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Figure 3 | Molecular Medicine

Figure 3

From: Cordycepin Blocks Lung Injury-Associated Inflammation and Promotes BRCA1-Deficient Breast Cancer Cell Killing by Effectively Inhibiting PARP

Figure 3

Cordycepin reduces H2O2-induced PARP activation and inhibits in vitro PARP-1 activity with great potency. (A) A549 cells were treated with H2O2 for 10 min in the presence or absence of 10 or 20 μmol/L cordycepin. A portion of the H2O2-treated cells also was treated with the known PARP inhibitors NU1025 (100 μmol/L) or TIQ-A (2 μmol/L). Protein extracts were prepared and subjected to immunoblot analysis with antibodies to poly(ADP-ribose) (PAR) or GAPDH. (B) A549 cells were treated with H2O2 for 10 min in the presence or absence of 10 μmol/L cordycepin. Cells were then fixed and subjected to immunofluorescence with antibodies to poly(ADP-ribose). Note that the cytosolic staining is non-specific as it appears in all cells. (C) A549 cells were treated the same as in (B) after which they were washed with and incubated in complete medium for 1 h. Cells were then treated once more with H2O2 for 10 min in the absence of any additional cordycepin. Cells were then fixed and subjected to immunofluorescence with antibodies to poly(ADP-ribose). (D) Recombinant human PARP-1 was incubated for 5 min in an in vitro poly(ADP-ribosyl)ation reaction in the absence or presence of different concentrations of cordycepin or in the presence of NU1025 or TIQ-A (both PARP inhibitors) with NAD+, activated (sonicated) DNA, and protein extracts from PARP-1−/−smooth muscle cells. The reactions were terminated with sample buffer and subjected to immunoblot analysis with antibodies to poly(ADP-ribose) (PAR) or actin.

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