Abstract
Cysteine (C)-X-C motif chemokine receptor 4 (CXCR4), the primary receptor for stromal cell-derived factor-1 (SDF-1), is involved in bone morphogenic protein 2 (BMP2)-induced osteogenic differentiation of mesenchymal progenitors. To target the in vivo function of CXCR4 in bone and explore the underlying mechanisms, we conditionally inactivated CXCR4 in osteoprecursors by crossing osterix (Osx)-Cre mice with floxed CXCR4 (CXCR4(fl/fl)) mice to generate knock-outs with CXCR4 deletion driven by the Osx promoter (Osx::CXCR4(fl/fl)). The Cre-mediated excision of CXCR4 occurred exclusively in bone of Osx::CXCR4(fl/fl) mice. When compared with littermate controls, Osx::CXCR4(fl/fl) mice developed smaller osteopenic skeletons as evidenced by reduced trabecular and cortical bone mass, lower bone mineral density, and a slower mineral apposition rate. In addition, Osx::CXCR4(fl/fl) mice displayed chondrocyte disorganization in the epiphyseal growth plate associated with decreased proliferation and collagen matrix syntheses. Moreover, mature osteoblast-related expression of type I collagen α1 and osteocalcin was reduced in bone of Osx::CXCR4(fl/fl) mice versus controls, suggesting that CXCR4 deficiency results in arrested osteoblast progression. Primary cultures for osteoblastic cells derived from Osx::CXCR4(fl/fl) mice also showed decreased proliferation and impaired osteoblast differentiation in response to BMP2 or BMP6 stimulation, and suppressed activation of intracellular BMP receptor-regulated Smads (R-Smads) and Erk1/2 was identified in CXCR4-deficient cells and bone tissues. These findings provide the first in vivo evidence that CXCR4 functions in postnatal bone development by regulating osteoblast development in cooperation with BMP signaling. Thus, CXCR4 acts as an endogenous signaling component necessary for bone formation.
Highlights
Stromal cell-derived factor-1 (SDF-1; named CXCL12 or pre-B cell stimulating factor) [1,2,3,4]
CXCR4 in Bone Development mesenchymal stem cell induction of ectopic bone formation in nude mice [27]. In mesenchymal cultures, such as human and mouse bone marrow-derived stromal cells [33] and C2C12 and ST2 cells [25], we demonstrated that blocking the stromal cell-derived factor-1 (SDF-1)/ CXCR4 signal axis inhibits the differentiation of these cells toward the osteoblastic lineage in response to bone morphogenic protein 2 (BMP2) stimulation
Growth Plate Disorganization and Abnormal Osteoblast Development in Osx::CXCR4fl/fl Mice—To understand molecular and cellular changes associated with bone defects in Osx::CXCR4fl/fl mice, we examined the epiphyseal growth plate where bone formation occurs on a cartilaginous template [44, 45]
Summary
Antibodies and Reagents—Anti-CXCR4 and anti-Osx antibodies were purchased from eBioscience (San Diego, CA) and Abcam (Cambridge, MA), respectively. We first generated double heterozygous mice for Cre and floxed CXCR4 (Osx-Cre; CXCR4fl/ϩ) that were bred to CXCR4fl/fl mice via a backmating strategy to generate excised floxed CXCR4 homozygous (Osx::CXCR4fl/fl; used as conditional knock-outs), heterozygous for floxed CXCR4 (Osx-Cre;CXCR4fl/ϩ), and Cre-null mice (CXCR4fl/ϩ and CXCR4fl/fl; used as littermate controls throughout the study). Quantitative volumetric analyses of cortical and trabecular microstructures were performed on entire parietal calvariae and on a region of 200 microtomographic slices (2.1 mm) at the proximal tibia with an isotropic resolution of 6 m. These analyses measured the cortical and trabecular bone volume fraction, thickness, and bone mineral density as well as trabecular number, separation, and connectivity density.
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
