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E Raz

Publications and source records attributed to E Raz.

At least 55 records · Page 3Linked to original sources
PubMed1998

Immunostimulatory DNA is a potent mucosal adjuvant.

Most proteins delivered to mucosal surfaces fail to induce mucosal or systemic immune responses. We demonstrate that a single intranasal (i.n.) coadministration of a model antigen (beta-galactosidase, beta-gal) with immunostimulatory sequence oligodeoxynucleotide (ISS-ODN) induces a mucosal IgA response equivalent to that induced by i.n. codelivery of beta-gal with cholera toxin (CT). Furthermore, i.n. and intradermal (i.d.) delivery of the beta-gal/ISS-ODN mix stimulates equivalent Th1-biased systemic immune responses with high-level cytotoxic T lymphocyte (CTL) activity. In contrast, i.n. immunization with beta-gal and CT results in a Th2-biased systemic immune response with poor CTL activity. Our data show that i.n. delivery of ISS-ODN provides effective adjuvant activity for the induction of both mucosal and systemic Th1-biased immune responses. This immunization approach deserves consideration in the development of vaccines against mucosal pathogens.

Adjuvants, Immunologic↗
PubMed1998

beta-lactamase as a marker for gene expression in live zebrafish embryos.

In this report we describe the development of a sensitive assay for gene expression in zebrafish embryos using beta-lactamase as a reporter gene. We show that injection of a green fluorescent substrate for beta-lactamase allows the detection of reporter gene expression in live embryos. The beta-lactamase enzyme catalyzes the hydrolysis of the substrate, thereby disrupting fluorescence resonance energy transfer from the donor to the acceptor dye in the molecule. As a result, a blue fluorescent product is produced and retained specifically in cells within which the enzyme is expressed. beta-Lactamase is therefore suitable for monitoring spatially restricted patterns of gene expression in the early embryo. We suggest that this new reporter system provides a major advancement in sensitivity over the existing methods for monitoring gene expression in vivo during early embryogenesis.

Animals↗
PubMed1998

Transposition of the nematode Caenorhabditis elegans Tc3 element in the zebrafish Danio rerio.

BACKGROUND: Transposable elements of the Tc1/mariner family are found in many species of the animal kingdom. It has been suggested that the widespread distribution of this transposon family resulted from horizontal transmission among different species. RESULTS: To test the ability of Tc1/mariner to cross species barriers, as well as to develop molecular genetic tools for studying zebrafish development, we determined the ability of the Tc3 transposon, a member of the Tc1/mariner family, to function in zebrafish. Tc3 transposons carrying sequences encoding the green fluorescent protein (GFP) were able to integrate in the fish genome by transposition. Integrated transposons expressed the GFP marker after germline transmission, and were capable of being mobilized upon introduction of transposase protein in trans. CONCLUSIONS: Our findings support models of horizontal transmission of Tc1/mariner elements between species. The work also establishes the basis for a novel method of transposon-mediated genetic transformation and for transposon-mediated genetic screens in zebrafish and other organisms.

Animals↗
PubMed1998

DNA immunotherapeutics: new potential treatment modalities for allergic disease.

Genetic immunization is a relatively new approach to vaccination, one that has generated considerable interest for its potential to prevent or treat a number of types of disease processes. Although initial work focused on potential applications in infectious disease, it was soon recognized that the antigen-specific Th1 response typically generated by genetic immunization protocols could also be useful for the treatment of allergic disease. In this review we present a summary of genetic vaccination and the related topic of immunostimulatory DNA sequences - which we collectively designate DNA immunotherapeutics - and discuss their potential for the prevention and therapy of allergic disease.

Animals↗
PubMed1997

Immunostimulatory DNA sequences function as T helper-1-promoting adjuvants.

An adjuvant role for certain short bacterial immunostimulatory DNA sequences (ISSs) has recently been proposed on the basis of their ability to stimulate T helper-1 (Th1) responses in gene-vaccinated animals. We report here that noncoding, ISS-enriched plasmid DNAs or ISS oligonucleotides (ISS-ODNs) potently stimulate immune responses to coadministered antigens. The ISS-DNAs suppress IgE synthesis, but promote IgG and interferon-gamma (IFN-gamma) production. They furthermore initiate the production of IFN-gamma, IFN-alpha, IFN-beta, and interleukins 12 and 18, all of which foster Th1 responses and enhance cell-mediated immunity. Consideration should be given to adding noncoding DNA adjuvants to inactivated or subunit viral vaccines that, by themselves, provide only partial protection from infection.

Adjuvants, Immunologic↗
PubMed1997

Spitz and Wingless, emanating from distinct borders, cooperate to establish cell fate across the Engrailed domain in the Drosophila epidermis.

A key step in development is the establishment of cell type diversity across a cellular field. Segmental patterning within the Drosophila embryonic epidermis is one paradigm for this process. At each parasegment boundary, cells expressing the Wnt family member Wingless confront cells expressing the homeoprotein Engrailed. The Engrailed-expressing cells normally differentiate as one of two alternative cell types. In investigating the generation of this cell type diversity among the 2-cell-wide Engrailed stripe, we previously showed that Wingless, expressed just anterior to the Engrailed cells, is essential for the specification of anterior Engrailed cell fate. In a screen for additional mutations affecting Engrailed cell fate, we identified anterior open/yan, a gene encoding an inhibitory ETS-domain transcription factor that is negatively regulated by the Rasl-MAP kinase signaling cascade. We find that Anterior Open must be inactivated for posterior Engrailed cells to adopt their correct fate. This is achieved by the EGF receptor (DER), which is required autonomously in the Engrailed cells to trigger the Ras1-MAP kinase pathway. Localized activation of DER is accomplished by restricted processing of the activating ligand, Spitz. Processing is confined to the cell row posterior to the Engrailed domain by the restricted expression of Rhomboid. These cells also express the inhibitory ligand Argos, which attenuates the activation of DER in cell rows more distant from the ligand source. Thus, distinct signals flank each border of the Engrailed domain, as Wingless is produced anteriorly and Spitz posteriorly. Since we also show that En cells have the capacity to respond to either Wingless or Spitz, these cells must choose their fate depending on the relative level of activation of the two pathways.

Animals↗
PubMed1996

Immunostimulatory DNA sequences necessary for effective intradermal gene immunization.

Vaccination with naked DNA elicits cellular and humoral immune responses that have a T helper cell type 1 bias. However, plasmid vectors expressing large amounts of gene product do not necessarily induce immune responses to the encoded antigens. Instead, the immunogenicity of plasmid DNA (pDNA) requires short immunostimulatory DNA sequences (ISS) that contain a CpG dinucleotide in a particular base context. Human monocytes transfected with pDNA or double-stranded oligonucleotides containing the ISS, but not those transfected with ISS-deficient pDNA or oligonucleotides, transcribed large amounts of interferon-alpha, interferon-beta, and interleukin-12. Although ISS are necessary for gene vaccination, they down-regulate gene expression and thus may interfere with gene replacement therapy by inducing proinflammatory cytokines.

Amino Acid Sequence↗
PubMed1996

Preferential induction of a Th1 immune response and inhibition of specific IgE antibody formation by plasmid DNA immunization.

We compared the antigen-specific antibody isotypes and lymphokine secretion by CD4+ T cells in BALB/c mice immunized intradermally with either Escherichia coli beta-galactosidase (beta-gal) or plasmid DNA (pDNA) encoding beta-gal in a cytomegalovirus-based expression vector (pCMV-LacZ). pCMV-LacZ induced mainly IgG2a, whereas beta-gal in saline or alum induced IgG1 and IgE beta-gal-specific antibodies. In addition, splenic CD4+ T helper (Th) cells isolated from pDNA-immunized mice secreted interferon-gamma but not interleukin (IL)-4 and IL-5, whereas Th cells from beta-gal-injected mice secreted IL-4 and IL-5 but not interferon-gamma after in vitro stimulation with antigen. Together these data demonstrate that pDNA immunization induced a T helper type 1 (Th1) response, whereas protein immunization induced a T helper type 2 (Th2) response to the same antigen. Interestingly, priming of mice with pCMV-LacZ prevented IgE antibody formation to a subsequent i.p. beta-gal in alum injection. This effect was antigen-specific, because priming with pCMV-LacZ did not inhibit IgE anti-ovalbumin antibody formation. Most importantly, intradermal immunization with pCMV-LacZ (but not pCMV-OVA) of beta-gal in alum-primed mice caused a 66-75% reduction of the IgE anti-beta-gal titer in 6 weeks. Also, pCMV-LacZ induced specific IgG2a antibody titers and interferon-gamma secretion by Th cells in the beta-gal in alum-primed mice. The data demonstrate that gene immunization induces a Th1 response that dominates over an ongoing protein-induced Th2 response in an antigen-specific manner. This suggests that immunization with pDNA encoding for allergens may provide a novel type of immunotherapy for allergic diseases.

Animals↗
PubMed1996

Green fluorescent protein marks skeletal muscle in murine cell lines and zebrafish.

The green fluorescent protein (GFP) acts as a vital dye upon the absorption of blue light. When the gfp gene is expressed in bacteria, flies or nematodes, green fluorescence can be directly observed in the living organism. We inserted the cDNA encoding this 238-amino-acid (aa) jellyfish protein into an expression vector containing the rat myosin light-chain enhancer (MLC-GFP) to evaluate its ability to serve as a muscle-specific marker. Transiently, as well as stably, transfected C2C12 cell lines produced high levels of GFP distributed homogeneously throughout the cytoplasm and was not toxic through several cell passages. Expression of MLC-GFP was strictly muscle-specific, since Cos 7 fibroblasts transfected with MLC-GFP did not fluoresce. When GFP and beta Gal markers were compared, the GFP signal was visible in the cytoplasm of the living cell, whereas visualization of beta Gal required fixation and resulted in deformation of the cells. When the MLC-GFP construct was injected into zebrafish embryos, muscle-specific gfp expression was apparent within 24 h of development. gfp expression was never observed in non-muscle tissues using the MLC-GFP construct. Transgenic fish continued to express high levels of gfp in skeletal muscle at 1.5 months, demonstrating that GFP is an effective marker of muscle cells in vivo.

Animals↗
PubMed1996

The Drosophila embryonic midline is the site of Spitz processing, and induces activation of the EGF receptor in the ventral ectoderm.

The Drosophila EGF receptor (DER) is activated by secreted Spitz to induce different cell fates in the ventral ectoderm. Processing of the precursor transmembrane Spitz to generate the secreted form was shown to be the limiting event, but the cells in which processing takes place and the mechanism that may generate a gradient of secreted Spitz in the ectoderm were not known. The ectodermal defects in single minded (sim) mutant embryos, in which the midline fails to develop, suggested that the midline cells contribute to patterning of the ventral ectoderm. This work shows that the midline provides the site for Spitz expression and processing. The Rhomboid and Star proteins are also expressed and required in the midline. The ectodermal defects of spitz, rho or Star mutant embryos could be rescued by inducing the expression of the respective normal genes only in the midline cells. Rho and Star thus function non-autonomously, and may be required for the production or processing of the Spitz precursor. Secreted Spitz is the only sim-dependent contribution of the midline to patterning the ectoderm, since the ventral defects observed in sim mutant embryos can be overcome by expression of secreted Spitz in the ectoderm. While ectopic expression of secreted Spitz in the ectoderm or mesoderm gave rise to ventralization of the embryo, increased expression of secreted Spitz in the midline did not lead to alterations in ectoderm patterning. A mechanism for adjustment to variable levels of secreted Spitz emanating from the midline may be provided by Argos, which forms an inhibitory feedback loop for DER activation. The production of secreted Spitz in the midline, may provide a stable source for graded DER activation in the ventral ectoderm.

Animals↗
PubMed1995

Modulation of disease activity in murine systemic lupus erythematosus by cytokine gene delivery.

Somatic gene therapy is a novel approach with the potential to achieve prolonged increases in circulating levels of peptide hormones and cytokines. The present study evaluates the effects of monthly, intramuscular injections of cDNA expression vectors encoding for transforming growth factor beta (TGF beta) or interleukin 2 (IL-2) on disease activity in the MRL/lpr/lpr murine model of systemic lupus erythematosus (SLE). Monthly injections of plasmids cDNA between 6 and 26 weeks significantly elevated the serum levels of TGF beta (P < 0.005) and IL-2 (P < 0.05) compared with a control plasmid without insert. TGF beta encoding plasmid had beneficial effects in murine SLE with a prolonged survival of 70% at 26 weeks compared with 40% in the control group, decreased anti-chromatin and rheumatoid factor antibodies and a 50% decrease in total IgG production. Renal function was improved with reduced BUN levels and kidney inflammation as estimated by an histologic score. Those beneficial effects occurred in the apparent absence of local or systemic side-effects. In contrast, IL-2 cDNA injections appeared harmful with a decreased survival to 20% at 26 weeks, enhanced total IgG synthesis and autoantibodies production with a 4.5-fold increase in antichromatin antibodies. These results indicate that somatic gene therapy may provide a simple, inexpensive and effective mechanism for the long-term control of autoimmune diseases.

Animals↗