Effect of astragaloside Ⅳ on SK ⁃ N ⁃ SH cells damage induced by 1 ⁃ methyl ⁃ 4 ⁃ phenylpyridine in Parkinson's disease

Abstract

Objective To investigate the effect of astragaloside Ⅳ on the injury of SK⁃N⁃SH cell in Parkinson's disease (PD) induced by 1 ⁃ methy ⁃ 4 ⁃ phenylpyridine (MPP＋) and its mechanism. Methods Human neuroblastoma cell line SK ⁃N ⁃SH was cultured to logarithmic growth stage, which were randomized to routine culture (control group), MPP＋ induction (MPP＋ group), astragaloside Ⅳ 10 mg/ml + MPP＋ induction (astragaloside Ⅳ 10 mg/ml group), astragaloside Ⅳ 30 mg/ml + MPP＋ induction (astragaloside Ⅳ 30 mg/ml group), astragaloside Ⅳ 30 mg/ml + MPP＋ induction + Janus kinase 2 (JAK2) inhibitor AG490 (JAK2 inhibitor group). Cell survival was detected by methyl thiazolyl tetrazolium (MTT) assay, cell apoptosis was detected by flow cytometry, reactive oxygen species (ROS) level was detected by DCFH ⁃ DA fluorescence probe, lactate dehydrogenase (LDH) and superoxide dismutase (SOD) activities were detected by enzyme method. And the relative expression levels of phosphorylated Janus kinase 2 (pJAK2) and phosphorylated signal transducer and activator of transcription 3 (pSTAT3) proteins in JAK2 ⁃ STAT3 signal transducer pathway were detected by Western blotting. Results There were significant differences in cell survival rate (P = 0.000), apoptosis rate (P = 0.000), ROS content (P = 0.000), LDH activity (P = 0.000), SOD activity (P = 0.003), pJAK2 (P = 0.000) and pSTAT3 (P = 0.000) proteins relative expression levels among different SK ⁃ N ⁃ SH groups. Further pairwise comparison showed that compared with the control group, the cell survival rate (P = 0.000, 0.001, 0.049, 0.000), SOD activity (P = 0.000, 0.002, 0.012, 0.000), relative expression levels of pJAK2 (P = 0.003, 0.006, 0.036, 0.002) and pSTAT3 (P = 0.001, 0.002, 0.024, 0.001) proteins were decreased in MPP＋ group, astragaloside Ⅳ 10 mg/ml and 30 mg/ml groups, JAK2 inhibitor group, but the cell apoptosis rate (P = 0.001, 0.001, 0.001, 0.000), ROS content (P = 0.000, 0.001, 0.002, 0.000) and LDH activity (P = 0.000, 0.002, 0.038, 0.000) were increased. Compared with MPP＋ group, the survival rate (P = 0.016, 0.000), SOD activity (P = 0.003, 0.001), relative expression of pJAK2 (P = 0.013, 0.002) and pSTAT3 (P = 0.018, 0.002) proteins increased in astragaloside Ⅳ 10 mg/ml and 30 mg/ml groups, which the apoptosis rate (P = 0.021, 0.008), ROS content (P = 0.031, 0.003) and LDH activity (P = 0.001, 0.000) decreased. Compared with astragaloside Ⅳ 10 mg/ml group, the cell survival rate (P = 0.002), SOD activity (P = 0.027), relative expression of pJAK2 (P = 0.007) and pSTAT3 (P = 0.006) proteins were increased in astragaloside Ⅳ 30 mg/ml group, but ROS content (P = 0.019) and LDH activity (P = 0.011) decreased, while apoptosis rate (P = 0.016), ROS content (P = 0.030) and LDH activity (P = 0.004) increased in JAK2 inhibitor group, and SOD activity (P = 0.004), the relative expression of pJAK2 (P = 0.001) and pSTAT3 (P = 0.005) proteins were decreased. Compared with astragaloside Ⅳ 30 mg/ml group, cell survival rate (P = 0.001), SOD activity (P = 0.001), the relative expression of pJAK2 (P = 0.000) and pSTAT3 (P = 0.001) proteins in JAK2 inhibitor group were decreased, but the apoptosis rate (P = 0.004), ROS content (P = 0.002) and LDH activity (P = 0.001) were increased. Conclusions Astragaloside Ⅳ can reduce MPP＋ induced SK ⁃ N ⁃ SH cell injury in PD, which may be related to the activation of JAK2 ⁃ STAT3 signal transducer pathway.

Keywords

• parkinson disease
• astragalus membranaceus
• janus kinase 2
• activating transcription factor 3
• tumor cells
• cultured