β-Estradiol antagonizes the inhibitory effects of caffeine in BMMSCs via the ERβ-mediated cAMP-dependent PKA pathway

Caffeine negatively mediates bone homeostasis to cause bone loss and even osteoporosis. This phenomenon occurs in postmenopausal women with estrogen deficiency but not in healthy young women. In this study, we determined whether the effects of caffeine on bone homeostasis were antagonized by estrogen and the underlying mechanisms. In particular, because high levels of cAMP, an important second messenger, have been observed in postmenopausal women suffering from osteoporosis, we examined the role of cAMP in the effects of caffeine on bone homeostasis. In vivo study showed that caffeine accelerated bone loss in osteoporotic rats, whereas β-estradiol blunted the negative effect of caffeine on bone. In vitro study, we harvested bone marrow-derived mesenchymal stromal cells (BMMSCs) from osteoporotic rats. We found that caffeine and β-estradiol inversely affected BMMCSs proliferation, apoptosis, osteogenic lineage commitment, extracellular matrix synthesis and mineralization. These parameters were assessed according to the expression levels of osteogenic markers, alkaline phosphatase activity and Alizarin red staining. The deleterious effects of caffeine on BMMSCs were blunted by β-estradiol. The cAMP-dependent PKA pathway was found to be involved in regulating caffeine/β-estradiol-mediated cell growth, survival and osteogenesis. Additionally, after estrogen receptor (ER) β knockdown, the antagonistic effects of β-estradiol on caffeine were nearly abolished. These results indicated that by binding to ERβ, β-estradiol antagonizes the negative impacts of caffeine on cell growth and osteogenic differentiation in BMMSCs through the cAMP-dependent PKA signaling pathway.