Submitted to: Journal of Pharmacology and Experimental Therapeutics
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 10/2/2007
Publication Date: 1/15/2008
Citation: Chen, J., Shankar, K., Nagarajan, S., Badger, T.M., Ronis, M.J. 2008. Protective effects of estradiol on ethanol-induced bone loss involves inhibition of reactive oxygen species generation in osteoblasts and downstream activation of the extracellular signal-regulated kinase/signal transducer and activator of transceiption 3/receptor activator of nuclear factor-kB ligand signaling cascade. Journal of Pharmacology and Experimental Therapeutics. 324(1):50-59.
Interpretive Summary: It has been known that chronic alcohol intake will cause bone loss in females and males. We have previously found that the sex hormone estrogen can prevent bone loss due to chronic alcohol intake. The mechanism by which chronic alcohol consumption can cause bone loss and how estrogen can prevent chronic induced bone loss are unknown. In this study, we have conducted lab experiments that have answered the questions we raised above. We found that alcohol given to bone forming cells could promote alcohol byproducts to accumulate into cells and cause gene over expression. Over-expressed genes may cause more bone resorbing cells to form and therefore resorb normal and well-built bone. All these effects of alcohol on bone were blocked by treatment of cells with estrogen. We can conclude from these studies that estrogen can control genes that are over-expressed in alcohol-treated bone-forming cells, thus providing explanation for results of previous study and providing evidence for clinical treatment of alcohol-induced bone loss.
Technical Abstract: Bone loss occurs following chronic ethanol (EtOH) consumption in males and cycling females in part as a result of increased bone resorption. We have demonstrated in vivo that estradiol treatment can reverse this effect. Using osteoclast precursors from bone marrow and osteoblast/pre-osteoclast co-culture, we found that EtOH-induced RANKL expression in osteoblasts was able to promote osteoclastogenesis. These effects were blocked by pre-treatment of cells with either 17 beta-estradiol (E2) or the antioxidant N-acetyl cysteine (NAC). EtOH treatment of stromal osteoblasts increased the intracellular level of reactive oxygen species (ROS). This was associated with induction of NADPH (nicotinamide adenine dinucleotide phosphate) oxidase (NOX), and a downstream signaling cascade involving sustained activation of extracellular signal-regulated kinase (ERK) and activation of signal transducers and activators of transcription 3 (STAT3) resulting in increased gene expression of RANKL. In the presence of EtOH, sustained nuclear ERK translocation > 24 h was observed in calvarial osteoblasts and UMR-106 cells transfected with GFP-ERK2 plasmid. This was abolished by pretreatment with either E2 or NAC. NOX subtypes 1, 2, 4, but not 3 were expressed in stromal osteoblasts. Chemical inhibition of NOX by diphenylene iodonium also reversed the ability of EtOH to phosphorylate ERK and induce RANKL mRNA expression. Down-regulation of EtOH-induced ROS generation in osteoblasts was also observed following treatment with E2 or NAC. These data suggest that the molecular mechanisms whereby E2 prevents EtOH-induced bone loss involve interference with ROS generation and cytoplasmic kinase activation.