Alpha-lipoic acid (ALA) is a naturally occurring antioxidant and has been previously used to treat diabetes and cardiovascular disease. However, the autophagy effects of ALA against oxidative stress-induced dopaminergic neuronal cell injury remain unclear. The aim of this study was to investigate the role of ALA in autophagy and apoptosis against oxidative stress in the SH-SY5Y human dopaminergic neuronal cell line. We examined SH-SY5Y phenotypes using the 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide assay (cell viability/proliferation), 4′,6-diamidino-2-phenylindole dihydrochloride nuclear staining, Live/Dead cell assay, cellular reactive oxygen species (ROS) assay, immunoblotting, and immunocytochemistry. Our data showed ALA attenuated hydrogen peroxide (H2O2)-induced ROS generation and cell death. ALA effectively suppressed Bax up-regulation and Bcl-2 and BclxL down-regulation. Furthermore, ALA increased the expression of the antioxidant enzyme, heme oxygenase-1. Moreover, the expression of Beclin-1 and LC-3 autophagy biomarkers was decreased by ALA in our cell model. Combined, these data suggest ALA protects human dopaminergic neuronal cells against H2O2-induced cell injury by inhibiting autophagy and apoptosis.

Introduction
Materials and Methods
    1. Materials
    2. Cell culture and cell treatments
    3. Cytotoxicity assay
    4. Cell Live/Dead assay
    5. DAPI staining
    6. Measurement of intracellular ROS
    7. Immunoblotting
    8. Haematoxylin and eosin staining
    9. Immunocytochemistry
    10. Statistical analysis
Results
    1. ALA protects SH-SY5Y cells against H2O2-inducedcytotoxicity
    2. ALA inhibited H2O2-induced apoptosis and ROSproduction in SH-SY5Y cells
    3. ALA protects H2O2-induced cell death throughdown-regulating oxidative stress
    4. ALA inhibits autophagy in H2O2-induced SH-SY5Ycell injury
Discussion
References

• Kyeong-Rok Kang(The Institute of Dental Science, Chosun University)
• Jae-Sung Kim(The Institute of Dental Science, Chosun University)
• Tae-Hyeon Kim(The Institute of Dental Science, Chosun University)
• Jeong-Yeon Seo(Department of Integrative Biological Sciences & BK21 FOUR Educational Research Group for Age-associated Disorder Control Technology, Chosun University)
• HyangI Lim(The Institute of Dental Science, Chosun University)
• Jong-Hyun Park(The Institute of Dental Science, Chosun University)
• Kwang Yeol Yang(The Institute of Dental Science, Chosun University)
• Sun-Kyoung Yu(The Institute of Dental Science, Chosun University)
• Heung-Joong Kim(The Institute of Dental Science, Chosun University)
• Chun Sung Kim(The Institute of Dental Science, Chosun University)
• Hong Sung Chun(Department of Integrative Biological Sciences & BK21 FOUR Educational Research Group for Age-associated Disorder Control Technology, Chosun University)
• Dong-Seol Lee(Laboratory for the Study of Regenerative Dental Medicine, Department of Oral Histology-Developmental Biology, School of Dentistry and Dental Research Institute, Seoul National University)
• Joo-Cheol Park(Laboratory for the Study of Regenerative Dental Medicine, Department of Oral Histology-Developmental Biology, School of Dentistry and Dental Research Institute, Seoul National University)
• Do Kyung Kim(The Institute of Dental Science, Chosun University) Correspondence to