To investigate the effect of wogonin on hypoxia/reoxygenation (H/R)-induced neuronal injury based on the C-C chemokine ligand 2-C-C chemokine receptor 2 (CCL2-CCR2) pathway.

PC12 cells were cultured and divided into five groups, control group (normal culture), model group (H/R injury), experimental group (H/R+50 μmol·L^-1 wogonin pretreatment for 1 h), antagonist group (H/R+10 μmol·L^-1 CCR2 antagonist RS102895 pretreatment for 1 h), and agonist group (H/R+50 μmol·L^-1 wogonin+20 μmol·L^-1 CCL2 recombinant protein pretreatment for 1 h). Cell viability and cytotoxicity were assessed using the cell counting kit-8 (CCK-8) and lactate dehydrogenase (LDH) assay kits. Apoptosis was detected by Annexin V-fluorescein isothiocyanate/propidium iodide (Annexin V-FITC/PI) double staining and terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) staining. Oxidative stress and inflammatory responses were evaluated by enzyme-linked immunosorbent assay (ELISA) and quantitative real-time polymerase chain reaction (RT-qPCR). Protein expression levels related to the CCL2-CCR2 signaling pathway were measured by Western blot.

The viability of PC12 cells in control group, model group, experimental group, antagonist group and agonist group were 0.89±0.06, 0.32±0.04, 0.69±0.05, 0.65±0.07 and 0.46±0.04, respectively; the SOD activities were (45.11±3.02), (10.05±1.05), (33.62±3.15), (30.15±2.87) and (21.05±2.17) U·mL^-1, respectively; the Annexin V FITC/PI double staining apoptosis rates were (3.21±0.68)%, (29.65±1.59)%, (8.99±0.82)%, (9.87±0.85)% and (18.65±1.06)%, respectively; the TUNEL positive rates were (5.66±0.89)%, (33.65±2.42)%, (13.25±1.05)%, (11.66±1.87)% and (24.06±1.16)%, respectively; the relative expression levels of tumor necrosis factor-alpha(TNF-α) mRNA were 1.02±0.15, 3.05±0.32, 1.59±0.16, 2.98±0.24 and 2.66±0.20, respectively; the relative expression levels of interleukin-6 (IL-6) mRNA were 1.05±0.12, 4.54±0.39, 1.87±0.15, 1.75±0.14 and 3.08±0.32, respectively; the relative expression levels of CCL2 mRNA were 1.01±0.07, 3.99±0.32, 1.64±0.15, 3.90±0.33 and 2.84±0.23, respectively; the relative expression levels of CCR2 mRNA were 1.03±0.10, 5.21±0.51, 2.03±0.26, 2.11±0.20 and 3.85±0.32, respectively; the relative expression levels of phosphorylation (p)p65/p65 were 0.12±0.02, 0.75±0.06, 0.24±0.03, 0.22±0.01 and 0.52±0.05, respectively; the leakage rates of LDH were (18.32±1.15)%, (58.64±5.16)%, (29.02±2.16)%, (26.98±3.02)% and (40.24±3.54)%, respectively; the malondialdehyde (MDA) contents were (50.16±5.24), (169.65±10.16), (91.32±7.12), (86.21±5.16) and (134.16±11.87) nmol·mL^-1, respectively. The differences of above indicators between model group and control group, between experimental group and model group, and between the antagonist, agonist group and experimental group were all statistically significant (all P<0.05).

Wogonin exerts a protective effect against H/R-induced neuronal injury by inhibiting the CCL2-CCR2 signaling pathway.