Advanced search
1 file | 558.05 KB

Efficient genetic modification of murine dendritic cells by electroporation with mRNA

(2002) CANCER GENE THERAPY. 9(9). p.787-797
Author
Organization
Abstract
Recently, human dendritic cells (DCs) pulsed with mRNA encoding a broad range of tumor antigens have proven to be potent activators of a primary anti-tumor-specific T-cell response in vitro. The aim of this study was to improve the mRNA pulsing of murine DC. Compared to a standard lipofection protocol and passive pulsing, electroporation was, in our hands, the most efficient method. The optimal conditions to electroporate murine bone marrow-derived DCs with mRNA were determined using enhanced green fluorescent protein and a truncated form of the nerve growth factor receptor. We could obtain high transfection efficiencies around 70-80% with a mean fluorescence intensity of 100-200. A maximal expression level was reached 3 hours after electroporation. A clear dose-response effect was seen depending on the amount of mRNA used. Importantly, the electroporation process did not affect the viability nor the allostimulatory capacity or phenotype of the DC. To study the capacity of mRNA-electroporated DCs to present antigen in the context of MHC classes I and II, we made use of chimeric constructs of ovalbumin. The dose-dependent response effect and the duration of presentation were also determined. Together, these results demonstrate that mRNA electroporation is a useful method to generate genetically modified murine DC, which can be used for preclinical studies testing immunotherapeutic approaches.
Keywords
INVARIANT CHAIN, CANCER-IMMUNOTHERAPY, N-END RULE, THERAPEUTIC ANTITUMOR IMMUNITY, RESPONSES IN-VITRO, CYTOTOXIC T-LYMPHOCYTES, cancer vaccine, antigen presentation, mRNA electroporation, dendritic cells, TUMOR-IMMUNITY, BONE-MARROW, CYTOPLASMIC TAIL, SOLUBLE-PROTEIN

Downloads

  • (...).pdf
    • full text
    • |
    • UGent only
    • |
    • PDF
    • |
    • 558.05 KB

Citation

Please use this url to cite or link to this publication:

Chicago
Van Meirvenne, Sonja, Lieven Straetman, Carlo Heirman, Melissa Dullaers, Catherine De Greef, Viggo Van Tendeloo, and Kris Thielemans. 2002. “Efficient Genetic Modification of Murine Dendritic Cells by Electroporation with mRNA.” Cancer Gene Therapy 9 (9): 787–797.
APA
Van Meirvenne, S., Straetman, L., Heirman, C., Dullaers, M., De Greef, C., Van Tendeloo, V., & Thielemans, K. (2002). Efficient genetic modification of murine dendritic cells by electroporation with mRNA. CANCER GENE THERAPY, 9(9), 787–797.
Vancouver
1.
Van Meirvenne S, Straetman L, Heirman C, Dullaers M, De Greef C, Van Tendeloo V, et al. Efficient genetic modification of murine dendritic cells by electroporation with mRNA. CANCER GENE THERAPY. 2002;9(9):787–97.
MLA
Van Meirvenne, Sonja, Lieven Straetman, Carlo Heirman, et al. “Efficient Genetic Modification of Murine Dendritic Cells by Electroporation with mRNA.” CANCER GENE THERAPY 9.9 (2002): 787–797. Print.
@article{1227832,
  abstract     = {Recently, human dendritic cells (DCs) pulsed with mRNA encoding a broad range of tumor antigens have proven to be potent activators of a primary anti-tumor-specific T-cell response in vitro. The aim of this study was to improve the mRNA pulsing of murine DC. Compared to a standard lipofection protocol and passive pulsing, electroporation was, in our hands, the most efficient method. The optimal conditions to electroporate murine bone marrow-derived DCs with mRNA were determined using enhanced green fluorescent protein and a truncated form of the nerve growth factor receptor. We could obtain high transfection efficiencies around 70-80\% with a mean fluorescence intensity of 100-200. A maximal expression level was reached 3 hours after electroporation. A clear dose-response effect was seen depending on the amount of mRNA used. Importantly, the electroporation process did not affect the viability nor the allostimulatory capacity or phenotype of the DC. To study the capacity of mRNA-electroporated DCs to present antigen in the context of MHC classes I and II, we made use of chimeric constructs of ovalbumin. The dose-dependent response effect and the duration of presentation were also determined. Together, these results demonstrate that mRNA electroporation is a useful method to generate genetically modified murine DC, which can be used for preclinical studies testing immunotherapeutic approaches.},
  author       = {Van Meirvenne, Sonja and Straetman, Lieven and Heirman, Carlo and Dullaers, Melissa and De Greef, Catherine and Van Tendeloo, Viggo and Thielemans, Kris},
  issn         = {0929-1903},
  journal      = {CANCER GENE THERAPY},
  keyword      = {INVARIANT CHAIN,CANCER-IMMUNOTHERAPY,N-END RULE,THERAPEUTIC ANTITUMOR IMMUNITY,RESPONSES IN-VITRO,CYTOTOXIC T-LYMPHOCYTES,cancer vaccine,antigen presentation,mRNA electroporation,dendritic cells,TUMOR-IMMUNITY,BONE-MARROW,CYTOPLASMIC TAIL,SOLUBLE-PROTEIN},
  language     = {eng},
  number       = {9},
  pages        = {787--797},
  title        = {Efficient genetic modification of murine dendritic cells by electroporation with mRNA},
  url          = {http://dx.doi.org/10.1038/sj.cgt.7700499},
  volume       = {9},
  year         = {2002},
}

Altmetric
View in Altmetric
Web of Science
Times cited: