Inorganic phosphate induces cancer cell mediated angiogenesis dependent on forkhead box protein C2 (FOXC2) regulated osteopontin expression.

Recent studies in both rodents and humans suggest that elevated serum phosphorus, in the context of normal renal function, potentiates, or exacerbates pathologies associates with cardiovascular disease, bone metabolism, and cancer. Our recent microarray studies identified the potent stimulation of pro-angiogenic genes such as forkhead box protein C2 (FOXC2), osteopontin, and Vegfα, among others in response to elevated inorganic phosphate (Pi). Increased angiogenesis and neovascularization are important events in tumor growth and the progression to malignancy and FOXC2 has recently been identified as a potential transcriptional regulator of these processes. In this study we addressed the possibility that a high Pi environment would increase the angiogenic potential of cancer cells through a mechanism requiring FOXC2. Our studies utilized lung and breast cancer cell lines in combination with the human umbilical vascular endothelial cell (HUVEC) vessel formation model to better understand the mechanism(s) by which a high Pi environment might alter cancer progression. Exposure of cancer cells to elevated Pi stimulated expression of FOXC2 and conditioned medium from the Pi-stimulated cancer cells stimulated migration and tube formation in the HUVEC model. Mechanistically, we define the requirement of FOXC2 for Pi-induced osteopontin (OPN) expression and secretion from cancer cells as necessary for the angiogenic response. These studies reveal for the first time that cancer cells grown in a high Pi environment promote migration of endothelial cells and tube formation and in so doing identify a novel potential therapeutic target to reduce tumor progression.

Bioactive silica nanoparticles reverse age-associated bone loss in mice.

We recently reported that in vitro, engineered 50nm spherical silica nanoparticles promote the differentiation and activity of bone building osteoblasts but suppress bone-resorbing osteoclasts. Furthermore, these nanoparticles promote bone accretion in young mice in vivo. We have now investigated the capacity of these nanoparticles to reverse bone loss in aged mice, a model of human senile osteoporosis. Aged mice received nanoparticles weekly and bone mineral density (BMD), bone structure, and bone turnover were quantified. Our data revealed a significant increase in BMD, bone volume, and biochemical markers of bone formation. Biochemical and histological examinations failed to identify any abnormalities caused by nanoparticle administration. Our studies demonstrate that silica nanoparticles effectively blunt and reverse age-associated bone loss in mice by a mechanism involving promotion of bone formation. The data suggest that osteogenic silica nanoparticles may be a safe and effective therapeutic for counteracting age-associated bone loss. FROM THE CLINICAL EDITOR: Osteoporosis poses a significant problem in the society. Based on their previous in-vitro findings, the authors' group investigated the effects of spherical silica nanoparticles in reversing bone loss in a mouse model of osteoporosis. The results showed that intra-peritoneal injections of silica nanoparticles could increase bone mineral density, with little observed toxic side effects. This novel method may prove important in future therapy for combating osteoporosis.

An integrated understanding of the physiological response to elevated extracellular phosphate.

Recent studies have suggested that changes in serum phosphate levels influence pathological states associated with aging such as cancer, bone metabolism, and cardiovascular function, even in individuals with normal renal function. The causes are only beginning to be elucidated but are likely a combination of endocrine, paracrine, autocrine, and cell autonomous effects. We have used an integrated quantitative biology approach, combining transcriptomics and proteomics to define a multi-phase, extracellular phosphate-induced, signaling network in pre-osteoblasts as well as primary human and mouse mesenchymal stromal cells. We identified a rapid mitogenic response stimulated by elevated phosphate that results in the induction of immediate early genes including c-fos. The mechanism of activation requires FGF receptor signaling followed by stimulation of N-Ras and activation of AP-1 and serum response elements. A distinct long-term response also requires FGF receptor signaling and results in N-Ras activation and expression of genes and secretion of proteins involved in matrix regulation, calcification, and angiogenesis. The late response is synergistically enhanced by addition of FGF23 peptide. The intermediate phase results in increased oxidative phosphorylation and ATP production and is necessary for the late response providing a functional link between the phases. Collectively, the results define elevated phosphate, as a mitogen and define specific mechanisms by which phosphate stimulates proliferation and matrix regulation. Our approach provides a comprehensive understanding of the cellular response to elevated extracellular phosphate, functionally connecting temporally coordinated signaling, transcriptional, and metabolic events with changes in long-term cell behavior.

Bioactive silica nanoparticles target autophagy, NF-κB, and MAPK pathways to inhibit osteoclastogenesis.

Spherical 50 nm silica-based nanoparticles (SiNPs) promote healthy bone homeostasis and maintenance by supporting bone forming osteoblast lineage cells while simultaneously inhibiting the differentiation of bone resorbing osteoclasts. Previous work demonstrated that an intraperitoneal injection of SiNPs in healthy mice - both young and old - increased bone density and quality, suggesting the possibility that SiNPs represent a dual action therapeutic. However, the underlying mechanisms governing the osteoclast response to SiNPs have yet to be fully explored and defined. Therefore, the goals of this study were to investigate the cellular and molecular mechanisms by which SiNPs inhibit osteoclastogenesis. SiNPs strongly inhibited RANKL-induced osteoclast differentiation within the first hours and concomitantly inhibited early transcriptional regulators such as Nfatc1. SiNPs simultaneously stimulated expression of autophagy related genes p62 and LC3ß dependent on ERK1/2 signaling pathway. Intriguingly, SiNPs were found to stimulate autophagosome formation while inhibiting the autophagic flux necessary for RANKL-stimulated osteoclast differentiation, resulting in the inhibition of both the canonical and non-canonical NF-κB signaling pathways and stabilizing TRAF3. These results suggest a model in which SiNPs inhibit osteoclastogenesis by inhibiting the autophagic machinery and RANKL-dependent functionality. This mechanism of action defines a novel therapeutic strategy for inhibiting osteoclastogenesis.

Dietary phosphorus consumption alters T cell populations, cytokine production, and bone volume in mice.

The intake of dietary phosphate far exceeds recommended levels; however, the long-term health consequences remain relatively unknown. Here, the chronic physiological response to sustained elevated and reduced dietary phosphate consumption was investigated in mice. Although serum phosphate levels were brought into homeostatic balance, the prolonged intake of a high-phosphate diet dramatically and negatively impacted bone volume; generated a sustained increase in the phosphate responsive circulating factors FGF23, PTH, osteopontin and osteocalcin; and produced a chronic low-grade inflammatory state in the BM, marked by increased numbers of T cells expressing IL-17a, RANKL, and TNF-α. In contrast, a low-phosphate diet preserved trabecular bone while increasing cortical bone volume over time, and it reduced inflammatory T cell populations. Cell-based studies identified a direct response of T cells to elevated extracellular phosphate. Neutralizing antibodies against proosteoclastic cytokines RANKL, TNF-α, and IL-17a blunted the high-phosphate diet-induced bone loss identifying bone resorption as a regulatory mechanism. Collectively, this study illuminates that habitual consumption of a high-phosphate diet in mice induces chronic inflammation in bone, even in the absence of elevated serum phosphate. Furthermore, the study supports the concept that a reduced phosphate diet may be a simple yet effective strategy to reduce inflammation and improve bone health during aging.

Sclerostin small-molecule inhibitors promote osteogenesis by activating canonical Wnt and BMP pathways.

Background: The clinical healing environment after a posterior spinal arthrodesis surgery is one of the most clinically challenging bone-healing environments across all orthopedic interventions due to the absence of a contained space and the need to form de novo bone. Our group has previously reported that sclerostin in expressed locally at high levels throughout a developing spinal fusion. However, the role of sclerostin in controlling bone fusion remains to be established. Methods: We computationally identified two FDA-approved drugs, as well as a single novel small-molecule drug, for their ability to disrupt the interaction between sclerostin and its receptor, LRP5/6. The drugs were tested in several in vitro biochemical assays using murine MC3T3 and MSCs, assessing their ability to (1) enhance canonical Wnt signaling, (2) promote the accumulation of the active (non-phosphorylated) form of ß-catenin, and (3) enhance the intensity and signaling duration of BMP signaling. These drugs were then tested subcutaneously in rats as standalone osteoinductive agents on plain collagen sponges. Finally, the top drug candidates (called VA1 and C07) were tested in a rabbit posterolateral spine fusion model for their ability to achieve a successful fusion at 6 wk. Results: We show that by controlling GSK3b phosphorylation our three small-molecule inhibitors (SMIs) simultaneously enhance canonical Wnt signaling and potentiate canonical BMP signaling intensity and duration. We also demonstrate that the SMIs produce dose-dependent ectopic mineralization in vivo in rats as well as significantly increase posterolateral spine fusion rates in rabbits in vivo, both as standalone osteogenic drugs and in combination with autologous iliac crest bone graft. Conclusions: Few if any osteogenic small molecules possess the osteoinductive potency of BMP itself - that is, the ability to form de novo ectopic bone as a standalone agent. Herein, we describe two such SMIs that have this unique ability and were shown to induce de novo bone in a stringent in vivo environment. These SMIs may have the potential to be used in novel, cost-effective bone graft substitutes for either achieving spinal fusion or in the healing of critical-sized fracture defects. Funding: This work was supported by a Veteran Affairs Career Development Award (IK2-BX003845).

Tissue-specific gene targeting by the multiprotein mammalian DREAM complex.

The mammalian DP, RB-like, E2F, and MuvB-like proteins (DREAM) complex, whose key components include p130 and E2F4, plays a fundamental role in repression of cell cycle-specific genes during growth arrest. Mammalian DREAM is well conserved with Drosophila and Caenorhabditis elegans complexes that repress pivotal developmental genes, but the mammalian complex has been thought to exist only in quiescent cells and not to be linked with development. However, new findings here identify tissue-specific promoters repressed by DREAM in proliferating precursors, revealing a new connection between control of growth arrest and terminal differentiation. Mechanistically, tissue-specific promoter occupation by DREAM is dependent on the integrity of a repressor form of the SWI/SNF chromatin-remodeling complex.

Bioactive silica-based nanoparticles stimulate bone-forming osteoblasts, suppress bone-resorbing osteoclasts, and enhance bone mineral density in vivo.

Bone is a dynamic tissue that undergoes renewal throughout life in a process whereby osteoclasts resorb worn bone and osteoblasts synthesize new bone. Imbalances in bone turnover lead to bone loss and development of osteoporosis and ultimately fracture, a debilitating condition with high morbidity and mortality. Silica is a ubiquitous biocontaminant that is considered to have high biocompatibility. The authors report that silica nanoparticles (NPs) mediate potent inhibitory effects on osteoclasts and stimulatory effects on osteoblasts in vitro. The mechanism of bioactivity is a consequence of an intrinsic capacity to antagonize activation of NF-κB, a signal transduction pathway required for osteoclastic bone resorption but inhibitory to osteoblastic bone formation. We further demonstrate that silica NPs promote a significant enhancement of bone mineral density (BMD) in mice in vivo, providing a proof of principle for the potential application of silica NPs as a pharmacological agent to enhance BMD and protect against bone fracture.

Modulating phosphate consumption, a novel therapeutic approach for the control of cancer cell proliferation and tumorigenesis.

Phosphorus, often in the form of inorganic phosphate (Pi), is critical to cellular function on many levels; it is required as an integral component of kinase signaling, in the formation and function of DNA and lipids, and energy metabolism in the form of ATP. Accordingly, crucial aspects of cell mitosis - such as DNA synthesis and ATP energy generation - elevate the cellular requirement for Pi, with rapidly dividing cells consuming increased levels. Mechanisms to sense, respond, acquire, accumulate, and potentially seek Pi have evolved to support highly proliferative cellular states such as injury and malignant transformation. As such, manipulating Pi availability to target rapidly dividing cells presents a novel strategy to reduce or prevent unrestrained cell growth. Currently, limited knowledge exists regarding how modulating Pi consumption by pre-cancerous cells might influence the initiation of aberrant growth during malignant transformation, and if reducing the bioavailability or suppressing Pi consumption by malignant cells could alter tumorigenesis. The concept of targeting Pi-regulated pathways and/or consumption by pre-cancerous or tumor cells represents a novel approach to cancer prevention and control, although current data remains insufficient as to rigorously assess the therapeutic value and physiological relevance of this strategy. With this review, we present a critical evaluation of the paradox of how an element critical to essential cellular functions can, when available in excess, influence and promote a cancer phenotype. Further, we conjecture how Pi manipulation could be utilized as a therapeutic intervention, either systemically or at the cell level, to ultimately suppress or treat cancer initiation and/or progression.

Toxicity and clearance of intratracheally administered multiwalled carbon nanotubes from murine lung.

Carbon nanotubes (CNT) are known to have widespread industrial applications; however, several reports indicated that these compounds may be associated with adverse effects in humans. In this study, multiwalled carbon nanotubes were administered to murine lungs intratracheally to determine whether acute and chronic pulmonary toxicity occurred. In particular, pristine multiwalled carbon nanotubes (PMWCNT) and acid-treated multiwalled carbon nanotubes (TMWCNT) were used in this study. In broncheoalveolar lavage fluid (BALF) cell analysis, PMWCNT induced more severe acute inflammatory cell recruitment than TMWCNT. Histopathologically, both PMWCNT and TMWCNT induced multifocal inflammatory granulomas in a dose-dependent manner. The observed granulomas were reversible, with TMWCNT-induced granulomas diminishing faster than PMWCNT-induced granulomas. Although the area of granuloma reduced with time, hyperplasia and dysplastic characteristics such as mitotic figures, anisokaryosis, and anisocytosis were still observed. These findings demonstrate that MWCNT induces granulomatous inflammation, and the duration and pattern of inflammation seem to vary depending upon the types of MWCNT to which mice are exposed. Therefore, toxicity studies on various types of CNT are needed as the responsiveness to these compounds differs.

Repeated aerosol delivery of carboxyl-terminal modulator protein suppresses tumor in the lungs of K-rasLA1 mice.

RATIONALE: Difficulties in achieving long-term survival of patients with lung cancer treated with conventional therapies suggest that novel approaches are required. Recent advances in aerosol-mediated gene delivery have provided the possibility of an alternative for the safe and effective treatment of lung cancer. OBJECTIVES: To investigate the repeated effect of carboxyl-terminal modulator protein (CTMP) on multistage lung tumorigenesis. In this study, we addressed this question by studying the effects of lentivirus-based CTMP in the lungs of 9- and 13-week-old K-ras(LA1) mice, a model of lung cancer. METHODS: An aerosol of lentivirus-based CTMP was delivered into 9- and 13-week-old K-ras(LA1) mice, a model of lung cancer, through a nose-only inhalation system twice a week for 4 weeks. The effects of CTMP on lung cancer progression and Akt-related signals were evaluated. MEASUREMENTS AND MAIN RESULTS: Long-term repeated delivery of CTMP effectively reduced tumor progression in the lungs at different stages of development. Lentiviral-CTMP inhibited protein synthesis and cell cycle and altered Akt signaling pathway in the lungs of 9-week-old K-ras(LA1) mice, and increased apoptosis was observed in the lungs of 13-week-old K-ras(LA1) mice. CONCLUSIONS: Long-term repeated viral delivery of CTMP may provide a useful tool for designing lung tumor treatment.

High dietary inorganic phosphate increases lung tumorigenesis and alters Akt signaling.

RATIONALE: Phosphate (Pi) is an essential nutrient to living organisms. Recent surveys indicate that the intake of Pi has increased steadily. Our previous studies have indicated that elevated Pi activates the Akt signaling pathway. An increased knowledge of the response of lung cancer tissue to high dietary Pi may provide an important link between diet and lung tumorigenesis. OBJECTIVES: The current study was performed to elucidate the potential effects of high dietary Pi on lung cancer development. METHODS: Experiments were performed on 5-week-old male K-ras(LA1) lung cancer model mice and 6-week-old male urethane-induced lung cancer model mice. Mice were fed a diet containing 0.5% Pi (normal Pi) and 1.0% Pi (high Pi) for 4 weeks. At the end of the experiment, all mice were killed. Lung cancer development was evaluated by diverse methods. MEASUREMENT AND MAIN RESULTS: A diet high in Pi increased lung tumor progression and growth compared with normal diet. High dietary Pi increased the sodium-dependent inorganic phosphate transporter-2b protein levels in the lungs. High dietary consumption of Pi stimulated pulmonary Akt activity while suppressing the protein levels of tumor suppressor phosphatase and tensin homolog deleted on chromosome 10 as well as Akt binding partner carboxyl-terminal modulator protein, resulting in facilitated cap-dependent protein translation. In addition, high dietary Pi significantly stimulated cell proliferation in the lungs of K-ras(LA1) mice. CONCLUSIONS: Our results showed that high dietary Pi promoted tumorigenesis and altered Akt signaling, thus suggesting that careful regulation of dietary Pi may be critical for lung cancer prevention as well as treatment.

Effects of phosphorus and calcium to phosphorus consumption ratio on mineral metabolism and cardiometabolic health.

Phosphorus is a common additive used in food processing that is typically consumed in excess of the recommended daily allowance; however, our knowledge of its effects on health, in the context of normal renal function, is limited. Unlike phosphorus, calcium intake is generally less than recommended, and it has been hypothesized that the calcium to phosphorus ratio may be partly responsible for the proposed negative health consequences. Therefore, this study sought to determine the effects of increased phosphorus additive intake, in the context of high calcium consumption, on endocrine markers of mineral metabolism and cardiometabolic health. An outpatient feeding study was performed in which healthy adults were fed a run-in control diet for 2 weeks followed by a phosphorus additive enhanced diet with supplemental calcium to an approximate ratio of 1 (experimental diet) for 2 weeks. Blood and urine samples were collected, and participants had brachial flow-mediated dilatation measured, with analyses comparing follow-up measures to baseline. Two weeks of experimental diet increased serum fibroblast growth factor 23 concentrations but lowered body weight and serum leptin; however, other phosphorus responsive factors such as osteopontin and osteocalcin did not increase. A complementary study in male mice also demonstrated that the regulation of known dietary phosphorus responsive factors was mostly abrogated when dietary calcium was raised in parallel with phosphorus. In conclusion, the study identifies weight, leptin and insulin as responsive to dietary phosphorus and that certain aspects of the systemic phosphorus response are attenuated by a corresponding high calcium intake.

New method to prepare very stable and biocompatible fluorescent silica nanoparticles.

A new synthetic method has been developed to prepare fluorescent silica nanoparticles without employing isothiocyanated dye molecules and (3-aminopropyl)triethoxysilane (APS) for the thiourea linkage formation; the resulting fluorescent silica nanoparticles show excellent photochemical, thermal and pH stabilities and a good biocompatibility with over 85% viability from various cell types.

Poly(ester amine)-mediated, aerosol-delivered Akt1 small interfering RNA suppresses lung tumorigenesis.

RATIONALE: The low efficiency of conventional therapies in achieving long-term survival of patients with lung cancer calls for the development of novel therapeutic options. Recent advances in aerosol-mediated gene delivery have provided the possibility of an alternative for the safe and effective treatment of lung cancer. OBJECTIVES: To demonstrate the feasibility and emphasize the importance of noninvasive aerosol delivery of Akt1 small interfering RNA (siRNA) as an effective and selective option for lung cancer treatment. METHODS: Nanosized poly(ester amine) polymer was synthesized and used as a gene carrier. An aerosol of poly(ester amine)/Akt1 siRNA complex was delivered into K-ras(LA1) and urethane-induced lung cancer models through a nose-only inhalation system. The effects of Akt1 siRNA on lung cancer progression and Akt-related signals were evaluated. MEASUREMENTS AND MAIN RESULTS: The aerosol-delivered Akt1 siRNA suppressed lung tumor progression significantly through inhibiting Akt-related signals and cell cycle. CONCLUSIONS: The use of poly(ester amine) serves as an effective carrier, and aerosol delivery of Akt1 siRNA may be a promising approach for lung cancer treatment and prevention.

Dominant-negative activator protein 1 (TAM67) targets cyclooxygenase-2 and osteopontin under conditions in which it specifically inhibits tumorigenesis.

Activation of activator protein 1 (AP-1) and nuclear factor kappaB (NFkappaB)-dependent transcription is required for tumor promotion in cell culture models and transgenic mice. Dominant-negative c-Jun (TAM67) blocks AP-1 activation by dimerizing with Jun or Fos family proteins and blocks NFkappaB activation by interacting with NFkappaB p65. Two-stage [7,12-dimethylbenz(a)anthracene (DMBA)/12-O-tetradecanoylphorbol-13-acetate (TPA)] skin carcinogenesis experiments in a model relevant to human cancer risk, transgenic mice expressing human papillomavirus 16 E7 oncogene (K14-HPV16-E7), show E7-enhanced tumor promotion. A cross to K14-TAM67-expressing mice results in dramatic inhibition of tumor promoter-induced AP-1 luciferase reporter activation and papillomagenesis. Epithelial specific TAM67 expression inhibits tumorigenesis without affecting TPA- or E7-induced hyperproliferation of the skin. Thus, the mouse model enriches for TAM67 targets relevant to tumorigenesis rather than to general cell proliferation or hyperplasia, implicating a subset of AP-1- and/or NFkappaB-dependent genes. The aim of the present study was to identify target genes responsible for TAM67 inhibition of DMBA-TPA-induced tumorigenesis. Microarray expression analysis of epidermal tissues revealed small sets of genes in which expression is both up-regulated by tumor promoter and down-regulated by TAM67. Among these, cyclooxygenase-2 (Cox-2/Ptgs2) and osteopontin (Opn/Spp1) are known to be functionally significant in driving carcinogenesis. Results identify both Cox-2 and Opn as transcriptional targets of TAM67 with CRE, but not NFkappaB sites important in the Cox-2 promoter and an AP-1 site important in the Opn promoter.

Low dietary inorganic phosphate affects the brain by controlling apoptosis, cell cycle and protein translation.

Inorganic phosphate (Pi) plays a key role in diverse physiologic functions. In a previous study, we showed that high dietary Pi perturbs brain growth through Akt/ERK signaling in developing mice. However, no study has investigated the response of the brain to low dietary Pi. In this study, we addressed this question by studying the effects of low dietary Pi on the cerebrum of developing mice. Two-week-old weaned mice were fed with a low phosphate diet for 4 weeks. At the end of the study, their cerebrum was dissected and signals important for protein translation, apoptosis and cell cycle were examined. The low phosphate diet did not cause physiologically significant changes; it increased the protein expression of phosphatase and tensin homolog deleted on chromosome 10 but decreased Akt activity. In addition, expression of eukaryotic translation initiation factor binding protein coupled with increased complex formation of eukaryotic translation initiation factor 4E/eukaryotic translation initiation factor binding protein 1 was induced in the cerebrum by low phosphate, leading to reduced cap-dependent protein translation. Finally, low phosphate facilitated apoptosis and suppressed signals important for the cell cycle in the cerebrum of dual-luciferase reporter mice. In summary, our results showed that a low phosphate diet affects the brain by controlling protein translation, apoptosis and cell cycle in developing mice. Our results support the hypothesis that Pi works as a stimulus capable of increasing or decreasing several pivotal genes for normal development and suggest that regulation of Pi consumption is important in maintaining a healthy life.

Analysis of the extracellular matrix and secreted vesicle proteomes by mass spectrometry.

The extracellular matrix (ECM) and secreted vesicles are unique structures outside of cells that carry out dynamic biological functions. ECM is created by most cell types and is responsible for the three-dimensional structure of the tissue or organ in which they are originated. Many cells also produce or secrete specialized vesicles into the ECM, which are thought to influence the extracellular environment. ECM is not s a physical structure to connect cells in a tissue or organ. The proteins in ECM and secreted vesicles are critical to cell function, differentiation, motility, and cell-to-cell interaction. Although a number of major structural proteins of ECM and secreted vesicles have long been known, an appreciation of the role of less-abundant non-collagenous proteins has just begun to emerge. This chapter outlines a series of methods used to isolate and enrich ECM constituents and secreted vesicles from bone-forming osteoblast cells, enabling comprehensive profiles of their proteomes to be obtained by mass spectrometry. These methods can be easily adapted to study ECM and secreted vesicles in other cell types, primary cell cultures derived from animal models, or tissue specimens.

The c-myc gene is a direct target of mammalian SWI/SNF-related complexes during differentiation-associated cell cycle arrest.

The activity of mammalian SWI/SNF-related chromatin remodeling complexes is crucial for differentiation, development, and tumor suppression. Cell cycle-regulating activities dependent on the complexes include induction of the p21(WAF1/CIP1) kinase inhibitor and repression of E2F-responsive promoters. These responses are linked through effects on pRb phosphorylation, but the direct role of the SWI/SNF-related complexes in their regulation is not fully understood. Results presented here reveal that the complexes are required for regulation of a distinct pathway of proliferation control involving repression of c-myc expression in differentiating cells. This involves direct promoter targeting of the c-myc gene by the complexes. Induction of p21(WAF1/CIP1) is specifically dependent on prior repression of c-myc, but repression of E2F-responsive genes is dissociable from the regulation of c-myc and p21(WAF1/CIP1).

Bioactive effects of silica nanoparticles on bone cells are size, surface, and composition dependent.

Silica based nanoparticles have been demonstrated to have intrinsic biologic activity towards the skeleton and to function by promoting the differentiation of bone forming osteoblasts while inhibiting the differentiation of bone resorbing osteoclasts. The excitement surrounding nanomedicine in part revolves around the almost unlimited possibilities for varying the physicochemical properties including size, composition, and surface charge. To date few studies have attempted to manipulate these characteristics in concert to optimize a complex biologic outcome. Towards this end, spherical silica nanoparticles of various sizes (50-450 nm), of different surface properties (OH, CO2H, NR4+, mNH2), and of different composition (silica, gold, and polystyrene) were synthesized and evaluated for biological activity toward skeletal cells. Osteoblast activity was most influenced by composition and size variables, whereas osteoclasts were most affected by surface property variation. The study also establishes nanoparticle mediated suppression of Nfatc1, a key transcriptional regulator for osteoclast differentiation, identifying a novel mechanism of action. Collectively, the study highlights how during the design of bioactive nanoparticles, it is vital to consider not only the myriad of physical properties that can be manipulated, but also that the characteristics of the target cell plays an equally integral role in determining biological outcome. STATEMENT OF SIGNIFICANCE: Silica nanomaterials represent a promising biomaterial for beneficial effects on bone mass and quality as well as regenerative tissue engineering and are currently being investigated for intrinsic bioactivity towards the primary cells responsible for skeletal homeostasis; osteoblasts and osteoclasts. The goal of the current study was to assess the physical properties of silica nanoparticles that impart intrinsic bioactivity by evaluating size, surface charge, and composition. Results reveal differential influences of the physical properties of nanoparticles towards osteoblasts and osteoclasts. This study provides new insights into the design of nanoparticles to specifically target different aspects of bone metabolism and highlights the opportunities provided by nanotechnology to modulate a range of cell specific biological responses for therapeutic benefit.