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In vivo gene transfer using cetylated polyethylenimine.

100%

Sochanik A., Cichoń T., Makselon M., Stróżyk M., Smolarczyk R., Jazowiecka-Rakus J., Szala S.

Acta Biochimica Polonica

2004

vol. 51

issue 3

693-702

This report describes gene transfer in vitro as well as in vivo using cetylated low-molecular mass (600 Da) polyethylenimine (28% of amine groups substituted with cetyl moieties), termed CT-PEI. This compound is hydrophobic and has to be incorporated into liposomes in order to be suitable for gene transfer studies. Serum-induced plasmid DNA degradation assay demonstrated that CT-PEI-containing liposomal carriers could protect complexed DNA (probably via condensation). In vitro luciferase gene expression achieved using medium supplemented with 10% serum was comparable to that achieved in serum-reduced medium and was highest for CT-PEI/cholesterol liposomes, followed by CT-PEI/dioleoylphosphatidylcholine liposomes and PEI 600 Da (uncetylated) carrier. In vivo systemic transfer into mice was most efficient when liposome formulations contained CT-PEI and cholesterol. Higher luciferase expression was then observed in lungs than in liver. In conclusion: liposomes containing cetylated polyethylenimine and cholesterol are a suitable vehicle for investigating systemic plasmid DNA transfer into lungs.

Doping in Sport: New Developments

100%

Krych K., Goździcka-Józefiak A.

Human Movement

2008

vol. 9

issue 1

62-75

Gene doping is defined by the World Anti-Doping Agency (WADA) as "the non-therapeutic use of genes, genetic elements and/or cells that have the capacity to enhance athletic performance." The rapid development of molecular biology has enabled not only treatment of many diseases, but also improvement of athletes' fitness. Gene therapy methods can be used to modify the athlete's body by inserting genes into the target tissue. It is very possible that in near future, many genes will be used in gene doping, e.g. erythropoietin, growth hormone, insulin-like growth hormone and vascular endothelial growth factor. Functional tests conducted by many independent laboratories proved that products of these genes exert a crucial influence on the body's adaptation to exercise. The risk of gene doping is enormous. Gene therapy is currently in the phase of clinical tests so it is impossible to predict what kind of side effects it may produce. Studies on animal models showed that the uncontrolled transgene expression and insertional mutagenesis can even lead to death. At present the detection of gene doping is very difficult for a variety of reasons. The main problem is the identification of the transgene and endogenously produced protein. The only possible detection is the biopsy of the target tissue, where the exogenous genes were inserted.

Nonviral transfection of human umbilical cord blood dendritic cells is feasible, but the yield of dendritic cells with transgene expression limits the application of this method in cancer immunotherapy

75%

Markowicz S., Niedzielska J., Kruszewski M., Ołdak T., Gajkowska A., Machaj E. K., Skurzak H., Pojda Z.

Acta Biochimica Polonica

2006

vol. 53

issue 1

203-212

Dendritic cells (DC) generated from human umbilical cord blood might replace patients' DC in attempts to elicit tumor-specific immune response in cancer patients. We studied the efficiency of transfection of human cord blood DC with plasmid DNA carrying the enhanced version of green fluorescent protein (EGFP) as a reporter gene, to test if nonviral gene transfer would be a method to load DC with protein antigens for immunotherapy purposes. Cord blood mononuclear cells were cultured in serum-free medium in the presence of granulocyte-monocyte colony stimulating factor (GM-CSF), stem cell factor (SCF) and Flt-3 ligand (FL), to generate DC from their precursors, and thereafter transfected by electroporation. Maturation of DC was induced by stimulation with GM-CSF, SCF, FL and phorbol myristate acetate (PMA). Transfected DC strongly expressed EGFP, but transfection efficiency of DC, defined as HLA-DR+ cells lacking lineage-specific markers, did not exceed 2.5%. Expression of the reporter gene was also demonstrated in the DC generated from transfected, purified CD34+ cord blood cells, by stimulation with GM-CSF, SCF, FL, and tumor necrosis factor α (TNF-α). Transfection of CD34+ cells was very efficient, but proliferation of the transfected cells was much reduced as compared to the untransfected cells. Therefore, the yield of transgene-expressing DC was relatively low. In conclusion, nonviral transfection of cord blood DC proved feasible, but considering the requirements for immunotherapy in cancer patients, transfection of differentiated DC or generation of DC from transfected hematopoietic stem cells provide only a limited number of DC expressing the transgene.

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2 Acta Biochimica Polonica

1 Human Movement

1 2008

1 2006

1 2004

1 Cichoń T.

1 Gajkowska A.

1 Goździcka-Józefiak A.

1 Jazowiecka-Rakus J.

1 Kruszewski M.

1 Krych K.

1 Machaj E. K.

1 Makselon M.

1 Markowicz S.

1 Niedzielska J.

1 Ołdak T.

1 Pojda Z.

1 Skurzak H.

1 Smolarczyk R.

1 Sochanik A.

1 Stróżyk M.

1 Szala S.

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In vivo gene transfer using cetylated polyethylenimine.

Doping in Sport: New Developments

Nonviral transfection of human umbilical cord blood dendritic cells is feasible, but the yield of dendritic cells with transgene expression limits the application of this method in cancer immunotherapy