Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0432222 (SEM)
 47,337 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The ability of the N2 retrovirus to introduce the selectable neo gene into (transform) caprine hematopoietic cells (CHC) was evaluated. Helper-free amphotropic retrovirus producing cells (RPC) were plated at approximately 1.0 x 10(5) cells per 25 cm2 tissue culture flask, cultured to 80% confluence and irradiated (1,500 rads) prior to CHC incubation. The CHC collected from three donor goats were washed and cultured for 24 h in either the RPC or control flasks. Cells were cultured in Dulbecco's Modified Eagles Medium containing 10% fetal calf serum (FCS) (DMEM) and 4 micrograms polybrene/ml. After 48 h of culture in fresh DMEM, cells were recovered and suspended in Iscove's DMEM supplemented with .3% gar, 12% FCS and 400 micrograms geneticin/ml (G418; neomycin) and transferred to 35-mm petri dishes (7.5 x 10(5) cells) for selection of G418 resistant cells. After 17 d of culture, plates were evaluated for total number of colony forming units (CFU, greater than 10 cells). Total number of CFU was greater (P less than .01) in treatment samples (means = 175, SEM = 70) than in control cultures (mean = 0, SEM = 0). The N2 retrovirus appears to be an effective vector for the transformation of CHC and may provide a means to introduce gene(s) into cells of domestic animals.
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PMID:Incorporation and expression of a neo gene after transfer to caprine hematopoietic cells using a modified retrovirus. 231 32

To help establish an effective gene therapy protocol for patients with congenital metabolic diseases, we evaluated retrovirus-mediated transduction and long-term (LT) expression of the NeoR gene in cryopreserved and thawed CD34+ cells purified from granulocyte colony-stimulating factor (G-CSF)-mobilized peripheral blood (PB) of infant and cord blood (CB). The results were compared with those in bone marrow (BM) CD34+ cells. The final purity of the CD34+-enriched fraction from PB, CB, and BM, based on FACS analysis, was 88 +/- 14%, 73 +/- 13%, and 68 +/- 19% (mean +/- SEM), respectively. Cells were then cultured for 96 hours with supernatant containing the vector in the presence of interleukin (IL)-3, IL-6, and stem cell factor (SCF). The average efficiency of gene transfer into mobilized PB (n = 5) or CB CD34+ cells (n = 6) was significantly higher than that into BM CD34+ cells, as measured by G418-resistant colony-forming units for granulocyte/macrophage (CFU-GM; 59% or 58% vs. 39%; p < 0.05) and PCR-positive CFU-GM (83% or 79% vs. 53%; p < 0.05). When the evaluation was made in an LT culture system with irradiated allogeneic marrow stroma, these efficiencies were, respectively, 74% or 61% vs. 34% (p < 0.005 or < 0.02) for G418-resistant CFU-GM at week 5 of long-term culture, and 88% or 83% vs. 63% (p < 0.05) for PCR-positive CFU-GM. Fluorometric examination was performed for cell-cycle analysis before and after culture, and the results showed that the fraction of cycling cells was largest in freshly prepared BM (18%), whereas only a small portion of PB (4.6%) and CB (2%) was cycling. However, this value was 17% in BM, 22% in PB, and 13% in CB after culture. These results suggest that mobilized PB from small children and CB cells are suitable and realistic targets for clinical gene therapy and that tandem transduction procedures can be achieved by combining CB and PB.
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PMID:Transduction of retrovirus-mediated NeoR gene into CD34+ cells purified from granulocyte colony-stimulating factor (G-CSF)-mobilized infant and cord blood. 950 13