E el m0006c

Mouse TNF-α (E-EL-M0049c), MCP-1 (E-EL-M0006c), IL-6 (E-EL-M0044c) and IL-10 (E-EL-M0045c, Elabscience, Wuhan) enzyme-linked immunosorbent assay (ELISA) kits were used to measure the concentrations of TNF-α, MCP-1, IL-6, and IL-10 in the supernatant according to the manufacturer’s instructions.

Serum CCL-2 was measured by a mouse ELISA kit (Cat. No. E-EL-M0006c; Elabscience Biotechnology Co., Ltd, China). Glutamic oxaloacetic transaminase (GOT) (Cat. No. c010-1) and glutamic pyruvic transaminase (GPT) (Cat. No. c009-1) were determined using assay kits (Nanjing Jiancheng Biology Engineering Institute, China). In brief, 5 μL serum and 25 μL GOT or GPT substrates were added to 96-well plates at 37°C for 30 min. Then, 2,4-dinitrophenylhydrazine (25 μL) was added to all wells at 37°C for 30 min. Finally, sodium hydroxide (0.4 mol/L; 250 μL) was added to stop the reactions at room temperature for 15 min, and the absorbance was measured at 510 nm. Serum creatinine (Cre; Cat. No. C011-1; Nanjing Jiancheng Biology Engineering Institute, China) and blood urea nitrogen (BUN; Cat. No. C013-2; Nanjing Jiancheng Biology Engineering Institute) levels were measured using an autoanalyzer and an enzymatic kinetic method using commercial kits following the manufacturer’s protocol with a SpectraMax M5 ELISA plate reader (Molecular Devices, LLC, USA).