Antioxidant activity test

The antioxidant activity of MSC-EVs on H₂O₂-stimulated primary cultures of hippocampal neurons was tested according to the procedures described in our recent study ²¹. The cell supernatant in each group (n = 6~9 samples per group) was collected by centrifugation at 12000 g for 5 min, and the samples and reaction reagents, including those from the ferric ion reducing antioxidant power (FRAP) (A015-3, Nanjing Jiancheng Bioengineering Institute, China), catalase (CAT) (S0051, Beyotime, China), glutathione peroxidase (GSH-PX) (A005, Nanjing Jiancheng Bioengineering Institute, China) and superoxide dismutase (SOD) (A001-3, Nanjing Jiancheng Bioengineering Institute, China) kits, were added to 96-well flat-bottom plates. The antioxidant effect or enzyme activity was detected using a microplate reader (Bio-Rad Laboratories Inc., Hercules, CA, USA) per the manufacturer's instructions. Each test was replicated 3 times.