Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: DrugBank:EXPT01586 (G418)
 2,237 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Transformation of secondary Sprague-Dawley rat embryo (RE) cells with type 5 adenovirus (Ad5) results in morphologically transformed cells which can undergo a series of sequential changes resulting in enhanced expression of the transformed phenotype, a process termed progression. Selection for a progressed phenotype often occurs after growth in agar or tumor formation in nude mice, and this process is reversible following treatment of cells with 5-azacytidine. In the present study we have analyzed a series of clonal populations of Ad5-transformed RE cells representing different stages in a defined progression lineage. Progression was not associated with alterations in the steady-state levels of mRNA produced by the viral transforming genes, E1A and E1B, or the cellular gene, c-myc. In addition, the tumor-promoting agent 12-O-tetradecanoyl-phorbol-13-acetate (TPA), which induces expression of a progressed phenotype in Ad5-transformed RE cells, did not significantly alter the RNA transcription rates of the Ad5 E1A or E1B genes, the TPA-inducible gene TPA-S1 or the TPA-responsive genes Pro1 or protein kinase C. TPA did, however, increase by 1 h the steady-state level of c-fos mRNA, but this effect was similar in both progressed and unprogressed cells. Progression also did not involve a change in the RNA transcription rate of a number of cellular and viral genes, including actin, c-Ha-ras, c-myc, v-fos, erbB, TGF-alpha, TGF-beta, Pro-2, transin, TPA-R1, v-myb and c-mos, or other adenovirus genes in addition to E1A and E1B, including E2A and E4. Immunoblotting analysis using E1B polyclonal antiserum further indicated that progression was not associated with changes in the levels of an Mr 21,000 polypeptide encoded by E1B. Similarly, immunoprecipitation analysis with an Ad2 E1A monoclonal antibody indicated similar levels of the Mr 55,000 and 48,000 E1A polypeptides, as well as coprecipitated proteins of Mr 300,000, 107,000 and 105,000 [which is the retinoblastoma (Rb) protein], in E11 and E11-NMT cells. Immunoprecipitation of cell lysates with a monoclonal antibody specific for the Mr 105,000 Rb protein further demonstrated that progression also was not associated with a change in the level or state of phosphorylation of the Rb protein. However, transfection of a human Rb gene (also containing a neomycin resistance gene) into Ad5-transformed RE cells was more inhibitory, with respect to formation of G418-resistant colonies, in unprogressed than in progressed Ad5-transformed RE cells.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Analysis of viral and cellular gene expression during progression and suppression of the transformed phenotype in type 5 adenovirus-transformed rat embryo cells. 192 6

A TGF-beta 1 gene expression plasmid was constructed by inserting the porcine 1.7 Kb TGF-beta 1 cDNA into BamHI site of retrovirus vector Dol. The plasmid DNA was introduced into mouse embryonic stem cells (ES-5 line) by calcium phosphate mediated transfection, and transfected ES-5 cells were then selected by stepwise increase in G418 concentration. Finally, we obtained 21 clones that could be stably grown in culture medium with G418 at 500 micrograms/ml and were designated as ES-T cells. Dot blot and Northern analysis of total RNA and polyA+ RNA extracted from those ES-T cells were shown in FIg. 2 and 3, demonstrating that 6 clones could express exogenous porcine TGF-beta 1 mRNA. The stronger hybridized signal in two clones (ES-T6 and ES-T 16) of them were further proved by southern hybridization of genomic DNA from these ES-T cells with 1.7 Kb TGF-beta 1 cDNA probe (Fig. 4). The product of TGF-beta 1 gene overexpression in ES-T 6 cells was shown in Fig. 5 and 6 by SE-LISA for TGF-beta 1 immunoreactivity to TGF-beta 1 antibodies and biological assay for CCL/64 cell growth inhibition, respectively. With respect to some biological characteristics, ES-T 6 cells, like their parent ES-5 cells, retained their pluripotent properties and positive SSEA-1 antigen (Plate I, Fig. 1). ES-T6 cells were expanded and used for studies of in vitro differentiation. Both of ES-T 6 cells and control ES-5 cells could form a lot of simple aggregates and differentiate into embryoid bodies by hanging drop culture for 3 days in the presence of retinoic acid (RA) at 10(-9) mol/L. Then individual embryoid bodies were plated on gelatinized tissue culture wells. On the third day of further culture without RA, a large amounts of epithelial-like and round cells occurred around the embryoid bodies formed either from ES-T 6 cells or ES-5 cells (Plate I, Fig. 4). However, with further culture of embryoid bodies, only the cells differentiated from ES-T6 embryoid bodies could arrange themselves and differentiate into a lot of radially arranged tubular structures (Plate II, Fig. 5). The frequency of tubular structures present in ES-T6 embryoid bodies were about 95.5%, but in ES-5 group there was only about 17.8% cases giving less defined tubular structures (Plate III, Fig. 8).(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:[Expression of exogenous porcine transforming growth factor beta-1 gene in ES cells and its effect on their differentiation in vitro]. 757 52

We have recently shown that TGF-beta-treated normal fibroblasts are able to induce apoptosis of transformed fibroblasts, leading to their elimination. Here we describe a test system that allows the quantitative analysis of the elimination of G418-resistant transformed cells by TGF-beta-treated normal cells. This assay system was used to screen for substances that interfere with the elimination of transformed cells. Catechol and hydroquinone, but not resorcinol, were found to represent potent antagonists of TGF-beta-induced elimination of transformed cells by normal cells. Protection of transformed cells from negative effects derived from their cellular environment defines a hitherto unrecognized crucial mechanism for the survival of transformed cells. The protective effect of catechol as seen in this experimental system may act in concert with its co-carcinogenic and promoting activities during carcinogenesis.
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PMID:Catechol interferes with TGF-beta-induced elimination of transformed cells by normal cells: implications for the survival of transformed cells during carcinogenesis. 782 67

We investigated whether transduction of human cord blood progenitor cells can be increased by spinoculation in fibronectin fragment CH-296 (FN)-coated tubes. Bicistronic vectors PA317/LgEIN, containing the enhanced green fluorescent protein (EGFP) and neomycin phosphotransferase (neo) genes, and PG13/LgDIN, containing the dihydrofolate reductase and neo genes, were used to transduce CD34-enriched human cord blood cells. Transduction by spinoculation in FN-coated tubes (spin/FN+) was compared with spinoculation in noncoated tubes (spin/FN-) and transduction in plates coated with FN (plate/FN+). Antibody to TGF-beta was added to spin/FN+ to evaluate its impact on transduction. Using producer cell line PA317/LgEIN for transduction of CD34+ cord blood cells, FACS analysis for expression of EGFP revealed mean transduction of 30.6+/-4.3, 9.1+/-1.6, and 21.1+/-6.5% of CD34+ cells in the spin/FN+, spin/FN-, and plate/FN+ arms, respectively. Transduction of CD+CD38low cells was also higher in the spin/FN+ arm as compared with transduction in the spin/FN- arm. These results were corroborated by colony-forming assays. Antibody to TGF-beta did not further increase transduction. Using a different producer cell line, PG13/pLgDIN, a higher number of G418-resistant CFU-GM was observed in the spin/FN+ as compared with the plate/FN+ and spin/FN-arms. NOD/SCID mice were transplanted with transduced, CD34-enriched human cord blood cells, and persistence of transduced human cells was analyzed in the mice marrows after 6-8 weeks: 32.8, 6.0, and 23.9% human G418-resistant CFU-GM colonies were observed in the spin/FN+, spin/FN-, and plate/FN+ arms, respectively. These results suggest that spinoculation in FN-coated tubes increases transduction of early human cord blood progenitor cells as compared with spinoculation in noncoated tubes.
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PMID:Increased gene transfer into human cord blood cells by centrifugation-enhanced transduction in fibronectin fragment-coated tubes. 1058 31

The feasibility of using gene therapy to treat full-thickness articular cartilage defects was investigated with respect to the transfection and expression of exogenous transforming growth factor (TGF)-beta 1 genes in bone marrow-derived mesenchymal stem cells (MSCs) in vitro. The full-length rat TGF-beta 1 cDNA was transfected to MSCs mediated by lipofectamine and then selected with G418, a synthetic neomycin analog. The transient and stable expression of TGF-beta 1 by MSCs was detected by using immunohistochemical staining. The lipofectamine-mediated gene therapy efficiently transfected MSCs in vitro with the TGF-beta 1 gene causing a marked up-regulation in TGF-beta 1 expression as compared with the vector-transfected control groups, and the increased expression persisted for at least 4 weeks after selected with G418. It was suggested that bone marrow-derived MSCs were susceptible to in vitro lipofectamine mediated TGF-beta 1 gene transfer and that transgene expression persisted for at least 4 weeks. Having successfully combined the existing techniques of tissue engineering with the novel possibilities offered by modern gene transfer technology, an innovative concept, i.e. molecular tissue engineering, are put forward for the first time. As a new branch of tissue engineering, it represents both a new area and an important trend in research. Using this technique, we have a new powerful tool with which: (1) to modify the functional biology of articular tissue repair along defined pathways of growth and differentiation and (2) to affect a better repair of full-thickness articular cartilage defects that occur as a result of injury and osteoarthritis.
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PMID:Expression of transforming growth factor beta 1 in mesenchymal stem cells: potential utility in molecular tissue engineering for osteochondral repair. 1265 48

BMP6 is a member of TGF-beta superfamily, represent more effective osteogenic activity. Two recombinant plasmids were constructed to expression rhBMP6 in mammalian cells, one contained the cDNA encoding the signal peptide, propeptide and mature peptide of human BMP6, wich was named pcDNA-BMP6, the other contained the recombinant DNA encoding the signal peptide, propeptide of human BMP2 and the mature peptide of BMP6, which was named pcDNA-BMP2/6. Transient expression in Cos7 cells demonstrated that the pcDNA-BMP2/6 produced more rhBMP6 than pcDNA-BMP6. For stable expression, the CHO-dhfr- cells were transfected with pcDNA-BMP2/6 and pSV2-dhfr, then screened by G418 and treated with MTX for targeting gene amplification. The partially purified rhBMP6 by heparin affinity chromatography was shown to possess bone induction activity tested by the induction of alkaline phosphatase activity in C2C12 cells.
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PMID:[Expression of recombinant human BMP6 in CHO cells by fused to the signal peptide and propeptide of another homologue protein]. 1757 85