Anandamide Protects HT22 Cells Exposed to Hydrogen Peroxide by Inhibiting CB1 Receptor-Mediated Type 2 NADPH Oxidase

Figure 9

Nox2 expression was inhibited in the presence of AEA via CB1. (a) Nox2 expression was increased in HT22 cells exposed to H₂O₂ in a time-dependent manner. Then the cells were divided into five groups, Control: cells cultured in drug-free medium; H₂O₂: cells exposed to 200 μM H₂O₂ for 3 h; AEA + H₂O₂: cells exposed to 10 μM AEA plus 200 μM H₂O₂ for 3 h; AM251 + AEA + H₂O₂: cells exposed to 10 μM AEA plus 10 μM CB1 antagonist AM251 in the presence of 200 μM H₂O₂ for 3 h; AM251 + H₂O₂: cells exposed to 10 μM AM251 plus 200 μM H₂O₂ for 3 h. Nox2 protein expression (b) and mRNA transcription (c) were evaluated by western blotting and real-time PCR, respectively. (d) Incubation with CB1-siRNA for 5 h abolished the AEA-induced inhibition of Nox2 mRNA transcription. Results are expressed as means ± SD (𝑛=4). ^∗𝑃<0.05 versus the control (no H₂O₂, no AEA, and no AM251 or siRNA), ^#𝑃<0.05 versus the cells exposed to H₂O₂ alone, and ^∧𝑃<0.05 versus the cells exposed to AEA plus H₂O₂.

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