Objective: To examine the effect of silencing SKP2 on chemosensitivity of human glioma cells U251 to temozolomide (TMZ).
Methods: Adenoviruses harbouring shRNA targeting SKP2 (i.e. Ad-shSKP2) and non-targeting scrambled shRNA (i.e. Ad-shNC) were used to infect U251 cells. The transduced cells were then treated with TMZ. Cell viability after treatment was assayed using CCK8; while cell cycle and apoptosis were examined using flow cytometry. To study the effect of silencing SKP2 on autophagy in U251, we co-transduced the cells with Ad-mRFP-LC3 and Ad-shSKP2/Ad-shNC. The expression of autophagy marker LC3 after TMZ treatment was studied using microscopy and Western blotting assays.
Results: The cytotoxicity of TMZ (i.e. 20-100 µM) was more significantly seen in Ad-shSKP2-transduced U251 cells than in the Ad-shNC-transduced U251 cells. The IC50 values in shSKP2-U251 were significantly lower than those of the shNC-U251 (P < 0.05). Both TMZ and Ad-shSKP2 alone increased apoptosis and promoted expression of LC3 in U251. Combined treatment of Ad-shSKP2 and TMZ further elevated apoptosis and LC3 expression.
Conclusion: Silencing SKP2 in U251 cells increased chemosensitivity to TMZ that was accompanied with enhanced apoptosis and autophagy. Targeting SKP2 may be a potential approach to potentiate TMZ treatment in patients with glioma.
Keywords: Glioma; S-phase kinase-associated protein 2 (SKP2); autophagy; chemosensitivity; temozolomide (TMZ).