Transfection of mammalian cells by the methods of receptor mediated gene transfer and particle bombardment.
The efficacy of currently developed methods for gene transfer into mammalian cells depends primarily on the transfection technique, and also on the type of targeted cells. Considering the importance of gene transfer in the creation of gene therapies, our study was aimed at the assessment of transfection capacity of receptor mediated gene transfer method (RMGT), and method of particle bombardment (helios gene gun system--HGG) in different normal and malignant mammalian cells ex vivo. In addition, the HGG was also assessed for its ability to transfect tumor cells of subcutaneous (s.c.) tumors in C57Bl/6 mice in vivo. Using RMGT an average ex vivo transfection rate of 35.7%, and 20.4% was achieved in malignant melanoma B-16, and human breast adenocarcinoma MCF7, respectively. However, in normal fibroblast L929 cells the transfection by RMGT succeeded only in 2.1% of the cells. On the other hand, the transfection efficacy of HGG was comparable in both malignant cell lines resulting in an average gene transfer to 9.6% of B-16 and 10.5% of MCF7 cells, while only 3.9% of normal fibroblasts were successfully transfected. Application of HGG for an in vivo gene transfer into s.c. B-16 melanoma tumors in C57Bl/6 mice resulted in a successful but limited transfection of the epithelium as well as of the superficially sited tumor cells. Taking into consideration both methods, RMGT is more appropriate for ex vivo transfection of cells, under the condition that target cells express a specific receptor for the molecule attached to the carrier. On the other hand, HGG is not complicated to use, no requirements for specific structures on target cells are necessary (potentially usable in different cells), and it has applicability in direct in vivo transfection processes.
Novaković
S
S
Dept. of Tumor Biology, Institute of Oncology, Ljubljana, Slovenia.
Knezević
M
M
Golouh
R
R
Jezersek
B
B
D000230:Adenocarcinoma; D000818:Animals; D019470:Biolistics / Q000379:methods*; D001943:Breast Neoplasms; D005260:Female; D018014:Gene Transfer Techniques; D017930:Genes, Reporter; D006801:Humans; D007739:L Cells; D008546:Melanoma, Experimental / Q000473:pathology*; D051379:Mice; D008810:Mice, Inbred C57BL; D011990:Receptors, Transferrin / Q000235:genetics / Q000502:physiology; D014162:Transfection / Q000379:methods*; D014407:Tumor Cells, Cultured; D001616:beta-Galactosidase / Q000032:analysis / Q000235:genetics
D011990:Receptors, Transferrin; D001616:beta-Galactosidase