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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)

HSV-1 amplicon vectors were used to express either a cytoplasmic (beta-galactosidase) or a membrane targeted protein (TIMP-Thy1) in primary neuronal cultures, and a human astrocytoma cell line. Whereas some cells became infected by vector particles alone others were simultaneously infected by both vector and helper particles. Our results show that IEHCMV and HSV-1 IE3 promoters are able to direct transgene expression in these cells in the absence of synthesis of helper virus transacting proteins, and stress the need of monitoring expression from both partners of an amplicon population, in order to differentiate transgene expression in cells singly infected with amplicon particles, from those infected by both amplicon and helper particles.
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PMID:Simultaneous detection of amplicon and HSV-1 helper encoded proteins reveals that neurons and astrocytoma cells do express amplicon-borne transgenes in the absence of synthesis of virus immediate early proteins. 760 39

To identify a good system to introduce foreign genes into normal and tumoral astrocytes, we studied the efficiency of two chemical methods, calcium phosphate precipitation and lipofection, and of a viral-mediated transfer by a vector derived from the highly attenuated modified vaccinia virus Ankara (MVA). Using the beta-galactosidase (beta-gal) gene (lacZ) as reporter, we searched for optimal experimental conditions to obtain an efficient gene transfer into human embryonic and neonatal rat astrocytes and into a human astrocytoma cell line (U373 MG). The beta-gal protein production was evaluated by cytochemical staining and enzymatic activity assay. Among chemical methods, lipofection was the most efficient system to transfect astrocytes in providing up to 60% of beta-gal-positive cells in all the cell types analyzed. MVA infection also proved to be an efficient system to introduce heterologous genes into human embryonic astrocytes that appeared 80-100% positive 48-96 hr after an infection at a multiplicity of 1-10. In contrast, only a limited infection was observed with rat astrocytes, human astrocytoma cells, and human leptomeningeal cells. A recombinant MVA vector expressing the human immunodeficiency virus-1 (HIV-1) regulatory protein Nef was used to transfect human embryonic astrocytes, and the resulting Nef expression was compared with that detected after lipofection in the same cells. By Western blot analysis, Nef expression was observed in human astrocytes 24-96 hr after infection and was similar to that present in stably HIV-1-infected astrocytoma cells. Lipofection resulted in lower Nef expression. In spite of these promising results, the negative effects of MVA infection on cell viability and the possibility that a productive infection occurs in human embryonic astrocytes limit the use of this vector for gene delivery in developmentally immature human glial cells.
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PMID:Gene transfer in astrocytes: comparison between different delivering methods and expression of the HIV-1 protein Nef. 1008 79

Astrocytic tumors frequently exhibit defects in the expression or activity of proteins that control cell-cycle progression. Inhibition of kinase activity associated with cyclin/cyclin-dependent kinase co-complexes by cyclin-dependent kinase inhibitors is an important mechanism by which the effects of growth signals are down-regulated. We undertook the present study to determine the role of p57(KIP2) (p57) in human astrocytomas. We demonstrate here that whereas p57 is expressed in fetal brain tissue, specimens of astrocytomas of varying grade and permanent astrocytoma cell lines do not express p57, and do not contain mutations of the p57 gene by multiplex-heteroduplex analysis. However, the inducible expression of p57 in three well-characterized human astrocytoma cell lines (U343 MG-A, U87 MG, and U373 MG) using the tetracycline repressor system leads to a potent proliferative block in G(1) as determined by growth curve and flow cytometric analyses. After the induction of p57, retinoblastoma protein, p107, and E2F-1 levels diminish, and retinoblastoma protein is shifted to a hypophosphorylated form. Morphologically, p57-induced astrocytoma cells became large and flat with an expanded cytoplasm. The inducible expression of p57 leads to the accumulation of senescence-associated beta-galactosidase marker within all astrocytoma cell lines such that approximately 75% of cells were positive at 1 week after induction. Induction of p57 in U373 astrocytoma cells generated a small population of cells ( approximately 15%) that were nonviable, contained discrete nuclear fragments on Hoechst 33258 staining, and demonstrated ultrastructural features characteristic of apoptosis. Examination of bax and poly-(ADP ribose) polymerase levels showed no change in bax, but decreased expression of poly-(ADP ribose) polymerase after p57 induction in all astrocytoma cell lines. These data demonstrate that the proliferative block imposed by p57 on human astrocytoma cells results in changes in the expression of a number of cell cycle regulatory factors, cell morphology, and a strong stimulus to cell senescence.
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PMID:Expression of p57(KIP2) potently blocks the growth of human astrocytomas and induces cell senescence. 1098 Jan 31