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

Arylsulfatase A (ASA) and cerebroside-beta-galactosidase activities in leukocytes serve as a diagnostic tool for determining the presence of metachromatic leukodystrophy and globoid cell leukodystrophy, respectively. It has not been demonstrated whether a delay in blood processing and the presence of mixed cell types in different proportions in leukocytes affect the activities of the two enzymes in these cells. We have in the present study determined the specific activity in leukocytes and lymphocytes (T-cells) prepared from blood samples processed immediately after, 4, and 24 h after collection. In order to determine whether the enzyme activities in lymphocytes reflect expression of genetic trait, and not environmental or "state" influence, the activities of the two enzymes in interleukin 2-stimulated T-cells and resting T-cells were compared. A delay of up to 24 h in blood processing did not significantly change the specific activities of the two enzymes in both leukocytes and lymphocytes. The specific activity of ASA and beta-galactosidase in lymphocytes was 1.4-1.8 times that in leukocytes. The activities of the two enzymes in interleukin 2-stimulated T-cells did not differ from those in resting T-cells. These results indicate that blood-processing delay had no significant effects on ASA and beta-galactosidase activity. The data further indicate that the ASA and beta-galactosidase activity in interleukin 2-stimulated T-cells was not significantly different from resting lymphocytes from either normal or psychiatric subjects exposed to various medications. The activity levels in lymphocytes from psychiatric subjects thus reflect expression of genetic trait, rather than environmental or state influence.
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PMID:Arylsulfatase A and beta-galactosidase activities in leukocytes and lymphocytes from normal and psychiatric subjects. Effects of blood-processing delay and interleukin-2 stimulation. 775 46

To better understand the events occurring during immunotherapy of liver metastases with effector cells, we have developed a clinically relevant animal model in which both effector-tumor cell interactions and survival can be evaluated. A cell line of human gastric carcinoma (HR) metastatic to the liver has been established from a patient's liver biopsy. HR cells (10 x 10(6)) injected intrasplenically metastasize into the liver of immunosuppressed nude mice, with micrometastases detectable histologically by day 4 and macrometastases by day 7. The animals subsequently develop ascites and die between days 30 and 40 after tumor injection. To investigate early metastatic events in the liver, HR cells were transduced with a plasmid containing both the lacZ gene under the control of the CMV promoter and NeoR gene. Transfectants selected for neomycin resistance were lacZ gene positive and stained blue in the presence of a beta-galactosidase substrate, 5-bromo-4-chloro-3-indolyl-beta-D-galactopyranoside (X-Gal). These transfectants (HRLZ) remained lacZ gene positive for at least 25 passages in vitro. Injected intrasplenically, an HRLZ clone grew invasively in nude mice and formed liver metastases comparably to parental tumor cells. The number and localization of blue X-Gal-positive tumor cells were followed in liver tissues of animals sacrificed at various times, from 1 h to 28 days postinjection of HRLZ cells. HRLZ cells were seen in liver blood vessels and sinusoids within 1 h after injection, and the progressive growth of micrometastases and macrometastases could be followed with precision by X-Gal staining. On day 3 after injection of HRLZ cells, numerous micrometastases were established containing 12-16 tumor cells. When these 3-day established HRLZ micrometastases were treated by the intrasplenic infusion of interleukin 2 (IL2)-activated human natural killer (NK) cells selected by IL2-induced adherence to plastic (A-NK) and systemic IL2, nearly all liver micrometastases were eliminated within 24 h after a single transfer of A-NK cells (P < 0.001). This xenogeneic model was also used for adoptive immunotherapy of 7-day established liver macrometastases with human A-NK cells injected intrasplenically and exogenous IL2 given i.p. A significant decrease in the number of hepatic metastases and the weight of livers (P < 0.003) in comparison with those of control mice was observed.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Immunotherapy of liver metastases of human gastric carcinoma with interleukin 2-activated natural killer cells. 803

In T lymphocytes, intracellular Ca2+ concentration ([Ca2+]i) rises within seconds of T-cell antigen-receptor stimulation and initiates the synthesis and secretion of interleukin 2, a cytokine essential for T-cell proliferation and the immune response. Using video-imaging techniques, we tracked [Ca2+]i signals in individual T cells and measured subsequent expression of a beta-galactosidase reporter gene (lacZ) controlled by the NF-AT element of the interleukin 2 enhancer. [Ca2+]i spikes elicited by monoclonal antibody binding to the CD3 epsilon subunit of the T-cell receptor were positively correlated with gene expression, but varied widely between individual cells and were therefore difficult to relate quantitatively to lacZ expression. The [Ca2+]i dependence of NF-AT-regulated gene expression was determined by elevating [Ca2+]i with either thapsigargin or ionomycin and then "clamping" [Ca2+]i to various, stable levels by altering either extracellular [Ca2+] or extracellular [K+]. Raising [Ca2+]i from resting levels of 70 nM to between 200 nM and 1.6 microM increased the fraction of cells expressing lacZ, with Kd approximately 1 microM. Activation of protein kinase C enhanced the [Ca2+]i sensitivity of gene expression (Kd = 210 nM), whereas stimulation of protein kinase A inhibited [Ca2+]i-dependent gene expression. The experiments described here provide single-cell measurements linking a second messenger to gene expression in individual cells.
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PMID:Intracellular calcium dependence of gene expression in single T lymphocytes. 814 3

Like interleukin 2 (IL-2), interferon gamma (IFN-gamma) is an early response gene in T cells and both are prototypical T helper cell type 1 (Th-1) lymphokines. Yet IL-2 and IFN-gamma production are independently regulated, as demonstrated by their differential expression in certain T cell subsets, suggesting that the regulatory elements in these two genes must differ. To explore this possibility, the 5' flank of the human IFN-gamma gene was analyzed. Expression of IFN-gamma promoter-driven beta-galactosidase reporter constructs containing 538 bp of 5' flank was similar to that by constructs driven by the IL-2 promoter in activated Jurkat T cells; expression nearly as great was observed with the construct containing only 108 bp of IFN-gamma 5' flank. These IFN-gamma promoter constructs faithfully mirrored expression of the endogenous gene, in that expression required activation both with ionomycin and PMA, was inhibited by cyclosporin A, and was not observed in U937 or THP-1 cells. The region between -108 and -40 bp in the IFN-gamma promoter was required for promoter function and contained two elements that are conserved across species. Deletion of 10 bp within either element reduced promoter function by 70%, whereas deletions in nonconserved portions of this region had little effect on promoter function. The distal conserved element (-96 to -80 bp) contained a consensus GATA motif and a potential regulatory motif found in the promoter regions of the GM-CSF and macrophage inflammatory protein (MIP) genes. Factors binding to this element, including GATA-3, were found in Jurkat nuclear extracts by electromobility shift assays and two of the three complexes observed were altered in response to activation. One or both of these motifs are present in the 5' flank of multiple, other lymphokine genes, including IL-3, IL-4, IL-5, and GM-CSF, but neither is present in the promoter of the IL-2 gene. The proximal conserved element (-73 to -48 bp) shares homology with the NFIL-2A element in the IL-2 promoter; these elements compete for binding of factors in Jurkat nuclear extracts, although the NFIL-2A element but not the IFN-gamma element binds Oct-1. Factors binding to this element in the IFN-gamma gene were present in extracts from resting and activated Jurkat T cells. However, by in vivo footprinting of intact cells, this element was protected from methylation only with activation.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Two essential regulatory elements in the human interferon gamma promoter confer activation specific expression in T cells. 822 2

The mechanism of action of the immunosuppressive drug cyclosporin A (CsA) is the inactivation of the Ca2+/calmodulin-dependent serine-threonine phosphatase calcineurin by the drug-immunophilin complex. Inactive calcineurin is unable to activate the nuclear factor of activated T cells (NFAT), a transcription factor required for expression of the interleukin 2 (IL-2) gene. IL-2 production by CsA-treated cells is therefore dramatically reduced. We demonstrate here, however, that NFAT can be activated, and significant levels of IL-2 can be produced by the CsA-resistant CD28-signaling pathway. In transient transfection assays, both multicopy NFAT- and IL-2 promoter-beta-galactosidase reporter gene constructs could be activated by phorbol 12-myristate 13-acetate (PMA)/alpha-CD28 stimulation, and this activation was resistant to CsA. Electrophoretic mobility shift assay showed the induction of a CsA-resistant NFAT complex in the nuclear extracts of peripheral blood T cells stimulated with PMA plus alphaCD28. Peripheral blood T cells stimulated with PMA/alphaCD28 produced IL-2 in the presence of CsA. Collectively, these data suggest that NFAT can be activated and IL-2 can be produced in a calcineurin independent manner.
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PMID:Activation of nuclear factor of activated T cells in a cyclosporin A-resistant pathway. 863 9

Myoblasts were grown from monkey muscle biopsies and infected in vitro with a defective retroviral vector containing a cytoplasmic beta-galactosidase (beta-gal) gene. These myoblasts were then transplanted to 14 different monkeys, 6 of which were immunosuppressed with FK506. Without immunosuppression, only a few myoblasts and myotubes expressing beta-gal were observed 1 week after the transplantation, but no cells expressing beta-gal were observed after 4 weeks. This result was attributed to immune responses since infiltration by CD4+ or CD8+ lymphocytes was abundant 1 week after transplantation but not after 4 weeks. The expression of interleukin 6 (IL-6), interleukin 2 (IL-2), granulocyte/macrophage colony stimulating factor (GM-CSF), transforming growth factor-beta (TGF-beta) and granzyme B mRNAs was increased in the myoblast-injected muscle indicating that the infiltrating lymphocytes were activated. Moreover, antibodies against the donor myoblasts were detected in 3 out of 6 cases. When the monkeys were immunosuppressed with FK506, muscle fibers expressing beta-galactosidase (beta-gal) were present 1, 4 and 12 weeks after the transplantation. There was neither significant infiltration by CD4 or CD8 lymphocytes, nor antibodies detected. The mRNA expression of most cytokines was significantly reduced as compared to the nonimmunosuppressed monkeys. These results indicate that FK506 is effective in controlling short-term immune reactions following myoblast transplantation in monkeys and suggest that it may prove useful for myoblast transplantation in Duchenne Muscular Dystrophy patients.
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PMID:Myoblast transplantation in monkeys: control of immune response by FK506. 864 94

Cloned high-metastatic Lewis lung carcinoma. A11 cells were retrovirally transduced with either granulocyte macrophage-colony stimulating factor (GM-CSF) or beta-galactosidase gene and examined for their tumorigenicity. GM-CSF-engineered A11 cells produced a much higher amount of GM-CSF than the parental and control cells. Unexpectedly, GM-CSF-engineered A11 cells grew more rapidly than the control cells, while in vitro growth rates of these cells were almost the same. The enhanced tumor growth seemed to be unique to GM-CSF among various cytokines, because interleukin 2 (IL-2), interleukin 4 (IL-4) and interleukin 6 (IL-6) producer cells exhibited suppressed tumor growth.
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PMID:Augmentation of in vivo growth of Lewis lung carcinoma cells transduced with granulocyte macrophage-colony stimulating factor gene. 868 29

One of the most widely used methods for the study of T-helper (T(h)) cell activity is the interleukin 2 (IL-2) assay. Typically, this assay is a two-step process involving (a) the activation of T(h) cells in vitro and (b) the testing of IL-2-dependent indicator cells for growth in the presence of T(h) cell supernatants. The assay has served to quantify and characterize T(h) responses to a variety of unique pathogens and immunization regimens. A one-step, single-cell assay is also available for the testing of mouse T(h) cell hybridomas. In this assay, cells fused with the BWZ.36 parent line are scored for positive responses based on their expression of a beta-galactosidase gene linked to an IL-2 enhancer element. The experiments described in this report were designed to examine the conventional and single-cell assays in a side-by-side comparison. Results revealed a striking difference between the two assays. The single-cell assay proved to be extremely sensitive and identified T(h) responses that were altogether missed by the conventional test. Based on these results, we suggest that (i) the conventional and single-cell assays should not be used interchangeably, and (ii) negative results in the conventional IL-2 assay should be considered preliminary until the more sensitive, single-cell assay can be performed.
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PMID:A highly sensitive single-cell assay detects T-helper cell responses missed by conventional interleukin-2-based methods. 1179 96

Tissue-specific gene transfer remains one of the main challenges to deliver genes into designated and/or disseminated cells. We have previously shown successful gene transfer with a nonviral gene delivery system based on the simple chemical conjugation of plasmid DNA with antibody. However, this approach was hampered by low efficiency due to the poor translocation rate of DNA to the nucleus. To improve this approach, we have modified our vector by introducing noncovalent binding between the antibody and DNA, allowing the possibility to introduce different important molecules. The noncovalent association was achieved with neutravidin and biotinylated components: (1) biotinylated antibodies; (2) a biotinylated hemagglutinin fusogenic peptide of influenza virus to favor endosomal escape; and (3) biotinylated histone H1 to compact, protect, and associate DNA to the complex. We report here that this delivery system can be internalized by tumor cells targeted by a specific monoclonal antibody, permits the protection of the transfected DNA, and allows its subsequent transfer into the nucleus after escape from the endosomal compartment. We also demonstrate that, in vitro, gene transfer with this vector showed much higher reporter activity in cells (15 vs. 0.5%) and a stronger production of murine interleukin 2 as compared with our previous vector. In vivo, a single intravenous injection of the vector containing an antibody directed to the G250 renal cell carcinoma-associated antigen led to beta-galactosidase expression in engrafted tumor bearing G250 but not in G250-negative tumor or in other tissues. Altogether, these results indicate that our antibody-based vector is suitable to promote gene delivery in vitro and in vivo in tumor cells.
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PMID:In vivo-targeted gene delivery using antibody-based nonviral vector. 1206 43

We conducted two phase 1 trials of direct intratumoral injection of a recombinant E1E3-deleted adenovirus (AdR) encoding either the bacterial enzyme beta-galactosidase (Ad.RSVbetagal) or interleukin 2 (IL2, AdTG5327) into primary nonsmall-cell lung cancers of 21 patients. We report here virus shedding and the duration of virus expression in the tumor after intrabronchial injection of 10(7), 10(8) or 10(9) PFU of adenovirus. The infectious AdR and the viral DNA were detected in PBL, plasma, stool and aerodigestive samples in a dose-dependent manner, since cell cultures and PCRs were found to be positive mainly for samples from patients who received the highest AdR dose (10(9) PFU). We detected beta-galactosidase activity in the tumor biopsy samples of 66% of the patients, seemingly dose related, and only low levels of IL2 mRNA could be detected in tumor biopsy samples. E1 sequences were not detected by PCR in any of the PBL and bronchial samples collected after virus delivery, except in one patient. In this patient, E1 sequences were detected in PBL as well as in tumor biopsy samples collected at days 8, 30 and 60 and were correlated with longer beta-galactosidase expression in tumor samples. PBL tested before and after virus delivery contained both E1 sequences indicating that they did not result from replication-competent adenovirus (RCA) E1 sequences present in the inoculum. In addition, only on the day of the injection was Ad.RSVbetagal also detected in E1-positive PBL, indicating that virus replication in blood was very unlikely.
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PMID:Recombinant adenovirus shedding after intratumoral gene transfer in lung cancer patients. 1260 93


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