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)

Peptides of melanosomal proteins have recently been shown to be recognized in an HLA-restricted mode by specific cytolytic T lymphocytes in melanoma patients. Dendritic antigen-presenting cells (DC) are considered to be the most effective stimulators of T cell responses, and the use of these cells has therefore been proposed to generate therapeutic responses to tumor antigens in cancer patients. We, therefore, generated DC from peripheral blood of normal donors in the presence of granulocyte/macrophage colony-stimulating factor and interleukin-4. Flow cytometric analysis of the cells during a 2-week culture revealed a loss of CD14 and CD34 expression, a concomittent increase of CD1a, CD11a,b and c, CD44, CD45, CD54, HLA-class I and II, and intermediate levels of CD26, CD80 and CD86. Cultured DC stimulated proliferation of allogeneic T cells and induced a marked, up to 20-fold, stimulation of T cell proliferation after pulsing with tetanus toxoid. To achieve independence of already-identified antigenic peptides presented in HLA class I-restricted fashion, which limits the general applicability of such peptides for vaccination of melanoma patients, we tested whether DC are transfectable with eukaryotic expression plasmids. DC transfected with two reporter genes (CAT, beta-galactosidase) using a liposome-based transfection technique, exhibited only low levels of enzymatically active proteins, but were able to degrade rapidly intracellular proteins and to process peptides efficiently. Chloramphenicol acetyltransferase as well as tyrosinase mRNA were detectable after transfection by reverse-transcriptase-polymerase chain reaction, and enzyme activities became measurable. Furthermore, DC transfected with the tyrosinase gene were able to induce specific T cell activation in vitro, indicating appropriate peptide processing and presentation in DC after transfection. These data suggest new approaches to future tumor vaccination strategies.
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PMID:Dendritic cells generated from peripheral blood transfected with human tyrosinase induce specific T cell activation. 748 49

Three major inhibitors of the NF-kappaB/Rel family of transcription factors, IkappaBalpha, IkappaBbeta, and IkappaBepsilon, have been described. To examine the in vivo role of the most recently discovered member of the IkappaB family, IkappaBepsilon, we generated a null allele of the murine IkappaBepsilon gene by replacement of all coding sequences with nlslacZ. Unlike IkappaBalpha nullizygous mice, mice lacking IkappaBepsilon are viable, fertile, and indistinguishable from wild-type animals in appearance and histology. Analysis of beta-galactosidase expression pattern revealed that IkappaBepsilon is mainly expressed in T cells in the thymus, spleen, and lymph nodes. Flow cytometric analysis of immune cell populations from the bone marrow, thymus, spleen, and lymph nodes did not show any specific differences between the wild-type and the mutant mice, with the exception of a reproducible 50% reduction of the CD44-CD25+ T cell subspecies. The IkappaBepsilon-null mice present constitutive up-regulation of IgM and IgG1 Ig isotypes together with a further increased synthesis of these two isotypes after immunization against T cell-dependent or independent Ags. The failure of observable augmentation of constitutive nuclear NF-kappaB/Rel-binding activity is probably due to compensatory mechanisms involving IkappaBalpha and IkappaBbeta, which are up-regulated in several organs. RNase-mapping analysis indicated that IL-1alpha, IL-1beta, IL-1Ra, and IL-6 mRNA levels are constitutively elevated in thioglycolate-elicited IkappaBepsilon-null macrophages in contrast to GM-CSF, G-CSF, and IFN-gamma, which remain undetectable.
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PMID:IkappaBepsilon-deficient mice: reduction of one T cell precursor subspecies and enhanced Ig isotype switching and cytokine synthesis. 1057 Feb 87

To analyze the NF-kappaB/Rel activity pattern in a living organism, we previously generated transgenic mice carrying a kappaB-dependent lacZ gene. In situ analysis of both primary and secondary lymphoid organs revealed a strong NF-kappaB transcriptional activity in antigen-presenting cells, some endothelial cells and sinus lining cells of the lymph node capsula with very little activity in lymphocytes and thymocytes. Using fluorescein-di-beta-D-galactopyranoside (FDG) as a vital substrate for the beta-galactosidase, we re-examined by flow cytometry the NF-kappaB/Rel transcriptional activity in our mouse model. We report here that such constitutive NF-kappaB/Rel activity was significantly detected in thymocytes at the CD44+CD25(-) stage. This constitutive activity extended with CD25 expression to the majority of the CD44(-)CD25(+) thymocytes and was then restricted to a few mature T cells. In the spleen, constitutive NF-kappaB/Rel activity was found in most B cells, unlike T cells which were largely negative. Virgin IgD(+) B cells expressed higher levels of NF-kappaB transcriptional activity than other B cell types. Altogether, these results suggest that NF-kappaB/Rel complexes are key players in the in vivo differentiation of IgD(+) B lymphocytes and possibly CD25(+) thymocytes.
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PMID:In vivo identification of lymphocyte subsets exhibiting transcriptionally active NF-kappaB/Rel complexes. 1078 7

Activated T cells recognize Ag in the retina, an immune privileged tissue, and may mediate autoimmune disease. In contrast, this report asks if resting, Ag-specific CD4(+) CD44(+) T cells can recognize Ag expressed in the retina. As a probe for Ag, 3E9 T cells specific for an immunodominant epitope of beta-galactosidase (beta-gal) were transferred to transgenic (Tg) mice expressing beta-gal in retinal photoreceptor cells, or to ROSA26 mice which express beta-gal widely. The survival, phenotype, and responsiveness of transferred 3E9 T cells were unaffected by the presence of retinal beta-gal, but altered by recognition of beta-gal in the ROSA26 mice. Inoculation or induction of activated T cells with specificity for this epitope produced autoimmune uveoretinitis, showing that the retinal beta-gal is expressed at immunologically significant levels. We conclude that sequestration provides a substantial barrier to recognition of Ag in quiet retina, and that insufficient Ag leaves the retina for detectable immune recognition outside of the retina.
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PMID:Failure of memory (CD44 high) CD4 T cells to recognize their target antigen in retina. 1169 17

Although several observations show local T cell recognition of retinal Ag, there has been no direct demonstration that the APC were retinal derived, rather than recruited. In this study, CD45(+) cells isolated from immunologically quiescent murine retina were tested in vitro for functional evidence of Ag presentation to naive and Ag-experienced CD4 T cells specific for beta-galactosidase. Because CD45(+) cells from brain have been reported to be efficient APC, they were included for comparison. Measures of activation included changes in CD4, CD25, CD44, CD45RB, CD62L, CD69, caspase-3 activation, CFSE dilution, size, number of cells recovered, and cytokine production. Retinal CD45(+) cells gave no evidence of Ag-dependent TCR ligation in naive T cells, unlike splenic APC and CD45(+) cells from brain, which supported potent responses. Instead, addition of retinal CD45(+) cells to cocultures of naive 3E9 T cells plus splenic APC reduced the yield of activated T cells and cytokine production by limiting T cell activation at early time points. Ag-experienced T cells responded weakly to Ag presented by retinal CD45(+) cells. Activating the retinal cells with IFN-gamma, anti-CD40, or LPS incrementally increased their APC activity. Addition of neutralizing Abs to TGF-beta did not reveal suppressed retinal APC activity. Because retina lacks tissue equivalents of meninges and choroid plexus, rich sources of dendritic cells in brain, cells from retina may better represent the APC activity of fresh, adult CNS parenchymal and perivascular cells. The activity of the retinal CD45(+) cells appears to be directed to limiting T cell responses.
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PMID:The antigen-presenting activity of fresh, adult parenchymal microglia and perivascular cells from retina. 1515 73

Recently, it was shown that MYCN amplified cells spontaneously expulse extrachromosomally amplified gene copies by micronuclei formation. Furthermore, it was shown that these cells lose their malignant phenotype and start to age. We tested whether it is possible to encourage neuroblastoma tumor cells to enter the senescence pathway by low concentrations of the micronuclei-inducing drug hydroxyurea (HU). We studied the effect of HU on 12 neuroblastoma cell lines with extra- or intrachromosomally amplified MYCN copies and without amplification. Two extrachromosomally amplified neuroblastoma cell lines (with double minutes) were investigated in detail. Already after 3 weeks of HU treatment, the BrdU uptake dropped to 25% of the starting cells. After 4 weeks, enlarged and flattened cells (F-cells) and increased granularity in the majority of cells were observed. A drastic reduction of the MYCN copy number-down to one copy per cell-associated with CD44 and MHCI upregulation in up to 100% of the HU treated neuroblastoma cells was found after 5-8 weeks. Telomere length was reduced to half the length within 8 weeks of HU treatment, and telomerase activity was not detectable at this time, while being strongly expressed at the beginning. All these features and the expression of senescence-associated-beta-galactosidase (SA-beta-GAL) in up to 100% of the cells support the hypothesis that these cells entered the senescence pathway. Thus, low-dose HU is a potent senescence elicitor for tumor cells with gene amplification, possibly representing an attractive additional strategy for treatment of this subset of tumors.
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PMID:Induction of senescence in MYCN amplified neuroblastoma cell lines by hydroxyurea. 1710 70

Primary cultures of human mammary epithelial cells underwent significant morphological and functional changes during the aging process between passage 12 (P12) and passage 16 (P16). Concomitant with a progressive and significant expression of senescence-associated beta-galactosidase as aging marker, the cells restructured their attachment, increased in size and ceased to divide. Young HMEC until P11 demonstrated a nearly 100% expression of distinct adhesion molecules such as CD24, integrin beta1 (CD29) and CD44 similar to the human mammary tumor cell line MCF-7. In parallel with the aging-associated alterations of the cell adhesion, expression of CD24 and CD44 dropped in senescent P16 HMECs. However, levels of CD29 remained unchanged during the aging process. The tumor-associated Muc-1 (CD227), which was expressed to about 100% in the tumorigenic MCF-7 cells, was detectable in 51% of young HMEC in P11 and declined to 37% in aged HMEC in P16. In association with the remodeling of cell shape, expression levels of distinct matrix metalloproteinases including MMP-7 markedly decreased in aging HMEC. In contrast, MMP-1, MMP-2 and MMP-9 remained unchanged indicating a possible functional role of MMP-7 during the HMEC aging process. Indeed, down-modulation of MMP-7 by RNAi revealed a significantly elevated G(2)/M cell cycle arrest and a 2- to 3-fold enhanced senescence-associated beta-galactosidase expression as compared to control siRNA transfectants and control HMEC, respectively. Together, these findings suggested that decreasing MMP-7 expression contributes to accelerated aging of human mammary epithelial cells.
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PMID:MMP-7 is involved in the aging of primary human mammary epithelial cells (HMEC). 1820 46

Life cycle limitation hampers the production of high amounts of primary human mesenchymal stroma-/stem-like cells (MSC) and limits cell source reproducibility for clinical applications. The characterization of permanently growing MSC544 revealed some differentiation capacity and the simultaneous presence of known MSC markers CD73, CD90, and CD105 even after continuous long-term culture for more than one year and 32 passages. The expression of CD13, CD29, CD44, and CD166 were identified as further surface proteins, all of which were also simultaneously detectable in various other types of primary MSC populations derived from the umbilical cord, bone marrow, and placenta suggesting MSC-like properties in the cell line. Proliferating steady state MSC544 exhibited immune-modulatory activity similar to a subpopulation of long-term growth-inhibited MSC544 after 189d of continuous culture in confluency. This confluent connective cell layer with fibroblast-like morphology can spontaneously contract and the generated space is subsequently occupied by new cells with regained proliferative capacity. Accordingly, the confluent and senescence-associated beta-galactosidase-positive MSC544 culture with about 95% G0/G1 growth-arrest resumed re-entry into the proliferative cell cycle within 3d after sub-confluent culture. The MSC544 cells remained viable during confluency and throughout this transition which was accompanied by marked changes in the release of proteins. Thus, expression of proliferation-associated genes was down-modulated in confluent MSC544 and re-expressed following sub-confluent conditions whilst telomerase (hTERT) transcripts remained detectable at similar levels in both, confluent growth-arrested and proliferating MSC544. Together with the capability of connective cell layer formation for potential therapeutic approaches, MSC544 provide a long term reproducible human cell source with constant properties.
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PMID:Reversible Growth-Arrest of a Spontaneously-Derived Human MSC-Like Cell Line. 3263 95