Myd88

Cells were lysed and soluble proteins were extracted using the radio immunoprecipitation assay (RIPA) buffer (Beyotime, Shanghai, China) containing a protease inhibitor cocktail and sodium orthovanadate (Santa Cruz Biotechnology, Dallas, TX). Protein concentration was determined using a BCA protein assay kit (Pierce Biotechnology, Rockford, IL). Protein samples were then run on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) with 10% polyacrylamide, and electro-transferred onto polyvinyl difluoride (PVDF) membranes (Millipore, Billerica, MA). The membranes were probed with specific primary and secondary antibodies, and developed with chemiluminescence (ECL, Thermo Fisher) using the ChemiDoc Touch Imaging System (BioRad). Quantification of bands was achieved by a densitometry, with β-actin as a reference control. Specific primary antibodies against the following proteins were used: c-MYC (SAB, 1:1000), SOX2 (R&D Systems, 1:1000), OCT4 (Cell Signaling, 1:1000), NANOG (SAB, 1:1000), STAT3 (SAB, 1:1000), pSTAT3 (SAB, 1:1000), MYD88 (R&D Systems, 1:1000), β-actin (Santa Cruz Biotechnology, 1:2000). Species-specific horseradish peroxidase (HRP)-conjugated anti-rabbit IgG (Jackson ImmunoResearch) and HRP-conjugated anti-mouse IgG (Jackson ImmunoResearch) were used as secondary antibodies.