Notoginsenoside r1

Modeling: All mice were injected intraperitoneally with a solution of urethane (China National Chemical Reagent Corporation, Ltd.) configurated by double distilled water (600 mg/kg), once a week for 10 weeks.
Administration: administration of intragastric administration was started on the day of modeling. The drugs were all dissolved with 0.5% CMC-Na. The shikonin group was given a dose of 2 mg/kg of shikonin (Sigma-Aldrich Company, USA), and the aconitine group was given a dose of 0.2 mg/kg of aconitine (Sigma-Aldrich Company, USA), and the notoginsenoside R1 group was administered with a dose of 20 mg/kg of notoginsenoside R1 (Sigma-Aldrich Company, USA), and the compound group was given a dose of compound of the three: 2 mg/kg of shikonin, 0.2 mg/kg of aconitine and 20 mg/kg. The control group was given an equal dose of 0.5% CMC-Na. Once a day for 20 weeks. The autonomic activity and mortality of the mice were continuously recorded weekly. The score of mouse's autonomic activity was measured by YLS-1A multifunctional mouse autonomic activity recorder (Anhui Huaibei Zhenghua biological Instrument Co., Ltd., China). The higher the score was, the more autonomic activity the mouse had.

Notoginsenoside R1 was acquired from Sigma Aldrich (St. Louis, USA). We used a High-Capacity cDNA Reverse Transcription Kit (Thermo-Fisher, USA) and a Master Mix Rotor-Gene SYBR^® Green PCR Kit (Qiagen, USA). Oxidative marker enzymes and antioxidant enzymes were obtained from Cayman Chemicals (USA). Primary antibodies for iNOS and COX-2, HRP conjugated secondary antibody were obtained from Santa Cruz Biotechnology, Inc. (USA). ELISA assay kits for Cystatin-C, NGAL, β2-microglobulin, NAG, IL-18, kim-1, TNF-α, IL-2, IL-17, IL-8, IL-6, and IL-1β were obtained from Fine Biotech, China. All other chemicals used were of reagent grade.