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Biomedical subjects

O Gaspar

Publications and source records attributed to O Gaspar.

6 recordsLinked to original sources
PubMed1996

Transient protection of human T-cells from human immunodeficiency virus type 1 infection by transduction with adeno-associated viral vectors which express RNA decoys.

RNA decoys are oligonucleotides corresponding to the TAR and RRE sequences of HIV which inhibit the HIV-encoded regulatory proteins Tat and Rev, respectively. Adeno-associated viral vectors encoding RNA decoys stably transduced into the human T-cell line CEM-SS expressed transactivating region (TAR) and Rev-responsive element (RRE) RNA decoys from tRNA polIII promoters at high levels, without any apparent deleterious effects on cell growth or expression of CD4. DNA blot analysis indicated that RNA decoy-encoding vectors were not rearranged and were integrated into the genomic DNA of selected cell lines. Vector DNA with the appropriate TAR and RRE sequences was isolated from transduced cell lines after prolonged growth in culture, further confirming that the vector DNA was present in a stable form through multiple cell cycles. Cell lines expressing TAR and RRE decoys transiently inhibited HIV gene expression and replication by 70-99% as determined by measurement of intracellular and extracellular HIV p24 production. Adeno-associated vectors encoding RNA decoys may be useful for gene therapy of HIV infection.

Cell Line↗
PubMed1996

High-efficiency retroviral vector mediated gene transfer into human peripheral blood CD4+ T lymphocytes.

Genetic modification of peripheral blood T lymphocytes has been proposed as a therapeutic strategy for treating congenital disorders, cancer and viral diseases. Central to all T lymphocyte-based gene therapy strategies is the ability to efficiently and stably deliver genes into primary T lymphocytes. In this study, we sought to increase the gene transfer efficiency in CD4+ peripheral blood T lymphocytes using procedures which could be utilized in clinical applications. In order to quantity the gene transfer efficiency in primary CD4+ T cells, a high-titer retroviral vector which efficiently expresses a truncated version of the human low-affinity nerve growth factor receptor (delta LNGFR) was constructed. Transduced cells were then accurately enumerated with immunofluorescence staining and fluorescence activated cell sorting (FACS) analysis and rapidly isolated at high purity for further analysis. Using this system, a supernatant-based gene transfer procedure was developed which routinely yields gene transfer efficiencies of 25-40% into a wide repertoire of both freshly obtained and cryopreserved peripheral blood CD4+ T lymphocytes.

3T3 Cells↗
PubMed1995

Inhibition of HIV-1 in CEM cells by a potent TAR decoy.

TAR decoys are short RNA oligonucleotides, corresponding to the HIV TAR sequence, which inhibit HIV expression and replication by blocking the binding of the HIV regulatory protein Tat to the authentic TAR region. In previous studies, TAR decoys expressed from a tRNA polIII promoter were moderately effective at inhibiting HIV in isolated human T cell lines and less effective at inhibiting HIV in peripheral blood CD4+ T cells. In this study, a series of modifications was introduced into the tRNA expression cassette in order to improve their effectiveness. These modifications included the addition of sequences which are predicted to have stem-loop secondary structures and addition of a wild-type tRNA processing site. TAR decoy RNA expressed in CEM cells from modified tRNA-based expression cassettes yielded five- to 20-fold more TAR transcripts than unmodified tRNA-based expression cassettes. HIV replication, as measured by a flow cytometric method to quantify intracellular viral p24 expression, was significantly reduced in polyclonal populations of CEM cells expressing a modified tRNA-TAR transcript that contains a wild-type tRNA processing site and stem-loops 5' and 3' to the TAR sequence. Similar modifications to the tRNA expression cassette also increased the intracellular concentration of a random test oligonucleotide, indicating that this improved expression system may also be useful for antisense and ribozyme based gene inhibition strategies.

Antiviral Agents↗
PubMed1985

Hickman catheter tip displacement.

In a series of 60 consecutive patients in whom Hickman catheters were placed for treatment of malignancy, four properly positioned catheter tips migrated secondarily from the superior vena cava to the ipsilateral jugular vein 2, 4, 21, and 25 days after placement. Three of the four patients had begun to have catheter dysfunction when the displacement was diagnosed. No satisfactory explanation for this rarely reported complication was evident in three of the cases. Maneuvers such as coughing, Valsalva's maneuver, and forceful heparin flushing produced no motion in three normally directed catheter tips in other patients observed under fluoroscopy. The phenomenon may be more common than previously reported. Evaluation of any new Hickman catheter dysfunction should include a chest x-ray film to ascertain the position of the catheter.

Adult↗