Targeted Disruption of the CSF-1 Gene Leads to Osteopetrosis and Osteoblast Defects .
Sherry Abboud Werner, Diane Horn, Kathleen Woodruff, Roberto Fajardo, Marie Harris, Stephen Harris. University of Texas Health Science Center, San Antonio
Background: CSF-1, a key determinant of osteoclast-mediated bone remodeling, is highly expressed by osteoblasts and osteocytes. CSF-1 deficiency in spontaneous mutant op/op mice decreases macrophages/osteoclasts and leads to osteopetrosis and a pleiotropic phenotype. The effect of CSF-1 knockout (KO) in all tissues or conditional KO of CSF-1 using a Cre/loxP system has not been explored. Objective: To determine the effect of inactivation of CSF-1 and develop a strategy for examining the biologic effect of CSF-1 KO in bone using Cre-lox technology.
Design: A targeting vector for generating a conditional KO allele for CSF-1 (deleting exons 4,5,6) was used to generate heterozygous mice harboring the floxed construct without a neo cassette (fx allele). CSF-1fx/CSF-1fx mice were bred with Meox2Cre mice to produce mice homozygous for the KO allele(hCSF-1KO). (Meox2 promoter drives expression of Cre throughout the epiblast). At 3 weeks, hCSF-1KO and WT littermates were analyzed for CSF-1 protein and F4/80+ macrophages. Hind limbs were examined by x-ray, microCT, histology and stained with TRAP and CD31 to identify osteoclasts and vessels, respectively.
Results: Homozygous (h)CSF-1KO showed absence of CSF-1 in all tissues, reduced macrophages in most organs, decreased circulating monocytes and osteopetrosis with failure of tooth eruption similar to op/op mice. Radiographs of hCSF-1KO showed marked skeletal sclerosis and, by microCT, %BV/TV and trabecular number were increased and cortical bone thickness was decreased; bone mineral density was increased by DEXA scan. Histologically, hCSF-1KO bones showed an expanded growth plate, increased bone trabeculae with prominent cartilage cores replacing the marrow cavity, narrow vascular sinusoids and poorly formed cortical bone. Numerous TRAP+ osteoclasts were identified in WT, whereas rare mononucleated osteoclast-like cells were detected in CSF-1KO bone. In CSF-1KO, osteoblasts showed loss of polarity; matrix formation and collagen fibrils were disorganized and abnormal clusters of osteocytes entrapped in matrix were identified.
Conclusions: Results provide the first evidence that global CSF-1KO using a Meox2Cre-based system leads to osteopetrosis and alters osteoblast function. Conditional KO of CSF-1 in bone cells is feasible and will be crucial for elucidating the mechanisms by which CSF-1 exerts pleiotropic effects and regulates bone turnover and repair.
Wednesday, March 2, 2011 9:30 AM
Poster Session V # 231, Wednesday Morning