银杏总黄酮对IL-1β诱导的髓核细胞凋亡及Nrf2/ARE信号通路的影响

doi:10.3969/j.issn.1006-6233.2021.10.001

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Abstract Objective: To investigate the effect of total flavonoids of Ginkgo biloba (TFG) on apoptosis of nucleus pulposus cells induced by interleukin-1β (IL-1β), and its regulation on nuclear factor E2 related factor 2 (Nrf2)/antioxidant response element (ARE) signaling pathway. Methods: Human nucleus pulposus cells were divided into normal group, IL-1β induced group, dimethyl sulfoxide (DMSO) group and TFG groups with different concentrations (5, 10, 20, 40, 80, 160 mg/mL), in addition to the normal group, all groups were induced with IL-1β (10 ng/mL) for 24 hours, and CCK-8 method was used to detect the cell proliferation activity. The nucleus pulposus cells were divided into normal control group, IL-1β induced group, TFG low (20 mg/mL), high (80 mg/mL) dose groups, Nrf2 pathway inhibitor group (Brusatol group, 0.2 μg/mL) group, TFG + Brusatol group (80 mg/mL + 0.2 μg/mL). CCK-8 assay was used to detect cell viability; hochest 33258 staining was used to detect apoptosis; immunofluorescence was used to detect the content of reactive oxygen species (ROS); detection of Nrf2 entry by immunofluorescence cofocal method; Western blot was used to detect the expression level of Nrf2, ARE, cleaved caspase-3, tumor necrosis factor-α (TNF-α), heme oxygenase 1 (HO-1) protein. Results: Compared with that in the normal group, the proliferation activity of IL-1β induced group was lower (P<0.05). Compared with the IL-1β induced group, the proliferation activity increased with the increase of TFG concentration, it was stable at 80 mg/mL-160 mg/mL, and 20 mg/mL and 80 mg/mL were selected as TFG low and high dose groups for follow-up test. Compared with the normal control group, the cell proliferation activity of IL-1β induced group was decreased (P<0.05), and the apoptosis rate, ROS content, Nrf2 number, Nrf2, ARE, cleaved caspase-3, HO-1 and TNF-α protein expression levels were increased (P<0.05). Compared with IL-1β induced group, cell proliferation activity, Nrf2 number, Nrf2, ARE and HO-1 protein expression levels of TFG low dose and high dose groups were increased (P<0.05), cell apoptosis rate, ROS content, cleaved caspase-3 and TNF-α protein expression levels were decreased (P<0.05); and the cell proliferation activity, Nrf2 number, Nrf2, ARE and HO-1 protein expression levels in TFG high dose group were higher than those in TFG low dose group (P<0.05), and the apoptosis rate, ROS content, cleaved caspase-3 and TNF-α protein expression levels in TFG high dose group were lower than those in TFG low dose group (P<0.05); the cell proliferation activity, Nrf2 number, Nrf2, ARE and HO-1 protein expression levels of Brusatol group were decreased (P<0.05), while the apoptosis rate, ROS content, cleaved caspase-3 and TNF-α protein expression levels were increased (P<0.05). Compared with the TFG high dose group, the cell proliferation activity, Nrf2 number, Nrf2, ARE and HO-1 protein expression levels of TFG + Brusatol group were decreased (P<0.05), and the apoptosis rate, ROS content, cleaved caspase-3 and TNF-α protein expression levels were increased (P<0.05). Conclusion: TFG can play an anti-inflammatory, anti-oxidation and anti-apoptosis of nucleus pulposus cells effect by activating Nrf2/ARE pathway.

Key words： Total flavonoids of Ginkgo biloba Nucleus pulposus cells Nuclear factor E2 related factor 2/antioxidant response element signaling pathway Interleukin-1β Apoptosis

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http://www.hbyxzzs.cn/EN/10.3969/j.issn.1006-6233.2021.10.001 OR http://www.hbyxzzs.cn/EN/Y2021/V27/I10/1585

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