Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.2.1.23 (beta-galactosidase)
 14,648 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A recombinant strain of Listeria monocytogenes that stably and constitutively expresses Escherichia coli beta-galactosidase was used as a live vaccine vector. BALB/c mice were immunized orally or parenterally with the recombinant L. monocytogenes, and their cellular and humoral immune responses to beta-galactosidase were measured. Spleen cells taken 1 week after oral inoculation or 5 weeks after oral or parenteral inoculation (with a boost at 4 weeks) showed beta-galactosidase-specific CTL responses. The CTL line derived from mice immunized i.p. was also shown to be class I restricted and Thy-1.2+, CD8+, and TCR alpha beta+. All mice immunized with the recombinant L. monocytogenes had positive delayed-type hypersensitivity responses to heat-killed L. monocytogenes, but only 15% had a positive delayed-type hypersensitivity reaction to beta-galactosidase. Individual serum samples from mice immunized i.p. or i.v. were tested for antibody to beta-galactosidase. Approximately 11% had low positive titers for beta-galactosidase antibodies. These results demonstrate that both oral and parenteral immunization with recombinant L. monocytogenes results in a cellular immune response to the foreign protein, which is primarily a specific CD8+ CTL response.
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PMID:Induction of a cellular immune response to a foreign antigen by a recombinant Listeria monocytogenes vaccine. 160 62

Measles virus (MV)-specific murine helper T cell clones (Thy-1.2+, CD4+, CD8-) were generated from mice immunized with MV-infected mouse brain homogenate by limiting dilution and in vitro stimulation of spleen cells with UV-inactivated MV Ag. The protein specificity of 7 out of 37 stable T cell clones, which displayed MHC-restricted MV Ag recognition, could be assessed by using purified MV proteins. Two fusion (F) protein-specific, two hemagglutinin-specific, and three nucleoprotein- or matrix protein-specific clones were shown to be established. The F protein-specific T cell clones together with a panel of previously generated F protein-specific T cell clones were characterized for their fine specificity by using beta-galactosidase fusion products, which contained different parts of the F protein. It was shown that at least two epitopes on the major part of the F protein (amino acid 2-513) can be recognized by mouse T cells. Functional characterization of three T cell clones showed that they were able to assist MV-specific B cells and bystander B cells for antibody production. Furthermore, they were shown to produce the lymphokines IL-2 and IFN-gamma. It was also shown that these T cell clones induced a MV-specific delayed type hypersensitivity response. These observations suggest that all of the T cell clones characterized belong to the TH1 helper subset.
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PMID:Measles virus-specific murine T cell clones: characterization of fine specificity and function. 252 70

A solid-phase, indirect beta-galactosidase-linked immunoassay (ELISA) is described for screening large numbers of monoclonal antibodies that recognize cell surface antigens of primary monolayer cerebellar cultures. Target cultures were prepared from perikaryal suspensions of postnatal rodent cerebellum seeded into poly-L-lysine pre-coated, flat-bottom microtiter wells and fixed with glutaraldehyde after growth in vitro. Hybridoma supernatants were then incubated on these cultures. After the addition of beta-galactosidase-linked anti-mouse IgG F(ab')2 fragments, antigen-positive supernatants were detected with the enzyme substrate o-nitrophenyl-beta-D-galactopyranoside. Using a monoclonal antibody specific for rat brain Thy-1 glycoprotein, this solid-phase ELISA was found to be useful in quantifying changes in the developmental expression of cerebellar surface antigens in these cultures.
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PMID:A solid-phase beta-galactosidase ELISA for detecting and quantifying monoclonal antibody binding to dissociated cell cultures of postnatal rodent cerebellum. 641 Jan 26

We constructed three recombinant vectors derived from the herpes simplex virus type 1 mutant tsK, each of which contained a different transgene under the control of the herpes simplex virus type 1 immediate early 3 promoter inserted into the thymidine kinase locus: the prokaryotic enzymes beta-galactosidase and chloramphenicol acetyl transferase, and a fusion gene consisting of human tissue inhibitor of metalloproteinases linked to the last exon of Thy-1, which encodes for a glycosyl-phosphatidyl-inositol membrane anchor. Infection of postmitotic neocortical and hippocampal neurons in low-density primary cultures with these vectors, achieved reliable expression of all three foreign gene products in various neocortical cell types, e.g. pyramidal neurons, non-pyramidal neurons, and glial cells. The percentage of neurons expressing transgenes ranged from 1 to 46% depending on the multiplicity of infection (highest assayed = 5); the percentage of glial cells expressing transgenes ranged from 0.5 to 98% (highest multiplicity assayed = 3.4). Expression of transgenes could be detected for up to three days in approximately 20% of neurons infected at a multiplicity of infection of 1. Infection of neurons with tk K-derived recombinant vectors inhibited their protein synthesis by 40-50% at a multiplicity of infection of 10, but no effect was observed at a multiplicity of infection of 1. Infection of glial cells with the same vectors at a multiplicity of infection of 1 inhibited protein synthesis by more than 90%. Analysis of neuronal viability at different times post-infection indicated that more than 98% of neurons expressing transgenes 48 h post-infection were viable. Thus, low-density neuronal cultures can be used to assess the efficiency of herpes simplex virus type 1-derived gene transfer vectors and transgene expression in developing cortical postmitotic cells, before and after they establish polarity. In addition, we show that two cytoplasmic enzymes, beta-galactosidase and chloramphenicol acetyl transferase, are able to diffuse freely in the cytoplasm reaching even growth cones in young neurons, while the chimeric protein tissue inhibitor of metalloproteinases/Thy-1 is correctly targeted to the plasma membrane via a glycosyl-phosphatidylinositol anchor. This model system should be useful for investigation of cellular and molecular aspects of the development and establishment of neuronal polarity, as well as for analysis of signals involved in protein targeting in postmitotic neurons.
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PMID:Use of recombinant vectors derived from herpes simplex virus 1 mutant tsK for short-term expression of transgenes encoding cytoplasmic and membrane anchored proteins in postmitotic polarized cortical neurons and glial cells in vitro. 793 6

A transgenic mouse system has been established to follow the pattern of IL-2 expression at the level of single T cells. This was achieved by introducing a human IL-2 promoter-driven reporter gene (Escherichia coli lacZ) into the germline of mice and monitoring its product, beta-galactosidase (beta-gal), by FACS analysis. Ex vivo experiments confirmed that the regulated expression of the transgene is comparable with that of the endogenous IL-2 gene. Transgene expression is inducible by mitogens, restricted to T cells, and diminished by immunosuppressive agents, such as cyclosporin A, at concentrations known to suppress IL-2 transcription. Depending on the mitogens used, 30-50% of peripheral T cells produced IL-2 with an asynchronous induction pattern, as measured by transgenic beta-gal activity. Both helper (CD4+CD8-) and cytotoxic T cells (CD4-CD8+) respond with comparable heterogenous expression levels but they show different frequencies of beta-gal production. Transgenic beta-gal-producing T cells were detectable as early as 2 h after mitogen stimulation. These cells represent a transitional IL-2 secreting, IL-2 receptor alpha-chain negative T cell population, which occurs in the autocrine process of T cell activation. Administration of staphylococcal enterotoxin A (SEA), a bacterial superantigen, resulted in a T cell specific (Thy-1.2) increase (2.5-fold) of reporter gene expression in vivo. In summary, we could demonstrate that IL-2 promoter-driven reporter gene expression in transgenic mice is a sensitive tool to characterize IL-2 expressing cells phenotypically.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:IL-2 promoter-driven lacZ expression as a monitoring tool for IL-2 expression in primary T cells of transgenic mice. 815 96

In this study, naturally-occurring, monoclonal IgM kappa anti-thymocyte autoantibodies from the neonatal inbred Balb/c mouse-derived hybridoma NMT-1 (NMT-1 mAb), previously reported to identify a restricted CD4+CD8+CD3/lo/int thymocyte subpopulation, have been shown to exhibit extensive polyspecificity. Using immunofluorescence, immunoblotting and antibody titration and competition ELISAs, NMT-1 mAbs exhibited polyspecific binding to 12 apparently structurally unrelated self and non-self antigens. The autoreactive component of the polyspecificity profile of NMT-1 mAbs encompassed reactivity to developmentally-related 14.5 and 18.3 kDa Thy-1 glycoforms expressed on a CD4+CD8+CD3-/lo/int thymocyte subpopulation. The autoreactivity profile of NMT-1 mAbs also included recognition of the heavy and light chains of mouse IgG1 and mouse cytokeratins within thymic medullary epithelium and basal epithelial cells of stratified squamous epithelium of mouse tongue, oesophagus, stomach, skin and vagina. Examination of the polyspecificity profile of NMT-1 mAbs was also undertaken using a panel of 23 antigens including heterologous proteins, phospholipids, haptens and bacterial antigens by antibody titration and competition ELISAs. Antibody titration ELISAs demonstrated that NMT-1 mAbs bound nine antigens including bovine carbonic anhydrase, ovalbumin, cardiolipin, phosphatidylserine, the haptens, DNP and FITC and the bacterial antigens including Escherichia coli beta-galactosidase and the toxoids from Corynebacterium tetani and Clostridium diphtheria. Competition ELISAs, based on the inhibition of NMT-1 mAb binding to antigens adsorbed to ELISA plate surfaces by inhibitor antigens in solution, demonstrated that NMT-1 mAb interactions were not dependent on multivalent binding. In these assays, NMT-1 mAbs recognized unmodified (native) epitopes on the solution phase forms of the protein antigens, including E. coli beta-galactosidase and toxoids from Corynebacterium tetani and Clostridium diphtheria, providing further evidence for the hypothesis that the binding of multiple, apparently unrelated, antigens by NMT-1 mAbs occurs via unique polyspecific antigen combining sites.
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PMID:Naturally-occurring anti-thymocyte autoantibody which identifies a restricted CD4+CD8+CD3-/lo/int thymocyte subpopulation exhibits extensive polyspecificity. 872 53

Stem cells of fetal origin lie between embryonic and adult stem cells in terms of potentiality. Because of the ethical controversy surrounding embryonic stem cells and the relatively inferior quality of adult stem cells, the use of fetal stem cells would be an attractive option in future therapeutic applications. Here, we have investigated primitive characteristics of human umbilical-cord-derived fetal mesenchymal stem cells (UC fMSCs) during extensive expansion. We have successfully isolated and cultured UC fMSCs from all UC samples, but with two early fungal contaminations. UC fMSCs proliferated without significant evidence of morphological changes, and the average cumulative population-doubling level was over 25 for about 3 months. UC fMSCs showed the positive expression of several CD markers, known to be related to MSCs, including CD73 (SH-3, 4), CD90 (Thy-1), CD105 (SH-2), CD117 (c-kit), and CD166 (ALCAM). They demonstrated primitive properties throughout the expansion period: multilineage differentiation potentials examined by functional assays, a variety of pluripotent stem cell markers including Nanog, Oct-4, Sox-2, Rex-1, SSEA-3, SSEA-4, Tra-1-60, and Tra-1-81, minimal evidence of senescence as shown by beta-galactosidase staining, and the consistent expression of telomerase activity. These results suggest that UC fMSCs have more primitive properties than adult MSCs, which might make them a useful source of MSCs for clinical applications.
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PMID:Fetal mesenchymal stem cells derived from human umbilical cord sustain primitive characteristics during extensive expansion. 1894 82