Glial cell line-derived neurotrophic factor (GDNF) is a proliferation factor for rat C6 glioma cells: evidence from antisense experiments

Growth factors play an important role in proliferation and differentiation of malignant brain gliomas in humans. Glial cell line-derived neurotrophic factor (GDNF) has been shown recently to be highly expressed in human gliob lastomas and in rat glial cell lines B49 and C6. The aim of the present stu dy was to knockdown GDNF, its receptor GFR-alpha1, and the related family m ember persephin by using antisense oligonucleotides and to observe the effe cts on cell proliferation. To enhance cellular uptake into C6 glioma cells, 15-mer phosphorothioate oligonucleotides were complexed with the cationic lipid Lipofectamine(TM). The complex was applied for 3 x 12 hours to C6 gli oma cells, and cells were allowed to recover for 24 hours after each transf ection and then analyzed. This protocol markedly reduced GDNF and GFR-alpha 1 protein levels in C6 glioma cells compared with control oligonucleotides, Knockdown of C6 cells with GDNF and GFR-alpha1 but not with persephin anti sense oligonucleotides significantly decreased the number of C6 glioma cell s and also inhibited the incorporation of bromodeoxyuridine as a sign of re duced DNA synthesis. In conclusion, it is shown that GDNF but not persephin is a potent proliferation factor for rat glioma cells. Knockdown of GDNF u sing antisense oligonucleotides complexed with lipids as carriers may be us eful in gene therapeutic approaches in vitro and possibly also in vivo.