Raloxifene

T47D cells were obtained from the American Type Culture Collection/National Cancer Institute (ATCC/NCI) Breast Cancer SPORE program, and MCF7 cells were purchased from the ATCC. Both cell lines were authenticated at the University of Arizona Genetics Core. T47D and MCF7 cells were maintained in RPMI 1640 medium + 10% FBS and DMEM + 5% FBS, respectively. For hormone treatment experiments, cells were deprived in phenol-red-free IMEM with 10% and 5% CSS for T47D and MCF7, respectively. CSS was purchased from Hyclone (#SH30068) and Gibco (#12676). 17β-estradiol (E2) and 4-hydroxytamoxifen (4OHT) were obtained from Sigma, and fulvestrant (Ful) and raloxifene were purchased from Tocris. AZD9496 recently described in Weir et al. [11 (link)] was kindly provided by AstraZeneca.

All experiments were conducted in female mice maintained in specific pathogen-free animal facility in individually ventilated cages with controlled temperature (22 ± 1°C) and light (12 h light/dark cycle). Animal care and use were in accordance with the guidelines of the institution. The study protocol has been approved by ethics committee of local institution (Approval No. XHEC-F-2021-012). Pax7-CreERT2 mice were purchased from Jackson Laboratory, and Esr1^f/f mice were purchased from Gem Pharmatech. Muscle stem cell specific gene knockout was induced by i.p. injection of 100 μL of 10 mg/mL tamoxifen (ABCONE, cat#T56488) every 2 days for a week.
Bipedal mice were established to simulate upright posture as described previously^{51 (link)}. Briefly, forelimb and tail of mice were firstly removed at 3 weeks old. With progressively raising water and food, these bipedal mice will gradually maintain standing posture. Unilateral intramuscular injection of 100 μL 100 nM MPP (Tocris, cat#1991) at the left side of the para-spinal muscle was performed in 3-week-old mice twice every week for 3 weeks.
In the scoliosis mouse model, intramuscular injections of 10 nM Raloxifene (Tocris, cat#2280) were performed at the concave para-spinal muscle once every 3.5 days for 2 weeks. The spinal alignment and para-spinal muscle size were then analyzed at 2 weeks after Raloxifene injection.

Fulvestrant (CAS No: 129453–61-8, > 99% purity) was purchased from MedChem Express. Estradiol (E2) (E8875), lasofoxifene (SML1026), 4-hydroxytamoxifen (H7904), and tamoxifen (T5648) were purchased from Sigma. Raloxifene (2280) was purchased from Tocris. GDC-0810 (S7855), bazedoxifene (S2128), and AZD9496 (S8372) were purchased from Selleckchem. GW5638 (5638), GW7604 (7604), and RU 58,668 (RU) were provided by Donald McDonnell (Duke University). G1T48 was provided by G1 Therapeutics, Inc., as analytical grade compound.