Homocysteine enhances bone resorption by stimulation of osteoclast formation and activity through increased intracellular ROS generation.
Hyperhomocystinemia is a modifiable risk factor for osteoporosis and fracture. Physiologic concentrations of Hcy directly activate osteoclast formation and activity through stimulation of p38 MAPK and integrin beta3. The effects of Hcy were mediated by generation of intracellular ROS.


BACKGROUND
Hyperhomocysteinemia is a modifiable risk factor for osteoporosis and its related bone fractures. It has been reported that bone resorption and turnover rate were increased in hyperhomocystinemia. Using mouse bone marrow cells, we examined the direct effects of homocysteine (Hcy) on osteoclast formation and activity.


METHODS
Osteoclast formation was determined by TRACP staining and TRACP activity. Intracellular reactive oxygen species (ROS) generation was measured using a fluorescent probe, dichlorodihydrofluorescein diacetate. Intracellular signaling cascades of p38 mitogen-activated protein kinase (MAPK), extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK), and NF-kappaB were measured by Western blotting. Integrin beta3 mRNA levels were measured by RT-PCR. Actin ring formation and bone resorption assays were also performed.


RESULTS
Physiologic concentrations of Hcy upregulated TRACP+ multinucleated cells and TRACP activity, stimulated actin ring formation, and increased the number of nuclei per cell and the level of expression of integrin beta3 mRNA. In addition, Hcy increased bone resorption and stimulated p38 MAPK activity and intracellular reactive oxygen species (ROS) generation. All of these Hcy-induced changes were blocked by pretreatment with the antioxidant, N-acetyl cysteine.


CONCLUSIONS
Hcy directly activates osteoclast formation and activity through increased generation of intracellular ROS. These findings suggest that, in individuals with mild to moderate hyperhomocystinemia, increased bone resorption by osteoclasts may contribute to osteoporosis and that an antioxidant may attenuate bone loss in these individuals.
Koh
Jung-Min
JM
Division of Endocrinology and Metabolism, Asan Medical Center, University of Ulsan College of Medicine, Songpa-Gu, Seoul, Korea.
Lee
Young-Sun
YS
Kim
Yang Soon
YS
Kim
Duk Jae
DJ
Kim
Hong-Hee
HH
Park
Joong-Yeol
JY
Lee
Ki-Up
KU
Kim
Ghi Su
GS
D000135:Acid Phosphatase / Q000378:metabolism; D000818:Animals; D001854:Bone Marrow Cells / Q000378:metabolism*; D001862:Bone Resorption / Q000188:drug therapy*; D002454:Cell Differentiation / Q000187:drug effects*; D002478:Cells, Cultured; D050723:Fractures, Bone / Q000378:metabolism; D006710:Homocysteine / Q000378:metabolism / Q000494:pharmacology*; D020138:Hyperhomocysteinemia / Q000150:complications / Q000378:metabolism; D007527:Isoenzymes / Q000378:metabolism; D020935:MAP Kinase Signaling System / Q000187:drug effects; D051379:Mice; D008813:Mice, Inbred ICR; D010010:Osteoclasts / Q000378:metabolism*; D010024:Osteoporosis / Q000209:etiology / Q000378:metabolism; D011494:Protein Kinases / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism*; D012307:Risk Factors; D000071681:Tartrate-Resistant Acid Phosphatase
D007527:Isoenzymes; D017382:Reactive Oxygen Species; D006710:Homocysteine; D011494:Protein Kinases; D000135:Acid Phosphatase; D000071681:Tartrate-Resistant Acid Phosphatase