Loading buffer 5

Whey protein isolate (WPI) was bought from Fonterra Co. Ltd. (Auckland, New Zealand) and the protein content was 93% as stated by the manufacturer. Pea protein isolate (PPI, ≥80%) was purchased from Yuanye Bio-tech Co. Ltd. (Shanghai, China) . The SDS-PAGE gel preparation kit, loading buffer (5×), and Coomassie blue fast staining solution were obtained from Beyotime Institute of Biotechnology (Nanjing, China). Nile blue was obtained from Aladdin (Shanghai, China). The 1-Anilinonaphthalene-8-sulfonic acid (ANS) was purchased from Solarbio (Beijing, China). Soybean oil was purchased from a local market (Changchun, China). All other chemicals were bought from Beijing Chemical Works (Beijing, China).

Whey protein isolate with a protein purity of 93.14% was obtained from Fonterra Co-operative Group. Cannabidiol (purity of 99%) was purchased from Macklin Biochemical Co. Ltd. Maltodextrin (corn starch origin) with a dextrose equivalent of 16 to 20% was purchased from Kemai Biochemistry. The SDS-PAGE gel preparation kit, loading buffer (5×), and Coomassie blue fast staining solution were purchased from Beyotime Institute of Biotechnology. Ortho-phthalaldehyde (OPA) and 1-anilinonaphthalene-8-sulfonic acid (ANS) were purchased from Solarbio. Medium-chain triglyceride (MCT) was purchased from Danisco Foods International Inc.

The isolated spinal cords were homogenated with RIPA buffer (Beyotime Biotechnology, Shanghai, China) containing 1% protease inhibitor (Beyotime Biotechnology, Shanghai, China) for 30 minutes on ice. The homogenates were centrifuged at 12000 g for 25 minutes at 4°C and supernatants were collected. NanoDrop 2000C (Thermo Scienti c, USA) was then used for protein concentrations determination. The homogenates and loading buffer 5× (Beyotime Biotechnology, Shanghai, China) were mixed at a ratio of 4: 1, denatured in the metal bath (100°C, 8~10 mins), cooled to room temperature and loaded. Protein samples were subjected to 12.5% SDS-PAGE (Epizyme Biotech, Shanghai, China) and then transferred to polyvinylidene uoride (PVDF) membranes. Membranes were blocked in protein free rapid blocking buffer (Epizyme Biotech, Shanghai, China) for 15 minutes at room temperature. After blocking, membranes were incubated with the primary antibodies against ATF3 (DF6660, A nity Biosciences), XBP1 (WL00708, WanleiBio), HMOX1 (WL02400, WanleiBio), DDIT3 (GADD153, Proteintech), CHAC1 (DF9353, A nity Biosciences) and GAPDH (AF7021, A nity Biosciences) overnight at 4°C. Membranes were incubated with the secondary antibody at room temperature for 1.5 h. Protein bands were captured using an ECL chemiluminescence system (Epizyme Biotech, Shanghai, China).