Reagents

Anti-human PTEN monoclonal antibody (cat. no. M3627; 1:1,000 dilution) was obtained from Dako (Carpinteria, CA, USA). Monoclonal anti-mouse IgG horseradish peroxidase conjugate (cat. no. NXA931; 1:2,000 dilution) was purchased from GE Healthcare (Little Chalfont, UK). Monoclonal anti-human HMGB1 (cat. no. H9537; 1:2,000) and β-actin (cat. no. A5316; 1:10,000) antibodies were purchased from Sigma-Aldrich (St. Louis, MO, USA). Human recombinant HMGB-1 expressed in Escherichia coli was from Sigma-Aldrich (cat no. H4652; St. Louis, MO, USA). miRIDIAN Hairpin Inhibitor hsa-miRNA-221 (cat. no. IH300578-07-005) and hsa-miRNA-222 (cat. no. IH301176-02 −005) and miRIDIAN microRNA Hairpin Inhibitor Transfection Control with Dy547 (cat. no. FE5IP0045000105) (control oligonucleotide) and DharmaFECT 1 Transfection Reagent (cat. no. FE5T200103) were purchased from GE Healthcare Dharmacon Inc. (Lafayette, CO, USA). The mature miRNA-221 sequence was 5′-AGCUACAUUGUCUGCUGGGUUUC-3′ and the mature miRNA-222 sequence was 5′-CUCAGUAGCCAGUGUAGAUCCU-3′. Reverse transcription (RT) primers and TaqMan probes were obtained from TaqMan miRNA assays (Applied Biosystems, Foster City, CA, USA).