Figure 1. Silencing GAPDH and its effect on CSF-1 mRNA and protein expression.
(A) Immunoblot analysis of total protein extracts from control or GAPDH siRNA treated NOSE.1 or Hey cells. The extracts were harvested 48 hours post-siRNA transfection. The membrane was probed with antibodies to GAPDH and β-actin. GAPDH siRNA treatment significantly reduced or abolished GAPDH protein. (B) RT-PCR analysis of the CSF-1 and β-actin RNA expression in GAPDH or control siRNA treated NOSE.1 or Hey cells. The RNAs were harvested 24 hours post-siRNA transfection. GAPDH siRNA significantly down-regulated CSF-1 RNA compared to controls. (C) Levels of CSF-1 secreted in the conditioned media of GAPDH or control siRNA treated NOSE.1 or Hey cells (ng/ml). The media was harvested 57 hours post-siRNA transfection. There was no significant effect of control siRNA on secreted CSF-1 levels compared to untreated Hey cells (p=0.15). GAPDH siRNA was able to significantly decrease secreted CSF-1 protein levels when compared to controls.