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Protective effects of astragaloside IV against amyloid beta1-42 neurotoxicity by inhibiting the mitochondrial permeability transition pore opening.

Sun Q, Jia N, Wang W, Jin H, Xu J, Hu H - PLoS ONE (2014)

Bottom Line: The results showed that pretreatment of AS-IV significantly increased the viability of neuronal cells, reduced apoptosis, decreased the generation of intracellular reactive oxygen species (ROS) and decreased mitochondrial superoxide in the presence of Aβ1-42.Moreover, pretreatment of AS-IV reduced the expression of Bax and cleaved caspase-3 and increased the expression of Bcl-2 in an Aβ1-42 rich environment.These results provide novel insights of AS-IV for the prevention and treatment of neurodegenerative disorders such as AD.

View Article: PubMed Central - PubMed

Affiliation: Department of Human Anatomy and Histo-Embryology, Xi'an Jiaotong University Health Science Center, Xi'an, Shaanxi, China.

ABSTRACT
Mitochondrial dysfunction caused by amyloid β-peptide (Aβ) plays an important role in the pathogenesis of Alzheimer disease (AD). Substantial evidence has indicated that the mitochondrial permeability transition pore (mPTP) opening is involved in Aβ-induced neuronal death and reactive oxygen species (ROS) generation. Astragaloside IV (AS-IV), one of the major active constituents of Astragalus membranaceus, has been reported as an effective anti-oxidant for treating neurodegenerative diseases. However, the molecular mechanisms still need to be clarified. In this study, we investigated whether AS-IV could prevent Aβ1-42-induced neurotoxicity in SK-N-SH cells via inhibiting the mPTP opening. The results showed that pretreatment of AS-IV significantly increased the viability of neuronal cells, reduced apoptosis, decreased the generation of intracellular reactive oxygen species (ROS) and decreased mitochondrial superoxide in the presence of Aβ1-42. In addition, pretreatment of AS-IV inhibited the mPTP opening, rescued mitochondrial membrane potential (ΔΨm), enhanced ATP generation, improved the activity of cytochrome c oxidase and blocked cytochrome c release from mitochondria in Aβ1-42 rich milieu. Moreover, pretreatment of AS-IV reduced the expression of Bax and cleaved caspase-3 and increased the expression of Bcl-2 in an Aβ1-42 rich environment. These data indicate that AS-IV prevents Aβ1-42-induced SK-N-SH cell apoptosis via inhibiting the mPTP opening and ROS generation. These results provide novel insights of AS-IV for the prevention and treatment of neurodegenerative disorders such as AD.

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AS-IV attenuated Aβ1-42-induced mitochondrial dysfunction.A. A representative image of mitochondrial membrane potential detection by TMRM staining (red fluorescence). Mitochondria were counterstained with Mitotracker Green (green fluorescence). (a) vehicle; (b) 50 µM AS-IV; (c) 5 µM Aβ1-42; (d, e, f) 10, 25, 50 µM AS-IV+ 5 µM Aβ1-42, respectively. B. Quantification analysis of ΔΨm. #P<0.01 vs vehicle; *P<0.01 vs Aβ1-42 (n = 6). C. ATP detection. D. Cytochrome c oxidase activity detection. Aβ1-42 significantly decreased both CcO activity and ATP which was reversed by pretreatment with AS-IV (25, 50 µM). #P<0.01 vs vehicle; *P<0.01 vs Aβ1-42 (n = 6). Scale bar = 10 µm.
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pone-0098866-g003: AS-IV attenuated Aβ1-42-induced mitochondrial dysfunction.A. A representative image of mitochondrial membrane potential detection by TMRM staining (red fluorescence). Mitochondria were counterstained with Mitotracker Green (green fluorescence). (a) vehicle; (b) 50 µM AS-IV; (c) 5 µM Aβ1-42; (d, e, f) 10, 25, 50 µM AS-IV+ 5 µM Aβ1-42, respectively. B. Quantification analysis of ΔΨm. #P<0.01 vs vehicle; *P<0.01 vs Aβ1-42 (n = 6). C. ATP detection. D. Cytochrome c oxidase activity detection. Aβ1-42 significantly decreased both CcO activity and ATP which was reversed by pretreatment with AS-IV (25, 50 µM). #P<0.01 vs vehicle; *P<0.01 vs Aβ1-42 (n = 6). Scale bar = 10 µm.

Mentions: To explore the potential role of AS-IV in Aβ1-42-induced neuronal cell death, we examined the mitochondrial function by testing mitochondrial membrane potential (ΔΨm), ATP level and cytochrome c oxidase (CcO) in SK-N-SH cells. Firstly, we employed TMRM as an indicator of mitochondrial membrane potential (ΔΨm). Aβ1-42 treated SK-N-SH cells showed a significant decrease in red fluorescence intensity compared with cells from the vehicle group (P<0.01). Cells pretreated with 25 or 50 µM AS-IV at showed higher red fluorescence intensity compared with cells treated with Aβ1-42 alone for 24h (P<0.01). There was no significant difference between the 10 µM AS-IV pretreatment group and the Aβ1-42 group (P<0.01) (Fig. 3A, B). Secondly, we measured ATP level. As shown in Fig. 3C, ATP generation in the Aβ1-42 group was decreased compared with the vehicle group. In the presence of AS-IV at 25 or 50 µM, ATP level was significantly increased compared with that in the Aβ1-42 group (P<0.01). Application of 10 µM AS-IV did not recover ATP level (P>0.05). Thirdly, the CcO activity was examined. Cells treated with 5 µM Aβ1-42 demonstrated significantly decreased CcO activity compared with that in the vehicle group (P<0.01). Notably, cells in the presence of AS-IV exhibited significantly increased CcO activity in a dose-dependent manner compared with cells treated with Aβ1-42 alone for 24 h (P<0.01). Pretreatment of 25 and 50 µM AS-IV significantly increased CcO activity in a dose-dependent manner. AS-IV at 10 µM did not show significant difference compared with that in the Aβ1-42 group (P>0.05) (Fig. 3D). Statistical analysis showed that the mitochondrial function of cells pretreated with AS-IV did not recovery fully to the level of the vehicle group. In the experiments to detect ΔΨm, ATP level or CcO activity, 50 µM AS-IV alone treatment did not show an insult to SK-N-SH cells.


Protective effects of astragaloside IV against amyloid beta1-42 neurotoxicity by inhibiting the mitochondrial permeability transition pore opening.

Sun Q, Jia N, Wang W, Jin H, Xu J, Hu H - PLoS ONE (2014)

AS-IV attenuated Aβ1-42-induced mitochondrial dysfunction.A. A representative image of mitochondrial membrane potential detection by TMRM staining (red fluorescence). Mitochondria were counterstained with Mitotracker Green (green fluorescence). (a) vehicle; (b) 50 µM AS-IV; (c) 5 µM Aβ1-42; (d, e, f) 10, 25, 50 µM AS-IV+ 5 µM Aβ1-42, respectively. B. Quantification analysis of ΔΨm. #P<0.01 vs vehicle; *P<0.01 vs Aβ1-42 (n = 6). C. ATP detection. D. Cytochrome c oxidase activity detection. Aβ1-42 significantly decreased both CcO activity and ATP which was reversed by pretreatment with AS-IV (25, 50 µM). #P<0.01 vs vehicle; *P<0.01 vs Aβ1-42 (n = 6). Scale bar = 10 µm.
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Related In: Results  -  Collection

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pone-0098866-g003: AS-IV attenuated Aβ1-42-induced mitochondrial dysfunction.A. A representative image of mitochondrial membrane potential detection by TMRM staining (red fluorescence). Mitochondria were counterstained with Mitotracker Green (green fluorescence). (a) vehicle; (b) 50 µM AS-IV; (c) 5 µM Aβ1-42; (d, e, f) 10, 25, 50 µM AS-IV+ 5 µM Aβ1-42, respectively. B. Quantification analysis of ΔΨm. #P<0.01 vs vehicle; *P<0.01 vs Aβ1-42 (n = 6). C. ATP detection. D. Cytochrome c oxidase activity detection. Aβ1-42 significantly decreased both CcO activity and ATP which was reversed by pretreatment with AS-IV (25, 50 µM). #P<0.01 vs vehicle; *P<0.01 vs Aβ1-42 (n = 6). Scale bar = 10 µm.
Mentions: To explore the potential role of AS-IV in Aβ1-42-induced neuronal cell death, we examined the mitochondrial function by testing mitochondrial membrane potential (ΔΨm), ATP level and cytochrome c oxidase (CcO) in SK-N-SH cells. Firstly, we employed TMRM as an indicator of mitochondrial membrane potential (ΔΨm). Aβ1-42 treated SK-N-SH cells showed a significant decrease in red fluorescence intensity compared with cells from the vehicle group (P<0.01). Cells pretreated with 25 or 50 µM AS-IV at showed higher red fluorescence intensity compared with cells treated with Aβ1-42 alone for 24h (P<0.01). There was no significant difference between the 10 µM AS-IV pretreatment group and the Aβ1-42 group (P<0.01) (Fig. 3A, B). Secondly, we measured ATP level. As shown in Fig. 3C, ATP generation in the Aβ1-42 group was decreased compared with the vehicle group. In the presence of AS-IV at 25 or 50 µM, ATP level was significantly increased compared with that in the Aβ1-42 group (P<0.01). Application of 10 µM AS-IV did not recover ATP level (P>0.05). Thirdly, the CcO activity was examined. Cells treated with 5 µM Aβ1-42 demonstrated significantly decreased CcO activity compared with that in the vehicle group (P<0.01). Notably, cells in the presence of AS-IV exhibited significantly increased CcO activity in a dose-dependent manner compared with cells treated with Aβ1-42 alone for 24 h (P<0.01). Pretreatment of 25 and 50 µM AS-IV significantly increased CcO activity in a dose-dependent manner. AS-IV at 10 µM did not show significant difference compared with that in the Aβ1-42 group (P>0.05) (Fig. 3D). Statistical analysis showed that the mitochondrial function of cells pretreated with AS-IV did not recovery fully to the level of the vehicle group. In the experiments to detect ΔΨm, ATP level or CcO activity, 50 µM AS-IV alone treatment did not show an insult to SK-N-SH cells.

Bottom Line: The results showed that pretreatment of AS-IV significantly increased the viability of neuronal cells, reduced apoptosis, decreased the generation of intracellular reactive oxygen species (ROS) and decreased mitochondrial superoxide in the presence of Aβ1-42.Moreover, pretreatment of AS-IV reduced the expression of Bax and cleaved caspase-3 and increased the expression of Bcl-2 in an Aβ1-42 rich environment.These results provide novel insights of AS-IV for the prevention and treatment of neurodegenerative disorders such as AD.

View Article: PubMed Central - PubMed

Affiliation: Department of Human Anatomy and Histo-Embryology, Xi'an Jiaotong University Health Science Center, Xi'an, Shaanxi, China.

ABSTRACT
Mitochondrial dysfunction caused by amyloid β-peptide (Aβ) plays an important role in the pathogenesis of Alzheimer disease (AD). Substantial evidence has indicated that the mitochondrial permeability transition pore (mPTP) opening is involved in Aβ-induced neuronal death and reactive oxygen species (ROS) generation. Astragaloside IV (AS-IV), one of the major active constituents of Astragalus membranaceus, has been reported as an effective anti-oxidant for treating neurodegenerative diseases. However, the molecular mechanisms still need to be clarified. In this study, we investigated whether AS-IV could prevent Aβ1-42-induced neurotoxicity in SK-N-SH cells via inhibiting the mPTP opening. The results showed that pretreatment of AS-IV significantly increased the viability of neuronal cells, reduced apoptosis, decreased the generation of intracellular reactive oxygen species (ROS) and decreased mitochondrial superoxide in the presence of Aβ1-42. In addition, pretreatment of AS-IV inhibited the mPTP opening, rescued mitochondrial membrane potential (ΔΨm), enhanced ATP generation, improved the activity of cytochrome c oxidase and blocked cytochrome c release from mitochondria in Aβ1-42 rich milieu. Moreover, pretreatment of AS-IV reduced the expression of Bax and cleaved caspase-3 and increased the expression of Bcl-2 in an Aβ1-42 rich environment. These data indicate that AS-IV prevents Aβ1-42-induced SK-N-SH cell apoptosis via inhibiting the mPTP opening and ROS generation. These results provide novel insights of AS-IV for the prevention and treatment of neurodegenerative disorders such as AD.

Show MeSH
Related in: MedlinePlus