IL-13 Gene Translation is Arrested by a Novel Oligonucleotide in Cultured Human B-Lymphocytes.
Antisense oligodeoxynucleotides (oligos) are the tools that bind to complementary sequence of targeted mRNA and block specifically protein translation. In the present study, a novel 20 mer oligo as an antisense for human IL-13 is introduced. This oligo is designed according to the IL-13 mRNA coding region and synthesized in two HPLC purified and FITC conjugated forms. Fluorescence oligo cell uptake is confirmed using flowcytometry and confocal microscopy, and cytotoxicity evaluation is performed using BrdU proliferation assay. Human tonsilar B-lymphocytes are purified by positive selection using magnetic cell sorting method and cultured with anti CD40 monoclonal antibody plus rIL-4 to induce IL-13 production. IL-13 antisense is added to medium and Real Time PCR for mRNA, and ELISA for protein assays. Data indicate that antisense application leads to down regulation and complete suppression of IL-13 protein with no significant effects on mRNA, suggesting in vitro protein translation arrest. Since Il-13 is a crucial cytokine in allergic conditions, we conclude that interference with the protein synthesis by a nontoxic and efficient antisense oligo can provide an available tool for the investigators on allergic diseases.
Mousavi
Tahere
T
Biotechnology department, Razi Institute, Iran. mousavi36@yahoo.com.
Mazer
Bruce
B