Zinc ion mediates neuroprotective effect of astragaloside IV on endoplasmic reticulum stress-induced PC12 cells
10.3760/cma.j.cn.115354-20230303-00119
- VernacularTitle:黄芪甲苷通过锌离子抑制内质网应激发挥PC12细胞神经保护作用
- Author:
Xinyi ZHAO
1
;
Ying YANG
;
Luyao HUANG
;
Jinkun XI
;
Yonggui HE
Author Information
1. 华北理工大学附属医院神经内科，唐山 063000
- Keywords:
Astragaloside IV;
Cerebral ischemia-reperfusion injury;
Zinc ion;
Apoptosis
- From:
Chinese Journal of Neuromedicine
2023;22(6):566-574
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To investigate whether astragaloside IV (AS-IV) exerts neuroprotective effects via zinc ion (Zn 2+) modulation of mitochondria-associated endoplasmic reticulum membranes (MAMs) and to elucidate the possible mechanisms. Methods:PC12 cells (rat adrenal pheochromocytoma cells) were routinely cultured; they were divided into 7 groups: control group (routinely cultured), 2-DG group (treated with 2-DG at 50 μmol/L for 30 min), astragaloside IV+2-DG group (treated with 50 μmol/L astragaloside IV for 20 min, and then treated with 50 μmol/L 2-DG for 30 min), astragaloside IV group (treated with 50 μmol/L astragaloside IV for 20 min), N,N,N',N'-tetrakis(2-pyridylmethyl)ethylenediamine (a zinc chelator, TPEN)+astragaloside IV+2-DG group (treated with 10 μmol/L TPEN for 10 min, 50 μmol/L astragaloside IV for 20 min, and then 50 μmol/L 2-DG for 30 min); TPEN group (treated with 10 μmol/L TPEN for 10 min), TPEN+2-DG group (treated with 10 μmol/L TPEN for 10 min and then treated with 50 μmol/L 2-DG for 30 min). The expressions of glucose-regulated protein (GRP)78, GRP94, cysteine-containing aspartate proteolytic enzyme caspases-8, B cell receptor associated protein 31 (BAP31) and mitochondrial fission protein 1 (Fis1) were detected by Western blotting. Annexin V-FITC/PI kit was used to detect the apoptosis level. Immunofluorescence was used to detect the Fis1 protein expression, and mitochondrial fluorescence probe TMRE was used to detect the opening of mitochondrial permeability transition pore (mPTP).Results:(1) Compared with the control group, the 2-DG group had significantly increased expressions of GRP78, GRP94, Caspase-8, Bap31 and Fis1, and green fluorescent intensity of Annexin V-FITC ( P<0.05); compared with the 2-DG group, the astragaloside IV+2-DG group had significantly decreased expressions of GRP78, GRP94, Caspase-8, Bap31 and Fis1, and green fluorescent intensity of Annexin V-FITC ( P<0.05); compared with the astragaloside IV+2-DG group, the TPEN+astragaloside IV+2-DG group had significantly increased expressions of GRP78, GRP94, Caspase-8, Bap31 and Fis1, and green fluorescent intensity of Annexin V-FITC ( P<0.05). (2) Compared with the control group, the 2-DG group had significantly enhanced Fis1 protein fluorescent intensity ( P<0.05); the astragaloside IV+2-DG group had significantly decreased Fis1 protein fluorescent intensity compared with 2-DG group ( P<0.05); Compared with astragaloside IV+2-DG group, TPEN+astragaloside IV+2-DG group had significantly enhanced Fis1 protein fluorescence intensity ( P<0.05); compared with the control group, the TPEN+2-DG group had significantly enhanced Fis1 protein fluorescent intensity ( P<0.05). (3) TMRE fluorescence intensity in 2-DG group was significantly decreased compared with control group ( P<0.05); TMRE fluorescence intensity in astragaloside IV+2-DG group was significantly enhanced compared with 2-DG group ( P<0.05); TMRE fluorescence intensity in TPEN+astragaloside IV+2-DG group was significantly decreased compared with astragaloside IV+2-DG group (P<0.05); TMRE fluorescence intensity in TPEN+2-DG group was significantly decreased compared with the control group ( P<0.05). Conclusion:Astragaloside IV can exert neuroprotective effects through Zn 2+ inhibiting endoplasmic reticulum stress-induced apoptosis and preventing mPTP opening in PC12 cells, whose mechanism may be related to Fis1 and Bap31 expressions.
