Apoptosis is implicated in the generation and resolution of inflammation in response to bacterial pathogens. All bacterial pathogens produce lipoproteins (BLPs), which trigger the innate immune response. BLPs were found to induce apoptosis in THP-1 monocytic cells through human Toll-like receptor-2 (hTLR2). BLPs also initiated apoptosis in an epithelial cell line transfected with hTLR2. In addition, BLPs stimulated nuclear factor-kappaB, a transcriptional activator of multiple host defense genes, and activated the respiratory burst through hTLR2. Thus, hTLR2 is a molecular link between microbial products, apoptosis, and host defense mechanisms.

Appreciation of the role of the gut microbiome in regulating vertebrate metabolism has exploded recently. However, the effects of gut microbiota on skeletal growth and homeostasis have only recently begun to be explored. Here, we report that colonization of sexually mature germ-free (GF) mice with conventional specific pathogen-free (SPF) gut microbiota increases both bone formation and resorption, with the net effect of colonization varying with the duration of colonization. Although colonization of adult mice acutely reduces bone mass, in long-term colonized mice, an increase in bone formation and growth plate activity predominates, resulting in equalization of bone mass and increased longitudinal and radial bone growth. Serum levels of insulin-like growth factor 1 (IGF-1), a hormone with known actions on skeletal growth, are substantially increased in response to microbial colonization, with significant increases in liver and adipose tissue IGF-1 production. Antibiotic treatment of conventional mice, in contrast, decreases serum IGF-1 and inhibits bone formation. Supplementation of antibiotic-treated mice with short-chain fatty acids (SCFAs), products of microbial metabolism, restores IGF-1 and bone mass to levels seen in nonantibiotic-treated mice. Thus, SCFA production may be one mechanism by which microbiota increase serum IGF-1. Our study demonstrates that gut microbiota provide a net anabolic stimulus to the skeleton, which is likely mediated by IGF-1. Manipulation of the microbiome or its metabolites may afford opportunities to optimize bone health and growth.

Breast cancer is one of the most common cancers in humans and will on average affect up to one in eight women in their lifetime in the United States and Europe1. The Women’s Health Initiative and the Million Women Study have shown that hormone replacement therapy is associated with an increased risk of incident and fatal breast cancer2,3. In particular, synthetic progesterone derivatives (progestins) such as medroxyprogesterone acetate (MPA), used in millions of women for hormone replacement therapy and contraceptives, markedly increase the risk of developing breast cancer. Here we show that the in vivo administration of MPA triggers massive induction of the key osteoclast differentiation factor RANKL (receptor activator of NF-κB ligand) in mammary-gland epithelial cells. Genetic inactivation of the RANKL receptor RANK in mammary-gland epithelial cells prevents MPA-induced epithelial proliferation, impairs expansion of the CD49fhi stem-cell-enriched population, and sensitizes these cells to DNA-damage-induced cell death. Deletion of RANK from the mammary epithelium results in a markedly decreased incidence and delayed onset of MPA-driven mammary cancer. These data show that the RANKL/RANK system controls the incidence and onset of progestin-driven breast cancer.

Quiescent adult stem cells reside in specialized niches where they become activated to proliferate and differentiate during tissue homeostasis and injury. How stem cell quiescence is governed is poorly understood. We report here that NFATc1 is preferentially expressed by hair follicle stem cells in their niche, where its expression is activated by BMP signaling upstream and it acts downstream to transcriptionally repress CDK4 and maintain stem cell quiescence. As stem cells become activated during hair growth, NFATc1 is downregulated, relieving CDK4 repression and activating proliferation. When calcineurin/NFATc1 signaling is suppressed, pharmacologically or via complete or conditional NFATc1 gene ablation, stem cells are activated prematurely, resulting in precocious follicular growth. Our findings may explain why patients receiving cyclosporine A for immunosuppressive therapy display excessive hair growth, and unveil a functional role for calcium-NFATc1-CDK4 circuitry in governing stem cell quiescence.

contributed equally to this workThe innate immune system uses Toll family receptors to signal for the presence of microbes and initiate host defense. Bacterial lipoproteins (BLPs), which are expressed by all bacteria, are potent activators of Toll-like receptor-2 (TLR2). Here we show that the adaptor molecule, myeloid differentiation factor 88 (MyD88), mediates both apoptosis and nuclear factorkB (NF-kB) activation by BLP-stimulated TLR2. Inhibition of the NF-kB pathway downstream of MyD88 potentiates apoptosis, indicating that these two pathways bifurcate at the level of MyD88. TLR2 signals for apoptosis through MyD88 via a pathway involving Fas-associated death domain protein (FADD) and caspase 8. Moreover, MyD88 binds FADD and is suf®cient to induce apoptosis. These data indicate that TLR2 is a novel`death receptor' that engages the apoptotic machinery without a conventional cytoplasmic death domain. Through TLR2, BLP induces the synthesis of the precursor of the pro-in¯ammatory cytokine interleukin-1b (IL-1b). Interestingly, BLP also activates caspase 1 through TLR2, resulting in proteolysis and secretion of mature IL-1b. These results indicate that caspase activation is an innate immune response to microbial pathogens, culminating in apoptosis and cytokine production.

Osteoporosis results from an imbalance in skeletal remodeling that favors bone resorption over bone formation. Bone matrix is degraded by osteoclasts, which differentiate from myeloid precursors in response to the cytokine RANKL. To gain insight into the transcriptional regulation of bone resorption during growth and disease, we generated a conditional knockout of the transcription factor nuclear factor of activated T cells c1 (Nfatc1). Deletion of Nfatc1 in young mice resulted in osteopetrosis and inhibition of osteoclastogenesis in vivo and in vitro. Transcriptional profiling revealed NFATc1 as a master regulator of the osteoclast transcriptome, promoting the expression of numerous genes needed for bone resorption. In addition, NFATc1 directly repressed osteoclast progenitor expression of osteoprotegerin, a decoy receptor for RANKL previously thought to be an osteoblast-derived inhibitor of bone resorption. "Cherubism mice", which carry a gain-of-function mutation in SH3-domain binding protein 2 (Sh3bp2), develop osteoporosis and widespread inflammation dependent on the proinflammatory cytokine, TNF-α. Interestingly, deletion of Nfatc1 protected cherubism mice from systemic bone loss but did not inhibit inflammation. Taken together, our study demonstrates that NFATc1 is required for remodeling of the growing and adult skeleton and suggests that NFATc1 may be an effective therapeutic target for osteoporosis associated with inflammatory states.

As the only cells definitively shown to degrade bone, osteoclasts are key mediators of skeletal diseases including osteoporosis. Bone forming osteoblasts, and hematopoietic and immune system cells, each influence osteoclast formation and function, but the reciprocal impact of osteoclasts on these cells is less well appreciated. Here, we highlight functions osteoclasts perform beyond bone resorption. First, we consider how osteoclast signals may contribute to bone formation by osteoblasts and the pathology of bone lesions, such as fibrous dysplasia and giant cell tumors. Second, we review the interaction of osteoclasts with the hematopoietic system, including the stem cell niche and adaptive immune cells. Connections between osteoclasts and other cells in the bone microenvironment are discussed within a clinically relevant framework. Keywordsosteoclast; osteoblast; bone remodeling; osteopetrosis; osteoporosis; PTH Bone remodeling and osteoclasts 101Bone is a composite tissue of protein and mineral, which undergoes continual remodeling to grow, heal damage, and regulate calcium and phosphate metabolism. This remodeling process is executed by the concerted and sequential effort of bone resorbing osteoclasts and bone forming osteoblasts, acting in what has been termed the basic multicellular unit (BMU) ( Figure 1A). Osteocytes, long-lived osteoblast-derived cells that reside within the bone matrix, monitor bone quality and stress, and coordinate remodeling through membrane bound and secreted factors. Skeletal integrity is maintained throughout the lifespan by matching bone formation and resorption, a process referred to as osteoclast:osteoblast 'coupling.' Coupling is thoroughly summarized in recent excellent reviews [1,2] and in Figure 1.Osteoclasts are multinucleated giant cells that differentiate from myeloid precursors under the influence of the cytokines macrophage colony stimulating factor (MCSF) and receptor © 2014 Elsevier Ltd. All rights reserved.Corresponding Author: Antonios O. Aliprantis, M.D., Ph.D., (AALIPRANTIS@PARTNERS.ORG). Publisher's Disclaimer: This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final citable form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. NIH Public Access NIH-PA Author ManuscriptNIH-PA Author Manuscript NIH-PA Author Manuscript activator of NF-κB ligand (RANKL) supplied by osteoblasts and/or osteocytes ( Figure 1A) [3]. A decoy receptor for RANKL called osteoprotegrin (OPG), which is also made by the osteoblast lineage, tempers osteoclast differentiation [4]. Osteoclasts degrade bone by the polarized secretion of proteolytic enzymes (e.g. cathepsin K) and acid, which hydrolyze and solubilize the organic and inorganic...

Clustering based on clinicophysiologic parameters yielded 4 stable and reproducible clusters that associate with different pathobiological pathways.