Fig. 7: Cldn11 positively regulates OBs through interaction with the EphB4 receptor.

a The expression of Cldn11, EphrinB2, and EphB4 and co-localization of these proteins in the differentiated OBs were observed using a confocal imaging assay. b Primary OB precursor cells were cultured in osteogenic medium containing AA (50 μg/mL) and β-gly (10 mM) for the indicated times, and mRNA levels of EphrinB2 and EphB4 were analyzed by quantitative real time RT-PCR. c OBs were cultured in the absence of AA and β-gly for 1 day prior to cell lysis. Lysates were immunoprecipitated with EphrinB2 or EphB4 antibody and subsequently immunoblotted with Cldn11 antibody. d BMMs transfected with siControl or siCldn11 were cultured and lysed in the presence of AA and β-gly for 0, 2, 4, and 8 days. Lysates were immunoprecipitated with EphB4 antibody and subsequently immunoblotted with Cldn11 antibody. c, d Co-immunoprecipitated samples (top) or total cell extracts (bottom) were subjected to western blotting to detect Cldn11, EphrinB2, EphB4, and β-actin. e, f OBs were divided into siControl, siCldn11, and siCldn11 treated with EphrinB2-Fc groups and cultured with AA and β-gly for e 7 days to perform the ALP assay and ALP activity assay and f 21 days to conduct the ARS assay. **P < 0.01; ***P < 0.001 vs. the control (0 day) or siControl, ^##P < 0.01; ^###P < 0.001 vs. the siControl