Fig. 2

SLIT3 deficiency increases matured OCPs and upregulates RANK expression with LPS stimulation. A-B Bone marrow (BM) cells obtained from WT or Slit3^−/− mice that had received two LPS (5 mg/kg) intraperitoneal injections were analyzed by fluorescence-activated cell sorting (FACS) (A). Isolated BM cells were gated on ckit⁺CD45⁺, then cfms⁺CD11b^low and cfms⁺CD11b^hi populations were shown (B). C OCPs obtained from WT or Slit3^−/− mice were incubated with LPS (10 ng/ml) and cell surface RANK⁺ cell was observed by FACS analysis. D-F BM cells were incubated with M-CSF (30 ng/mL) and RANKL (10 ng/mL) for six days. Cells were then stained for TRAP (D) and the number of TRAP⁺ multinucleated cells (MNCs) containing more than three nuclei (E). The increase of OC numbers in Slit3^−/− (Slit3^−/− / WT) was calculated in PBS or LPS injection (F). Scale bar, 100 μm. G OCPs isolated from WT or Slit3^−/− mice were incubated with or without 10 ng/mL LPS and RANK expression was detected and quantified by immunoblotting against RANK- and β-actin-specific antibodies. Densitometry quantification of RANK compared to Actin is represented (right). Data are shown as mean ± s.d., *P < 0.05, **P < 0.005, ***P < 0.0001. All representative data from three independent experiments are shown