Children's everyday exposure to food marketing: an objective analysis using wearable cameras.

BACKGROUND: Over the past three decades the global prevalence of childhood overweight and obesity has increased by 47%. Marketing of energy-dense nutrient-poor foods and beverages contributes to this worldwide increase. Previous research on food marketing to children largely uses self-report, reporting by parents, or third-party observation of children's environments, with the focus mostly on single settings and/or media. This paper reports on innovative research, Kids'Cam, in which children wore cameras to examine the frequency and nature of everyday exposure to food marketing across multiple media and settings. METHODS: Kids'Cam was a cross-sectional study of 168 children (mean age 12.6 years, SD = 0.5) in Wellington, New Zealand. Each child wore a wearable camera on four consecutive days, capturing images automatically every seven seconds. Images were manually coded as either recommended (core) or not recommended (non-core) to be marketed to children by setting, marketing medium, and product category. Images in convenience stores and supermarkets were excluded as marketing examples were considered too numerous to count. RESULTS: On average, children were exposed to non-core food marketing 27.3 times a day (95% CI 24.8, 30.1) across all settings. This was more than twice their average exposure to core food marketing (12.3 per day, 95% CI 8.7, 17.4). Most non-core exposures occurred at home (33%), in public spaces (30%) and at school (19%). Food packaging was the predominant marketing medium (74% and 64% for core and non-core foods) followed by signs (21% and 28% for core and non-core). Sugary drinks, fast food, confectionary and snack foods were the most commonly encountered non-core foods marketed. Rates were calculated using Poisson regression. CONCLUSIONS: Children in this study were frequently exposed, across multiple settings, to marketing of non-core foods not recommended to be marketed to children. The study provides further evidence of the need for urgent action to reduce children's exposure to marketing of unhealthy foods, and suggests the settings and media in which to act. Such action is necessary if the Commission on Ending Childhood Obesity's vision is to be achieved.

Animal models of maternal high fat diet exposure and effects on metabolism in offspring: a meta-regression analysis.

Animal models of maternal high fat diet (HFD) demonstrate perturbed offspring metabolism although the effects differ markedly between models. We assessed studies investigating metabolic parameters in the offspring of HFD fed mothers to identify factors explaining these inter-study differences. A total of 171 papers were identified, which provided data from 6047 offspring. Data were extracted regarding body weight, adiposity, glucose homeostasis and lipidaemia. Information regarding the macronutrient content of diet, species, time point of exposure and gestational weight gain were collected and utilized in meta-regression models to explore predictive factors. Publication bias was assessed using Egger's regression test. Maternal HFD exposure did not affect offspring birthweight but increased weaning weight, final bodyweight, adiposity, triglyceridaemia, cholesterolaemia and insulinaemia in both female and male offspring. Hyperglycaemia was found in female offspring only. Meta-regression analysis identified lactational HFD exposure as a key moderator. The fat content of the diet did not correlate with any outcomes. There was evidence of significant publication bias for all outcomes except birthweight. Maternal HFD exposure was associated with perturbed metabolism in offspring but between studies was not accounted for by dietary constituents, species, strain or maternal gestational weight gain. Specific weaknesses in experimental design predispose many of the results to bias.

High-fat diet disrupts metabolism in two generations of rats in a parent-of-origin specific manner.

Experimental and epidemiological evidence demonstrate that ancestral diet might contribute towards offspring health. This suggests that nutrition may be able to modify genetic or epigenetic information carried by germ cells (GCs). To examine if a parental high fat diet (HFD) influences metabolic health in two generations of offspring, GC-eGFP Sprague Dawley rats were weaned onto HFD (45% fat) or Control Diet (CD; 10% fat). At 19 weeks, founders (F0) were bred with controls, establishing the F1 generation. HFD resulted in 9.7% and 14.7% increased weight gain in male and female F0 respectively. F1 offspring of HFD mothers and F1 daughters of HFD-fed fathers had increased weight gain compared to controls. F1 rats were bred with controls at 19 weeks to generate F2 offspring. F2 male offspring derived from HFD-fed maternal grandfathers exhibited increased adiposity, plasma leptin and luteinising hormone to testosterone ratio. Despite transmission via the founding male germline, we did not find significant changes in the F0 intra-testicular GC transcriptome. Thus, HFD consumption by maternal grandfathers results in a disrupted metabolic and reproductive hormone phenotype in grandsons in the absence of detectable changes in the intra-testicular GC transcriptome.

Wnt signalling in pituitary development and tumorigenesis.

Wnt signalling is activated in both pituitary organogenesis and its mature function. Wnt ligands and Wnt signalling pathways are critical for the regulation of the formation of the pituitary. In the mature pituitary, Wnt signalling pathways control cell activity and may stimulate cell proliferation in both physiological and pathological processes. This review compares Wnt signalling pathways active in the developing and mature pituitary and explores how this gives us further insight into the development of pituitary adenomas.

How are osteoclasts induced to resorb bone?

Although much is known about how osteoclasts are formed, we know little about how they are activated, or how they recognize bone as the substrate appropriate for resorption. Bone mineral is considered to be essential to this recognition process, but a "mineral receptor" has never been identified. Recently, we found that resorptive behavior, as judged by the formation of ruffled borders and actin rings, occurs on ordinary tissue culture substrates if they are first coated with vitronectin. Similarly, vitronectin-coated substrates induce osteoclasts to secrete tartrate-resistant acid phosphatase and to form podosome belts, and to make resorption trails in the protein that coat the substrate. The same applies to bone mineral, which only induces resorptive behavior if coated with vitronectin. In contrast, fibronectin has none of these effects, despite inducing adhesion and spreading. It appears that osteoclasts recognize bone as the substrate appropriate for resorption through the high affinity of vitronectin-receptor ligands for bone mineral.

The birth of the osteoclast.

Thirty-five years ago it had become clear that the osteoclast was not a bone cell but an immigrant into bone, derived from the hemopoietic system. Among hemopoietic cells, mononuclear phagocytes seemed the most likely precursors. However, for the progeny of wandering cells such as those to achieve nonrandom localization implies control by the local bone cells (cells of the osteoblastic lineage). To test this idea, we extracted osteoclasts from bone and observed their behavior in culture. We noted that calcitonin induced a striking shape change, which reflected suppression of cytoplasmic motility. Because bone resorption is likely to depend on motile processes, we used this response to infer the regulation of osteoclasts by systemic and local hormones and osteoblastic cells. We went on to provide direct evidence for the predominantly osteoblastic regulation of osteoclasts by measuring the ability of isolated osteoclasts to resorb the surface of bone slices.

Tumor necrosis factor-alpha mediates osteopenia caused by depletion of antioxidants.

We recently found that estrogen deficiency leads to a lowering of thiol antioxidant defenses in rodent bone. Moreover, administration of agents that increase the concentration in bone of glutathione, the main intracellular antioxidant, prevented estrogen-deficiency bone loss, whereas depletion of glutathione by buthionine sulfoximine (BSO) administration provoked substantial bone loss. It has been shown that the estrogen-deficiency bone loss is dependent on TNFalpha signaling. Therefore, a model in which estrogen deficiency causes bone loss by lowering antioxidant defenses predicts that the osteopenia caused by lowering antioxidant defenses should similarly depend on TNFalpha signaling. We found that the loss of bone caused by either BSO administration or ovariectomy was inhibited by administration of soluble TNFalpha receptors and abrogated in mice deleted for TNFalpha gene expression. In both circumstances, lack of TNFalpha signaling prevented the increase in bone resorption and the deficit in bone formation that otherwise occurred. Thus, depletion of thiol antioxidants by BSO, like ovariectomy, causes bone loss through TNFalpha signaling. Furthermore, in ovariectomized mice treated with soluble TNFalpha receptors, thiol antioxidant defenses in bone remained low, despite inhibition of bone loss. This suggests that the low levels of antioxidants in bone seen after ovariectomy are the cause, rather than the effect, of the increased resorption. These experiments are consistent with a model for estrogen-deficiency bone loss in which estrogen deficiency lowers thiol antioxidant defenses in bone cells, thereby increasing reactive oxygen species levels, which in turn induce expression of TNFalpha, which causes loss of bone.

Parathyroid hormone activates adhesion in bone marrow stromal precursor cells.

The ability of parathyroid hormone (PTH) to enhance bone formation has recently been exploited in the treatment of osteoporosis. However, the underlying mechanisms are unknown. Osteoblasts, the bone-forming cells, derive from multipotential bone marrow stromal precursors called colony-forming units-fibroblastic (CFU-F) upon culture ex vivo. Adhesion of such stromal precursors to bone is likely to be an early event in the anabolic response of bone to PTH. To test this, we measured the number of CFU-F that could be extracted from murine bone marrow after administration of an anabolic dose of PTH. We found that a very early response is a dramatic reduction, starting within 2 h, in the number of CFU-F that could be extracted from their bone marrow. We then tested whether PTH has the ability to activate adhesion of CFU-F in vitro. For this, bone marrow cells were incubated in PTH for varying times. Non-adherent cells were then removed, and the adherent cells were incubated in PTH-free medium for 14 days to assess, as colony formation, the number of CFU-F that had adhered in the preceding period. We found that incubation in PTH caused a substantial increase in the number of CFU-F that adhered within 24 h. This increase was abrogated by peptidic inhibitors of integrins. The increase did not seem to be mediated through a PTH-induced increase in interleukin-6, since interleukin-6 had no effect on CFU-F numbers when substituted for PTH. Similarly, adhesion was unaffected by incubation of bone marrow cells in dibutyryl cyclic AMP, nor by inhibitors or donors of nitric oxide. However, activation of CFU-F in vitro by PTH was strongly inhibited by indomethacin and mimicked by prostaglandin E(2), and indomethacin reversed the PTH-mediated reduction of CFU-F that could be extracted from mouse bone marrow. These results suggested that PTH rapidly activates adhesion of CFU-F to plastic or bone surfaces. This activation may represent an early event in the anabolic response of bone cells to PTH.

Mitf-PU.1 interactions with the tartrate-resistant acid phosphatase gene promoter during osteoclast differentiation.

It has been postulated that the transcription factors micropthalmia associated factor (Mitf) and PU.1 interact with the tartrate-resistant acid phosphatase (TRAP) gene promoter and activate TRAP gene expression in osteoclasts. However, studies on the interaction of these factors with the TRAP promoter employing nuclear extracts from osteoclasts and osteoclast precursors have not been reported. We therefore treated murine mononuclear phagocyte cells with various cytokines to generate cultures of osteoclasts and macrophagic cells with high or low potential to form osteoclasts. The presence of Mitf and PU.1 in nuclear extracts from these cultures and the ability of these factors to bind to the TRAP promoter was then assessed. We demonstrate that Mitf and a related factor, TFE3, are present in nuclear extracts from all cultures and bind the TRAP promoter. While PU.1 is present in nuclear extracts from all cultures, it does not significantly interact with a putative binding site in the TRAP promoter. These results suggest Mitf and PU.1 interactions with the TRAP promoter are not responsible for the specific activation of TRAP gene expression in osteoclasts.

FLT3 ligand can substitute for macrophage colony-stimulating factor in support of osteoclast differentiation and function.

Although bone resorption and osteoclast numbers are reduced in osteopetrotic (op/op) mice, osteoclasts are nevertheless present and functional, despite the absence of macrophage colony-stimulating factor (M-CSF). This suggests that alternative factors can partly compensate for the crucial actions of M-CSF in osteoclast induction. It was found that when nonadherent bone marrow cells were incubated in RANKL with Flt3 ligand (FL) without exogenous M-CSF, tartrate-resistance acid phosphatase (TRAP)-positive cells were formed, and bone resorption occurred. Without FL, only macrophagelike TRAP-negative cells were present. Granulocyte-macrophage CSF, stem cell factor, interleukin-3, and vascular endothelial growth factor could not similarly replace the need for M-CSF. TRAP-positive cell induction in FL was not due to synergy with M-CSF produced by the bone marrow cells themselves because FL also enabled their formation from the hemopoietic cells of op/op mice, which lack any M-CSF. FL appeared to substitute for M-CSF by supporting the differentiation of adherent cells that express mRNA for RANK and responsiveness to RANKL. To determine whether FL can account for the compensation for M-CSF deficiency that occurs in vivo, FL signaling was blockaded in op/op mice by the injection of soluble recombinant Flt3. It was found that the soluble receptor induced a substantial decrease in osteoclast number, strongly suggesting that FL is responsible for the partial compensation for M-CSF deficiency that occurs in these mice.

Neuroadapted yellow fever virus 17D: genetic and biological characterization of a highly mouse-neurovirulent virus and its infectious molecular clone.

A neuroadapted strain of yellow fever virus (YFV) 17D derived from a multiply mouse brain-passaged virus (Porterfield YF17D) was additionally passaged in SCID and normal mice. The virulence properties of this virus (SPYF) could be distinguished from nonneuroadapted virus (YF5.2iv, 17D infectious clone) by decreased average survival time in SCID mice after peripheral inoculation, decreased average survival time in normal adult mice after intracerebral inoculation, and occurrence of neuroinvasiveness in normal mice. SPYF exhibited more efficient growth in peripheral tissues of SCID mice than YF5.2iv, resulting in a more rapid accumulation of virus burden, but with low-titer viremia, at the time of fatal encephalitis. In cell culture, SPYF was less efficient in replication than YF5.2iv in all cell lines tested. The complete nucleotide sequence of SPYF revealed 29 nucleotide substitutions relative to YF5.2iv, and these were distributed throughout the genome. There were a total of 13 predicted amino acid substitutions, some of which correspond to known differences among the Asibi, French viscerotropic virus, French neurotropic vaccine, and YF17D vaccine strains. The envelope (E) protein contained five substitutions, within all three functional domains. Substitutions were also present in regions encoding the NS1, NS2A, NS4A, and NS5 proteins and in the 3' untranslated region (UTR). Construction of YFV harboring all of the identified coding nucleotide substitutions and those in the 3' UTR yielded a virus whose cell culture and pathogenic properties, particularly neurovirulence and neuroinvasiveness for SCID mice, generally resembled those of the original SPYF isolate. These findings implicate the E protein and possibly other regions of the genome as virulence determinants during pathogenesis of neuroadapted YF17D virus in mice. The determinants affect replication efficiency in both neural and extraneural tissues of the mouse and confer some limited host-range differences in cultured cells of nonmurine origin.

Yellow fever virus encephalitis: properties of the brain-associated T-cell response during virus clearance in normal and gamma interferon-deficient mice and requirement for CD4+ lymphocytes.

Viral encephalitis caused by neuroadapted yellow fever 17D virus (PYF) was studied in parental and gamma interferon (IFN-gamma)-deficient (IFN-gamma knockout [GKO]) C57BL/6 mice. The T-cell responses which enter the brain during acute fatal encephalitis of nonimmunized mice, as well as nonfatal encephalitis of immunized mice, were characterized for relative proportions of CD4+ and CD8+ cells, their proliferative responses, and antigen-specific expression of cytokines during stimulation in vitro. Unimmunized mice accumulated only low levels of T cells within the brain during fatal disease, whereas the brains of immunized mice contained higher levels of both T-cell subsets in response to challenge, with CD8+ cells increased relative to the CD4+ subset. The presence of T cells correlated with the time at which virus was cleared from the central nervous system in both parental and GKO mice. Lymphocytes isolated from the brains of challenged immunized mice failed to proliferate in vitro in response to T-cell mitogens or viral antigens; however, IFN-gamma, interleukin 4 (IL-4), and, to a lesser extent, IL-2 were detectable after stimulation. The levels of IFN-gamma, but not IL-2 or IL-4, were augmented in response to viral antigen, and this specificity was detectable in the CD4+ compartment. When tested for the ability to survive both immunization and challenge with PYF virus, GKO and CD8 knockout mice did not differ from parental mice (80 to 85% survival), although GKO mice exhibited a defect in virus clearance. In contrast, CD4 knockout and Igh-6 mice were unable to resist challenge. The data implicate antibody in conjunction with CD4+ lymphocytes bearing a Th1 phenotype as the critical factors involved in virus clearance in this model.

Molecular basis for attenuation of neurovirulence of a yellow fever Virus/Japanese encephalitis virus chimera vaccine (ChimeriVax-JE).

A yellow fever virus (YFV)/Japanese encephalitis virus (JEV) chimera in which the structural proteins prM and E of YFV 17D are replaced with those of the JEV SA14-14-2 vaccine strain is under evaluation as a candidate vaccine against Japanese encephalitis. The chimera (YFV/JEV SA14-14-2, or ChimeriVax-JE) is less neurovirulent than is YFV 17D vaccine in mouse and nonhuman primate models (F. Guirakhoo et al., Virology 257:363-372, 1999; T. P. Monath et al., Vaccine 17:1869-1882, 1999). Attenuation depends on the presence of the JEV SA14-14-2 E protein, as shown by the high neurovirulence of an analogous YFV/JEV Nakayama chimera derived from the wild JEV Nakayama strain (T. J. Chambers, A. Nestorowicz, P. W. Mason, and C. M. Rice, J. Virol. 73:3095-3101, 1999). Ten amino acid differences exist between the E proteins of ChimeriVax-JE and the YFV/JEV Nakayama virus, four of which are predicted to be neurovirulence determinants based on various sequence comparisons. To identify residues that are involved in attenuation, a series of intratypic YFV/JEV chimeras containing either single or multiple amino acid substitutions were engineered and tested for mouse neurovirulence. Reversions in at least three distinct clusters were required to restore the neurovirulence typical of the YFV/JEV Nakayama virus. Different combinations of cluster-specific reversions could confer neurovirulence; however, residue 138 of the E protein (E(138)) exhibited a dominant effect. No single amino acid reversion produced a phenotype significantly different from that of the ChimeriVax-JE parent. Together with the known genetic stability of the virus during prolonged cell culture and mouse brain passage, these findings support the candidacy of this experimental vaccine as a novel live-attenuated viral vaccine against Japanese encephalitis.

Interferon-gamma directly inhibits TRANCE-induced osteoclastogenesis.

The immune system has profound effects on bone remodeling. IFN-gamma, a major product of immune cells, potently inhibits bone resorption, but its mechanism of action is unknown. We found in cultures of stroma-free mononuclear precursors that IFN-gamma strongly suppresses TRANCE/RANKL-induced osteoclast formation in a dose-dependent manner. This direct effect on osteoclast progenitors was not due to stimulation of NO production by IFN-gamma, as the NOS inhibitors 1400W and L-NAME were unable to reverse the suppression. However, TGFbeta(1), which has opposing actions to IFN-gamma on diverse cellular functions, was able to antagonize the effect of IFN-gamma. This suggests that IFN-gamma prevents osteoclast formation by actively directing the differentiation of osteoclastic progenitors toward an alternative cytocidal lineage to the osteoclast.

TGF-beta 1 and IFN-gamma direct macrophage activation by TNF-alpha to osteoclastic or cytocidal phenotype.

TNF-related activation-induced cytokine (TRANCE; also called receptor activator of NF-kappaB ligand (RANKL), osteoclast differentiation factor (ODF), osteoprotegerin ligand (OPGL), and TNFSF11) induces the differentiation of progenitors of the mononuclear phagocyte lineage into osteoclasts in the presence of M-CSF. Surprisingly, in view of its potent ability to induce inflammation and activate macrophage cytocidal function, TNF-alpha has also been found to induce osteoclast-like cells in vitro under similar conditions. This raises questions concerning both the nature of osteoclasts and the mechanism of lineage choice in mononuclear phagocytes. We found that, as with TRANCE, the macrophage deactivator TGF-beta(1) strongly promoted TNF-alpha-induced osteoclast-like cell formation from immature bone marrow macrophages. This was abolished by IFN-gamma. However, TRANCE did not share the ability of TNF-alpha to activate NO production or heighten respiratory burst potential by macrophages, or induce inflammation on s.c. injection into mice. This suggests that TGF-beta(1) promotes osteoclast formation not only by inhibiting cytocidal behavior, but also by actively directing TNF-alpha activation of precursors toward osteoclasts. The osteoclast appears to be an equivalent, alternative destiny for precursors to that of cytocidal macrophage, and may represent an activated variant of scavenger macrophage.

Yellow fever virus NS2B-NS3 protease: charged-to-alanine mutagenesis and deletion analysis define regions important for protease complex formation and function.

Charged-to-alanine substitutions and deletions within the yellow fever virus NS2B-NS3(181) protease were analyzed for effects on protease function. During cell-free translation of NS2B-3(181) polyproteins, mutations at three charge clusters markedly impaired cis cleavage activity: a single N-terminal cluster in the conserved domain of NS2B (residues ELKK(52-55)) and two in NS3 (ED(21-22), and residue H(47)). These mutations inhibited other protease-dependent cleavages of a transiently expressed nonstructural polyprotein, although differential effects occurred. NS2B and NS3(181) proteins harboring these mutations were impaired in their ability to associate for trans cleavage activity. N-terminal deletions in NS3 also implicated residues ED(21-22) in the association with NS2B. Deletions within NS2B revealed that the conserved domain alone provided minimal cofactor activity, with optimal function requiring both flanking hydrophobic regions. NS2B-3(181)- and NS3(181)-green fluorescent protein fusion proteins were used to determine the intracellular distribution of the protease complex. The former localized in membrane-based vesicular structures, whereas the latter localized poorly. The data suggest that NS2B-NS3 complex formation requires charge interactions involving the N-terminus of the conserved domain of NS2B and 22 N-terminal residues of NS3. A role for the putative transmembrane regions of NS2B in targeting of NS3 to intracellular membranes is also suggested.

Regulation of the differentiation and function of osteoclasts.

The osteoclast is the cell that resorbs bone. It has been known for many years that its formation and function are regulated by cells of the osteoblastic lineage. Recently the molecular basis for this regulation was identified; osteoblastic cells induce osteoclastic differentiation and resorptive activity through expression of tumour necrosis factor (TNF) activation-induced cytokine (TRANCE) (also known as RANKL, ODF, OPGL, and TNFSF11), a novel membrane-inserted member of the TNF superfamily. Osteoclastic regulation is assisted through secretion of an inhibitor, osteoprotegerin (OPG) (OCIF, TNFRSF11B), a soluble (decoy) receptor for TRANCE. Osteoclast formation and survival also depend on and are substantially enhanced by transforming growth factor-beta (TGF-beta), which is abundant in bone matrix. Surprisingly, not only TRANCE but also TNF-alpha can induce osteoclast formation in vitro from bone marrow-derived mononuclear phagocytes, especially in the presence of TGF-beta. Whether or not TNF-alpha does the same in vivo, its ability to generate osteoclasts in vitro has significant implications regarding the nature of osteoclasts and their relationship to other mononuclear phagocytes, and a possible wider role for TRANCE in macrophage pathobiology. A hypothesis is presented in which the osteoclast is a mononuclear phagocyte directed towards a debriding function by TGF-beta, activated for this function by TRANCE, and induced to become specifically osteoclastic by the characteristics of the substrate or signals from bone cells that betoken such characteristics.

Activation of osteoclasts by interleukin-1: divergent responsiveness in osteoclasts formed in vivo and in vitro.

Recently, it has been found that osteoclasts are induced and activated by osteoblastic cells through expression of receptor activator NF-kB ligand (RANKL), and that soluble recombinant RANKL, with M-CSF, can replace the need for osteoblastic cells in osteoclast formation. We exploited this opportunity to compare the responsiveness of osteoclast-like cells (OCL) formed in vitro in the absence of osteoblasts, with that of osteoclasts ex vivo. We found that while OCL responded to several hormones and cytokines like ex vivo osteoclasts, their responsiveness to interleukin-1 (IL-1) was fundamentally different: IL1 directly stimulated actin ring formation in OCL, but had no effect on actin rings or survival in osteoclasts ex vivo unless osteoblastic cells were present. This difference could not be attributed to the use of plastic culture substrates for OCL formation, nor to osteoblastic contamination, and did not seem to be mediated by the macrophages that form in OCL cultures. To understand the mechanisms by which IL-1 induces bone loss, it will need to be determined whether or not IL-1-responsive OCLs have a counterpart in vivo. Whichever is the case, our data suggest that the behavior of osteoclasts formed in culture will not always predict that of osteoclasts in vivo.

Osteoclast lineage commitment of bone marrow precursors through expression of membrane-bound TRANCE.

Osteoclast formation from hemopoietic precursors is induced by TRANCE (also called RANKL, ODF, and OPGL), a membrane-bound ligand expressed by bone marrow stromal cells. Because soluble recombinant TRANCE is a suboptimal osteoclastogenic stimulus, and to eliminate the need for such dependence on stromal cells, membrane-bound TRANCE was expressed in hematopoietic precursors using retroviral gene transfer. Four TRANCE-expressing osteoclast cell lines were established that continuously generate large numbers of multinucleated cells and express tartrate-resistant acid phosphatase and calcitonin receptors. The multinuclear cells are long-lived and either fuse continuously with each other and with mononuclear cells to form enormous syncytia, or separate to form daughter multinuclear cells. When formed on bone, but not on plastic, the majority of multinuclear cells develop actin rings on bone, and resorb bone, suggesting that bone matrix may provide additional signals that facilitate osteoclastic functional maturation. Surprisingly, multinuclear cells originate from fusion of proliferating mononuclear cells that strongly express the mature macrophage markers F4/80 and Fc receptor, which are not expressed by osteoclasts. These results indicate that osteoclasts can be derived from F4/80-positive and Fc receptor-positive cells, and that TRANCE induces osteoclastic differentiation partly by suppressing the macrophage phenotype.