scholarly journals Skin-Derived Precursor Cells as an In Vitro Modelling Tool for the Study of Type 1 Neurofibromatosis

2012 ◽  
Vol 2012 ◽  
pp. 1-9 ◽  
Author(s):  
Araika Gutiérrez-Rivera ◽  
Haizea Iribar ◽  
Anna Tuneu ◽  
Ander Izeta
Keyword(s):  
In Vitro Study ◽  
Neurofibromatosis Type ◽  
Precursor Cells ◽  
Cell Of Origin ◽  
Wild Type ◽  
Multipotent Progenitor Cells ◽  
Clonal Origin ◽  
Lines Of Evidence

The most characteristic feature of neurofibromatosis type 1 (NF1) is the development of neurofibromas. It has been suggested that these tumors are caused by somatic inactivation of the wild-typeNF1allele, but the cell that originally suffers this mutation remains controversial. Several lines of evidence support the clonal origin of these tumors, and it has been recently suggested that skin-derived precursor cells (SKPs) could be the cell of origin of dermal neurofibromas. Nullizygous (NF1−/−) SKPs do give rise to neurofibromas when transplanted to heterozygous mice. Moreover, a nullizygous population of cells that is S100βnegative is present in human neurofibromas, andNF1+/−multipotent progenitor cells are seemingly recruited to the tumor. This evidence supports the neurofibroma stem cell hypothesis and a putative involvement of SKPs in the aetiopathogenesis of the disease, suggesting that SKPs could become a valuable tool for the in vitro study of NF1.

scholarly journals Construction of an Excisable Bacterial Artificial Chromosome Containing a Full-Length Infectious Clone of Herpes Simplex Virus Type 1: Viruses Reconstituted from the Clone Exhibit Wild-Type Properties In Vitro and In Vivo

2003 ◽  
Vol 77 (2) ◽  
pp. 1382-1391 ◽  
Author(s):  
Michiko Tanaka ◽  
Hiroyuki Kagawa ◽  
Yuji Yamanashi ◽  
Tetsutaro Sata ◽  
Yasushi Kawaguchi
Keyword(s):  
Vero Cells ◽  
Infectious Molecular Clone ◽  
Wild Type ◽  
Molecular Clone ◽  
Viral Genes ◽  
Simplex Virus ◽  
Hsv 1

ABSTRACT In recent years, several laboratories have reported on the cloning of herpes simplex virus type 1 (HSV-1) genomes as bacterial artificial chromosomes (BACs) in Escherichia coli and on procedures to manipulate these genomes by using the bacterial recombination machinery. However, the HSV-BACs reported so far are either replication incompetent or infectious, with a deletion of one or more viral genes due to the BAC vector insertion. For use as a multipurpose clone in research on HSV-1, we attempted to generate infectious HSV-BACs containing the full genome of HSV-1 without any loss of viral genes. Our results were as follows. (i) E. coli (YEbac102) harboring the full-length HSV-1 genome (pYEbac102) in which a BAC flanked by loxP sites was inserted into the intergenic region between UL3 and UL4 was constructed. (ii) pYEbac102 was an infectious molecular clone, given that its transfection into rabbit skin cells resulted in production of infectious virus (YK304). (iii) The BAC vector sequence was almost perfectly excisable from the genome of the reconstituted virus YK304 by coinfection of Vero cells with YK304 and a recombinant adenovirus, AxCANCre, expressing Cre recombinase. (iv) As far as was examined, the reconstituted viruses from pYEbac102 could not be phenotypically differentiated from wild-type viruses in vitro and in vivo. Thus, the viruses grew as well in Vero cells as did the wild-type virus and exhibited wild-type virulence in mice on intracerebral inoculation. (v) The infectious molecular clone pYEbac102 is in fact useful for mutagenesis of the HSV-1 genome by bacterial genetics, and a recombinant virus carrying amino acid substitutions in both copies of the α0 gene was generated. pYEbac102 will have multiple applications to the rapid generation of genetically engineered HSV-1 recombinants in basic research into HSV-1 and in the development of HSV vectors in human therapy.

scholarly journals Distinct progenitor behavior underlying neocortical gliogenesis related to tumorigenesis

2020 ◽  
Author(s):  
Zhongfu Shen ◽  
Yang Lin ◽  
Jiajun Yang ◽  
David J. Jörg ◽  
Yuwei Peng ◽  
...  
Keyword(s):  
Progenitor Cell ◽  
Neurofibromatosis Type ◽  
Primary Brain Tumor ◽  
Precursor Cells ◽  
Cell Behavior ◽  
Neocortical Neurons ◽  
Glial Progenitors ◽  
Radial Glial ◽  
Oligodendrocyte Precursor

SUMMARYRadial glial progenitors (RGPs) are responsible for producing the vast majority of neurons and glia in the neocortex. While RGP behavior and progressive generation of neocortical neurons have been delineated, the exact process of neocortical gliogenesis remains elusive. Here, we report the precise progenitor cell behavior and gliogenesis program at single-cell resolution in the mouse neocortex. RGPs transition from neurogenesis to gliogenesis progressively, producing astrocytes, oligodendrocytes, or both in well-defined propensities of 60%:15%:25%, respectively, via fate-restricted “intermediate” precursor cells. While the total number of precursor cells generated by individual RGPs appears stochastic, the output of individual precursor cells exhibit clear patterns in number and subtype, and form discrete local subclusters. Clonal loss of tumor suppressor Neurofibromatosis type 1 leads to excessive production of glia selectively, especially oligodendrocyte precursor cells. These results delineate the cellular program of neocortical gliogenesis quantitatively and suggest the cellular and lineage origin of primary brain tumor.

scholarly journals Codon Substitution Mutations at Two Positions in the L Polymerase Protein of Human Parainfluenza Virus Type 1 Yield Viruses with a Spectrum of Attenuation In Vivo and Increased Phenotypic Stability In Vitro

2004 ◽  
Vol 78 (4) ◽  
pp. 2029-2036 ◽  
Author(s):  
Josephine M. McAuliffe ◽  
Sonja R. Surman ◽  
Jason T. Newman ◽  
Jeffrey M. Riggs ◽  
Peter L. Collins ◽  
...  
Keyword(s):  
Amino Acid ◽  
Nucleotide Substitution ◽  
Wild Type ◽  
Phenotypic Stability ◽  
Single Nucleotide ◽  
Codon Substitution ◽  
Substitution Mutations ◽  
Human Parainfluenza Virus

ABSTRACT The Y942H and L992F temperature-sensitive (ts) and attenuating amino acid substitution mutations, previously identified in the L polymerase of the HPIV3cp45 vaccine candidate, were introduced into homologous positions of the L polymerase of recombinant human parainfluenza virus type 1 (rHPIV1). In rHPIV1, the Y942H mutation specified the ts phenotype in vitro and the attenuation (att) phenotype in hamsters, whereas the L992F mutation specified neither phenotype. Each of these codon mutations was generated by a single nucleotide substitution and therefore had the potential to readily revert to a codon specifying the wild-type amino acid residue. We introduced alternative amino acid assignments at codon 942 or 992 as a strategy to increase genetic stability and to generate mutants that exhibit a range of attenuation. Twenty-three recombinants with codon substitutions at position 942 or 992 of the L protein were viable. One highly ts and att mutant, the Y942A virus, which had a difference of three nucleotides from the codon encoding a wild-type tyrosine, also possessed a high level of genetic and phenotypic stability upon serial passage in vitro at restrictive temperatures compared to that of the parent Y942H virus, which possessed a single nucleotide substitution. We obtained mutants with substitutions at position 992 that, in contrast to the L992F virus, possessed the ts and att phenotypes. These findings identify the use of alternative codon substitution mutations as a method that can be used to generate candidate vaccine viruses with increased genetic stability and/or a modified level of attenuation.

scholarly journals Thermolabile L-A virus-like particles from pet18 mutants of Saccharomyces cerevisiae.

1986 ◽  
Vol 6 (2) ◽  
pp. 404-410 ◽  
Author(s):  
T Fujimura ◽  
R B Wickner
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Rna Polymerase ◽  
Polymerase Activity ◽  
Nonpermissive Temperature ◽  
Wild Type ◽  
Rna Dependent Rna Polymerase ◽  
Virus Like Particles ◽  
Rna Polymerase Activity ◽  
Lines Of Evidence

pet18 mutations in Saccharomyces cerevisiae confer on the cell the inability to maintain either L-A or M double-stranded RNAs (dsRNAs) at the nonpermissive temperature. In in vitro experiments, we examined the effects of pet18 mutations on the RNA-dependent RNA polymerase activity associated with virus-like particles (VLPs). pet18 mutations caused thermolabile RNA polymerase activity of L-A VLPs, and this thermolability was found to be due to the instability of the L-A VLP structure. The pet18 mutations did not affect RNA polymerase activity of M VLPs. Furthermore, the temperature sensitivity of wild-type L-A RNA polymerase differed substantially from that of M RNA polymerase. From these results, and from other genetic and biochemical lines of evidence which suggest that replication of M dsRNA requires the presence of L-A dsRNA, we propose that the primary effect of the pet18 mutation is on the L-A VLP structure and that the inability of pet18 mutants to maintain M dsRNA comes from the loss of L-A dsRNA.

Mutations of the NF1 Gene in Children With Juvenile Myelomonocytic Leukemia Without Clinical Evidence of Neurofibromatosis, Type 1

Blood ◽  
1998 ◽  
Vol 92 (1) ◽  
pp. 267-272 ◽  
Author(s):  
Lucy E. Side ◽  
Peter D. Emanuel ◽  
Brigit Taylor ◽  
Janet Franklin ◽  
Patricia Thompson ◽  
...  
Keyword(s):  
Clinical Diagnosis ◽  
Neurofibromatosis Type 1 ◽  
Neurofibromatosis Type ◽  
Juvenile Myelomonocytic Leukemia ◽  
Translation System ◽  
Polymorphism Analysis ◽  
Single Stranded Conformational Polymorphism ◽  
Myelomonocytic Leukemia

Juvenile myelomonocytic leukemia (JMML) is a pediatric myelodysplastic syndrome that is associated with neurofibromatosis, type 1 (NF1). The NF1 tumor suppressor gene encodes neurofibromin, which regulates the growth of immature myeloid cells by accelerating guanosine triphosphate hydrolysis on Ras proteins. The purpose of this study was to determine if the NF1gene was involved in the pathogenesis of JMML in children without a clinical diagnosis of NF1. An in vitro transcription and translation system was used to screen JMML marrows from 20 children for NF1mutations that resulted in a truncated protein. Single-stranded conformational polymorphism analysis was used to detect RASpoint mutations in these samples. We confirmed mutations of NF1in three leukemias, one of which also showed loss of the normalNF1 allele. An NF1 mutation was detected in normal tissue from the only patient tested and this suggests that JMML may be the presenting feature of NF1 in some children. Activating RASmutations were found in four patients; as expected, none of these samples harbored NF1 mutations. Because 10% to 14% of children with JMML have a clinical diagnosis of NF1, these data are consistent with the existence of NF1 mutations in approximately 30% of JMML cases.

Activation of human aortic endothelial cells by LDL from Type 1 diabetic patients: an in vitro study

2002 ◽  
Vol 165 (1) ◽  
pp. 69-77 ◽  
Author(s):  
Rosa A Rabini ◽  
Arianna Vignini ◽  
Eleonora Salvolini ◽  
Roberto Staffolani ◽  
Daniela Martarelli ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
In Vitro Study ◽  
Vitro Study ◽  
Diabetic Patients ◽  
Aortic Endothelial Cells ◽  
Type 1 Diabetic Patients ◽  
Human Aortic Endothelial Cells ◽  
Aortic Endothelial

Antiviral Activity of Dicrocephala Integrifolia (Kuntze) Against Herpes Simplex Type-1 Virus: An in vitro Study

2018 ◽  
Vol 8 (2) ◽  
pp. 81-85 ◽  
Author(s):  
Abdi Hussein Hadun ◽  
James Mucunu Mbaria ◽  
Gabriel Oluga Aboge ◽  
Mitchel Otieno Okumu ◽  
Antony Letoyah Yiaile
Keyword(s):  
Herpes Simplex ◽  
Antiviral Activity ◽  
In Vitro Study ◽  
Vitro Study ◽  
Herpes Simplex Type

scholarly journals Regions of the Herpes Simplex Virus Type 1 Latency-Associated Transcript That Protect Cells from Apoptosis In Vitro and Protect Neuronal Cells In Vivo

2002 ◽  
Vol 76 (2) ◽  
pp. 717-729 ◽  
Author(s):  
Maryam Ahmed ◽  
Martin Lock ◽  
Cathie G. Miller ◽  
Nigel W. Fraser
Keyword(s):  
Herpes Simplex ◽  
Trigeminal Ganglia ◽  
Wild Type ◽  
Exon 1 ◽  
Induced Apoptosis ◽  
Simplex Virus ◽  
In Cells

ABSTRACT Recent studies have suggested that the latency-associated transcript (LAT) region of herpes simplex virus type 1 (HSV-1) is effective at blocking virus-induced apoptosis both in vitro and in the trigeminal ganglia of acutely infected rabbits (Inman et al., J. Virol. 75:3636–3646, 2001; Perng et al., Science 287:1500–1503, 2000). By transfecting cells with a construct expressing the Pst-Mlu segment of the LAT, encompassing the LAT exon 1, the stable 2.0-kb intron, and 5′ part of exon 2, we confirmed that this region was able to diminish the onset of programmed cell death initiated by anti-Fas and camptothecin treatment. In addition, caspase 8-induced apoptosis was specifically inhibited in cells expressing the Pst-Mlu LAT fragment. To further delineate the minimal region of LAT that is necessary for this antiapoptotic function, LAT mutants were used in our cotransfection assays. In HeLa cells, the plasmids lacking exon sequences were the least effective at blocking apoptosis. However, similar to previous work (Inman et al., op. cit.), our data also indicated that the 5′ end of the stable 2.0-kb LAT intron appeared to contribute to the promotion of cell survival. Furthermore, cells productively infected with the 17N/H LAT mutant virus, a virus deleted in the LAT promoter, exon 1, and about half of the intron, exhibited a greater degree of DNA fragmentation than cells infected with wild-type HSV-1. These data support the finding that the exon 1 and 2.0-kb intron region of the LAT transcription unit display an antiapoptotic function both in transfected cells and in the context of the virus infection in vitro. In trigeminal ganglia of mice acutely infected with the wild-type virus, 17, and 17ΔSty, a virus lacking most of exon 1, apoptosis was not detected in cells that were positive for virus particles. However, dual staining was observed in cells from mice infected with 17N/H virus, indicating that the LAT antiapoptotic function demonstrated in cells transfected by LAT-expressing constructs may also play a role in protecting cells from virus-induced apoptosis during acute viral infection in vivo.

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