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Abstract: Background: Osteoblast-derived matrix metalloproteinse-1 (MMP-1), MMP-2 and tissue inhibitor of metalloproteinase-1 (TIMP-1) play a role in bone metabolism by degrading bone matrix. Methods: We measured MMP-1, MMMP-2, TIMP-1 and associated results with age and bone metabolism in 591 Chinese women aged 20–80 y. Results: Serum MMP-1, MMP-2, and TIMP-1 concentrations exhibited positive correlation with age. Serum concentrations of MMP-1 were higher in 40–69 y old women. The concentrations of MMP-2 were significantly increased in the 50–69 y olds. Serum TIMP-1 concentrations were significantly lower in women aged 30–59 y, and then these were followed by an increase at >60 y olds. We found a significant negative weaker correlation between MMP-2 and BMD. But multiple linear stepwise regression analysis showed that MMP-2 was not a determinant factor for BMD. There were significant positive correlations between MMP-2 and bone alkaline phosphatase (BAP), osteocalcin (OC), and cross-linked N-telopeptides of type I collagen (NTX). Conclusions: The serum concentrations of MMP-1, MMP-2, and TIMP-1 exhibit age-related changes, and circulating MMP-2 and bone turnover are related. [Copyright &y& Elsevier]

Abstract: Adipocytes can highly and specifically express adiponectin, and the adiponectin receptor (AdipoR) has been detected in bone-forming cells. The present study was undertaken to investigate the action of adiponectin on osteoblast proliferation and differentiation. AdipoR1 protein was detected in human osteoblasts. Adiponectin promoted osteoblast proliferation and resulted in a dose- and time-dependent increase in alkaline phosphatase (ALP) activity, osteocalcin and type I collagen production, and an increase in mineralized matrix. Suppression of AdipoR1 with small-interfering RNA (siRNA) abolished the adiponectin-induced cell proliferation and ALP expression. Adiponectin induces activation of p38 mitogen-activated protein kinase (MAPK) and c-jun N-terminal Kinase (JNK), but not ERK1/2 in osteoblasts, and these effects were blocked by suppression of AdipoR1 with siRNA. Furthermore, pretreatment of osteoblasts with the JNK inhibitor SP600125 abolished the adiponectin-induced cell proliferation. p38 inhibitor SB203580 blocked the adiponectin-induced ALP activity. These data indicate that adiponectin induces human osteoblast proliferation and differentiation, and the proliferation response is mediated by the AdipoR/JNK pathway, while the differentiation response is mediated via the AdipoR/p38 pathway. These findings suggest that osteoblasts are the direct targets of adiponectin. [Copyright &y& Elsevier]

Estriol has been showed to prevent bone loss in osteoporotic rats and postmenopausal women, but the mechanisms remain unclear. In the present study, we evaluated the effect of estriol on osteoblastic MG-63 cells in vitro, and compared its action with 17β-estradiol (E2). Cell proliferation was determined by measuring total cell numbers and [3H]thymidine incorporation. Cell function was studied by measuring alkaline phosphatase (ALP) activity and secreted osteocalcin. Our data showed that estriol stimulated MG-63 cells proliferation in a dose-dependent manner, but had no influence on ALP activity in MG-63 cells and osteocalcin production. Compared with estriol treatment, E2 showed a stronger proliferation. Estrogen receptor (ER) α and β expression in MG-63 cells can be detected by Western immunoblot analysis, and the proliferative response to E2 and estriol can be all abrogated by ER antagonist ICI 182,780. In conclusion, estriol stimulates osteoblastic MG-63 cells proliferation, but has no effects on differentiation. The proliferative response to estriol is mediated by the ER. These results suggest that estriol has an effect on osteoblastic proliferation, and this may contribute to its actions on prevention of bone loss. [ABSTRACT FROM AUTHOR]

Recently, membrane-type matrix metalloproteinase-1 (MT1-MMP) was identified, and found that it can activate proMMP-2 on the cell membrane, degrade bone matrix, and participate in bone formation. Since bone is a target tissue of 1α,25-dihydroxyvitamin D3 [1α,25(OH)2D3], in the present study we observed the effects of 1α,25(OH)2D3 on MT1-MMP expression, and proMMP-2 activation in normal human osteoblast-like cells (hOB). Western immunoblots showed 1α,25(OH)2D3 time and dose dependently stimulated MT1-MMP production. By ELISA, we found that the activation of proMMP-2 in cultures of hOB was intensified by 1α,25(OH)2D3. Our studies suggest that 1α,25(OH)2D3 induces MMP-2 activation in part by up-regulating MT1-MMP expression, and since MT1-MMP plays a role in bone metabolism, the induction of MT1-MMP levels by 1α,25(OH)2D3 in hOB cells may contribute to a new mechanism by which 1α,25(OH)2D3 promotes bone formation and stimulates bone resorption. [ABSTRACT FROM AUTHOR]

Objective. Recently our studies have shown that nylestriol in combination with levonorgestrel prevented bone loss, decreased bone turnover rate and increased the maximal loading of bone without obvious side effects in retinoic acid (RA) induced osteoporotic rats. In addition to the animal experiments, we evaluate the effect of Compound Nylestriol Tablet (CNT) on bone mineral density (BMD) in women with postmenopausal osteoporosis. Compound Nylestriol Tablet, which contains 0.5 mg of nylestriol (cyclopentylethinyl estriol) and 0.15 mg of levonorgestrel per tablet, was authorized as a new anti-osteoporotic agent for clinical trial in postmenopausal osteoporosis. Methods. One year's clinical observation was performed in 191 eligible patients who were randomly divided into two groups (A and B). In group A, 119 patients were treated for one year with CNT (one tablet per week) and in group B, 72 patients with placebo. Bone mineral density of lumbar antero-posterior spine (L1-L4), lateral spine, total hip and total forearm positions including radius+ulna at the ultra distal areas, mid areas, and one-third areas, were measured before and after treatment. Biochemical parameters and effects of CNT on uterus, and breast were observed. Results. We found that patients treated with CNT had a significant decrease of bone loss in total forearm, including radius+ulna at the ultra distal, mid, and 1/3 areas compared with control subjects (all P <0.05). An improved BMD tendency could be seen at the lumbar spine. There were no differences in the observed biochemical variables. No side-effects on uterus, or mammary glands were observed. None of the patients had uterine bleeding or vertebral fractures during one year's CNT treatment. Conclusion. These data suggested that CNT is effective, safe and convenient in treating postmenopausal osteoporosis. [ABSTRACT FROM AUTHOR]

Objective. To determine an optimal dosage combination for a nylestriol/levonorgestrel (NYL/LNG) regimen in the treatment of female Sprague-Dawley (SD) rats with retinoic acid (RA)-induced osteoporosis in order to examine the rationale of NYL/LNG use for postmenopausal women. Methods: This study included two sets of experiments; one was designed to confirm the success of the RA-induced osteoporosis model involving 48 SD rats, and the other to determine the optimal dosage combination of NYL/LNG. In the second set of experiments, a total of 160 female SD rats at 7 months of age were randomly divided according to their basal body weights into 16 groups (10 in each). Treatment dosages of NYL and LNG were arrayed in a 2 factor-4 level (2[sup 4]) factorial design, i.e., NYL level I (N1 0.015 mg/kg), level II (N2 0.05 mg/kg), level III (N3 0.15 mg/kg) and level IV (N4 0.5 mg/kg) and LNG level I (L1 0.005 mg/kg), level II (L2 0.015 mg/kg), level III (L3 0.05 mg/kg) and level IV (L4 0.15 mg/kg). For 14 d, 70 mg/kg/d RA was given intragastrically to establish the osteoporofic model and NYL and LNG were then administered in different dosages on alternate days over a 3 wk period. Subsequently, body weight, uterine weight, endometrial status, Bone mineral density (BMD), bone turnover, and morphometry as well as parameters of biomechanics, serum sex hormones and lipids and estrogen receptor-α expression were determined for these rats. Results. Uterine weights at L2, L3, and L4 dosage levels were significantly lower than those at L1 (P < 0.05). Serum estradiol at N4 and progesterone at N2, N3, and N4 had decreased. Bone mineral density at distal tibiae (R5) in L3 and L4, and at proximal femora (R3) in L4 had increased. The peak loading of lumbar vertebrae at N3 was higher than that at N1, N2, and N4 dosage levels and that of femora higher at N3 than that at N1 and N4. Serum alkaline phosphatase (ALP) levels at N3, N4, L2, L3, and L4 dosage levels were lower than those at... [ABSTRACT FROM AUTHOR]

Osteoblasts rely on glucose as their primary energy source both for differentiation and to maintain function. Oxidative phosphorylation (OXPHOS) predominantly facilitates osteoclast differentiation, while glycolysis is the primary energy source for bone resorption by osteoclasts. Bone releases a series of endocrine factors to regulate systemic energy homeostasis. Disturbances in bone metabolism resulting from osteoporosis lead to imbalances in energy metabolism, further expediting the advance of osteoporosis. The bone serves as an energy reservoir and actively engages in whole-body energy metabolism. Numerous studies have determined fuel requirements and bioenergetic properties of bone under physiological conditions as well as the dysregulation of energy metabolism associated with bone metabolic diseases. Here, we review the main sources of energy in bone cells and their regulation, as well as the endocrine role of the bone in systemic energy homeostasis. Moreover, we discuss metabolic changes that occur as a result of osteoporosis. Exploration in this area will contribute to an enhanced comprehension of bone energy metabolism, presenting novel possibilities to address metabolic diseases. [ABSTRACT FROM AUTHOR]

Senescence and altered differentiation potential of bone marrow stromal cells (BMSCs) lead to age‐related bone loss. As an important posttranscriptional regulatory pathway, alternative splicing (AS) regulates the diversity of gene expression and has been linked to induction of cellular senescence. However, the role of splicing factors in BMSCs during aging remains poorly defined. Herein, we found that the expression of the splicing factor Y‐box binding protein 1 (YBX1) in BMSCs decreased with aging in mice and humans. YBX1 deficiency resulted in mis‐splicing in genes linked to BMSC osteogenic differentiation and senescence, such as Fn1, Nrp2, Sirt2, Sp7, and Spp1, thus contributing to BMSC senescence and differentiation shift during aging. Deletion of Ybx1 in BMSCs accelerated bone loss in mice, while its overexpression stimulated bone formation. Finally, we identified a small compound, sciadopitysin, which attenuated the degradation of YBX1 and bone loss in old mice. Our study demonstrated that YBX1 governs cell fate of BMSCs via fine control of RNA splicing and provides a potential therapeutic target for age‐related osteoporosis. Synopsis: Alternative splicing has been shown to regulate cellular senescence and differentiation. This study links the reduced expression of the splicing regulator YBX1 in aging bone marrow stromal cells to senescence and aging‐related bone loss. The expression level of YBX1 in BMSCs decreases during aging.Ybx1 overexpression in BMSCs stimulates bone formation, while its deletion accelerates bone loss.YBX1 suppresses BMSC senescence and modulates BMSC differentiation by controlling RNA splicing.The small compound sciadopitysin attenuates YBX1 degradation and bone loss in old mice. [ABSTRACT FROM AUTHOR]

Abstract: Background: The relationship between the levels of gonadotropic hormones and bone metabolism-related cytokines in Chinese women is unclear. We investigated the relationship between FSH and LH and OPG, leptin, TGF-β1, and TGF-β2 in Chinese women. Methods: A cross-sectional study of 694 Chinese women, aged 20 to 82y was conducted. Levels of serum FSH, LH, OPG, leptin, TGF-β1, and TGF-β2 were determined. Results: In premenopausal females, serum follicle-stimulating hormone (FSH) and luteinizing hormone (LH) levels seemly showed no correlation with the cytokine levels. In perimenopausal females, serum FSH and LH levels showed significant positive correlation with osteoprotegerin (OPG) and transforming growth factor-β2 (TGF-β2) levels (r =0.286 to 0.405, all P =0.000), whereas they showed negative correlation with TGF-β1 levels (r =−0.413 and −0.354, all P =0.000). In postmenopausal females, FSH and LH levels showed positive correlation with OPG levels (r =0.247 and 0.241, all P =0.000), negative correlation with leptin and TGF-β1 levels (r =−0.234 to −0.319, all P =0.000), and no correlation with TGF-β2 levels. Multiple linear regression stepwise analysis revealed the following results. In premenopausal females, 2.0% and 1.5% of the changes in LH could be explained by OPG and leptin, respectively, while 1.9% of the changes in OPG could be explained by LH. In perimenopausal females, the determinants of OPG and TGF-β1 on FSH were 10.9% and 17.0%, respectively, and the determinants of OPG, TGF-β1 and TGF-β2 on LH were 4.5%, 4.9% and 16.4%, respectively. The determinants of FSH and LH on OPG were 14.5% and 2.5%, respectively. The determinant of FSH on TGF-β1 was 4.5%, while the determinant of LH on TGF-β2 was 16.4%. In postmenopausal females, the determinants of leptin and OPG on FSH were 10.2% and 2.8%, respectively, and the determinants of OPG and TGF-β1 on LH were 5.8% and 2.3%, respectively. The determinant of FSH on OPG, leptin and TGF-β1 were 6.1%, 3.4% and 9.2%. Conclusions: These results indicate that age-related gonadotropic hormone levels are associated with changes in OPG, TGF-β1, TGF-β2 and leptin, and change with menopausal status. [Copyright &y& Elsevier]

Autophagy is an evolutionarily conserved intracellular process and is considered one of the main catabolism pathways. In the process of autophagy, cells are digested nonselectively or selectively to recover nutrients and energy, so it is regarded as an antiaging process. In addition to the essential role of autophagy in cellular homeostasis, autophagy is a stress response mechanism for cell survival. Here, we review recent literature describing the pathway of autophagy and its role in different bone cell types, including osteoblasts, osteoclasts, and osteocytes. Also discussed is the mechanism of autophagy in bone diseases associated with bone homeostasis, including osteoporosis and Paget's disease. Finally, we discuss the application of autophagy regulators in bone diseases. This review aims to introduce autophagy, summarize the understanding of its relevance in bone physiology, and discuss its role and therapeutic potential in the pathogenesis of bone diseases such as osteoporosis. [ABSTRACT FROM AUTHOR]

Objectives: CD31hiEMCNhi vessels (CD31, also known as PECAM1 [platelet and endothelial cell adhesion molecule 1]; EMCN, endomucin), which are strongly positive for CD31 and endomucin, couple angiogenesis and osteogenesis. However, the role of CD31hiEMCNhi vessels in bone regeneration remains unknown. In the present study, we investigated the role of CD31hiEMCNhi vessels in the process of bone regeneration. Materials and Methods: We used endothelial‐specific Krüppel like factor 3 (Klf3) knockout mice and ophiopogonin D treatment to interfere with CD31hiEMCNhi vessel formation. We constructed a bone regeneration model by surgical ablation of the trabecular bone. Immunofluorescence and micro‐computed tomography (CT) were used to detect CD31hiEMCNhi vessels and bone formation. Results: CD31hiEMCNhi vessels participate in the process of bone regeneration, such that endothelial‐specific Klf3 knockout mice showed increased CD31hiEMCNhi vessels and osteoprogenitors in the bone regeneration area, and further accelerated bone formation. We also demonstrated that the natural compound, ophiopogonin D, acts as a KLF3 inhibitor to promote vessels formation both in vitro and in vivo. Administration of ophiopogonin D increased the abundance of CD31hiEmcnhi vessels and accelerated bone healing. Conclusions: Our findings confirmed the important role of CD31hiEmcnhi vessels in bone regeneration and provided a new target to treat bone fracture or promote bone regeneration. [ABSTRACT FROM AUTHOR]

Bone marrow mesenchymal stem cells (BMSCs) exhibit an age-related lineage switch between osteogenic and adipogenic fates, which contributes to bone loss and adiposity. Here we identified a long noncoding RNA, Bmncr, which regulated the fate of BMSCs during aging. Mice depleted of Bmncr (Bmncr-KO) showed decreased bone mass and increased bone marrow adiposity, whereas transgenic overexpression of Bmncr (Bmncr-Tg) alleviated bone loss and bone marrow fat accumulation. Bmncr regulated the osteogenic niche of BMSCs by maintaining extracellular matrix protein fibromodulin (FMOD) and activation of the BMP2 pathway. Bmncr affected local 3D chromatin structure and transcription of Fmod. The absence of Fmod modified the bone phenotype of Bmncr-Tg mice. Further analysis revealed that Bmncr would serve as a scaffold to facilitate the interaction of TAZ and ABL, and thus facilitate the assembly of the TAZ and RUNX2/PPARG transcriptional complex, promoting osteogenesis and inhibiting adipogenesis. Adeno-associated viral-mediated overexpression of Taz in osteoprogenitors alleviated bone loss and marrow fat accumulation in Bmncr-KO mice. Furthermore, restoring BMNCR levels in human BMSCs reversed the age-related switch between osteoblast and adipocyte differentiation. Our findings indicate that Bmncr is a key regulator of the age-related osteogenic niche alteration and cell fate switch of BMSCs. [ABSTRACT FROM AUTHOR]

Femoral neck geometric parameters (FNGPs) are closely related to the strength of the femoral neck and the risk of fragility fractures. No reference database is available for FNGPs for Chinese population, and gender-related differences in FNGPs as well as their association with the risk of femoral neck fractures are unknown. This investigation aimed to set up reference databases for FNGPs, understand gender-related differences in FNGPs, and examine the association between FNGPs and the risk of osteoporotic fractures of the femoral neck. This study included 5268 females and 2156 males (aged 15–91 years) from Chinese population. A total of 384 patients (282 females and 102 males) had sustained femoral neck fractures; 384 age- and sex-matched individuals without any fractures served as controls. Femoral neck DXA images were used to measure bone mineral density (BMD) and eight FNGPs. Our results showed that the age-related trends of FNGPs were fitted with the best goodness-of-fit by applying the cubic regression model. The trends shown by FNGPs were significantly different between male and female subjects, and the fitting curves were significantly higher in male subjects. After adjustments were made for age, height, weight, and body mass index, Cox regression analysis showed that changes in all FNGPs were related to increased hazard ratios (HRs) of femoral neck fractures. After further adjustment was made for BMD of the femoral neck, the HRs related to a cortical thickness (CT) decrease and buckling ratio (BR) increase in females went up by 3.35-folds (95% CI: 2.75–4.07) and 1.86-folds (95% CI: 1.33–2.60), respectively. In males, the HRs related to the decrease in CT and cross-sectional area (CSA) increased by 3.21-folds (95% CI: 2.32–4.45) and 1.88-folds (95% CI: 1.03–3.44), respectively. In conclusions, the reference databases of FNGPs established in this study will assist in the evaluation and prediction of femoral neck fracture risk in the clinic. The decrease in CT and increase in BR of the femoral neck were independent risk factors for osteoporotic fractures of the femoral neck in females from mainland China, while a decrease in CT and CSA were risk factors in male. [ABSTRACT FROM AUTHOR]

Growth differentiation factor 11 (GDF11) is a member of the transforming growth factor-β superfamily. Recent studies confirmed that GDF11 plays an important role in regulating the regeneration of brain, skeletal muscle, and heart during aging; however, its role in bone metabolism remains unclear. Thus, the aim of this study was to determine the effects of GDF11 on bone metabolism, including bone formation and bone resorption, both in vitro and in vivo. Our results showed that GDF11 inhibited osteoblastic differentiation of bone marrow mesenchymal stem cells in vitro. Mechanistically, GDF11 repressed Runx2 expression by inducing SMAD2/3 phosphorylation during osteoblast differentiation. Moreover, intraperitoneal injection of GDF11 inhibited bone formation and accelerated age-related bone loss in mice. Our results also showed that GDF11 had no effect on osteoclast differentiation or bone resorption both in vitro and in vivo. These results provide a further rationale for the therapeutic targeting of GDF11 for the treatment of age-related osteoporosis. [ABSTRACT FROM AUTHOR]

We previously reported that Runx2/miR-3960/miR-2861 regulatory feedback loop stimulates osteoblast differentiation. However, the effect of this feedback loop on the osteogenic transdifferentiation of vascular smooth muscle cells (VSMCs) remains unclear. Our recent study showed that miR-2861 and miR-3960 expression increases significantly during β-glycerophosphate-induced osteogenic transdifferentiation of VSMCs. Overexpression of miR-2861 or miR-3960 in VSMCs enhances β-glycerophosphate-induced osteoblastogenesis, whereas inhibition of miR-2861 or miR-3960 expression attenuates it. MiR-2861 or miR-3960 promotes osteogenic transdifferentiation of VSMCs by targeting histone deacetylase 5 or Homeobox A2, respectively, resulting in increased runt-related transcription factor 2 (Runx2) protein production. Furthermore, overexpression of Runx2 induces miR-2861 and miR-3960 transcription, and knockdown of Runx2 attenuates β-glycerophosphate-induced miR-2861 and miR-3960 transcription in VSMCs. Thus, our data show that Runx2/miR-3960/miR-2861 positive feedback loop plays an important role in osteogenic transdifferentiation of VSMCs and contributes to vascular calcification. [ABSTRACT FROM AUTHOR]

Abstract: Background: Cytokines including transforming growth factor beta 1 (TGF-β1) and osteoprotegerin (OPG) are closely related to bone metabolism. However, the relationships between TGF-β1, OPG and risk of osteoporosis in native Chinese women are unknown. Our research indicated that there is a positive correlation between TGF-β1 and bone mineral density (BMD) T-score, and a negative correlation between OPG and T-score. The risk of osteoporosis is reduced as TGF-β1 increases and increases as OPG was raised. We investigated correlations of BMD T-scores with circulating TGF-β1 and BMD T-scores with circulating OPG in healthy native Chinese women, and to study the effects of changes in TGF-β1 and OPG on osteoporosis. Methods: This was a cross-sectional study of 691 healthy native Chinese women aged 20–80years. Concentrations of serum TGF-β1 and OPG were determined. BMD T-scores at the posteroanterior spine, left hip, and forearm were measured by dual-energy X-ray absorptiometry. Results: There were positive correlations between serum TGF-β1 and T-scores at the various skeletal sites (r =0.167–0.285, all P =0.000) and negative correlations between serum OPG and T-scores (r =−0.179 to −0.270, all P =0.007–0.000). After adjusting for age and BMI, correlations between TGF-β1 and T-score at the lumbar vertebrae and ultradistal forearm, and between OPG and T-score at the ultradistal forearm in premenopausal subjects, remained statistically significant. Multivariate linear stepwise analysis showed that TGF-β1 could explain 1.9–8.3% of T-score variation at each skeletal site. OPG could explain 2.4–4.4% of T-score variation. When TGF-β1 and OPG concentrations were grouped according to quartile intervals, T-score and the prevalence and risk of osteoporosis varied with changes in the cytokines. Conclusions: The risk of osteoporosis in native Chinese women increased as circulating TGF-β1 was reduced and OPG was raised. [Copyright &y& Elsevier]

Abstract: Background: It remains unclear whether gonadotropins or estrogen is responsible for early bone mineral density (BMD) decrease in Chinese women. Methods: A cross-sectional study was conducted on 368 healthy adult women, aged 35–60years. We measured BMD, calculated BMD decrease rates (BDRs) and assessed serum follicle-stimulating hormone (FSH), luteinizing hormone (LH) and estradiol (E2) levels. Results: BDR was significantly negatively correlated with serum FSH (r =−0.429 to −0.622, all p =0.000) and LH (r =−0.359 to −0.526, all p =0.000). After adjustment for age and body mass index, the negative correlations of serum FSH and LH with BDR persisted, but there was no overall correlation between serum E2 and BDR. Multiple linear stepwise regression analysis suggested that serum FSH is a negative determinant of BDR. Serum E2 seems to be a positive determinant of BDR in a few parts of the skeleton. Conclusions: The decrease of BMD during the menopause is associated with FSH and LH levels, rather than E2 in Chinese women. [Copyright &y& Elsevier]

Abstract: Adiponectin may exert a negative effect on bone metabolism by regulating osteoprotegerin (OPG) and receptor activator of nuclear factor-κB ligand (RANKL) expression. However, the action of adiponectin on bone may be influenced by estrogen in women. The present study was undertaken to investigate the effects of 17β-estradiol (E2) on adiponectin-regulated OPG and RANKL expression in human osteoblast. Human osteoblasts were treated with α-MEM containing 10μg/ml adiponectin alone or together with 10−10 to 10−8 M E2 for 12–48h. Cells were also treated with α-MEM containing 10μg/ml adiponectin together with 10−8 M E2 plus p38 agonist-anisomycin or estrogen receptor (ER) antagonist ICI182780 for 48h. The effects of E2 were also investigated by knockdown of ERs or overexpression of p38 MAPK in osteoblasts. Further, we examined the effects of E2 on adiponectin-dependent osteoclastogenesis by the co-culture systems of osteoblast and CD14+ peripheral blood monocytes (PBMCs). Real-time quantitative PCR (RT-PCR) and ELISA were used to detect OPG/RANKL mRNA and their corresponding protein expression, Western Blot was used to analyze the phosphorylated p38 (p-p38) levels. The results showed that E2 blocked adiponectin-induced p38 phosphorylation, decreased adiponectin-regulated OPG/RANKL mRNA and protein expression in a dose- and time-dependent manner. ICI182780 or knockdown of ERs abolished the effects of E2 on adiponectin-dependent p38 phosphorylation and OPG/RANKL expression. Furthermore, anisomycin or overexpression of p38 also reserved the effects of E2 on adiponectin-dependent p38 phosphorylation and OPG/RANKL expression. E2 inhibited adiponectin-dependent osteoclastogenesis in the co-culture systems of osteoblast and CD14+ PBMCs, whereas anisomycin, ICI182780, knockdown of ERs and overexpression of p38 significantly reversed this response. In conclusions, our findings demonstrated, through blocking the activation of adiponectin-induced p38 MAPK, E2 suppressed the adiponectin-regulated OPG/RANKL expression and then inhibited osteoclastogenesis, which suggested that estrogen would suppress the effect of adiponectin on bone metabolism. [Copyright &y& Elsevier]

Aims Omentin-1 (also known as intelectin-1) is a recently identified visceral adipose tissue-derived cytokine that is inversely related to obesity. Our previous study showed that omentin-1 inhibits osteoblastic differentiation of calcifying vascular smooth muscle cells (CVSMCs) in vitro. This study was undertaken to investigate the effects of omentin-1 on arterial calcification and bone metabolism in vivo. Methods and results In vitro, omentin-1 stimulated production of osteoprotegerin (OPG) and inhibited production of receptor activator for nuclear factor κB ligand (RANKL) in both CVSMCs and osteoblasts. In vivo, adenovirus-mediated over-expression of omentin-1 attenuated arterial calcification and bone loss in OPG−/− mice. All these in vitro and in vivo actions were abrogated by blockade of the PI3K–Akt signalling pathway. Furthermore, omentin-1 reduced serum levels of RANKL, tartarate-resistant acid phosphatase-5b and osteocalcin, all of which are increased dramatically in OPG−/− mice. Conclusion These data suggest that omentin-1 ameliorates arterial calcification and bone loss in vivo through the regulation of the RANK signalling pathway. [ABSTRACT FROM PUBLISHER]

Abstract: Background: Adiponectin and leptin, as the main circulating peptides secreted by adipose tissue, are potential contributors to bone metabolism. However, their association with bone mineral density (BMD) is unknown. We investigated whether these serum adipocytokines concentrations are associated with BMD and bone turnover markers. Methods: Serum adiponectin, leptin concentrations, bone turnover biochemical markers, and BMD were determined in 265 premenopausal and 336 postmenopausal Chinese women. Results: In postmenopausal Chinese women, the multiple linear stepwise regression analysis showed that year since menopause, lean mass, estradiol, and adiponectin, but not fat mass, leptin, were independent predictors of BMD in postmenopausal Chinese women. However, in premenopausal Chinese women, adiponectin was not the predictor of BMD. The significant positive correlations between adiponectin and bone-specific alkaline phosphatase (BAP), bone cross-linked N-telopeptides of type I collagen (NTX) were found only in postmenopausal women. Serum BAP, and NTX, but not adiponectin, decreased in response to alendronate therapy. Conclusions: Adiponectin was an independent predictor of BMD, and positively correlated with bone turnover biochemical markers in postmenopausal Chinese women, but not premenopausal women. It suggested that adiponectin may exert a negative effect on bone mass by promoting excessive bone resorption associated with bone loss. However, these effects may be mediated by menopausal status. [Copyright &y& Elsevier]

Abstract: Background: Transforming growth factor-β 1 (TGF-β1), TGF-β2, osteoprotegerin (OPG), and leptin are important cytokines in the regulation of bone remodeling. We investigated the relationship of TGF-β1 and TGF-β2 concentrations with those of OPG and leptin in Chinese females. Methods: The serum concentrations of TGF-β1, TGF-β2, OPG, and leptin were measured by ELISA in 459 healthy Chinese females aged 25–80 y. Results: The mean values (±SD) of the serum concentrations of TGF-β1, TGF-β2, OPG, and leptin in Chinese females were 29.7±1.69 μg/l, 13.7±3.86 μg/l, 3.81±1.96 pmol/l, and 10.5±2.01 μg/l, respectively. Further, the serum TGF-β1 concentrations of postmenopausal women were significantly lower than those of perimenopausal and premenopausal women (24.3±1.59 vs 33.4±1.69 and 37.6±1.64, respectively), while the TGF-β2 concentrations of postmenopausal women were significantly higher than those of perimenopausal and premenopausal women (14.6±3.91 vs 13.5±3.93 and 11.7±2.68, respectively). The serum TGF-β1 concentration was found to be significantly negatively correlated with age (r =−0.335, P =0.000) and the TGF-β2 concentration, to be significantly positively correlated with age (r =0.230, P =0.000). The TGF-β1 concentration was found to be significantly negatively correlated with both TGF-β2 (r =−0.261, P =0.000) and OPG (r =−0.313, P =0.000) concentrations; a significantly positive correlation was found between the TGF-β1 and leptin concentrations (r =0.164, P =0.000) and between TGF-β2 and OPG concentrations (r =0.432, P =0.000). Conclusion: These results provide age-related reference values of TGF-β1 and TGF-β2 in Chinese adult women, and reveal the relationships between these cytokines. [Copyright &y& Elsevier]

Abstract: Background: Follicle-stimulating hormone (FSH) and luteinizing hormone (LH) may play an important role in bone mass regulation in postmenopausal women. Methods: A cross-sectional study of 699 healthy Chinese women, aged 20 to 82 y, was conducted. Serum FSH and LH and BMD were measured at the posteroanterior (PA) spine, lateral spine, total hip, and distal forearm. Results: The geometric mean values (±SD) of serum FSH and LH in premenopausal women were 3.94±2.08 and 7.51±2.58 IU/l, respectively, and in postmenopausal women were 28.8±1.88 and 25.6±1.95 IU/l, respectively. The correlation of FSH to BMD at different skeletal regions (r =−0.597–−0.492, P =0.000) was higher than that of LH to BMD (r =−0.452–−0.332, P =0.000). The prevalences of osteoporosis for the quartiles of FSH at various skeletal sites were 0.57%, 0.43%, 27.1%, and 30.9%, respectively; and of LH were 2.14%, 4.43%, 19.5%, and 26.0%, respectively. The prevalence of osteoporosis in 3rd and 4th quartile was more significantly increased than the 1st and 2nd quartile. Conclusions: These data suggest that FSH and LH levels in circulation are associated with BMD changes and osteoporosis occurrence in Chinese women. [Copyright &y& Elsevier]

Abstract: Background: Osteoprotegerin (OPG) and leptin are important cellular factors in the regulation of bone remodeling. We investigated the serum OPG and leptin in Chinese women. Methods: The serum OPG and leptin in 690 Chinese women aged 20–81y were measured by an ELISA. The values of OPG and leptin in women of other races were acquired from previous reports on the same. Results: The geometric mean values (± SD) of the serum OPG and leptin in Chinese women were 3.42±1.91pmol/l and 10.5±1.99μg/l, respectively. Further, the serum OPG (4.39±1.85 vs 2.74±1.81) and leptin (11.4±2.21 vs 9.68±1.81) in postmenopausal women were significantly higher than in premenopausal women. The serum OPG in middle-aged Chinese women was significantly higher than that in middle-aged Austrian and Icelandic women; however, this is quite contrary to the results obtained in the case of old-aged women. The values of serum leptin in Chinese women were significantly lower than those in white, black, and Mexican American women. Conclusions: These results provide reliable reference values for OPG and leptin in Chinese adult women. The serum of OPG and leptin differ with ethnicity. [Copyright &y& Elsevier]

Abstract: WISP3 is essential for maintaining cartilage integrity mainly by regulating the expression of collagen II, and mutations of WISP3 linked to spondyloepiphyseal dysplasia tarda with progressive arthropathy (SEDT-PA) can compromise this function and lead to cartilage loss. The aim of this study was to evaluate the effect of WISP3 on insulin-like growth factor (IGF) signaling in human chondrocytes, investigate whether WISP3 up-regulates collagen II through the IGF signaling pathway, and compare IGF signaling between wild-type and mutant WISP3. Experimental results suggest that WISP3 up-regulates collagen II expression and inhibits the activation of IGF-IR, IRS-1, and ERK kinase in human chondrocytes, and mutation of WISP3 augments IGF signaling in human chondrocytes. In addition to the IGF signaling pathway, WISP3 might up-regulate collagen II expression through an IGF-independent signaling cascade. [Copyright &y& Elsevier]

Abstract: The aim of this study was to investigate the effects of apelin on proliferation and apoptosis of mouse osteoblastic MC3T3-E1 cells. APJ was expressed in MC3T3-E1 cells. Apelin did not affect Runx2 expression, alkaline phosphatase (ALP) activity, osteocalcin and type I collagen secretion, suggesting that it has no effect on osteoblastic differentiation of MC3T3-E1 cells. However, apelin stimulated MC3T3-E1 cell proliferation and inhibited cell apoptosis induced by serum deprivation. Our study also shows that apelin decreased cytochrome c release and caspase-3, capase-8 and caspase-9 activation in serum-deprived MC3T3-E1 cells. Apelin activated c-Jun N-terminal kinase (JNK) and Akt (phosphatidylinositol 3-kinase downstream effector), and the JNK inhibitor SP600125, the phosphatidylinositol 3-kinase (PI3-K) inhibitor LY294002 or the Akt inhibitor 1L-6-hydroxymethyl-chiro-inositol 2-(R)-2-O-methyl-3-O-octadecylcarbonate (HIMO) inhibited its effects on proliferation and serum deprivation-induced apoptosis. Furthermore, apelin protected against apoptosis induced by the glucocorticoid dexamethasone or TNF-α. Apelin stimulates proliferation and suppresses serum deprivation-induced apoptosis of MC3T3-E1 cells and these actions are mediated via JNK and PI3-K/Akt signaling pathways. [Copyright &y& Elsevier]

Abstract: Objectives: Apelin is a recently discovered peptide that is the endogenous ligand for the orphan G-protein-coupled receptor APJ. Adipocytes can express and secrete apelin. The aim of this study was to characterize apelin and APJ expression in human osteoblasts and to investigate the effects of apelin on osteoblasts. Results: Apelin and APJ were expressed in human osteoblasts. Apelin stimulated proliferation of human osteoblasts, but had no effect on alkaline phosphatase (ALP) activity, osteocalcin and type I collagen production in human osteoblasts. Suppression of APJ with small-interfering RNA (siRNA) abolished the apelin-induced cell proliferation. Apelin induced activation of Akt (Phosphatidylinositol-3 kinase downstream effector), but not MAPKs, such as c-jun N-terminal Kinase (JNK), p38 and ERK1/2 in human osteoblasts. This effect was blocked by suppression of APJ with siRNA. Furthermore, LY294002 (PI3 kinase inhibitor) blocked the activation of Akt by apelin and abolished the apelin-induced cell proliferation. Conclusions: Human osteoblasts express apelin and APJ and apelin enhances human osteoblast proliferation, but has no effect on osteoblast differentiation, and APJ/PI3 kinase/Akt pathway is involved in the proliferation response. These findings suggest that apelin may function as a mitogenic agent for osteoblasts. [Copyright &y& Elsevier]

Abstract: Background: Previous studies have shown that bone turnover rate changes with age. At the same time, there is no definitive research regarding age-related changes of bone turnover level and its association with bone mineral density (BMD) in Chinese mainland women. Methods: In a cohort of 663 Chinese mainland women aged 20–70 years, serum bone alkaline phosphatase (BAP) and serum cross-linked C-telopeptides of type I collagen (sCTX) were measured to evaluate the state of bone formation and resorption, respectively. BMD was measured in the posteroanterior spine, supine lateral spine, hip and forearm using a dual-energy X-ray absorptiometry. Results: The cubic polynomial regression model best fit age-related changes in serum BAP (R 2 =0.398, p <0.001) and sCTX concentrations (R 2 =0.148, p <0.001) with largest R 2 from comparison 8 different regression models. Their values reached a minimal level in the 30–39 years age group, and increased dramatically in the 40–59 years groups. There was a decreasing trend of BAP in women >60 years. The levels of BAP and sCTX were inversely correlated to BMD in various skeletal regions over the entire population (r =−0.096 to −0.357, p <0.05). sCTX was a significant predictor of a T-score≤−2.5 of BMD in postmenopausal women with sCTX levels above mean+2 SD of women aged 30–39 years compared with other postmenopausal women, which indicated by odds ratios 1.9–3.7 (p <0.05) for various skeletal regions, especially for the lateral lumbar spine (2.2, p <0.01), Ward''s triangle (3.7, p <0.01), and ultradistal end of radius+ulna (2.8, p <0.001). Conclusions: Age-dependent serum BAP and sCTX were inversely correlated to BMD, and sCTX was a useful parameter for the prediction of a low T-score of BMD at skeletal sites with abundant cancellous bone in postmenopausal women. [Copyright &y& Elsevier]

Abstract: Background: Serum leptin concentration is associated with age, fat and bone mineral density (BMD), and there are ethnic differences in physique and BMD values. The relationship between serum leptin concentration and BMD in Chinese women is presently unknown. We examined the relationship of serum leptin concentration with age, body weight, BMI and BMD in mainland Chinese women. Methods: Serum leptin concentration in a population of 676 Chinese females, aged 20–80 years (45.4±14.8 years, mean±S.D.), was measured using an enzyme-linked immunosorbent assay (ELISA). BMD values were measured by dual-energy X-ray absorptiometry (DXA) at a number of sites: the posteroanterior lumbar spine (PA, L1–L4), lateral lumbar spine (Lat, L2–L4), hip (including the femoral neck (FN) and total hip (T-hip)), and forearm (one-third region (RU1/3) and total region (RUT)). The relationship between changes in serum leptin concentration with age, body weight, height, body mass index (BMI) and BMD values at six sites were analyzed using 10 different regression models, and the models giving the best fit were selected. Results: The cubic regression model best described the changes in serum leptin concentration with age (R2=0.048, p<0.001) and body weight (R2=0.352, p<0.001), while the quadratic regression model was best for serum leptin concentration changes with BMI (R2=0.410, p<0.001); serum leptin concentration was not correlated with height. Serum leptin concentration was significantly higher in postmenopausal than premenopausal women (p<0.001). Serum leptin concentration was correlated with Lat BMD (R2=0.012, p<0.018), FN BMD (R2=0.006, p<0.041) and T-hip BMD (R2=0.013, p<0.004) in the whole population. In premenopausal women, leptin was positively associated with BMD except for Lat (R2=0.029–0.055, p<0.008); in postmenopausal women, leptin was also positively associated with AP, FN, T-hip BMD (R2=0.026–0.042, p<0.007). However, after adjusting for BMI not for body weight, there was no association between serum leptin concentration and age. Meanwhile, after adjustment for age, body weight and BMI, there was no association between serum leptin concentration and BMD values in premenopausal and postmenopausal women. Conclusions: Age-related changes in serum leptin concentration is dependent on BMI, but not a direct determinant of BMD in Chinese females. [Copyright &y& Elsevier]

Receptor activator of NF-κB(RANK) ligand(RANKL), expressed by cells of the osteoblast lineage binds to RANK, induces signaling and a gene expression cascade that leads to osteoclast differentiation and activation. Recently, osteoblast-derived membrane-type matrix metalloproteinases-1(MT1-MMP) have been implicated in the process of bone resorption by degrading bone matrix. In the present study, we investigated the effects of parathyroid hormone[PTH(1–34)] on RANKL and MT1-MMP production in cultured normal human osteoblast-like cells(hOB). In reverse transcription polymerase chain reaction studies, we observed that PTH(1–34) induced RANKL messenger ribonucleic acid(mRNA) expression. Activity assays demonstrated that PTH(1–34) simultaneously inhibited MT1-MMP protein expression in a dose- and time-dependent manner. The effect of PTH(1–34) on MT1-MMP production was parallel to that on RANKL expression, suggesting a tight inverse relationship between MT1-MMP and RANKL expression. Our findings indicated that the decreased MT1-MMP expression by PTH may be involved in RANKL signaling in osteoblasts and activation of osteoclasts. [ABSTRACT FROM AUTHOR]

OBJECTIVE: To study the effects of progesterone (P) and 18-methyl levonorgestrel (LNG) on the expression of P receptors (PRA and PRB), c-fos, c-jun, and osteocalcin in the human osteosarcoma cell line MG-63 and human normal osteoblasts in order to understand the mechanism of progestin action on the proliferation and differentiation of osteoblasts. METHODS: Cell proliferation was tested by MTT assay. The expression of PR, c-fos, c-jun, and osteocalcin was measured by semi-quantitative RT-PCR, Western immunoblot or immunocytochemistry. RESULTS: Progesterone and LNG (10−10–10−6 M) stimulated cell proliferation in a dose-dependent manner. Progesterone and LNG did not affect the expression of PRA and PRB mRNA and protein, but c-fos and c-jun mRNA and protein were upregulated in a dose-dependent manner. The expression of osteocalcin mRNA was also increased in human osteoblasts in a dose and time-dependent manner with greater effects of LNG than P, while the expression of osteocalcin mRNA in MG-63 cells was not changed by P or LNG. CONCLUSION: Progesterone and LNG promote osteocalcin gene transcription by stimulating the expression of c-fos and c-jun, and result in osteoblast proliferation and differentiation. [ABSTRACT FROM AUTHOR]

With the increasing aging population, aging‐associated diseases are becoming epidemic worldwide, including aging‐associated metabolic dysfunction. However, the underlying mechanisms are poorly understood. In the present study, we aimed to investigate the role of microRNA miR‐188 in the aging‐associated metabolic phenotype. The results showed that the expression of miR‐188 increased gradually in brown adipose tissue (BAT) and inguinal white adipose tissue (iWAT) of mice during aging. MiR‐188 knockout mice were resistant to the aging‐associated metabolic phenotype and had higher energy expenditure. Meanwhile, adipose tissue‐specific miR‐188 transgenic mice displayed the opposite phenotype. Mechanistically, we identified the thermogenic‐related gene Prdm16 (encoding PR domain containing 16) as the direct target of miR‐188. Notably, inhibition of miR‐188 expression in BAT and iWAT of aged mice by tail vein injection of antagomiR‐188 ameliorated aging‐associated metabolic dysfunction significantly. Taken together, our findings suggested that miR‐188 plays an important role in the regulation of the aging‐associated metabolic phenotype, and targeting miR‐188 could be an effective strategy to prevent aging‐associated metabolic dysfunction. [ABSTRACT FROM AUTHOR]