Extra d glucose

Y. pestis KIM6+ and E. coli DH5α strains transformed with plasmids carrying terZAB or terCDE genes were examined under light microscope to evaluate the morphological changes caused by expression of these genes. Overnight cultures of Y. pestis KIM6+ or E. coli DH5α strains were diluted to 1×10⁶ CFU/mL in LB broth containing 1% extra D-glucose (Sigma-Aldrich) or 0.05 or 1 mM IPTG and 100 μg/mL ampicillin as the final concentrations. These cultures were incubated at 26°C for Y. pestis strains and at 37°C for E. coli DH5α strains for 5 h with 160 rpm shaking. Subsequently, the samples were spun on glass slides using cytocentrifuge, stained with Wright Giemsa stain and observed under light microscope at 1,000× magnification

MC3T3-E1 cells originated from Zhong Qiao Xin Zhou Biotechnology Co., Ltd. (Shanghai, China). Minimum essential medium-alpha modification (α-MEM) and foetal bovine serum (FES) were provided by Gibco (Gaithersburg, USA). MC3T3-E1 cells were cultured with α-MEM supplemented with 10% FBS at 37 °C in a 5% CO2 atmosphere.
High-glucose (25.0 mmol/L) medium was produced by adding extra D-glucose purchased from Sigma–Aldrich Co., Ltd. (Shanghai, China) to the ordinary medium. β-glycerophosphate and ascorbic acid were purchased from Aladdin Co., Ltd. (Shanghai, China), and β-glycerophosphate (10 mM) and ascorbic acid (50 mg/mL) dissolved in α-MEM supplemented with 10% FBS acted as the osteogenesis medium. Anti-GSK3β, anti-p-GSK3β (S9), anti-β-catenin, anti-Cyclin D1, anti-Cmyc, and anti-Runx2 were purchased from Abcam (Cambridge, UK). Alizarin Red S staining solution, propidium iodide solution and DAPI solution were purchased from Solarbio Co., Ltd. (Beijing, China). Furthermore, Beyotime Biotechnology (Suzhou, China) provided an EdU Cell Proliferation Kit with Alexa Fluor 555, BCIP/NBT Alkaline Phosphatase Colour Development Kit, Alkaline Phosphatase Assay Kit, anti-β-actin, HRP-labelled secondary antibody and Enhanced Chemiluminescence Kit. XAV939 was purchased from MedChemExpress Company (New Jersey, USA).