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)

Our previous study demonstrated the positive relationship between the gene introduction rate into hematopoietic cell lines by electroporation and the percentage of cells in S-phase. In the present study, granulocyte-macrophage progenitor cells (CFU-C) rich marrow cell fraction were cultured in suspension with IL-3, GM-CSF and G-CSF for 4 days. The number of CFU-C were increased three times after the culture, and 3H-thymidine suicide tests of cultured cells demonstrated that the proportion of CFU-C in S-phase was increased by two to four times. The efficiency of gene transfer into CFU-C with the plasmid pMoZtk (containing the beta-galactosidase gene) by electroporation was nearly doubled by culturing marrow cells with these growth factors. These findings confirm that the introduction rate of the gene into CFU-C by electroporation is more efficient in cell populations with a higher percentage of CFU-C in S-phase.
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PMID:Gene introduction into granulocyte-macrophage progenitor cells by electroporation: the relationship between introduction efficiency and the proportion of cells in S-phase. 152 64

To examine the potential use of adenovirus vectors in cancer gene therapy as a mechanism for purging bone marrow cells of possible breast cancer contaminants, we compared the infection efficiency of adenovirus and the transfection efficiency of plasmid DNA in the presence of adenovirus in human breast cancer and bone marrow cells. Following infection of breast cancer cells with an adenovirus expressing beta-galactosidase gene, high levels of beta-galactoside activity were observed. No beta-galactosidase activity was observed in low-density human bone marrow cells. A replication-deficient adenovirus mutant dl312 enhanced the transfection efficiency of a plasmid DNA-expressing beta-galactosidase gene into breast cancer cells, and addition of a liposome, lipofectamine, further enhanced the transfection efficiency. In contrast, human bone marrow cells treated under the same conditions expressed very low levels of transfected beta-galactosidase DNA. Transfection of cells with plasmid DNA expressing a truncated but fully active Pseudomonas exotoxin gene in the presence of dl312 and lipofectamine resulted in marked breast cancer cell killing, whereas colony-forming unit granulocyte-macrophage (CFU-GM) were relatively resistant to these treatments. A recombinant adenovirus expressing human wild-type p53 protein (AdWTp53) was also highly cytotoxic to breast tumor cells. Infection of breast cancer cells with AdWTp53 (100 plaque-forming units/cell) resulted in 100% loss of the clonogenicity of breast tumor cells. However, colony formation from CFU-GM was relatively resistant to the cytotoxic effects of AdWTp53 alone or in the presence of pULI100 plasmid and lipofectamine. On the basis of these results, it is proposed that human adenoviruses are potentially useful for cancer gene therapy and bone marrow purging.
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PMID:Adenovirus-mediated gene transfer to human breast tumor cells: an approach for cancer gene therapy and bone marrow purging. 864 Aug 24

One of the major questions in any gene therapy approach is the selection of the appropriate vector system. Here, the optimization of a gene transfer protocol for renal cell carcinoma using lipofection as a nonviral gene transduction system was evaluated. To select the promoter which gives the highest expression, different plasmids which are able to express Escherichia coli beta-galactosidase gene as a reporter gene under the control of different promoters were tested: human cytomegalovirus promoter (pCMVbeta), simian virus 40 promoter (pSVbeta), adenovirus promoter (ADbeta), and herpes simplex virus thymidine kinase promoter (TKbeta). The pCMVbeta revealed the highest expression of the beta-gal gene in the renal cell carcinoma (RCC) lines. Thus this CMV promoter was selected for the expression of the granulocyte-macrophage colony stimulator factor (GM-CSF) gene. Three different lipids (LipofectAmine, LipofectAce, and Lipofectin) were compared for their transduction efficiency, and the optimal conditions for quantitatively high lipofection rates were established. The consistently best results regarding gene expression as well as viability of the RCC lines were obtained when Lipofectin was used. Gene expression was monitored by a specific enzyme-linked immunosorbent assay and functionally validated by a cell proliferation test. The GM-CSF expression profile showed a peak at 48 hours after transfection and was still detectable after 5 days. Here the feasibility of efficient lipofection of the GM-CSF gene into RCC lines is demonstrated. Most importantly, considerable differences in the relative quantity of GM-CSF gene transfer into the different RCC lines was observed here. This may be of critical relevance for the design of any clinical gene transduction protocol in tumor cell vaccination attempts.
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PMID:Differential transfection efficiency rates of the GM-CSF gene into human renal cell carcinoma lines by lipofection. 901 52

Tumor cells genetically modified to coexpress certain cytokines (such as IL-7 or IL-4) and B7.1 have increased immunogenicity. Since tumor Ags can be presented either directly by tumor cells or indirectly by host APC (cross-priming), we asked whether B7.1 and IL-7 or IL-4 complemented each other by improving preferentially one or both pathways of Ag presentation. We used TS/A (H-2d) tumor cells and their IL-7, B7, and IL-7/B7 transfectants, and MCA205 (H-2b) tumor cells and their IL-4 and B7 transfectants. beta-galactosidase (beta-gal) was chosen as surrogate tumor Ag. beta-gal has different predominant MHC class I epitopes in H-2d and H-2b mice. Immunization of (H-2b x d)F1 mice with TS/A/beta-gal transfectants showed that both IL-7 and B7.1 and, as control, granulocyte-macrophage CSF augmented cross-priming and rejection of a challenge with MCA205/beta-gal (H-2b). Similarly, immunization with MCA205/beta-gal B7.1 or IL-4 transfectants enhanced cross-priming and rejection of a challenge with TS/A/beta-gal. beta-gal-specific rejection was confirmed by CTL assay. However, direct Ag presentation by tumor cells was enhanced only by B7.1, and not IL-7. For this study, H-2b nu/nu mice reconstituted with F1 lymphocytes were immunized with H-2d TS/A/beta-gal transfectants and challenged with TS/A/beta-gal. In conclusion, indirect Ag presentation was augmented by B7, IL-7, and IL-4, while direct Ag presentation was improved only by B7.
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PMID:Influence of gene-modified (IL-7, IL-4, and B7) tumor cell vaccines on tumor antigen presentation. 905 19

To determine the kinetics of tissue macrophage and microglial engraftment after bone marrow (BM) transplantation, we have developed a model using the ROSA 26 mouse. Transplanted ROSA 26 cells can be precisely identified in recipient animals because they constitutively express beta-galactosidase (beta-gal) and neomycin resistance. B6/129 F2 mice were irradiated and transplanted with BM from ROSA 26 donors and their tissues (spleen, marrow, brain, liver, and lung) examined at various time points to determine the kinetics of engraftment. Frozen sections from transplanted animals were stained histochemically for beta-gal to identify donor cells. At 1, 2, 6, and 12 months posttransplantation, 98% to 100% of granulocyte-macrophage colonies were of donor (ROSA 26) origin determined by beta-gal staining and by neomycin resistance. Splenic monocytes/macrophages were 89% donor origin by 1 month confirming quick and complete engraftment of hematopoietic tissues. At this time, only rare ROSA 26 tissue macrophages or microglia were observed. Alveolar macrophage engraftment was evident by 2 months and had increased to 61% of total tissue macrophages at 1 year posttransplantation. The kinetics of liver Kupffer cell engraftment were similar to those seen in the lung. However, donor microglial engraftment remained only 23% of total microglia at 6 months and increased to only 30% by 1 year. Also, donor microglia were predominantly seen at perivascular and leptomeningeal, and not parenchymal, sites. The data show that microglia derive from BM precursors but turn over at a significantly slower rate than other tissue macrophages. No clinical or histological graft-versus-host disease was observed in the recipients of ROSA 26 BM. These kinetics may impact strategies for the gene therapy of lysosomal storage diseases. Because individual donor cells can be identified in situ, the ROSA 26 model should have many applications in transplantation biology including studies of homing and differentiation.
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PMID:Kinetics of central nervous system microglial and macrophage engraftment: analysis using a transgenic bone marrow transplantation model. 924 27