LPS-mediated bone erosion mouse model and micro-CT and histological analysis

Five-week-old male ICR mice were purchased from Samtako Inc. (Osan, Korea). The mice were kept under controlled temperature (22–24 °C) and humidity (55–60 %) with a 12 h light/dark cycle. The use of experimental animals was reviewed by the institutional animal care and use committee (IACUC) and approved under WKU15-91. To examine the effect of EEST on LPS-induced bone destruction, 5-week-old male ICR mice were randomly divided into 3 groups (5 mice per group): Control (treated with PBS), LPS (treated only with LPS), and LPS + ST (treated with both LPS and EEST). EEST (200 mg/kg) or PBS was administered orally 1 day before LPS injection (5 mg/kg), and then every other 8 days. LPS was injected intraperitoneally on day 2 and 6. All mice were sacrificed after 8 days and femur of each mouse were examined by high-resolution micro-CT analysis and histological analysis including hematoxylin and eosin (H&E) and TRAP staining as described previously [17]. Briefly, micro-CT analysis was performed using bone-related parameters, including bone volume fraction (BV/TV), trabecular thickness (Tb.Th), trabecular separation (Tb.Sp), and trabecular number (Tb.N) which are minimal set of variables that should be investigated for trabecular bone regions [18]. Nomenclature, symbols, and units used in this study were recommended by the American Society for Bone Mineral Research (ASBMR) Nomenclature Committee.