## 4 Discussion and Future Directions  
Despite the exponential increase in reports on the study of estrogen signaling over the past twenty years, the precise mechanisms of action in exerting the different effects in the musculoskeletal system are still to be determined. The ability of estrogen to signal not only through its nuclear receptors, ER\(\alpha\) and ER\(\beta\), but also through membrane-bound receptors, e.g., GPER1/GP30, the possible crosstalk with other signaling pathways, e.g., BMP4 signaling pathway, epigenetic regulation of estrogen receptor itself, and recruitment of coactivators that are able to modify DNA methylation and histone arrangement might shed a light on explaining the wide effects of estrogen in bone. Additionally, new evidence is emerging that ERs can be activated without ligand stimulation. While this newly discovered mechanism of action is becoming a hot topic of study in the cancer field, there are not many reports in the bone field regarding this new ER signaling. In 2014, [PERSON] et al.  
reported that unliganded ER\(\alpha\) stimulates bone sialoprotein (BSP) a tissue-specific protein expressed by osteoblasts, odontoblasts, and cementoblasts during mineralization [18].  
Estrogen depletion due to menopause or hysterectomy, or due to genetic disruption in its receptors, is detrimental to the whole body resulting in several physical and psychological changes. The systemic impact of estrogen depletion makes it impossible to understand the effect of estrogen in one specific organ due to the potential effects of its depletion in other endocrine organs. Therefore, one question that still needs to be answered is does impairment of estrogen signaling in one specific organ, like muscle or brain, affects other tissues such as bone? Answers to these questions can be obtained using the LoxP/Cre system. Several original articles have been published using animal models developed with this approach. However, most studies focus on deleting one of the receptors, ER\(\alpha\) in most studies, in a specific bone cell and observing the effects in the skeletal tissue itself. However, until now very few reports focus on analyzing the effects of deletion of estrogen receptors in bone and the effects on other organs. Another area of high importance to fully understand the role of estrogen in the musculoskeletal system is the analysis of both genders in the same study. It is well accepted that estrogen plays key roles in both female and male health. However, some studies pursuing to understand the role of estrogen focus on the female group alone, missing important observations in the males. Furthermore, these studies must be done in the context of aging and especially in the context of sudden estrogen depletion such as menopause or hysterectomy. The results from these important studies can greatly help in the prevention, treatment, and management of age-related musculoskeletal diseases such as osteoporosis and sarcopenia. More reproducible, comprehensive, and conclusive experiments are needed to have a clear understanding of the specific role and the precise mechanism of estrogen in the musculoskeletal system and its crosstalk with other organs.  
This work was supported by National Institute on Aging grants PO1 AG039355 and 2 PO1 AG039355-06. The author would like to thank [PERSON] for editorial support. Figures were created with BioRender.com (accessed on 3 May 2021). The author declares no conflict of interest. The funders had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript, or in the decision to publish the results.