Becline 1

Propranolol was obtained from the Chinese materials research center (Beijing, China). Fetal bovine serum (FBS) was got from Gibco, Gaithersburg, MD, USA). N-Acetyl-L-cysteine (NAC), 3-(4, 5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT), Hoechst 33258 staining, and Annexin V-PE Apoptosis Detection Kit were purchased from Beyotime Institute of Biotechnology (Shanghai, China). The enhanced chemiluminescence (ECL) were purchased from Beyotime Institute of Biotechnology (Shanghai, China). JNK inhibitor (SP600125) and 3-MA (3-Methyladenine) were obteined from sigma (St. Louis, MI, USA). Antibodies against Cyclin B1, phospho-cdc2, cdc2, JNK, phosphorylated JNK, and p21 were obtained from Cell Signaling Technology (Beverly, MA, USA). Antibodies against Cdk1, Bcl-2, cleavage caspase-3, cleavage caspase -8, cleavage caspase -9, BAX, LC3-I, LC3- II, Becline-1, p62, GAPDH were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA). All other common chemicals and buffers were from Boster (Wuhan, China).

The lumbar enlargement (L4-L6) regions of the spinal cord from WT and TLR4 KO mice were dissected and homogenized in lysis buffer. The lysates of the spinal cord (20 μg) were separated by 12 or 15% sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and transferred to nitrocellulose membranes. The blots were probed with the following primary antibodies: Becline 1 (1:1000; #sc-11,427, Santa Cruz Biotechnology, Santa Cruz, CA, USA), LC3 (1:1000; #L8918, Sigma-Aldrich), p62 (1:1000; #P0067, Sigma-Aldrich), PINK1 (1:1000; #NBP1–39667, Novus Biologicals, Littleton, CO, USA), and β-actin (1:500; #2965, Cell Signaling Technology, Danvers, MA, USA). The immune complexes were identified using an enhanced chemiluminescence (ECL) detection system (Habersham, Little Chalfont, United Kingdom).