25 cluster and up-regulation of it's target genes p21 and BIM via MYC.

Calcium/calmodulin-dependent protein kinase II activity regulates the proliferative potential of growth plate chondrocytes.

PubMed

Li, Yuwei; Ahrens, Molly J; Wu, Amy; Liu, Jennifer; Dudley, Andrew T

2011-01-01

For tissues that develop throughout embryogenesis and into postnatal life, the generation of differentiated cells to promote tissue growth is at odds with the requirement to maintain the stem cell/progenitor cell population to preserve future growth potential. In the growth plate cartilage, this balance is achieved in part by establishing a proliferative phase that amplifies the number of progenitor cells prior to terminal differentiation into hypertrophic chondrocytes. Here, we show that endogenous calcium/calmodulin-dependent protein kinase II (CamkII, also known as Camk2) activity is upregulated prior to hypertrophy and that loss of CamkII function substantially blocks the transition from proliferation to hypertrophy. Wnt signaling and Pthrp-induced phosphatase activity negatively regulate CamkII activity. Release of this repression results in activation of multiple effector pathways, including Runx2- and β-catenin-dependent pathways. We present an integrated model for the regulation of proliferation potential by CamkII activity that has important implications for studies of growth control and adult progenitor/stem cell populations.

Arsenic trioxide-mediated growth inhibition in gallbladder carcinoma cells via down-regulation of Cyclin D1 transcription mediated by Sp1 transcription factor

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ai, Zhilong; Lu, Weiqi; Ton, Saixiong

2007-08-31

Gallbladder carcinoma (GBC), an aggressive and mostly lethal malignancy, is known to be resistant to a number of drug stimuli. Here, we demonstrated that arsenic trioxide inhibited the proliferation of gallbladder carcinoma in vivo and in vitro as well as the transcription of cell cycle-related protein Cyclin D1. And, Cyclin D1 overexpression inhibited the negative role of arsenic trioxide in cell cycle progression. We further explored the mechanisms by which arsenic trioxide affected Cyclin D1 transcription and found that the Sp1 transcription factor was down-regulated by arsenic trioxide, with a corresponding decrease in Cyclin D1 promoter activity. Taken together, thesemore » results suggested that arsenic trioxide inhibited gallbladder carcinoma cell proliferation via down-regulation of Cyclin D1 transcription in a Sp1-dependent manner, which provided a new mechanism of arsenic trioxide-involved cell proliferation and may have important therapeutic implications in gallbladder carcinoma patients.« less

The C. elegans SoxC protein SEM-2 opposes differentiation factors to promote a proliferative blast cell fate in the postembryonic mesoderm

PubMed Central

Tian, Chenxi; Shi, Herong; Colledge, Clark; Stern, Michael; Waterston, Robert; Liu, Jun

2011-01-01

The proper development of multicellular organisms requires precise regulation and coordination of cell fate specification, cell proliferation and differentiation. Abnormal regulation and coordination of these processes could lead to disease, including cancer. We have examined the function of the sole C. elegans SoxC protein, SEM-2, in the M lineage, which produces the postembryonic mesoderm. We found that SEM-2/SoxC is both necessary and sufficient to promote a proliferating blast cell fate, the sex myoblast fate, over a differentiated striated bodywall muscle fate. A number of factors control the specific expression of sem-2 in the sex myoblast precursors and their descendants. This includes direct control of sem-2 expression by a Hox-PBC complex. The crucial nature of the HOX/PBC factors in directly enhancing expression of this proliferative factor in the C. elegans M lineage suggests a possible more general link between Hox-PBC factors and SoxC proteins in regulating cell proliferation. PMID:21307099

Microtubule-regulating proteins and cAMP-dependent signaling in neuroblastoma differentiation.

PubMed

Muñoz-Llancao, Pablo; de Gregorio, Cristian; Las Heras, Macarena; Meinohl, Christopher; Noorman, Kevin; Boddeke, Erik; Cheng, Xiaodong; Lezoualc'h, Frank; Schmidt, Martina; Gonzalez-Billault, Christian

2017-03-01

Neurons are highly differentiated cells responsible for the conduction and transmission of information in the nervous system. The proper function of a neuron relies on the compartmentalization of their intracellular domains. Differentiated neuroblastoma cells have been extensively used to study and understand the physiology and cell biology of neuronal cells. Here, we show that differentiation of N1E-115 neuroblastoma cells is more pronounced upon exposure of a chemical analog of cyclic AMP (cAMP), db-cAMP. We next analysed the expression of key microtubule-regulating proteins in differentiated cells and the expression and activation of key cAMP players such as EPAC, PKA and AKAP79/150. Most of the microtubule-promoting factors were up regulated during differentiation of N1E-115 cells, while microtubule-destabilizing proteins were down regulated. We observed an increase in tubulin post-translational modifications related to microtubule stability. As expected, db-cAMP increased PKA- and EPAC-dependent signalling. Consistently, pharmacological modulation of EPAC activity instructed cell differentiation, number of neurites, and neurite length in N1E-115 cells. Moreover, disruption of the PKA-AKAP interaction reduced these morphometric parameters. Interestingly, PKA and EPAC act synergistically to induce neuronal differentiation in N1E-115. Altogether these results show that the changes observed in the differentiation of N1E-115 cells proceed by regulating several microtubule-stabilizing factors, and the acquisition of a neuronal phenotype is a process involving concerted although independent functions of EPAC and PKA. © 2017 Wiley Periodicals, Inc.

Quantitative Evaluation of CART-Containing Cells in Urinary Bladder of Rats with Renovascular Hypertension

PubMed Central

Janiuk, I.; Kasacka, I.

2015-01-01

Recent biological advances make it possible to discover new peptides associated with hypertension. The cocaine- and amphetamine-regulated transcript (CART) is a known factor in appetite and feeding behaviour. Various lines of evidence suggest that this peptide participates not only in control of feeding behaviour but also in the regulation of the cardiovascular and sympathetic systems and blood pressure. The role of CART in blood pressure regulation led us to undertake a study aimed at analysing quantitative changes in CART-containing cells in urinary bladders (UB) of rats with renovascular hypertension. We used the Goldblatt model of arterial hypertension (two-kidney, one clip) to evaluate quantitative changes. This model provides researchers with a commonly used tool to analyse the renin-angiotensin system of blood pressure control and, eventually, to develop drugs for the treatment of chronic hypertension. The study was performed on sections of urinary bladders of rats after 3-, 14-, 28-, 42 and 91 days from hypertension induction. Immunohistochemical identification of CART cells was performed on paraffin for the UBs of all the study animals. CART was detected in the endocrine cells, especially numerous in the submucosa and muscularis layers, with a few found in the transitional epithelium and only occasionally in serosa. Hypertension significantly increased the number of CART-positive cells in the rat UBs. After 3 and 42 days following the procedure, statistically significantly higher numbers of CART-positive cells were identified in comparison with the control animals. The differences between the hypertensive rats and the control animals concerned not only the number density of CART-immunoreactive cells but also their localization. After a 6-week period, each of the rats subjected to the renal artery clipping procedure developed stable hypertension. CART appeared in numerous transitional epithelium cells. As this study provides novel findings, the question appears about the type of connection between hypertension and the functioning and activity of CART in the urinary tract (UT). The study gives rise to the assumption that high blood pressure can be a factor that intensifies CART secretion. In conclusion, the endocrine system of the urinary tract is modified by renovascular hypertension. This may affect the production of hormones and biologically active substances and contribute to the development of possible hypertension complications. In order to fully comprehend the role of the CART peptide in blood pressure regulation, further analyses are necessary. PMID:26150151

Quantitative evaluation of CART-containing cells in urinary bladder of rats with renovascular hypertension.

PubMed

Janiuk, I; Kasacka, I

2015-04-13

Recent biological advances make it possible to discover new peptides associated with hypertension. The cocaine- and amphetamine-regulated transcript (CART) is a known factor in appetite and feeding behaviour. Various lines of evidence suggest that this peptide participates not only in control of feeding behaviour but also in the regulation of the cardiovascular and sympathetic systems and blood pressure. The role of CART in blood pressure regulation led us to undertake a study aimed at analysing quantitative changes in CART-containing cells in urinary bladders (UB) of rats with renovascular hypertension. We used the Goldblatt model of arterial hypertension (two-kidney, one clip) to evaluate quantitative changes. This model provides researchers with a commonly used tool to analyse the renin-angiotensin system of blood pressure control and, eventually, to develop drugs for the treatment of chronic hypertension. The study was performed on sections of urinary bladders of rats after 3-, 14-, 28-, 42 and 91 days from hypertension induction. Immunohistochemical identification of CART cells was performed on paraffin for the UBs of all the study animals. CART was detected in the endocrine cells, especially numerous in the submucosa and muscularis layers, with a few found in the transitional epithelium and only occasionally in serosa. Hypertension significantly increased the number of CART-positive cells in the rat UBs. After 3 and 42 days following the procedure, statistically significantly higher numbers of CART-positive cells were identified in comparison with the control animals. The differences between the hypertensive rats and the control animals concerned not only the number density of CART-immunoreactive cells but also their localization. After a 6-week period, each of the rats subjected to the renal artery clipping procedure developed stable hypertension. CART appeared in numerous transitional epithelium cells. As this study provides novel findings, the question appears about the type of connection between hypertension and the functioning and activity of CART in the urinary tract (UT). The study gives rise to the assumption that high blood pressure can be a factor that intensifies CART secretion. In conclusion, the endocrine system of the urinary tract is modified by renovascular hypertension. This may affect the production of hormones and biologically active substances and contribute to the development of possible hypertension complications. In order to fully comprehend the role of the CART peptide in blood pressure regulation, further analyses are necessary.

Independent regulation of the two Pax5 alleles during B-cell development.

PubMed

Nutt, S L; Vambrie, S; Steinlein, P; Kozmik, Z; Rolink, A; Weith, A; Busslinger, M

1999-04-01

The developmental control genes of the Pax family are frequently associated with mouse mutants and human disease syndromes. The function of these transcription factors is sensitive to gene dosage, as mutation of one allele or a modest increase in gene number results in phenotypic abnormalities. Pax5 has an important role in B-cell and midbrain development. By following the expression of individual Pax5 alleles at the single-cell level, we demonstrate here that Pax5 is subject to allele-specific regulation during B-lymphopoiesis. Pax5 is predominantly transcribed from only one allele in early progenitors and mature B cells, whereas it switches to a biallelic transcription mode in immature B cells. The allele-specific regulation of Pax5 is stochastic, reversible, independent of parental origin and correlates with synchronous replication, in contrast with imprinted and other monoallelically expressed genes. As a consequence, B-lymphoid tissues are mosaics with respect to the transcribed Pax5 allele, and thus mutation of one allele in heterozygous mice results in deletion of the cell population expressing the mutant allele due to loss of Pax5 function at the single-cell level. Similar allele-specific regulation may be a common mechanism causing the haploinsufficiency and frequent association of other Pax genes with human disease.

Microenvironmental Regulation of Mammary Carcinogenesis

DTIC Science & Technology

2009-06-01

cells, leukocytes 16. SECURITY CLASSIFICATION OF: 17. LIMITATION OF ABSTRACT 18 . NUMBER OF PAGES 19a. NAME OF RESPONSIBLE PERSON...metastatic spread to sentinel LNs and increased primary tumor size13. Perhaps more significant, the ratio of CD4+ to CD8+ T cells or TH2 to TH1 cells...present in primary tumors, where CD4+ or TH2 cells are more frequent than CD8+ or TH1 cells, correlates with LN metastasis and reduced overall patient

Caspase cascade regulated mitochondria mediated apoptosis in monocrotophos exposed PC12 cells.

PubMed

Kashyap, M P; Singh, A K; Siddiqui, M A; Kumar, V; Tripathi, V K; Khanna, V K; Yadav, S; Jain, S K; Pant, A B

2010-11-15

Monocrotophos (MCP) is a commonly used organophosphorus (OP) pesticide. We studied apoptotic changes in PC12 cells exposed to MCP. A significant induction in reactive oxygen species (ROS), lipid peroxide (LPO), and the ratio of glutathione disulfide (GSSG)/reduced glutathione (GSH) was observed in cells exposed to selected doses of MCP. Following the exposure of PC12 cells to MCP, the levels of protein and mRNA expressions of Caspase-3, Caspase-9, Bax, p53, P(21), Puma, and cytochrome-c were significantly upregulated, whereas the levels of Bcl(2), Bcl(w), and Mcl1 were downregulated. TUNEL assay, DNA laddering, and micronuclei induction show that long-term exposure of PC12 cells to MCP at higher concentration (10(-5) M) decreases the number of apoptotic events due to an increase in the number of necrotic cells. MCP-induced translocation of Bax and cytochrome-c proteins between the cytoplasm and mitochondria confirmed the role of p53 and Puma in mitochondrial membrane permeability. Mitochondria mediated apoptosis induction was confirmed by the increased activity of caspase cascade. We believe that this is the first report showing MCP-induced apoptosis in PC12 cells, which is mitochondria mediated and regulated through the caspase cascade. Our data demonstrates that MCP induced the apoptotic cell death in neuronal cells and identifies the possible cellular and molecular mechanisms of organophosphate pesticide-induced apoptosis in neuronal cells.

Expression of regulatory neuropeptides in the hypothalamus of red deer (Cervus elaphus) reveals anomalous relationships in the seasonal control of appetite and reproduction.

PubMed

Barrell, G K; Ridgway, M J; Wellby, M; Pereira, A; Henry, B A; Clarke, I J

2016-04-01

Red deer are seasonal with respect to reproduction and food intake, so we tested the hypothesis that their brains would show seasonal changes in numbers of cells containing hypothalamic neuropeptides that regulate these functions. We examined the brains of male and female deer in non-breeding and breeding seasons to quantify the production of kisspeptin, gonadotropin inhibitory hormone (GnIH), neuropeptide Y (NPY) and γ-melanocyte stimulating hormone (γ-MSH - an index of pro-opiomelanocortin production), using immunohistochemistry. These neuropeptides are likely to be involved in the regulation of reproductive function and appetite. During the annual breeding season there were more cells producing kisspeptin in the arcuate nucleus of the hypothalamus than during the non-breeding season in males and females whereas there was no seasonal difference in the expression of GnIH. There were more cells producing the appetite stimulating peptide, NPY, in the arcuate/median eminence regions of the hypothalamus of females during the non-breeding season whereas the levels of an appetite suppressing peptide, γ-MSH, were highest in the breeding season. Male deer brains exhibited the converse, with NPY cell numbers highest in the breeding season and γ-MSH levels highest in the non-breeding season. These results support a role for kisspeptin as an important stimulatory regulator of seasonal breeding in deer, as in other species, but suggest a lack of involvement of GnIH in the seasonality of reproduction in deer. In the case of appetite regulation, the pattern exhibited by females for NPY and γ-MSH was as expected for the breeding and non-breeding seasons, based on previous studies of these peptides in sheep and the seasonal cycle of appetite reported for various species of deer. An inverse result in male deer most probably reflects the response of appetite regulating cells to negative energy balance during the mating season. Differences between the sexes in the seasonal changes in appetite regulating peptide cells of the hypothalamus present an interesting model for future studies. Copyright © 2016 Elsevier Inc. All rights reserved.

Circadian clock regulation of the cell cycle in the zebrafish intestine.

PubMed

Peyric, Elodie; Moore, Helen A; Whitmore, David

2013-01-01

The circadian clock controls cell proliferation in a number of healthy tissues where cell renewal and regeneration are critical for normal physiological function. The intestine is an organ that typically undergoes regular cycles of cell division, differentiation and apoptosis as part of its role in digestion and nutrient absorption. The aim of this study was to explore circadian clock regulation of cell proliferation and cell cycle gene expression in the zebrafish intestine. Here we show that the zebrafish gut contains a directly light-entrainable circadian pacemaker, which regulates the daily timing of mitosis. Furthermore, this intestinal clock controls the expression of key cell cycle regulators, such as cdc2, wee1, p21, PCNA and cdk2, but only weakly influences cyclin B1, cyclin B2 and cyclin E1 expression. Interestingly, food deprivation has little impact on circadian clock function in the gut, but dramatically reduces cell proliferation, as well as cell cycle gene expression in this tissue. Timed feeding under constant dark conditions is able to drive rhythmic expression not only of circadian clock genes, but also of several cell cycle genes, suggesting that food can entrain the clock, as well as the cell cycle in the intestine. Rather surprisingly, we found that timed feeding is critical for high amplitude rhythms in cell cycle gene expression, even when zebrafish are maintained on a light-dark cycle. Together these results suggest that the intestinal clock integrates multiple rhythmic cues, including light and food, to function optimally.

Circadian Clock Regulation of the Cell Cycle in the Zebrafish Intestine

PubMed Central

Peyric, Elodie; Moore, Helen A.; Whitmore, David

2013-01-01

The circadian clock controls cell proliferation in a number of healthy tissues where cell renewal and regeneration are critical for normal physiological function. The intestine is an organ that typically undergoes regular cycles of cell division, differentiation and apoptosis as part of its role in digestion and nutrient absorption. The aim of this study was to explore circadian clock regulation of cell proliferation and cell cycle gene expression in the zebrafish intestine. Here we show that the zebrafish gut contains a directly light-entrainable circadian pacemaker, which regulates the daily timing of mitosis. Furthermore, this intestinal clock controls the expression of key cell cycle regulators, such as cdc2, wee1, p21, PCNA and cdk2, but only weakly influences cyclin B1, cyclin B2 and cyclin E1 expression. Interestingly, food deprivation has little impact on circadian clock function in the gut, but dramatically reduces cell proliferation, as well as cell cycle gene expression in this tissue. Timed feeding under constant dark conditions is able to drive rhythmic expression not only of circadian clock genes, but also of several cell cycle genes, suggesting that food can entrain the clock, as well as the cell cycle in the intestine. Rather surprisingly, we found that timed feeding is critical for high amplitude rhythms in cell cycle gene expression, even when zebrafish are maintained on a light-dark cycle. Together these results suggest that the intestinal clock integrates multiple rhythmic cues, including light and food, to function optimally. PMID:24013905

Cathelin-related antimicrobial peptide differentially regulates T- and B-cell function

PubMed Central

Kin, Nicholas W.; Chen, Yao; Stefanov, Emily K.; Gallo, Richard L.; Kearney, John F.

2011-01-01

Mammalian antimicrobial peptides (AMPs) play an important role in host defense via direct antimicrobial activity as well as immune regulation. The mouse cathelin-related antimicrobial peptide (mCRAMP), produced from the mouse gene Camp, is the only mouse cathelicidin identified and the ortholog of the human gene encoding the peptide LL-37. This study tested the hypothesis that mouse B and T cells produce and respond to mCRAMP. We show that all mature mouse B-cell subsets, including follicular (FO), marginal zone (MZ), B1a, and B1b cells, as well as CD4+ and CD8+ T cells produce Camp mRNA and mCRAMP protein. Camp−/− B cells produced equivalent levels of IgM, IgG3, and IgG2c but less IgG1 and IgE, while Camp−/− CD4+ T cells cultured in Th2-inducing conditions produced more IL-4-expressing cells when compared with WT cells, effects that were reversed upon addition of mCRAMP. In vivo, Camp−/− mice immunized with TNP-OVA absorbed in alum produced an enhanced TNP-specific IgG1 response when compared with WT mice. ELISpot analysis revealed increased numbers of TNP-specific IgG1-secreting splenic B cells and FACS analysis revealed increased CD4+ T-cell IL-4 expression. Our results suggest that mCRAMP differentially regulates B- and T-cell function and implicate mCRAMP in the regulation of adaptive immune responses. PMID:21773974

In tobacco BY-2 cells xyloglucan oligosaccharides alter the expression of genes involved in cell wall metabolism, signalling, stress responses, cell division and transcriptional control.

PubMed

González-Pérez, Lien; Perrotta, Lara; Acosta, Alexis; Orellana, Esteban; Spadafora, Natasha; Bruno, Leonardo; Bitonti, Beatrice M; Albani, Diego; Cabrera, Juan Carlos; Francis, Dennis; Rogers, Hilary J

2014-10-01

Xyloglucan oligosaccharides (XGOs) are breakdown products of XGs, the most abundant hemicelluloses of the primary cell walls of non-Poalean species. Treatment of cell cultures or whole plants with XGOs results in accelerated cell elongation and cell division, changes in primary root growth, and a stimulation of defence responses. They may therefore act as signalling molecules regulating plant growth and development. Previous work suggests an interaction with auxins and effects on cell wall loosening, however their mode of action is not fully understood. The effect of an XGO extract from tamarind (Tamarindus indica) on global gene expression was therefore investigated in tobacco BY-2 cells using microarrays. Over 500 genes were differentially regulated with similar numbers and functional classes of genes up- and down-regulated, indicating a complex interaction with the cellular machinery. Up-regulation of a putative XG endotransglycosylase/hydrolase-related (XTH) gene supports the mechanism of XGO action through cell wall loosening. Differential expression of defence-related genes supports a role for XGOs as elicitors. Changes in the expression of genes related to mitotic control and differentiation also support previous work showing that XGOs are mitotic inducers. XGOs also affected expression of several receptor-like kinase genes and transcription factors. Hence, XGOs have significant effects on expression of genes related to cell wall metabolism, signalling, stress responses, cell division and transcriptional control.

Genome-wide differential gene expression in immortalized DF-1 chicken embryo fibroblast cell line

PubMed Central

2011-01-01

Background When compared to primary chicken embryo fibroblast (CEF) cells, the immortal DF-1 CEF line exhibits enhanced growth rates and susceptibility to oxidative stress. Although genes responsible for cell cycle regulation and antioxidant functions have been identified, the genome-wide transcription profile of immortal DF-1 CEF cells has not been previously reported. Global gene expression in primary CEF and DF-1 cells was performed using a 4X44K chicken oligo microarray. Results A total of 3876 differentially expressed genes were identified with a 2 fold level cutoff that included 1706 up-regulated and 2170 down-regulated genes in DF-1 cells. Network and functional analyses using Ingenuity Pathways Analysis (IPA, Ingenuity® Systems, http://www.ingenuity.com) revealed that 902 of 3876 differentially expressed genes were classified into a number of functional groups including cellular growth and proliferation, cell cycle, cellular movement, cancer, genetic disorders, and cell death. Also, the top 5 gene networks with intermolecular connections were identified. Bioinformatic analyses suggested that DF-1 cells were characterized by enhanced molecular mechanisms for cell cycle progression and proliferation, suppressing cell death pathways, altered cellular morphogenesis, and accelerated capacity for molecule transport. Key molecules for these functions include E2F1, BRCA1, SRC, CASP3, and the peroxidases. Conclusions The global gene expression profiles provide insight into the cellular mechanisms that regulate the unique characteristics observed in immortal DF-1 CEF cells. PMID:22111699

RNA splicing and splicing regulator changes in prostate cancer pathology.

PubMed

Munkley, Jennifer; Livermore, Karen; Rajan, Prabhakar; Elliott, David J

2017-09-01

Changes in mRNA splice patterns have been associated with key pathological mechanisms in prostate cancer progression. The androgen receptor (abbreviated AR) transcription factor is a major driver of prostate cancer pathology and activated by androgen steroid hormones. Selection of alternative promoters by the activated AR can critically alter gene function by switching mRNA isoform production, including creating a pro-oncogenic isoform of the normally tumour suppressor gene TSC2. A number of androgen-regulated genes generate alternatively spliced mRNA isoforms, including a prostate-specific splice isoform of ST6GALNAC1 mRNA. ST6GALNAC1 encodes a sialyltransferase that catalyses the synthesis of the cancer-associated sTn antigen important for cell mobility. Genetic rearrangements occurring early in prostate cancer development place ERG oncogene expression under the control of the androgen-regulated TMPRSS2 promoter to hijack cell behaviour. This TMPRSS2-ERG fusion gene shows different patterns of alternative splicing in invasive versus localised prostate cancer. Alternative AR mRNA isoforms play a key role in the generation of prostate cancer drug resistance, by providing a mechanism through which prostate cancer cells can grow in limited serum androgen concentrations. A number of splicing regulator proteins change expression patterns in prostate cancer and may help drive key stages of disease progression. Up-regulation of SRRM4 establishes neuronal splicing patterns in neuroendocrine prostate cancer. The splicing regulators Sam68 and Tra2β increase expression in prostate cancer. The SR protein kinase SRPK1 that modulates the activity of SR proteins is up-regulated in prostate cancer and has already given encouraging results as a potential therapeutic target in mouse models.

Hypoxia regulates microRNA expression in the human carotid body

DOE Office of Scientific and Technical Information (OSTI.GOV)

Mkrtchian, Souren, E-mail: souren.mkrtchian@ki.se; Lee, Kian Leong, E-mail: csilkl@nus.edu.sg; Kåhlin, Jessica

The carotid body (CB) is the key sensing organ for physiological oxygen levels in the body. Under conditions of low oxygen (hypoxia), the CB plays crucial roles in signaling to the cardiorespiratory center in the medulla oblongata for the restoration of oxygen homeostasis. How hypoxia regulates gene expression in the human CB remains poorly understood. While limited information on transcriptional regulation in animal CBs is available, the identity and impact of important post-transcriptional regulators such as non-coding RNAs, and in particular miRNAs are not known. Here we show using ex vivo experiments that indeed a number of miRNAs are differentiallymore » regulated in surgically removed human CB slices when acute hypoxic conditions were applied. Analysis of the hypoxia-regulated miRNAs shows that they target biological pathways with upregulation of functions related to cell proliferation and immune response and downregulation of cell differentiation and cell death functions. Comparative analysis of the human CB miRNAome with the global miRNA expression patterns of a large number of different human tissues showed that the CB miRNAome had a unique profile which reflects its highly specialized functional status. Nevertheless, the human CB miRNAome is most closely related to the miRNA expression pattern of brain tissues indicating that they may have the most similar developmental origins. - Highlights: • Hypoxia triggers differential expression of many miRNAs in the human carotid body. • This can lead to the upregulation of proliferation and immune response functions. • CB expression profile in the carotid body resembles the miRNA expression pattern in the brain. • miRNAs are involved in the regulation of carotid body functions including oxygen sensing.« less

Targeted deletion of c-Met in thymic epithelial cells leads to an autoimmune phenotype

PubMed Central

Su, Min; Hu, Rong; Song, Yinhong; Liu, Yalan; Lai, Laijun

2017-01-01

Hepatocyte growth factor (HGF) and its receptor c-Met signaling have been implicated in regulating various types of cells including epithelial cells. We have previously reported that c-Met is expressed by thymic epithelial cells (TECs), and that in vivo administration of hybrid cytokines containing IL-7 and the beta- or alpha-chain of HGF significantly increase the number of TECs. In order to study the role of c-Met signaling in TECs, we generated conditional knockout (cKO) mice in which c-Met was specifically deleted in TECs using a Foxn1-Cre transgene. We show here that c-Met deficiency in TECs results in age-progressive reduction in TEC number and reduced number of regulatory T cells. Consequently, c-Met TEC cKO mice displayed an autoimmune phenotype. Thus, c-Met signaling in TECs is important for the maintenance of TECs and immune self-tolerance. PMID:29363160

PRMT5 is essential for the maintenance of chondrogenic progenitor cells in the limb bud

PubMed Central

Norrie, Jacqueline L.; Li, Qiang; Co, Swanie; Huang, Bau-Lin; Ding, Ding; Uy, Jann C.; Ji, Zhicheng; Mackem, Susan; Bedford, Mark T.; Galli, Antonella; Ji, Hongkai

2016-01-01

During embryonic development, undifferentiated progenitor cells balance the generation of additional progenitor cells with differentiation. Within the developing limb, cartilage cells differentiate from mesodermal progenitors in an ordered process that results in the specification of the correct number of appropriately sized skeletal elements. The internal pathways by which these cells maintain an undifferentiated state while preserving their capacity to differentiate is unknown. Here, we report that the arginine methyltransferase PRMT5 has a crucial role in maintaining progenitor cells. Mouse embryonic buds lacking PRMT5 have severely truncated bones with wispy digits lacking joints. This novel phenotype is caused by widespread cell death that includes mesodermal progenitor cells that have begun to precociously differentiate into cartilage cells. We propose that PRMT5 maintains progenitor cells through its regulation of Bmp4. Intriguingly, adult and embryonic stem cells also require PRMT5 for maintaining pluripotency, suggesting that similar mechanisms might regulate lineage-restricted progenitor cells during organogenesis. PMID:27827819

PRMT5 is essential for the maintenance of chondrogenic progenitor cells in the limb bud.

PubMed

Norrie, Jacqueline L; Li, Qiang; Co, Swanie; Huang, Bau-Lin; Ding, Ding; Uy, Jann C; Ji, Zhicheng; Mackem, Susan; Bedford, Mark T; Galli, Antonella; Ji, Hongkai; Vokes, Steven A

2016-12-15

During embryonic development, undifferentiated progenitor cells balance the generation of additional progenitor cells with differentiation. Within the developing limb, cartilage cells differentiate from mesodermal progenitors in an ordered process that results in the specification of the correct number of appropriately sized skeletal elements. The internal pathways by which these cells maintain an undifferentiated state while preserving their capacity to differentiate is unknown. Here, we report that the arginine methyltransferase PRMT5 has a crucial role in maintaining progenitor cells. Mouse embryonic buds lacking PRMT5 have severely truncated bones with wispy digits lacking joints. This novel phenotype is caused by widespread cell death that includes mesodermal progenitor cells that have begun to precociously differentiate into cartilage cells. We propose that PRMT5 maintains progenitor cells through its regulation of Bmp4 Intriguingly, adult and embryonic stem cells also require PRMT5 for maintaining pluripotency, suggesting that similar mechanisms might regulate lineage-restricted progenitor cells during organogenesis. © 2016. Published by The Company of Biologists Ltd.

Regulation of exosome release from mammary epithelial and breast cancer cells - a new regulatory pathway.

PubMed

Riches, Andrew; Campbell, Elaine; Borger, Eva; Powis, Simon

2014-03-01

Exosomes are small 50-100nm sized extracellular vesicles released from normal and tumour cells and are a source of a new intercellular communication pathway. Tumour exosomes promote tumour growth and progression. What regulates the release and homoeostatic levels of exosomes, in cancer, in body fluids remains undefined. We utilised a human mammary epithelial cell line (HMEC B42) and a breast cancer cell line derived from it (B42 clone 16) to investigate exosome production and regulation. Exosome numbers were quantified using a Nanosight LM10 and measured in culture supernatants in the absence and presence of exosomes in the medium. Concentrated suspensions of exosomes from the normal mammary epithelial cells, the breast cancer cells and bladder cancer cells were used. The interaction of exosomes with tumour cells was also investigated using fluorescently labelled exosomes. Exosome release from normal human mammary epithelial cells and breast cancer cells is regulated by the presence of exosomes, derived from their own cells, in the extracellular environment of the cells. Exosomes from normal mammary epithelial cells also inhibit exosome secretion by breast cancer cells, which occurs in a tissue specific manner. Labelled exosomes from mammary epithelial cells are internalised into the tumour cells implicating a dynamic equilibrium and suggesting a mechanism for feedback control. These data suggest a previously unknown novel feedback regulatory mechanism for controlling exosome release, which may highlight a new therapeutic approach to controlling the deleterious effects of tumour exosomes. This regulatory mechanism is likely to be generic to other tumours. Copyright © 2014 Elsevier Ltd. All rights reserved.

Amplification and up-regulation of MIR30D was associated with disease progression of cervical squamous cell carcinomas.

PubMed

Zhou, You; Hao, Yinghua; Li, Yuxia; Li, Ruizhen; Wu, Ruifang; Wang, Shubin; Fang, Zhengyu

2017-03-29

Cervical squamous cell carcinoma (CSCC) is the most frequent type among cervical cancers. Although the altered miRNA miR-30d expression and the amplified chromosome locus of MIR30D, 8q24, have been reported in somatic cancers, the definitive functional impact of such region especially in CSCC remains under-investigated. One hundred thirty-six cases of CSCC tissues and matched adjacent normal ovarian epithelial tissues were assessed in this study. FISH and qPCR were performed to detect the copy number and microRNA expression of MIR30D gene in the collected samples. In in-vitro study, proliferation of CSCC cells were analyzed using WST-1 assay and invasion abilities of CSCC cells were evaluated by transwell assay. In-vivo study using a model of nude mice bearing tumor was also performed. Copy number gains of MIR30D were detected in 22.8% (31 out of 136) of CSCC samples. Copy number of MIR30D was positively correlated with tumor progression. CSCCs with lymph node metastases (LNM) also showed more frequencies (36.4%) of MIR30D amplification than those without LNM (18.4%, p < 0.05). CSCCs with increased copy number of MIR30D also showed a positive correlation with miR-30d up-regulation. Inhibition of miR-30d in CSCC cells led to impaired tumor growth and migration. Copy number amplifications of MIR30D gene and enhanced expression of miR-30d were positively correlated with tumor progression in CSCCs, indicating miR-30d might play an oncomiric role in the progression of CSCC.

A potential inhibitory function of draxin in regulating mouse trunk neural crest migration.

PubMed

Zhang, Sanbing; Su, Yuhong; Gao, Jinbao; Zhang, Chenbing; Tanaka, Hideaki

2017-01-01

Draxin is a repulsive axon guidance protein that plays important roles in the formation of three commissures in the central nervous system and dorsal interneuron 3 (dI3) in the chick spinal cord. In the present study, we report the expression pattern of mouse draxin in the embryonic mouse trunk spinal cord. In the presence of draxin, the longest net migration length of a migrating mouse trunk neural crest cell was significantly reduced. In addition, the relative number of apolar neural crest cells increased as the draxin treatment time increased. Draxin caused actin cytoskeleton rearrangement in the migrating trunk neural crest cells. Our data suggest that draxin may regulate mouse trunk neural crest cell migration by the rearrangement of cell actin cytoskeleton and by reducing the polarization activity of these cells subsequently.

The TCP4 transcription factor regulates trichome cell differentiation by directly activating GLABROUS INFLORESCENCE STEMS in Arabidopsis thaliana.

PubMed

Vadde, Batthula Vijaya Lakshmi; Challa, Krishna Reddy; Nath, Utpal

2018-01-01

Trichomes are the first cell type to be differentiated during the morphogenesis of leaf epidermis and serve as an ideal model to study cellular differentiation. Many genes involved in the patterning and differentiation of trichome cells have been studied over the past decades, and the majority of these genes encode transcription factors that specifically regulate epidermal cell development. However, the upstream regulators of these genes that link early leaf morphogenesis with cell type differentiation are less studied. The TCP proteins are the plant-specific transcription factors involved in regulating diverse aspects of plant development including lateral organ morphogenesis by modulating cell proliferation and differentiation. Here, we show that the miR319-regulated class II TCP proteins, notably TCP4, suppress trichome branching in Arabidopsis leaves and inflorescence stem by direct transcriptional activation of GLABROUS INFLORESCENCE STEMS (GIS), a known negative regulator of trichome branching. The trichome branch number is increased in plants with reduced TCP activity and decreased in the gain-of-function lines of TCP4. Biochemical analyses show that TCP4 binds to the upstream regulatory region of GIS and activates its expression. Detailed genetic analyses show that GIS and TCP4 work in same pathway and GIS function is required for TCP4-mediated regulation of trichome differentiation. Taken together, these results identify a role for the class II TCP genes in trichome differentiation, thus providing a connection between organ morphogenesis and cellular differentiation. © 2017 The Authors The Plant Journal © 2017 John Wiley & Sons Ltd.

Peripheral cannabinoid receptor, CB2, regulates bone mass

PubMed Central

Ofek, Orr; Karsak, Meliha; Leclerc, Nathalie; Fogel, Meirav; Frenkel, Baruch; Wright, Karen; Tam, Joseph; Attar-Namdar, Malka; Kram, Vardit; Shohami, Esther; Mechoulam, Raphael; Zimmer, Andreas; Bab, Itai

2006-01-01

The endogenous cannabinoids bind to and activate two G protein-coupled receptors, the predominantly central cannabinoid receptor type 1 (CB1) and peripheral cannabinoid receptor type 2 (CB2). Whereas CB1 mediates the cannabinoid psychotropic, analgesic, and orectic effects, CB2 has been implicated recently in the regulation of liver fibrosis and atherosclerosis. Here we show that CB2-deficient mice have a markedly accelerated age-related trabecular bone loss and cortical expansion, although cortical thickness remains unaltered. These changes are reminiscent of human osteoporosis and may result from differential regulation of trabecular and cortical bone remodeling. The CB2–/– phenotype is also characterized by increased activity of trabecular osteoblasts (bone-forming cells), increased osteoclast (the bone-resorbing cell) number, and a markedly decreased number of diaphyseal osteoblast precursors. CB2 is expressed in osteoblasts, osteocytes, and osteoclasts. A CB2-specific agonist that does not have any psychotropic effects enhances endocortical osteoblast number and activity and restrains trabecular osteoclastogenesis, apparently by inhibiting proliferation of osteoclast precursors and receptor activator of NF-κB ligand expression in bone marrow-derived osteoblasts/stromal cells. The same agonist attenuates ovariectomy-induced bone loss and markedly stimulates cortical thickness through the respective suppression of osteoclast number and stimulation of endocortical bone formation. These results demonstrate that the endocannabinoid system is essential for the maintenance of normal bone mass by osteoblastic and osteoclastic CB2 signaling. Hence, CB2 offers a molecular target for the diagnosis and treatment of osteoporosis, the most prevalent degenerative disease in developed countries. PMID:16407142

Isolation of a novel cell wall architecture mutant of rice with defective Arabidopsis COBL4 ortholog BC1 required for regulated deposition of secondary cell wall components.

PubMed

Sato, Kanna; Suzuki, Ryu; Nishikubo, Nobuyuki; Takenouchi, Sachi; Ito, Sachiko; Nakano, Yoshimi; Nakaba, Satoshi; Sano, Yuzou; Funada, Ryo; Kajita, Shinya; Kitano, Hidemi; Katayama, Yoshihiro

2010-06-01

The plant secondary cell wall is a highly ordered structure composed of various polysaccharides, phenolic components and proteins. Its coordinated regulation of a number of complex metabolic pathways and assembly has not been resolved. To understand the molecular mechanisms that regulate secondary cell wall synthesis, we isolated a novel rice mutant, cell wall architecture1 (cwa1), that exhibits an irregular thickening pattern in the secondary cell wall of sclerenchyma, as well as culm brittleness and reduced cellulose content in mature internodes. Light and transmission electron microscopy revealed that the cwa1 mutant plant has regions of local aggregation in the secondary cell walls of the cortical fibers in its internodes, showing uneven thickness. Ultraviolet microscopic observation indicated that localization of cell wall phenolic components was perturbed and that these components abundantly deposited at the aggregated cell wall regions in sclerenchyma. Therefore, regulation of deposition and assembly of secondary cell wall materials, i.e. phenolic components, appear to be disturbed by mutation of the cwa1 gene. Genetic analysis showed that cwa1 is allelic to brittle culm1 (bc1), which encodes the glycosylphosphatidylinositol-anchored COBRA-like protein specifically in plants. BC1 is known as a regulator that controls the culm mechanical strength and cellulose content in the secondary cell walls of sclerenchyma, but the precise function of BC1 has not been resolved. Our results suggest that CWA1/BC1 has an essential role in assembling cell wall constituents at their appropriate sites, thereby enabling synthesis of solid and flexible internodes in rice.

Signatures of combinatorial regulation in intrinsic biological noise

PubMed Central

Warmflash, Aryeh; Dinner, Aaron R.

2008-01-01

Gene expression is controlled by the action of transcription factors that bind to DNA and influence the rate at which a gene is transcribed. The quantitative mapping between the regulator concentrations and the output of the gene is known as the cis-regulatory input function (CRIF). Here, we show how the CRIF shapes the form of the joint probability distribution of molecular copy numbers of the regulators and the product of a gene. Namely, we derive a class of fluctuation-based relations that relate the moments of the distribution to the derivatives of the CRIF. These relations are useful because they enable statistics of naturally arising cell-to-cell variations in molecular copy numbers to substitute for traditional manipulations for probing regulatory mechanisms. We demonstrate that these relations can distinguish super- and subadditive gene regulatory scenarios (molecular analogs of AND and OR logic operations) in simulations that faithfully represent bacterial gene expression. Applications and extensions to other regulatory scenarios are discussed. PMID:18981421

Eosinophils Regulate Peripheral B Cell Numbers in Both Mice and Humans

PubMed Central

Wong, Tina W.; Doyle, Alfred D.; Lee, James J.; Jelinek, Diane F.

2014-01-01

The view of eosinophils (Eos) as solely effector cells involved in host parasite defense and in the pathophysiology of allergic diseases has been challenged in recent years. In fact, there is a growing realization that these cells interact with other components of innate and adaptive immunity. For example, mouse Eos were recently demonstrated to promote plasma cell retention in the bone marrow. However, it remains unknown whether Eos influence the biology of normal B lymphocytes. In this study, we specifically assessed the effect of Eos on B cell survival, proliferation, and immunoglobulin secretion. Our data first revealed that the genetic deletion of Eos from NJ1638 IL-5 transgenic hypereosinophilic mice (previously shown to display profound B cell expansion) resulted in the near abolishment of the B cell lymphocytosis. In vitro studies using human tissues demonstrated Eos’ proximity to B cell follicles and their ability to promote B cell survival, proliferation, and immunoglobulin secretion via a contact-independent mechanism(s). Additionally, this ability of Eos to enhance B cell responsiveness was observed in both T-independent and T-dependent B cell activation and appears to be independent of the Eos’ activation state. Finally, a retrospective clinical study of hypereosinophilic patients revealed for the first time a direct correlation between peripheral blood eosinophil levels and B cell numbers. Taken together, our study identifies a novel role for Eos in the regulation of humoral immunity via their impact on B cell homeostasis and proliferation upon activation. PMID:24616476

Tumor necrosis factor-alpha inhibits stem cell factor-induced proliferation of human bone marrow progenitor cells in vitro. Role of p55 and p75 tumor necrosis factor receptors.

PubMed Central

Rusten, L S; Smeland, E B; Jacobsen, F W; Lien, E; Lesslauer, W; Loetscher, H; Dubois, C M; Jacobsen, S E

1994-01-01

Stem cell factor (SCF), a key regulator of hematopoiesis, potently synergizes with a number of hematopoietic growth factors. However, little is known about growth factors capable of inhibiting the actions of SCF. TNF-alpha has been shown to act as a bidirectional regulator of myeloid cell proliferation and differentiation. This study was designed to examine interactions between TNF-alpha and SCF. Here, we demonstrate that TNF-alpha potently and directly inhibits SCF-stimulated proliferation of CD34+ hematopoietic progenitor cells. Furthermore, TNF-alpha blocked all colony formation stimulated by SCF in combination with granulocyte colony-stimulating factor (CSF) or CSF-1. The synergistic effect of SCF observed in combination with GM-CSF or IL-3 was also inhibited by TNF-alpha, resulting in colony numbers similar to those obtained in the absence of SCF. These effects of TNF-alpha were mediated through the p55 TNF receptor, whereas little or no inhibition was signaled through the p75 TNF receptor. Finally, TNF-alpha downregulated c-kit cell-surface expression on CD34+ bone marrow cells, and this was predominantly a p55 TNF receptor-mediated event as well. Images PMID:7518828

Poly(ADP-ribose) polymerase-1 (Parp-1)-deficient mice demonstrate abnormal antibody responses

PubMed Central

Ambrose, Helen E; Willimott, Shaun; Beswick, Richard W; Dantzer, Françoise; de Murcia, Josiane Ménissier; Yelamos, José; Wagner, Simon D

2009-01-01

Poly(ADP-ribosylation) of acceptor proteins is an epigenetic modification involved in DNA strand break repair, recombination and transcription. Here we provide evidence for the involvement of poly(ADP-ribose) polymerase-1 (Parp-1) in antibody responses. Parp-1−/− mice had increased numbers of T cells and normal numbers of total B cells. Marginal zone B cells were mildly reduced in number, and numbers of follicular B cells were preserved. There were abnormal levels of basal immunoglobulins, with reduced levels of immunoglobulin G2a (IgG2a) and increased levels of IgA and IgG2b. Analysis of specific antibody responses showed that T cell-independent responses were normal but T cell-dependent responses were markedly reduced. Germinal centres were normal in size and number. In vitro purified B cells from Parp-1−/− mice proliferated normally and showed normal IgM secretion, decreased switching to IgG2a but increased IgA secretion. Collectively our results demonstrate that Parp-1 has essential roles in normal T cell-dependent antibody responses and the regulation of isotype expression. We speculate that Parp-1 forms a component of the protein complex involved in resolving the DNA double-strand breaks that occur during class switch recombination. PMID:18778284

Multi-scale computational study of the mechanical regulation of cell mitotic rounding in epithelia

PubMed Central

Xu, Zhiliang; Zartman, Jeremiah J.; Alber, Mark

2017-01-01

Mitotic rounding during cell division is critical for preventing daughter cells from inheriting an abnormal number of chromosomes, a condition that occurs frequently in cancer cells. Cells must significantly expand their apical area and transition from a polygonal to circular apical shape to achieve robust mitotic rounding in epithelial tissues, which is where most cancers initiate. However, how cells mechanically regulate robust mitotic rounding within packed tissues is unknown. Here, we analyze mitotic rounding using a newly developed multi-scale subcellular element computational model that is calibrated using experimental data. Novel biologically relevant features of the model include separate representations of the sub-cellular components including the apical membrane and cytoplasm of the cell at the tissue scale level as well as detailed description of cell properties during mitotic rounding. Regression analysis of predictive model simulation results reveals the relative contributions of osmotic pressure, cell-cell adhesion and cortical stiffness to mitotic rounding. Mitotic area expansion is largely driven by regulation of cytoplasmic pressure. Surprisingly, mitotic shape roundness within physiological ranges is most sensitive to variation in cell-cell adhesivity and stiffness. An understanding of how perturbed mechanical properties impact mitotic rounding has important potential implications on, amongst others, how tumors progressively become more genetically unstable due to increased chromosomal aneuploidy and more aggressive. PMID:28531187

Endothelial basement membrane limits tip cell formation by inducing Dll4/Notch signalling in vivo.

PubMed

Stenzel, Denise; Franco, Claudio A; Estrach, Soline; Mettouchi, Amel; Sauvaget, Dominique; Rosewell, Ian; Schertel, Andreas; Armer, Hannah; Domogatskaya, Anna; Rodin, Sergey; Tryggvason, Karl; Collinson, Lucy; Sorokin, Lydia; Gerhardt, Holger

2011-10-28

How individual components of the vascular basement membrane influence endothelial cell behaviour remains unclear. Here we show that laminin α4 (Lama4) regulates tip cell numbers and vascular density by inducing endothelial Dll4/Notch signalling in vivo. Lama4 deficiency leads to reduced Dll4 expression, excessive filopodia and tip cell formation in the mouse retina, phenocopying the effects of Dll4/Notch inhibition. Lama4-mediated Dll4 expression requires a combination of integrins in vitro and integrin β1 in vivo. We conclude that appropriate laminin/integrin-induced signalling is necessary to induce physiologically functional levels of Dll4 expression and regulate branching frequency during sprouting angiogenesis in vivo.

Endothelial basement membrane limits tip cell formation by inducing Dll4/Notch signalling in vivo

PubMed Central

Stenzel, Denise; Franco, Claudio A; Estrach, Soline; Mettouchi, Amel; Sauvaget, Dominique; Rosewell, Ian; Schertel, Andreas; Armer, Hannah; Domogatskaya, Anna; Rodin, Sergey; Tryggvason, Karl; Collinson, Lucy; Sorokin, Lydia; Gerhardt, Holger

2011-01-01

How individual components of the vascular basement membrane influence endothelial cell behaviour remains unclear. Here we show that laminin α4 (Lama4) regulates tip cell numbers and vascular density by inducing endothelial Dll4/Notch signalling in vivo. Lama4 deficiency leads to reduced Dll4 expression, excessive filopodia and tip cell formation in the mouse retina, phenocopying the effects of Dll4/Notch inhibition. Lama4-mediated Dll4 expression requires a combination of integrins in vitro and integrin β1 in vivo. We conclude that appropriate laminin/integrin-induced signalling is necessary to induce physiologically functional levels of Dll4 expression and regulate branching frequency during sprouting angiogenesis in vivo. PMID:21979816

Poisons, ruffles and rockets: bacterial pathogens and the host cell cytoskeleton.

PubMed

Steele-Mortimer, O; Knodler, L A; Finlay, B B

2000-02-01

The cytoskeleton of eukaryotic cells is affected by a number of bacterial and viral pathogens. In this review we consider three recurring themes of cytoskeletal involvement in bacterial pathogenesis: 1) the effect of bacterial toxins on actin-regulating small GTP-binding proteins; 2) the invasion of non-phagocytic cells by the bacterial induction of ruffles at the plasma membrane; 3) the formation of actin tails and pedestals by intracellular and extracellular bacteria, respectively. Considerable progress has been made recently in the characterization of these processes. It is becoming clear that bacterial pathogens have developed a variety of sophisticated mechanisms for utilizing the complex cytoskeletal system of host cells. These bacterially-induced processes are now providing unique insights into the regulation of fundamental eukaryotic mechanisms.

Unusual Suspects in the Development of Obesity-Induced Inflammation and Insulin Resistance: NK cells, iNKT cells, and ILCs.

PubMed

Bonamichi, Beatriz Dal Santo Francisco; Lee, Jongsoon

2017-08-01

The notion that obesity-induced inflammation mediates the development of insulin resistance in animal models and humans has been gaining strong support. It has also been shown that immune cells in local tissues, in particular in visceral adipose tissue, play a major role in the regulation of obesity-induced inflammation. Specifically, obesity increases the numbers and activation of proinflammatory immune cells, including M1 macrophages, neutrophils, Th1 CD4 T cells, and CD8 T cells, while simultaneously suppressing anti-inflammatory cells such as M2 macrophages, CD4 regulatory T cells, regulatory B cells, and eosinophils. Recently, however, new cell types have been shown to participate in the development of obesity-induced inflammation and insulin resistance. Some of these cell types also appear to regulate obesity. These cells are natural killer (NK) cells and innate lymphoid cells (ILCs), which are closely related, and invariant natural killer T (iNKT) cells. It should be noted that, although iNKT cells resemble NK cells in name, they are actually a completely different cell type in terms of their development and functions in immunity and metabolism. In this review, we will focus on the roles that these relatively new players in the metabolism field play in obesity-induced insulin resistance and the regulation of obesity. Copyright © 2017 Korean Diabetes Association.

Production of large numbers of hybridomas producing monoclonal antibodies against rat IgE using mast cell-deficient w/wv and sl/sld strains of mice.

PubMed

Rup, B J

1989-08-15

A number of different mouse strains and immunization protocols were used to attempt to make monoclonal antibodies against rat IgE for use in studies of the structure, biological activities and regulation of this class of antibody. Successful production of large numbers of monoclonal antibodies was achieved when mast cell deficient (w/wv and sl/sld) but not conventional (BALB/c, CAF1 or SJL) mice were used. These results suggest that the poor response of conventional strains of mice to rat IgE may be due to the presence of mast cells bearing high affinity receptors for IgE in these mice.

The Transcription Factor Bright Plays a Role in Marginal Zone B Lymphocyte Development and Autoantibody Production

PubMed Central

Oldham, Athenia L.; Miner, Cathrine A.; Wang, Hong-Cheng; Webb, Carol F.

2011-01-01

Previous data suggested that constitutive expression of the transcription factor Bright (B cell regulator of immunoglobulin heavy chain transcription), normally tightly regulated during B cell differentiation, was associated with autoantibody production. Here we show that constitutive Bright expression results in skewing of mature B lineage subpopulations toward marginal zone cells at the expense of the follicular subpopulation. C57Bl/6 transgenic mice constitutively expressing Bright in B lineage cells generated autoantibodies that were not the result of global increases in immunoglobulin or of breaches in key tolerance checkpoints typically defective in other autoimmune mouse models. Rather, autoimmunity correlated with increased numbers of marginal zone B cells and alterations in the phenotype and gene expression profiles of lymphocytes within the follicular B cell compartment. These data suggest a novel role for Bright in the normal development of mature B cell subsets and in autoantibody production. PMID:21963220

MATHEMATICAL ANALYSIS OF STEADY-STATE SOLUTIONS IN COMPARTMENT AND CONTINUUM MODELS OF CELL POLARIZATION

PubMed Central

ZHENG, ZHENZHEN; CHOU, CHING-SHAN; YI, TAU-MU; NIE, QING

2013-01-01

Cell polarization, in which substances previously uniformly distributed become asymmetric due to external or/and internal stimulation, is a fundamental process underlying cell mobility, cell division, and other polarized functions. The yeast cell S. cerevisiae has been a model system to study cell polarization. During mating, yeast cells sense shallow external spatial gradients and respond by creating steeper internal gradients of protein aligned with the external cue. The complex spatial dynamics during yeast mating polarization consists of positive feedback, degradation, global negative feedback control, and cooperative effects in protein synthesis. Understanding such complex regulations and interactions is critical to studying many important characteristics in cell polarization including signal amplification, tracking dynamic signals, and potential trade-off between achieving both objectives in a robust fashion. In this paper, we study some of these questions by analyzing several models with different spatial complexity: two compartments, three compartments, and continuum in space. The step-wise approach allows detailed characterization of properties of the steady state of the system, providing more insights for biological regulations during cell polarization. For cases without membrane diffusion, our study reveals that increasing the number of spatial compartments results in an increase in the number of steady-state solutions, in particular, the number of stable steady-state solutions, with the continuum models possessing infinitely many steady-state solutions. Through both analysis and simulations, we find that stronger positive feedback, reduced diffusion, and a shallower ligand gradient all result in more steady-state solutions, although most of these are not optimally aligned with the gradient. We explore in the different settings the relationship between the number of steady-state solutions and the extent and accuracy of the polarization. Taken together these results furnish a detailed description of the factors that influence the tradeoff between a single correctly aligned but poorly polarized stable steady-state solution versus multiple more highly polarized stable steady-state solutions that may be incorrectly aligned with the external gradient. PMID:21936604

Cell motility and ECM proteolysis regulate tumor growth and tumor relapse by altering the fraction of cancer stem cells and their spatial scattering

NASA Astrophysics Data System (ADS)

Kumar, Sandeep; Kulkarni, Rahul; Sen, Shamik

2016-06-01

Tumors consist of multiple cell sub-populations including cancer stem cells (CSCs), transiently amplifying cells and terminally differentiated cells (TDCs), with the CSC fraction dictating the aggressiveness of the tumor and drug sensitivity. In epithelial cancers, tumor growth is influenced greatly by properties of the extracellular matrix (ECM), with cancer progression associated with an increase in ECM density. However, the extent to which increased ECM confinement induced by an increase in ECM density influences tumor growth and post treatment relapse dynamics remains incompletely understood. In this study, we use a cellular automata-based discrete modeling approach to study the collective influence of ECM density, cell motility and ECM proteolysis on tumor growth, tumor heterogeneity, and tumor relapse after drug treatment. We show that while increased confinement suppresses tumor growth and the spatial scattering of CSCs, this effect can be reversed when cells become more motile and proteolytically active. Our results further suggest that, in addition to the absolute number of CSCs, their spatial positioning also plays an important role in driving tumor growth. In a nutshell, our study suggests that, in confined environments, cell motility and ECM proteolysis are two key factors that regulate tumor growth and tumor relapse dynamics by altering the number and spatial distribution of CSCs.

Parallel RNAi screens across different cell lines identify generic and cell type-specific regulators of actin organization and cell morphology.

PubMed

Liu, Tao; Sims, David; Baum, Buzz

2009-01-01

In recent years RNAi screening has proven a powerful tool for dissecting gene functions in animal cells in culture. However, to date, most RNAi screens have been performed in a single cell line, and results then extrapolated across cell types and systems. Here, to dissect generic and cell type-specific mechanisms underlying cell morphology, we have performed identical kinome RNAi screens in six different Drosophila cell lines, derived from two distinct tissues of origin. This analysis identified a core set of kinases required for normal cell morphology in all lines tested, together with a number of kinases with cell type-specific functions. Most significantly, the screen identified a role for minibrain (mnb/DYRK1A), a kinase associated with Down's syndrome, in the regulation of actin-based protrusions in CNS-derived cell lines. This cell type-specific requirement was not due to the peculiarities in the morphology of CNS-derived cells and could not be attributed to differences in mnb expression. Instead, it likely reflects differences in gene expression that constitute the cell type-specific functional context in which mnb/DYRK1A acts. Using parallel RNAi screens and gene expression analyses across cell types we have identified generic and cell type-specific regulators of cell morphology, which include mnb/DYRK1A in the regulation of protrusion morphology in CNS-derived cell lines. This analysis reveals the importance of using different cell types to gain a thorough understanding of gene function across the genome and, in the case of kinases, the difficulties of using the differential gene expression to predict function.

Microparticles released by vascular endothelial cells increase hypoxia inducible factor expression in human proximal tubular HK-2 cells.

PubMed

Fernandez-Martínez, Ana Belen; Torija, Ana Valdehita; Carracedo, Julia; Ramirez, Rafael; de Lucio-Cazaña, Francisco Javier

2014-08-01

Microparticles are produced by vesiculation of the cell plasma membrane and serve as vectors of cell-to-cell communication. Co-culture experiments have shown that hypoxia-inducible factor-α (HIF-α)-regulated-genes are up-regulated in human renal proximal tubular HK-2 cells by endothelial cell factors which might be transported inside endothelial microparticles (EMP). Here we aimed to study in HK-2 cells the effect of EMP, produced by activated endothelial cells, on HIF-α and HIF-α-regulated vascular endothelial growth factor-A (VEGF-A). EMP, at a concentration much lower than that found in plasma, increased the expression of HIF-α/VEGF-A in a COX-2/EP2 receptor dependent manner. Since the EMP/cells ratio was ∼1/1000, we hypothesized that paracrine mediators produced by HK-2 cells amplified the initial signal. This hypothesis was confirmed by two facts which also suggested that the mediators were conveyed by particles released by HK-2 cells: (i) HIF-α was up-regulated in HK-2 cells treated with the pellet obtained from the conditioned medium of the EMP-treated HK-2 cells. (ii) In transwell experiments, EMP-treated cells increased the expression of HIF-α in untreated HK-2 cells. Interestingly, we detected these cells, particles that were released by EMP-treated HK-2 cells. Depending on the pathological context, activation of HIF-α and VEGF-A signaling in renal tissue/cells may have either beneficial or harmful effects. Therefore, our results suggest that their presence in the urinary space of EMP produced by activated endothelial cells may influence the outcome of a number of renal diseases. Copyright © 2014 Elsevier Ltd. All rights reserved.

Advances and challenges in logical modeling of cell cycle regulation: perspective for multi-scale, integrative yeast cell models

PubMed Central

Todd, Robert G.; van der Zee, Lucas

2016-01-01

Abstract The eukaryotic cell cycle is robustly designed, with interacting molecules organized within a definite topology that ensures temporal precision of its phase transitions. Its underlying dynamics are regulated by molecular switches, for which remarkable insights have been provided by genetic and molecular biology efforts. In a number of cases, this information has been made predictive, through computational models. These models have allowed for the identification of novel molecular mechanisms, later validated experimentally. Logical modeling represents one of the youngest approaches to address cell cycle regulation. We summarize the advances that this type of modeling has achieved to reproduce and predict cell cycle dynamics. Furthermore, we present the challenge that this type of modeling is now ready to tackle: its integration with intracellular networks, and its formalisms, to understand crosstalks underlying systems level properties, ultimate aim of multi-scale models. Specifically, we discuss and illustrate how such an integration may be realized, by integrating a minimal logical model of the cell cycle with a metabolic network. PMID:27993914

Neutral buoyancy and sleep-deprived serum factors alter expression of cytokines regulating osteogenesis

NASA Astrophysics Data System (ADS)

Gorczynski, Reginald M.; Gorczynski, Christopher P.; Gorczynski, Laura Y.; Hu, Jiang; Lu, Jin; Manuel, Justin; Lee, Lydia

2005-05-01

We examined expression of genes associated with cytokine production, and genes implicated in regulating bone metabolism, in bone stromal and osteoblast cells incubated under standard ground conditions and under conditions of neutral buoyancy, and in the presence/absence of serum from normal or sleep-deprived mice. We observed a clear interaction between these two conditions (exposure to neutral buoyancy and serum stimulation) in promoting enhanced osteoclastogenesis. Both conditions independently altered expression of a number of cytokines implicated in the regulation of bone metabolism. However, using stromal cells from IL-1 and TNF α cytokine r KO mice, we concluded that the increased bone loss under microgravity conditions was not primarily cytokine mediated.

Stem cell research and therapies in Argentina: the legal and regulatory approach.

PubMed

de Arzuaga, Fabiana C

2013-12-01

Argentina has a significant number of researchers in public and private institutions conducting research in regenerative medicine and stem cells. There is not specific legislation in this area; however, the National Ministry of Health has issued regulations under the scope of the Transplant Act and the Medicines Act. Alongside the groups doing research, it is possible to find professionals offering experimental stem cell therapies to patients. These professionals take refuge in the term "medical practice" and sell experimental treatment to patients with no guarantee of safety and security given that they were not tested in clinical research. These practices offered to patients in a scheme, apparently legal, are generating an important number of judicial actions requesting the payment of said treatments. The decisions of the courts ordering payment in most cases are generating a transfer of funds from patients, social welfare systems, and the state to medical centers offering stem cell experimental therapies. This article describes the current regulations as well as the course of action to solve the emerging problems of these new technologies at legislative level.

Foxg1 regulates retinal axon pathfinding by repressing an ipsilateral program in nasal retina and by causing optic chiasm cells to exert a net axonal growth-promoting activity.

PubMed

Tian, Natasha M; Pratt, Thomas; Price, David J

2008-12-01

Mammalian binocular vision relies on the divergence of retinal ganglion cell axons at the optic chiasm, with strictly controlled numbers projecting contralaterally and ipsilaterally. In mouse, contralateral projections arise from the entire retina, whereas ipsilateral projections arise from ventrotemporal retina. We investigate how development of these patterns of projection is regulated by the contralateral determinant Foxg1, a forkhead box transcription factor expressed in nasal retina and at the chiasm. In nasal retina, loss of Foxg1 causes increased numbers of ipsilateral projections and ectopic expression of the ipsilateral determinants Zic2, Ephb1 and Foxd1, indicating that nasal retina is competent to express an ipsilateral program that is normally suppressed by Foxg1. Using co-cultures that combine Foxg1-expressing with Foxg1-null retinal explants and chiasm cells, we provide functional evidence that Foxg1 promotes contralateral projections through actions in nasal retina, and that in chiasm cells, Foxg1 is required for the generation of a hitherto unrecognized activity supporting RGC axon growth.

DUBbing Cancer: Deubiquitylating Enzymes Involved in Epigenetics, DNA Damage and the Cell Cycle As Therapeutic Targets.

PubMed

Pinto-Fernandez, Adan; Kessler, Benedikt M

2016-01-01

Controlling cell proliferation is one of the hallmarks of cancer. A number of critical checkpoints ascertain progression through the different stages of the cell cycle, which can be aborted when perturbed, for instance by errors in DNA replication and repair. These molecular checkpoints are regulated by a number of proteins that need to be present at the right time and quantity. The ubiquitin system has emerged as a central player controlling the fate and function of such molecules such as cyclins, oncogenes and components of the DNA repair machinery. In particular, proteases that cleave ubiquitin chains, referred to as deubiquitylating enzymes (DUBs), have attracted recent attention due to their accessibility to modulation by small molecules. In this review, we describe recent evidence of the critical role of DUBs in aspects of cell cycle checkpoint control, associated DNA repair mechanisms and regulation of transcription, representing pathways altered in cancer. Therefore, DUBs involved in these processes emerge as potentially critical targets for the treatment of not only hematological, but potentially also solid tumors.

DUBbing Cancer: Deubiquitylating Enzymes Involved in Epigenetics, DNA Damage and the Cell Cycle As Therapeutic Targets

PubMed Central

Pinto-Fernandez, Adan; Kessler, Benedikt M.

2016-01-01

Controlling cell proliferation is one of the hallmarks of cancer. A number of critical checkpoints ascertain progression through the different stages of the cell cycle, which can be aborted when perturbed, for instance by errors in DNA replication and repair. These molecular checkpoints are regulated by a number of proteins that need to be present at the right time and quantity. The ubiquitin system has emerged as a central player controlling the fate and function of such molecules such as cyclins, oncogenes and components of the DNA repair machinery. In particular, proteases that cleave ubiquitin chains, referred to as deubiquitylating enzymes (DUBs), have attracted recent attention due to their accessibility to modulation by small molecules. In this review, we describe recent evidence of the critical role of DUBs in aspects of cell cycle checkpoint control, associated DNA repair mechanisms and regulation of transcription, representing pathways altered in cancer. Therefore, DUBs involved in these processes emerge as potentially critical targets for the treatment of not only hematological, but potentially also solid tumors. PMID:27516771

A diffusible signal derived from hematopoietic cells supports the survival and proliferation of regenerative cells during zebrafish fin fold regeneration.

PubMed

Hasegawa, Tomoya; Nakajima, Teruhiro; Ishida, Takashi; Kudo, Akira; Kawakami, Atsushi

2015-03-01

Multicellular organisms maintain body integrity by constantly regenerating tissues throughout their lives; however, the overall mechanism for regulating regeneration remains an open question. Studies of limb and fin regeneration in teleost fish and urodeles have shown the involvement of a number of locally activated signals at the wounded site during regeneration. Here, we demonstrate that a diffusible signal from a distance also play an essential role for regeneration. Among a number of zebrafish mutants, we found that the zebrafish cloche (clo) and tal1 mutants, which lack most hematopoietic tissues, displayed a unique regeneration defect accompanying apoptosis in primed regenerative tissue. Our analyses of the mutants showed that the cells in the primed regenerative tissue are susceptible to apoptosis, but their survival is normally supported by the presence of hematopoietic tissues, mainly the myeloid cells. We further showed that a diffusible factor in the wild-type body fluid mediates this signal. Thus, our study revealed a novel mechanism that the hematopoietic tissues regulate tissue regeneration through a diffusible signal. Copyright © 2014 Elsevier Inc. All rights reserved.

Involvement of Heterogeneous Ribonucleoprotein F in the Regulation of Cell Proliferation via the Mammalian Target of Rapamycin/S6 Kinase 2 Pathway

PubMed Central

Goh, Eddy T. H.; Pardo, Olivier E.; Michael, Nicholas; Niewiarowski, Andrew; Totty, Nick; Volkova, Dariya; Tsaneva, Irina R.; Seckl, Michael J.; Gout, Ivan

2010-01-01

The S6 kinases (S6Ks) have been linked to a number of cellular processes, including translation, insulin metabolism, cell survival, and RNA splicing. Signaling via the phosphotidylinositol 3-kinase and mammalian target of rapamycin (mTOR) pathways is critical in regulating the activity and subcellular localization of S6Ks. To date, nuclear functions of both S6K isoforms, S6K1 and S6K2, are not well understood. To better understand S6K nuclear roles, we employed affinity purification of S6Ks from nuclear preparations followed by mass spectrometry analysis for the identification of novel binding partners. In this study, we report that in contrast to S6K1, the S6K2 isoform specifically associates with a number of RNA-binding proteins, including heterogeneous ribonucleoproteins (hnRNPs). We focused on studying the mechanism and physiological relevance of the S6K2 interaction with hnRNP F/H. Interestingly, the S6K2-hnRNP F/H interaction was not affected by mitogenic stimulation, whereas mTOR binding to hnRNP F/H was induced by serum stimulation. In addition, we define a new role of hnRNP F in driving cell proliferation, which could be partially attenuated by rapamycin treatment. S6K2-driven cell proliferation, on the other hand, could be blocked by small interfering RNA-mediated down-regulation of hnRNP F. These results demonstrate that the specific interaction between mTOR and S6K2 with hnRNPs is implicated in the regulation of cell proliferation. PMID:20308064

Regulation of cytokine signaling by B cell antigen receptor and CD40-controlled expression of heparan sulfate proteoglycans.

PubMed

van der Voort, R; Keehnen, R M; Beuling, E A; Spaargaren, M; Pals, S T

2000-10-16

Recently, biochemical, cell biological, and genetic studies have converged to reveal that integral membrane heparan sulfate proteoglycans (HSPGs) are critical regulators of growth and differentiation of epithelial and connective tissues. As a large number of cytokines involved in lymphoid tissue homeostasis or inflammation contain potential HS-binding domains, HSPGs presumably also play important roles in the regulation of the immune response. In this report, we explored the expression, regulation, and function of HSPGs on B lymphocytes. We demonstrate that activation of the B cell antigen receptor (BCR) and/or CD40 induces a strong transient expression of HSPGs on human tonsillar B cells. By means of these HSPGs, the activated B cells can bind hepatocyte growth factor (HGF), a cytokine that regulates integrin-mediated B cell adhesion and migration. This interaction with HGF is highly selective since the HSPGs did not bind the chemokine stromal cell-derived factor (SDF)-1 alpha, even though the affinities of HGF and SDF-1alpha for heparin are similar. On the activated B cells, we observed induction of a specific HSPG isoform of CD44 (CD44-HS), but not of other HSPGs such as syndecans or glypican-1. Interestingly, the expression of CD44-HS on B cells strongly promotes HGF-induced signaling, resulting in an HS-dependent enhanced phosphorylation of Met, the receptor tyrosine kinase for HGF, as well as downstream signaling molecules including Grb2-associated binder 1 (Gab1) and Akt/protein kinase B (PKB). Our results demonstrate that the BCR and CD40 control the expression of HSPGs, specifically CD44-HS. These HSPGs act as functional coreceptors that selectively promote cytokine signaling in B cells, suggesting a dynamic role for HSPGs in antigen-specific B cell differentiation.

Indoleamine 2,3-dioxygenase and regulatory t cells in intestinal mucosa in children with inflammatory bowel disease.

PubMed

Sznurkowska, K; Żawrocki, A; Sznurkowski, J; Iżycka-Świeszewska, E; Landowski, P; Szlagatys-Sidorkiewicz, A; Plata-Nazar, K; Kamińska, B

2017-01-01

Impaired immune regulation has been suggested as an underlying mechanism of inflammatory bowel disease. Indoleamine 2,3-dioxygenase (IDO) and regulatory T cells expressing FOXP3 are crucial elements of immune regulation. Conversion of FOXP3- lymphocytes to Tregs is one of the functions of IDO. The aim of this study was to evaluate the number of cells expressing FOXP3 and IDO in the lamina propria of intestinal mucosa and to evaluate correlations between these parameters and disease activity. Sixty-six children newly diagnosed with inflammatory bowel disease (41 patients with ulcerative colitis and 25 patients with Crohn’s disease) were included in the study. Clinical activity of the disease was assessed by the Pediatric Ulcerative Colitis Activity Index and the Pediatric Crohn’s Disease Activity Index. Histopathological activity was scored according to the system described by Geboes. The infiltration of FOXP3+ and IDO+ cells was evaluated by immunohistochemistry. Sixteen patients with a diagnosis of irritable bowel syndrome (IBS) served as a control group. Lamina propria demonstrated a significantly higher infiltration of FOXP3+ and IDO+ cells in inflammatory bowel disease compared to the control group (p=0.001, p=0.004, respectively). The number of IDO+ and FOXP3+ cells correlated with clinical and histopathologic activity of Crohn’s disease. A positive correlation between the number of IDO+ and FOXP3+ cells was found in both types of inflammatory disease but not in patients with IBS. We conclude that indoleamine dioxygenase and FOXP3+ cells are upregulated in the intestinal mucosa of children with inflammatory bowel disease. IDO mediated conversion of FOXP3 -T cells to Tregs predominantly occurs in inflammation.

In Vitro Cardiomyogenic Potential of Human Amniotic Fluid Stem Cells

PubMed Central

Guan, Xuan; Delo, Dawn M.; Atala, Anthony; Soker, Shay

2010-01-01

Stem cell therapy for damaged cardiac tissue is currently limited by a number of factors, including the inability to obtain sufficient cell numbers, the potential tumorigenicity of certain types of stem cells, and the possible link between stem cell therapy and the development of malignant arrhythmias. In this study, we investigated whether human amniotic fluid-derived stem (hAFS) cells could be a potential source of cells for cardiac cell therapy by testing the in vitro differentiation capabilities. Undifferentiated hAFS cells express several cardiac genes, including the transcription factor mef2, the gap junction connexin43, and H- and N-cadherin. A 24-hour incubation with 5-aza-2′–deoxycytidine (5-AZA-dC) induced hAFS cell differentiation along the cardiac lineage. Evidence for this differentiation included morphological changes, up-regulation of cardiac-specific genes (cardiac troponin I and cardiac troponin T) and redistribution of connexin43, as well as down-regulation of the stem cell marker SRY-box 2 (sox2). When co-cultured with neonatal rat cardiomyocytes (NRCs), hAFS cells formed both mechanical and electrical connections with the NRCs. Dye transfer experiments showed that calcein dye could be transferred from NRCs to hAFS cells through cellular connections. The gap junction connexin 43 likely involved in the communication between the two cell types, because 12-O-Tetradecanoylphorbol 13-acetate (TPA) could partially block cellular crosstalk. We conclude that hAFS cells can be differentiated into a cardiomyocyte-like phenotype and can establish functional communication with NRCs. Thus, hAFS cells may potentially be used for cardiac cell therapy. PMID:20687122

Regulation of Fanconi anemia protein FANCD2 monoubiquitination by miR-302

DOE Office of Scientific and Technical Information (OSTI.GOV)

Suresh, Bharathi; College of Medicine, Hanyang University, Seoul; Kumar, A. Madhan

2015-10-16

Fanconi anemia (FA) is a recessively inherited multigene disease characterized by congenital defects, progressive bone marrow failure, and heightened cancer susceptibility. Monoubiquitination of the FA pathway member FANCD2 contributes to the repair of replication stalling DNA lesions. However, cellular regulation of FANCD2 monoubiquitination remains poorly understood. In the present study, we identified the miR-302 cluster as a potential regulator of FANCD2 by bioinformatics analysis. MicroRNAs (miRNAs) are the major posttranscriptional regulators of a wide variety of biological processes, and have been implicated in a number of diseases. Expression of the exogenous miR-302 cluster (without miR-367) reduced FANCD2 monoubiquitination and nuclearmore » foci formation. Furthermore, miR-302 cells showed extensive chromosomal breakage upon MMC treatment when compared to mock control cells. Taken together, our results suggest that overexpression of miR-302 plays a critical role in the regulation of FANCD2 monoubiquitination, resulting in characteristic defects in DNA repair within cells. - Highlights: • miR-302 binds to the 3′UTR promoter of the FANCD2 gene to regulate gene expression. • miR-302 cluster down-regulates FANCD2 protein expression. • miR-302 cluster reduces FANCD2 monoubiquitination and nuclear foci formation. • miR-302 exhibits the characteristic defects in DNA repair in cells.« less

Defective interleukin-4/Stat6 activity correlates with increased constitutive expression of negative regulators SOCS-3, SOCS-7, and CISH in colon cancer cells.

PubMed

Liu, Xiao Hong; Xu, Shuang Bing; Yuan, Jia; Li, Ben Hui; Zhang, Yan; Yuan, Qin; Li, Pin Dong; Li, Feng; Zhang, Wen Jie

2009-12-01

Interleukin-4 (IL-4)-induced Stat6 activities (phenotypes) vary among human cancer cells, of which the HT-29 cell line carries an active Stat6(high) phenotype, while Caco-2 carries a defective Stat6(null) phenotype, respectively. Cancer cells with Stat6(high) show resistance to apoptosis and exaggerated metastasis, suggesting the clinical significance of Stat6 phenotypes. We previously showed that Stat6(high) HT-29 cells exhibited low constitutive expression of Stat6-negative regulators SOCS-1 and SHP-1 because of gene hypermethylation. This study further examined the constitutive expression of other closely related SOCS family numbers including SOCS-3, SOCS-5, SOCS-7, and CISH using RT-PCR. Similar to SOCS-1 and SHP-1, Stat6(high) HT-29 cells expressed low constitutive mRNA of SOCS-3, SOCS-7, and CISH than Stat6(null) Caco-2 cells. Interestingly, DNA demethylation using 5-aza-2'-deoxycytidine in HT-29 cells up-regulated mRNA expression of the above genes, indicating a hypermethylation status, which was confirmed by methylation-specific sequencing in selected SOCS-3 gene. Furthermore, defective Stat6(null) Caco-2 exhibited impaired phosphorylation of Stat6 after IL-4 stimulation by flow cytometry, in keeping with the notion of an over-performed negative regulation. The findings that IL-4/Stat6 phenotypes show differential expression of multiple negative regulators suggest a model that a collective force of powerful negative regulators, directly and indirectly, acts on Stat6 activation, which may result in differential Stat6 phenotypes.

Ontogenesis of muscle and adipose tissues and their interactions in ruminants and other species.

PubMed

Bonnet, M; Cassar-Malek, I; Chilliard, Y; Picard, B

2010-07-01

The lean-to-fat ratio, that is, the relative masses of muscle and adipose tissue, is a criterion for the yield and quality of bovine carcasses and meat. This review describes the interactions between muscle and adipose tissue (AT) that may regulate the dynamic balance between the number and size of muscle v. adipose cells. Muscle and adipose tissue in cattle grow by an increase in the number of cells (hyperplasia), mainly during foetal life. The total number of muscle fibres is set by the end of the second trimester of gestation. By contrast, the number of adipocytes is never set. Number of adipocytes increases mainly before birth until 1 year of age, depending on the anatomical location of the adipose tissue. Hyperplasia concerns brown pre-adipocytes during foetal life and white pre-adipocytes from a few weeks after birth. A decrease in the number of secondary myofibres and an increase in adiposity in lambs born from mothers severely underfed during early pregnancy suggest a balance in the commitment of a common progenitor into the myogenic or adipogenic lineages, or a reciprocal regulation of the commitment of two distinct progenitors. The developmental origin of white adipocytes is a subject of debate. Molecular and histological data suggested a possible transdifferentiation of brown into white adipocytes, but this hypothesis has now been challenged by the characterization of distinct precursor cells for brown and white adipocytes in mice. Increased nutrient storage in fully differentiated muscle fibres and adipocytes, resulting in cell enlargement (hypertrophy), is thought to be the main mechanism, whereby muscle and fat masses increase in growing cattle. Competition or prioritization between adipose and muscle cells for the uptake and metabolism of nutrients is suggested, besides the successive waves of growth of muscle v. adipose tissue, by the inhibited or delayed adipose tissue growth in bovine genotypes exhibiting strong muscular development. This competition or prioritization occurs through cellular signalling pathways and the secretion of proteins by adipose tissue (adipokines) and muscle (myokines), putatively regulating their hypertrophy in a reciprocal manner. Further work on the mechanisms underlying cross-talk between brown or white adipocytes and muscle fibres will help to achieve better understanding as a prerequisite to improving the control of body growth and composition in cattle.

Physalis floridana Cell Number Regulator1 encodes a cell membrane-anchored modulator of cell cycle and negatively controls fruit size

PubMed Central

Li, Zhichao; He, Chaoying

2015-01-01

Physalis species show a significant variation in berry size; however, the underlying molecular basis is unknown. In this work, we showed that cell division difference in the ovaries might contribute to the ultimate berry size variation within Physalis species, and that mRNA abundance of Physalis floridana Cell Number Regulator1 (PfCNR1), the putative orthologue of the tomato fruit weight 2.2 (FW2.2), was negatively correlated with cell division in the ovaries. Moreover, heterochronic expression variation of the PfCNR1 genes in the ovaries concomitantly correlated with berry weight variation within Physalis species. In transgenic Physalis, multiple organ sizes could be negatively controlled by altering PfCNR1 levels, and cell division instead of cell expansion was primarily affected. PfCNR1 was shown to be anchored in the plasma membrane and to interact with PfAG2 (an AGAMOUS-like protein determining ovary identity). The expression of PfCYCD2;1, a putative orthologue of the mitosis-specific gene CyclinD2;1 in the cell cycle was negatively correlated with the PfCNR1 mRNA levels. PfAG2 was found to selectively bind to the CArG-box in the PfCYCD2;1 promoter and to repress PfCYCD2;1 expression, thus suggesting a PfAG2-mediated pathway for PfCNR1 to regulate cell division. The interaction of PfCNR1 with PfAG2 enhanced the repression of PfCYCD2;1 expression. The nuclear import of PfAG2 was essential in the proposed pathway. Our data provide new insights into the developmental pathways of a cell membrane-anchored protein that modulates cell division and governs organ size determination. This study also sheds light on the link between organ identity and organ growth in plants. PMID:25305759

FGF signaling supports Drosophila fertility by regulating development of ovarian muscle tissues

PubMed Central

Irizarry, Jihyun; Stathopoulos, Angelike

2015-01-01

The thisbe (ths) gene encodes a Drosophila fibroblast growth factor (FGF), and mutant females are viable but sterile suggesting a link between FGF signaling and fertility. Ovaries exhibit abnormal morphology including lack of epithelial sheaths, muscle tissues that surround ovarioles. Here we investigated how FGF influences Drosophila ovary morphogenesis and identified several roles. Heartless (Htl) FGF receptor was found expressed within somatic cells at the larval and pupal stages, and phenotypes were uncovered using RNAi. Differentiation of terminal filament cells was affected, but this effect did not alter ovariole number. In addition, proliferation of epithelial sheath progenitors, the apical cells, was decreased in both htl and ths mutants, while ectopic expression of the Ths ligand led to these cells’ over-proliferation suggesting that FGF signaling supports ovarian muscle sheath formation by controlling apical cell number in the developing gonad. Additionally, live imaging of adult ovaries was used to show that htl RNAi mutants, hypomorphic mutants in which epithelial sheaths are present, exhibit abnormal muscle contractions. Collectively, our results demonstrate that proper formation of ovarian muscle tissues is regulated by FGF signaling in the larval and pupal stages through control of apical cell proliferation and is required to support fertility. PMID:25958090

Loss of β-arrestin 2 exacerbates experimental autoimmune encephalomyelitis with reduced number of Foxp3+ CD4+ regulatory T cells

PubMed Central

Zhang, Yu; Liu, Chang; Wei, Bin; Pei, Gang

2013-01-01

β-Arrestins are well-known regulators and mediators of G protein-coupled receptor signalling, and accumulating evidence reveals that they are functionally involved in inflammation and autoimmune diseases. Of the two β-arrestins, β-arrestin 1 is documented to play regulatory roles in an animal model of multiple sclerosis (MS), whereas the role of β-arrestin 2 is less clear. Here, we show that β-arrestin 2-deficient mice displayed the exacerbated and sustained symptoms of experimental autoimmune encephalomyelitis (EAE), an animal model of MS. At the cellular level, deficiency of β-arrestin 2 led to a decreased number of Foxp3+ CD4+ regulatory T (Treg) cells in peripheral lymphoid organs of EAE mice. Consistently, our in vitro observations also revealed that loss of β-arrestin 2 impaired the conversion of Foxp3− CD4+ T cells into Foxp3+ CD4+ inducible Treg cells. Taken together, our data suggest that β-arrestin 2 plays a regulatory role in MS, that is opposite to that of β-arrestin 1, in autoimmune diseases such as MS, which is at least partially through regulation of iTreg cell differentiation. PMID:23859136

Ectopic Expression of Capsicum-Specific Cell Wall Protein Capsicum annuum Senescence-Delaying 1 (CaSD1) Delays Senescence and Induces Trichome Formation in Nicotiana benthamiana

PubMed Central

Seo, Eunyoung; Yeom, Seon-In; Jo, SungHwan; Jeong, Heejin; Kang, Byoung-Cheorl; Choi, Doil

2012-01-01

Secreted proteins are known to have multiple roles in plant development, metabolism, and stress response. In a previous study to understand the roles of secreted proteins, Capsicum annuum secreted proteins (CaS) were isolated by yeast secretion trap. Among the secreted proteins, we further characterized Capsicum annuum senescence-delaying 1 (CaSD1), a gene encoding a novel secreted protein that is present only in the genus Capsicum. The deduced CaSD1 contains multiple repeats of the amino acid sequence KPPIHNHKPTDYDRS. Interestingly, the number of repeats varied among cultivars and species in the Capsicum genus. CaSD1 is constitutively expressed in roots, and Agrobacterium-mediated transient overexpression of CaSD1 in Nicotiana benthamiana leaves resulted in delayed senescence with a dramatically increased number of trichomes and enlarged epidermal cells. Furthermore, senescence- and cell division-related genes were differentially regulated by CaSD1-overexpressing plants. These observations imply that the pepper-specific cell wall protein CaSD1 plays roles in plant growth and development by regulating cell division and differentiation. PMID:22441673

From single cells to tissue architecture-a bottom-up approach to modelling the spatio-temporal organisation of complex multi-cellular systems.

PubMed

Galle, J; Hoffmann, M; Aust, G

2009-01-01

Collective phenomena in multi-cellular assemblies can be approached on different levels of complexity. Here, we discuss a number of mathematical models which consider the dynamics of each individual cell, so-called agent-based or individual-based models (IBMs). As a special feature, these models allow to account for intracellular decision processes which are triggered by biomechanical cell-cell or cell-matrix interactions. We discuss their impact on the growth and homeostasis of multi-cellular systems as simulated by lattice-free models. Our results demonstrate that cell polarisation subsequent to cell-cell contact formation can be a source of stability in epithelial monolayers. Stroma contact-dependent regulation of tumour cell proliferation and migration is shown to result in invasion dynamics in accordance with the migrating cancer stem cell hypothesis. However, we demonstrate that different regulation mechanisms can equally well comply with present experimental results. Thus, we suggest a panel of experimental studies for the in-depth validation of the model assumptions.

Polymer microarray technology for stem cell engineering

PubMed Central

Coyle, Robert; Jia, Jia; Mei, Ying

2015-01-01

Stem cells hold remarkable promise for applications in tissue engineering and disease modeling. During the past decade, significant progress has been made in developing soluble factors (e.g., small molecules and growth factors) to direct stem cells into a desired phenotype. However, the current lack of suitable synthetic materials to regulate stem cell activity has limited the realization of the enormous potential of stem cells. This can be attributed to a large number of materials properties (e.g., chemical structures and physical properties of materials) that can affect stem cell fate. This makes it challenging to design biomaterials to direct stem cell behavior. To address this, polymer microarray technology has been developed to rapidly identify materials for a variety of stem cell applications. In this article, we summarize recent developments in polymer array technology and their applications in stem cell engineering. Statement of significance Stem cells hold remarkable promise for applications in tissue engineering and disease modeling. In the last decade, significant progress has been made in developing chemically defined media to direct stem cells into a desired phenotype. However, the current lack of the suitable synthetic materials to regulate stem cell activities has been limiting the realization of the potential of stem cells. This can be attributed to the number of variables in material properties (e.g., chemical structures and physical properties) that can affect stem cells. Polymer microarray technology has shown to be a powerful tool to rapidly identify materials for a variety of stem cell applications. Here we summarize recent developments in polymer array technology and their applications in stem cell engineering. PMID:26497624

Cyclin-dependent kinase 5 mediates adult OPC maturation and myelin repair through modulation of Akt and GsK-3β signaling.

PubMed

Luo, FuCheng; Burke, Kathryn; Kantor, Christopher; Miller, Robert H; Yang, Yan

2014-07-30

Failure of remyelination in diseases, such as multiple sclerosis (MS), leads to permanent axonal damage and irreversible functional loss. The mechanisms controlling remyelination are currently poorly understood. Recent studies implicate the cyclin-dependent kinase 5 (Cdk5) in regulating oligodendrocyte (OL) development and myelination in CNS. In this study, we show that Cdk5 is also an important regulator of remyelination. Pharmacological inhibition of Cdk5 inhibits repair of lysolecithin lesions. This inhibition is a consequence of Cdk5 disruption in neural cells because remyelination in slice cultures is blocked by Cdk5 inhibitors, whereas specific deletion of Cdk5 in OLs inhibits myelin repair. In CNP-Cre;Cdk5(fl/fl) conditional knock-out mouse (Cdk5 cKO), myelin repair was delayed significantly in response to focal demyelinating lesions compared with wild-type animals. The lack of myelin repair was reflected in decreased expression of MBP and proteolipid protein and a reduction in the total number of myelinated axons in the lesion. The number of CC1(+) cells in the lesion sites was significantly reduced in Cdk5 cKO compared with wild-type animals although the total number of oligodendrocyte lineage cells (Olig2(+) cells) was increased, suggesting that Cdk5 loss perturbs the transition of early OL lineage cell into mature OL and subsequent remyelination. The failure of remyelination in Cdk5 cKO animals was associated with a reduction in signaling through the Akt pathway and an enhancement of Gsk-3β signaling pathways. Together, these data suggest that Cdk5 is critical in regulating the transition of adult oligodendrocyte precursor cells to mature OLs that is essential for myelin repair in adult CNS. Copyright © 2014 the authors 0270-6474/14/3410415-15$15.00/0.

Genomic instability of osteosarcoma cell lines in culture: impact on the prediction of metastasis relevant genes.

PubMed

Muff, Roman; Rath, Prisni; Ram Kumar, Ram Mohan; Husmann, Knut; Born, Walter; Baudis, Michael; Fuchs, Bruno

2015-01-01

Osteosarcoma is a rare but highly malignant cancer of the bone. As a consequence, the number of established cell lines used for experimental in vitro and in vivo osteosarcoma research is limited and the value of these cell lines relies on their stability during culture. Here we investigated the stability in gene expression by microarray analysis and array genomic hybridization of three low metastatic cell lines and derivatives thereof with increased metastatic potential using cells of different passages. The osteosarcoma cell lines showed altered gene expression during in vitro culture, and it was more pronounced in two metastatic cell lines compared to the respective parental cells. Chromosomal instability contributed in part to the altered gene expression in SAOS and LM5 cells with low and high metastatic potential. To identify metastasis-relevant genes in a background of passage-dependent altered gene expression, genes involved in "Pathways in cancer" that were consistently regulated under all passage comparisons were evaluated. Genes belonging to "Hedgehog signaling pathway" and "Wnt signaling pathway" were significantly up-regulated, and IHH, WNT10B and TCF7 were found up-regulated in all three metastatic compared to the parental cell lines. Considerable instability during culture in terms of gene expression and chromosomal aberrations was observed in osteosarcoma cell lines. The use of cells from different passages and a search for genes consistently regulated in early and late passages allows the analysis of metastasis-relevant genes despite the observed instability in gene expression in osteosarcoma cell lines during culture.

The Cryptococcus neoformans Capsule: a Sword and a Shield

PubMed Central

O'Meara, Teresa R.

2012-01-01

Summary: The human fungal pathogen Cryptococcus neoformans is characterized by its ability to induce a distinct polysaccharide capsule in response to a number of host-specific environmental stimuli. The induction of capsule is a complex biological process encompassing regulation at multiple steps, including the biosynthesis, transport, and maintenance of the polysaccharide at the cell surface. By precisely regulating the composition of its cell surface and secreted polysaccharides, C. neoformans has developed intricate ways to establish chronic infection and dormancy in the human host. The plasticity of the capsule structure in response to various host conditions also underscores the complex relationship between host and parasite. Much of this precise regulation of capsule is achieved through the transcriptional responses of multiple conserved signaling pathways that have been coopted to regulate this C. neoformans-specific virulence-associated phenotype. This review focuses on specific host stimuli that trigger the activation of the signal transduction cascades and on the downstream transcriptional responses that are required for robust encapsulation around the cell. PMID:22763631

Decreased IL-10-producing regulatory B cells in patients with advanced mycosis fungoides.

PubMed

Akatsuka, Taro; Miyagaki, Tomomitsu; Nakajima, Rina; Kamijo, Hiroaki; Oka, Tomonori; Takahashi, Naomi; Suga, Hiraku; Yoshizaki, Ayumi; Asano, Yoshihide; Sugaya, Makoto; Sato, Shinichi

2018-06-28

Historically, B cells have been considered as positive regulators of humoral immune responses. Specific B-cell subsets, however, negatively regulate immune responses and are termed "regulatory B cells" (Bregs). Recently, Bregs have been linked to not only inflammatory and autoimmune diseases, but also malignancies via suppressing anti-tumour immunity. To investigate the involvement of Bregs in advanced mycosis fungoides (MF). The frequency of CD19 + CD24 hi CD27 + memory B cells and CD19 + CD24 hi CD38 hi transitional B cells (which enrich IL-10-producing Bregs) was examined in peripheral blood from patients with advanced MF (n = 11) and healthy controls (n = 9) by flow cytometry. The frequency of IL-10-producing Bregs was also measured by flow cytometry. The correlation between frequency or number of B-cell subsets and disease severity markers was also analysed. The frequency of CD19 + CD24 hi CD27 + B cells, CD19 + CD24 hi CD38 hi B cells, and IL-10-producing B cells was decreased in peripheral blood of advanced MF patients. The frequency and number of these B-cell subsets inversely correlated with serum soluble IL-2 receptor and serum lactate dehydrogenase levels. The development of IL-10-producing Bregs is impaired in patients with advanced MF and a decrease in IL-10-producing Bregs may play an important role in the progression of advanced MF.

p21 as a Transcriptional Co-Repressor of S-Phase and Mitotic Control Genes

PubMed Central

Ferrándiz, Nuria; Caraballo, Juan M.; García-Gutierrez, Lucía; Devgan, Vikram; Rodriguez-Paredes, Manuel; Lafita, M. Carmen; Bretones, Gabriel; Quintanilla, Andrea; Muñoz-Alonso, M. Jose; Blanco, Rosa; Reyes, Jose C.; Agell, Neus; Delgado, M. Dolores; Dotto, G. Paolo; León, Javier

2012-01-01

It has been previously described that p21 functions not only as a CDK inhibitor but also as a transcriptional co-repressor in some systems. To investigate the roles of p21 in transcriptional control, we studied the gene expression changes in two human cell systems. Using a human leukemia cell line (K562) with inducible p21 expression and human primary keratinocytes with adenoviral-mediated p21 expression, we carried out microarray-based gene expression profiling. We found that p21 rapidly and strongly repressed the mRNA levels of a number of genes involved in cell cycle and mitosis. One of the most strongly down-regulated genes was CCNE2 (cyclin E2 gene). Mutational analysis in K562 cells showed that the N-terminal region of p21 is required for repression of gene expression of CCNE2 and other genes. Chromatin immunoprecipitation assays indicated that p21 was bound to human CCNE2 and other p21-repressed genes gene in the vicinity of the transcription start site. Moreover, p21 repressed human CCNE2 promoter-luciferase constructs in K562 cells. Bioinformatic analysis revealed that the CDE motif is present in most of the promoters of the p21-regulated genes. Altogether, the results suggest that p21 exerts a repressive effect on a relevant number of genes controlling S phase and mitosis. Thus, p21 activity as inhibitor of cell cycle progression would be mediated not only by the inhibition of CDKs but also by the transcriptional down-regulation of key genes. PMID:22662213

Lactic Acid Bacteria Improves Peyer's Patch Cell-Mediated Immunoglobulin A and Tight-Junction Expression in a Destructed Gut Microbial Environment.

PubMed

Kim, Sung Hwan; Jeung, Woonhee; Choi, Il-Dong; Jeong, Ji-Woong; Lee, Dong Eun; Huh, Chul-Sung; Kim, Geun-Bae; Hong, Seong Soo; Shim, Jae-Jung; Lee, Jung Lyoul; Sim, Jae-Hun; Ahn, Young-Tae

2016-06-28

To evaluate the effects of lactic acid bacteria (LAB) on Peyer's patch cells, mice were treated with a high dose of kanamycin to disturb the gut microbial environment. The overarching goal was to explore the potential of LAB for use as a dietary probiotic that buffers the negative consequences of antibiotic treatment. In vitro, LAB stimulated the production of immunoglobulin A (IgA) from isolated Peyer's patch cells. Inflammation-related genes (TNF-α, IL-1β, and IL-8) were up-regulated in Caco-2 cells stimulated with lipopolysaccharide (LPS), while tight-junction-related genes (ZO-1 and occludin) were down-regulated; the effects of LPS on inflammatory gene and tight-junction gene expression were reversed by treatment with LAB. Mice treated with a high dose of kanamycin showed increased serum IgE levels and decreases in serum IgA and fecal IgA levels; the number of Peyer's patch cells decreased with kanamycin treatment. However, subsequent LAB treatment was effective in reducing the serum IgE level and recovering the serum IgA and fecal IgA levels, as well as the number of Peyer's patch cells. In addition, ZO-1 and occludin mRNA levels were up-regulated in the ileum tissues of mice receiving LAB treatment. Lactic acid bacteria can enhance the intestinal immune system by improving the integrity of the intestinal barrier and increasing the production of IgA in Peyer's patches. Lactic acid bacteria should be considered a potential probiotic candidate for improving intestinal immunity, particularly in mitigating the negative consequences of antibiotic use.

SN-38 Acts as a Radiosensitizer for Colorectal Cancer by Inhibiting the Radiation-induced Up-regulation of HIF-1α.

PubMed

Okuno, Takayuki; Kawai, Kazushige; Hata, Keisuke; Murono, Koji; Emoto, Shigenobu; Kaneko, Manabu; Sasaki, Kazuhito; Nishikawa, Takeshi; Tanaka, Toshiaki; Nozawa, Hiroaki

2018-06-01

Hypoxia offers resistance to therapy in human solid tumors. The aim of the study was to investigate whether SN-38, the active metabolite of irinotecan, acts as a radiosensitizer through inhibition of hypoxia-inducible factor (HIF)-1α in the human colorectal cancer (CRC) cells. HT29 and SW480 cells were cultured with SN-38 (0-4 μM) immediately after irradiation (0-8 Gy). HIF-1α expression was assessed using flow-cytometry and western blot analysis. Cell proliferation was evaluated by the calcein assay. Apoptosis and cell cycle were determined by flow-cytometry. Radiation up-regulated HIF-1α, and SN-38 inhibited the radiation-induced HIF-1α. The combination of radiation and SN-38 inhibited cell proliferation more than radiation alone; treatment with SN-38 after radiation exposure did not increase the number of apoptotic cells, whereas, it enhanced the S and G 2 /M cell-cycle arrest and decreased the population of cells in G 1 Conclusion: SN-38 inhibits the radiation-induced up-regulation of HIF-1α and acts as a radiosensitizer by inducing cell-cycle arrest in CRC cells. Copyright© 2018, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

GLI1 inhibitor GANT61 exhibits antitumor efficacy in T-cell lymphoma cells through down-regulation of p-STAT3 and SOCS3

PubMed Central

Geng, Lingyun; Lu, Kang; Li, Peipei; Li, Xinyu; Zhou, Xiangxiang; Li, Ying; Wang, Xin

2017-01-01

T-cell lymphomas are lymphoid malignancies with aggressive clinical course and poor prognosis. Increasing evidences suggest that deregulation of signal transducer and activator of transcription-3 (STAT3) and suppressor of cytokine signaling 3 (SOCS3) is associated with the pathogenesis of T-cell lymphomas. The hedgehog (Hh)/glioma-associated oncogene-1 (GLI1) pathway, aberrantly activated in a number of tumors, has also been extensively studied. We found that protein expressions of GL11, p-STAT3, STAT3, and SOCS3 were up-regulated in T-cell lymphoma tissues and cell lines. Moreover, the protein expressions of p-STAT3 and SOCS3 were positively correlated with GLI1 in T-cell lymphomas. GLI1 inhibitor GANT61 and lentivirus-mediated siGLI1 exhibited inhibitory effects in the three T-cell lines (Jurkat, Karpass299 and Myla3676 cells). The protein expressions of p-STAT3 and SOCS3 were decreased accompanied with the inhibition of GLI1. These findings indicated that GANT61 is a promising agent against T-cell lymphoma and the antitumor activity might be partly mediated by down-regulating p-STAT3 and SOCS3. PMID:27275540

Roles of Stat3 and ERK in G-CSF signaling.

PubMed

Kamezaki, Kenjirou; Shimoda, Kazuya; Numata, Akihiko; Haro, Takashi; Kakumitsu, Haruko; Yoshie, Masumi; Yamamoto, Masahiro; Takeda, Kiyoshi; Matsuda, Tadashi; Akira, Shizuo; Ogawa, Katsuhiro; Harada, Mine

2005-02-01

G-CSF specifically stimulates the proliferation and differentiation of cells that are committed to the neutrophil-granulocyte lineage. Although Stat3 was thought to be essential for the transduction of G-CSF-induced cell proliferation and differentiation signals, mice deficient for Stat3 in hematopoietic cells show neutrocytosis and infiltration of cells into the digestive tract. The number of progenitor cells in the neutrophil lineage is not changed, and G-CSF-induced proliferation of progenitor cells and prolonged neutrophil survival were observed in Stat3-deficient mice. In hematopoietic cells from Stat3-deficient mice, trace levels of SOCS3, a negative regulator of granulopoiesis, were observed, and SOCS3 expression was not induced by G-CSF stimulation. Stat3-null bone marrow cells displayed a significant activation of extra-cellular regulated kinase 1 (ERK1)/ERK2 under basal conditions, and the activation of ERK was enhanced and sustained by G-CSF stimulation. Furthermore, the augmented proliferation of Stat3-deficient bone marrow cells in response to G-CSF was dramatically decreased by addition of a MEK1 inhibitor. These results indicate that Stat3 functions as a negative regulator of G-CSF signaling by inducing SOCS3 expression and that ERK activation is the major factor responsible for inducing the proliferation of hematopoietic cells in response to G-CSF.

Epigenetic suppression of neprilysin regulates breast cancer invasion

PubMed Central

Stephen, H M; Khoury, R J; Majmudar, P R; Blaylock, T; Hawkins, K; Salama, M S; Scott, M D; Cosminsky, B; Utreja, N K; Britt, J; Conway, R E

2016-01-01

In women, invasive breast cancer is the second most common cancer and the second cause of cancer-related death. Therefore, identifying novel regulators of breast cancer invasion could lead to additional biomarkers and therapeutic targets. Neprilysin, a cell-surface enzyme that cleaves and inactivates a number of substrates including endothelin-1 (ET1), has been implicated in breast cancer, but whether neprilysin promotes or inhibits breast cancer cell progression and metastasis is unclear. Here, we asked whether neprilysin expression predicts and functionally regulates breast cancer cell invasion. RT–PCR and flow cytometry analysis of MDA-MB-231 and MCF-7 breast cancer cell lines revealed decreased neprilysin expression compared with normal epithelial cells. Expression was also suppressed in invasive ductal carcinoma (IDC) compared with normal tissue. In addition, in vtro invasion assays demonstrated that neprilysin overexpression decreased breast cancer cell invasion, whereas neprilysin suppression augmented invasion. Furthermore, inhibiting neprilysin in MCF-7 breast cancer cells increased ET1 levels significantly, whereas overexpressing neprilysin decreased extracellular-signal related kinase (ERK) activation, indicating that neprilysin negatively regulates ET1-induced activation of mitogen-activated protein kinase (MAPK) signaling. To determine whether neprilysin was epigenetically suppressed in breast cancer, we performed bisulfite conversion analysis of breast cancer cells and clinical tumor samples. We found that the neprilysin promoter was hypermethylated in breast cancer; chemical reversal of methylation in MDA-MB-231 cells reactivated neprilysin expression and inhibited cancer cell invasion. Analysis of cancer databases revealed that neprilysin methylation significantly associates with survival in stage I IDC and estrogen receptor-negative breast cancer subtypes. These results demonstrate that neprilysin negatively regulates the ET axis in breast cancer, and epigenetic suppression of neprilysin in invasive breast cancer cells enables invasion. Together, this implicates neprilysin as an important regulator of breast cancer invasion and clarifies its utility as a potential biomarker for invasive breast cancer. PMID:26950599

Epigenetic suppression of neprilysin regulates breast cancer invasion.

PubMed

Stephen, H M; Khoury, R J; Majmudar, P R; Blaylock, T; Hawkins, K; Salama, M S; Scott, M D; Cosminsky, B; Utreja, N K; Britt, J; Conway, R E

2016-03-07

In women, invasive breast cancer is the second most common cancer and the second cause of cancer-related death. Therefore, identifying novel regulators of breast cancer invasion could lead to additional biomarkers and therapeutic targets. Neprilysin, a cell-surface enzyme that cleaves and inactivates a number of substrates including endothelin-1 (ET1), has been implicated in breast cancer, but whether neprilysin promotes or inhibits breast cancer cell progression and metastasis is unclear. Here, we asked whether neprilysin expression predicts and functionally regulates breast cancer cell invasion. RT-PCR and flow cytometry analysis of MDA-MB-231 and MCF-7 breast cancer cell lines revealed decreased neprilysin expression compared with normal epithelial cells. Expression was also suppressed in invasive ductal carcinoma (IDC) compared with normal tissue. In addition, in vtro invasion assays demonstrated that neprilysin overexpression decreased breast cancer cell invasion, whereas neprilysin suppression augmented invasion. Furthermore, inhibiting neprilysin in MCF-7 breast cancer cells increased ET1 levels significantly, whereas overexpressing neprilysin decreased extracellular-signal related kinase (ERK) activation, indicating that neprilysin negatively regulates ET1-induced activation of mitogen-activated protein kinase (MAPK) signaling. To determine whether neprilysin was epigenetically suppressed in breast cancer, we performed bisulfite conversion analysis of breast cancer cells and clinical tumor samples. We found that the neprilysin promoter was hypermethylated in breast cancer; chemical reversal of methylation in MDA-MB-231 cells reactivated neprilysin expression and inhibited cancer cell invasion. Analysis of cancer databases revealed that neprilysin methylation significantly associates with survival in stage I IDC and estrogen receptor-negative breast cancer subtypes. These results demonstrate that neprilysin negatively regulates the ET axis in breast cancer, and epigenetic suppression of neprilysin in invasive breast cancer cells enables invasion. Together, this implicates neprilysin as an important regulator of breast cancer invasion and clarifies its utility as a potential biomarker for invasive breast cancer.

Hormonal regulation of steroid receptor coactivator-1 mRNA in the male and female green anole brain.

PubMed

Kerver, H N; Wade, J

2015-03-01

Green anole lizards are seasonal breeders, with male sexual behaviour primarily regulated by an annual increase in testosterone. Morphological, biochemical and behavioural changes associated with reproduction are activated by testosterone, generally with a greater effect in the breeding season (BS) than in the nonbreeding season (NBS). The present study investigates the possibility that differences in a steroid receptor coactivator may regulate this seasonal difference in responsiveness to testosterone. In situ hybridisation was used to examine the expression of steroid receptor coactivator-1 (SRC-1) in the brains of gonadally intact male and female green anoles across breeding states. A second experiment examined gonadectomised animals with and without testosterone treatment. Gonadally intact males had more SRC-1 expressing cells in the preoptic area and larger volumes of this region as defined by these cells than females. Main effects of both sex and season (males > females and BS > NBS) were present in cell number and volume of the ventromedial hypothalamus. An interaction between sex and season suggested that high expression in BS males was driving these effects. In hormone-manipulated animals, testosterone treatment increased both the number of SRC-1 expressing cells in and volumes of the preoptic area and amygdala. These results suggest that testosterone selectively regulates SRC-1, and that this coactivator may play a role in facilitating reproductive behaviours across both sexes. However, changes in SRC-1 expression are not likely responsible for the seasonal change in responsiveness to testosterone. © 2014 British Society for Neuroendocrinology.

Glucocorticoid-induced TNF receptor family-related protein ligand regulates the migration of monocytes to the inflamed intestine

PubMed Central

Liao, Gongxian; van Driel, Boaz; Magelky, Erica; O'Keeffe, Michael S.; de Waal Malefyt, Rene; Engel, Pablo; Herzog, Roland W.; Mizoguchi, Emiko; Bhan, Atul K.; Terhorst, Cox

2014-01-01

Glucocorticoid-induced TNF receptor family-related protein (GITR) regulates the function of both T cells and antigen-presenting cells (APCs), while the function of GITR ligand (GITR-L) is largely unknown. Here we evaluate the role of GITR-L, whose expression is restricted to APCs, in the development of enterocolitis. On injecting naive CD4+ T cells, GITR-L−/−Rag−/− mice develop a markedly milder colitis than Rag−/− mice, which correlates with a 50% reduction of Ly6C+CD11b+MHCII+ macrophages in the lamina propria and mesenteric lymph nodes. The same result was observed in αCD40-induced acute colitis and during peritonitis, suggesting an altered monocyte migration. In line with these observations, the number of nondifferentiated monocytes was approximately 3-fold higher in the spleen of GITR-L−/−Rag−/− mice than in Rag−/− mice after αCD40 induction. Consistent with the dynamic change in the formation of an active angiotensin II type 1 receptor (AT1) dimer in GITR-L−/− splenic monocytes during intestinal inflammation, the migratory capability of splenic monocytes from GITR-L-deficient mice was impaired in an in vitro transwell migration assay. Conversely, αGITR-L reduces the number of splenic Ly6Chi monocytes, concomitantly with an increase in AT1 dimers. We conclude that GITR-L regulates the number of proinflammatory macrophages in sites of inflammation by controlling the egress of monocytes from the splenic reservoir.—Liao, G., van Driel, B., Magelky, E., O'Keeffe, M. S., de Waal Malefyt, R., Engel, P., Herzog, R. W., Mizoguchi, E., Bhan, A. K., Terhorst, C. Glucocorticoid-induced TNF receptor family-related protein ligand regulates the migration of monocytes to the inflamed intestine. PMID:24107315

The expanding universe of T-cell subsets: Th1, Th2 and more.

PubMed

Mosmann, T R; Sad, S

1996-03-01

Since their discovery nearly ten years ago, T helper 1 (Th1) and Th2 subsets have been implicated in the regulation of many immune responses. In this article, Tim Mosmann and Subash Sad discuss the increasing number of T-cell subsets defined by cytokine patterns; the differentiation pathways of CD4+ and CD8+ T cells; the contribution of other cell types to these patterns; and the cytokine interactions during infection and pregnancy.

Epidermal stem cells: location, potential and contribution to cancer.

PubMed

Ambler, C A; Määttä, A

2009-01-01

Epidermal stem cells have been classically characterized as slow-cycling, long-lived cells that reside in discrete niches in the skin. Gene expression studies of niche-resident cells have revealed a number of stem cell markers and regulators, including the Wnt/beta-catenin, Notch, p63, c-Myc and Hedgehog pathways. A new study challenges the traditional developmental paradigm of slow-cycling stem cells and rapid-cycling transit amplifying cells in some epidermal regions, and there is mounting evidence to suggest that multi-lineage epidermal progenitors can be isolated from highly proliferative, non-niche regions. Whether there is a unique microenvironment surrounding these progenitors remains to be determined. Interestingly, cancer stem cells derived from epidermal tumours exist independent of the classic skin stem cell niche, yet also have stem cell properties, including multi-lineage differentiation. This review summarizes recent studies identifying the location and regulators of mouse and human epidermal stem cells and highlights the strategies used to identify cancer stem cells, including expression of normal epidermal stem cell markers, expression of cancer stem cell markers identified in other epidermal tumours and characterization of side-population tumour cells.

Genetic Analysis of Growth-Regulator-Induced Parthenocarpy in Arabidopsis1

PubMed Central

Vivian-Smith, Adam; Koltunow, Anna M.

1999-01-01

In Arabidopsis, seedless silique development or parthenocarpy can be induced by the application of various plant growth regulators (PGRs) to unfertilized pistils. Ecotype-specific responses were observed in the Arabidopsis ecotypes Columbia and Landsberg relative to the type of PGR and level applied. The parthenocarpic response was greatest in ecotype Landsberg, and comparisons of fruit growth and morphology were studied primarily in this ecotype. Gibberellic acid application (10 μmol pistil−1) caused development similar to that in pollinated pistils, while benzyladenine (1 μmol pistil−1) and naphthylacetic acid (10 μmol pistil−1) treatment produced shorter siliques. Naphthylacetic acid primarily modified mesocarp cell expansion. Arabidopsis mutants were employed to examine potential dependencies on gibberellin biosynthesis (ga1-3, ga4-1, and ga5-1) and perception (spy-4 and gai) during parthenocarpic silique development. Emasculated spy-4 pistils were neither obviously parthenocarpic nor deficient in PGR perception. By contrast, emasculated gai mutants did not produce parthenocarpic siliques following gibberellic acid application, but silique development occurred following pollination or application of auxin and cytokinin. Pollinated gai siliques had decreased cell numbers and morphologically resembled auxin-induced parthenocarpic siliques. This shows that a number of independent and possibly redundant pathways can direct hormone-induced parthenocarpy, and that endogenous gibberellins play a role in regulating cell expansion and promoting cell division in carpels. PMID:10517835

Cellular and Molecular Players in Adipose Tissue Inflammation in the Development of Obesity-induced Insulin Resistance

PubMed Central

Lee, Byung-Cheol; Lee, Jongsoon

2013-01-01

There is increasing evidence showing that inflammation is an important pathogenic mediator of the development of obesity-induced insulin resistance. It is now generally accepted that tissue-resident immune cells play a major role in the regulation of this obesity-induced inflammation. The roles that adipose tissue (AT)-resident immune cells play have been particularly extensively studied. AT contains most types of immune cells and obesity increases their numbers and activation levels, particularly in AT macrophages (ATMs). Other pro-inflammatory cells found in AT include neutrophils, Th1 CD4 T cells, CD8 T cells, B cells, DCs, and mast cells. However, AT also contains anti-inflammatory cells that counter the pro-inflammatory immune cells that are responsible for the obesity-induced inflammation in this tissue. These anti-inflammatory cells include regulatory CD4 T cells (Tregs), Th2 CD4 T cells, and eosinophils. Hence, AT inflammation is shaped by the regulation of pro- and anti-inflammatory immune cell homeostasis, and obesity skews this balance towards a more pro-inflammatory status. Recent genetic studies revealed several molecules that participate in the development of obesity-induced inflammation and insulin resistance. In this review, the cellular and molecular players that participate in the regulation of obesity-induced inflammation and insulin resistance are discussed, with particular attention being placed on the roles of the cellular players in these pathogeneses. PMID:23707515

Chromatin Remodeling Function of BRCA1 and Its Implication in Regulation of DNA Replication

DTIC Science & Technology

2001-09-01

Remodeling Function of BRCAI and its Implication in Regulation of DNA Replication PRINCIPAL INVESTIGATOR: Rong Li, Ph.D. CONTRACTING ORGANIZATION: University...of BRCAI and its Implication DAMD17-99-1-9572 in Regulation of DNA Replication 6. AUTHOR(S) Rong Li, Ph.D. 7. PERFORMING ORGANIZATION NAME(S) AND...1-mediated nuclear functions. 14. SUBJECT TERMS 15. NUMBER OF PAGES Breast Cancer, DNA replication , chromatin remodeling, transcription, 19 cell cycle

MicroRNA-99 family members suppress Homeobox A1 expression in epithelial cells.

PubMed

Chen, Dan; Chen, Zujian; Jin, Yi; Dragas, Dragan; Zhang, Leitao; Adjei, Barima S; Wang, Anxun; Dai, Yang; Zhou, Xiaofeng

2013-01-01

The miR-99 family is one of the evolutionarily most ancient microRNA families, and it plays a critical role in developmental timing and the maintenance of tissue identity. Recent studies, including reports from our group, suggested that the miR-99 family regulates various physiological processes in adult tissues, such as dermal wound healing, and a number of disease processes, including cancer. By combining 5 independent genome-wide expression profiling experiments, we identified a panel of 266 unique transcripts that were down-regulated in epithelial cells transfected with miR-99 family members. A comprehensive bioinformatics analysis using 12 different sequence-based microRNA target prediction algorithms revealed that 81 out of these 266 down-regulated transcripts are potential direct targets for the miR-99 family. Confirmation experiments and functional analyses were performed to further assess 6 selected miR-99 target genes, including mammalian Target of rapamycin (mTOR), Homeobox A1 (HOXA1), CTD small phosphatase-like (CTDSPL), N-myristoyltransferase 1 (NMT1), Transmembrane protein 30A (TMEM30A), and SWI/SNF-related matrix-associated actin-dependent regulator of chromatin subfamily A member 5 (SMARCA5). HOXA1 is a known proto-oncogene, and it also plays an important role in embryonic development. The direct targeting of the miR-99 family to two candidate binding sequences located in the HOXA1 mRNA was confirmed using a luciferase reporter gene assay and a ribonucleoprotein-immunoprecipitation (RIP-IP) assay. Ectopic transfection of miR-99 family reduced the expression of HOXA1, which, in consequence, down-regulated the expression of its downstream gene (i.e., Bcl-2) and led to reduced proliferation and cell migration, as well as enhanced apoptosis. In summary, we identified a number of high-confidence miR-99 family target genes, including proto-oncogene HOXA1, which may play an important role in regulating epithelial cell proliferation and migration during physiological disease processes, such as dermal wound healing and tumorigenesis.

Programmed cell death acts at different stages of Drosophila neurodevelopment to shape the central nervous system

PubMed Central

Desplan, Claude

2016-01-01

Nervous system development is a process that integrates cell proliferation, differentiation and programmed cell death (PCD). PCD is an evolutionary conserved mechanism and a fundamental developmental process by which the final cell number in a nervous system is established. In vertebrates and invertebrates, PCD can be determined intrinsically by cell lineage and age, as well as extrinsically by nutritional, metabolic and hormonal states. Drosophila has been an instrumental model for understanding how this mechanism is regulated. We review the role of PCD in Drosophila central nervous system development from neural progenitors to neurons, its molecular mechanism and function, how it is regulated and implemented, and how it ultimately shapes the fly central nervous system from the embryo to the adult. Finally, we discuss ideas that emerge while integrating this information. PMID:27404003

Innervation regulates synaptic ribbons in lateral line mechanosensory hair cells

PubMed Central

Pujol, Remy; Cunningham, Dale E.; Hailey, Dale W.; Prendergast, Andrew; Rubel, Edwin W.; Raible, David W.

2016-01-01

ABSTRACT Failure to form proper synapses in mechanosensory hair cells, the sensory cells responsible for hearing and balance, leads to deafness and balance disorders. Ribbons are electron-dense structures that tether synaptic vesicles to the presynaptic zone of mechanosensory hair cells where they are juxtaposed with the post-synaptic endings of afferent fibers. They are initially formed throughout the cytoplasm, and, as cells mature, ribbons translocate to the basolateral membrane of hair cells to form functional synapses. We have examined the effect of post-synaptic elements on ribbon formation and maintenance in the zebrafish lateral line system by observing mutants that lack hair cell innervation, wild-type larvae whose nerves have been transected and ribbons in regenerating hair cells. Our results demonstrate that innervation is not required for initial ribbon formation but suggest that it is crucial for regulating the number, size and localization of ribbons in maturing hair cells, and for ribbon maintenance at the mature synapse. PMID:27103160

Innervation regulates synaptic ribbons in lateral line mechanosensory hair cells.

PubMed

Suli, Arminda; Pujol, Remy; Cunningham, Dale E; Hailey, Dale W; Prendergast, Andrew; Rubel, Edwin W; Raible, David W

2016-06-01

Failure to form proper synapses in mechanosensory hair cells, the sensory cells responsible for hearing and balance, leads to deafness and balance disorders. Ribbons are electron-dense structures that tether synaptic vesicles to the presynaptic zone of mechanosensory hair cells where they are juxtaposed with the post-synaptic endings of afferent fibers. They are initially formed throughout the cytoplasm, and, as cells mature, ribbons translocate to the basolateral membrane of hair cells to form functional synapses. We have examined the effect of post-synaptic elements on ribbon formation and maintenance in the zebrafish lateral line system by observing mutants that lack hair cell innervation, wild-type larvae whose nerves have been transected and ribbons in regenerating hair cells. Our results demonstrate that innervation is not required for initial ribbon formation but suggest that it is crucial for regulating the number, size and localization of ribbons in maturing hair cells, and for ribbon maintenance at the mature synapse. © 2016. Published by The Company of Biologists Ltd.

Extracellular IL-33 cytokine, but not endogenous nuclear IL-33, regulates protein expression in endothelial cells.

PubMed

Gautier, Violette; Cayrol, Corinne; Farache, Dorian; Roga, Stéphane; Monsarrat, Bernard; Burlet-Schiltz, Odile; Gonzalez de Peredo, Anne; Girard, Jean-Philippe

2016-10-03

IL-33 is a nuclear cytokine from the IL-1 family that plays important roles in health and disease. Extracellular IL-33 activates a growing number of target cells, including group 2 innate lymphoid cells, mast cells and regulatory T cells, but it remains unclear whether intracellular nuclear IL-33 has additional functions in the nucleus. Here, we used a global proteomic approach based on high-resolution mass spectrometry to compare the extracellular and intracellular roles of IL-33 in primary human endothelial cells, a major source of IL-33 protein in human tissues. We found that exogenous extracellular IL-33 cytokine induced expression of a distinct set of proteins associated with inflammatory responses in endothelial cells. In contrast, knockdown of endogenous nuclear IL-33 expression using two independent RNA silencing strategies had no reproducible effect on the endothelial cell proteome. These results suggest that IL-33 acts as a cytokine but not as a nuclear factor regulating gene expression in endothelial cells.

The GIT–PIX complexes regulate the chemotactic response of rat basophilic leukaemia cells

PubMed Central

Gavina, Manuela; Za, Lorena; Molteni, Raffaella; Pardi, Ruggero; Curtis, Ivan de

2009-01-01

Background information. Cell motility entails the reorganization of the cytoskeleton and membrane trafficking for effective protrusion. The GIT–PIX protein complexes are involved in the regulation of cell motility and adhesion and in the endocytic traffic of members of the family of G-protein-coupled receptors. We have investigated the function of the endogenous GIT complexes in the regulation of cell motility stimulated by fMLP (formyl-Met-Leu-Phe) peptide, in a rat basophilic leukaemia RBL-2H3 cell line stably expressing an HA (haemagglutinin)-tagged receptor for the fMLP peptide. Results. Our analysis shows that RBL cells stably transfected with the chemoattractant receptor expressed both GIT1–PIX and GIT2–PIX endogenous complexes. We have used silencing of the different members of the complex by small interfering RNAs to study the effects on a number of events linked to agonist-induced cell migration. We found that cell adhesion was not affected by depletion of any of the proteins of the GIT complex, whereas agonist-enhanced cell spreading was inhibited. Analysis of agonist-stimulated haptotactic cell migration indicated a specific positive effect of GIT1 depletion on trans-well migration. The internalization of the formyl-peptide receptor was also inhibited by depletion of GIT1 and GIT2. The effects of the GIT complexes on trafficking of the receptors was confirmed by an antibody-enhanced agonist-induced internalization assay, showing that depletion of PIX, GIT1 or GIT2 protein caused decreased perinuclear accumulation of internalized receptors. Conclusions. Our results show that endogenous GIT complexes are involved in the regulation of chemoattractant-induced cell motility and receptor trafficking, and support previous findings indicating an important function of the GIT complexes in the regulation of different G-protein-coupled receptors. Our results also indicate that endogenous GIT1 and GIT2 regulate distinct subsets of agonist-induced responses and suggest a possible functional link between the control of receptor trafficking and the regulation of cell motility by GIT proteins. PMID:19912111

Cooperative functions of Hes/Hey genes in auditory hair cell and supporting cell development.

PubMed

Tateya, Tomoko; Imayoshi, Itaru; Tateya, Ichiro; Ito, Juichi; Kageyama, Ryoichiro

2011-04-15

Notch-mediated lateral inhibition has been reported to regulate auditory hair cell and supporting cell development from common precursors. While the Notch effector genes Hes1, Hes5 and Hey1 are expressed in the developing cochlea, inactivation of either of them causes only mild abnormality, suggesting their functional redundancy. To explore the roles of Hes/Hey genes in cochlear development, we examined compound heterozygous or homozygous mutant mice that lacked Hes1, Hes5 and Hey1 alleles. We found that a reduction in Hes/Hey gene dosage led to graded increase of hair cell formation. However, if at least one allele of Hes1, Hes5 or Hey1 was intact, excessive hair cells were accompanied by overproduction of supporting cells, suggesting that the hair cell increase does not occur at the expense of supporting cells, and that each Hes/Hey gene functions to induce supporting cells. By contrast, when all alleles of Hes1, Hes5 and Hey1 were inactivated, the number of hair cells increased more drastically, whereas that of supporting cells was unchanged compared with control, suggesting that supporting cell formation was balanced by their overproduction and fate conversion into hair cells. The increase of the cell numbers seemed to occur after the prosensory domain formation in the mutants because the proliferation state and the size of the prosensory domain were not affected. Thus, Hes1, Hes5 and Hey1 cooperatively inhibit hair cell formation, and one allele of Hes1, Hes5 or Hey1 is sufficient for supporting cell production probably by lateral inhibition in the sensory epithelium. Strikingly, Hes/Hey mutations lead to disorganized cell alignment and polarity and to hearing loss despite hair cell overproduction. These results suggest that Hes/Hey gene dosage is essential not only for generation of appropriate numbers of hair cells and supporting cells by controlling cell proliferation and lateral inhibition but also for the hearing ability by regulating the cell alignment and polarity. Copyright © 2011 Elsevier Inc. All rights reserved.

In situ monitoring of PTHLH secretion in neuroblastoma cells cultured onto nanoporous membranes.

PubMed

de la Escosura-Muñiz, Alfredo; Espinoza-Castañeda, Marisol; Chamorro-García, Alejandro; Rodríguez-Hernández, Carlos J; de Torres, Carmen; Merkoçi, Arben

2018-06-01

In this work, we propose for the first time the use of anodic aluminum oxide (AAO) nanoporous membranes for in situ monitoring of parathyroid hormone-like hormone (PTHLH) secretion in cultured human cells. The biosensing system is based on the nanochannels blockage upon immunocomplex formation, which is electrically monitored through the voltammetric oxidation of Prussian blue nanoparticles (PBNPs). Models evaluated include a neuroblastoma cell line (SK-N-AS) and immortalized keratinocytes (HaCaT) as a control of high PTHLH production. The effect of total number of seeded cells and incubation time on the secreted PTHLH levels is assessed, finding that secreted PTHLH levels range from approximately 60 to 400 ng/mL. Moreover, our methodology is also applied to analyse PTHLH production following PTHLH gene knockdown upon transient cell transfection with a specific silencing RNA (siRNA). Given that inhibition of PTHLH secretion reduces cell proliferation, survival and invasiveness in a number of tumors, our system provides a powerful tool for the preclinical evaluation of therapies that regulate PTHLH production. This nanoporous membrane - based sensing technology might be useful to monitor the active secretion of other proteins as well, thus contributing to characterize their regulation and function. Copyright © 2018 Elsevier B.V. All rights reserved.

Lipocalin 2 functions as a negative regulator of red blood cell production in an autocrine fashion.

PubMed

Miharada, Ken-ichi; Hiroyama, Takashi; Sudo, Kazuhiro; Nagasawa, Toshiro; Nakamura, Yukio

2005-11-01

Members of the lipocalin protein family are typically small, secreted proteins that possess a variety of functions. Although the physiological role of lipocalin 2 remains to be fully elucidated, a few pivotal functions have recently been reported, e.g., regulation of the apoptosis of leukocytes. Unexpectedly, lipocalin 2 is abundantly expressed in erythroid progenitor cells. An in vitro culture experiment demonstrated that lipocalin 2 induces apoptosis and inhibits differentiation of erythroid progenitor cells. During acute anemia the expression of lipocalin 2 was reduced in erythroid cells by a feedback system. Furthermore, injection of recombinant lipocalin 2 into mice suffering from acute anemia retarded the recovery of red blood cell (RBC) numbers, suggesting the importance of reduced expression of lipocalin 2 for the efficient recovery of RBC numbers. These results indicate that lipocalin 2 suppresses RBC production in an autocrine fashion. Hence, anemia arising from pathological conditions, such as chronic inflammation, might be partly due to increased levels of lipocalin 2 secreted from expanded leukocytes and/or macrophages. Also, anemia arising from malignancies might be partly due to the abundant secretion of lipocalin 2 from tumor cells. Thus, lipocalin 2 may represent an attractive therapeutic target for anemia under certain pathological conditions.

Sexual experience affects reproductive behavior and preoptic androgen receptors in male mice

PubMed Central

Swaney, William T.; Dubose, Brittany N.; Curley, James P.; Champagne, Frances A.

2012-01-01

Reproductive behavior in male rodents is made up of anticipatory and consummatory elements which are regulated in the brain by sensory systems, reward circuits and hormone signaling. Gonadal steroids play a key role in the regulation of male sexual behavior via steroid receptors in the hypothalamus and preoptic area. Typical patterns of male reproductive behavior have been characterized, however these are not fixed but are modulated by adult experience. We assessed the effects of repeated sexual experience on male reproductive behavior of C57BL/6 mice; including measures of olfactory investigation of females, mounting, intromission and ejaculation. The effects of sexual experience on the number of cells expressing either androgen receptor (AR) or estrogen receptor alpha (ERα) in the primary brain nuclei regulating male sexual behavior was also measured. Sexually experienced male mice engaged in less sniffing of females before initiating sexual behavior and exhibited shorter latencies to mount and intromit, increased frequency of intromission, and increased duration of intromission relative to mounting. No changes in numbers of ERα-positive cells were observed, however sexually experienced males had increased numbers of AR-positive cells in the medial preoptic area (MPOA); the primary regulatory nucleus for male sexual behavior. These results indicate that sexual experience results in a qualitative change in male reproductive behavior in mice that is associated with increased testosterone sensitivity in the MPOA and that this nucleus may play a key integrative role in mediating the effects of sexual experience on male behavior. PMID:22266118

DOE Office of Scientific and Technical Information (OSTI.GOV)

Buttner, William; Rivkin, C.; Burgess, R.

Here, the United Nations Economic Commission for Europe Global Technical Regulation (GTR) Number 13 ( Global Technical Regulation on Hydrogen and Fuel Cell Vehicles) is the defining document regulating safety requirements in hydrogen vehicles, and in particular, fuel cell electric vehicles (FCEVs). GTR Number 13 has been formally adopted and will serve as the basis for the national regulatory standards for FCEV safety in North America (led by the United States), Japan, Korea, and the European Union. The GTR defines safety requirements for these vehicles, including specifications on the allowable hydrogen levels in vehicle enclosures during in-use and post-crash conditionsmore » and on the allowable hydrogen emissions levels in vehicle exhaust during certain modes of normal operation. However, in order to be incorporated into national regulations, that is, to be legally binding, methods to verify compliance with the specific requirements must exist. In a collaborative program, the Sensor Laboratories at the National Renewable Energy Laboratory in the United States and the Joint Research Centre, Institute for Energy and Transport in the Netherlands have been evaluating and developing analytical methods that can be used to verify compliance with the hydrogen release requirements as specified in the GTR.« less

Identification of genetic loci that modulate cell proliferation in the adult rostral migratory stream using the expanded panel of BXD mice.

PubMed

Poon, Anna; Goldowitz, Daniel

2014-03-19

Adult neurogenesis, which is the continual production of new neurons in the mature brain, demonstrates the strikingly plastic nature of the nervous system. Adult neural stem cells and their neural precursors, collectively referred to as neural progenitor cells (NPCs), are present in the subgranular zone (SGZ) of the dentate gyrus, the subventricular zone (SVZ), and rostral migratory stream (RMS). In order to harness the potential of NPCs to treat neurodegenerative diseases and brain injuries, it will be important to understand the molecules that regulate NPCs in the adult brain. The genetic basis underlying NPC proliferation is still not fully understood. From our previous quantitative trait locus (QTL) analysis, we had success in using a relatively small reference population of recombinant inbred strains of mice (AXBXA) to identify a genetic region that is significantly correlated with NPC proliferation in the RMS. In this study, we expanded our initial QTL mapping of RMS proliferation to a far richer genetic resource, the BXD RI mouse strains. A 3-fold difference in the number of proliferative, bromodeoxyuridine (BrdU)-labeled cells was quantified in the adult RMS of 61 BXD RI strains. RMS cell proliferation is highly dependent on the genetic background of the mice with an estimated heritability of 0.58. Genome-wide mapping revealed a significant QTL on chromosome (Chr) 6 and a suggestive QTL on Chr 11 regulating the number of NPCs in the RMS. Composite interval analysis further revealed secondary QTLs on Chr 14 and Chr 18. The loci regulating RMS cell proliferation did not overlap with the suggestive loci modulating cell proliferation in the SGZ. These mapped loci serve as starting points to identify genes important for this process. A subset of candidate genes in this region is associated with cell proliferation and neurogenesis. Interconnectivity of these candidate genes was demonstrated using pathway and transcriptional covariance analyses. Differences in RMS cell proliferation across the BXD RI strains identifies genetic loci that serve to provide insights into the interplay of underlying genes that may be important for regulating NPC proliferation in the adult mouse brain.

Hypocotyl Transcriptome Reveals Auxin Regulation of Growth-Promoting Genes through GA-Dependent and -Independent Pathways

PubMed Central

Castillejo, Cristina; Sartor, Ryan; Bialy, Agniezska; Sun, Tai-ping; Estelle, Mark

2012-01-01

Many processes critical to plant growth and development are regulated by the hormone auxin. Auxin responses are initiated through activation of a transcriptional response mediated by the TIR1/AFB family of F-box protein auxin receptors as well as the AUX/IAA and ARF families of transcriptional regulators. However, there is little information on how auxin regulates a specific cellular response. To begin to address this question, we have focused on auxin regulation of cell expansion in the Arabidopsis hypocotyl. We show that auxin-mediated hypocotyl elongation is dependent upon the TIR1/AFB family of auxin receptors and degradation of AUX/IAA repressors. We also use microarray studies of elongating hypocotyls to show that a number of growth-associated processes are activated by auxin including gibberellin biosynthesis, cell wall reorganization and biogenesis, and others. Our studies indicate that GA biosynthesis is required for normal response to auxin in the hypocotyl but that the overall transcriptional auxin output consists of PIF-dependent and -independent genes. We propose that auxin acts independently from and interdependently with PIF and GA pathways to regulate expression of growth-associated genes in cell expansion. PMID:22590525

Role of ER Export Signals in Controlling Surface Potassium Channel Numbers

NASA Astrophysics Data System (ADS)

Ma, Dzwokai; Zerangue, Noa; Lin, Yu-Fung; Collins, Anthony; Yu, Mei; Jan, Yuh Nung; Yeh Jan, Lily

2001-01-01

Little is known about the identity of endoplasmic reticulum (ER) export signals and how they are used to regulate the number of proteins on the cell surface. Here, we describe two ER export signals that profoundly altered the steady-state distribution of potassium channels and were required for channel localization to the plasma membrane. When transferred to other potassium channels or a G protein-coupled receptor, these ER export signals increased the number of functional proteins on the cell surface. Thus, ER export of membrane proteins is not necessarily limited by folding or assembly, but may be under the control of specific export signals.

Notochordal cells in the adult intervertebral disc: new perspective on an old question.

PubMed

Risbud, Makarand V; Shapiro, Irving M

2011-01-01

The intervertebral disc is a tissue positioned between each of the vertebrae that accommodates applied biomechanical forces to the spine. The central compartment of the disc contains the nucleus pulposus (NP) which is enclosed by the annulus fibrosus and the endplate cartilage.The NP is derived from the notochord, a rod-like structure of mesodermal origin. Development of the notochord is tightly regulated by interactive transcription factors and target genes. Since a number of these molecules are unique they have be used for cell lineage and fate mapping studies of tissues of the intervertebral disc. These studies have shown that in a number of species including human, NP tissue retains notochordal cells throughout life. In the adult NP, there are present both large and small notochordal cells, as well as a progenitor cell population which can differentiate along the mesengenic pathway. Since tissue renewal in the intervertebral disc is dependent on the ability of these cells to commit to the NP lineage and undergo terminal differentiation, studies have been performed to assess which signaling pathways may regulate these activities. The notch signaling pathway is active in the intervertebral disc and is responsive to hypoxia, probably through HIF-1a. From a disease viewpoint, it is hypothesized that an oxemic shift, possibly mediated by alterations in the vascular supply to the tissues of the disc would be expected to lead to a failure in notochordal progenitor cell activation and a decrease in the number of differentiated cells. In turn, this would lead to decrements in function and enhancement of the effect of agents that are known to promote disc degeneration.

mRNA expression profiling of laser microbeam microdissected cells from slender embryonic structures.

PubMed

Scheidl, Stefan J; Nilsson, Sven; Kalén, Mattias; Hellström, Mats; Takemoto, Minoru; Håkansson, Joakim; Lindahl, Per

2002-03-01

Microarray hybridization has rapidly evolved as an important tool for genomic studies and studies of gene regulation at the transcriptome level. Expression profiles from homogenous samples such as yeast and mammalian cell cultures are currently extending our understanding of biology, whereas analyses of multicellular organisms are more difficult because of tissue complexity. The combination of laser microdissection, RNA amplification, and microarray hybridization has the potential to provide expression profiles from selected populations of cells in vivo. In this article, we present and evaluate an experimental procedure for global gene expression analysis of slender embryonic structures using laser microbeam microdissection and laser pressure catapulting. As a proof of principle, expression profiles from 1000 cells in the mouse embryonic (E9.5) dorsal aorta were generated and compared with profiles for captured mesenchymal cells located one cell diameter further away from the aortic lumen. A number of genes were overexpressed in the aorta, including 11 previously known markers for blood vessels. Among the blood vessel markers were endoglin, tie-2, PDGFB, and integrin-beta1, that are important regulators of blood vessel formation. This demonstrates that microarray analysis of laser microbeam micro-dissected cells is sufficiently sensitive for identifying genes with regulative functions.

Causality, randomness, intelligibility, and the epistemology of the cell.

PubMed

Dougherty, Edward R; Bittner, Michael L

2010-06-01

Because the basic unit of biology is the cell, biological knowledge is rooted in the epistemology of the cell, and because life is the salient characteristic of the cell, its epistemology must be centered on its livingness, not its constituent components. The organization and regulation of these components in the pursuit of life constitute the fundamental nature of the cell. Thus, regulation sits at the heart of biological knowledge of the cell and the extraordinary complexity of this regulation conditions the kind of knowledge that can be obtained, in particular, the representation and intelligibility of that knowledge. This paper is essentially split into two parts. The first part discusses the inadequacy of everyday intelligibility and intuition in science and the consequent need for scientific theories to be expressed mathematically without appeal to commonsense categories of understanding, such as causality. Having set the backdrop, the second part addresses biological knowledge. It briefly reviews modern scientific epistemology from a general perspective and then turns to the epistemology of the cell. In analogy with a multi-faceted factory, the cell utilizes a highly parallel distributed control system to maintain its organization and regulate its dynamical operation in the face of both internal and external changes. Hence, scientific knowledge is constituted by the mathematics of stochastic dynamical systems, which model the overall relational structure of the cell and how these structures evolve over time, stochasticity being a consequence of the need to ignore a large number of factors while modeling relatively few in an extremely complex environment.

Copper Regulates Maturation and Expression of an MITF:Tryptase Axis in Mast Cells.

PubMed

Hu Frisk, Jun Mei; Kjellén, Lena; Kaler, Stephen G; Pejler, Gunnar; Öhrvik, Helena

2017-12-15

Copper has previously been implicated in the regulation of immune responses, but the impact of this metal on mast cells is poorly understood. In this article, we address this issue and show that copper starvation of mast cells causes increased granule maturation, as indicated by higher proteoglycan content, stronger metachromatic staining, and altered ultrastructure in comparison with nontreated cells, whereas copper overload has the opposite effects. In contrast, copper status did not impact storage of histamine in mast cells, nor did alterations in copper levels affect the ability of mast cells to degranulate in response to IgER cross-linking. A striking finding was decreased tryptase content in mast cells with copper overload, whereas copper starvation increased tryptase content. These effects were associated with corresponding shifts in tryptase mRNA levels, suggesting that copper affects tryptase gene regulation. Mechanistically, we found that alterations in copper status affected the expression of microphthalmia-associated transcription factor, a transcription factor critical for driving tryptase expression. We also found evidence supporting the concept that the effects on microphthalmia-associated transcription factor are dependent on copper-mediated modulation of MAPK signaling. Finally, we show that, in MEDNIK syndrome, a condition associated with low copper levels and a hyperallergenic skin phenotype, including pruritis and dermatitis, the number of tryptase-positive mast cells is increased. Taken together, our findings reveal a hitherto unrecognized role for copper in the regulation of mast cell gene expression and maturation. Copyright © 2017 by The American Association of Immunologists, Inc.

The histone H2A deubiquitinase Usp16 regulates hematopoiesis and hematopoietic stem cell function.

PubMed

Gu, Yue; Jones, Amanda E; Yang, Wei; Liu, Shanrun; Dai, Qian; Liu, Yudong; Swindle, C Scott; Zhou, Dewang; Zhang, Zhuo; Ryan, Thomas M; Townes, Tim M; Klug, Christopher A; Chen, Dongquan; Wang, Hengbin

2016-01-05

Epigenetic mechanisms play important regulatory roles in hematopoiesis and hematopoietic stem cell (HSC) function. Subunits of polycomb repressive complex 1 (PRC1), the major histone H2A ubiquitin ligase, are critical for both normal and pathological hematopoiesis; however, it is unclear which of the several counteracting H2A deubiquitinases functions along with PRC1 to control H2A ubiquitination (ubH2A) level and regulates hematopoiesis in vivo. Here we investigated the function of Usp16 in mouse hematopoiesis. Conditional deletion of Usp16 in bone marrow resulted in a significant increase of global ubH2A level and lethality. Usp16 deletion did not change HSC number but was associated with a dramatic reduction of mature and progenitor cell populations, revealing a role in governing HSC lineage commitment. ChIP- and RNA-sequencing studies in HSC and progenitor cells revealed that Usp16 bound to many important hematopoietic regulators and that Usp16 deletion altered the expression of genes in transcription/chromosome organization, immune response, hematopoietic/lymphoid organ development, and myeloid/leukocyte differentiation. The altered gene expression was partly rescued by knockdown of PRC1 subunits, suggesting that Usp16 and PRC1 counterbalance each other to regulate cellular ubH2A level and gene expression in the hematopoietic system. We further discovered that knocking down Cdkn1a (p21cip1), a Usp16 target and regulated gene, rescued the altered cell cycle profile and differentiation defect of Usp16-deleted HSCs. Collectively, these studies identified Usp16 as one of the histone H2A deubiquitinases, which coordinates with the H2A ubiquitin ligase PRC1 to regulate hematopoiesis, and revealed cell cycle regulation by Usp16 as key for HSC differentiation.

Regulation of early Xenopus development by ErbB signaling

PubMed Central

Nie, Shuyi; Chang, Chenbei

2008-01-01

ErbB signaling has long been implicated in cancer formation and progression and is shown to regulate cell division, migration and death during tumorigenesis. The functions of the ErbB pathway during early vertebrate embryogenesis, however, are not well understood. Here we report characterization of ErbB activities during early frog development. Gain-of-function analyses show that EGFR, ErbB2 and ErbB4 induce ectopic tumor-like cell mass that contains increased numbers of mitotic cells. Both the muscle and the neural markers are expressed in these ectopic protrusions. ErbBs also induce mesodermal markers in ectodermal explants. Loss-of-function studies using carboxyl terminal-truncated dominant-negative ErbB receptors demonstrate that blocking ErbB signals leads to defective gastrulation movements and malformation of the embryonic axis with a reduction in the head structures in early frog embryos. These data, together with the observation that ErbBs are expressed early during frog embryogenesis, suggest that ErbBs regulate cell proliferation, movements and embryonic patterning during early Xenopus development. PMID:16258939

The transcription factor Th-POK negatively regulates Th17 differentiation in Vα14i NKT cells

PubMed Central

Engel, Isaac; Zhao, Meng; Kappes, Dietmar; Taniuchi, Ichiro

2012-01-01

The majority of mouse Vα14 invariant natural killer T (Vα14i NKT) cells produce several cytokines, including IFNγ and IL-4, very rapidly after activation. A subset of these cells, known as NKT17 cells, however, differentiates in the thymus to preferentially produce IL-17. Here, we show that the transcription factor—known as T helper, Poxviruses, and Zinc-finger and Krüppel family, (Th-POK)—represses the formation of NKT17 cells. Vα14i NKT cells from Th-POK–mutant helper deficient (hd/hd) mice have increased transcripts of genes normally expressed by Th17 and NKT17 cells, and even heterozygosity for this mutation leads to dramatically increased numbers of Vα14i NKT cells that are poised to express IL-17, especially in the thymus and lymph nodes. In addition, using gene reporter mice, we demonstrate that NKT17 cells from wild-type mice express lower amounts of Th-POK than the majority population of Vα14i NKT cells. We also show that retroviral transduction of Th-POK represses the expression of the Th17 master regulator RORγT in Vα14i NKT-cell lines. Our data suggest that NKT17-cell differentiation is intrinsically regulated by Th-POK activity, with only low levels of Th-POK permissive for the differentiation of NKT17 cells. PMID:23034280

Organtropic Metastatic Secretomes and Exosomes in Breast Cancer

DTIC Science & Technology

2015-10-01

AWARD NUMBER: W81XWH-13-1-0427 TITLE: Organtropic Metastatic Secretomes and Exosomes in Breast Cancer PRINCIPAL INVESTIGATOR: Dr. David Lyden...4. TITLE AND SUBTITLE 5a. CONTRACT NUMBER Organtropic Metastatic Secretomes and Exosomes in Breast Cancer 5b. GRANT NUMBER W81XWH-13-1-0427 5c...understanding of secreted metastasis regulators (extracellular proteins, cell-free nucleic acids and small vesicles – exosomes -) has tremendous

Active chinese mistletoe lectin-55 enhances colon cancer surveillance through regulating innate and adaptive immune responses

PubMed Central

Ma, Yan-Hui; Cheng, Wei-Zhi; Gong, Fang; Ma, An-Lun; Yu, Qi-Wen; Zhang, Ji-Ying; Hu, Chao-Ying; Chen, Xue-Hua; Zhang, Dong-Qing

2008-01-01

AIM: To investigate the potential role of Active Chinese mistletoe lectin-55 (ACML-55) in tumor immune surveillance. METHODS: In this study, an experimental model was established by hypodermic inoculating the colon cancer cell line CT26 (5 × 105 cells) into BALB/c mice. The experimental treatment was orally administered with ACML-55 or PBS, followed by the inoculation of colon cancer cell line CT26. Intracellular cytokine staining was used to detect IFN-γ production by tumor antigen specific CD8+ T cells. FACS analysis was employed to profile composition and activation of CD4+, CD8+, γδ T and NK cells. RESULTS: Our results showed, compared to PBS treated mice, ACML-55 treatment significantly delayed colon cancer development in colon cancer -bearing Balb/c mice in vivo. Treatment with ACML-55 enhanced both Ag specific activation and proliferation of CD4+ and CD8+ T cells, and increased the number of tumor Ag specific CD8+ T cells. It was more important to increase the frequency of tumor Ag specific IFN-γ producing-CD8+ T cells. Interestingly, ACML-55 treatment also showed increased cell number of NK, and γδT cells, indicating the role of ACML-55 in activation of innate lymphocytes. CONCLUSION: Our results demonstrate that ACML-55 therapy can enhance function in immune surveillance in colon cancer-bearing mice through regulating both innate and adaptive immune responses. PMID:18785279

SPSB1, a Novel Negative Regulator of the Transforming Growth Factor-β Signaling Pathway Targeting the Type II Receptor.

PubMed

Liu, Sheng; Nheu, Thao; Luwor, Rodney; Nicholson, Sandra E; Zhu, Hong-Jian

2015-07-17

Appropriate cellular signaling is essential to control cell proliferation, differentiation, and cell death. Aberrant signaling can have devastating consequences and lead to disease states, including cancer. The transforming growth factor-β (TGF-β) signaling pathway is a prominent signaling pathway that has been tightly regulated in normal cells, whereas its deregulation strongly correlates with the progression of human cancers. The regulation of the TGF-β signaling pathway involves a variety of physiological regulators. Many of these molecules act to alter the activity of Smad proteins. In contrast, the number of molecules known to affect the TGF-β signaling pathway at the receptor level is relatively low, and there are no known direct modulators for the TGF-β type II receptor (TβRII). Here we identify SPSB1 (a Spry domain-containing Socs box protein) as a novel regulator of the TGF-β signaling pathway. SPSB1 negatively regulates the TGF-β signaling pathway through its interaction with both endogenous and overexpressed TβRII (and not TβRI) via its Spry domain. As such, TβRII and SPSB1 co-localize on the cell membrane. SPSB1 maintains TβRII at a low level by enhancing the ubiquitination levels and degradation rates of TβRII through its Socs box. More importantly, silencing SPSB1 by siRNA results in enhanced TGF-β signaling and migration and invasion of tumor cells. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

Cell-derived microparticles: new targets in the therapeutic management of disease.

PubMed

Roseblade, Ariane; Luk, Frederick; Rawling, Tristan; Ung, Alison; Grau, Georges E R; Bebawy, Mary

2013-01-01

Intercellular communication is essential to maintain vital physiological activities and to regulate the organism's phenotype. There are a number of ways in which cells communicate with one another. This can occur via autocrine signaling, endocrine signaling or by the transfer of molecular mediators across gap junctions. More recently communication via microvesicular shedding has gained important recognition as a significant pathway by which cells can coordinate the spread and dominance of selective traits within a population. Through this communication apparatus, cells can now acquire and secure a survival advantage, particularly in the context of malignant disease. This review aims to highlight some of the functions and implications of microparticles in physiology of various disease states, and present a novel therapeutic strategy through the regulation of microparticle production.

Programmed Cell Death and Caspase Functions During Neural Development.

PubMed

Yamaguchi, Yoshifumi; Miura, Masayuki

2015-01-01

Programmed cell death (PCD) is a fundamental component of nervous system development. PCD serves as the mechanism for quantitative matching of the number of projecting neurons and their target cells through direct competition for neurotrophic factors in the vertebrate peripheral nervous system. In addition, PCD plays roles in regulating neural cell numbers, canceling developmental errors or noise, and tissue remodeling processes. These findings are mainly derived from genetic studies that prevent cells from dying by apoptosis, which is a major form of PCD and is executed by activation of evolutionarily conserved cysteine protease caspases. Recent studies suggest that caspase activation can be coordinated in time and space at multiple levels, which might underlie nonapoptotic roles of caspases in neural development in addition to apoptotic roles. © 2015 Elsevier Inc. All rights reserved.

Deregulation of cell growth and malignant transformation.

PubMed

Sulić, Sanda; Panić, Linda; Dikić, Ivan; Volarević, Sinisa

2005-08-01

Cell growth and cell division are fundamental aspects of cell behavior in all organisms. Recent insights from many model organisms have shed light on the molecular mechanisms that control cell growth and cell division. A significant body of evidence has now been accumulated, showing a direct link between deregulation of components of cell cycle machinery and cancer. In addition, defects in one or more steps that control growth are important for malignant transformation, as many tumor suppressors and proto-oncogenes have been found to regulate cell growth. The importance of cell growth in tumor development is further supported by the discovery that rapamycin, an effective anticancer drug, inhibits a key regulator of protein synthetic machinery and cell growth, mammalian target of rapamycin (mTOR). In most cases, cell growth and cell division are coupled, thereby maintaining cell size within physiological limits. We believe that, in a long-term perspective, understanding how these two processes are coordinated in vivo and how their interplay is deregulated in a number of diseases, including cancer, may have a direct impact on the efficiency of modern therapeutics.

Small intestinal eosinophils regulate Th17 cells by producing IL-1 receptor antagonist.

PubMed

Sugawara, Reiko; Lee, Eun-Jung; Jang, Min Seong; Jeun, Eun-Ji; Hong, Chun-Pyo; Kim, Jung-Hwan; Park, Areum; Yun, Chang Ho; Hong, Sung-Wook; Kim, You-Me; Seoh, Ju-Young; Jung, YunJae; Surh, Charles D; Miyasaka, Masayuki; Yang, Bo-Gie; Jang, Myoung Ho

2016-04-04

Eosinophils play proinflammatory roles in helminth infections and allergic diseases. Under steady-state conditions, eosinophils are abundantly found in the small intestinal lamina propria, but their physiological function is largely unexplored. In this study, we found that small intestinal eosinophils down-regulate Th17 cells. Th17 cells in the small intestine were markedly increased in the ΔdblGATA-1 mice lacking eosinophils, and an inverse correlation was observed between the number of eosinophils and that of Th17 cells in the small intestine of wild-type mice. In addition, small intestinal eosinophils suppressed the in vitro differentiation of Th17 cells, as well as IL-17 production by small intestinal CD4(+)T cells. Unlike other small intestinal immune cells or circulating eosinophils, we found that small intestinal eosinophils have a unique ability to constitutively secrete high levels of IL-1 receptor antagonist (IL-1Ra), a natural inhibitor of IL-1β. Moreover, small intestinal eosinophils isolated from IL-1Ra-deficient mice failed to suppress Th17 cells. Collectively, our results demonstrate that small intestinal eosinophils play a pivotal role in the maintenance of intestinal homeostasis by regulating Th17 cells via production of IL-1Ra. © 2016 Sugawara et al.

Small intestinal eosinophils regulate Th17 cells by producing IL-1 receptor antagonist

PubMed Central

Sugawara, Reiko; Lee, Eun-Jung; Jang, Min Seong; Jeun, Eun-Ji; Hong, Chun-Pyo; Kim, Jung-Hwan; Park, Areum; Yun, Chang Ho; Hong, Sung-Wook; Kim, You-Me; Seoh, Ju-Young; Jung, YunJae; Surh, Charles D.; Miyasaka, Masayuki

2016-01-01

Eosinophils play proinflammatory roles in helminth infections and allergic diseases. Under steady-state conditions, eosinophils are abundantly found in the small intestinal lamina propria, but their physiological function is largely unexplored. In this study, we found that small intestinal eosinophils down-regulate Th17 cells. Th17 cells in the small intestine were markedly increased in the ΔdblGATA-1 mice lacking eosinophils, and an inverse correlation was observed between the number of eosinophils and that of Th17 cells in the small intestine of wild-type mice. In addition, small intestinal eosinophils suppressed the in vitro differentiation of Th17 cells, as well as IL-17 production by small intestinal CD4+ T cells. Unlike other small intestinal immune cells or circulating eosinophils, we found that small intestinal eosinophils have a unique ability to constitutively secrete high levels of IL-1 receptor antagonist (IL-1Ra), a natural inhibitor of IL-1β. Moreover, small intestinal eosinophils isolated from IL-1Ra−deficient mice failed to suppress Th17 cells. Collectively, our results demonstrate that small intestinal eosinophils play a pivotal role in the maintenance of intestinal homeostasis by regulating Th17 cells via production of IL-1Ra. PMID:26951334

Down-regulation of KIAA1199/CEMIP by miR-216a suppresses tumor invasion and metastasis in colorectal cancer.

PubMed

Zhang, Dejun; Zhao, Lei; Shen, Qiong; Lv, Qing; Jin, Min; Ma, Hong; Nie, Xiu; Zheng, Xiumei; Huang, Shaoyi; Zhou, Pengfei; Wu, Gang; Zhang, Tao

2017-05-15

Colorectal cancer is one of the major causes of death from cancer. Metastasis is the leading cause of treatment failure, in which cancer stem cells and circulating tumor cells play crucial roles. Identifying the involved metastatic biomarkers and clarifying the regulation mechanisms are of great importance for targeting tumor metastasis. In the current research, we discovered that KIAA1199, a cell-migration inducing protein, showed higher expression in CD44+ cancer cells from metastatic compared with the paired primary tissues, and was upregulated in colorectal cancer and positively correlated with numbers and mesenchymal phenotype of circulating tumor cells, and predicted shorter progress-free survival. Moreover, we indicated that down-regulation of KIAA1199 suppressed migration and invasion of colorectal cancer cells in vitro, and inhibited metastasis in vivo. Furthermore, we demonstrated that KIAA1199 was one of the direct and functional targets of miR-216a, and miR-216a overexpression led to decreased migration and invasion of colorectal cancer cells in vitro, and inhibited metastasis in vivo. Collectively, KIAA1199 plays a critical role in maintaining an aggressive phenotype of tumor cells, and suppression of KIAA1199-related motilities of tumor cells contributes to reduced tumor metastasis in colorectal cancer. © 2017 UICC.

Dehydrated human amnion/chorion membrane regulates stem cell activity in vitro.

PubMed

Massee, Michelle; Chinn, Kathryn; Lei, Jennifer; Lim, Jeremy J; Young, Conan S; Koob, Thomas J

2016-10-01

Human-derived placental tissues have been shown in randomized clinical trials to be effective for healing chronic wounds, and have also demonstrated the ability to recruit stem cells to the wound site in vitro and in vivo. In this study, PURION(®) Processed dehydrated human amnion/chorion membrane allografts (dHACM, EpiFix(®) , MiMedx Group, Marietta, GA) were evaluated for their ability to alter stem cell activity in vitro. Human bone marrow mesenchymal stem cells (BM-MSCs), adipose derived stem cells (ADSCs), and hematopoietic stem cells (HSCs) were treated with soluble extracts of dHACM tissue, and were evaluated for cellular proliferation, migration, and cytokine secretion. Stem cells were analyzed for cell number by DNA assay after 24 h, closure of an acellular zone using microscopy over 3 days, and soluble cytokine production in the medium of treated stem cells was analyzed after 3 days using a multiplex ELISA array. Treatment with soluble extracts of dHACM tissue stimulated BM-MSCs, ADSCs, and HSCs to proliferate with a significant increase in cell number after 24 h. dHACM treatment accelerated closure of an acellular zone by ADSCs and BM-MSCs after 3 days, compared to basal medium. BM-MSCs, ADSCs, and HSCs also modulated endogenous production of a number of various soluble signals, including regulators of inflammation, mitogenesis, and wound healing. dHACM treatment promoted increased proliferation and migration of ADSCs, BM-MSCs, and HSCs, along with modulation of secreted proteins from those cells. Therefore, dHACM may impact wound healing by amplifying host stem cell populations and modulating their responses in treated wound tissues. © 2015 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 104B: 1495-1503, 2016. © 2015 The Authors. Journal of Biomedical Materials Research Part A Published by Wiley Periodicals, Inc.

Cerebellar neurodegeneration in the absence of microRNAs

PubMed Central

Schaefer, Anne; O'Carroll, Dónal; Tan, Chan Lek; Hillman, Dean; Sugimori, Mutsuyuki; Llinas, Rodolfo; Greengard, Paul

2007-01-01

Genome-encoded microRNAs (miRNAs) are potent regulators of gene expression. The significance of miRNAs in various biological processes has been suggested by studies showing an important role of these small RNAs in regulation of cell differentiation. However, the role of miRNAs in regulation of differentiated cell physiology is not well established. Mature neurons express a large number of distinct miRNAs, but the role of miRNAs in postmitotic neurons has not been examined. Here, we provide evidence for an essential role of miRNAs in survival of differentiated neurons. We show that conditional Purkinje cell–specific ablation of the key miRNA-generating enzyme Dicer leads to Purkinje cell death. Deficiency in Dicer is associated with progressive loss of miRNAs, followed by cerebellar degeneration and development of ataxia. The progressive neurodegeneration in the absence of Dicer raises the possibility of an involvement of miRNAs in neurodegenerative disorders. PMID:17606634

Activity-dependent self-regulation of viscous length scales in biological systems

NASA Astrophysics Data System (ADS)

Nandi, Saroj Kumar

2018-05-01

The cellular cortex, which is a highly viscous thin cytoplasmic layer just below the cell membrane, controls the cell's mechanical properties, which can be characterized by a hydrodynamic length scale ℓ . Cells actively regulate ℓ via the activity of force-generating molecules, such as myosin II. Here we develop a general theory for such systems through a coarse-grained hydrodynamic approach including activity in the static description of the system providing an experimentally accessible parameter and elucidate the detailed mechanism of how a living system can actively self-regulate its hydrodynamic length scale, controlling the rigidity of the system. Remarkably, we find that ℓ , as a function of activity, behaves universally and roughly inversely proportional to the activity of the system. Our theory rationalizes a number of experimental findings on diverse systems, and comparison of our theory with existing experimental data shows good agreement.

Capture of microRNA-bound mRNAs identifies the tumor suppressor miR-34a as a regulator of growth factor signaling.

PubMed

Lal, Ashish; Thomas, Marshall P; Altschuler, Gabriel; Navarro, Francisco; O'Day, Elizabeth; Li, Xiao Ling; Concepcion, Carla; Han, Yoon-Chi; Thiery, Jerome; Rajani, Danielle K; Deutsch, Aaron; Hofmann, Oliver; Ventura, Andrea; Hide, Winston; Lieberman, Judy

2011-11-01

A simple biochemical method to isolate mRNAs pulled down with a transfected, biotinylated microRNA was used to identify direct target genes of miR-34a, a tumor suppressor gene. The method reidentified most of the known miR-34a regulated genes expressed in K562 and HCT116 cancer cell lines. Transcripts for 982 genes were enriched in the pull-down with miR-34a in both cell lines. Despite this large number, validation experiments suggested that ~90% of the genes identified in both cell lines can be directly regulated by miR-34a. Thus miR-34a is capable of regulating hundreds of genes. The transcripts pulled down with miR-34a were highly enriched for their roles in growth factor signaling and cell cycle progression. These genes form a dense network of interacting gene products that regulate multiple signal transduction pathways that orchestrate the proliferative response to external growth stimuli. Multiple candidate miR-34a-regulated genes participate in RAS-RAF-MAPK signaling. Ectopic miR-34a expression reduced basal ERK and AKT phosphorylation and enhanced sensitivity to serum growth factor withdrawal, while cells genetically deficient in miR-34a were less sensitive. Fourteen new direct targets of miR-34a were experimentally validated, including genes that participate in growth factor signaling (ARAF and PIK3R2) as well as genes that regulate cell cycle progression at various phases of the cell cycle (cyclins D3 and G2, MCM2 and MCM5, PLK1 and SMAD4). Thus miR-34a tempers the proliferative and pro-survival effect of growth factor stimulation by interfering with growth factor signal transduction and downstream pathways required for cell division.

The microbiota regulates neutrophil homeostasis and host resistance to Escherichia coli K1 sepsis in neonatal mice.

PubMed

Deshmukh, Hitesh S; Liu, Yuhong; Menkiti, Ogechukwu R; Mei, Junjie; Dai, Ning; O'Leary, Claire E; Oliver, Paula M; Kolls, Jay K; Weiser, Jeffrey N; Worthen, G Scott

2014-05-01

Neonatal colonization by microbes, which begins immediately after birth, is influenced by gestational age and the mother's microbiota and is modified by exposure to antibiotics. In neonates, prolonged duration of antibiotic therapy is associated with increased risk of late-onset sepsis (LOS), a disorder controlled by neutrophils. A role for the microbiota in regulating neutrophil development and susceptibility to sepsis in the neonate remains unclear. We exposed pregnant mouse dams to antibiotics in drinking water to limit transfer of maternal microbes to the neonates. Antibiotic exposure of dams decreased the total number and composition of microbes in the intestine of the neonates. This was associated with decreased numbers of circulating and bone marrow neutrophils and granulocyte/macrophage-restricted progenitor cells in the bone marrow of antibiotic-treated and germ-free neonates. Antibiotic exposure of dams reduced the number of interleukin-17 (IL-17)-producing cells in the intestine and production of granulocyte colony-stimulating factor (G-CSF). Granulocytopenia was associated with impaired host defense and increased susceptibility to Escherichia coli K1 and Klebsiella pneumoniae sepsis in antibiotic-treated neonates, which could be partially reversed by administration of G-CSF. Transfer of a normal microbiota into antibiotic-treated neonates induced IL-17 production by group 3 innate lymphoid cells (ILCs) in the intestine, increasing plasma G-CSF levels and neutrophil numbers in a Toll-like receptor 4 (TLR4)- and myeloid differentiation factor 88 (MyD88)-dependent manner and restored IL-17-dependent resistance to sepsis. Specific depletion of ILCs prevented IL-17- and G-CSF-dependent granulocytosis and resistance to sepsis. These data support a role for the intestinal microbiota in regulation of granulocytosis, neutrophil homeostasis and host resistance to sepsis in neonates.

Use of whole genome expression analysis in the toxicity screening of nanoparticles

DOE Office of Scientific and Technical Information (OSTI.GOV)

Fröhlich, Eleonore, E-mail: eleonore.froehlich@medunigraz.at; Meindl, Claudia; Wagner, Karin

2014-10-15

The use of nanoparticles (NPs) offers exciting new options in technical and medical applications provided they do not cause adverse cellular effects. Cellular effects of NPs depend on particle parameters and exposure conditions. In this study, whole genome expression arrays were employed to identify the influence of particle size, cytotoxicity, protein coating, and surface functionalization of polystyrene particles as model particles and for short carbon nanotubes (CNTs) as particles with potential interest in medical treatment. Another aim of the study was to find out whether screening by microarray would identify other or additional targets than commonly used cell-based assays formore » NP action. Whole genome expression analysis and assays for cell viability, interleukin secretion, oxidative stress, and apoptosis were employed. Similar to conventional assays, microarray data identified inflammation, oxidative stress, and apoptosis as affected by NP treatment. Application of lower particle doses and presence of protein decreased the total number of regulated genes but did not markedly influence the top regulated genes. Cellular effects of CNTs were small; only carboxyl-functionalized single-walled CNTs caused appreciable regulation of genes. It can be concluded that regulated functions correlated well with results in cell-based assays. Presence of protein mitigated cytotoxicity but did not cause a different pattern of regulated processes. - Highlights: • Regulated functions were screened using whole genome expression assays. • Polystyrene particles regulated more genes than short carbon nanotubes. • Protein coating of polystyrene particles did not change regulation pattern. • Functions regulated by microarray were confirmed by cell-based assay.« less

Arf6 guanine-nucleotide exchange factor, cytohesin-2, interacts with actinin-1 to regulate neurite extension.

PubMed

Torii, Tomohiro; Miyamoto, Yuki; Nakamura, Kazuaki; Maeda, Masahiro; Yamauchi, Junji; Tanoue, Akito

2012-09-01

Proper regulation of morphological changes in neuronal cells is essential for their differentiation. Complex signaling mechanisms mediate a variety of morphological changes such as formation of neurites. It is well established that a number of small GTPases control neurite behavior before the connection with the target tissue. However, their regulatory mechanisms remain to be fully understood. Here, we show that the Arf6 guanine-nucleotide exchange factor (GEF), cytohesin-2 (CYTH2), interacts with the cytoskeletal protein actinin-1 (ACTN1) and regulates neurite extension in N1E-115 cells used as the model. Knockdown of ACTN1, as well as that of CYTH2, in cells inhibits cellular Arf6 activity and neurite extension. The C-terminal polybasic region of CYTH2 participates in interacting directly with the EFh2 domain of ACTN1. Expression of CYTH2 mutant deficient of the EFh2 domain in cells also inhibits Arf6 activation and neurite extension. Furthermore, FRET analysis detects that the respective interactive region peptides, tagged with cell-permeable short peptides, greatly decrease Arf6 activation at growth cones in a time-dependent manner. Collectively, the signaling through CYTH2 and ACTN1 properly regulates neurite extension in N1E-115 cells, demonstrating the unexpected interaction of CYTH2 and ACTN1 in the regulation of cellular Arf6 activity involved in neurite extension. Copyright © 2012 Elsevier Inc. All rights reserved.

DEPTOR regulates vascular endothelial cell activation and proinflammatory and angiogenic responses.

PubMed

Bruneau, Sarah; Nakayama, Hironao; Woda, Craig B; Flynn, Evelyn A; Briscoe, David M

2013-09-05

The maintenance of normal tissue homeostasis and the prevention of chronic inflammatory disease are dependent on the active process of inflammation resolution. In endothelial cells (ECs), proinflammation results from the activation of intracellular signaling responses and/or the inhibition of endogenous regulatory/pro-resolution signaling networks that, to date, are poorly defined. In this study, we find that DEP domain containing mTOR interacting protein (DEPTOR) is expressed in different microvascular ECs in vitro and in vivo, and using a small interfering RNA (siRNA) knockdown approach, we find that it regulates mammalian target of rapamycin complex 1 (mTORC1), extracellular signal-regulated kinase 1/2, and signal transducer and activator of transcription 1 activation in part through independent mechanisms. Moreover, using limited gene arrays, we observed that DEPTOR regulates EC activation including mRNA expression of the T-cell chemoattractant chemokines CXCL9, CXCL10, CXCL11, CX3CL1, CCL5, and CCL20 and the adhesion molecules intercellular adhesion molecule-1 and vascular cell adhesion molecule-1 (P < .05). DEPTOR siRNA-transfected ECs also bound increased numbers of peripheral blood mononuclear cells (P < .005) and CD3+ T cells (P < .005) in adhesion assays in vitro and had increased migration and angiogenic responses in spheroid sprouting (P < .01) and wound healing (P < .01) assays. Collectively, these findings define DEPTOR as a critical upstream regulator of EC activation responses and suggest that it plays an important role in endogenous mechanisms of anti-inflammation and pro-resolution.

A Systems Biology Approach Reveals that Tissue Tropism to West Nile Virus Is Regulated by Antiviral Genes and Innate Immune Cellular Processes

PubMed Central

Suthar, Mehul S.; Brassil, Margaret M.; Blahnik, Gabriele; McMillan, Aimee; Ramos, Hilario J.; Proll, Sean C.; Belisle, Sarah E.; Katze, Michael G.; Gale, Michael

2013-01-01

The actions of the RIG-I like receptor (RLR) and type I interferon (IFN) signaling pathways are essential for a protective innate immune response against the emerging flavivirus West Nile virus (WNV). In mice lacking RLR or IFN signaling pathways, WNV exhibits enhanced tissue tropism, indicating that specific host factors of innate immune defense restrict WNV infection and dissemination in peripheral tissues. However, the immune mechanisms by which the RLR and IFN pathways coordinate and function to impart restriction of WNV infection are not well defined. Using a systems biology approach, we defined the host innate immune response signature and actions that restrict WNV tissue tropism. Transcriptional profiling and pathway modeling to compare WNV-infected permissive (spleen) and nonpermissive (liver) tissues showed high enrichment for inflammatory responses, including pattern recognition receptors and IFN signaling pathways, that define restriction of WNV replication in the liver. Assessment of infected livers from Mavs−/−×Ifnar−/− mice revealed the loss of expression of several key components within the natural killer (NK) cell signaling pathway, including genes associated with NK cell activation, inflammatory cytokine production, and NK cell receptor signaling. In vivo analysis of hepatic immune cell infiltrates from WT mice demonstrated that WNV infection leads to an increase in NK cell numbers with enhanced proliferation, maturation, and effector action. In contrast, livers from Mavs−/−×Ifnar−/− infected mice displayed reduced immune cell infiltration, including a significant reduction in NK cell numbers. Analysis of cocultures of dendritic and NK cells revealed both cell-intrinsic and -extrinsic roles for the RLR and IFN signaling pathways to regulate NK cell effector activity. Taken together, these observations reveal a complex innate immune signaling network, regulated by the RLR and IFN signaling pathways, that drives tissue-specific antiviral effector gene expression and innate immune cellular processes that control tissue tropism to WNV infection. PMID:23544010

The effects and regulatory mechanism of RIP3 on RGC-5 necroptosis following elevated hydrostatic pressure.

PubMed

Shang, Lei; Ding, Wei; Li, Na; Liao, Lvshuang; Chen, Dan; Huang, Jufang; Xiong, Kun

2017-02-06

Necroptosis is a type of regulated cell death that has been implicated in various diseases. Receptor-interacting protein 3 (RIP3), a member of the RIP family, is an important mediator of the necroptotic pathway. Cleavage of RIP3 at Asp328 by caspase-8 abolishes the kinase activity of RIP3, which is critical for necroptosis. Moreover, RIP3 is significantly upregulated during the early stages of acute high intra-ocular pressure and oxygen glucose deprivation. In this study, the effects of RIP3 during elevated hydrostatic pressure (EHP) were investigated and the possible mechanism through which caspase-8 regulated RIP3 cleavage was explored. Flow cytometry analysis revealed that the number of EHP-induced necrotic retinal ganglion cell 5 (RGC-5) cells was reduced after RIP3-knockdown. Furthermore, malondialdehyde (MDA) levels and glycogen phosphorylase (PYGL) activity in normal RGC-5 cells were much higher than those in RIP3-knockdown cells after EHP. EHP-induced RGC-5 necrosis was significantly reduced after treatment with butylated hydroxyanisole (BHA), a reactive oxygen species (ROS) scavenger. MDA levels and PYGL activity were lower in normal RGC-5 cells than those in cells with caspase-8 inhibition after EHP. Western blot analysis demonstrated that the RIP3 cleavage product was upregulated in cells with caspase-8 inhibition. Additionally, flow cytometry analysis revealed that the number of EHP-induced necrotic RGC-5 cells was increased after caspase-8 inhibition. Our results suggested that RGC-5 necroptosis following EHP was mediated by RIP3 through induction of PYGL activity and subsequent ROS accumulation. Thus, caspase-8 may participate in the regulation of RGC-5 necroptosis via RIP3 cleavage. © The Author 2016. Published by Oxford University Press on behalf of the Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Preliminary study of the role of gastrointestinal endocrine cells in the maintenance of villous structure following X-irradiation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wyatt, M.G.; Hume, S.P.; Carr, K.E.

The mechanism of gastrointestinal villous damage following ionizing irradiation is complex. Various compartments within the gastrointestinal tract have in turn been considered important for the maintenance of normal villous structure. To date, however, evidence for a single overriding regulator of epithelial well-being is lacking. In this study, the role of the gastro-intestinal (enteroendocrine) cells is explored and comparison made between endocrine cell number and villous structure. Experiments were organized using both control and irradiated groups of mice. Two time points (1 and 3 days) and three radiation doses (6, 10 and 18Gy) were employed. A simple method for endocrine cellmore » identification and subsequent quantification is described. Endocrine cell number was then compared with villous surface detail, as seen with a scanning electron microscope (SEM). Results indicated a decrease in the endocrine cell number at all three radiation doses. Whereas at low doses endocrine cell recovery occurred between 1 and 3 days, at medium and high doses further decline was noticed. A similar pattern was seen when considering villous surface structure. It is suggested that both scanning electron microscopy and endocrine cell number provide a more sensitive indicator of gastrointestinal radiation damage than do current crypt counting techniques. In addition, a link between endocrine cell number and villous structure is proposed.« less

Insulin-producing cells could not mimic the physiological regulation of insulin secretion performed by pancreatic beta cells

PubMed Central

2013-01-01

Objective The aim of this study was to compare the difference between insulin-producing cells (IPCs) and normal human pancreatic beta cells both in physiological function and morphological features in cellular level. Methods The levels of insulin secretion were measured by enzyme-linked immunosorbent assay. The insulin gene expression was determined by real-time quantitative polymerase chain reaction. The morphological features were detected by atomic force microscopy (AFM) and laser confocal scanning microscopy. Results IPCs and normal human pancreatic beta cells were similar to each other under the observation in AFM with the porous structure features in the cytoplasm. Both number of membrane particle size and average roughness of normal human beta cells were higher than those of IPCs. Conclusions Our results firstly revealed that the cellular ultrastructure of IPCs was closer to that of normal human pancreatic beta cells, but they still could not mimic the physiological regulation of insulin secretion performed by pancreatic beta cells. PMID:23421382

Fatty acid trafficking in starved cells: regulation by lipid droplet lipolysis, autophagy, and mitochondrial fusion dynamics.

PubMed

Rambold, Angelika S; Cohen, Sarah; Lippincott-Schwartz, Jennifer

2015-03-23

Fatty acids (FAs) provide cellular energy under starvation, yet how they mobilize and move into mitochondria in starved cells, driving oxidative respiration, is unclear. Here, we clarify this process by visualizing FA trafficking with a fluorescent FA probe. The labeled FA accumulated in lipid droplets (LDs) in well-fed cells but moved from LDs into mitochondria when cells were starved. Autophagy in starved cells replenished LDs with FAs, increasing LD number over time. Cytoplasmic lipases removed FAs from LDs, enabling their transfer into mitochondria. This required mitochondria to be highly fused and localized near LDs. When mitochondrial fusion was prevented in starved cells, FAs neither homogeneously distributed within mitochondria nor became efficiently metabolized. Instead, FAs reassociated with LDs and fluxed into neighboring cells. Thus, FAs engage in complex trafficking itineraries regulated by cytoplasmic lipases, autophagy, and mitochondrial fusion dynamics, ensuring maximum oxidative metabolism and avoidance of FA toxicity in starved cells. Copyright © 2015 Elsevier Inc. All rights reserved.

Natural Compounds as Modulators of Cell Cycle Arrest: Application for Anticancer Chemotherapies

PubMed Central

Bailon-Moscoso, Natalia; Cevallos-Solorzano, Gabriela; Romero-Benavides, Juan Carlos; Orellana, Maria Isabel Ramirez

2017-01-01

Natural compounds from various plants, microorganisms and marine species play an important role in the discovery novel components that can be successfully used in numerous biomedical applications, including anticancer therapeutics. Since uncontrolled and rapid cell division is a hallmark of cancer, unraveling the molecular mechanisms underlying mitosis is key to understanding how various natural compounds might function as inhibitors of cell cycle progression. A number of natural compounds that inhibit the cell cycle arrest have proven effective for killing cancer cells in vitro, in vivo and in clinical settings. Significant advances that have been recently made in the understanding of molecular mechanisms underlying the cell cycle regulation using the chemotherapeutic agents is of great importance for improving the efficacy of targeted therapeutics and overcoming resistance to anticancer drugs, especially of natural origin, which inhibit the activities of cyclins and cyclin-dependent kinases, as well as other proteins and enzymes involved in proper regulation of cell cycle leading to controlled cell proliferation. PMID:28367072

Up-regulated expression of substance P in CD8+ T cells and NK1R on monocytes of atopic dermatitis.

PubMed

Zhang, Zenan; Zheng, Wenjiao; Xie, Hua; Chai, Ruonan; Wang, Junling; Zhang, Huiyun; He, Shaoheng

2017-05-01

Large numbers of CD8 + T cells were observed in atopic dermatitis (AD) skin, and monocytes from AD patients showed increased prostaglandin E2 production. However, little is known about the expression of substance P (SP) and its receptor NK1R in blood leukocytes of patients with AD. To explore the expression of SP and NK1R in leukocytes of AD and the influence of allergens on SP and NK1R expression. The expression levels of SP and NK1R in patients with AD were examined by flow cytometry, ELISA and a mouse AD model. The plasma SP level was 4.9-fold higher in patients with AD than in HC subjects. Both the percentage of SP expression in the population and mean fluorescence intensity (MFI) of SP expression were elevated in CD8 + T cells in the blood of AD patients. However, both the CD14 + NK1R + population and MFI of NK1R expression on CD14 + cells were enhanced in the blood of AD patients. Allergens ASWE, HDME and PPE failed to up-regulate SP expression in CD8 + T cells. However, allergens ASWE and HDME both enhanced NK1R expression on CD14 + blood leukocytes regardless of AD or HC subjects. OVA-sensitized AD mice showed an elevated proportion and MFI of SP-expressing CD8 + T cells in the blood, which agrees with the SP expression situation in human AD blood. Injection of SP into mouse skin did not up-regulate NK1R expression on monocytes. An elevated plasma SP level, up-regulated expression of SP and NK1R indicate that the SP/NK1R complex is important in the development of AD. Therefore, SP and NK1R antagonist or blocker agents may help to treat patients with AD. Trial registration Registration number: ChiCTR-BOC-16010279; Registration date: Dec., 28, 2016; retrospectively registered.

A novel mode of regulation of the Staphylococcus aureus Vancomycin-resistance-associated response regulator VraR mediated by Stk1 protein phosphorylation.

PubMed

Canova, Marc J; Baronian, Grégory; Brelle, Solène; Cohen-Gonsaud, Martin; Bischoff, Markus; Molle, Virginie

2014-04-25

The Staphylococcus aureus Vancomycin-resistance-associated response regulator VraR is known as an important response regulator, member of the VraTSR three-component signal transduction system that modulates the expression of the cell wall stress stimulon in response to a number of different cell wall active antibiotics. Given its crucial role in regulating gene expression in response to antibiotic challenges, VraR must be tightly regulated. We report here for the first time in S. aureus convergence of two major signal transduction systems, serine/threonine protein kinase and two (three)-component systems. We demonstrate that VraR can be phosphorylated by the staphylococcal Ser/Thr protein kinase Stk1 and that phosphorylation negatively affects its DNA-binding properties. Mass spectrometric analyses and site-directed mutagenesis identified Thr106, Thr119, Thr175 and Thr178 as phosphoacceptors. A S. aureus ΔvraR mutant expressing a VraR derivative that mimics constitutive phosphorylation, VraR_Asp, still exhibited markedly decreased antibiotic resistance against different cell wall active antibiotics, when compared to the wild-type, suggesting that VraR phosphorylation may represent a novel and presumably more general mechanism of regulation of the two (three)-component systems in staphylococci. Copyright © 2014 Elsevier Inc. All rights reserved.

Quantitative Cell Cycle Analysis Based on an Endogenous All-in-One Reporter for Cell Tracking and Classification.

PubMed

Zerjatke, Thomas; Gak, Igor A; Kirova, Dilyana; Fuhrmann, Markus; Daniel, Katrin; Gonciarz, Magdalena; Müller, Doris; Glauche, Ingmar; Mansfeld, Jörg

2017-05-30

Cell cycle kinetics are crucial to cell fate decisions. Although live imaging has provided extensive insights into this relationship at the single-cell level, the limited number of fluorescent markers that can be used in a single experiment has hindered efforts to link the dynamics of individual proteins responsible for decision making directly to cell cycle progression. Here, we present fluorescently tagged endogenous proliferating cell nuclear antigen (PCNA) as an all-in-one cell cycle reporter that allows simultaneous analysis of cell cycle progression, including the transition into quiescence, and the dynamics of individual fate determinants. We also provide an image analysis pipeline for automated segmentation, tracking, and classification of all cell cycle phases. Combining the all-in-one reporter with labeled endogenous cyclin D1 and p21 as prime examples of cell-cycle-regulated fate determinants, we show how cell cycle and quantitative protein dynamics can be simultaneously extracted to gain insights into G1 phase regulation and responses to perturbations. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.

Oscillatory Protein Expression Dynamics Endows Stem Cells with Robust Differentiation Potential

PubMed Central

Kaneko, Kunihiko

2011-01-01

The lack of understanding of stem cell differentiation and proliferation is a fundamental problem in developmental biology. Although gene regulatory networks (GRNs) for stem cell differentiation have been partially identified, the nature of differentiation dynamics and their regulation leading to robust development remain unclear. Herein, using a dynamical system modeling cell approach, we performed simulations of the developmental process using all possible GRNs with a few genes, and screened GRNs that could generate cell type diversity through cell-cell interactions. We found that model stem cells that both proliferated and differentiated always exhibited oscillatory expression dynamics, and the differentiation frequency of such stem cells was regulated, resulting in a robust number distribution. Moreover, we uncovered the common regulatory motifs for stem cell differentiation, in which a combination of regulatory motifs that generated oscillatory expression dynamics and stabilized distinct cellular states played an essential role. These findings may explain the recently observed heterogeneity and dynamic equilibrium in cellular states of stem cells, and can be used to predict regulatory networks responsible for differentiation in stem cell systems. PMID:22073296

Regulation of miR-21 expression in human melanoma via UV-ray-induced melanin pigmentation.

PubMed

Lin, Kuan-Yu; Chen, Chien-Min; Lu, Cheng-You; Cheng, Chun-Yuan; Wu, Yu-Hsin

2017-08-01

Excessive environmental ultraviolet (UV) radiation produces genetic mutations that can lead to skin cancer. This study was designed to assess the potential inhibitory activity of microRNA-21 (miR-21) on the UV irradiation-stimulated melanogenesis signal pathway in melanoma cells. The molecular mechanism of miR-21-induced inhibitory activity on UV-ray-stimulated melanogenesis-regulating proteins was examined in A375.S2 human melanoma and B16F10 mouse melanoma cells. UV irradiation for 30 min induced melanogenesis signal pathway by increasing melanin production and the number of A375.S2 cells. Similarly, UV radiation increased the expression of α-melanocyte-stimulating hormone (α-MSH) protein and decreased the melanogenesis-regulating signal, such as EGFR and Akt phosphorylation. Notably, miR-21 overexpression in UV-ray-stimulated A375.S2 cells decreased α-MSH expression and increased EGFR and Akt phosphorylation levels. Furthermore, miR-21 on UV-ray- induced melanogenesis was down-regulated by the Akt inhibitor and the EGFR inhibitor (Gefitinib). Results suggest that the suppressive activity of miR-21 on UV-ray-stimulated melanogenesis may involve the down-regulation of α-MSH and the activation in both of EGFR and Akt. © 2017 Wiley Periodicals, Inc.

Keratinocyte differentiation is regulated by the Rho and ROCK signaling pathway.

PubMed

McMullan, Rachel; Lax, Siân; Robertson, Vicki H; Radford, David J; Broad, Simon; Watt, Fiona M; Rowles, Alison; Croft, Daniel R; Olson, Michael F; Hotchin, Neil A

2003-12-16

The epidermis comprises multiple layers of specialized epithelial cells called keratinocytes. As cells are lost from the outermost epidermal layers, they are replaced through terminal differentiation, in which keratinocytes of the basal layer cease proliferating, migrate upwards, and eventually reach the outermost cornified layers. Normal homeostasis of the epidermis requires that the balance between proliferation and differentiation be tightly regulated. The GTP binding protein RhoA plays a fundamental role in the regulation of the actin cytoskeleton and in the adhesion events that are critically important to normal tissue homeostasis. Two central mediators of the signals from RhoA are the ROCK serine/threonine kinases ROCK-I and ROCK-II. We have analyzed ROCK's role in the regulation of epidermal keratinocyte function by using a pharmacological inhibitor and expressing conditionally active or inactive forms of ROCK-II in primary human keratinocytes. We report that blocking ROCK function results in inhibition of keratinocyte terminal differentiation and an increase in cell proliferation. In contrast, activation of ROCK-II in keratinocytes results in cell cycle arrest and an increase in the expression of a number of genes associated with terminal differentiation. Thus, these results indicate that ROCK plays a critical role in regulating the balance between proliferation and differentiation in human keratinocytes.

Generating autologous hematopoietic cells from human-induced pluripotent stem cells through ectopic expression of transcription factors.

PubMed

Hwang, Yongsung; Broxmeyer, Hal E; Lee, Man Ryul

2017-07-01

Hematopoietic cell transplantation (HCT) is a successful treatment modality for patients with malignant and nonmalignant disorders, usually when no other treatment option is available. The cells supporting long-term reconstitution after HCT are the hematopoietic stem cells (HSCs), which can be limited in numbers. Moreover, finding an appropriate human leukocyte antigen-matched donor can be problematic. If HSCs can be stably produced in large numbers from autologous or allogeneic cell sources, it would benefit HCT. Induced pluripotent stem cells (iPSCs) established from patients' own somatic cells can be differentiated into hematopoietic cells in vitro. This review will highlight recent methods for regulating human (h) iPSC production of HSCs and more mature blood cells. Advancements in transcription factor-mediated regulation of the developmental stages of in-vivo hematopoietic lineage commitment have begun to provide an understanding of the molecular mechanism of hematopoiesis. Such studies involve not only directed differentiation in which transcription factors, specifically expressed in hematopoietic lineage-specific cells, are overexpressed in iPSCs, but also direct conversion in which transcription factors are introduced into patient-derived somatic cells which are dedifferentiated to hematopoietic cells. As iPSCs derived from patients suffering from genetically mutated diseases would express the same mutated genetic information, CRISPR-Cas9 gene editing has been utilized to differentiate genetically corrected iPSCs into normal hematopoietic cells. IPSCs provide a model for molecular understanding of disease, and also may function as a cell population for therapy. Efficient differentiation of patient-specific iPSCs into HSCs and progenitor cells is a potential means to overcome limitations of such cells for HCT, as well as for providing in-vitro drug screening templates as tissue-on-a-chip models.

Random Signal Fluctuations Can Reduce Random Fluctuations in Regulated Components of Chemical Regulatory Networks

NASA Astrophysics Data System (ADS)

Paulsson, Johan; Ehrenberg, Måns

2000-06-01

Many intracellular components are present in low copy numbers per cell and subject to feedback control. We use chemical master equations to analyze a negative feedback system where species X and S regulate each other's synthesis with standard intracellular kinetics. For a given number of X-molecules, S-variation can be significant. We show that this signal noise does not necessarily increase X-variation as previously thought but, surprisingly, can be necessary to reduce it below a Poissonian limit. The principle resembles Stochastic Resonance in that signal noise improves signal detection.

Polyploidization of liver cells.

PubMed

Celton-Morizur, Séverine; Desdouets, Chantal

2010-01-01

Eukaryotic organisms usually contain a diploid complement of chromosomes. However, there are a number of exceptions. Organisms containing an increase in DNA content by whole number multiples of the entire set of chromosomes are defined as polyploid. Cells that contain more than two sets of chromosomes were first observed in plants about a century ago and it is now recognized that polyploidy cells form in many eukaryotes under a wide variety of circumstance. Although it is less common in mammals, some tissues, including the liver, show a high percentage of polyploid cells. Thus, during postnatal growth, the liver parenchyma undergoes dramatic changes characterized by gradual polyploidization during which hepatocytes of several ploidy classes emerge as a result of modified cell-division cycles. This process generates the successive appearance of tetraploid and octoploid cell classes with one or two nuclei (mononucleated or binucleated). Liver cells polyploidy is generally considered to indicate terminal differentiation and senescence and to lead both to the progressive loss of cell pluripotency and a markedly decreased replication capacity. In adults, liver polyploidization is differentially regulated upon loss of liver mass and liver damage. Interestingly, partial hepatectomy induces marked cell proliferation followed by an increase in liver ploidy. In contrast, during hepatocarcinoma (HCC), growth shifts to a nonpolyploidizing pattern and expansion of the diploid hepatocytes population is observed in neoplastic nodules. Here we review the current state of understanding about how polyploidization is regulated during normal and pathological liver growth and detail by which mechanisms hepatocytes become polyploid.

Leukocyte-specific protein 1 regulates T-cell migration in rheumatoid arthritis

PubMed Central

Hwang, Seong-Hye; Jung, Seung-Hyun; Lee, Saseong; Choi, Susanna; Yoo, Seung-Ah; Park, Ji-Hwan; Hwang, Daehee; Shim, Seung Cheol; Sabbagh, Laurent; Kim, Ki-Jo; Park, Sung Hwan; Cho, Chul-Soo; Kim, Bong-Sung; Leng, Lin; Montgomery, Ruth R.; Bucala, Richard; Chung, Yeun-Jun; Kim, Wan-Uk

2015-01-01

Copy number variations (CNVs) have been implicated in human diseases. However, it remains unclear how they affect immune dysfunction and autoimmune diseases, including rheumatoid arthritis (RA). Here, we identified a novel leukocyte-specific protein 1 (LSP1) deletion variant for RA susceptibility located in 11p15.5. We replicated that the copy number of LSP1 gene is significantly lower in patients with RA, which correlates positively with LSP1 protein expression levels. Differentially expressed genes in Lsp1-deficient primary T cells represent cell motility and immune and cytokine responses. Functional assays demonstrated that LSP1, induced by T-cell receptor activation, negatively regulates T-cell migration by reducing ERK activation in vitro. In mice with T-cell–dependent chronic inflammation, loss of Lsp1 promotes migration of T cells into the target tissues as well as draining lymph nodes, exacerbating disease severity. Moreover, patients with RA show diminished expression of LSP1 in peripheral T cells with increased migratory capacity, suggesting that the defect in LSP1 signaling lowers the threshold for T-cell activation. To our knowledge, our work is the first to demonstrate how CNVs result in immune dysfunction and a disease phenotype. Particularly, our data highlight the importance of LSP1 CNVs and LSP1 insufficiency in the pathogenesis of RA and provide previously unidentified insights into the mechanisms underlying T-cell migration toward the inflamed synovium in RA. PMID:26554018

YAP regulates neuronal differentiation through Sonic hedgehog signaling pathway

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lin, Yi-Ting; Ding, Jing-Ya; Li, Ming-Yang

2012-09-10

Tight regulation of cell numbers by controlling cell proliferation and apoptosis is important during development. Recently, the Hippo pathway has been shown to regulate tissue growth and organ size in Drosophila. In mammalian cells, it also affects cell proliferation and differentiation in various tissues, including the nervous system. Interplay of several signaling cascades, such as Notch, Wnt, and Sonic Hedgehog (Shh) pathways, control cell proliferation during neuronal differentiation. However, it remains unclear whether the Hippo pathway coordinates with other signaling cascades in regulating neuronal differentiation. Here, we used P19 cells, a mouse embryonic carcinoma cell line, as a model tomore » study roles of YAP, a core component of the Hippo pathway, in neuronal differentiation. P19 cells can be induced to differentiate into neurons by expressing a neural bHLH transcription factor gene Ascl1. Our results showed that YAP promoted cell proliferation and inhibited neuronal differentiation. Expression of Yap activated Shh but not Wnt or Notch signaling activity during neuronal differentiation. Furthermore, expression of Yap increased the expression of Patched homolog 1 (Ptch1), a downstream target of the Shh signaling. Knockdown of Gli2, a transcription factor of the Shh pathway, promoted neuronal differentiation even when Yap was over-expressed. We further demonstrated that over-expression of Yap inhibited neuronal differentiation in primary mouse cortical progenitors and Gli2 knockdown rescued the differentiation defect in Yap over-expressing cells. In conclusion, our study reveals that Shh signaling acts downstream of YAP in regulating neuronal differentiation. -- Highlights: Black-Right-Pointing-Pointer YAP promotes cell proliferation and inhibits neuronal differentiation in P19 cells. Black-Right-Pointing-Pointer YAP promotes Sonic hedgehog signaling activity during neuronal differentiation. Black-Right-Pointing-Pointer Knockdown of Gli2 rescues the Yap-overexpression phenotype in P19 cells. Black-Right-Pointing-Pointer Knockdown of Gli2 rescues the Yap-overexpression phenotype in cortical progenitors.« less

Expression and regulation of proton-coupled oligopeptide transporters in colonic tissue and immune cells of mice.

PubMed

Wang, Yuqing; Hu, Yongjun; Li, Ping; Weng, Yayun; Kamada, Nobuhiko; Jiang, Huidi; Smith, David E

2018-02-01

A number of studies have implicated proton-coupled oligopeptide transporters (POTs) in the initiation and/or progression of inflammatory bowel disease and immune cell signaling. With this in mind, the aim of this study was to delineate the expression of POTs in mouse colonic tissues and immune cells, and characterize the potential role of these transporters in nucleotide-binding oligomerization domain (NOD) signaling. Using a dextran sodium sulfate (DSS)-induced colitis mouse model, we found that DSS down regulated Pht1 gene expression and up regulated Pht2 gene expression in colonic tissue and immune cells. In contrast, PEPT1 protein was absent from the colonic tissue and immune cells of normal and DSS-treated mice. NOD ligands, muramyl dipeptide (MDP) and l-Ala-γ-d-Glu-meso-diaminopimelic acid (tri-DAP), were shown to be substrates of PHT2 in MDCK-hPHT2 19,20AA cells. Subsequent studies revealed that the immune response of lamina propia mononuclear cells may be regulated by PHT1 and PHT2, and that PHT2 facilitated the NOD-dependent immune response in RAW264.7 macrophages. These results clarified the expression of POTs in mouse colonic segments, cells and subtypes, and the role of increased Pht2 expression during chemically-induced colitis in facilitating NOD-dependent immune response. The findings further suggest that intestinal PHT2 may serve as a therapeutic target for IBD therapy. Copyright © 2018 Elsevier Inc. All rights reserved.

Methods to study maternal regulation of germ cell specification in zebrafish

PubMed Central

Kaufman, O.H.; Marlow, F.L.

2016-01-01

The process by which the germ line is specified in the zebrafish embryo is under the control of maternal gene products that were produced during oogenesis. Zebrafish are highly amenable to microscopic observation of the processes governing maternal germ cell specification because early embryos are transparent, and the germ line is specified rapidly (within 4–5 h post fertilization). Advantages of zebrafish over other models used to study vertebrate germ cell formation include their genetic tractability, the large numbers of progeny, and the easily manipulable genome, all of which make zebrafish an ideal system for studying the genetic regulators and cellular basis of germ cell formation and maintenance. Classical molecular biology techniques, including expression analysis through in situ hybridization and forward genetic screens, have laid the foundation for our understanding of germ cell development in zebrafish. In this chapter, we discuss some of these classic techniques, as well as recent cutting-edge methodologies that have improved our ability to visualize the process of germ cell specification and differentiation, and the tracking of specific molecules involved in these processes. Additionally, we discuss traditional and novel technologies for manipulating the zebrafish genome to identify new components through loss-of-function studies of putative germ cell regulators. Together with the numerous aforementioned advantages of zebrafish as a genetic model for studying development, we believe these new techniques will continue to advance zebrafish to the forefront for investigation of the molecular regulators of germ cell specification and germ line biology. PMID:27312489

Bone Morphogenetic Protein Regulation of Enteric Neuronal Phenotypic Diversity: Relationship to Timing of Cell Cycle Exit

PubMed Central

Chalazonitis, Alcmène; Pham, Tuan.D.; Li, Zhishan; Roman, Daniel; Guha, Udayan; Gomes, William; Kan, Lixin; Kessler, John A.; Gershon, Michael D.

2008-01-01

The effects of bone morphogenetic protein (BMP) signaling on enteric neuron development were examined in transgenic mice over expressing either the BMP inhibitor, noggin, or BMP4 under control of the neuron specific enolase (NSE) promoter. Noggin antagonism of BMP signaling increased total numbers of enteric neurons and those of subpopulations derived from precursors that exit the cell cycle early in neurogenesis (serotonin, calretinin, calbindin). In contrast, noggin overexpression decreased numbers of neurons derived from precursors that exit the cell cycle late (γ-aminobutyric acid, tyrosine hydroxylase [TH], dopamine transporter, calcitonin gene related peptide, TrkC). Numbers of TH- and TrkC-expressing neurons were increased by overexpression of BMP4. These observations are consistent with the idea that phenotypic expression in the enteric nervous system (ENS) is determined, in part, by the number of proliferative divisions neuronal precursors undergo before their terminal mitosis. BMP signaling may thus regulate enteric neuronal phenotypic diversity by promoting the exit of precursors from the cell cycle. BMP2 increased the numbers of TH- and TrkC-expressing neurons developing in vitro from immunoselected enteric crest-derived precursors; BMP signaling may thus also specify or promote the development of dopaminergic TrkC/NT-3-dependent neurons. The developmental defects in the ENS of noggin overexpressing mice caused a relatively mild disturbance of motility (irregular rapid transit and increased stool frequency, weight, and water content). Although the function of the gut thus displays a remarkable tolerance for ENS defects, subtle functional abnormalities in motility or secretion may arise when ENS defects short of aganglionosis occur during development. PMID:18537141

Glycogen synthase kinase-3α/β inhibition promotes in vivo amplification of endogenous mesenchymal progenitors with osteogenic and adipogenic potential and their differentiation to the osteogenic lineage.

PubMed

Gambardella, Alessandra; Nagaraju, Chandan K; O'Shea, Patrick J; Mohanty, Sindhu T; Kottam, Lucksy; Pilling, James; Sullivan, Michael; Djerbi, Mounira; Koopmann, Witte; Croucher, Peter I; Bellantuono, Ilaria

2011-04-01

Small molecules are attractive therapeutics to amplify and direct differentiation of stem cells. They also can be used to understand the regulation of their fate by interfering with specific signaling pathways. Mesenchymal stem cells (MSCs) have the potential to proliferate and differentiate into several cell types, including osteoblasts. Activation of canonical Wnt signaling by inhibition of glycogen synthase kinase 3 (GSK-3) has been shown to enhance bone mass, possibly by involving a number of mechanisms ranging from amplification of the mesenchymal stem cell pool to the commitment and differentiation of osteoblasts. Here we have used a highly specific novel inhibitor of GSK-3, AR28, capable of inducing β-catenin nuclear translocation and enhanced bone mass after 14 days of treatment in BALB/c mice. We have shown a temporally regulated increase in the number of colony-forming units-osteoblast (CFU-O) and -adipocyte (CFU-A) but not colony-forming units-fibroblast (CFU-F) in mice treated for 3 days. However, the number of CFU-O and CFU-A returned to normal levels after 14 days of treatment, and the number of CFU-F was decreased significantly. In contrast, the number of osteoblasts increased significantly only after 14 days of treatment, and this was seen together with a significant decrease in bone marrow adiposity. These data suggest that the increased bone mass is the result of an early temporal wave of amplification of a subpopulation of MSCs with both osteogenic and adipogenic potential, which is driven to osteoblast differentiation at the expense of adipogenesis. Copyright © 2011 American Society for Bone and Mineral Research.

gone early, a Novel Germline Factor, Ensures the Proper Size of the Stem Cell Precursor Pool in the Drosophila Ovary

PubMed Central

Matsuoka, Shinya; Gupta, Swati; Suzuki, Emiko; Hiromi, Yasushi; Asaoka, Miho

2014-01-01

In order to sustain lifelong production of gametes, many animals have evolved a stem cell–based gametogenic program. In the Drosophila ovary, germline stem cells (GSCs) arise from a pool of primordial germ cells (PGCs) that remain undifferentiated even after gametogenesis has initiated. The decision of PGCs to differentiate or remain undifferentiated is regulated by somatic stromal cells: specifically, epidermal growth factor receptor (EGFR) signaling activated in the stromal cells determines the fraction of germ cells that remain undifferentiated by shaping a Decapentaplegic (Dpp) gradient that represses PGC differentiation. However, little is known about the contribution of germ cells to this process. Here we show that a novel germline factor, Gone early (Goe), limits the fraction of PGCs that initiate gametogenesis. goe encodes a non-peptidase homologue of the Neprilysin family metalloendopeptidases. At the onset of gametogenesis, Goe was localized on the germ cell membrane in the ovary, suggesting that it functions in a peptidase-independent manner in cell–cell communication at the cell surface. Overexpression of Goe in the germline decreased the number of PGCs that enter the gametogenic pathway, thereby increasing the proportion of undifferentiated PGCs. Inversely, depletion of Goe increased the number of PGCs initiating differentiation. Excess PGC differentiation in the goe mutant was augmented by halving the dose of argos, a somatically expressed inhibitor of EGFR signaling. This increase in PGC differentiation resulted in a massive decrease in the number of undifferentiated PGCs, and ultimately led to insufficient formation of GSCs. Thus, acting cooperatively with a somatic regulator of EGFR signaling, the germline factor goe plays a critical role in securing the proper size of the GSC precursor pool. Because goe can suppress EGFR signaling activity and is expressed in EGF-producing cells in various tissues, goe may function by attenuating EGFR signaling, and thereby affecting the stromal environment. PMID:25420147

Failure To Recruit Anti-Inflammatory CD103+ Dendritic Cells and a Diminished CD4+ Foxp3+ Regulatory T Cell Pool in Mice That Display Excessive Lung Inflammation and Increased Susceptibility to Mycobacterium tuberculosis

PubMed Central

Leepiyasakulchai, Chaniya; Ignatowicz, Lech; Pawlowski, Andrzej; Källenius, Gunilla

2012-01-01

Susceptibility to Mycobacterium tuberculosis is characterized by excessive lung inflammation, tissue damage, and failure to control bacterial growth. To increase our understanding of mechanisms that may regulate the host immune response in the lungs, we characterized dendritic cells expressing CD103 (αE integrin) (αE-DCs) and CD4+ Foxp3+ regulatory T (Treg) cells during M. tuberculosis infection. In resistant C57BL/6 and BALB/c mice, the number of lung αE-DCs increased dramatically during M. tuberculosis infection. In contrast, highly susceptible DBA/2 mice failed to recruit αE-DCs even during chronic infection. Even though tumor necrosis factor alpha (TNF-α) is produced by multiple DCs and macrophage subsets and is required for control of bacterial growth, αE-DCs remained TNF-α negative. Instead, αE-DCs contained a high number of transforming growth factor beta-producing cells in infected mice. Further, we show that Treg cells in C57BL/6 and DBA/2 mice induce gamma interferon during pulmonary tuberculosis. In contrast to resistant mice, the Treg cell population was diminished in the lungs, but not in the draining pulmonary lymph nodes (PLN), of highly susceptible mice during chronic infection. Treg cells have been reported to inhibit M. tuberculosis-specific T cell immunity, leading to increased bacterial growth. Still, despite the reduced number of lung Treg cells in DBA/2 mice, the bacterial load in the lungs was increased compared to resistant animals. Our results show that αE-DCs and Treg cells that may regulate the host immune response are increased in M. tuberculosis-infected lungs of resistant mice but diminished in infected lungs of susceptible mice. PMID:22215739

Regulation of the expression of the cell-cycle gene ftsZ by DicF antisense RNA. Division does not require a fixed number of FtsZ molecules.

PubMed

Tétart, F; Bouché, J P

1992-03-01

We show that the 53-nucleotide RNA molecule encoded by gene dicF blocks cell division in Escherichia coli by inhibiting the translation of ftsZ mRNA. Such a role for dicF had been predicted on the basis of the complementarity of DicF RNA with the ribosome-binding region of the ftsZ mRNA. An analysis of ftsZ expression at its chromosomal locus, and of an ftsZ-lacZ translational fusion controlled by promoters ftsZ1p and ftsZ2p only, indicates that ftsZ is not autoregulated. Partial inhibition of FtsZ synthesis leads to increased cell size. However, the number of FtsZ molecules per cell can be reduced threefold without affecting the division rate significantly. Our results suggest that septation is not triggered by a fixed number of newly synthesized FtsZ molecules per cell.

Regulation of CD4+ T-Cell Function by Membrane Cholesterol

DTIC Science & Technology

2012-03-13

hypercholesterolemia in mice was associated with an increased number of 97 splenic CD4 T-cells [65]. In contrast, Atorvastatin -induced inhibition of HMG...Mauri C, Ehrenstein MR (2006) Atorvastatin restores Lck expression and lipid raft-associated signaling in T cells from patients with systemic lupus...J, Rehm C, et al. (2011) High dose atorvastatin decreases cellular markers of immune activation without affecting HIV-1 RNA levels: results of a

Folic acid induces cell type-specific changes in the transcriptome of breast cancer cell lines: a proof-of-concept study.

PubMed

Price, R Jordan; Lillycrop, Karen A; Burdge, Graham C

2016-01-01

The effect of folic acid (FA) on breast cancer (BC) risk is uncertain. We hypothesised that this uncertainty may be due, in part, to differential effects of FA between BC cells with different phenotypes. To test this we investigated the effect of treatment with FA concentrations within the range of unmetabolised FA reported in humans on the expression of the transcriptome of non-transformed (MCF10A) and cancerous (MCF7 and Hs578T) BC cells. The total number of transcripts altered was: MCF10A, seventy-five (seventy up-regulated); MCF7, twenty-four (fourteen up-regulated); and Hs578T, 328 (156 up-regulated). Only the cancer-associated gene TAGLN was altered by FA in all three cell lines. In MCF10A and Hs578T cells, FA treatment decreased pathways associated with apoptosis, cell death and senescence, but increased those associated with cell proliferation. The folate transporters SLC19A1, SLC46A1 and FOLR1 were differentially expressed between cell lines tested. However, the level of expression was not altered by FA treatment. These findings suggest that physiological concentrations of FA can induce cell type-specific changes in gene regulation in a manner that is consistent with proliferative phenotype. This has implications for understanding the role of FA in BC risk. In addition, these findings support the suggestion that differences in gene expression induced by FA may involve differential activities of folate transporters. Together these findings indicate the need for further studies of the effect of FA on BC.

Appetitive and consummatory sexual and agonistic behaviour elicits FOS expression in aromatase and vasotocin neurones within the preoptic area and bed nucleus of the stria terminalis of male domestic chickens.

PubMed

Xie, J; Kuenzel, W J; Sharp, P J; Jurkevich, A

2011-03-01

Some components of male sexual and agonistic behaviours are considered to be regulated by the same neurocircuitry in the medial preoptic nucleus (POM) and the medial portion of bed nucleus of the stria terminalis (BSTM). To better understand this neurocircuitry, numbers of aromatase- (ARO) or arginine vasotocin- (AVT) immunoreactive (ir) neurones expressing immediate early gene protein FOS were compared in the POM and BSTM of male chickens following sexual or agonistic behaviours. Observations were made on males showing: (i) appetitive (courtship) and consummatory (copulation) sexual behaviours; (ii) only appetitive sexual behaviour, or (iii) displaying agonistic behaviour toward other males. Control males were placed on their own in the observation pen, or only handled. In the POM, appetitive sexual behaviour increased ARO+FOS colocalisation, whereas agonistic behaviour decreased the number of visible ARO-ir cells. In the dorsolateral subdivision of BSTM (BSTM1), appetitive sexual behaviour also increased ARO+FOS colocalisation, although the numbers of visible ARO-ir and AVT-ir cells were not altered by sexual or agonistic behaviours. In the ventromedial BSTM (BSTM2), appetitive sexual behaviour increased ARO+FOS and AVT+FOS colocalisation, and all behaviours decreased the number of visible ARO-ir cells, particularly in males expressing consummatory sexual behaviour. Positive correlations were found between numbers of cells with ARO+FOS and AVT+FOS colocalisation in both subdivisions of the BSTM. Waltzing frequency was positively correlated with ARO+FOS colocalisation in the lateral POM, and in both subdivisions of the BSTM in males expressing sexual behaviour. Waltzing frequency in males expressing agonistic behaviour was negatively correlated with the total number of visible ARO-ir cells in the lateral POM and BSTM2. These observations suggest a key role for ARO and AVT neurones in BSTM2 in the expression of appetitive sexual behaviour, and differential roles for ARO cells in the POM and BSTM in the regulation of components of sexual and agonistic behaviours. © 2011 The Authors. Journal of Neuroendocrinology © 2011 Blackwell Publishing Ltd.

The Coordinated Actions of TIM-3 on Cancer and Myeloid Cells in the Regulation of Tumorigenicity and Clinical Prognosis in Clear Cell Renal Cell Carcinomas.

PubMed

Komohara, Yoshihiro; Morita, Tomoko; Annan, Dorcas A; Horlad, Hasita; Ohnishi, Koji; Yamada, Sohsuke; Nakayama, Toshiyuki; Kitada, Shohei; Suzu, Shinya; Kinoshita, Ichiro; Dosaka-Akita, Hirotoshi; Akashi, Koichi; Takeya, Motohiro; Jinushi, Masahisa

2015-09-01

Clear cell renal cell carcinoma (ccRCC) is one of most common cancers in urogenital organs. Although recent experimental and clinical studies have shown the immunogenic properties of ccRCC as illustrated by the clinical sensitivities to various immunotherapies, the detailed immunoregulatory machineries governing the tumorigenicity of human ccRCC remain largely obscure. In this study, we demonstrated the clinical significance and functional relevance of T-cell immunoglobulin and mucin domain-containing molecule-3 (TIM-3) expressed on tumor cells and myeloid cells in patients with ccRCC. TIM-3 expression was detected on cancer cells and CD204(+) tumor-associated macrophages (TAM), and higher expression level of TIM-3 was positively correlated with shorter progression-free survival (PFS) in patients with ccRCC. We found that TIM-3 expression was detected on a large number of tumors, and there was significant correlation between an increased number of TAMs and high expression level of TIM-3 in patients with ccRCC. Furthermore, TIM-3 rendered RCC cells with the ability to induce resistance to sunitinib and mTOR inhibitors, the standard regimen for patients with ccRCC, as well as stem cell activities. TIM-3 expression was induced on CD14(+) monocytes upon long-term stimulation with RCC cells, and TIM-3-expressing myeloid cells play a critical role in augmenting tumorigenic activities of TIM-3-negative RCC cells. More importantly, treatment with anti-TIM-3 mAb suppressed its tumorigenic effects in in vitro and in vivo settings. These findings indicate the coordinated action of TIM-3 in cancer cells and in myeloid cells regulates the tumorigenicity of human RCC. ©2015 American Association for Cancer Research.

Vectors to Increase Production Efficiency of Inducible Pluripotent Stem Cell (iPSC) | NCI Technology Transfer Center | TTC

Cancer.gov

This invention describes the discovery that specific p53 isoform increase the number of inducible pluripotent stem cells (iPS). It is known that the activity of p53 regulates the self-renewal and pluripotency of normal and cancer stem cells, and also affects re-programming efficiency of iPS cells. This p53 isoform-based technology provides a more natural process of increasing iPS cell production than previous methods of decreasing p53. NCI seeks licensees for this technology.

Pleiotrophin regulates the expansion and regeneration of hematopoietic stem cells.

PubMed

Himburg, Heather A; Muramoto, Garrett G; Daher, Pamela; Meadows, Sarah K; Russell, J Lauren; Doan, Phuong; Chi, Jen-Tsan; Salter, Alice B; Lento, William E; Reya, Tannishtha; Chao, Nelson J; Chute, John P

2010-04-01

Hematopoietic stem cell (HSC) self-renewal is regulated by both intrinsic and extrinsic signals. Although some of the pathways that regulate HSC self-renewal have been uncovered, it remains largely unknown whether these pathways can be triggered by deliverable growth factors to induce HSC growth or regeneration. Here we show that pleiotrophin, a neurite outgrowth factor with no known function in hematopoiesis, efficiently promotes HSC expansion in vitro and HSC regeneration in vivo. Treatment of mouse bone marrow HSCs with pleiotrophin caused a marked increase in long-term repopulating HSC numbers in culture, as measured in competitive repopulating assays. Treatment of human cord blood CD34(+)CDCD38(-)Lin(-) cells with pleiotrophin also substantially increased severe combined immunodeficient (SCID)-repopulating cell counts in culture, compared to input and cytokine-treated cultures. Systemic administration of pleiotrophin to irradiated mice caused a pronounced expansion of bone marrow stem and progenitor cells in vivo, indicating that pleiotrophin is a regenerative growth factor for HSCs. Mechanistically, pleiotrophin activated phosphoinositide 3-kinase (PI3K) signaling in HSCs; antagonism of PI3K or Notch signaling inhibited pleiotrophin-mediated expansion of HSCs in culture. We identify the secreted growth factor pleiotrophin as a new regulator of both HSC expansion and regeneration.

Senescence-associated microRNAs target cell cycle regulatory genes in normal human lung fibroblasts.

PubMed

Markopoulos, Georgios S; Roupakia, Eugenia; Tokamani, Maria; Vartholomatos, George; Tzavaras, Theodore; Hatziapostolou, Maria; Fackelmayer, Frank O; Sandaltzopoulos, Raphael; Polytarchou, Christos; Kolettas, Evangelos

2017-10-01

Senescence recapitulates the ageing process at the cell level. A senescent cell stops dividing and exits the cell cycle. MicroRNAs (miRNAs) acting as master regulators of transcription, have been implicated in senescence. In the current study we investigated and compared the expression of miRNAs in young versus senescent human fibroblasts (HDFs), and analysed the role of mRNAs expressed in replicative senescent HFL-1 HDFs. Cell cycle analysis confirmed that HDFs accumulated in G 1 /S cell cycle phase. Nanostring analysis of isolated miRNAs from young and senescent HFL-1 showed that a distinct set of 15 miRNAs were significantly up-regulated in senescent cells including hsa-let-7d-5p, hsa-let-7e-5p, hsa-miR-23a-3p, hsa-miR-34a-5p, hsa-miR-122-5p, hsa-miR-125a-3p, hsa-miR-125a-5p, hsa-miR-125b-5p, hsa-miR-181a-5p, hsa-miR-221-3p, hsa-miR-222-3p, hsa-miR-503-5p, hsa-miR-574-3p, hsa-miR-574-5p and hsa-miR-4454. Importantly, pathway analysis of miRNA target genes down-regulated during replicative senescence in a public RNA-seq data set revealed a significant high number of genes regulating cell cycle progression, both G 1 /S and G 2 /M cell cycle phase transitions and telomere maintenance. The reduced expression of selected miRNA targets, upon replicative and oxidative-stress induced senescence, such as the cell cycle effectors E2F1, CcnE, Cdc6, CcnB1 and Cdc25C was verified at the protein and/or RNA levels. Induction of G1/S cell cycle phase arrest and down-regulation of cell cycle effectors correlated with the up-regulation of miR-221 upon both replicative and oxidative stress-induced senescence. Transient expression of miR-221/222 in HDFs promoted the accumulation of HDFs in G1/S cell cycle phase. We propose that miRNAs up-regulated during replicative senescence may act in concert to induce cell cycle phase arrest and telomere erosion, establishing a senescent phenotype. Copyright © 2017 Elsevier Inc. All rights reserved.

Glial Cells in the Genesis and Regulation of Circadian Rhythms

PubMed Central

Chi-Castañeda, Donají; Ortega, Arturo

2018-01-01

Circadian rhythms are biological oscillations with a period of ~24 h. These rhythms are orchestrated by a circadian timekeeper in the suprachiasmatic nucleus of the hypothalamus, the circadian “master clock,” which exactly adjusts clock outputs to solar time via photic synchronization. At the molecular level, circadian rhythms are generated by the interaction of positive and negative feedback loops of transcriptional and translational processes of the so-called “clock genes.” A large number of clock genes encode numerous proteins that regulate their own transcription and that of other genes, collectively known as “clock-controlled genes.” In addition to the sleep/wake cycle, many cellular processes are regulated by circadian rhythms, including synaptic plasticity in which an exquisite interplay between neurons and glial cells takes place. In particular, there is compelling evidence suggesting that glial cells participate in and regulate synaptic plasticity in a circadian fashion, possibly representing the missing cellular and physiological link between circadian rhythms with learning and cognition processes. Here we review recent studies in support of this hypothesis, focusing on the interplay between glial cells, synaptic plasticity, and circadian rhythmogenesis. PMID:29483880

Blue light inhibits proliferation of melanoma cells

NASA Astrophysics Data System (ADS)

Becker, Anja; Distler, Elisabeth; Klapczynski, Anna; Arpino, Fabiola; Kuch, Natalia; Simon-Keller, Katja; Sticht, Carsten; van Abeelen, Frank A.; Gretz, Norbert; Oversluizen, Gerrit

2016-03-01

Photobiomodulation with blue light is used for several treatment paradigms such as neonatal jaundice, psoriasis and back pain. However, little is known about possible side effects concerning melanoma cells in the skin. The aim of this study was to assess the safety of blue LED irradiation with respect to proliferation of melanoma cells. For that purpose we used the human malignant melanoma cell line SK-MEL28. Cell proliferation was decreased in blue light irradiated cells where the effect size depended on light irradiation dosage. Furthermore, with a repeated irradiation of the melanoma cells on two consecutive days the effect could be intensified. Fluorescence-activated cell sorting with Annexin V and Propidium iodide labeling did not show a higher number of dead cells after blue light irradiation compared to non-irradiated cells. Gene expression analysis revealed down-regulated genes in pathways connected to anti-inflammatory response, like B cell signaling and phagosome. Most prominent pathways with up-regulation of genes were cytochrome P450, steroid hormone biosynthesis. Furthermore, even though cells showed a decrease in proliferation, genes connected to the cell cycle were up-regulated after 24h. This result is concordant with XTT test 48h after irradiation, where irradiated cells showed the same proliferation as the no light negative control. In summary, proliferation of melanoma cells can be decreased using blue light irradiation. Nevertheless, the gene expression analysis has to be further evaluated and more studies, such as in-vivo experiments, are warranted to further assess the safety of blue light treatment.

A novel MDSC-induced PD-1-PD-L1+ B-cell subset in breast tumor microenvironment possesses immuno-suppressive properties.

PubMed

Shen, Meng; Wang, Jian; Yu, Wenwen; Zhang, Chen; Liu, Min; Wang, Kaiyuan; Yang, Lili; Wei, Feng; Wang, Shizhen Emily; Sun, Qian; Ren, Xiubao

2018-01-01

Myeloid-derived suppressor cells (MDSCs) are a heterogeneous group of myeloid cells that suppress T-cell activity in a tumor microenvironment. However, the suppressive function of MDSCs on B cells and its underlying mechanism remain unclear. Here, we show that in 4T1 breast cancer mice, a significantly increased number of MDSCs, in parallel with splenic B cells, are accumulated when compared to normal mice. In the presence of MDSCs, the surface molecules of B cells are remolded, with checkpoint-related molecules such as PD-1 and PD-L1 changing prominently. MDSCs also emerge as vital regulators in B-cell immune functions such as proliferation, apoptosis and the abilities to secrete antibodies and cytokines. Our study further identifies that MDSCs can transform normal B cells to a subtype of immuno- regulatory B cells (Bregs) which inhibit T-cell response. Furthermore, we identified a novel kind of Bregs with a specific phenotype PD-1 - PD-L1 + CD19 + , which exert the greatest suppressive effects on T cells in comparison with the previously reported Bregs characterized as CD1d + CD5 + CD19 + , CD5 + CD19 + and Interleukin (IL)-10-secreting B cells. Our results highlight that MDSCs regulate B-cell response and may serve as a therapeutic approach in anti-tumor treatment. Investigation of this new Breg subtype extends our understanding of regulation of T-cell response and sheds new light on anti-tumor immunity and immune therapy.

Functional roles of cell surface peptidases in reproductive organs

PubMed Central

2004-01-01

A number of biologically active peptides have been proposed to regulate function and differentiation of reproductive organs in an autocrine and/or paracrine fashion. Regulation of the local concentrations of these peptides is one of the important factors influencing their physiological effects on target cells. Membrane‐bound cell surface peptidases can activate or inactivate biologically active peptides before peptide factors access their receptors on the cell surface. Aminopeptidase A (EC 3.4.11.7), placental leucine aminopeptidase (EC 3.4.11.3), aminopeptidase‐N/CD13 (EC 3.4.11.2), dipeptidyl peptidases IV/CD26 (EC.3.4.14.5), carboxypeptidase‐M (EC 3.4.17.12), neutral endopeptidase/CD10 (EC 3.4.24.11) and endothelin converting enzyme‐1 (EC 3.4.23) are differentially expressed on the ovary, endometrium and placenta. The inhibition of enzyme activity affects steroid hormone production by granulosa and thecal cells, decidualization of endometrium and migration of extravillous trophoblasts. These findings suggest that membrane‐bound cell surface peptidases are local regulators for cellular growth and differentiation in reproductive organs by controlling extracellular concentration of peptide factors. (Reprod Med Biol 2004; 3: 165 –176) PMID:29662383

Impaired mitotic progression and preimplantation lethality in mice lacking OMCG1, a new evolutionarily conserved nuclear protein.

PubMed

Artus, Jérôme; Vandormael-Pournin, Sandrine; Frödin, Morten; Nacerddine, Karim; Babinet, Charles; Cohen-Tannoudji, Michel

2005-07-01

While highly conserved through evolution, the cell cycle has been extensively modified to adapt to new developmental programs. Recently, analyses of mouse mutants revealed that several important cell cycle regulators are either dispensable for development or have a tissue- or cell-type-specific function, indicating that many aspects of cell cycle regulation during mammalian embryo development remain to be elucidated. Here, we report on the characterization of a new gene, Omcg1, which codes for a nuclear zinc finger protein. Embryos lacking Omcg1 die by the end of preimplantation development. In vitro cultured Omcg1-null blastocysts exhibit a dramatic reduction in the total cell number, a high mitotic index, and the presence of abnormal mitotic figures. Importantly, we found that Omcg1 disruption results in the lengthening of M phase rather than in a mitotic block. We show that the mitotic delay in Omcg1-/- embryos is associated with neither a dysfunction of the spindle checkpoint nor abnormal global histone modifications. Taken together, these results suggest that Omcg1 is an important regulator of the cell cycle in the preimplantation embryo.

Impaired Mitotic Progression and Preimplantation Lethality in Mice Lacking OMCG1, a New Evolutionarily Conserved Nuclear Protein†

PubMed Central

Artus, Jérôme; Vandormael-Pournin, Sandrine; Frödin, Morten; Nacerddine, Karim; Babinet, Charles; Cohen-Tannoudji, Michel

2005-01-01

While highly conserved through evolution, the cell cycle has been extensively modified to adapt to new developmental programs. Recently, analyses of mouse mutants revealed that several important cell cycle regulators are either dispensable for development or have a tissue- or cell-type-specific function, indicating that many aspects of cell cycle regulation during mammalian embryo development remain to be elucidated. Here, we report on the characterization of a new gene, Omcg1, which codes for a nuclear zinc finger protein. Embryos lacking Omcg1 die by the end of preimplantation development. In vitro cultured Omcg1-null blastocysts exhibit a dramatic reduction in the total cell number, a high mitotic index, and the presence of abnormal mitotic figures. Importantly, we found that Omcg1 disruption results in the lengthening of M phase rather than in a mitotic block. We show that the mitotic delay in Omcg1−/− embryos is associated with neither a dysfunction of the spindle checkpoint nor abnormal global histone modifications. Taken together, these results suggest that Omcg1 is an important regulator of the cell cycle in the preimplantation embryo. PMID:15988037

A Legume TOR Protein Kinase Regulates Rhizobium Symbiosis and Is Essential for Infection and Nodule Development1[OPEN

PubMed Central

Blanco, Lourdes; Quinto, Carmen

2016-01-01

The target of rapamycin (TOR) protein kinase regulates metabolism, growth, and life span in yeast, animals, and plants in coordination with nutrient status and environmental conditions. The nutrient-dependent nature of TOR functionality makes this kinase a putative regulator of symbiotic associations involving nutrient acquisition. However, TOR’s role in these processes remains to be understood. Here, we uncovered the role of TOR during the bean (Phaseolus vulgaris)-Rhizobium tropici (Rhizobium) symbiotic interaction. TOR was expressed in all tested bean tissues, with higher transcript levels in the root meristems and senesced nodules. We showed TOR promoter expression along the progressing infection thread and in the infected cells of mature nodules. Posttranscriptional gene silencing of TOR using RNA interference (RNAi) showed that this gene is involved in lateral root elongation and root cell organization and also alters the density, size, and number of root hairs. The suppression of TOR transcripts also affected infection thread progression and associated cortical cell divisions, resulting in a drastic reduction of nodule numbers. TOR-RNAi resulted in reduced reactive oxygen species accumulation and altered CyclinD1 and CyclinD3 expression, which are crucial factors for infection thread progression and nodule organogenesis. Enhanced expression of TOR-regulated ATG genes in TOR-RNAi roots suggested that TOR plays a role in the recognition of Rhizobium as a symbiont. Together, these data suggest that TOR plays a vital role in the establishment of root nodule symbiosis in the common bean. PMID:27698253

Macrophage-Mediated Glial Cell Elimination in the Postnatal Mouse Cochlea

PubMed Central

Brown, LaShardai N.; Xing, Yazhi; Noble, Kenyaria V.; Barth, Jeremy L.; Panganiban, Clarisse H.; Smythe, Nancy M.; Bridges, Mary C.; Zhu, Juhong; Lang, Hainan

2017-01-01

Hearing relies on the transmission of auditory information from sensory hair cells (HCs) to the brain through the auditory nerve. This relay of information requires HCs to be innervated by spiral ganglion neurons (SGNs) in an exclusive manner and SGNs to be ensheathed by myelinating and non-myelinating glial cells. In the developing auditory nerve, mistargeted SGN axons are retracted or pruned and excessive cells are cleared in a process referred to as nerve refinement. Whether auditory glial cells are eliminated during auditory nerve refinement is unknown. Using early postnatal mice of either sex, we show that glial cell numbers decrease after the first postnatal week, corresponding temporally with nerve refinement in the developing auditory nerve. Additionally, expression of immune-related genes was upregulated and macrophage numbers increase in a manner coinciding with the reduction of glial cell numbers. Transient depletion of macrophages during early auditory nerve development, using transgenic CD11bDTR/EGFP mice, resulted in the appearance of excessive glial cells. Macrophage depletion caused abnormalities in myelin formation and transient edema of the stria vascularis. Macrophage-depleted mice also showed auditory function impairment that partially recovered in adulthood. These findings demonstrate that macrophages contribute to the regulation of glial cell number during postnatal development of the cochlea and that glial cells play a critical role in hearing onset and auditory nerve maturation. PMID:29375297

Enhancement of high glucose-induced PINK1 expression by melatonin stimulates neuronal cell survival: Involvement of MT2 /Akt/NF-κB pathway.

PubMed

Onphachanh, Xaykham; Lee, Hyun Jik; Lim, Jae Ryong; Jung, Young Hyun; Kim, Jun Sung; Chae, Chang Woo; Lee, Sei-Jung; Gabr, Amr Ahmed; Han, Ho Jae

2017-09-01

Hyperglycemia is a representative hallmark and risk factor for diabetes mellitus (DM) and is closely linked to DM-associated neuronal cell death. Previous investigators reported on a genome-wide association study and showed relationships between DM and melatonin receptor (MT), highlighting the role of MT signaling by assessing melatonin in DM. However, the role of MT signaling in DM pathogenesis is unclear. Therefore, we investigated the role of mitophagy regulators in high glucose-induced neuronal cell death and the effect of melatonin against high glucose-induced mitophagy regulators in neuronal cells. In our results, high glucose significantly increased PTEN-induced putative kinase 1 (PINK1) and LC-3B expressions; as well it decreased cytochrome c oxidase subunit 4 expression and Mitotracker™ fluorescence intensity. Silencing of PINK1 induced mitochondrial reactive oxygen species (ROS) accumulation and mitochondrial membrane potential impairment, increased expressions of cleaved caspases, and increased the number of annexin V-positive cells. In addition, high glucose-stimulated melatonin receptor 1B (MTNR1B) mRNA and PINK1 expressions were reversed by ROS scavenger N-acetyl cysteine pretreatment. Upregulation of PINK1 expression in neuronal cells is suppressed by pretreatment with MT 2 receptor-specific inhibitor 4-P-PDOT. We further showed melatonin stimulated Akt phosphorylation, which was followed by nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) phosphorylation and nuclear translocation. Silencing of PINK1 expression abolished melatonin-regulated mitochondrial ROS production, cleaved caspase-3 and caspase-9 expressions, and the number of annexin V-positive cells. In conclusion, we have demonstrated the melatonin stimulates PINK1 expression via an MT 2 /Akt/NF-κB pathway, and such stimulation is important for the prevention of neuronal cell apoptosis under high glucose conditions. © 2017 The Authors. Journal of Pineal Research Published by John Wiley & Sons Ltd.

Consequences of endogenous and exogenous WNT signaling for development of the preimplantation bovine embryo.

PubMed

Tribulo, Paula; Leão, Beatriz Caetano da Silva; Lehloenya, Khoboso C; Mingoti, Gisele Zoccal; Hansen, Peter J

2017-06-01

The specific role of WNT signaling during preimplantation development remains unclear. Here, we evaluated consequences of activation and inhibition of β-catenin (CTNNB1)-dependent and -independent WNT signaling in the bovine preimplantation embryo. Activation of CTNNB1-mediated WNT signaling by the agonist 2-amino-4-(3,4-(methylenedioxy)benzylamino)-6-(3-methoxyphenyl)pyrimidine (AMBMP) and a glycogen synthase kinase 3 inhibitor reduced development to the blastocyst stage. Moreover, the antagonist of WNT signaling, dickkopf-related protein 1 (DKK1), alleviated the negative effect of AMBMP on development via reduction of CTNNB1. Based on labeling for phospho c-Jun N-terminal kinase, there was no evidence that DKK1 activated the planar cell polarity (PCP) pathway. Inhibition of secretion of endogenous WNTs did not affect development but increased number of cells in the inner cell mass (ICM). In contrast, DKK1 did not affect number of ICM or trophectoderm (TE) cells, suggesting that embryo-derived WNTs regulate ICM proliferation through a mechanism independent of CTNNB1. In addition, DKK1 did not affect the number of cells positive for the transcription factor yes-associated protein 1 (YAP1) involved in TE formation. In fact, DKK1 decreased YAP1. In contrast, exposure of embryos to WNT family member 7A (WNT7A) improved blastocyst development, inhibited the PCP pathway, and did not affect amounts of CTNNB1. Results indicate that embryo-derived WNTs are dispensable for blastocyst formation but participate in regulation of ICM proliferation, likely through a mechanism independent of CTNNB1. The response to AMBMP and WNT7A leads to the hypothesis that maternally derived WNTs can play a positive or negative role in regulation of preimplantation development. © The Authors 2017. Published by Oxford University Press on behalf of Society for the Study of Reproduction.

T cell regulation in microgravity - The current knowledge from in vitro experiments conducted in space, parabolic flights and ground-based facilities

NASA Astrophysics Data System (ADS)

Hauschild, Swantje; Tauber, Svantje; Lauber, Beatrice; Thiel, Cora S.; Layer, Liliana E.; Ullrich, Oliver

2014-11-01

Dating back to the Apollo and Skylab missions, it has been reported that astronauts suffered from bacterial and viral infections during space flight or after returning to Earth. Blood analyses revealed strongly reduced capability of human lymphocytes to become active upon mitogenic stimulation. Since then, a large number of in vitro studies on human immune cells have been conducted in space, in parabolic flights, and in ground-based facilities. It became obvious that microgravity affects cell morphology and important cellular functions. Observed changes include cell proliferation, the cytoskeleton, signal transduction and gene expression. This review gives an overview of the current knowledge of T cell regulation under altered gravity conditions obtained by in vitro studies with special emphasis on the cell culture conditions used. We propose that future in vitro experiments should follow rigorous standardized cell culture conditions, which allows better comparison of the results obtained in different flight- and ground-based experiment platforms.

CXCR2 and CXCL4 regulate survival and self-renewal of hematopoietic stem/progenitor cells.

PubMed

Sinclair, Amy; Park, Laura; Shah, Mansi; Drotar, Mark; Calaminus, Simon; Hopcroft, Lisa E M; Kinstrie, Ross; Guitart, Amelie V; Dunn, Karen; Abraham, Sheela A; Sansom, Owen; Michie, Alison M; Machesky, Laura; Kranc, Kamil R; Graham, Gerard J; Pellicano, Francesca; Holyoake, Tessa L

2016-07-21

The regulation of hematopoietic stem cell (HSC) survival and self-renewal within the bone marrow (BM) niche is not well understood. We therefore investigated global transcriptomic profiling of normal human HSC/hematopoietic progenitor cells [HPCs], revealing that several chemokine ligands (CXCL1-4, CXCL6, CXCL10, CXCL11, and CXCL13) were upregulated in human quiescent CD34(+)Hoescht(-)Pyronin Y(-) and primitive CD34(+)38(-), as compared with proliferating CD34(+)Hoechst(+)Pyronin Y(+) and CD34(+)38(+) stem/progenitor cells. This suggested that chemokines might play an important role in the homeostasis of HSCs. In human CD34(+) hematopoietic cells, knockdown of CXCL4 or pharmacologic inhibition of the chemokine receptor CXCR2, significantly decreased cell viability and colony forming cell (CFC) potential. Studies on Cxcr2(-/-) mice demonstrated enhanced BM and spleen cellularity, with significantly increased numbers of HSCs, hematopoietic progenitor cell-1 (HPC-1), HPC-2, and Lin(-)Sca-1(+)c-Kit(+) subpopulations. Cxcr2(-/-) stem/progenitor cells showed reduced self-renewal capacity as measured in serial transplantation assays. Parallel studies on Cxcl4 demonstrated reduced numbers of CFC in primary and secondary assays following knockdown in murine c-Kit(+) cells, and Cxcl4(-/-) mice showed a decrease in HSC and reduced self-renewal capacity after secondary transplantation. These data demonstrate that the CXCR2 network and CXCL4 play a role in the maintenance of normal HSC/HPC cell fates, including survival and self-renewal. © 2016 by The American Society of Hematology.

CXCR2 and CXCL4 regulate survival and self-renewal of hematopoietic stem/progenitor cells

PubMed Central

Sinclair, Amy; Park, Laura; Shah, Mansi; Drotar, Mark; Calaminus, Simon; Hopcroft, Lisa E. M.; Kinstrie, Ross; Guitart, Amelie V.; Dunn, Karen; Abraham, Sheela A.; Sansom, Owen; Michie, Alison M.; Machesky, Laura; Kranc, Kamil R.; Graham, Gerard J.; Pellicano, Francesca

2016-01-01

The regulation of hematopoietic stem cell (HSC) survival and self-renewal within the bone marrow (BM) niche is not well understood. We therefore investigated global transcriptomic profiling of normal human HSC/hematopoietic progenitor cells [HPCs], revealing that several chemokine ligands (CXCL1-4, CXCL6, CXCL10, CXCL11, and CXCL13) were upregulated in human quiescent CD34+Hoescht−Pyronin Y− and primitive CD34+38−, as compared with proliferating CD34+Hoechst+Pyronin Y+ and CD34+38+ stem/progenitor cells. This suggested that chemokines might play an important role in the homeostasis of HSCs. In human CD34+ hematopoietic cells, knockdown of CXCL4 or pharmacologic inhibition of the chemokine receptor CXCR2, significantly decreased cell viability and colony forming cell (CFC) potential. Studies on Cxcr2−/− mice demonstrated enhanced BM and spleen cellularity, with significantly increased numbers of HSCs, hematopoietic progenitor cell-1 (HPC-1), HPC-2, and Lin−Sca-1+c-Kit+ subpopulations. Cxcr2−/− stem/progenitor cells showed reduced self-renewal capacity as measured in serial transplantation assays. Parallel studies on Cxcl4 demonstrated reduced numbers of CFC in primary and secondary assays following knockdown in murine c-Kit+ cells, and Cxcl4−/− mice showed a decrease in HSC and reduced self-renewal capacity after secondary transplantation. These data demonstrate that the CXCR2 network and CXCL4 play a role in the maintenance of normal HSC/HPC cell fates, including survival and self-renewal. PMID:27222476

MicroRNA-211 Regulates Oxidative Phosphorylation and Energy Metabolism in Human Vitiligo.

PubMed

Sahoo, Anupama; Lee, Bongyong; Boniface, Katia; Seneschal, Julien; Sahoo, Sanjaya K; Seki, Tatsuya; Wang, Chunyan; Das, Soumen; Han, Xianlin; Steppie, Michael; Seal, Sudipta; Taieb, Alain; Perera, Ranjan J

2017-09-01

Vitiligo is a common chronic skin disorder characterized by loss of epidermal melanocytes and progressive depigmentation. Vitiligo has complex immune, genetic, environmental, and biochemical causes, but the exact molecular mechanisms of vitiligo development and progression, particularly those related to metabolic control, are poorly understood. In this study we characterized the human vitiligo cell line PIG3V and the normal human melanocyte line HEM-l by RNA sequencing, targeted metabolomics, and shotgun lipidomics. Melanocyte-enriched microRNA-211, a known metabolic switch in nonpigmented melanoma cells, was severely down-regulated in vitiligo cell line PIG3V and skin biopsy samples from vitiligo patients, whereas its predicted targets PPARGC1A, RRM2, and TAOK1 were reciprocally up-regulated. microRNA-211 binds to PGC1-α 3' untranslated region locus and represses it. Although mitochondrial numbers were constant, mitochondrial complexes I, II, and IV and respiratory responses were defective in vitiligo cells. Nanoparticle-coated microRNA-211 partially augmented the oxygen consumption rate in PIG3V cells. The lower oxygen consumption rate, changes in lipid and metabolite profiles, and increased reactive oxygen species production observed in vitiligo cells appear to be partly due to abnormal regulation of microRNA-211 and its target genes. These genes represent potential biomarkers and therapeutic targets in human vitiligo. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

Impact of branching on the elasticity of actin networks

PubMed Central

Pujol, Thomas; du Roure, Olivia; Fermigier, Marc; Heuvingh, Julien

2012-01-01

Actin filaments play a fundamental role in cell mechanics: assembled into networks by a large number of partners, they ensure cell integrity, deformability, and migration. Here we focus on the mechanics of the dense branched network found at the leading edge of a crawling cell. We develop a new technique based on the dipolar attraction between magnetic colloids to measure mechanical properties of branched actin gels assembled around the colloids. This technique allows us to probe a large number of gels and, through the study of different networks, to access fundamental relationships between their microscopic structure and their mechanical properties. We show that the architecture does regulate the elasticity of the network: increasing both capping and branching concentrations strongly stiffens the networks. These effects occur at protein concentrations that can be regulated by the cell. In addition, the dependence of the elastic modulus on the filaments’ flexibility and on increasing internal stress has been studied. Our overall results point toward an elastic regime dominated by enthalpic rather than entropic deformations. This result strongly differs from the elasticity of diluted cross-linked actin networks and can be explained by the dense dendritic structure of lamellipodium-like networks. PMID:22689953

Abdominal-B and caudal inhibit the formation of specific neuroblasts in the Drosophila tail region

PubMed Central

Birkholz, Oliver; Vef, Olaf; Rogulja-Ortmann, Ana; Berger, Christian; Technau, Gerhard M.

2013-01-01

The central nervous system of Drosophila melanogaster consists of fused segmental units (neuromeres), each generated by a characteristic number of neural stem cells (neuroblasts). In the embryo, thoracic and anterior abdominal neuromeres are almost equally sized and formed by repetitive sets of neuroblasts, whereas the terminal abdominal neuromeres are generated by significantly smaller populations of progenitor cells. Here we investigated the role of the Hox gene Abdominal-B in shaping the terminal neuromeres. We show that the regulatory isoform of Abdominal-B (Abd-B.r) not only confers abdominal fate to specific neuroblasts (e.g. NB6-4) and regulates programmed cell death of several progeny cells within certain neuroblast lineages (e.g. NB3-3) in parasegment 14, but also inhibits the formation of a specific set of neuroblasts in parasegment 15 (including NB7-3). We further show that Abd-B.r requires cooperation of the ParaHox gene caudal to unfold its full competence concerning neuroblast inhibition and specification. Thus, our findings demonstrate that combined action of Abdominal-B and caudal contributes to the size and composition of the terminal neuromeres by regulating both the number and lineages of specific neuroblasts. PMID:23903193

Investigation of fusion gene expression in HCT116 cells.

PubMed

Zhang, Yanmei; Ren, Juan; Fang, Mengdie; Wang, Xiaoju

2017-12-01

Colon cancer is the most common type of gastrointestinal cancer. A number of specific and sensitive biomarkers facilitate the diagnosis and monitoring of patients with colon cancer. Fusion genes are typically identified in cancer and a majority of the newly identified fusion genes are oncogenic in nature. Therefore, fusion genes are potential biomarkers and/or therapy targets in cancer. In the present study, the regulation of specific candidate fusion genes were investigated using Brother of the Regulator of Imprinted Sites (BORIS) in the HCT116 colon cancer cell line, which is a paralog of the fusion gene regulator CCCTC-binding factor (CTCF). The copy number of BORIS increased correspondingly to the progression of colorectal carcinoma from the M0 to the M1a stage. It was identified that EIF3E(e1)-RSPO2(e2) , EIF3E(e1)-RSPO2(e3) , PTPRK(e1)-RSPO3(e2) , PTPRK(e7)-RSPO3(e2), TADA2A-MEF2B and MED13L-CD4 are fusion transcripts present in the transcriptome of the HCT116 colon cancer cell line. CDC42SE2-KIAAO146 is a genomic fusion transcript, which originates from DNA arrangement in HCT116 cells. BORIS suppresses the expression of EIF3E , RSPO2 , PTPRK , RSPO3 , TADA2A and CD4 to inhibit the expression of fusion transcripts in HCT116 cells. It was hypothesized that the fusion transcripts investigated in the present study may not be oncogenic in HCT116 cells. As BORIS is not colorectal carcinoma-specific, the fusion genes investigated may be a biomarker assemblage for monitoring the progression of colorectal carcinoma.

Investigation of fusion gene expression in HCT116 cells

PubMed Central

Zhang, Yanmei; Ren, Juan; Fang, Mengdie; Wang, Xiaoju

2017-01-01

Colon cancer is the most common type of gastrointestinal cancer. A number of specific and sensitive biomarkers facilitate the diagnosis and monitoring of patients with colon cancer. Fusion genes are typically identified in cancer and a majority of the newly identified fusion genes are oncogenic in nature. Therefore, fusion genes are potential biomarkers and/or therapy targets in cancer. In the present study, the regulation of specific candidate fusion genes were investigated using Brother of the Regulator of Imprinted Sites (BORIS) in the HCT116 colon cancer cell line, which is a paralog of the fusion gene regulator CCCTC-binding factor (CTCF). The copy number of BORIS increased correspondingly to the progression of colorectal carcinoma from the M0 to the M1a stage. It was identified that EIF3E(e1)-RSPO2(e2), EIF3E(e1)-RSPO2(e3), PTPRK(e1)-RSPO3(e2), PTPRK(e7)-RSPO3(e2), TADA2A-MEF2B and MED13L-CD4 are fusion transcripts present in the transcriptome of the HCT116 colon cancer cell line. CDC42SE2-KIAAO146 is a genomic fusion transcript, which originates from DNA arrangement in HCT116 cells. BORIS suppresses the expression of EIF3E, RSPO2, PTPRK, RSPO3, TADA2A and CD4 to inhibit the expression of fusion transcripts in HCT116 cells. It was hypothesized that the fusion transcripts investigated in the present study may not be oncogenic in HCT116 cells. As BORIS is not colorectal carcinoma-specific, the fusion genes investigated may be a biomarker assemblage for monitoring the progression of colorectal carcinoma. PMID:29181107

Function of OPG as a traffic regulator for RANKL is crucial for controlled osteoclastogenesis.

PubMed

Aoki, Shigeki; Honma, Masashi; Kariya, Yoshiaki; Nakamichi, Yuko; Ninomiya, Tadashi; Takahashi, Naoyuki; Udagawa, Nobuyuki; Suzuki, Hiroshi

2010-09-01

The amount of the receptor activator of NF-κB ligand (RANKL) on the osteoblastic cell surface is considered to determine the magnitude of the signal input to osteoclast precursors and the degree of osteoclastogenesis. Previously, we have shown that RANKL is localized predominantly in lysosomal organelles, but little is found on the osteoblastic cell surface, and consequently, the regulated subcellular trafficking of RANKL in osteoblastic cells is important for controlled osteoclastogenesis. Here we have examined the involvement of osteoprotegerin (OPG), which is currently recognized as a decoy receptor for RANKL, in the regulation of RANKL behavior. It was suggested that OPG already makes a complex with RANKL in the Golgi apparatus and that the complex formation is necessary for RANKL sorting to the secretory lysosomes. It was also shown that each structural domain of OPG is indispensable for exerting OPG function as a traffic regulator. In particular, the latter domains of OPG, whose physiologic functions have been unclear, were indicated to sort RANKL molecules to lysosomes from the Golgi apparatus. In addition, the overexpression of RANK-OPG chimeric protein, which retained OPG function as a decoy receptor but lost the function as a traffic regulator, inhibited endogenous OPG function as a traffic regulator selectively in osteoblastic cells and resulted in the upregulation of osteoclastogenic ability despite the increased number of decoy receptor molecules. Conclusively, OPG function as a traffic regulator for RANKL is crucial for regulating osteoclastogenesis at least as well as that as a decoy receptor. © 2010 American Society for Bone and Mineral Research.

IL-10 down-regulates T cell activation by antigen-presenting liver sinusoidal endothelial cells through decreased antigen uptake via the mannose receptor and lowered surface expression of accessory molecules.

PubMed

Knolle, P A; Uhrig, A; Hegenbarth, S; Löser, E; Schmitt, E; Gerken, G; Lohse, A W

1998-12-01

Our study demonstrates that antigen-presenting liver sinusoidal endothelial cells (LSEC) induce production of interferon-gamma (IFN-gamma) from cloned Th1 CD4+ T cells. We show that LSEC used the mannose receptor for antigen uptake, which further strengthened the role of LSEC as antigen-presenting cell (APC) population in the liver. The ability of LSEC to activate cloned CD4+ T cells antigen-specifically was down-regulated by exogenous prostaglandin E2 (PGE2) and by IL-10. We identify two separate mechanisms by which IL-10 down-regulated T cell activation through LSEC. IL-10 decreased the constitutive surface expression of MHC class II as well as of the accessory molecules CD80 and CD86 on LSEC. Furthermore, IL-10 diminished mannose receptor activity in LSEC. Decreased antigen uptake via the mannose receptor and decreased expression of accessory molecules may explain the down-regulation of T cell activation through IL-10. Importantly, the expression of low numbers of antigen on MHC II in the absence of accessory signals on LSEC may lead to induction of anergy in T cells. Because PGE2 and IL-10 are released from LSEC or Kupffer cells (KC) in response to those concentrations of endotoxin found physiologically in portal venous blood, it is possible that the continuous presence of these mediators and their negative effect on the local APC may explain the inability of the liver to induce T cell activation and to clear chronic infections. Our results support the notion that antigen presentation by LSEC in the hepatic microenvironment contributes to the observed inability to mount an effective cell-mediated immune response in the liver.

Missing Value Monitoring Enhances the Robustness in Proteomics Quantitation.

PubMed

Matafora, Vittoria; Corno, Andrea; Ciliberto, Andrea; Bachi, Angela

2017-04-07

In global proteomic analysis, it is estimated that proteins span from millions to less than 100 copies per cell. The challenge of protein quantitation by classic shotgun proteomic techniques relies on the presence of missing values in peptides belonging to low-abundance proteins that lowers intraruns reproducibility affecting postdata statistical analysis. Here, we present a new analytical workflow MvM (missing value monitoring) able to recover quantitation of missing values generated by shotgun analysis. In particular, we used confident data-dependent acquisition (DDA) quantitation only for proteins measured in all the runs, while we filled the missing values with data-independent acquisition analysis using the library previously generated in DDA. We analyzed cell cycle regulated proteins, as they are low abundance proteins with highly dynamic expression levels. Indeed, we found that cell cycle related proteins are the major components of the missing values-rich proteome. Using the MvM workflow, we doubled the number of robustly quantified cell cycle related proteins, and we reduced the number of missing values achieving robust quantitation for proteins over ∼50 molecules per cell. MvM allows lower quantification variance among replicates for low abundance proteins with respect to DDA analysis, which demonstrates the potential of this novel workflow to measure low abundance, dynamically regulated proteins.

Benzophenone-3 increases metastasis potential in lung cancer cells via epithelial to mesenchymal transition.

PubMed

Phiboonchaiyanan, Preeyaporn Plaimee; Busaranon, Kesarin; Ninsontia, Chuanpit; Chanvorachote, Pithi

2017-06-01

Exposure to compounds with cancer-potentiating effects can contribute to the progression of cancer. Herein we have discovered for the first time that benzophenone-3 (BP-3), a chemical used as sunscreen in various cosmetic products, enhances the ability of lung cancer cells to undergo metastasis. The exposure of the lung cancer cells to BP-3 at non-toxic concentrations significantly increased the number of anoikis resistant cells in a dose-dependent manner. Also, BP-3 increased the growth rate as well as the number of colonies accessed by anchorage-independent growth assay. We found that the underlying mechanisms of such behaviors were the epithelial to mesenchymal transition (EMT) process of cancer cells, and the increase in caveolin-1 (Cav-1) expression. As both mechanistic events mediated anoikis resistance via augmentation of cellular survival signals, our results further revealed that the BP-3 treatment significantly up-regulated extracellular-signal-regulated kinase (ERK). Also, such compounds increased the cellular levels of anti-apoptotic Bcl-2 and Mcl-1 proteins. As the presence of a substantial level of BP-3 in plasma of the consumers has been reported, this finding may facilitate further investigations that lead to better understanding and evidence concerning the safety of use in cancer patients.

Candida glabrata's Genome Plasticity Confers a Unique Pattern of Expressed Cell Wall Proteins.

PubMed

López-Fuentes, Eunice; Gutiérrez-Escobedo, Guadalupe; Timmermans, Bea; Van Dijck, Patrick; De Las Peñas, Alejandro; Castaño, Irene

2018-06-05

Candida glabrata is the second most common cause of candidemia, and its ability to adhere to different host cell types, to microorganisms, and to medical devices are important virulence factors. Here, we consider three characteristics that confer extraordinary advantages to C. glabrata within the host. (1) C. glabrata has a large number of genes encoding for adhesins most of which are localized at subtelomeric regions. The number and sequence of these genes varies substantially depending on the strain, indicating that C. glabrata can tolerate high genomic plasticity; (2) The largest family of CWPs (cell wall proteins) is the EPA (epithelial adhesin) family of adhesins. Epa1 is the major adhesin and mediates adherence to epithelial, endothelial and immune cells. Several layers of regulation like subtelomeric silencing, cis- acting regulatory regions, activators, nutritional signaling, and stress conditions tightly regulate the expression of many adhesin-encoding genes in C. glabrata , while many others are not expressed. Importantly, there is a connection between acquired resistance to xenobiotics and increased adherence; (3) Other subfamilies of adhesins mediate adherence to Candida albicans , allowing C. glabrata to efficiently invade the oral epithelium and form robust biofilms. It is noteworthy that every C. glabrata strain analyzed presents a unique pattern of CWPs at the cell surface.

[Mitochondria inheritance in yeast saccharomyces cerevisiae].

PubMed

Fizikova, A Iu

2011-01-01

The review is devoted to the main mechanisms of mitochondria inheritance in yeast Saccharonmyces cerevisiae. The genetic mechanisms of functionally active mitochondria inheritance in eukaryotic cells is one of the most relevant in modem researches. A great number of genetic diseases are associated with mitochondria dysfunction. Plasticity of eukaryotic cell metabolism according to the environmental changes is ensured by adequate mitochondria functioning by means of ATP synthesis coordination, reactive oxygen species accumulation, apoptosis regulation and is an important factor of cell adaptation to stress. Mitochondria participation in important for cell vitality processes masters the presence of accurate mechanisms of mitochondria functions regulation according to environment fluctuations. The mechanisms of mitochondria division and distribution are highly conserved. Baker yeast S. cerevisiae is an ideal model object for mitochondria researches due to energetic metabolism lability, ability to switch over respiration to fermentation, and petite-positive phenotype. Correction of metabolism according to the environmental changes is necessary for cell vitality. The influence of respiratory, carbon, amino acid and phosphate metabolism on mitochondria functions was shown. As far as the mechanisms that stabilize functions of mitochondria and mtDNA are highly conserve, we can project yeast regularities on higher eukaryotes systems. This makes it possible to approximate understanding the etiology and pathogenesis of a great number of human diseases.

Norisoboldine ameliorates collagen-induced arthritis through regulating the balance between Th17 and regulatory T cells in gut-associated lymphoid tissues

DOE Office of Scientific and Technical Information (OSTI.GOV)

Tong, Bei; Dou, Yannong; Wang, Ting

Norisoboldine (NOR), the main active ingredient of the dry root of Lindera aggregata, was previously proven to have substantial therapeutic effects on collagen-induced arthritis (CIA) in mice by oral administration. However, it exhibited a very poor bioavailability in normal rats. The pharmacokinetic–pharmacodynamics disconnection attracts us to explore its anti-arthritic mechanism in more detail. In this study, NOR, administered orally, markedly attenuated the pathological changes in CIA rats, which was accompanied by the down-regulation of pro-inflammatory cytokines and the up-regulation of anti-inflammatory cytokine IL-10. Pharmacokinetic studies demonstrated that the plasma concentration of NOR was moderately elevated in CIA rats compared withmore » normal rats, but it was still far lower than the minimal effective concentration required for inhibiting the proliferation and activation of T lymphocytes in vitro. Interestingly, NOR was shown to regulate the balance between Th17 and regulatory T (Treg) cells in the intestinal lymph nodes more strikingly than in other tissues. It could increase the expression of Foxp3 mRNA in both gut and joints, and markedly up-regulate the number of integrin α4β7 (a marker of gut source)-positive Foxp3{sup +} cells in the joints of CIA rats. These results suggest that the gut might be the primary action site of NOR, and NOR exerts anti-arthritis effect through regulating the balance between Th17 and Treg cells in intestinal lymph nodes and yielding a trafficking of lymphocytes (especially Treg cells) from the gut to joint. The findings of the present study also provide a plausible explanation for the anti-arthritic effects of poorly absorbed compounds like NOR. - Highlights: • Norisoboldine, administered orally, markedly attenuates the clinical signs of CIA. • Norisoboldine regulates the balance of Th17/Treg cells in the intestinal lymph node. • Norisoboldine induces the migration of Treg cells from the gut to joint.« less

Expression of the Argonaute protein PiwiL2 and piRNAs in adult mouse mesenchymal stem cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wu, Qiuling; Ma, Qi; Shehadeh, Lina A.

Piwi (P-element-induced wimpy testis) first discovered in Drosophila is a member of the Argonaute family of micro-RNA binding proteins with essential roles in germ-cell development. The murine homologue of PiwiL2, also known as Mili is selectively expressed in the testes, and mice bearing targeted mutations of the PiwiL2 gene are male-sterile. PiwiL2 proteins are thought to protect the germ line genome by suppressing retrotransposons, stabilizing heterochromatin structure, and regulating target genes during meiosis and mitosis. Here, we report that PiwiL2 and associated piRNAs (piRs) may play similar roles in adult mouse mesenchymal stem cells. We found that PiwiL2 is expressedmore » in the cytoplasm of metaphase mesenchymal stem cells from the bone marrow of adult and aged mice. Knockdown of PiwiL2 with a specific siRNA enhanced cell proliferation, significantly increased the number of cells in G1/S and G2/M cell cycle phases and was associated with increased expression of cell cycle genes CCND1, CDK8, microtubule regulation genes, and decreased expression of tumor suppressors Cables 1, LATS, and Cxxc4. The results suggest broader roles for Piwi in genome surveillance beyond the germ line and a possible role in regulating the cell cycle of mesenchymal stem cells.« less

CD22-Binding Synthetic Sialosides Regulate B Lymphocyte Proliferation Through CD22 Ligand-Dependent and Independent Pathways, and Enhance Antibody Production in Mice

PubMed Central

Matsubara, Naoko; Imamura, Akihiro; Yonemizu, Tatsuya; Akatsu, Chizuru; Yang, Hongrui; Ueki, Akiharu; Watanabe, Natsuki; Abdu-Allah, Hajjaj; Numoto, Nobutaka; Takematsu, Hiromu; Kitazume, Shinobu; Tedder, Thomas F.; Marth, Jamey D.; Ito, Nobutoshi; Ando, Hiromune; Ishida, Hideharu; Kiso, Makoto; Tsubata, Takeshi

2018-01-01

Sialic acid-binding immunoglobulin-like lectins (Siglecs) are expressed in various immune cells and most of them carry signaling functions. High-affinity synthetic sialoside ligands have been developed for various Siglecs. Therapeutic potentials of the nanoparticles and compounds that contain multiple numbers of these sialosides and other reagents such as toxins and antigens have been demonstrated. However, whether immune responses can be regulated by monomeric sialoside ligands has not yet been known. CD22 (also known as Siglec-2) is an inhibitory molecule preferentially expressed in B lymphocytes (B cells) and is constitutively bound and functionally regulated by α2,6 sialic acids expressed on the same cell (cis-ligands). Here, we developed synthetic sialosides GSC718 and GSC839 that bind to CD22 with high affinity (IC50 ~100 nM), and inhibit ligand binding of CD22. When B cells are activated by B cell antigen receptor (BCR) ligation, both GSC718 and GSC839 downregulate proliferation of B cells, and this regulation requires both CD22 and α2,6 sialic acids. This result suggests that these sialosides regulate BCR ligation-induced B cell activation by reversing endogenous ligand-mediated regulation of CD22. By contrast, GSC718 and GSC839 augment B cell proliferation induced by TLR ligands or CD40 ligation, and this augmentation requires CD22 but not α2,6 sialic acids. Thus, these sialosides appear to enhance B cell activation by directly suppressing the inhibitory function of CD22 independently of endogenous ligand-mediated regulation. Moreover, GSC839 augments B cell proliferation that depends on both BCR ligation and CD40 ligation as is the case for in vivo B cell responses to antigens, and enhanced antibody production to the extent comparable to CpG oligonuleotides or a small amount of alum. Although these known adjuvants induce production of the inflammatory cytokines or accumulation of inflammatory cells, CD22-binding sialosides do not. Thus, synthetic sialosides that bind to CD22 with high-affinity modulate B cell activation through endogenous ligand-dependent and independent pathways, and carry an adjuvant activity without inducing inflammation. PMID:29725338

CD22-Binding Synthetic Sialosides Regulate B Lymphocyte Proliferation Through CD22 Ligand-Dependent and Independent Pathways, and Enhance Antibody Production in Mice.

PubMed

Matsubara, Naoko; Imamura, Akihiro; Yonemizu, Tatsuya; Akatsu, Chizuru; Yang, Hongrui; Ueki, Akiharu; Watanabe, Natsuki; Abdu-Allah, Hajjaj; Numoto, Nobutaka; Takematsu, Hiromu; Kitazume, Shinobu; Tedder, Thomas F; Marth, Jamey D; Ito, Nobutoshi; Ando, Hiromune; Ishida, Hideharu; Kiso, Makoto; Tsubata, Takeshi

2018-01-01

Sialic acid-binding immunoglobulin-like lectins (Siglecs) are expressed in various immune cells and most of them carry signaling functions. High-affinity synthetic sialoside ligands have been developed for various Siglecs. Therapeutic potentials of the nanoparticles and compounds that contain multiple numbers of these sialosides and other reagents such as toxins and antigens have been demonstrated. However, whether immune responses can be regulated by monomeric sialoside ligands has not yet been known. CD22 (also known as Siglec-2) is an inhibitory molecule preferentially expressed in B lymphocytes (B cells) and is constitutively bound and functionally regulated by α2,6 sialic acids expressed on the same cell (cis-ligands). Here, we developed synthetic sialosides GSC718 and GSC839 that bind to CD22 with high affinity (IC 50 ~100 nM), and inhibit ligand binding of CD22. When B cells are activated by B cell antigen receptor (BCR) ligation, both GSC718 and GSC839 downregulate proliferation of B cells, and this regulation requires both CD22 and α2,6 sialic acids. This result suggests that these sialosides regulate BCR ligation-induced B cell activation by reversing endogenous ligand-mediated regulation of CD22. By contrast, GSC718 and GSC839 augment B cell proliferation induced by TLR ligands or CD40 ligation, and this augmentation requires CD22 but not α2,6 sialic acids. Thus, these sialosides appear to enhance B cell activation by directly suppressing the inhibitory function of CD22 independently of endogenous ligand-mediated regulation. Moreover, GSC839 augments B cell proliferation that depends on both BCR ligation and CD40 ligation as is the case for in vivo B cell responses to antigens, and enhanced antibody production to the extent comparable to CpG oligonuleotides or a small amount of alum. Although these known adjuvants induce production of the inflammatory cytokines or accumulation of inflammatory cells, CD22-binding sialosides do not. Thus, synthetic sialosides that bind to CD22 with high-affinity modulate B cell activation through endogenous ligand-dependent and independent pathways, and carry an adjuvant activity without inducing inflammation.

Ternary complex factor SAP-1 is required for Erk-mediated thymocyte positive selection.

PubMed

Costello, Patrick S; Nicolas, Robert H; Watanabe, Yasuyuki; Rosewell, Ian; Treisman, Richard

2004-03-01

Thymocyte selection and differentiation requires extracellular signal-regulated kinase (Erk) signaling, but transcription factor substrates of Erk in thymocytes are unknown. We have characterized the function of SAP-1 (Elk4), an Erk-regulated transcription factor, in thymocyte development. Early thymocyte development was normal, but single-positive thymocyte and peripheral T cell numbers were reduced, reflecting a T cell-autonomous defect. T cell receptor-induced activation of SAP-1 target genes such as Egr1 was substantially impaired in double-positive thymocytes, although Erk activation was normal. Analysis of T cell receptor transgenes showed that positive selection was reduced by 80-90% in SAP-1-deficient mice; heterozygous mice showed a moderate defect. Negative selection was unimpaired. SAP-1 thus directly links Erk signaling to the transcriptional events required for thymocyte positive selection.

Immune drug discovery from venoms.

PubMed

Jimenez, Rocio; Ikonomopoulou, Maria P; Lopez, J Alejandro; Miles, John J

2018-01-01

This review catalogues recent advances in knowledge on venoms as standalone therapeutic agents or as blueprints for drug design, with an emphasis on venom-derived compounds that affects the immune system. We discuss venoms and venom-derived compounds that affect total immune cell numbers, immune cell proliferation, immune cell migration, immune cell phenotype and cytokine secretion. Identifying novel compounds that 'tune' the system, up-regulating the immune response during infectious disease and cancer and down-regulating the immune response during autoimmunity, will greatly expand the tool kit of human immunotherapeutics. Targeting these pathways may also open therapeutic options that alleviate symptoms of envenomation. Finally, combining recent advances in venomics with progress in low cost, high-throughput screening platforms will no doubt yield hundreds of prototype immune modulating compounds in the coming years. Copyright © 2017 Elsevier Ltd. All rights reserved.

Regulation of colonic epithelial cell turnover by IDO contributes to the innate susceptibility of SCID mice to Trichuris muris infection.

PubMed

Bell, L V; Else, K J

2011-04-01

Tryptophan catabolism via the kynurenine pathway is dependent on the enzyme Indoleamine 2,3-dioxygenase (IDO). Expression of IDO is upregulated in a number of inflammatory settings such as wounding and infection, and the resulting local tryptophan depletion may inhibit the replication of intracellular pathogens. Indo gene expression is upregulated in the gut during chronic infection with the mouse whipworm Trichuris muris. We demonstrate an increase in the rate of colonic epithelial cell turnover after inhibition of IDO in T. muris-infected SCID mice, leading to a significant expulsion of parasite burden. We identify the goblet cell as a novel source of IDO and present data revealing a new role for IDO in the regulation of epithelial cell turnover post-infectious challenge. © 2011 Blackwell Publishing Ltd.

[Recent research progress in immunomodulatory effects of Chinese herbal medicine on asthma treatment].

PubMed

Zhou, Fang-Fang; Xu, Zhao-Xia; Adila, Aipire; Li, Jin-Yao

2017-10-01

Asthma is a kind of chronic respiratory inflammation, commonly with breathlessness, chest tightness, coughing, recurrent episodes of wheezing and airflow obstruction, severely affecting human health. A variety of immunocytes are involved in this chronic disease. Chinese herbal medicine(CHM) has a long history in the treatment of asthma. A large number of studies have shown that CHM could ameliorate asthma symptoms through regulating cellular immune responses. This paper reviewed the studies of CHM on the regulation of immunocytes and their mechanisms in recent years, including the count of inflammatory cells, maturation of dendritic cells, balance of helper T cell subtypes, induction of regulatory T cells and intracellular signaling pathways. We also proposed the future research directions about the effects of CHM on asthma treatment. Copyright© by the Chinese Pharmaceutical Association.

Molecular Genetic Traits Influencing Maize Endosperm Development and Value: Closeout Report for DOE Grant DE-FG02-96ER20242

DOE Office of Scientific and Technical Information (OSTI.GOV)

Brian A. Larkins

2012-09-12

Development of the endosperm in cereal grasses entails different phases characterized by cell division, endoreduplication, accumulation of storage metabolites and cell death, which need to be carried out in an orderly fashion. While correct regulation of the cell cycle plays an essential role in endosperm development, the key regulatory factors and how the cell cycle interfaces with other pathways in this developmental context are largely unknown. We investigated the cyclin-dependent kinase (CDK)-retinoblastoma pathway and how it controls the cell cycle and coordinates it with other processes during maize endosperm development. Retinoblastoma-related (RBR) proteins may be inactivated through CDK-mediated phosphorylation, butmore » the identity of the responsible kinase in maize is unknown. We have previously shown that down-regulation of CDKA;1 severely inhibits the endoreduplication cell cycle and suggested that CDK may be an up-stream regulator of the retinoblastoma pathway. We discovered two types of maize RBR genes, RBR1 and RBR3, which differ in terms of structure, regulation and function. Phylogenetic analyses indicate that these genes may be distinctive features of the Poaceae. We found that RBR3 plays a positive rather than a negative role in DNA replication, cell transformation, and the expression of the minichromosome maintenance (MCM)2-7 family of DNA replication factors. These features are a paradigm shift in RBR gene function and appear to be unique within the RBR gene family. They suggest the existence in maize and related cereal crops of specific RBR/E2F-dependent pathways impinging on the cell cycle and development. RBR1 was down-regulated in transgenic endosperm using RNAi approaches. This resulted in the de-repression of a number of down-stream E2F targets, including RBR3, the MCM2-7 gene family, DNA methyltransferase (MET)1, CDKB;1, and the recently identified RBR4 gene. It also increased endosperm ploidy levels, stimulated the production of a larger number of cells, reduced the average cell size, and promoted programmed cell death. To test whether CDKA;1 inhibits RBR1 (through phosphorylation) in the pathway that leads to DNA synthesis and endoreduplication, the two CDKA;1 and RBR1 down-regulated mutants were crossed and their progeny analyzed. Our results indicate that CDKA;1 controls endoreduplication through an RBR1-dependent pathway. However, the ability of RBR1 to repress gene expression programs is independent from CDKA1, suggesting the presence of two differently regulated RBR1 activities in developing endosperm. One type of RBR1 activity controls E2F-dependent gene expression and is largely independent from CDKA;1, while another suppresses endoreduplication and can be inhibited by CDKA;1. In addition, RBR1 is part of a regulatory feedback loop that impinges on CDK activity. Together, these results indicate that the CDKA;1-RBR1 pathway integrates and controls different processes associated with endosperm development. Genome-wide analyses of the transcriptome, metabolome, and epigenetic mechanisms to understand how the cell cycle is coordinated with other pathways at a systems biology level are currently underway.« less

Zebrafish grainyhead-like1 is a common marker of different non-keratinocyte epidermal cell lineages, which segregate from each other in a Foxi3-dependent manner

PubMed Central

JÄNICKE, MARTINA; RENISCH, BJÖRN; HAMMERSCHMIDT, MATTHIAS

2012-01-01

Grainyhead/CP2 transcription factor family members are widely conserved among the animal kingdom and have been implicated in different developmental processes. Thus far, nothing has been known about their roles in zebrafish. Here we identify seven zebrafish grainyhead-like (grhl) / cp2 genes, with focus on grhl1, which is expressed in the periderm and in epidermal ionocyte progenitors, but downregulated when ionocytes differentiate. In addition, expression was detected in other “non-keratinocyte” cell types of the epidermis, such as pvalb8-expressing cells, which according to our lineage tracing experiments are derived from the same pool of progenitor cells like keratinocytes and ionocytes. Antisense morpholino oligonucleotide-based loss-of-function analysis revealed that grhl1 is dispensable for the development and function of all investigated epidermal cell types, but required as a negative regulator of its own transcription during ionocyte differentiation. Knockdown of the transcription factor Foxi3a, which is expressed in a subset of the grhl1 population, caused a loss of ionocytes and a corresponding increase in the number of pvalb8-expressing cells, while leaving the number of grhl1-positive cells unaltered. We propose that grhl1 is a novel common marker of all or most “non-keratinocyte” epidermal progenitors, and that the sub-functionalisation of these cells is regulated by differential positive and negative effects of Foxi3 factors. PMID:19757382

Integrative Analysis Reveals an Outcome-associated and Targetable Pattern of p53 and Cell Cycle Deregulation in Diffuse Large B-cell Lymphoma

PubMed Central

Monti, Stefano; Chapuy, Bjoern; Takeyama, Kunihiko; Rodig, Scott J; Hao, Yangsheng; Yeda, Kelly T.; Inguilizian, Haig; Mermel, Craig; Curie, Treeve; Dogan, Ahmed; Kutok, Jeffery L; Beroukim, Rameen; Neuberg, Donna; Habermann, Thomas; Getz, Gad; Kung, Andrew L; Golub, Todd R; Shipp, Margaret A

2013-01-01

Summary Diffuse large B-cell lymphoma (DLBCL) is a clinically and biologically heterogeneous disease with a high proliferation rate. By integrating copy number data with transcriptional profiles and performing pathway analysis in primary DLBCLs, we identified a comprehensive set of copy number alterations (CNAs) that decreased p53 activity and perturbed cell cycle regulation. Primary tumors either had multiple complementary alterations of p53 and cell cycle components or largely lacked these lesions. DLBCLs with p53 and cell cycle pathway CNAs had decreased abundance of p53 target transcripts and increased expression of E2F target genes and the Ki67 proliferation marker. CNAs of the CDKN2A-TP53-RB-E2F axis provide a structural basis for increased proliferation in DLBCL, predict outcome with current therapy and suggest targeted treatment approaches. PMID:22975378

Chemically Induced Reprogramming of Somatic Cells to Pluripotent Stem Cells and Neural Cells.

PubMed

Biswas, Dhruba; Jiang, Peng

2016-02-06

The ability to generate transplantable neural cells in a large quantity in the laboratory is a critical step in the field of developing stem cell regenerative medicine for neural repair. During the last few years, groundbreaking studies have shown that cell fate of adult somatic cells can be reprogrammed through lineage specific expression of transcription factors (TFs)-and defined culture conditions. This key concept has been used to identify a number of potent small molecules that could enhance the efficiency of reprogramming with TFs. Recently, a growing number of studies have shown that small molecules targeting specific epigenetic and signaling pathways can replace all of the reprogramming TFs. Here, we provide a detailed review of the studies reporting the generation of chemically induced pluripotent stem cells (ciPSCs), neural stem cells (ciNSCs), and neurons (ciN). We also discuss the main mechanisms of actions and the pathways that the small molecules regulate during chemical reprogramming.

Quantitative analysis of taste bud cell numbers in fungiform and soft palate taste buds of mice.

PubMed

Ohtubo, Yoshitaka; Yoshii, Kiyonori

2011-01-07

Mammalian taste bud cells (TBCs) consist of several cell types equipped with different taste receptor molecules, and hence the ratio of cell types in a taste bud constitutes the taste responses of the taste bud. Here we show that the population of immunohistochemically identified cell types per taste bud is proportional to the number of total TBCs in the taste bud or the area of the taste bud in fungiform papillae, and that the proportions differ among cell types. This result is applicable to soft palate taste buds. However, the density of almost all cell types, the population of cell types divided by the area of the respective taste buds, is significantly higher in soft palates. These results suggest that the turnover of TBCs is regulated to keep the ratio of each cell type constant, and that taste responsiveness is different between fungiform and soft palate taste buds. Copyright © 2010 Elsevier B.V. All rights reserved.

Antral G-cell in gastrin and gastrin-cholecystokinin knockout animals.

PubMed

Friis-Hansen, Lennart; Wierup, Nils; Rehfeld, Jens F; Sundler, Frank

2005-07-01

The antral hormone gastrin is the key regulator of gastric acid secretion, mucosal growth and differentiation. Gastrin is synthesized in the endocrine G-cells in the antroduodenal mucosa. We have now examined the way in which the loss of gastrin alone or gastrin plus cholecystokinin (CCK) affects the antral G-cell. Immunohistochemistry, radioimmunoassay and quantitative real-time polymerase chain reaction techniques were employed to examine the expression of genes belonging to the G-cell secretory pathway in gastrin and gastrin-CCK knockout mice. Transmission electron microscopy was used to examine the ultrastructure of the G-cells. The number of G-cells increased but the secretory granules were few and abnormally small in the G-cells of both mouse models compared with wildtypes. Thus, gastrin is not necessary for the formation of G-cells as such but the lack of gastrin reduces the number and size of their secretory granules suggesting that gastrin is vital for the formation and/or maintenance of secretory granules in G-cells.

A dynamic T cell–limited checkpoint regulates affinity-dependent B cell entry into the germinal center

PubMed Central

Schwickert, Tanja A.; Victora, Gabriel D.; Fooksman, David R.; Kamphorst, Alice O.; Mugnier, Monica R.; Gitlin, Alexander D.; Dustin, Michael L.

2011-01-01

The germinal center (GC) reaction is essential for the generation of the somatically hypermutated, high-affinity antibodies that mediate adaptive immunity. Entry into the GC is limited to a small number of B cell clones; however, the process by which this limited number of clones is selected is unclear. In this study, we demonstrate that low-affinity B cells intrinsically capable of seeding a GC reaction fail to expand and become activated in the presence of higher-affinity B cells even before GC coalescence. Live multiphoton imaging shows that selection is based on the amount of peptide–major histocompatibility complex (pMHC) presented to cognate T cells within clusters of responding B and T cells at the T–B border. We propose a model in which T cell help is restricted to the B cells with the highest amounts of pMHC, thus allowing for a dynamic affinity threshold to be imposed on antigen-binding B cells. PMID:21576382

Mast Cells in Gastrointestinal Disease

PubMed Central

Ramsay, David B.; Stephen, Sindu; Borum, Marie; Voltaggio, Lysandra

2010-01-01

The function of mast cells in allergic inflammatory reactions is well documented in the literature. Mast cells also play an important role in the regulation of gastrointestinal visceral sensitivity and vascular permeability. Several studies have noted an increased number of mast cells in the mucosa of patients with gastrointestinal diseases such as irritable bowel syndrome, mastocytic enterocolitis, and systemic mastocytosis. The role of mast cells in the symptomatology of these and other diseases has only recently been fully appreciated and could provide avenues for new therapeutic opportunities. This paper examines studies that have evaluated the role of mast cells in various gastrointestinal diseases. PMID:21301631

ROCK2 signaling is required to induce a subset of T follicular helper cells through opposing effects on STATs in autoimmune settings.

PubMed

Weiss, Jonathan M; Chen, Wei; Nyuydzefe, Melanie S; Trzeciak, Alissa; Flynn, Ryan; Tonra, James R; Marusic, Suzana; Blazar, Bruce R; Waksal, Samuel D; Zanin-Zhorov, Alexandra

2016-07-19

Rho-associated kinase 2 (ROCK2) determines the balance between human T helper 17 (TH17) cells and regulatory T (Treg) cells. We investigated its role in the generation of T follicular helper (TFH) cells, which help to generate antibody-producing B cells under normal and autoimmune conditions. Inhibiting ROCK2 in normal human T cells or peripheral blood mononuclear cells from patients with active systemic lupus erythematosus (SLE) decreased the number and function of TFH cells induced by activation ex vivo. Moreover, inhibition of ROCK2 activity decreased the abundance of the transcriptional regulator Bcl6 (B cell lymphoma 6) and increased that of Blimp1 by reducing the binding of signal transducer and activator of transcription 3 (STAT3) and increasing that of STAT5 to the promoters of the genes Bcl6 and PRDM1, respectively. In the MRL/lpr murine model of SLE, oral administration of the selective ROCK2 inhibitor KD025 resulted in a twofold reduction in the numbers of TFH cells and antibody-producing plasma cells in the spleen, as well as a decrease in the size of splenic germinal centers, which are the sites of interaction between TFH cells and B cells. KD025-treated mice showed a substantial improvement in both histological and clinical scores compared to those of untreated mice and had reduced amounts of Bcl6 and phosphorylated STAT3, as well as increased STAT5 phosphorylation. Together, these data suggest that ROCK2 signaling plays a critical role in controlling the development of TFH cells induced by autoimmune conditions through reciprocal regulation of STAT3 and STAT5 activation. Copyright © 2016, American Association for the Advancement of Science.

Causality, Randomness, Intelligibility, and the Epistemology of the Cell

PubMed Central

Dougherty, Edward R; Bittner, Michael L

2010-01-01

Because the basic unit of biology is the cell, biological knowledge is rooted in the epistemology of the cell, and because life is the salient characteristic of the cell, its epistemology must be centered on its livingness, not its constituent components. The organization and regulation of these components in the pursuit of life constitute the fundamental nature of the cell. Thus, regulation sits at the heart of biological knowledge of the cell and the extraordinary complexity of this regulation conditions the kind of knowledge that can be obtained, in particular, the representation and intelligibility of that knowledge. This paper is essentially split into two parts. The first part discusses the inadequacy of everyday intelligibility and intuition in science and the consequent need for scientific theories to be expressed mathematically without appeal to commonsense categories of understanding, such as causality. Having set the backdrop, the second part addresses biological knowledge. It briefly reviews modern scientific epistemology from a general perspective and then turns to the epistemology of the cell. In analogy with a multi-faceted factory, the cell utilizes a highly parallel distributed control system to maintain its organization and regulate its dynamical operation in the face of both internal and external changes. Hence, scientific knowledge is constituted by the mathematics of stochastic dynamical systems, which model the overall relational structure of the cell and how these structures evolve over time, stochasticity being a consequence of the need to ignore a large number of factors while modeling relatively few in an extremely complex environment. PMID:21119887

A Clb/Cdk1-mediated regulation of Fkh2 synchronizes CLB expression in the budding yeast cell cycle.

PubMed

Linke, Christian; Chasapi, Anastasia; González-Novo, Alberto; Al Sawad, Istabrak; Tognetti, Silvia; Klipp, Edda; Loog, Mart; Krobitsch, Sylvia; Posas, Francesc; Xenarios, Ioannis; Barberis, Matteo

2017-01-01

Precise timing of cell division is achieved by coupling waves of cyclin-dependent kinase (Cdk) activity with a transcriptional oscillator throughout cell cycle progression. Although details of transcription of cyclin genes are known, it is unclear which is the transcriptional cascade that modulates their expression in a timely fashion. Here, we demonstrate that a Clb/Cdk1-mediated regulation of the Fkh2 transcription factor synchronizes the temporal mitotic CLB expression in budding yeast. A simplified kinetic model of the cyclin/Cdk network predicts a linear cascade where a Clb/Cdk1-mediated regulation of an activator molecule drives CLB3 and CLB2 expression. Experimental validation highlights Fkh2 as modulator of CLB3 transcript levels, besides its role in regulating CLB2 expression. A Boolean model based on the minimal number of interactions needed to capture the information flow of the Clb/Cdk1 network supports the role of an activator molecule in the sequential activation, and oscillatory behavior, of mitotic Clb cyclins. This work illustrates how transcription and phosphorylation networks can be coupled by a Clb/Cdk1-mediated regulation that synchronizes them.

Transcriptional regulation induced by cAMP elevation in mouse Schwann cells

PubMed Central

Schmid, Daniela; Zeis, Thomas; Schaeren-Wiemers, Nicole

2014-01-01

In peripheral nerves, Schwann cell development is regulated by a variety of signals. Some of the aspects of Schwann cell differentiation can be reproduced in vitro in response to forskolin, an adenylyl cyclase activator elevating intracellular cAMP levels. Herein, the effect of forskolin treatment was investigated by a comprehensive genome-wide expression study on primary mouse Schwann cell cultures. Additional to myelin-related genes, many so far unconsidered genes were ascertained to be modulated by forskolin. One of the strongest differentially regulated gene transcripts was the transcription factor Olig1 (oligodendrocyte transcription factor 1), whose mRNA expression levels were reduced in treated Schwann cells. Olig1 protein was localized in myelinating and nonmyelinating Schwann cells within the sciatic nerve as well as in primary Schwann cells, proposing it as a novel transcription factor of the Schwann cell lineage. Data analysis further revealed that a number of differentially expressed genes in forskolin-treated Schwann cells were associated with the ECM (extracellular matrix), underlining its importance during Schwann cell differentiation in vitro. Comparison of samples derived from postnatal sciatic nerves and from both treated and untreated Schwann cell cultures showed considerable differences in gene expression between in vivo and in vitro, allowing us to separate Schwann cell autonomous from tissue-related changes. The whole data set of the cell culture microarray study is provided to offer an interactive search tool for genes of interest. PMID:24641305

The Transcriptomic Response of Rat Hepatic Stellate Cells to Endotoxin: Implications for Hepatic Inflammation and Immune Regulation

PubMed Central

Tandon, Ashish; Gandhi, Chandrashekhar R.

2013-01-01

With their location in the perisinusoidal space of Disse, hepatic stellate cells (HSCs) communicate with all of the liver cell types both by physical association (cell body as well as cytosolic processes penetrating into sinusoids through the endothelial fenestrations) and by producing several cytokines and chemokines. Bacterial lipopolysaccharide (LPS), circulating levels of which are elevated in liver diseases and transplantation, stimulates HSCs to produce increased amounts of cytokines and chemokines. Although recent research provides strong evidence for the role of HSCs in hepatic inflammation and immune regulation, the number of HSC-elaborated inflammatory and immune regulatory molecules may be much greater then known at the present time. Here we report time-dependent changes in the gene expression profile of inflammatory and immune-regulatory molecules in LPS-stimulated rat HSCs, and their validation by biochemical analyses. LPS strongly up-regulated LPS-response elements (TLR2 and TLR7) but did not affect TLR4 and down-regulated TLR9. LPS also up-regulated genes in the MAPK, NFκB, STAT, SOCS, IRAK and interferon signaling pathways, numerous CC and CXC chemokines and IL17F. Interestingly, LPS modulated genes related to TGFβ and HSC activation in a manner that would limit their activation and fibrogenic activity. The data indicate that LPS-stimulated HSCs become a major cell type in regulating hepatic inflammatory and immunological responses by altering expression of numerous relevant genes, and thus play a prominent role in hepatic pathophysiology including liver diseases and transplantation. PMID:24349206

Rab5a‑mediated autophagy regulates the phenotype and behavior of vascular smooth muscle cells.

PubMed

Tan, Jin-Yun; Jia, Luo-Qi; Shi, Wei-Hao; He, Qing; Zhu, Lei; Yu, Bo

2016-11-01

Rab5a, a key member of the Rab family of GTPases, was determined to be a regulator of vascular smooth muscle cell (VSMC) proliferation and migration. However, the exact regulatory mechanism remains unclear. As Rab5a has been shown to be associated with autophagy, which is essential for the conversion of VSMCs from a contractile to a synthetic phenotype in order to prevent cell death due to oxidative stress. The present study hypothesized that autophagy may be responsible for the proliferation and migration of VSMCs via the Rab5a protein. The aim of the present study was to evaluate the effect of Rab5a on autophagy in VSMCs. The human aorta vascular smooth muscle cell line, T/G HA‑VSMCs, was treated with small interfering (si)RNA against Rab5a and/or platelet‑derived growth factor (PDGF). Following treatment, the phenotype transition of the VSMCs was evaluated by detecting the mRNA and protien expression levels of VSMC molecular markers using reverse transcription‑quantitative polymerase chain reaction and western blotting, respectively. In addition, autophagy in VSMCs was evaluated by western blotting for autophagy‑associated proteins, flow cytometry of acidic vesicular organelles, punctate fluorescence of microtubule associated protein light chain 3 and transmission electron microscopy of typical scattered double‑membrane vacuolar structures. Additionally, the proliferation, migration, cell cycle and apoptotic response of VSMCs were detected by sulforhodamine B assay, transwell assay and flow cytometry, respectively. The results revealed that transfection with siRNA against Rab5a led to a significant decrease in Rab5a protein expression, while the reduced expression trend of Rab5a was rescued by intervention with PDGF. Furthermore, cells transfected with siRNA against Rab5a inhibited the autophagy of VSMCs. Downregulated Rab5a inhibited the phenotype transition of VSMCs. Additionally, downregulated Rab5a led to slowed cell growth, decreased numbers of migrated cells, decreased numbers of cells at the G0‑G1 phase and a higher apoptosis rate. However, PDGF significantly rescued these phenomena caused by siRNA against Rab5a. These results indicated that Rab5a‑mediated autophagy may regulate the phenotype transition and cell behavior of VSMCs through the activation of the extracellular‑regulated kinase 1/2 signaling pathway.

DREAM mediates cAMP-dependent, Ca2+-induced stimulation of GFAP gene expression and regulates cortical astrogliogenesis.

PubMed

Cebolla, Beatriz; Fernández-Pérez, Antonio; Perea, Gertrudis; Araque, Alfonso; Vallejo, Mario

2008-06-25

In the developing mouse brain, once the generation of neurons is mostly completed during the prenatal period, precisely coordinated signals act on competent neural precursors to direct their differentiation into astrocytes, which occurs mostly after birth. Among these signals, those provided by neurotrophic cytokines and bone morphogenetic proteins appear to have a key role in triggering the neurogenic to gliogenic switch and in regulating astrocyte numbers. In addition, we have reported previously that the neurotrophic peptide pituitary adenylate cyclase-activating polypeptide (PACAP) is able to promote astrocyte differentiation of cortical precursors via activation of a cAMP-dependent pathway. Signals acting on progenitor cells of the developing cortex to generate astrocytes activate glial fibrillary acidic protein (GFAP) gene expression, but the transcriptional mechanisms that regulate this activation are unclear. Here, we identify the previously known transcriptional repressor downstream regulatory element antagonist modulator (DREAM) as an activator of GFAP gene expression. We found that DREAM occupies specific sites on the GFAP promoter before and after differentiation is initiated by exposure of cortical progenitor cells to PACAP. PACAP raises intracellular calcium concentration via a mechanism that requires cAMP, and DREAM-mediated transactivation of the GFAP gene requires the integrity of calcium-binding domains. Cortical progenitor cells from dream(-/-) mice fail to express GFAP in response to PACAP. Moreover, the neonatal cortex of dream(-/-) mice exhibits a reduced number of astrocytes and increased number of neurons. These results identify the PACAP-cAMP-Ca(2+)-DREAM cascade as a new pathway to activate GFAP gene expression during astrocyte differentiation.

Stochastic dynamics of genetic broadcasting networks

NASA Astrophysics Data System (ADS)

Potoyan, Davit; Wolynes, Peter

The complex genetic programs of eukaryotic cells are often regulated by key transcription factors occupying or clearing out of a large number of genomic locations. Orchestrating the residence times of these factors is therefore important for the well organized functioning of a large network. The classic models of genetic switches sidestep this timing issue by assuming the binding of transcription factors to be governed entirely by thermodynamic protein-DNA affinities. Here we show that relying on passive thermodynamics and random release times can lead to a ''time-scale crisis'' of master genes that broadcast their signals to large number of binding sites. We demonstrate that this ''time-scale crisis'' can be resolved by actively regulating residence times through molecular stripping. We illustrate these ideas by studying the stochastic dynamics of the genetic network of the central eukaryotic master regulator NFκB which broadcasts its signals to many downstream genes that regulate immune response, apoptosis etc.

Protein tyrosine phosphatase, PTP1B, expression and activity in rat corneal endothelial cells

PubMed Central

Harris, Deshea L.

2007-01-01

Purpose The current studies were conducted to determine whether the protein tyrosine phosphatase, PTP1B, plays a role in regulating epidermal growth factor receptor (EGFR) Tyr992 phosphorylation and cell cycle entry in rat corneal endothelial cells. Methods Corneas were obtained from male Sprague-Dawley rats. PTP1B mRNA and protein expression were compared in confluent and subconfluent cells by RT-PCR and western blots. Immunocytochemistry was used to determine the subcellular localization of both PTP1B and EGFR following epidermal growth factor (EGF) stimulation. Western blots were used to analyze the time-dependent effect of EGF on phosphorylation of EGFR Tyr992 plus or minus CinnGEL 2Me, an inhibitor of PTP1B activity. The effect of PTP1B inhibition on cell cycle entry was determined by calculating the percent of Ki67-positive cells following EGF treatment. Results PTP1B mRNA expression was similar in confluent and subconfluent cells, but PTP1B protein was expressed at 3 fold higher levels in subconfluent cells. Positive staining for PTP1B was localized in vesicular structures below the plasma membrane. EGFR staining was located at cell-cell borders in untreated endothelium, but was mainly cytoplasmic by 15 min after EGF treatment. In control cultures, phosphorylation of EGFR Tyr992 peaked by 5 min following EGF stimulation and rapidly decreased to basal levels by 30 min. In cultures pretreated with CinnGEL 2Me, Tyr992 phosphorylation peaked 2 min following EGF addition and was consistently sustained at a higher level than controls until 60 min after treatment. By 18 h following EGF treatment, cultures pretreated with CinnGEL 2Me exhibited a 1.7 fold increase in the number of Ki67-positive cells compared with control cultures. Conclusions Comparison of PTP1B mRNA and protein levels indicates that PTP1B expression is regulated mainly at the protein level and is higher in subconfluent cells. PTP1B was located in vesicles below the plasma membrane. The fact that EGFR is internalized in response to EGF stimulation suggests that it could interact with and be regulated by PTP1B. The ability of PTP1B inhibitor to sustain EGFR Tyr992 phosphorylation and increase the number of Ki67-positive cells indicates that PTP1B plays a role in the negative regulation of EGF-induced signaling and helps suppress cell cycle entry. PMID:17563729

Fibroblast growth factor-2 up-regulates the expression of nestin through the Ras–Raf–ERK–Sp1 signaling axis in C6 glioma cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chang, Kai-Wei; Huang, Yuan-Li; Wong, Zong-Ruei

Highlights: •Nestin expression in C6 glioma cells is induced by FGF-2. •Nestin expression is induced by FGF-2 via de novo RNA and protein synthesis. •The FGFR inhibitor SU5402 blocks the FGF-2-induced nestin expression. •The mRNA of FGFR1 and 3 are detected in C6 glioma cells. •Ras–Raf–ERK–Sp1 signaling pathway is responsibe for FGF-2-induced nestin expression. -- Abstract: Nestin is a 240-kDa intermediate filament protein expressed mainly in neural and myogenic stem cells. Although a substantial number of studies have focused on the expression of nestin during development of the central nervous system, little is known about the factors that induce andmore » regulate its expression. Fibroblast growth factor-2 (FGF-2) is an effective mitogen and stimulates the proliferation and differentiation of a subset of nestin-expressing cells, including neural progenitor cells, glial precursor cells, and smooth muscle cells. To assess whether FGF-2 is a potent factor that induces the expression of nestin, C6 glioma cells were used. The results showed that nestin expression was up-regulated by FGF-2 via de novo RNA and protein synthesis. Our RT-PCR results showed that C6 glioma cells express FGFR1/3, and FGFRs is required for FGF-2-induced nestin expression. Further signaling analysis also revealed that FGF-2-induced nestin expression is mediated through FGFR–MAPK–ERK signaling axis and the transcriptional factor Sp1. These findings provide new insight into the regulation of nestin in glial system and enable the further studies on the function of nestin in glial cells.« less

MicroRNA Regulation of CD44+ Prostate Tumor Stem/Progenitor Cells and Prostate Cancer Development/Metastasis

DTIC Science & Technology

2013-05-01

prostate cancer stem cells. Cancer Res. 65, 10946–10951 (2005). 18 . Patrawala, L. et al. Highly purified CD44+ prostate cancer cells from xenograft ... xenograft tumors were of the human origin and morphologically epithelial with detectable cytokeratin 8 and 18 . RT-PCR analysis detected AR whereas...miR-301 16. SECURITY CLASSIFICATION OF: 17. LIMITATION OF ABSTRACT 18 . NUMBER OF PAGES 19a. NAME OF RESPONSIBLE PERSON USAMRMC a. REPORT

Yeast Genetics for Delineating Bax/Bcl Pathway of Cell Death Regulation.

DTIC Science & Technology

1998-07-01

differences in tosol. The cytosol also became electron dense ("cyto- the copy number of the episomal plasmid from which solic condensation"), similar to...Cell Death & Differ . 3, 229-236. (1993). The C. eheans cell death gene ccd-3 encodes a protein similar ¶Xhitc. K., Tahaoglu, E., and Steller, H. (1996...components may be used in different functional contexts. Similar modules might exist in metazoan apoptotic pathways. Even though yeast does not contain

Gene expression profiles in chondrosarcoma cells subjected to cyclic stretching and hydrostatic pressure. A cDNA array study.

PubMed

Karjalainen, Hannu M; Sironen, Reijo K; Elo, Mika A; Kaarniranta, Kai; Takigawa, Masaharu; Helminen, Heikki J; Lammi, Mikko J

2003-01-01

Mechanical forces have a profound effect on cartilage tissue and chondrocyte metabolism. Strenuous loading inhibits the cellular metabolism, while optimal level of loading at correct frequency raises an anabolic response in chondrocytes. In this study, we used Atlas Human Cancer cDNA array to investigate mRNA expression profiles in human chondrosarcoma cells stretched 8% for 6 hours at a frequency of 0.5 Hz. In addition, cultures were exposed to continuous and cyclic (0.5 Hz) 5 MPa hydrostatic pressure. Cyclic stretch had a more profound effect on the gene expression profiles than 5 MPa hydrostatic pressure. Several genes involved with the regulation of cell cycle were increased in stretched cells, as well as mRNAs for PDGF-B, glucose-1-phosphate uridylyltransferase, Tiam1, cdc37 homolog, Gem, integrin alpha6, and matrix metalloproteinase-3. Among down-regulated genes were plakoglobin, TGF-alpha, retinoic acid receptor-alpha and Wnt8b. A smaller number of changes was detected after pressure treatments. Plakoglobin was increased under cyclic and continuous 5 MPa hydrostatic pressure, while mitogen-activated protein kinase-9, proliferating cell nuclear antigen, Rad6, CD9 antigen, integrins alphaE and beta8, and vimentin were decreased. Cyclic and continuous pressurization induces a number of specific changes. In conclusion, a different set of genes were affected by three different types of mechanical stimuli applied on chondrosarcoma cells.

LRIG1 modulates aggressiveness of head and neck cancers by regulating EGFR-MAPK-SPHK1 signaling and extracellular matrix remodeling.

PubMed

Sheu, J J-C; Lee, C-C; Hua, C-H; Li, C-I; Lai, M-T; Lee, S-C; Cheng, J; Chen, C-M; Chan, C; Chao, S C-C; Chen, J-Y; Chang, J-Y; Lee, C-H

2014-03-13

EGFR overexpression and chromosome 3p deletion are two frequent events in head and neck cancers. We previously mapped the smallest region of recurrent copy-number loss at 3p12.2-p14.1. LRIG1, a negative regulator of EGFR, was found at 3p14, and its copy-number loss correlated with poor clinical outcome. Inducible expression of LRIG1 in head and neck cancer TW01 cells, a line with low LRIG1 levels, suppressed cell proliferation in vitro and tumor growth in vivo. Gene expression profiling, quantitative RT-PCR, chromatin immunoprecipitation, and western blot analysis demonstrated that LRIG1 modulated extracellular matrix (ECM) remodeling and EGFR-MAPK-SPHK1 transduction pathway by suppressing expression of EGFR ligands/activators, MMPs and SPHK1. In addition, LRIG1 induction triggered cell morphology changes and integrin inactivation, which coupled with reduced SNAI2 expression. By contrast, knockdown of endogenous LRIG1 in TW06 cells, a line with normal LRIG1 levels, significantly enhanced cell proliferation, migration and invasiveness. Such tumor-promoting effects could be abolished by specific MAPK or SPHK1 inhibitors. Our data suggest LRIG1 as a tumor suppressor for head and neck cancers; LRIG1 downregulation in cancer cells enhances EGFR-MAPK-SPHK1 signaling and ECM remodeling activity, leading to malignant phenotypes of head and neck cancers.

Role of adrenal hormones in regulating distal nephron structure and ion transport.

PubMed

Stanton, B A

1985-08-01

Mineralocorticoid levels are an important determinant of membrane area and ion transport in the renal initial (ICT) and cortical (CCT) collecting tubules. Adrenalectomy leads to a dramatic and specific decrease in basolateral membrane area of principal (P) cells and depresses sodium reabsorption and potassium secretion. Although aldosterone replacement for 10 days restores basolateral membrane area and ATPase activity to control levels and dramatically elevates ion transport, glucocorticoids have no effect on basolateral membrane area, ion transport, or ATPase. It is suggested that the aldosterone-induced amplification of membrane area occurs as a mechanism whereby cells increase the number of ATPase pumps in the basolateral membrane. An acute (of 2-3 h) increase in aldosterone, but not dexamethasone, also stimulates potassium transport by the ICT. Future studies will have to establish whether the acute stimulation of transport by aldosterone involves a change in basolateral membrane area. It is concluded that mineralocorticoids, but not glucocorticoids, regulate sodium and potassium transport by P cells of the ICT and CCT, in part, by determining the number of ATPase pumps available for transport.

Gene Expression Patterns Define Key Transcriptional Events InCell-Cycle Regulation By cAMP And Protein Kinase A

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zambon, Alexander C.; Zhang, Lingzhi; Minovitsky, Simon

Although a substantial number of hormones and drugs increase cellular cAMP levels, the global impact of cAMP and its major effector mechanism, protein kinase A (PKA), on gene expression is not known. Here we show that treatment of murine wild-type S49 lymphoma cells for 24 h with 8-(4-chlorophenylthio)-cAMP (8-CPTcAMP), a PKA-selective cAMP analog, alters the expression of approx equal to 4,500 of approx. equal to 13,600 unique genes. By contrast, gene expression was unaltered in Kin- S49 cells (that lack PKA) incubated with 8-CPTcAMP. Changes in mRNA and protein expression of several cell cycle regulators accompanied cAMP-induced G1-phase cell-cycle arrestmore » of wild-type S49 cells. Within 2h, 8-CPT-cAMP altered expression of 152 genes that contain evolutionarily conserved cAMP-response elements within 5 kb of transcriptional start sites, including the circadian clock gene Per1. Thus, cAMP through its activation of PKA produces extensive transcriptional regulation in eukaryotic cells. These transcriptional networks include a primary group of cAMP-response element-containing genes and secondary networks that include the circadian clock.« less

Notch signaling patterns neurogenic ectoderm and regulates the asymmetric division of neural progenitors in sea urchin embryos.

PubMed

Mellott, Dan O; Thisdelle, Jordan; Burke, Robert D

2017-10-01

We have examined regulation of neurogenesis by Delta/Notch signaling in sea urchin embryos. At gastrulation, neural progenitors enter S phase coincident with expression of Sp-SoxC. We used a BAC containing GFP knocked into the Sp-SoxC locus to label neural progenitors. Live imaging and immunolocalizations indicate that Sp-SoxC-expressing cells divide to produce pairs of adjacent cells expressing GFP. Over an interval of about 6 h, one cell fragments, undergoes apoptosis and expresses high levels of activated Caspase3. A Notch reporter indicates that Notch signaling is activated in cells adjacent to cells expressing Sp-SoxC. Inhibition of γ-secretase, injection of Sp-Delta morpholinos or CRISPR/Cas9-induced mutation of Sp-Delta results in supernumerary neural progenitors and neurons. Interfering with Notch signaling increases neural progenitor recruitment and pairs of neural progenitors. Thus, Notch signaling restricts the number of neural progenitors recruited and regulates the fate of progeny of the asymmetric division. We propose a model in which localized signaling converts ectodermal and ciliary band cells to neural progenitors that divide asymmetrically to produce a neural precursor and an apoptotic cell. © 2017. Published by The Company of Biologists Ltd.

mRNA Expression Profiling of Laser Microbeam Microdissected Cells from Slender Embryonic Structures

PubMed Central

Scheidl, Stefan J.; Nilsson, Sven; Kalén, Mattias; Hellström, Mats; Takemoto, Minoru; Håkansson, Joakim; Lindahl, Per

2002-01-01

Microarray hybridization has rapidly evolved as an important tool for genomic studies and studies of gene regulation at the transcriptome level. Expression profiles from homogenous samples such as yeast and mammalian cell cultures are currently extending our understanding of biology, whereas analyses of multicellular organisms are more difficult because of tissue complexity. The combination of laser microdissection, RNA amplification, and microarray hybridization has the potential to provide expression profiles from selected populations of cells in vivo. In this article, we present and evaluate an experimental procedure for global gene expression analysis of slender embryonic structures using laser microbeam microdissection and laser pressure catapulting. As a proof of principle, expression profiles from 1000 cells in the mouse embryonic (E9.5) dorsal aorta were generated and compared with profiles for captured mesenchymal cells located one cell diameter further away from the aortic lumen. A number of genes were overexpressed in the aorta, including 11 previously known markers for blood vessels. Among the blood vessel markers were endoglin, tie-2, PDGFB, and integrin-β1, that are important regulators of blood vessel formation. This demonstrates that microarray analysis of laser microbeam micro-dissected cells is sufficiently sensitive for identifying genes with regulative functions. PMID:11891179

FLASH is essential during early embryogenesis and cooperates with p73 to regulate histone gene transcription.

PubMed

De Cola, A; Bongiorno-Borbone, L; Bianchi, E; Barcaroli, D; Carletti, E; Knight, R A; Di Ilio, C; Melino, G; Sette, C; De Laurenzi, V

2012-02-02

Replication-dependent histone gene expression is a fundamental process occurring in S-phase under the control of the cyclin-E/CDK2 complex. This process is regulated by a number of proteins, including Flice-Associated Huge Protein (FLASH) (CASP8AP2), concentrated in specific nuclear organelles known as HLBs. FLASH regulates both histone gene transcription and mRNA maturation, and its downregulation in vitro results in the depletion of the histone pull and cell-cycle arrest in S-phase. Here we show that the transcription factor p73 binds to FLASH and is part of the complex that regulates histone gene transcription. Moreover, we created a novel gene trap to disrupt FLASH in mice, and we show that homozygous deletion of FLASH results in early embryonic lethality, owing to arrest of FLASH(-/-) embryos at the morula stage. These results indicate that FLASH is an essential, non-redundant regulator of histone transcription and cell cycle during embryogenesis.

The apical scaffold big bang binds to spectrins and regulates the growth of Drosophila melanogaster wing discs.

PubMed

Forest, Elodie; Logeay, Rémi; Géminard, Charles; Kantar, Diala; Frayssinoux, Florence; Heron-Milhavet, Lisa; Djiane, Alexandre

2018-03-05

During development, cell numbers are tightly regulated, ensuring that tissues and organs reach their correct size and shape. Recent evidence has highlighted the intricate connections between the cytoskeleton and the regulation of the key growth control Hippo pathway. Looking for apical scaffolds regulating tissue growth, we describe that Drosophila melanogaster big bang (Bbg), a poorly characterized multi-PDZ scaffold, controls epithelial tissue growth without affecting epithelial polarity and architecture. bbg -mutant tissues are smaller, with fewer cells that are less apically constricted than normal. We show that Bbg binds to and colocalizes tightly with the β-heavy-Spectrin/Kst subunit at the apical cortex and promotes Yki activity, F-actin enrichment, and the phosphorylation of the myosin II regulatory light chain Spaghetti squash. We propose a model in which the spectrin cytoskeleton recruits Bbg to the cortex, where Bbg promotes actomyosin contractility to regulate epithelial tissue growth. © 2018 Forest et al.

B-cell homeostasis requires complementary CD22 and BLyS/BR3 survival signals.

PubMed

Smith, Susan H; Haas, Karen M; Poe, Jonathan C; Yanaba, Koichi; Ward, Christopher D; Migone, Thi-Sau; Tedder, Thomas F

2010-08-01

Peripheral B-cell numbers are tightly regulated by homeostatic mechanisms that influence the transitional and mature B-cell compartments and dictate the size and clonotypic diversity of the B-cell repertoire. B-lymphocyte stimulator (BLyS, a trademark of Human Genome Sciences, Inc.) plays a key role in regulating peripheral B-cell homeostasis. CD22 also promotes peripheral B-cell survival through ligand-dependent mechanisms. The B-cell subsets affected by the absence of BLyS and CD22 signals overlap, suggesting that BLyS- and CD22-mediated survival are intertwined. To examine this, the effects of BLyS insufficiency following neutralizing BLyS mAb treatment in mice also treated with CD22 ligand-blocking mAb were examined. Combined targeting of the BLyS and CD22 survival pathways led to significantly greater clearance of recirculating bone marrow, blood, marginal zone and follicular B cells than either treatment alone. Likewise, BLyS blockade further reduced bone marrow, blood and spleen B-cell numbers in CD22(-/-) mice. Notably, BLyS receptor expression and downstream signaling were normal in CD22(-/-) B cells, suggesting that CD22 does not directly alter BLyS responsiveness. CD22 survival signals were likewise intact in the absence of BLyS, as CD22 mAb treatment depleted blood B cells from mice with impaired BLyS receptor 3 (BR3) signaling. Finally, enforced BclxL expression, which rescues BR3 impairment, did not affect B-cell depletion following CD22 mAb treatment. Thus, the current studies support a model whereby CD22 and BLyS promote the survival of overlapping B-cell subsets but contribute to their maintenance through independent and complementary signaling pathways.

Diverse microRNAs with convergent functions regulate tumorigenesis.

PubMed

Zhu, Min-Yan; Zhang, Wei; Yang, Tao

2016-02-01

MicroRNAs (miRNAs) regulate several biological processes, including tumorigenesis. In order to comprehend the roles of miRNAs in cancer, various screens were performed to investigate the changes in the expression levels of miRNAs that occur in different types of cancer. The present review focuses on the results of five recent screens, whereby a number of overlapping miRNAs were identified to be downregulated or differentially regulated, whereas no miRNAs were observed to be frequently upregulated. Furthermore, the majority of the miRNAs that were common to >1 screen were involved in signaling networks, including wingless-related integration site, receptor tyrosine kinase and transforming growth factor-β, or in cell cycle checkpoint control. The present review will discuss the aforementioned miRNAs implicated in cell cycle checkpoint control and signaling networks.

Cancer growth and its inhibition in terms of coherence.

PubMed

Popp, Fritz-Albert

2009-01-01

It is shown that a molecular origin for growth inhibition is rather unlikely because the cross-sectional area of inhibitory forces in a cell population cannot exceed more than about 10(-8) Dalton. A model of the time dependence of cell number N(t), where t is the time, is based on biophotons and explains without any contradiction to known experimental results growth regulation in terms of the factor a = 1/T, which stimulates the cell division rate dN/dt and the factor b = dT/dN(1/T(2)), which inhibits cell division. It accounts for the total cell division rate dN/dt = aN(t) - bN(2)(t). For adults, T is the coherence time of about 10(6) s, corresponding to the longest lifetime of cell organelles in men, while dT/dN = 10(-7) s corresponds to the resolution time of the cell population which is always the average time interval between two cell loss events. Our model follows a stringently holistic approach to describing a cell population as an entity, regulated by a fully coherent (biophoton) field.

Critical issues for engineering cord blood stem cells to produce insulin.

PubMed

Denner, Larry; Urban, Randall J

2008-09-01

The objectives of using cord blood stem cells for treating type 1 diabetes are simple in principle yet complex in biological and molecular mechanisms. These are defined by the complexity of the insulin-producing unit of the pancreas, the islet. Islets are composed of various cell types that arise from diverse lineages and communicate by hormones, growth factors and small-molecule mediators. These processes are regulated by integration of signal transduction pathways. While advances have been made to engineer umbilical cord blood stem cells to produce insulin, these studies only illuminate the potential of such cells to fulfil a necessary, but not sufficient, requirement for transplantation. The challenges ahead demand detailed understanding of molecular mechanisms to move from an opportunistic, phenotypic approach to transplantation and amelioration of blood glucose, to an orderly and logical approach to a biologically and medically meaningful solution. The issues include expansion to generate large numbers of cells, self-renewal to regulate the destiny of cord blood stem cells to repopulate the hematopoietic system, and multipotency of stem cells to generate the distinct cell types of an islet.

Mechanisms of Aquaporin-Facilitated Cancer Invasion and Metastasis

NASA Astrophysics Data System (ADS)

De Ieso, Michael L.; Yool, Andrea J.

2018-04-01

Cancer is a leading cause of death worldwide, and its incidence is rising with numbers expected to increase 70% in the next two decades. The fact that current mainline treatments for cancer patients are accompanied by debilitating side effects prompts a growing demand for new therapies that not only inhibit growth and proliferation of cancer cells, but also control invasion and metastasis. One class of targets gaining international attention is the aquaporins, a family of membrane-spanning water channels with diverse physiological functions and extensive tissue-specific distributions in humans. Aquaporins -1, -2, -3, -4, -5, -8, and -9 have been linked to roles in cancer proliferation, invasion and metastasis, but their mechanisms of action remain to be fully defined. Aquaporins are implicated in the metastatic cascade in processes of angiogenesis, cellular dissociation, migration and invasion. Cancer invasion and metastasis are proposed to be potentiated by aquaporins in boosting tumor angiogenesis, enhancing cell volume regulation, regulating cell-cell and cell-matrix adhesions, interacting with actin cytoskeleton, regulating proteases and extracellular-matrix degrading molecules, contributing to the regulation of epithelial-mesenchymal transitions, and interacting with signaling pathways enabling motility and invasion. Pharmacological modulators of aquaporin channels are being identified and tested for therapeutic potential, including compounds derived from loop diuretics, metal-containing organic compounds, plant natural products, and other small molecules. Further studies on aquaporin-dependent functions in cancer metastasis are needed to define the differential contributions of different classes of aquaporin channels to regulation of fluid balance, cell volume, small solute transport, signal transduction, their possible relevance as rate limiting steps, and potential values as therapeutic targets for proliferation and invasion.

DEPTOR regulates vascular endothelial cell activation and proinflammatory and angiogenic responses

PubMed Central

Bruneau, Sarah; Nakayama, Hironao; Woda, Craig B.; Flynn, Evelyn A.

2013-01-01

The maintenance of normal tissue homeostasis and the prevention of chronic inflammatory disease are dependent on the active process of inflammation resolution. In endothelial cells (ECs), proinflammation results from the activation of intracellular signaling responses and/or the inhibition of endogenous regulatory/pro-resolution signaling networks that, to date, are poorly defined. In this study, we find that DEP domain containing mTOR interacting protein (DEPTOR) is expressed in different microvascular ECs in vitro and in vivo, and using a small interfering RNA (siRNA) knockdown approach, we find that it regulates mammalian target of rapamycin complex 1 (mTORC1), extracellular signal-regulated kinase 1/2, and signal transducer and activator of transcription 1 activation in part through independent mechanisms. Moreover, using limited gene arrays, we observed that DEPTOR regulates EC activation including mRNA expression of the T-cell chemoattractant chemokines CXCL9, CXCL10, CXCL11, CX3CL1, CCL5, and CCL20 and the adhesion molecules intercellular adhesion molecule-1 and vascular cell adhesion molecule-1 (P < .05). DEPTOR siRNA-transfected ECs also bound increased numbers of peripheral blood mononuclear cells (P < .005) and CD3+ T cells (P < .005) in adhesion assays in vitro and had increased migration and angiogenic responses in spheroid sprouting (P < .01) and wound healing (P < .01) assays. Collectively, these findings define DEPTOR as a critical upstream regulator of EC activation responses and suggest that it plays an important role in endogenous mechanisms of anti-inflammation and pro-resolution. PMID:23881914

BMP signaling in the human fetal ovary is developmentally regulated and promotes primordial germ cell apoptosis.

PubMed

Childs, Andrew J; Kinnell, Hazel L; Collins, Craig S; Hogg, Kirsten; Bayne, Rosemary A L; Green, Samira J; McNeilly, Alan S; Anderson, Richard A

2010-08-01

Primordial germ cells (PGCs) are the embryonic precursors of gametes in the adult organism, and their development, differentiation, and survival are regulated by a combination of growth factors collectively known as the germ cell niche. Although many candidate niche components have been identified through studies on mouse PGCs, the growth factor composition of the human PGC niche has not been studied extensively. Here we report a detailed analysis of the expression of components of the bone morphogenetic protein (BMP) signaling apparatus in the human fetal ovary, from postmigratory PGC proliferation to the onset of primordial follicle formation. We find developmentally regulated and reciprocal patterns of expression of BMP2 and BMP4 and identify germ cells to be the exclusive targets of ovarian BMP signaling. By establishing long-term cultures of human fetal ovaries in which PGCs are retained within their physiological niche, we find that BMP4 negatively regulates postmigratory PGC numbers in the human fetal ovary by promoting PGC apoptosis. Finally, we report expression of both muscle segment homeobox (MSX)1 and MSX2 in the human fetal ovary and reveal a selective upregulation of MSX2 expression in human fetal ovary in response to BMP4, suggesting this gene may act as a downstream effector of BMP-induced apoptosis in the ovary, as in other systems. These data reveal for the first time growth factor regulation of human PGC development in a physiologically relevant context and have significant implications for the development of cultures systems for the in vitro maturation of germ cells, and their derivation from pluripotent stem cells.

Circadian locomotor output cycles kaput affects the proliferation and migration of breast cancer cells by regulating the expression of E-cadherin via IQ motif containing GTPase activating protein 1.

PubMed

Li, Xiaoxue; Wang, Siyang; Yang, Shuhong; Ying, Junjie; Yu, Hang; Yang, Chunlei; Liu, Yanyou; Wang, Yuhui; Cheng, Shuting; Xiao, Jing; Guo, Huiling; Jiang, Zhou; Wang, Zhengrong

2018-05-01

The circadian rhythm regulates numerous physiological activities, including sleep and wakefulness, behavior, immunity and metabolism. Previous studies have demonstrated that circadian rhythm disorder is associated with the occurrence of tumors. Responsible for regulating a number of functions, the Circadian locomotor output cycles kaput ( Clock ) gene is one of the core regulatory genes of circadian rhythm. The Clock gene has also been implicated in the occurrence and development of tumors in previously studies. The present study evaluated the role of the Clock gene in the proliferation and migration of mouse breast cancer 4T1 cells, and investigated its possible regulatory pathways and mechanisms. It was reported that downregulation of Clock facilitated the proliferation and migration of breast cancer cells. Further investigation revealed the involvement of IQ motif containing GTPase activating protein 1 (IQGAP1) protein expression in the Clock regulatory pathway, further influencing the expression of E-cadherin, a known proprietor of tumor cell migration and invasion. To the best of our knowledge, the present study is the first to report that Clock , acting through the regulation of the scaffolding protein IQGAP1, regulates the downstream expression of E-cadherin, thereby affecting tumor cell structure and motility. These results confirmed the role of Clock in breast cancer tumor etiology and provide insight regarding the molecular avenues of its regulatory nature, which may translate beyond breast cancer into other known functions of the gene.

Functional study of miR-27a in human hepatic stellate cells by proteomic analysis: comprehensive view and a role in myogenic tans-differentiation.

PubMed

Ji, Yuhua; Zhang, Jinsheng; Wang, Wenwen; Ji, Juling

2014-01-01

We previous reported that miR-27a regulates lipid metabolism and cell proliferation during hepatic stellate cells (HSCs) activation. To further explore the biological function and underlying mechanisms of miR-27a in HSCs, global protein expression affected by overexpression of miR-27a in HSCs was analyzed by a cleavable isotope-coded affinity tags (cICAT) based comparative proteomic approach. In the present study, 1267 non-redundant proteins were identified with unique accession numbers (score ≥1.3, i.e. confidence ≥95%), among which 1171 were quantified and 149 proteins (12.72%) were differentially expressed with a differential expression ratio of 1.5. We found that up-regulated proteins by miR-27a mainly participate in cell proliferation and myogenesis, while down-regulated proteins were the key enzymes involved in de novo lipid synthesis. The expression of a group of six miR-27a regulated proteins was validated and the function of one miR-27a regulated protein was further validated. The results not only delineated the underlying mechanism of miR-27a in modulating fat metabolism and cell proliferation, but also revealed a novel role of miR-27a in promoting myogenic tans-differentiation during HSCs activation. This study also exemplified proteomics strategy as a powerful tool for the functional study of miRNA.

The R2R3 MYB Transcription Factors FOUR LIPS and MYB88 Regulate Female Reproductive Development

PubMed Central

Lamb, Rebecca S.

2012-01-01

Gamete formation is an important step in the life cycle of sexually reproducing organisms. In flowering plants, haploid spores are formed after the meiotic division of spore mother cells. These spores develop into male and female gametophytes containing gametes after undergoing mitotic divisions. In the female, the megaspore mother cell undergoes meiosis forming four megaspores, of which one is functional and three degenerate. The megaspore then undergoes three mitotic cycles thus generating an embryo sac with eight nuclei. The embryo sac undergoes cellularization to form the mature seven-celled female gametophyte. Entry into and progression through meiosis is essential for megasporogenesis and subsequent megagametogenesis, but control of this process is not well understood. FOUR LIPS (FLP) and its paralogue MYB88, encoding R2R3 MYB transcription factors, have been extensively studied for their role in limiting the terminal division in stomatal development by direct regulation of the expression of cell cycle genes. Here it is demonstrated that FLP and MYB88 also regulate female reproduction. Both FLP and MYB88 are expressed during ovule development and their loss significantly increases the number of ovules produced by the placenta. Despite the presence of excess ovules, single and double mutants exhibit reduced seed set due to reduced female fertility. The sterility results at least in part from defective meiotic entry and progression. Therefore, FLP and MYB88 are important regulators of entry into megasporogenesis, and probably act via the regulation of cell cycle genes. PMID:22915737

Zinc Oxide Nanoparticles Demoted MDM2 Expression to Suppress TSLP-Induced Mast Cell Proliferation.

PubMed

Kim, Min-Ho; Jeong, Hyun-Ja

2016-03-01

Activation of murine double minute 2 (MDM2) through thymic stromal lymphopoietin (TSLP)-induced signal transducers and activators of transcription (STAT6) phosphorylation plays a critical role in proliferation and survival of mast cells. Previously, we reported that zinc oxide nanoparticles (ZnO-NP) effectively decrease the mast cell-mediated allergic inflammatory reactions. Here, we evaluated the effect of ZnO-NP on TSLP-induced proliferation of mast cells. ZnO-NP significantly reduced the number of BrdU-incorporating mast cells increased by TSLP. ZnO-NP decreased the expression of MDM2 through the blockade of STAT6 phosphorylation. TSLP increased the production and mRNA expression of interleukin-13 (a growth factor of mast cells), its increase was significantly decreased by ZnO-NP (10 μg/mL). ZnO-NP induced the down-regulation of Bcl2 (an anti-apoptotic factor) and up-regulation of Bax (an apoptotic factor) through the stabilization of p53 protein. However, ZnO-NP has no effect on caspase-3 activation, cytochrome c release into cytosol, and apoptosis-inducing factor translocation into nucleus in TSLP-stimulated cells. The results of the present study demonstrated that ZnO-NP inhibited the proliferation of mast cells through the regulation of MDM2 and p53 protein levels. These finding suggest that ZnO-NP could be improved mast cell-mediated various diseases.

The Zinc-finger protein ASCIZ regulates B cell development via DYNLL1 and Bim

PubMed Central

Jurado, Sabine; Gleeson, Kimberly; O’Donnell, Kristy; Izon, David J.; Walkley, Carl R.; Strasser, Andreas; Tarlinton, David M.

2012-01-01

Developing B lymphocytes expressing defective or autoreactive pre-B or B cell receptors (BCRs) are eliminated by programmed cell death, but how the balance between death and survival signals is regulated to prevent immunodeficiency and autoimmunity remains incompletely understood. In this study, we show that absence of the essential ATM (ataxia telangiectasia mutated) substrate Chk2-interacting Zn2+-finger protein (ASCIZ; also known as ATMIN/ZNF822), a protein with dual functions in the DNA damage response and as a transcription factor, leads to progressive cell loss from the pre-B stage onwards and severely diminished splenic B cell numbers in mice. This lymphopenia cannot be suppressed by deletion of p53 or complementation with a prearranged BCR, indicating that it is not caused by impaired DNA damage responses or defective V(D)J recombination. Instead, ASCIZ-deficient B cell precursors contain highly reduced levels of DYNLL1 (dynein light chain 1; LC8), a recently identified transcriptional target of ASCIZ, and normal B cell development can be restored by ectopic Dynll1 expression. Remarkably, the B cell lymphopenia in the absence of ASCIZ can also be fully suppressed by deletion of the proapoptotic DYNLL1 target Bim. Our findings demonstrate a key role for ASCIZ in regulating the survival of developing B cells by activating DYNLL1 expression, which may then modulate Bim-dependent apoptosis. PMID:22891272

Heteromeric Slick/Slack K+ channels show graded sensitivity to cell volume changes

PubMed Central

Hashem, Nadia; Calloe, Kirstine; Klaerke, Dan A.

2017-01-01

Slick and Slack high-conductance K+ channels are found in the CNS, kidneys, pancreas, among other organs, where they play an important role in cell excitability as well as in ion transport processes. They are both activated by Na+ and Cl- but show a differential regulation by cell volume changes. Slick has been shown to be regulated by cell volume changes, whereas Slack is insensitive. α-subunits of these channels form homomeric as well as heteromeric channels. It is the aim of this work to explore whether the subunit composition of the Slick/Slack heteromeric channel affects the response to osmotic challenges. In order to provide with the adequate water permeability to the cell membrane of Xenopus laevis oocytes, mRNA of aquaporin 1 was co-expressed with homomeric or heteromeric Slick and Slack α-subunits. Oocytes were superfused with hypotonic or hypertonic buffers and changes in currents were measured by two-electrode voltage clamp. This work presents the first heteromeric K+ channel with a characteristic graded sensitivity to small and fast changes in cell volume. Our results show that the cell volume sensitivity of Slick/Slack heteromeric channels is dependent on the number of volume sensitive Slick α-subunits in the tetrameric channels, giving rise to graded cell volume sensitivity. Regulation of the subunit composition of a channel may constitute a novel mechanism to determine volume sensitivity of cells. PMID:28222129

Heteromeric Slick/Slack K+ channels show graded sensitivity to cell volume changes.

PubMed

Tejada, Maria A; Hashem, Nadia; Calloe, Kirstine; Klaerke, Dan A

2017-01-01

Slick and Slack high-conductance K+ channels are found in the CNS, kidneys, pancreas, among other organs, where they play an important role in cell excitability as well as in ion transport processes. They are both activated by Na+ and Cl- but show a differential regulation by cell volume changes. Slick has been shown to be regulated by cell volume changes, whereas Slack is insensitive. α-subunits of these channels form homomeric as well as heteromeric channels. It is the aim of this work to explore whether the subunit composition of the Slick/Slack heteromeric channel affects the response to osmotic challenges. In order to provide with the adequate water permeability to the cell membrane of Xenopus laevis oocytes, mRNA of aquaporin 1 was co-expressed with homomeric or heteromeric Slick and Slack α-subunits. Oocytes were superfused with hypotonic or hypertonic buffers and changes in currents were measured by two-electrode voltage clamp. This work presents the first heteromeric K+ channel with a characteristic graded sensitivity to small and fast changes in cell volume. Our results show that the cell volume sensitivity of Slick/Slack heteromeric channels is dependent on the number of volume sensitive Slick α-subunits in the tetrameric channels, giving rise to graded cell volume sensitivity. Regulation of the subunit composition of a channel may constitute a novel mechanism to determine volume sensitivity of cells.

N-Cadherin Maintains the Healthy Biology of Nucleus Pulposus Cells under High-Magnitude Compression.

PubMed

Wang, Zhenyu; Leng, Jiali; Zhao, Yuguang; Yu, Dehai; Xu, Feng; Song, Qingxu; Qu, Zhigang; Zhuang, Xinming; Liu, Yi

2017-01-01

Mechanical load can regulate disc nucleus pulposus (NP) biology in terms of cell viability, matrix homeostasis and cell phenotype. N-cadherin (N-CDH) is a molecular marker of NP cells. This study investigated the role of N-CDH in maintaining NP cell phenotype, NP matrix synthesis and NP cell viability under high-magnitude compression. Rat NP cells seeded on scaffolds were perfusion-cultured using a self-developed perfusion bioreactor for 5 days. NP cell biology in terms of cell apoptosis, matrix biosynthesis and cell phenotype was studied after the cells were subjected to different compressive magnitudes (low- and high-magnitudes: 2% and 20% compressive deformation, respectively). Non-loaded NP cells were used as controls. Lentivirus-mediated N-CDH overexpression was used to further investigate the role of N-CDH under high-magnitude compression. The 20% deformation compression condition significantly decreased N-CDH expression compared with the 2% deformation compression and control conditions. Meanwhile, 20% deformation compression increased the number of apoptotic NP cells, up-regulated the expression of Bax and cleaved-caspase-3 and down-regulated the expression of Bcl-2, matrix macromolecules (aggrecan and collagen II) and NP cell markers (glypican-3, CAXII and keratin-19) compared with 2% deformation compression. Additionally, N-CDH overexpression attenuated the effects of 20% deformation compression on NP cell biology in relation to the designated parameters. N-CDH helps to restore the cell viability, matrix biosynthesis and cellular phenotype of NP cells under high-magnitude compression. © 2017 The Author(s). Published by S. Karger AG, Basel.

The LIM protein LIMD1 influences osteoblast differentiation and function

DOE Office of Scientific and Technical Information (OSTI.GOV)

Luderer, Hilary F.; Bai Shuting; Longmore, Gregory D.

2008-09-10

The balance between bone resorption and bone formation involves the coordinated activities of osteoblasts and osteoclasts. Communication between these two cell types is essential for maintenance of normal bone homeostasis; however, the mechanisms regulating this cross talk are not completely understood. Many factors that mediate differentiation and function of both osteoblasts and osteoclasts have been identified. The LIM protein Limd1 has been implicated in the regulation of stress osteoclastogenesis through an interaction with the p62/sequestosome protein. Here we show that Limd1 also influences osteoblast progenitor numbers, differentiation, and function. Limd1{sup -/-} calvarial osteoblasts display increased mineralization and accelerated differentiation. Whilemore » no significant differences in osteoblast number or function were detected in vivo, bone marrow stromal cells isolated from Limd1{sup -/-} mice contain significantly more osteoblast progenitors compared to wild type controls when cultured ex vivo. Furthermore, we observed a significant increase in nuclear {beta}-catenin staining in differentiating Limd1{sup -/-} calvarial osteoblasts suggesting that Limd1 is a negative regulator of canonical Wnt signaling in osteoblasts. These results demonstrate that Limd1 influences not only stress osteoclastogenesis but also osteoblast function and osteoblast progenitor commitment. Together, these data identify Limd1 as a novel regulator of both bone osetoclast and bone osteoblast development and function.« less

Cellular Energy Pathways as Novel Targets for the Therapy of Autosomal Dominant Polycystic Kidney Disease

DTIC Science & Technology

2017-09-01

AWARD NUMBER: W81XWH-15-1-0419 TITLE: Cellular Energy Pathways as Novel Targets for the Therapy of Autosomal Dominant Polycystic Kidney Disease...COVERED 1 Sep 2016 - 31 Aug 2017 4. TITLE AND SUBTITLE 5a. CONTRACT NUMBER Cellular Energy Pathways as Novel Targets for the Therapy of Autosomal...inappropriate cell growth, fluid secretion, and dysregulation of cellular energy metabolism. The enzyme AMPK regulates a number of cellular pathways, including

Two Forkhead transcription factors regulate the division of cardiac progenitor cells by a Polo-dependent pathway

PubMed Central

Ahmad, Shaad M.; Tansey, Terese R.; Busser, Brian W.; Nolte, Michael T.; Jeffries, Neal; Gisselbrecht, Stephen S.; Rusan, Nasser M.; Michelson, Alan M.

2012-01-01

SUMMARY The development of a complex organ requires the specification of appropriate numbers of each of its constituent cell types, as well as their proper differentiation and correct positioning relative to each other. During Drosophila cardiogenesis, all three of these processes are controlled by jumeau (jumu) and Checkpoint suppressor homologue (CHES-1-like), two genes encoding forkhead transcription factors that we discovered utilizing an integrated genetic, genomic and computational strategy for identifying genes expressed in the developing Drosophila heart. Both jumu and CHES-1-like are required during asymmetric cell division for the derivation of two distinct cardiac cell types from their mutual precursor, and in symmetric cell divisions that produce yet a third type of heart cell. jumu and CHES-1-like control the division of cardiac progenitors by regulating the activity of Polo, a kinase involved in multiple steps of mitosis. This pathway demonstrates how transcription factors integrate diverse developmental processes during organogenesis. PMID:22814603

Expression and Regulation of Cholecystokinin Receptor in the Chicken's Immune Organs and Cells.

PubMed

El-Kassas, Seham; Odemuyiwa, Solomon; Hajishengallis, George; Connell, Terry D; Nashar, Toufic O

2016-12-01

Cholecystokinin (CCK) is a neuropeptide that affects growth rate in chickens by regulating appetite. CCK peptides exert their function by binding to two identified receptors, CCKAR and CCKBR in the GI tract and the brain, respectively, as well as in other organs. In mammals, CCK/CCKAR interactions affect a number of immunological parameters, including regulation of lymphocytes and functioning of monocytes. Thus, food intake and growth can potentially be altered by infection and the resulting inflammatory immune response. It is uncertain, however, whether chicken express CCKAR in immune organs and cells, and, if so, whether CCKAR expression is regulated by pathogen derived inflammatory stimuli. Herein, we identify expression of CCKAR protein in chicken peripheral blood mononuclear cells (PBMC) including monocytes, and expression of the CCKAR gene in PBMC, thymus, bursa, and spleen, in selected commercial and pure chicken breeds. Further, stimulation with various types of E. coli heat-labile enterotoxins or lipopolysaccharide significantly regulated expression of CCKAR on monocytes in the different breeds. Ligation of CCKAR with antibodies in PBMC induced mobilization of Ca 2+ , indicating that CCKAR is signal competent. Injection with polyinosinic: polycytidylic acid (poly I:C), a synthetic analogue of double stranded viral RNA that binds Toll-Like Receptor-3 (TLR3), also regulated gene expressions of CCKAR and proinflammatory cytokines, in the different breeds. Interestingly, variations in the expression levels of proinflammatory cytokines in the different breeds were highly correlated with CCKAR expression levels. Taken together, these findings indicate that the physiological function of CCKAR in the chicken is tightly regulated in immune organs and cells by external inflammatory stimuli, which in turn regulate growth. This is the first report CCKAR expression in immune organs and cells, in any species, and the initial observation that CCKAR is regulated by inflammatory stimuli associated with bacterial and viral infection.

Expression and Regulation of Cholecystokinin Receptor in the Chicken's Immune Organs and Cells

PubMed Central

El-Kassas, Seham; Odemuyiwa, Solomon; Hajishengallis, George; Connell, Terry D; Nashar, Toufic O

2017-01-01

Cholecystokinin (CCK) is a neuropeptide that affects growth rate in chickens by regulating appetite. CCK peptides exert their function by binding to two identified receptors, CCKAR and CCKBR in the GI tract and the brain, respectively, as well as in other organs. In mammals, CCK/CCKAR interactions affect a number of immunological parameters, including regulation of lymphocytes and functioning of monocytes. Thus, food intake and growth can potentially be altered by infection and the resulting inflammatory immune response. It is uncertain, however, whether chicken express CCKAR in immune organs and cells, and, if so, whether CCKAR expression is regulated by pathogen derived inflammatory stimuli. Herein, we identify expression of CCKAR protein in chicken peripheral blood mononuclear cells (PBMC) including monocytes, and expression of the CCKAR gene in PBMC, thymus, bursa, and spleen, in selected commercial and pure chicken breeds. Further, stimulation with various types of E. coli heat-labile enterotoxins or lipopolysaccharide significantly regulated expression of CCKAR on monocytes in the different breeds. Ligation of CCKAR with antibodies in PBMC induced mobilization of Ca2+, indicating that CCKAR is signal competent. Injection with polyinosinic: polycytidylic acid (poly I:C), a synthetic analogue of double stranded viral RNA that binds Toll-Like Receptor-3 (TLR3), also regulated gene expressions of CCKAR and proinflammatory cytokines, in the different breeds. Interestingly, variations in the expression levels of proinflammatory cytokines in the different breeds were highly correlated with CCKAR expression levels. Taken together, these findings indicate that the physiological function of CCKAR in the chicken is tightly regulated in immune organs and cells by external inflammatory stimuli, which in turn regulate growth. This is the first report CCKAR expression in immune organs and cells, in any species, and the initial observation that CCKAR is regulated by inflammatory stimuli associated with bacterial and viral infection. PMID:28149670