Anti sdf 1α

The SDF-1α, Arx, Pax4, Insulin and Glucagon expression were assessed by western blotting analysis and samples were normalized to GAPDH. Protein extraction was blocked with PBS-5% fat-free dried milk at room temperature for 1 h and incubated at 4°C overnight with anti-SDF-1α (1:1000, Santa cruz), anti-Arx (1:1000, Santa cruz), anti-Pax4 (1:1000, Santa Cruz), anti-Insulin (1:1000, Santa cruz), anti-Glucagon (1:1000, Santa cruz), anti-Aldh1a3 (1:1000, Novus), anti-Neurog3 (1:1000, Beta Cell Biology Consortium), anti-MafA (1:1000, Cell Signaling Technology), anti-Pdx1 (1:1000, Cell Signaling Technology), anti-NeuroD1 (1:1000, Cell Signaling Technology) and anti-XBP1 (1:1000, Santa cruz), anti-CD63 (1:2000, Abcam), anti-TSG101 (1:1000, Santa Cruz), anti-Ago2 (1: 1000, Santa Cruz) and anti-GAPDH (1:3000, Santa Cruz) antibodies respectively.

The H9c2 cells or heart tissues were lysed in RIPA buffer, and 30 µg of proteins were separated in a 12 % SDS–PAGE gel and transferred onto a nitrocellulose membrane (Millipore). The membrane was rinsed with Tris-buffered saline (TBS) containing 0.1 % Tween-20 (TBST) and blocked with 5 % fat free milk in TBS at room temperature for 1 h. The membrane was then incubated with anti-Akt, anti-phospho-Akt, anti-Bcl-2, anti-Bax, anti-beclin1, anti-p38MAPK, anti-ERK1/2, anti-pp38MAPK, anti-pERK1/2 (Cell Signaling Technology, 1:500), anti-SDF-1α, anti-HGF (Santa Cruz) or anti-α-Tubulin antibody (Sigma, 1:5000) and then incubated with horseradish peroxidase-conjugated secondary antibodies (1:10,000 dilution; Santa Cruz). The immunoblots were detected using enhanced chemiluminescence reaction (Amersham Pharmacia Biotech) and measured with densitometry.