Colony -stimulating factor-1 receptor-signaling in the development of the skeletal and the central nervous systems

Abstract

Background. Colony stimulating factor-1 (CSF-1) is the primary mononuclear phagocyte growth factor. All the effects of CSF-1 are mediated by the CSF-1 receptor tyrosine kinase (CSF-1R), which is expressed by the cells of monocyte/macrophage lineage, including macrophages, osteoclasts in bone and microglia in brain, as well as by some other cell types. CSF-1 is expressed as three distinct isoforms, membrane-spanning cell-surface glycoprotein (csCSF-1), secreted glycoprotein (sgCSF-1) and secreted chondroitin sulfate (Glycosaminoglycan or GAG) proteoglycan (spCSF-1), each containing the first 150 amino acids that are essential for in vitro biological activity. CSF1-deficient (Csf1op/op) mice are osteopetrotic, due to a paucity of osteoclasts and have reduced tissue macrophages. However, CSF-1R-deficient (Csf1r-/-) mice display a more severe skeletal phenotype and a second CSF-1R ligand, interleukin-34 (IL-34) was recently identified. Expression of all 3 isoforms together but not csCSF-1 alone, corrects the osteopetrotic defects of Csf1 op/op mice, suggesting a role for secreted CSF-1s in osteoclast development/function. The density of microglia in Csf1op/op brains is reduced and CSF-1 promotes neuronal survival and outgrowth in vitro, suggesting that CSF-1 is trophic for neural cells.;Aim. To examine the role of the CSF-1/CSF-1R in the development of skeleton (subaim 1) and brain (subaim 2).;Subaim 1. To determine the roles of the secreted CSF-1 isoforms and in particular, the spCSF-1 GAG chains in CSF-1-dependent skeletal development.;Methods. Transgenic mice expressing secreted CSF-1 precursors that differed only by presence or absence of the GAG addition site were characterized to determine the biological role of the CSF-1 GAG chains. The actions of both purified secreted CSF-1 isoforms on osteoclast development in vitro were also studied.;Results. Compared with mice exclusively expressing spCSF-1 precursor that generated both sp and sg CSF-1, mice expressing SpDeltaGAG and sg CSF-1, devoid of the GAG chains, failed to completely rescue the osteopetrotic phenotype of Csf1op/op mice, displaying reduced osteoclast polarization. However, in vitro, sgCSF-1 stimulated osteoclastogenesis more efficiently than spCSF-1.;Significance. These studies demonstrate an important in vivo role of the spCSF-1 GAG chains in the regulation of osteoclast function. In addition, distinct roles for secreted CSF-1s in osteoclast development are suggested, while sgCSF-1 may be important for the generation of osteoclasts, spCSF-1 is necessary for their function.;Subaim 2. To examine the role of CSF-1R-signaling in brain development.;Methods. Both Csf1op/op and Csf1r-/- mice were characterized to determine the roles of CSF-1 and CSF-1R in brain. CSF-1-reporter transgenic mice were used to determine CSF-1 mRNA localization and CSF-1R and IL-34 expression were determined immunohistochemically. FACS-purified neural stem cell (NSC) clonal cultures were used to assess the cell autonomous effects of CSF-1R in NSC.;Results. Csf1 promoter activity was detected in the post-mitotic neurons of the regional neurogenic niches. Csf1r-/- brains exhibited multiple morphogenetic alterations. They also lacked microglia and displayed increased proliferation and apoptosis of NSC and reduced numbers of mature oligodendrocytes and neurons. CSF-1R was expressed by NSC and by more lineage-restricted neuronal and glial precursors. Consistent with the existence of another ligand (IL-34) for the CSF-1R, the brain phenotype of Csf1op/op mice was less severe than the Csf1r-/- brain phenotype. Like CSF-1, IL-34 is expressed by neurons, but IL-34 expression is developmentally earlier and minimally overlaps with CSF-1 expression. In vitro, CSF-1 or IL-34 suppressed NSC self-renewal and enhanced neurogenesis, in the absence of microglia.;Significance. These results indicate that CSF-1R signaling is absolutely required for the development of microglia and, via the action of its ligands, plays additional roles in brain development by regulating NSC survival, proliferation and differentiation.