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Background MiR-17-92 cluster is coded by MIR17HG in chromosome 13, which is highly conserved in vertebrates. Published literatures have proved that miR-17-92 cluster critically regulates tumorigenesis and metastasis. Recent researches showed that the miR-17-92 cluster also plays novel functions in the endocrine system. Objective To summarize recent findings on the physiological and pathological roles of miR-17-92 cluster in bone, lipid and glucose metabolisms. Results MiR-17-92 cluster plays significant regulatory roles in bone development and metabolism through regulating the differentiation and function of osteoblasts and osteoclasts. In addition, miR-17- 92 cluster is nearly involved in every aspect of lipid metabolism. Last but not the least, the miR-17-92 cluster is closely bound up with pancreatic beta cell function, development of type 1 diabetes and insulin resistance. However, whether miR-17-92 cluster is involved in the communication among bone, fat and glucose metabolisms remains unknown. Conclusion Growing evidence indicates that miR-17-92 cluster plays significant roles in bone, lipid and glucose metabolisms through a variety of signaling pathways. Fully understanding its modulating mechanisms may necessarily facilitate to comprehend the clinical and molecule features of some metabolic disorders such as osteoporosis, arthrosclerosis and diabetes mellitus. It may provide new drug targets to prevent and cure these disorders.

Cellular senescence is the inability of cells to proliferate, which has both beneficial and detrimental effects on tissue development and homeostasis. Chronic accumulation of senescent cells is associated with age-related disease, including osteoarthritis, a common joint disease responsible for joint pain and disability in older adults. The pathology of this disease includes loss of cartilage, synovium inflammation, and subchondral bone remodeling. Senescent cells are present in the cartilage of people with advanced osteoarthritis, but the link between cellular senescence and this disease is unclear. In this review, we summarize current evidence for the role of cellular senescence of different cell types in the onset and progression of osteoarthritis. We focus on the underlying mechanisms of senescence in chondrocytes, which maintain the cartilage in joints, and review the role of the Forkhead family of transcription factors, which are involved in cartilage maintenance and osteoarthritis. Finally, we discuss the potential therapeutic value and implications of targeting senescent cells using senolytic agents or immune therapies, targeting the senescence-associated secretory phenotype of these cells using senomorphic agents, and renewing the plasticity of stem cells and chondrocytes. Our review highlights current gaps in understanding of the mechanism of senescence that may, when addressed, provided new options for modifying and treating disease in osteoarthritis.

Postmenopausal osteoporosis (PMOP) is considered to be a well-defined subject that has caused high morbidity and mortality. In elderly women diagnosed with PMOP, low bone mass and fragile bone strength have been proven to significantly increase risk of fragility fractures. Currently, various anabolic and anti-resorptive therapies have been employed in an attempt to retain healthy bone mass and strength. Pulsed electromagnetic fields (PEMFs), first applied in treating patients with delayed fracture healing and nonunions, may turn out to be another potential and effective therapy for PMOP. PEMFs can enhance osteoblastogenesis and inhibit osteoclastogenesis, thus contributing to an increase in bone mass and strength. However, accurate mechanisms of the positive effects of PEMFs on PMOP remain to be further elucidated. This review attempts to summarize recent advances of PEMFs in treating PMOP based on clinical trials, and animal and cellular studies. Possible mechanisms are also introduced, and the future possibility of application of PEMFs on PMOP are further explored and discussed. Bioelectromagnetics. 38:406-424, 2017. © 2017 Wiley Periodicals, Inc.

To compare the effects of high-fat diet (HFD) and high-fructose diet (HFrD) on bone metabolism at different time points, dynamically observe the bone histology and femur trabecular micro-architecture, and analyze the underlying mechanisms. Sixty–Five male 6- to 7-week-old C57BL/6J mice were given HFD, HFrD, or standard diets (SD) for 8, 16, and 24 weeks. Micro-computed tomography (μCT) and bone histology were used to measure bone mass and trabecular micro-structure. Quantitative real-time polymerase chain reaction (qRT-PCR) was used to determine the expression of genes related to bone and lipid metabolisms. Compared to SD mice, femoral trabecular bone mass was significantly increased in both HFrD mice and HFD mice at 8 weeks, it continued to be higher in HFrD mice at 16 and 24 weeks with the highest level at 16 weeks, but it was significantly decreased in HFD mice at 16 and 24 weeks. HFD mice showed more epididymal fat accumulation than HFrD mice. mRNA expression of Runx2 was up-regulated at 8 and 16 weeks, but down-regulated at 24 weeks similarly in both HFrD mice and HFD mice. mRNA expression of MMP9 and CTSK was up-regulated at 8 and 16 weeks in HFD mice, but down-regulated at 24 weeks in both HFrD mice and HFD mice. Our data indicated that the HFrD and HFD had different modulating effects on bone mass. After short-term feeding, both HFrD and HFD showed positive effects on bone mass; however, after long-term feeding, bone mass was decreased in HFD mice. In contrast, the bone mass was first increased and then decreased in the HFrD mice. On the basis of these findings, we speculated that chronic consumption of fat and fructose would exert detrimental effects on bone mass which might a combination action of body mass, fat mass, and bone formation/bone resorption along with proinflammatory factor and bone marrow environment.

Objective Osteoclastogenesis is a key process in osteolytic bone metastasis (BM). Previous studies indicated that some miRNAs could regulate cancers progression and osteoclastogenesis. Our purpose was to investigate the roles of lung cancer cells-derived circulating miR-21 on osteoclastogenesis and its clinical significance in BM patients. Materials and Methods The difference of miRNA expression in two lung cancer cell lines SBC-5 (with characteristic BM ability) and SBC-3 (without BM ability) were analyzed by microarray and qRT-PCR. Circulating miR-21 levels of lung cancer patients with or without BM were compared by qRT-PCR. The TRAP staining was used to investigate the effects of conditioned media from lung cancer cell lines or patients’ plasma with different miR-21 levels on osteoclastogenesis. ROC curve was used to evaluate the diagnostic performance of circulating miR-21 in BM patients. Results We found that miR-21 expression was specifically higher in SBC-5 than that in SBC-3 cells. The supernatants of SBC-5 cells with higher-level miR-21 promoted osteoclastogenesis. Moreover, we demonstrated that the circulating miR-21 level was significantly higher in BM patients than that in non-BM patients. The plasma from BM patients with higher-level miR-21 could also promote osteoclastogenesis. Mechanistically, lung cancer cells-derived circulating miR-21 could be transferred into osteoclast precursor cells and promote osteoclastogenesis probably by inhibiting PTEN. Finally, clinical data showed that circulating miR-21 had a potential for the diagnosis of BM. Conclusion Overall, our findings suggested that circulating miR-21 played important roles in osteoclastogenesis of lung cancer patients and may serve as a biomarker to diagnose BM of lung cancer.

Mucopolysaccharidosis type I (MPSI) is a rare autosomal recessive disorder caused by mutations in alpha-L-iduronidase (

Mucopolysaccharidosis type I (MPS I) is a rare autosomal recessive disorder caused by mutations in alpha-L-iduronidase (IDUA) gene. IDUA contributes to the degradation of the glycosaminoglycans, including heparan sulphate and dermatan sulphate. Deficient activity of IDUA generates accumulation of glycosaminoglycans in lysosomes leading to MPS I. Here, we identified two boys with MPS I caused by a compound heterozygote of a reported $$\hbox {c.265C}>\hbox {T}$$ (p.R89W) missense mutation in exon 2 and a novel $$\hbox {c.1633G}>\hbox {T}$$ (p.E545*, 109) nonsense mutation in exon 11 of IDUA gene in a Chinese family. R89 is close to the active site and its replacement will affect the structure and function of IDUA. Besides, termination from E545 deletes one of the prominent domains and alters the spatial structure of IDUA. In conclusion, our study demonstrates a previously unrecognized mutation in IDUA gene and this report adds to the mutational spectrum observed.

Osteoarthritis (OA) is the most common form of arthritic disease, leading to disability and impaired quality of life and no curative treatments exist. Increasing evidence indicates that low-grade inflammation plays a pivotal role in the onset and progression of OA. In this review, we summarize emerging findings on the pathological roles of synovial macrophages, adipose tissue macrophages, and osteoclasts in OA and their potential clinical implications from cell biology to preclinical and preliminary clinical trials. The failure of synovial macrophages to transition from pro-inflammatory M1 to anti-inflammatory M2 subtypes may contribute to the initiation and maintenance of synovitis in OA. M1 macrophages promote the inflammatory microenvironment and progression of OA through interactions with synovial fibroblasts and chondrocytes, thus increasing the secretion of matrix metalloproteinases. Direct inhibition of M1 or promotion of M2 polarization may be useful therapeutic interventions. Adipose tissue macrophages present in the infrapatella fat pad (IPFP) were involved in the progression of obesity-induced OA, which contributed to changes in the integrity of the IPFP. Furthermore, macrophages and osteoclasts in the subchondral bone were involved in bone remodeling and pain through uncoupled osteoclast/osteoblast activity and increased nociceptive signaling. Growing evidence has indicated an important role for macrophage-mediated low-grade inflammation in OA. Fully understanding the link between macrophages and other cells in joints will provide new insights into OA disease modification.

Background: Osteoclastogenesis is a key process in osteolytic bone metastasis (BM). Previous studies indicated that miRNAs could regulate cancer progression and osteoclastogenesis. Our purpose was to investigate the roles of lung cancer cells-derived circulating miR-21 on osteoclastogenesis and its clinical significance in BM patients. Methods: The difference of miRNA expression in two lung cancer cell lines SBC-5 (with characteristic BM ability) and SBC-3 (without BM ability) were analyzed. Circulating miR-21 levels of lung cancer patients with or without BM were compared. The effects of conditioned media from lung cancer cell lines or patients' plasma with different miR-21 levels on osteoclastogenesis were investigated. The diagnostic performance of circulating miR-21 in BM patients was evaluated. Findings: We found that miR-21 expression was specifically higher in SBC-5 than that in SBC-3 cells. The supernatants of SBC-5 cells with higher level miR-21 promoted osteoclastogenesis. Moreover, we demonstrated that the circulating miR-21 level was significantly higher in BM patients than that in non-BM patients. The plasma from BM patients with higher level miR-21 could also promot osteoclastogenesis. Mechanistically, lung cancer cells-derived circulating miR-21 could be transferred into osteoclast precursor cells and promot osteoclastogenesis probably by inhibiting PTEN (Phosphatase and tensin homologue). Finally, we indecated that circulating miR-21 had a potential for the diagnosis of BM. Interpretation: Our findings suggested circulating miR-21 played important roles in BM of lung cancer and may service as a biomarker to diagnose BM in lung cancer patients. Funding: This work was supported by grants from the National Natural Science Foundation of China (No. 81572639, 81770875, 81370969), Department of Science and Technology of Sichuan Province (2018SZ0142), and Sichuan University (2018SCUH0093). Declaration of Interest: No potential conflicts of interest were disclosed. Ethical Approval: The study was approved by the hospital research ethics committee of West China Hospital of Sichuan University and conducted in accordance with the International Ethical Guidelines for Biomedical Research Involving Human Subjects. All patients included in this study had signed the informed consent form.

GNAS is one of the most complex gene loci in the human genome and encodes multiple gene products. XLas, the extra-large isoform of alpha-subunit of the stimulatory guanine nucleotide-binding protein (Gas), is paternally inherited. Although XLas can mimic the action of Gas, its significance remains largely unknown in humans. Here we report a patient presented with increased bone mass, hypophosphatemia, and elevated parathyroid hormone levels. His serum calcium was in the lower limit of normal range. DEXA scan revealed progressive increase in the bone density of this patient. Whole exome sequencing of this subject found a novel nonsense mutation c.424G>T (p. G142X) in the first exon of XLas, which was inherited from his father and transmitted to his daughter. This mutation was predicted to exclusively influence the expression of XLas, while may have no significant effects on other gene products of this locus. SaOS2 cells transfected with mutant XLas failed to generate cAMP under parathyroid hormone stimulation, indicating skeletal resistance to this hormone. This subject showed higher circulating SOST, DKK1 and OPG levels, while lower RANKL levels and RANKL/OPG ratio, leading to reduced bone resorption. It is speculated that this patient may belong to a very rare type of pseudohypoparathyroidism with selective skeletal resistance but normal renal tubular response to parathyroid hormone. Our findings indicate that XLas plays a critical role in bone metabolism and GNAS locus should be considered as a candidate gene for high bone mass.

Osteoimmunology is the interdiscipline that focuses on the relationship between the skeletal and immune systems. They are interconnected by shared signal pathways and cytokines. Interferon-gamma (IFN-γ) plays important roles in immune responses and bone metabolism. IFN-γ enhances macrophage activation and antigen presentation. It regulates antiviral and antibacterial immunity as well as signal transduction. IFN-γ can promote osteoblast differentiation and inhibit bone marrow adipocyte formation. IFN-γ plays dual role in osteoclasts depending on its stage. Furthermore, IFN-γ is an important pathogenetic factor in some immune-mediated bone diseases including rheumatoid arthritis, postmenopausal osteoporosis, and acquired immunodeficiency syndrome. This review will discuss the contradictory findings of IFN-γ in osteoimmunology and its clinical application potential.

Lung cancer still remains the leading cause of cancer-related mortality worldwide. Bone is one of preferred metastatic sites for lung cancer cells. So far, both accurate diagnosis and effective treatment of lung cancer bone metastases are difficult.This review aimed to evaluate roles of bone turnover markers (BTMs), microRNAs (miRNAs), dickkopf1 (DKK1) and insulin like growth factor binding protein 3 (IGFBP-3) in lung cancer bone metastases.We searched articles about these four biomarkers in lung cancer bone metastases mainly in PubMed.The levels of bone specific alkaline phosphatase (BALP), cross-linked carboxy-terminal telopeptide of type-I collagen (ICTP) and N-terminal telopeptides of type-I collagen (NTX) were reported to be significantly increased in lung cancer patients with bone metastases. ALP, NTX and bone sialoprotein were thought to be associated with prognosis of lung cancer patients with bone metastases. MiRNA-335, miRNA-33a, miRNA-21, DKK1 and IGFBP-3 were revealed to be novel biomarkers in lung cancer bone metastases.Current researches have revealed that BTMs, miRNAs, DKK1 and IGFBP-3 may be useful in diagnosis, prognosis evaluation or treatment of lung cancer bone metastases. More studies about these biomarkers in lung cancer bone metastases are needed.

We examined the effects of tumor necrosis factor-α (TNFα) and interleukin-6 (IL6) gene knockout in preserving the bone loss induced by ovariectomy (OVX) and the mechanisms involved in bone metabolism. Twenty female wild-type (WT)

MicroRNAs (miRNAs) are a new class of small noncoding RNAs of 19-25 nucleotides that function as negative posttranscriptional gene regulators. MiRNAs hybridize to the 3' untranslated region (UTR) of target mRNAs and repress translation or mediate mRNA cleavage. MiRNAs critically regulate tumorigenesis and progression by targeting oncogenes, tumor suppressor genes, or genes related to proliferation, angiogenesis, and apoptosis. Different tumor types and tumors at different stages exhibit unique miRNA profiles. MiRNAs show promise as potential biomarkers for cancer diagnostics, progression, and response to treatment. The role of miRNAs in promoting bone metastases is under investigated. We summarize recent findings on the mechanisms by which miRNAs may regulate bone metastatic spread of breast cancer, prostate cancer, lung cancer and multiple myeloma. We review similarities and differences in miRNA profiles that may explain the variety of molecular pathways underling metastatic spread to the skeleton in different cancers. Finally, we discuss the exciting potential of using miRNAs as diagnostics and therapeutic targets to reduce the risk of bone metastases in cancer, from the perspective of data provided by recent pre-clinical and clinical studies.

Lung cancer is the leading cause of malignancy-related mortality worldwide. Metastases, which account for 90% of lung cancer deaths, frequently target the skeleton, leading to rapid deterioration in quality of life and premature death. The molecular mechanism underlying this progression, especially the development of bone metastases, is largely unknown. MicroRNAs (miRNAs) are small, endogenous, noncoding RNAs that function as negatively posttranscriptional gene regulators. Changes in miRNAs, which may exhibit either oncogenic or tumor suppressive activity, are common in lung cancer. Over-expressed miRNAs may contribute to oncogenesis by down-regulating tumor suppressors, whereas the loss of selected miRNAs may negatively regulate oncogenes or factors related to tumorigenesis and progression. MiRNAs may activate or repress metastases. Specific miRNA expression profiles may correlate with the response in treatment. We summarize recent findings and patents in the pathological roles of miRNAs in the progression and bone metastases in lung cancer, and discuss the diagnostic and therapeutic options in the clinical management of lung cancer.

Background High bone mass (HBM) disorders are a group of clinically and genetically heterogeneous bone diseases characterized by increased bone density on radiographs, due to progressive bone overgrowth or impaired bone resorption, or both. Some HBM cases are secondary to other diseases, such as chronic hepatitis C virus infection. Despite the great advance in gene diagnostic technology, the majority of HBM individuals remain undiagnosed. Objective In this review, we will summarize the clinical, radiological and biochemical characteristics of HBM cases due to varying etiologies, since these features are helpful in the differential diagnosis of HBM. Results Each subgroup of HBM cases shows distinctive clinical, radiological and biochemical characteristics. HBM, due to bone overgrowth, was designated as sclerosteosis, as a result of mutations located in genes critically involved in the Wnt/beta-catenin signal pathway. Mutations in genes encoding factors relevant to the differentiation and maturation of osteoclasts, or critical for the acidification and resorption of osteoclasts may lead to osteopetrosis. Hepatitis C associated osteosclerosis is characterized by a generalized increase in bone mass and markedly elevated serum levels of bone specific alkaline phosphatase. Conclusion The clarification of the etiologies of HBM may have a breakthrough role in understanding the molecular mechanisms involved in bone metabolism and may provide new pathways for the intervention of osteoporosis.

Background Research on the characteristics and mechanisms of diabetes in Tibetans is scant. Especially, there is no study on the relationship between osteocalcin and glucose metabolism. The objective of this study was to investigate the associations of serum total osteocalcin (tOC) and undercarboxylated osteocalcin (ucOC) with glucose and lipid metabolism in Chinese indigenous Tibetans with different degrees of glucose tolerance. Methods In this study, 160 middle-aged Tibetan men were involved, including 46 subjects with normal glucose tolerance (NGT), 52 subjects with impaired glucose regulation (IGR) and 62 subjects with type 2 diabetes. The homeostasis model assessment (HOMA) parameters, including HOMA-IR and HOMA-B, were used to estimate insulin resistance and β-cell function, respectively. Adiponectin, leptin, testosterone, 1,25-dihydroxyvitamin D, tOC and ucOC were measured using ELISA kits. Results After adjustment for age and body mass index, plasma tOC level was correlated negatively with fasting and 30-min post-OGTT glucose, HOMA-IR, leptin and testosterone; plasma ucOC level was correlated negatively with 30-min post-OGTT glucose, total cholesterol and 1,25-dihydroxyvitamin D; ucOC : tOC was correlated positively with leptin. The negative association between HOMA-IR and tOC remained significant after correcting for adiponectin; however, the association disappeared after correcting for leptin. HOMA-IR was correlated negatively with age, adiponectin and tOC, and positively with total cholesterol, triglyceride and leptin. Stepwise linear regression analysis revealed that total cholesterol, leptin and adiponectin were independent predictors for HOMA-IR in all subjects. Conclusions Our data support a link between osteocalcin and glucose metabolism in middle-aged Tibetan men. The improved glucose tolerance exerted by tOC may be related to improved insulin sensitivity rather than improved β-cell function. Copyright © 2013 John Wiley & Sons, Ltd.

Background: Bone metastasis is a common complication of certain types of cancer, and unfortunately, it is still incurable to date. Immune system is a major defence against tumor cells and bone metastasis. However, the immunodeficient mouse models are widely used in most researches of bone metastasis, thereby excluding the regulatory roles of the immune system in bone metastasis. Objective: To provide a comprehensive overview on the cellular and molecular mechanisms by which immune cells interact with tumor cells and bone cells, as well as recent related patents. Method: We performed a literature search of PubMed database for the current knowledge in the interaction between bone metastasis and immune system. Recent related patents were obtained from World Intellectual Property Organization (WIPO) website. Results: Immune cells in the bone-tumor microenvironment include dentritic cells, monocytes/macrophages, myeloid-derived suppressor cells, regulatory T cells, T helper cells, neutrophil, CD4+ T lyphocytes, CD8+cytotoxic T lymphocytes, and natural killer cells, et al. In addition to their well-known immunomodulatory function, recent studies revealed that immune cells could cooperate with tumor cells and bone cells to enhance bone metastasis and tumor progression. There are some patents targeted immune system to inhibit bone metastasis. Conclusion: Immune cells are instigated by tumor cells to enhance the occurrence and the development of bone metastasis. Taking full advantage of anti-tumor roles of immune cells may bring the promise of a possible cure for bone metastasis.

Rationale Currently, the relationship between heterozygous mutations in SLC34A1 and hypophosphatemia is controversial. Here we report an autosomal dominant hypophosphatemia pedigree carrying a novel heterozygous mutation in SLC34A1. Patient concerns The proband is a 32-year old young man, presented with progressive pain and weakness in his lower extremities for more than 5 years. The proband showed persistent hypophosphatemia and low TmPO4/GFR values, indicating renal phosphate leak. His grandfather, father, and one of his uncles showed the similar symptoms. Diagnoses Autosomal dominant hypophosphatemia. Interventions and outcomes Phosphorus supplement was prescribed to the proband and his affected uncle. Both their serum phosphorus levels recovered to normal and their symptoms such as back pain and lower extremity weakness were completely relieved. Whole exome sequencing was performed to identify disease-causing mutations in proband. Lessons A novel heterozygous missense mutation c.680A>G (p. N227S) in exon 7 of SLC34A1 was found in proband by whole exome sequencing, which was also found in other 4 family members of this pedigree. Our report of an autosomal dominant hypophosphatemia pedigree with 5 mutant carriers enriches the clinical phenotype caused by the SLC34A1 mutations and further affirms the heterozygous mutations are causative for hypophosphatemia.

MicroRNAs (miRNAs) have been identified to play important functions during osteoblast proliferation, differentiation, and apoptosis. The miR-17~92 cluster is highly conserved in all vertebrates. Loss-of-function of the miR-17-92 cluster results in smaller embryos and immediate postnatal death of all animals. Germline hemizygous deletions of MIR17HG are accounted for microcephaly, short stature, and digital abnormalities in a few cases of Feingold syndrome. These reports indicate that miR-17~92 may play important function in skeletal development and mature. To determine the functional roles of miR-17~92 in bone metabolism as well as osteoblast proliferation and differentiation. Murine embryonic stem cells D3 and osteoprogenitor cell line MC3T3-E1 were induced to differentiate into osteoblasts; the expression of miR-17-92 was assayed by quantitative real-time RT-PCR. The skeletal phenotypes were assayed in mice heterozygous for miR-17~92 (miR-17~92 +/Δ ). To determine the possibly direct function of miR-17~92 in bone cells, osteoblasts from miR-17~92 +/Δ mice were investigated by ex vivo cell culture. miR-17, miR-92a, and miR-20a within miR-17-92 cluster were expressed at high level in bone tissue and osteoblasts. The expression of miR-17-92 was down-regulated along with osteoblast differentiation, the lowest level was found in mature osteoblasts. Compared to wildtype controls, miR-17-92 +/Δ mice showed significantly lower trabecular and cortical bone mineral density, bone volume and trabecular number at 10 weeks old. mRNA expression of Runx2 and type I collagen was significantly lower in bone from miR-17-92 +/Δ mice. Osteoblasts from miR-17-92 +/Δ mice showed lower proliferation rate, ALP activity and less calcification. Our research suggests that the miR-17-92 cluster critically regulates bone metabolism, and this regulation is mostly through its function in osteoblasts.

Postmenopausal osteoporosis (PMOP) is considered to be a well-defined subject that has caused high morbidity and mortality. In elderly women diagnosed with PMOP, low bone mass and fragile bone strength have been proven to significantly increase risk of fragility fractures. Currently, various anabolic and anti-resorptive therapies have been employed in an attempt to retain healthy bone mass and strength. Pulsed electromagnetic fields (PEMFs), first applied in treating patients with delayed fracture healing and nonunions, may turn out to be another potential and effective therapy for PMOP. PEMFs can enhance osteoblastogenesis and inhibit osteoclastogenesis, thus contributing to an increase in bone mass and strength. However, accurate mechanisms of the positive effects of PEMFs on PMOP remain to be further elucidated. This review attempts to summarize recent advances of PEMFs in treating PMOP based on clinical trials, and animal and cellular studies. Possible mechanisms are also introduced, and the future possibility of application of PEMFs on PMOP are further explored and discussed. Bioelectromagnetics. 38:406-424, 2017. © 2017 Wiley Periodicals, Inc.

Studies suggest interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) may be beneficial in obesity-related disorders with inconsistent results. This study was designed to investigate and compare their pathophysiological roles in lipid metabolism with gene knockout approach. Male wild-type (WT), IL-6 knockout (IL-6-/-), and TNF-α knockout (TNF-α-/-) mice were maintained on either a chow diet or a high-fat diet (HFD) for 12 weeks. Indices of lipid metabolism in blood, adipose, and liver were determined. Our data showed that IL-6-/- and TNF-α-/- mice were more pronounced in body weight gains, hypercholesterolemia, fasting and post-load hyperglycemia on a chow diet or a HFD compared with WT mice. In WT mice feeding on a HFD, lipolysis and glyceroneogenesis were enhanced, lipogenesis was inhibited in adipose tissue; lipogenesis was increased in liver tissue. IL-6-/- mice on a chow diet or a HFD showed similar metabolic phenotypes as WT mice on a HFD, since those mice had similar expression profiles of lipid-related genes in adipose tissue and liver. However, TNF-α-/- mice were different. Therefore, IL-6-/- and TNF-α-/- mice showed different hepatic triglyceride infiltration in response to different diets. Our study suggests that complete blockage of IL-6 and TNF-α is unbeneficial in obesity and obesity-related metabolic disorders in mice.

The purpose of the study was to determine the roles of interleukin-6 (IL6) in fat and bone communication. Male wild-type (WT) mice and IL6 knockout (IL6−/−) mice were fed with either regular diet (RD) or high-fat diet (HFD) for 12 weeks. Bone mass and bone microstructure were evaluated by micro-computed tomography. Gene expression related to lipid and bone metabolisms was assayed with real-time quantitative polymerase chain reaction. Bone marrow cells from both genotypes were induced to differentiate into osteoblasts or osteoclasts, and treated with palmitic acid (PA). HFD increased the body weight and fat pad weight, and impaired lipid metabolism in both WT and IL6−/− mice. The dysregulation of lipid metabolism was more serious in IL6−/− mice. Trabecular bone volume fraction, trabecular bone number and trabecular bone thickness were significantly downregulated in WT mice after HFD than those in the RD (P −/− mice than those in WT mice on the HFD (P −/− osteoblasts displayed higher alkaline phosphatase (ALP) activity and higher mRNA levels of Runx2 and Colla1 than those in WT osteoblasts both in the control and PA treatment group (P −/− mice showed significantly lower mRNA levels of PPARγ and leptin and higher mRNA levels of adiponectin in comparison with WT mice on HFD. In conclusion, these findings suggested that IL6 gene deficiency antagonized HFD-induced bone loss. IL6 might bridge lipid and bone metabolisms and could be a new potential therapeutic target for lipid metabolism disturbance-related bone loss.

BACKGROUND Obesity was traditionally considered as a positive regulator on the strength of bone. With the in-depth study, obesity is considered as a major risk factor for osteoporosis. Some proinflammatory cytokines such as tumor necrosis factor-alpha (TNF-α) and interleukin-6 (IL-6) are the factors that fat uses to negatively regulate bone metabolism. OBJECTIVE This review was aimed to summarize and critically discuss the convincing evidence for the therapeutic effectiveness of pro-inflammatory cytokines for the treatment of obesity-related bone disorders. RESULTS Obese people and animals show a higher level of serum TNF-α and IL-6, which are produced by macrophages derived from adipose tissue. These pro-inflammatory cytokines regulate the proliferation and apoptosis of adipocyte, promote lipolysis, inhibit lipid synthesis and decrease blood lipids through autocrine and paracrine way. On the other hand, TNF-α and IL-6 regulate bone metabolism through the endocrine way. Several reports suggest that TNF-α is a negative regulator of osteoblast at some stages of differentiation and positively regulates osteoclast proliferation and differentiation. In contrast, IL-6 influences osteoblasts and osteoclasts through complex mechanisms, which reflect dual effects. In addition, TNF-α and IL-6 may regulate bone metabolism indirectly by regulating adiponectin and leptin released from adipocytes. CONCLUSION In this review, we first summarize the role of TNF-α and IL-6 in lipid and bone metabolisms. We further discuss how TNF-α and IL-6 regulate the communication between fat and bone, and their pathological roles in obesity-related bone disorders. Lastly, we discuss the possibility of using pro-inflammatory signaling pathway as a therapeutic target to develop drug for obesity-related bone disorders.

MiR-17-92 cluster contributes to the regulation of mammalian development, aging and tumorigenesis. The functional roles of miR-17-92 in pancreatic beta-cells are largely unknown. In this study, we found that conditional deletion of miR-17-92 in mouse pancreatic beta-cells (miR-17-92βKO) significantly reduces glucose tolerance and the first phase of insulin secretion, despite normal ad libitum fed and fasting glucose levels. Proliferation is down-regulated in pancreatic beta-cells after deleting miR-17-92. MiR-17-92βKO mice show higher phosphatase and tensin homologue (PTEN) and lower phosphorylated AKT in islets. Under high fat diet challenge for 16 weeks, miR-17-92βKO mice lose compensation and exhibit higher glucose levels, and lower insulin secretion. Collectively, these data suggest that miR-17-92 is a critical contributor to molecular mechanisms regulating glucose-stimulated insulin secretion and pancreatic beta-cell adaptation under metabolic stress.

Bone has emerged as a novel endocrine organ for its ability to produce hormones and involvement in several regulatory feedback loops. Osteocalcin (OCN) is released into bloodstream during bone resorption and has been demonstrated to exert endocrine regulation on islets, fat and male testis to form feedback loops. We hypothesize that bone delivers its energy metabolism signals to related energy-regulating organs through OCN based on the following evidence: First, OCN has close interactions with islets and fat, and it shows ability to stimulate islets and fat to secret insulin and adiponectin, respectively. Islets and fat are important organs involved in energy metabolism. Second, OCN undergoes physiological fluctuations during a lifetime. In children and adolescents, during the development of osteoporosis or after bone fracture, OCN level increases significantly. The elevated OCN at these stages represents enhanced bone turnover and metabolic activity, which require more energy supply. Therefore, the metabolic activity of bone and the energy-related organs like fat and islets are closely linked by circulating OCN. Through systemic release of OCN, bone delivers its energy-demanding information to other organs to satisfy its energy requirement.

Individuals with nerofibromatosis Type 1 (NF1) frequently suffer a spectrum of bone pathologies, such as abnormal skeletal development (scoliosis, congenital bowing, and congenital pseudoarthroses, etc), lower bone mineral density with increased fracture risk. These skeletal problems may result, in part, from abnormal osteoclastogenesis. Enhanced RAS/PI3K activity has been reported to contribute to abnormal osteoclastogenesis in Nf1 heterozygous (Nf1+/-) mice. However, the specific downstream pathways linked to NF1 abnormal osteoclastogenesis have not been defined. Our aim was to determine whether mammalian target of rapamycin (mTOR) was a key effector responsible for abnormal osteoclastogenesis in NF1. Primary osteoclast-like cells (OCLs) were cultured from Nf1 wild-type (Nf1+/+) and Nf1+/- mice. Compared to Nf1+/+ controls, there were 20% more OCLs induced from Nf1+/- mice. Nf1+/- OCLs were larger and contained more nuclei. Hyperactive mTOR signaling was detected in Nf1+/- OCLs. Inhibition of mTOR signaling by rapamycin in Nf1+/- OCLs abrogated abnormalities in cellular size and number. Moreover, we found that hyperactive mTOR signaling induced abnormal osteoclastogenesis major through hyper-proliferation. Our research suggests that neurofibromin directly regulates osteoclastogenesis through mTOR signaling pathway. Inhibiting mTOR may represent a viable strategy to treat NF1 bone diseases.

SHARPIN is a novel protein thought to interact with SHANK family and is widely expressed in multiple tissues/cells, including osteoblasts and osteoclasts. Loss-of-function of Sharpin develops the chronic proliferative dermatitis mutation (CPDM) in mice as well as a severe inflammation in other organs. The actual function of SHARPIN is poorly understood. Our aim was to determine the functional roles of SHARPIN in bone metabolism by using CPDM mice. The skeletal phenotypes were determined by peripheral quantitative computed tomography, micro-computed tomography, and quantitative real-time RT-PCR, the cellular functions of osteoblasts and osteoclasts were investigated by ex vivo cell culture. Compared to wild-type controls, CPDM mice demonstrated significantly lower total and cortical bone mineral content and bone mineral density, trabecular and cortical bone volume, and trabecular number. The mRNA expression of Runx2, osterix, type I collagen, and osteocalcin was significantly lower in the bone from CPDM mice. Osteoclasts and osteoblasts from CPDM mice were functionally defective. Our result suggests that SHARPIN plays important regulating roles in bone metabolism. These functional roles may either come from systemic chronic inflammatory or directly signaling pathway within bone cells.

The skeleton is frequently affected in individuals with neurofibromatosis type 1, and some of these bone manifestations can result in significant morbidity. The natural history and pathogenesis of the skeletal abnormalities of this disorder are poorly understood and consequently therapeutic options for these manifestations are currently limited. The Children's Tumor Foundation convened an International Neurofibromatosis Type 1 Bone Abnormalities Consortium to address future directions for clinical trials in skeletal abnormalities associated with this disorder. This report reviews the clinical skeletal manifestations and available preclinical mouse models and summarizes key issues that present barriers to optimal clinical management of skeletal abnormalities in neurofibromatosis type 1. These concepts should help advance optimal clinical management of the skeletal abnormalities in this disease and address major difficulties encountered for the design of clinical trials.

Unique among fibroblast growth factors (FGFs), FGF19, -21, and -23 act in an endocrine fashion to regulate energy, bile acid, glucose, lipid, phosphate, and vitamin D homeostasis. These FGFs require the presence of Klotho/betaKlotho in their target tissues. Here, we present the crystal structures of FGF19 alone and FGF23 in complex with sucrose octasulfate, a disaccharide chemically related to heparin. The conformation of the heparin-binding region between beta strands 10 and 12 in FGF19 and FGF23 diverges completely from the common conformation adopted by paracrine-acting FGFs. A cleft between this region and the beta1-beta2 loop, the other heparin-binding region, precludes direct interaction between heparin/heparan sulfate and backbone atoms of FGF19/23. This reduces the heparin-binding affinity of these ligands and confers endocrine function. Klotho/betaKlotho have evolved as a compensatory mechanism for the poor ability of heparin/heparan sulfate to promote binding of FGF19, -21, and -23 to their cognate receptors.

Individuals with neurofibromatosis type 1 (NF1) have a high incidence of osteoporosis and osteopenia. However, understanding of the cellular and molecular basis of these sequelae is incomplete. Osteoclasts are specialized myeloid cells that are the principal bone-resorbing cells of the skeleton. We found that Nf1(+/-) mice contain elevated numbers of multinucleated osteoclasts. Both osteoclasts and osteoclast progenitors from Nf1(+/-) mice were hyperresponsive to limiting concentrations of M-CSF and receptor activator of NF-kappaB ligand (RANKL) levels. M-CSF-stimulated p21(ras)-GTP and Akt phosphorylation was elevated in Nf1(+/-) osteoclasts associated with gains of function in survival, proliferation, migration, adhesion, and lytic activity. These gains of function are associated with more severe bone loss following ovariectomy as compared with that in syngeneic WT mice. Intercrossing Nf1(+/-) mice and mice deficient in class 1(A) PI3K (p85alpha) restored elevated PI3K activity and Nf1(+/-) osteoclast functions to WT levels. Furthermore, in vitro-differentiated osteoclasts from NF1 patients also displayed elevated Ras/PI3K activity and increased lytic activity analogous to those in murine Nf1(+/-) osteoclasts. Collectively, our results identify a what we believe to be a novel cellular and biochemical NF1-haploinsufficient phenotype in osteoclasts that has potential implications for the pathogenesis of NF1 bone disease.

The osteocyte, a terminally differentiated cell comprising 90%–95% of all bone cells1,2, may have multiple functions, including acting as a mechanosensor in bone (re)modeling3. Dentin matrix protein 1 (encoded by DMP1) is highly expressed in osteocytes4 and, when deleted in mice, results in a hypomineralized bone phenotype5. We investigated the potential for this gene not only to direct skeletal mineralization but also to regulate phosphate (Pi) homeostasis. Both Dmp1- null mice and individuals with a newly identified disorder, autosomal recessive hypophosphatemic rickets, manifest rickets and osteomalacia with isolated renal phosphate-wasting associated with elevated fibroblast growth factor 23 (FGF23) levels and normocalciuria. Mutational analyses showed that autosomal recessive hypophosphatemic rickets family carried a mutation affecting the DMP1 start codon, and a second family carried a 7-bp deletion disrupting the highly conserved DMP1 C terminus. Mechanistic studies using Dmp1-null mice demonstrated that absence of DMP1 results in defective osteocyte maturation and increased FGF23 expression, leading to pathological changes in bone mineralization. Our findings suggest a bone-renal axis that is central to guiding proper mineral metabolism.

Obesity and osteoporosis are two common chronic diseases, however, the basis for the correlation between them remains largely unknown. The pro-inflammation cytokine tumor necrosis factor-alpha (TNF-α) plays important roles in lipid and bone metabolisms, which may be a good candidate in the correlation between obesity and osteoporosis. We investigated the pathological roles of TNF-α in high-fat diet (HFD)-induced bone loss. Wild-type (WT) mice and TNF-α knockout (TNF-α(-/-)) mice were fed with the standard diet or the HFD for 12 weeks. Bone marrow stromal cells (BMSCs) from both genotypes were induced to differentiate into osteoblasts and treated with palmitic acid (PA). Bone mass and microstructure of femurs were evaluated by micro-CT. Lipid and bone metabolisms were investigated by histological and plasma analyses, and real-time PCR. On the HFD, both TNF-α(-/-) and WT mice presented notable visceral obesity, dyslipidemia. Adipogenesis and osteoclastogenesis were enhanced, while osteoblastogenesis was reduced in both genotypes. However, the changes were significantly different between TNF-α(-/-) and WT mice after the HFD. The gain of body and fat-pad weight was less and adipocyte area was smaller by 22 % in TNF-α(-/-) mice. Osteoclast numbers and plasma CTX level were lower by 40 % and by 23 % in TNF-α(-/-) mice. There were more ALP positive cells in the PA-treated TNF-α(-/-) BMSCs. mRNA expression of PPAR-γ was lower while that of Runx2 was higher in the bone from TNF-α(-/-) HFD group and in the PA-treated TNF-α(-/-) BMSCs, compared to WT on the same treatment. Furthermore, femoral trabecular bone mass and trabecular bone number were significantly decreased in WT mice on the HFD, whereas they were increased by 1.56-fold and 1.53-fold, respectively, in TNF-α(-/-) mice on the same diet (P0.05). Our results demonstrated that TNF-α gene knockout retained HFD-induced femoral trabecular bone loss mainly by suppressing adipogenesis and osteoclastogenesis, and enhancing osteoblastogenesis, which suggests that TNF-α plays a critical role in the development of HFD-related bone metabolic disorders and it may be a new potential therapeutic target for obesity-related bone loss.

Fibroblast growth factor (FGF)-23 has emerged as an endocrine regulator of phosphate and of vitamin D metabolism. It is produced in bone and, unlike other FGFs, circulates in the bloodstream to ultimately regulate phosphate handling and vitamin D production in the kidney. Presently, it is unknown which of the seven principal FGF receptors (FGFRs) transmits FGF23 biological activity. Furthermore, the molecular basis for the endocrine mode of FGF23 action is unclear. Herein, we performed surface plasmon resonance and mitogenesis experiments to comprehensively characterize receptor binding specificity. Our data demonstrate that FGF23 binds and activates the c splice isoforms of FGFR1–3, as well as FGFR4, but not the b splice isoforms of FGFR1–3. Interestingly, highly sulfated and longer glycosaminoglycan (GAG) species were capable of promoting FGF23 mitogenic activity. We also show that FGF23 induces tyrosine phosphorylation and inhibits sodium-phosphate cotransporter Npt2a mRNA expression using opossum kidney cells, a model kidney proximal tubule cell line. Removal of cell surface GAGs abolishes the effects of FGF23, and exogenous highly sulfated GAG is capable of restoring FGF23 activity, suggesting that proximal tubule cells naturally express GAGs that are permissive for FGF23 action. We propose that FGF23 signals through multiple FGFRs and that the unique endocrine actions of FGF23 involve escape from FGF23-producing cells and circulation to the kidney, where highly sulfated GAGs most likely act as cofactors for FGF23 activity. Our biochemical findings provide important insights into the molecular mechanisms by which dysregulated FGF23 signaling leads to disorders of hyper- and hypophosphatemia. (Endocrinology 146: 4647–4656, 2005)

Fibroblast growth factor-23 (FGF23) is a circulating factor that plays critical roles in phosphate and vitamin D metabolism. The goal of our studies was to dissect the pathways directing the vitamin D-phosphate-FGF23 homeostatic axis. To test the role of diet in the regulation of Fgf23, wild-type (WT) mice were fed either a standard (0.44% phosphorus) or a low-phosphate (0.02%) diet. WT mice on standard diet had a serum phosphate of 9.5 +/- 0.3 mg/dl and an Fgf23 concentration of 99.0 +/- 10.6 pg/ml; mice on the low-phosphate diet had a phosphate of 5.0 +/- 0.2 mg/dl (P0.01) and an Fgf23 of 10.6 +/- 3.7 pg/ml (P0.01). To genetically separate the effects of phosphate and vitamin D on Fgf23, we examined vitamin D receptor null (VDR(-/-)) mice, which are hypocalcemic and hypophosphatemic secondary to hyperparathyroidism. On standard diets, WT and VDR(+/-) mice had Fgf23 levels of 106.0 +/- 30.7 and 90.6 +/- 17.3 pg/ml, respectively, whereas Fgf23 was undetectable in the VDR(-/-). Animals were then placed on a diet that normalizes serum calcium and phosphorus. This 'rescue' increased Fgf23 in WT to 192.3 +/- 32.5 pg/ml and in VDR(+/-) to 388.2 +/- 89.6pg/ml, and importantly, in VDR(-/-) to 476.9 +/- 60.1 pg/ml (P0.01 vs. WT). In addition, renal vitamin D 1-alpha hydroxylase (1alpha-OHase) mRNA levels were corrected to WT levels in the VDR(-/-) mice. In summary, Fgf23 is suppressed in diet-induced hypophosphatemia and in hypophosphatemia associated with secondary hyperparathyroidism. Normalization of serum phosphate by diet in VDR(-/-) mice increases Fgf23. Thus, our results demonstrate that Fgf23 is independently regulated by phosphate and by vitamin D.

Gain-of-function mutations in fibroblast growth factor-23 (FGF23) are responsible for autosomal dominant hypophosphatemic rickets, a disorder of isolated renal phosphate wasting. Patients with the disorder display hypophosphatemia with normocalcemia as well as inappropriately normal 1,25-dihydroxyvitamin D [1,25(OH)2D3] concentrations. Reciprocally tumoral calcinosis (TC) patients are often hyperphosphatemic with inappropriately normal or elevated serum 1,25(OH)2D3 levels and have ectopic and vascular calcifications, a phenotype similar to that of Fgf23 null mice. Therefore, the goal of the present studies was to test whether FGF23 was a candidate gene for TC. Two sisters in a consanguineous TC family had hyperphosphatemia and normal 1,25(OH)2D3 levels with characteristic ectopic and vascular calcifications. Interestingly, these patients had low-normal intact serum FGF23 levels but demonstrated FGF23 concentrations approximately 40 times normal when assessed with a C-terminal FGF23 serum assay. Mutational analyses identified a homozygous S71G mutation in FGF23 in the TC patients, which was not found in control alleles. Finally, modeling demonstrated that the S71G mutation most likely destabilizes full-length FGF23. In summary, recessive FGF23 mutations can lead to TC. Additionally, our findings indicate that FGF23 may adopt an unstable conformation in some TC patients, possibly leading to nonfunctional FGF23 protein.

The association between lipid and bone metabolism has become an increasing focus of interest in recent years, and accumulating evidence has shown that atherosclerosis (AS) and osteoporosis (OP), a disorder of bone metabolism, frequently co-exist. Fat and bone are known to share a common progenitor cell: Multipotent mesenchymal stem cells (MSC) in the bone marrow (BM), which are able to differentiate into various cell phenotypes, including osteoblasts, adipocytes and chondrocytes. Laboratory-based and clinical trials have shown that increasing adipocytes are accompanied by a decrease in bone mineral density (BMD) and bone mass. Statins, lipid-lowering drugs used to treat hyperlipidemia, also provide benefit in the treatment of OP. There is thus evidence that the metabolism of lipids is correlated with that of bone, and that the two are mutually regulated. The present review primarily focuses on the potential association between lipid metabolism disturbance and OP, based on biological metabolism, pathophysiological processes, results from clinical and experimental animal studies, processes involved in the differentiation of adipocytes and osteoblasts, as well as pharmacological treatments of these diseases.

Type II autosomal dominant osteopetrosis (ADO2), which is the most common form of osteopetrosis, is caused by heterozygous mutations in the chloride channel 7 (CLCN7) gene. The osteopetrosis of ADO2 has been attributed to hypofunctional osteoclasts. The mechanism underlying the abnormality in osteoclast function remains largely unknown. This study was designed to investigate gene mutations and osteoclast function in a case that was clinically diagnosed as ADO2. Genomic DNA was extracted from blood samples of this patient, and the 25 exons of CLCN7 were amplified. Peripheral blood from the ADO2 subject and a healthy age- and sex-matched control was used to evaluate osteoclastogenesis, osteoclast morphology, and bone resorption. Analysis of DNA from the patient showed a germline heterozygous missense mutation, c.1856C>T (p.P619L), in exon 20 of CLCN7. A similar homozygous mutation at this site was previously reported in a patient with autosomal recessive osteopetrosis. When cultured, the peripheral blood mononuclear cells (PBMCs) from the ADO2 patient spontaneously differentiated into mature osteoclasts in vitro. The ADO2 patient’s PBMCs formed enhanced, but heterogeneous, osteoclasts in both the presence and absence of macrophage-colony stimulating factor, and nuclear factor-ĸB ligand. Bone resorption was reduced in the ADO2 patient’s osteoclasts, which exhibited aberrant morphology and abnormal distribution of integrin avβ3. Gene analysis found increased c-fos expression and reduced RhoA and integrin beta 3 expression in ADO2 cells. In conclusion, our data suggest that enhanced, heterogeneous osteoclast induction may be an intrinsic characteristic of ADO2. A mutation that reduces activity of bone-resorbing cells is responsible for a rare inherited disorder characterised by dense, brittle bones. Healthy bones undergo constant remodeling; osteoclasts resorb mature bone and osteoblasts generate new tissue. Patients with autosomal dominant osteopetrosis Type II (ADO2), the most common form of osteopetrosis, show impaired bone remodelling, resulting in overly dense, fragile bone tissue. No treatment is available, and although mutations in the chloride channel 7 gene (CLCN7) have been implicated, the mechanism underlying the disease remains unclear. Xijie Yu at Sichuan University and colleagues investigated gene mutations and osteoclast function in a recently diagnosed patient. The researchers found a single mutation of CLCN7 and higher than normal osteoclast generation; however, the osteoclasts were irregularly shaped and showed poor bone resorption. These results may help to identify treatments for osteopetrosis.

Research on the characteristics and mechanisms of diabetes in Tibetans is scant. Especially, there is no study on the relationship between osteocalcin and glucose metabolism. The objective of this study was to investigate the associations of serum total osteocalcin (tOC) and undercarboxylated osteocalcin (ucOC) with glucose and lipid metabolism in Chinese indigenous Tibetans with different degrees of glucose tolerance.In this study, 160 middle-aged Tibetan men were involved, including 46 subjects with normal glucose tolerance (NGT), 52 subjects with impaired glucose regulation (IGR) and 62 subjects with type 2 diabetes. The homeostasis model assessment (HOMA) parameters, including HOMA-IR and HOMA-B, were used to estimate insulin resistance and β-cell function, respectively. Adiponectin, leptin, testosterone, 1,25-dihydroxyvitamin D, tOC and ucOC were measured using ELISA kits.After adjustment for age and body mass index, plasma tOC level was correlated negatively with fasting and 30-min post-OGTT glucose, HOMA-IR, leptin and testosterone; plasma ucOC level was correlated negatively with 30-min post-OGTT glucose, total cholesterol and 1,25-dihydroxyvitamin D; ucOC : tOC was correlated positively with leptin. The negative association between HOMA-IR and tOC remained significant after correcting for adiponectin; however, the association disappeared after correcting for leptin. HOMA-IR was correlated negatively with age, adiponectin and tOC, and positively with total cholesterol, triglyceride and leptin. Stepwise linear regression analysis revealed that total cholesterol, leptin and adiponectin were independent predictors for HOMA-IR in all subjects.Our data support a link between osteocalcin and glucose metabolism in middle-aged Tibetan men. The improved glucose tolerance exerted by tOC may be related to improved insulin sensitivity rather than improved β-cell function.

Small cell lung cancer (SCLC) is a rapidly progressing, incurable cancer that frequently spreads to bone. New insights are needed to identify therapeutic targets to prevent or retard SCLC metastatic progression. Human SCLC SBC-5 cells in mouse xenograft models home to skeletal and nonskeletal sites, whereas human SCLC SBC-3 cells only pervade nonskeletal sites. Because microRNAs (miRNA) often act as tumor regulators, we investigated their role in preclinical models of SCLC. miRNA expression profiling revealed selective and reduced expression of miRNA (miR)-335 and miR-29a in SBC-5 cells, compared with SBC-3 cells. In SBC-5 cells, miR-335 expression correlated with bone osteolytic lesions, whereas miR-29a expression did not. Overexpression of miR-335 in SBC-5 cells significantly reduced cell migration, invasion, proliferation, colony formation, and osteoclast induction in vitro. Importantly, in miR-335 overexpressing SBC-5 cell xenografts (n = 10), there were minimal osteolytic lesions in the majority of mice and none in three mice. Expression of RANK ligand (RANKL) and insulin-like growth factor-I receptor (IGF-IR), key mediators of bone metastases, were elevated in SBC-5 as compared with SBC-3 cells. Mechanistically, overexpression of miR-335 in SBC-5 cells reduced RANKL and IGF-IR expression. In conclusion, loss of miR-335 promoted SCLC metastatic skeletal lesions via deregulation of IGF-IR and RANKL pathways and was associated with metastatic osteolytic skeletal lesions. Implications: These preclinical findings establish a need to pursue the role of miR-335 in human SCLC with metastatic skeletal disease. Mol Cancer Res; 12(1); 101–10. ©2013 AACR.

OBJECTIVE To summary the functional roles and molecular mechanisms of microRNA (miRNA) in osteoblast differentiation so as to supply information for basic and clinical researches.Recent literature concerning miRNA in osteoblast differentiation was reviewed. The information was classified and summarized.miRNAs critically regulate bone morphogenetic protein, transforming growth factor beta, and Wnt/beta-catenin signaling pathways during osteoblast differentiation. In pathological conditions, especially in some disorders of abnormal osteoblast differentiation, down-regulated miRNA expression has been observed.miRNA may represent a novel biomarker for diagnosis, and a candidate target therapies for the disorders with abnormal osteoblast differentiation.

Malignant peripheral nerve sheath tumors (MPNSTs) are highly aggressive soft tissue sarcomas accounting for 3%–10% of all soft tissue sarcomas. Neurofibromatosis type 1 (NF1) is the most important known risk factor. MPNSTs are often diagnosed at an advanced stage when distant metastases have developed. Although surgical resection remains the main treatment for MPNSTs, complete surgical resection is rarely possible. The prognosis for patients with MPNSTs is poor. There is an urgent need for improved therapies. To this end, we investigated whether microRNA (miR), specifically miR-204, might be implicated in MPNSTs because it is located at a cancer-associated genomic region exhibiting high frequency of loss of heterozygosity in tumors. We show that miR-204 expression is downregulated in NF1 and non-NF1 MPNST tumor tissues and in tumor cell lines. Restoring miR-204 expression in MPNST cell lines STS26T (non-NF1), ST88-14 (NF1), and T265p21 (NF1) significantly reduces cellular proliferation, migration, and invasion in vitro. Restoring miR-204 expression in STS26T decreases tumor growth and malignant progression in vivo. We also report that miR-204 inhibits Ras signaling and expression of high mobility group gene A2. These findings support the hypothesis that miR-204 plays critical roles in MPNST development and tumor progression. miR-204 may represent a novel biomarker for diagnosis and a candidate target with which to develop effective therapies for MPNSTs.

Metastases account for 90% of lung cancer mortalities, frequently target the skeleton and lead to rapid deterioration in quality of life. The molecular mechanism underlying bone metastases is largely unknown. Development of xenograft mouse models, such as the severe combined immunodeficient (SCID) CB-17 mouse and the non-obese diabetic (NOD)/SCID mouse, both of which lack functional B- and T-cells and are able to host allogeneic or xenogeneic tumor cells, has made great contributions in this area. However, residual natural killer (NK) cells in these models are able to significantly modify local tumor growth and metastasis. Treatment with anti-murine IL-2 receptor β chain Ab (TM-β1) antibody can abrogate NK cell activity in vivo; however, the antibody treatment may result in unexpected effects and the stability is hard to control. To overcome these shortcomings, we evaluated xenografts in NOD-scid IL2Rγ(null) immunodeficient mice that lacked mature T cells, B cells and functional NK cells. We compared the target tissue distribution of the human small cell lung cancer cell lines SBC-5 and SBC-3. Gross necropsy and whole skeletal X-ray film examination of the host mice were conducted 30 days post-tail vein injection. The SBC-5 cells colonized bone and formed lytic lesions. The cells also colonized liver, spleen and, less frequently, the pancreas, ovary and kidney. The SBC-3 cell xenografts formed easily visible tumor foci in the liver, pancreas, ovary/uterus and kidney, but not bone metastases. Our results showed that SBC-5 cells in NOD-scid IL2Rγ(null) immunodeficient mice provide a suitable xenograft model system for bone metastasis of human lung cancer. This novel animal model may therefore be used to study the molecular pathway of bone metastases and to evaluate targets for effective therapies.

Autosomal dominant hypophosphatemic rickets (ADHR) is unique among the disorders involving Fibroblast growth factor 23 (FGF23) because individuals with R176Q/W and R179Q/W mutations in the FGF23 176 RXXR 179 /S 180 proteolytic cleavage motif can cycle from unaffected status to delayed onset of disease. This onset may occur in physiological states associated with iron deficiency, including puberty and pregnancy. To test the role of iron status in development of the ADHR phenotype, WT and R176Q-Fgf23 knock-in (ADHR) mice were placed on control or low-iron diets. Both the WT and ADHR mice receiving low-iron diet had significantly elevated bone Fgf23 mRNA. WT mice on a low-iron diet maintained normal serum intact Fgf23 and phosphate metabolism, with elevated serum C-terminal Fgf23 fragments. In contrast, the ADHR mice on the low-iron diet had elevated intact and C-terminal Fgf23 with hypophosphatemic osteomalacia. We used in vitro iron chelation to isolate the effects of iron deficiency on Fgf23 expression. We found that iron chelation in vitro resulted in a significant increase in Fgf23 mRNA that was dependent upon Mapk. Thus, unlike other syndromes of elevated FGF23, our findings support the concept that late-onset ADHR is the product of gene–environment interactions whereby the combined presence of an Fgf23-stabilizing mutation and iron deficiency can lead to ADHR.

Individuals with nerofibromatosis Type 1 (NF1) frequently suffer a spectrum of bone pathologies, such as abnormal skeletal development (scoliosis, congenital bowing, and congenital pseudoarthroses, etc), lower bone mineral density with increased fracture risk. These skeletal problems may result, in part, from abnormal osteoclastogenesis. Enhanced RAS/PI3K activity has been reported to contribute to abnormal osteoclastogenesis in Nf1 heterozygous (Nf1+/-) mice. However, the specific downstream pathways linked to NF1 abnormal osteoclastogenesis have not been defined. Our aim was to determine whether mammalian target of rapamycin (mTOR) was a key effector responsible for abnormal osteoclastogenesis in NF1. Primary osteoclast-like cells (OCLs) were cultured from Nf1 wild-type (Nf1+/+) and Nf1+/- mice. Compared to Nf1+/+ controls, there were 20% more OCLs induced from Nf1+/- mice. Nf1+/- OCLs were larger and contained more nuclei. Hyperactive mTOR signaling was detected in Nf1+/- OCLs. Inhibition of mTOR signaling by rapamycin in Nf1+/- OCLs abrogated abnormalities in cellular size and number. Moreover, we found that hyperactive mTOR signaling induced abnormal osteoclastogenesis major through hyper-proliferation. Our research suggests that neurofibromin directly regulates osteoclastogenesis through mTOR signaling pathway. Inhibiting mTOR may represent a viable strategy to treat NF1 bone diseases.