Alpha-lipoic acid inhibits TNF-alpha-induced apoptosis in human bone marrow stromal cells.
TNF-alpha is an important mediator of bone loss. In the HS-5 hBMSC, TNF-alpha and H2O2 increased intracellular ROS levels and induced cell apoptosis through activation of caspases, JNK and NF-kappaB. alpha-Lipoic acid prevented these changes induced by TNF-alpha and H2O2, suggesting its potential therapeutic applications in attenuating bone loss.


BACKGROUND
Oxidative stress is an important mediator of bone loss. TNF-alpha, which plays a critical role in the bone loss after menopause, has been shown to increase intracellular oxidative stress. Because oxidative stress is associated with cell death, we analyzed the apoptotic effects of TNF-alpha and H2O2 on human bone marrow stromal cells (hBMSCs). We also examined the protective effects of an important biological thiol antioxidant, alpha-lipoic acid (alpha-LA), against TNF-alpha- and H2O2-induced apoptosis.


METHODS
Using the HS-5 hBMSC cell line, we tested whether TNF-alpha-induced apoptosis was mediated by the generation of excessive reactive oxygen species (ROS). Apoptosis was determined by 3-(4,5-dimethylthiazol-2-yl)2,5-diphenyltetrazolium bromide (MTT) assay, trypan blue exclusion assay, quantitation of histone-associated DNA fragments in cytosol, and the activation of caspases. The mechanisms mediating these apoptotic effects were determined by Western blotting and enzyme immunoassay.


RESULTS
Both TNF-alpha and H2O2 increased intracellular ROS levels, reduced total cellular glutathione levels, activated caspases-3, -9, and -8, and enhanced hBMSC apoptosis. The activation of c-jun N-terminal kinase (JNK) and NF-kappaB mediated these apoptotic effects. Pretreatment of cells with alpha-LA prevented these changes induced by TNF-alpha and H2O2.


CONCLUSIONS
Our data show that TNF-alpha increases intracellular ROS in hBMSC and that TNF-alpha and H2O2 induce apoptosis in hBMSC through the activation of JNK and NF-kappaB. Our findings also suggest that alpha-LA may have therapeutic applications in halting or attenuating bone loss associated with increased oxidative stress.
Byun
Chang-Hyun
CH
Asan Institute for Life Sciences, Seoul, Republic of Korea.
Koh
Jung-Min
JM
Kim
Dong Kwan
DK
Park
Seung-Il
SI
Lee
Ki-Up
KU
Kim
Ghi Su
GS
D000975:Antioxidants / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D001854:Bone Marrow Cells / Q000187:drug effects* / Q000378:metabolism; D001862:Bone Resorption / Q000188:drug therapy; D020169:Caspases / Q000378:metabolism; D005978:Glutathione / Q000378:metabolism; D006801:Humans; D006861:Hydrogen Peroxide / Q000037:antagonists & inhibitors / Q000494:pharmacology; D048031:JNK Mitogen-Activated Protein Kinases / Q000378:metabolism; D048670:MAP Kinase Kinase 4; D020929:Mitogen-Activated Protein Kinase Kinases / Q000378:metabolism; D016328:NF-kappa B / Q000378:metabolism; D010006:Osteoblasts / Q000187:drug effects; D018384:Oxidative Stress / Q000187:drug effects*; D017382:Reactive Oxygen Species / Q000378:metabolism; D015398:Signal Transduction; D017154:Stromal Cells / Q000187:drug effects / Q000378:metabolism; D008063:Thioctic Acid / Q000494:pharmacology*; D014409:Tumor Necrosis Factor-alpha / Q000037:antagonists & inhibitors* / Q000378:metabolism / Q000494:pharmacology
D000975:Antioxidants; D016328:NF-kappa B; D017382:Reactive Oxygen Species; D014409:Tumor Necrosis Factor-alpha; D008063:Thioctic Acid; D006861:Hydrogen Peroxide; D048031:JNK Mitogen-Activated Protein Kinases; D048670:MAP Kinase Kinase 4; D020929:Mitogen-Activated Protein Kinase Kinases; D020169:Caspases; D005978:Glutathione