The effects of colony-stimulating factor 1 (CSF-1), the primary regulator of mononuclear phagocyte production, are thought to be mediated by the CSF-1 receptor (CSF-1R), encoded by the c-fms proto-oncogene. To investigate the in vivo specificity of CSF-1 for the CSF-1R, the mouse IntroductionColony-stimulating factor 1 (CSF-1) regulates the survival, proliferation, and differentiation of mononuclear phagocytic cells and is the primary regulator of mononuclear phagocyte production in vivo. 1,2 However, CSF-1 also regulates cells of the female reproductive tract and plays an important role in fertility. 3,4 The effects of CSF-1 are mediated by a high-affinity receptor tyrosine kinase (CSF-1R) [5][6][7][8] encoded by the c-fms proto-oncogene. 9 The CSF-1R is expressed on primitive multipotent hematopoietic cells, 10,11 mononuclear phagocyte progenitor cells, 12 monoblasts, promonocytes, monocytes, 5,6 tissue macrophages, 6,13-15 osteoclasts, 16 B cells, 17,18 smooth muscle cells, 19 and neurons. 20,21 CSF-1R messenger RNA (mRNA) is expressed in Langerhans cells, 22 in the female reproductive tract, in oocytes and embryonic cells of the inner cell mass and trophectoderm, 23 in decidual cells, [24][25][26] and in cells of the trophoblast. 24,25 The expression of the CSF-1R on primitive hematopoietic cells that are unable to proliferate in vitro in response to CSF-1 alone 10,11 but are able to proliferate and differentiate if stimulated with combinations of CSF-1 and other hematopoietic growth factors 10,11,27 suggests that CSF-1R is involved in the regulation of more primitive hematopoietic cells than those that form macrophage colonies in vitro in response to CSF-1 alone.Mice homozygous for the mutation osteopetrotic 28 possess an inactivating mutation in the coding region of the CSF-1 gene and are devoid of detectable CSF-1. 29,30 These Csf1 op /Csf1 op mice are osteopetrotic because of an early and marked deficiency of osteoclasts 28 that spontaneously recovers with age, 31,32 probably because of the action of vascular endothelial growth factor. 33 However, the phenotype of these mice is pleiotropic. 3 They are toothless; have low body weight, low growth rate, and skeletal abnormalities; and are deficient in tissue macrophages. 2,28,30,34,35 They have defects in both male and female fertility, neural development, the dermis, and synovial membranes. 3 The pleiotropic phenotype of the Csf1 op /Csf1 op mouse may be due to a reduction in trophic and/or scavenger functions of the tissue macrophages regulated by CSF-1, secondary to the reduction of their concentration in tissues, 2 because outside the female reproductive tract the CSF-1R is primarily expressed in mononuclear phagocytes. 1,3 However, it is possible that some of these effects may also be due to loss of function of other cells such as neuronal cells and muscle precursors, which have also been reported to express the CSF-1R. 20,36 To address the questions of whether CSF-1 activates other receptors besides the CSF-1R and, conversely, whether the CSF-1R me...

Langerhans cells (LCs) are the only dendritic cells of the epidermis and constitute the first immunological barrier against pathogens and environmental insults. The factors regulating LC homeostasis remain elusive and the direct circulating LC precursor has not yet been identified in vivo. Here we report an absence of LCs in mice deficient in the receptor for colony-stimulating factor 1 (CSF-1) in steady-state conditions. Using bone marrow chimeric mice, we have established that CSF-1 receptor-deficient hematopoietic precursors failed to reconstitute the LC pool in inflamed skin. Furthermore, monocytes with high expression of the monocyte marker Gr-1 (also called Ly-6c/G) were specifically recruited to the inflamed skin, proliferated locally and differentiated into LCs. These results identify Gr-1 hi monocytes as the direct precursors for LCs in vivo and establish the importance of the CSF-1 receptor in this process.Langerhans cells (LCs) are members of a family of highly specialized antigen-presenting cells called dendritic cells (DCs). Uniquely present in the epidermis, LCs form a tight cellular network that covers the entire body surface, constituting the first immunological barrier against the external environment. LCs are well equipped to ingest antigens present in the skin and to migrate to the regional lymph node in both steady-state and inflammatory conditions 1,2 . After being activated, LCs increase their expression of major histocompatibility complex (MHC) class II and costimulatory molecules 3 and migrate to T cell areas 2 of draining lymph nodes, where they secrete T cell-attractant chemokines 4 and interact with antigen-specific T cells. Whether migrating LCs serve mainly to carry antigens to blood-derived DCs in the draining lymph nodes or whether they directly prime or tolerize antigen-specific T cells is still debated 5-8 .Given their importance in skin immunity, the mobilization of LCs to draining lymph nodes and the subsequent replacement of migrating cells by newly differentiated LCs must be tightly regulated events. Indeed, LC homeostasis is differentially regulated in quiescent and Reprints and permissions information is available online at http://npg.nature.com/reprintsandpermissions/ Correspondance should be addressed to M.M. (miriam.merad@mssm.edu).

The CSF-1 receptor (CSF-1R) regulates CNS microglial development. However, the localization and developmental roles of this receptor and its ligands, IL-34 and CSF-1, in the brain are poorly understood. Here we show that compared to wild type mice, CSF-1R-deficient (Csf1r−/−) mice have smaller brains of greater mass. They further exhibit an expansion of lateral ventricle size, an atrophy of the olfactory bulb and a failure of midline crossing of callosal axons. In brain, IL-34 exhibited a broader regional expression than CSF-1, mostly without overlap. Expression of IL-34, CSF-1 and the CSF-1R were maximal during early postnatal development. However, in contrast to the expression of its ligands, CSF-1R expression was very low in adult brain. Postnatal neocortical expression showed that CSF-1 was expressed in layer VI, whereas IL-34 was expressed in the meninges and layers II–V. The broader expression of IL-34 is consistent with its previously implicated role in microglial development. The differential expression of CSF-1R ligands, with respect to CSF-1R expression, could reflect their CSF-1R-independent signaling. Csf1r−/− mice displayed increased proliferation and apoptosis of neocortical progenitors and reduced differentiation of specific excitatory neuronal subtypes. Indeed, addition of CSF-1 or IL-34 to microglia-free, CSF-1R-expressing dorsal forebrain clonal cultures, suppressed progenitor self-renewal and enhanced neuronal differentiation. Consistent with a neural developmental role for the CSF-1R, ablation of the Csf1r gene in Nestin-positive neural progenitors led to a smaller brain size, an expanded neural progenitor pool and elevated cellular apoptosis in cortical forebrain. Thus our results also indicate novel roles for the CSF-1R in the regulation of corticogenesis.

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In mammals, the fetal liver is the first site of definitive erythropoiesis-the generation of mature, enucleated red cells. The functional unit for definitive erythropoiesis is the erythroblastic island, a multicellular structure composed of a central macrophage surrounded by erythroblasts at various stages of differentiation. Targeted disruption of the retinoblastoma (Rb) tumour suppressor gene in the mouse leads to embryonic death caused by failure of erythroblasts to enucleate. The erythroid defect has been attributed to loss of Rb in cells that support erythropoiesis, but the identity of these cells is unknown. Here we show that Rb-deficient embryos carry profound abnormalities of fetal liver macrophages that prevent physical interactions with erythroblasts. In contrast, wild-type macrophages bind Rb-deficient erythroblasts and lead them to terminal differentiation and enucleation. Loss of Id2, a helix-loop-helix protein that mediates the lethality of Rb-deficient embryos, rescues the defects of Rb-deficient fetal liver macrophages. Rb promotes differentiation of macrophages by opposing the inhibitory functions of Id2 on the transcription factor PU.1, a master regulator of macrophage differentiation. Thus, Rb has a cell autonomous function in fetal liver macrophages, and restrains Id2 in these cells in order to implement definitive erythropoiesis.

The primary macrophage growth factor, colony-stimulating factor 1 (CSF-1), is expressed as a secreted glycoprotein or proteoglycan found in the circulation or as a biologically active cell surface glycoprotein (csCSF-1). To investigate the in vivo roles of csCSF-1, we created mice that exclusively express csCSF-1, in a normal tissue-specific and developmental manner, by transgenic expression of csCSF-1 in the CSF-1-deficient osteopetrotic (Csf1 op / Csf1 op ) background. The gross defects of Csf1 op /Csf1 op mice, including growth retardation, failure of tooth eruption, and abnormal male and female reproductive functions were corrected. Macrophage densities in perinatal liver, bladder, sublinguinal salivary gland, kidney cortex, dermis, and synovial membrane were completely restored, whereas only partial or no restoration was achieved in adult liver, adrenal gland, kidney medulla, spleen, peritoneal cavity, and intestine. Residual osteopetrosis, significantly delayed trabecular bone resorption in the subepiphyseal region of the long bone, and incomplete correction of the hematologic abnormalities in the peripheral blood, bone marrow, and spleens of CSF-1-deficient mice were also found in mice exclusively expressing csCSF-1. These data suggest that although csCSF-1 alone is able to normalize several aspects of development in IntroductionColony-stimulating factor 1 (CSF-1), also known as macrophage CSF, is the primary regulator of the mononuclear phagocyte lineage and regulates cells of the female reproductive tract. [1][2][3][4][5][6] All effects of CSF-1 are mediated by a high-affinity receptor tyrosine kinase 7-10 encoded by the c-fms proto-oncogene. 11 At least 5 mature human or mouse CSF-1 mRNAs (4.0 kb, 3.0 kb, 2.3 kb, 1.9 kb, and 1.6 kb) resulting from alternative splicing in exon 6 and the alternative usages of the 3Ј-untranslated region exons 9 and 10, [12][13][14][15][16][17][18] have been shown to encode 3 isoforms of the CSF-1 protein: a secreted glycoprotein, 19-21 a secreted proteoglycan, 22,23 and a biologically active membrane-spanning cell surface glycoprotein 18,[24][25][26][27][28][29] (for a review, see Stanley 30 ).The primary source of the circulating proteoglycan and glycoprotein CSF-1 is thought to be the endothelial cells that line the small blood vessels (for a review, see Roth and Stanley 31 ). CSF-1 is also synthesized locally, 32 for example, by osteoblasts 33,34 and by uterine epithelial cells. 3 It has been suggested that regulation at particular tissue sites is mediated by local synthesis of the membrane-spanning, cell surface CSF-1 (csCSF-1), and/or selective sequestration of the secreted proteoglycan CSF-1 (spCSF-1). 22,23,35 The csCSF-1 is encoded by a truncated mRNA in which part of the exon 6 sequence encoding the fragment containing the unique glycosaminoglycan addition site and the proteolytic cleavage sites used to release the secreted isoforms has been spliced out ( Figure 1A). 18,27 csCSF-1 is expressed in all cell types examined that express soluble CSF-1, including fib...

CSF-1 is the major regulator of tissue macrophage development and function. A GM-CSF-dependent, CSF-1 receptor (CSF-1R)-deficient F4/80(hi)Mac-1(+)Gr1(-)CD11c(+) bone marrow macrophage (BMM) line (MacCsf1r-/-) was developed to study the roles of the eight intracellular CSF-1R tyrosines phosphorylated upon receptor activation. Retroviral expression of the wild-type CSF-1R rescued the CSF-1-induced survival, proliferation, differentiation, and morphological characteristics of primary BMM. Mutation of all eight tyrosines failed to rescue, whereas the individual Y --> F mutants (544, 559, 697, 706, 721, 807, 921, 974) rescued these CSF-1-inducible phenotypes to varying degrees. The juxtamembrane domain Y559F and activation loop Y807F mutations severely compromised proliferation and differentiation, whereas Y706, Y721F, and Y974F mutations altered morphological responses, and Y706F increased differentiation. Despite their retention of significant in vitro tyrosine kinase activity, Y559F and Y807F mutants exhibited severely impaired in vivo receptor tyrosine phosphorylation, consistent with the existence of cellular mechanisms inhibiting CSF-1R tyrosine phosphorylation that are relieved by phosphorylation of these two sites. The MacCsf1r-/- macrophage line will facilitate genetic and proteomic approaches to CSF-1R structure/function studies in the major disease-related CSF-1R-expressing cell type.

Glycogen synthase kinase-3β (GSK-3β), a serine/threonine protein kinase, has been reported to show essential roles in molecular pathophysiology of many diseases. Mitochondrion is a dynamic organelle for producing cellular energy and determining cell fates. Stress-induced translocated GSK-3β may interact with mitochondrial proteins, including PI3K-Akt, PGC-1α, HK II, PKCε, components of respiratory chain, and subunits of mPTP. Mitochondrial pool of GSK-3β has been implicated in mediation of mitochondrial functions. GSK-3β exhibits the regulatory effects on mitochondrial biogenesis, mitochondrial bioenergetics, mitochondrial permeability, mitochondrial motility, and mitochondrial apoptosis. The versatile functions of GSK-3β might be associated with its wide range of substrates. Accumulative evidence demonstrates that GSK-3β inactivation may be potentially developed as the promising strategy in management of many diseases, such as Alzheimer’s disease (AD) and Parkinson’s disease (PD). Intensive efforts have been made for exploring GSK-3β inhibitors. Natural products provide us a great source for screening new lead compounds in inactivation of GSK-3β. The key roles of GSK-3β in mediation of mitochondrial functions are discussed in this review.