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Novo Expression (novo + expression)
Kinds of Novo Expression Selected AbstractsComprehensive survey of carapacial ridge-specific genes in turtle implies co-option of some regulatory genes in carapace evolutionEVOLUTION AND DEVELOPMENT, Issue 1 2005Shigehiro Kuraku Summary The turtle shell is an evolutionary novelty in which the developmental pattern of the ribs is radically modified. In contrast to those of other amniotes, turtle ribs grow laterally into the dorsal dermis to form a carapace. The lateral margin of carapacial primordium is called the carapacial ridge (CR), and is thought to play an essential role in carapace patterning. To reveal the developmental mechanisms underlying this structure, we systematically screened for genes expressed specifically in the CR of the Chinese soft-shelled turtle, Pelodiscus sinensis, using microbead-based differential cDNA analysis and real-time reverse transcription-polymerase chain reaction. We identified orthologs of Sp5, cellular retinoic acid-binding protein-I (CRABP-I), adenomatous polyposis coli down-regulated 1 (APCDD1), and lymphoid enhancer-binding factor-1 (LEF-1). Although these genes are conserved throughout the major vertebrate lineages, comparison of their expression patterns with those in chicken and mouse indicated that these genes have acquired de novo expression in the CR in the turtle lineage. In association with the expression of LEF-1, the nuclear localization of ,-catenin protein was detected in the CR ectoderm, suggesting that the canonical Wnt signaling triggers carapace development. These findings indicate that the acquisition of the turtle shell did not involve the creation of novel genes, but was based on the co-option of pre-existing genes. [source] Activated B cells modified by electroporation of multiple mRNAs encoding immune stimulatory molecules are comparable to mature dendritic cells in inducing in vitro antigen-specific T-cell responsesIMMUNOLOGY, Issue 2 2008Jaewoo Lee Summary Ex-vivo -activated B cells are an alternative source of antigen-presenting cells (APCs) and a potential replacement for dendritic cells (DCs) in immunotherapy. However, the ability of ex-vivo -activated B cells to function as potent APCs has been a concern, especially when compared to DCs. Our study investigated whether modification of activated B cells with immune stimulatory molecules could enhance the ability of activated B cells to stimulate T cells. We show that murine splenic B cells, activated with a combination of Toll-like receptor agonist and agonistic anti-CD40, stimulated antigen-specific CD8+ T cells more efficiently than cells activated with Toll-like receptor agonist or anti-CD40 alone, probably by down-regulation of the immune regulatory cytokine interleukin-10 (IL-10). However, the activated B cells were still poor T-cell stimulators compared to mature DCs. Therefore, we modified the activated B cells by simultaneous electroporation of multiple messenger RNAs encoding costimulatory molecules (OX40L and 4-1BBL), cytokines (IL-12p35 and IL-12p40) and antigen. We found that de novo expression or overexpression of OX40L, 4-1BBL and IL-12p70 on activated B cells synergistically enhanced proliferation as well as IL-2 and interferon-, production by CD8+ T cells. Furthermore, the RNA-modified activated B cells induced antigen-specific cytotoxic T lymphocyte responses as efficiently as mature DCs in vitro. Unexpectedly, modified activated B cells were inferior to mature DCs at in vivo induction of CD8+ T-cell responses. In summary, activated B cells modified to express immune stimulatory molecules are a potent alternative to DCs in immunotherapy. [source] Induction of an antitumour adaptive immune response elicited by tumour cells expressing de novo B7-1 mainly depends on the anatomical site of their delivery: the dose applied regulates the expansion of the responseIMMUNOLOGY, Issue 4 2003Silvia Sartoris Summary De novo expression of costimulatory molecules in tumours generally increases their immunogenicity, but does not always induce a protective response against the parental tumour. This issue was addressed in the mouse Sp6 hybridoma model, comparing different immunization routes (subcutaneous, intraperitoneal and intravenous) and doses (0·5 × 106 and 5 × 106 cells) of Sp6 cells expressing de novo B7-1 (Sp6/B7). The results can be summarized as follows. First, de novo expression of B7-1 rendered Sp6 immunogenic, as it significantly reduced the tumour incidence to ,15% with all delivery routes and doses tested, whereas wild-type Sp6 was invariably tumorigenic (100% tumour incidence). Second, long-lasting protection against wild-type Sp6 was mainly achieved when immunization with Sp6/B7 was subcutaneous: a dose of 0·5 × 106 Sp6/B7 cells elicited protection that was confined to sites in the same anatomical quarter as the immunizing injection. Repeated injections of the same dose extended protection against wild-type Sp6 to other anatomical districts, as well as a single injection of a 10-fold higher dose (5 × 106 cells). Finally, Sp6-specific cytotoxic T-lymphocyte activity was detected in draining lymph nodes, and the splenic expansion of Sp6-specific cytotoxic T-lymphocyte precursors quantitatively correlated with the dose of antigen. We conclude that activation of a protective immune response against Sp6 depends on the local environment where the immunogenic form of the ,whole tumour cell antigen' is delivered. The antigen dose regulates the anatomical extent of the protective response. [source] Ion channels in toxicologyJOURNAL OF APPLIED TOXICOLOGY, Issue 6 2010Iván Restrepo-Angulo Abstract Ion channels play essential roles in human physiology and toxicology. Cardiac contraction, neural transmission, temperature sensing, insulin release, regulation of apoptosis, cellular pH and oxidative stress, as well as detection of active compounds from chilli, are some of the processes in which ion channels have an important role. Regulation of ion channels by several chemicals including those found in air, water and soil represents an interesting potential link between environmental pollution and human diseases; for instance, de novo expression of ion channels in response to exposure to carcinogens is being considered as a potential tool for cancer diagnosis and therapy. Non-specific binding of several drugs to ion channels is responsible for a huge number of undesirable side-effects, and testing guidelines for several drugs now require ion channel screening for pharmaceutical safety. Animal toxins targeting human ion channels have serious effects on the population and have also provided a remarkable tool to study the molecular structure and function of ion channels. In this review, we will summarize the participation of ion channels in biological processes extensively used in toxicological studies, including cardiac function, apoptosis and cell proliferation. Major findings on the adverse effects of drugs on ion channels as well as the regulation of these proteins by different chemicals, including some pesticides, are also reviewed. Association of ion channels and toxicology in several biological processes strongly suggests these proteins to be excellent candidates to follow the toxic effects of xenobiotics, and as potential early indicators of life-threatening situations including chronic degenerative diseases. Copyright © 2010 John Wiley & Sons, Ltd. [source] Circulating cell wall components derived from gram-negative, not gram-positive, bacteria cause a profound induction of the gene-encoding Toll-like receptor 2 in the CNSJOURNAL OF NEUROCHEMISTRY, Issue 3 2001Nathalie Laflamme The recent characterization of human homologs of Toll may be the missing link for the transduction events leading to nuclear factor-,B (NF-,B) activity and proinflammatory gene transcription during innate immune response. Mammalian cells may express as many as 10 distinct Toll-like receptors (TLRs), although TLR2 is a key receptor for recognizing cell wall components of Gram-positive bacteria. The present study investigated the effects of circulating bacterial cell wall components on the expression of the gene-encoding TLR2 across the mouse brain. Surprisingly, while Gram-negative components caused a robust increase in TLR2 transcription within the cerebral tissue, peptidoglycan (PGN) and lipoteichoic acid (LTA), either alone or combined, failed to modulate the receptor transcript. Indeed, the mRNA levels for TLR2 in the choroid plexus and few other regions of the brain remained similar between vehicle-, LTA-, PGN-, and LTA/PGN-administered mice at all the times evaluated (i.e. 30 min to 24 h post-intraperitoneal injection). This contrasts with the profound de novo expression of TLR2 following a single systemic injection of the lipopolysaccharide (LPS). The signal was first detected in regions devoid of blood,brain barrier and few blood vessels and microcapillaries. A second wave of TLR2 expression was also detected from these structures to their surrounding parenchymal cells that stained for a microglial marker iba1. The rapid induction of I,B, (index of NF-,B activity) and up-regulation of the adaptor protein MyD88 suggest that LPS-induced TLR2 transcription may be dependent on the NF-,B pathway. These data provide the evidence that TLR2 is not only present in the brain, but its encoding gene is regulated by cell wall components derived from Gram-negative, not Gram-positive, bacteria. The robust wave of TLR2-expressing microglial cells may have a determinant impact on the innate immune response that occurs in the brain during systemic infection by Gram-negative, not Gram-positive, bacteria. [source] Towards a therapeutic inhibition of dystrophin exon 23 splicing in mdx mouse muscle induced by antisense oligoribonucleotides (splicomers): target sequence optimisation using oligonucleotide arraysTHE JOURNAL OF GENE MEDICINE, Issue 10 2004Ian R. Graham Abstract Background The activity of synthetic antisense oligonucleotides (splicomers) designed to block pre-mRNA splicing at specific exons has been demonstrated in a number of model systems, including constitutively spliced exons in mouse dystrophin RNA. Splicomer reagents directed to Duchenne muscular dystrophy (DMD) RNAs might thus circumvent nonsense or frame-shifting mutations, leading to therapeutic expression of partially functional dystrophin, as occurs in the milder, allelic (Becker) form of the disease (BMD). Methods Functional and hybridisation array screens have been used to select optimised splicomers directed to exon 23 of dystrophin mRNA which carries a nonsense mutation in the mdx mouse. Splicomers were transfected into cultured primary muscle cells, and dystrophin mRNA assessed for exon exclusion. Splicomers were also administered to the muscles of mdx mice. Results Oligonucleotide array analyses with dystrophin pre-mRNA probes revealed strong and highly specific hybridisation patterns spanning the exon 23/intron 23 boundary, indicating an open secondary structure conformation in this region of the RNA. Functional screening of splicomer arrays by direct analysis of exon 23 RNA splicing in mdx muscle cultures identified a subset of biologically active reagents which target sequence elements associated with the 5, splice site region of dystrophin intron 23; splicomer-mediated exclusion of exon 23 was specific and dose-responsive up to a level exceeding 50% of dystrophin mRNA, and Western blotting demonstrated de novo expression of dystrophin protein at 2,5% of wild-type levels. Direct intramuscular administration of optimised splicomer reagents in vivo resulted in the reappearance of sarcolemmal dystrophin immunoreactivity in > 30% of muscle fibres in the mdx mouse Conclusions These results suggest that correctly designed splicomers may have direct therapeutic value in vivo, not only for DMD, but also for a range of other genetic disorders. Copyright © 2004 John Wiley & Sons, Ltd. [source] Renal ACE2 expression in human kidney disease,THE JOURNAL OF PATHOLOGY, Issue 5 2004AT Lely Abstract Angiotensin-converting enzyme 2 (ACE2) is a recently discovered homologue of angiotensin-converting enzyme (ACE) that is thought to counterbalance ACE. ACE2 cleaves angiotensin I and angiotensin II into the inactive angiotensin 1,9, and the vasodilator and anti-proliferative angiotensin 1,7, respectively. ACE2 is known to be present in human kidney, but no data on renal disease are available to date. Renal biopsies from 58 patients with diverse primary and secondary renal diseases were studied (hypertensive nephropathy n = 5, IgA glomerulopathy n = 8, minimal change nephropathy n = 7, diabetic nephropathy n = 8, focal glomerulosclerosis n = 5, vasculitis n = 7, and membranous glomerulopathy n = 18) in addition to 17 renal transplants and 18 samples from normal renal tissue. Immunohistochemical staining for ACE2 was scored semi-quantitatively. In control kidneys, ACE2 was present in tubular and glomerular epithelium and in vascular smooth muscle cells and the endothelium of interlobular arteries. In all primary and secondary renal diseases, and renal transplants, neo-expression of ACE2 was found in glomerular and peritubular capillary endothelium. There were no differences between the various renal disorders, or between acute and chronic rejection and control transplants. ACE inhibitor treatment did not alter ACE2 expression. In primary and secondary renal disease, and in transplanted kidneys, neo-expression of ACE2 occurs in glomerular and peritubular capillary endothelium. Further studies should elucidate the possible protective mechanisms involved in the de novo expression of ACE2 in renal disease. Copyright © 2004 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. [source] Transgenic neuronal nitric oxide synthase expression induces axotomy-like changes in adult motoneuronsTHE JOURNAL OF PHYSIOLOGY, Issue 18 2010Fernando Montero Dysregulation of protein expression, function and/or aggregation is a hallmark of a number of neuropathological conditions. Among them, upregulation and/or de novo expression of the neuronal isoform of nitric oxide (NO) synthase (nNOS) commonly occurs in diverse neurodegenerative diseases and in axotomized motoneurons. We used adenoviral (AVV) and lentiviral (LVV) vectors to study the effects of de novo nNOS expression on the functional properties and synaptic array of motoneurons. AVV-nNOS injection into the genioglossus muscle retrogradely transduced neonatal hypoglossal motoneurons (HMNs). Ratiometric real-time NO imaging confirmed that transduced HMNs generated NO gradients in brain parenchyma (space constant: ,12.3 ,m) in response to a glutamatergic stimulus. Unilateral AVV-nNOS microinjection in the hypoglossal nucleus of adult rats induced axotomy-like changes in HMNs. Specifically, we found alterations in axonal conduction properties and the recruitment order of motor units and reductions in responsiveness to synaptic drive and in the linear density of synaptophysin-positive puncta opposed to HMN somata. Functional alterations were fully prevented by chronic treatment with nNOS or soluble guanylyl cyclase inhibitors. Synaptic and functional changes were also completely avoided by prior intranuclear injection of a neuron-specific LVV system for miRNA-mediated nNOS knock-down (LVV-miR-shRNA/nNOS). Furthermore, synaptic and several functional changes evoked by XIIth nerve injury were to a large extent prevented by intranuclear administration of LVV-miR-shRNA/nNOS. We suggest that nNOS up-regulation creates a repulsive NO gradient for synaptic boutons underlying most of the functional impairment undergone by injured motoneurons. This further strengthens the case for nNOS targeting as a plausible strategy for treatment of peripheral neuropaties and neurodegenerative disorders. [source] De novo expression of Kv6.3 contributes to changes in vascular smooth muscle cell excitability in a hypertensive mice strainTHE JOURNAL OF PHYSIOLOGY, Issue 3 2009Alejandro Moreno-Domínguez Essential hypertension involves a gradual and sustained increase in total peripheral resistance, reflecting an increased vascular tone. This change associates with a depolarization of vascular myocytes, and relies on a change in the expression profile of voltage-dependent ion channels (mainly Ca2+ and K+ channels) that promotes arterial contraction. However, changes in expression and/or modulation of voltage-dependent K+ channels (Kv channels) are poorly defined, due to their large molecular diversity and their vascular bed-specific expression. Here we endeavor to characterize the molecular and functional expression of Kv channels in vascular smooth muscle cells (VSMCs) and their regulation in essential hypertension, by using VSMCs from resistance (mesenteric) or conduit (aortic) arteries obtained from a hypertensive inbred mice strain, BPH, and the corresponding normotensive strain, BPN. Real-time PCR reveals a differential distribution of Kv channel subunits in the different vascular beds as well as arterial bed-specific changes under hypertension. In mesenteric arteries, the most conspicuous change was the de novo expression of Kv6.3 (Kcng3) mRNA in hypertensive animals. The functional relevance of this change was studied by using patch-clamp techniques. VSMCs from BPH arteries were more depolarized than BPN ones, and showed significantly larger capacitance values. Moreover, Kv current density in BPH VSMCs is decreased mainly due to the diminished contribution of the Kv2 component. The kinetic and pharmacological profile of Kv2 currents suggests that the expression of Kv6.3 could contribute to the natural development of hypertension. [source] Ageing mechanisms: the role of telomere lossCLINICAL & EXPERIMENTAL DERMATOLOGY, Issue 7 2001P. Boukamp The ends of the chromosomes are capped by specialized structures, the telomeres. These are comprised of tracts of hexanucleotid sequences and, in combination with specific proteins, protect the chromosome against degradation, fusion events and as being recognized as 'damaged' DNA; thus, they guarantee chromosomal integrity. Due to deficiencies during DNA replication, the telomeres continuously loose part of their sequences and it has been proposed that this loss is the liming factor for the replicative capacity of a cell, i.e. telomeric loss is the counting mechanism - the internal clock of ageing. In order to proliferate indefinitely, the cells must prevent telomere erosion and this is mostly achieved by upregulation or de novo expression of the ribonucleoprotein complex telomerase. This enzyme, which has a reverse-transcriptase activity, is able to add telomeric sequences to the outer most ends off the telomeres and thereby stabilize or even elongate the telomeres. As telomerase is expressed in about 90% of all tumours while expression is absent in many somatic tissues, it is not surprising that the causal role of telomere erosion is presently the most favoured hypothesis of cellular ageing. [source] | |||||||||||||