Submitted to: Journal of Bone and Mineral Research
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 9/9/2008
Publication Date: 2/15/2009
Citation: Chen, J., Lazarenko, O.P., Haley, R.L., Blackburn, M.L., Badger, T.M., Ronis, M.J. 2009. Ethanol impairs estrogen receptor signaling resulting in accelerated activation of senescence pathways while estradiol attenuates the effects of ethanol in osteoblasts. Journal of Bone and Mineral Research. 24(2):221-230.
Interpretive Summary: Research from human and animals have suggested that chronic alcohol consumption is a major environmental factor for osteoporosis in later life. However, the reason or mechanisms regarding gradual bone mass reduction resulted from chronic alcohol drinking are largely unknown. In this study, we have utilized bone tissue from chronic ethanol (EtOH) infused female rats and isolated bone cells treated with alcohol. We have been able to determine the effects of EtOH and the interaction with sex steroid in bone cells. We found that EtOH increased some sex steroid-related gene expressions. Sex steroid itself has similar effects on those gene expressions. The interesting thing is when we give sex steroid and alcohol together, the results showed sex steroid and alcohol interacted with each other on those gene expressions mentioned above in bone cells. The other interesting thing is that chronic alcohol drinking caused bone cell senescence, and sex steroid reversed the alcohol-induced bone cell senescence. In other words, chronic alcohol drink will lead to bone cell being easy to get aged, and sex steroid can prevent this alcohol effect on those cells. Our data may explain the reason for bone weakness after chronic alcohol drink and why sex steroid can prevent this.
Technical Abstract: Epidemiological and animal studies suggest that chronic alcohol consumption increases the risk of osteoporosis. However, the mechanisms underlying alcohol-induced bone loss are largely unknown. Using bone from chronic ethanol (EtOH) infused cycling female rats and osteoblastic cells in vitro, we have been able to determine the direct effects of EtOH and the interaction with estrogen signaling pathways in osteoblasts. We found that EtOH increased (P<0.05) estrogen receptor alpha (ER alpha) and beta (ER beta) RNA and encoded ER alpha protein levels in bone in vivo and in osteoblasts in vitro when analyzed by real-time RT-PCR and Western blotting. Treatment with 17 beta-estradiol (E2) subcutaneously, in vivo, or pre-treatment of osteoblasts with E2, in vitro, antagonized all the effects of EtOH on ER expression in bone and osteoblastic cells (P<0.05). ER alpha agonist propylpyrazoletriol (PPT) and ER beta agonist diarylpropionitrile (DPN) attenuated (P<0.05) EtOH-induced ER alpha and ER beta gene over expression, respectively, indicating non-ER specific actions of EtOH on osteoblasts. In addition E2 attenuated EtOH-induced activation of p53 and p21 in bone tissue in vivo and osteoblasts in vitro. UMR-106 osteoblastic cells were transiently transfected with ER alpha-ECFP for ER alpha translocation analysis, and transiently transfected with ERE-TK-Luc or p21 promoter pGL2-p21-Luc reporter plasmids with or without co-transfection of ER alpha for expression analysis using luciferase assays. Similar to estrogen receptor antagonist ICI 182,780, EtOH blocked nuclear translocation of ER alpha-ECFP in the presence of E2. EtOH down-regulated (P<0.05) the ERE-luc reporter activity; however, E2 blocked this EtOH effect in osteoblasts. On the other hand, EtOH transactivated the luciferase activity of the p21 promoter region independent of additional exogenous ERa. EtOH activated (P<0.05) senescence-associated beta-galactosidase activity in rat stromal osteoblasts, while E2 attenuated this EtOH action. We conclude that inhibitory cross-talk between EtOH and E2 in osteoblasts on ERs, p53/p21 and cell senescence provides a pathophysiologic mechanism underlying bone loss and the protective effects of estrogens in alcohol-exposed females.