Aminobisphosphonates Cause Osteoblast Apoptosis and Inhibit Bone Nodule Formation In Vitro

Bisphosphonates are widely used for the treatment of bone diseases associated with increased osteoclastic bone resorption. Bisphosphonates are known to inhibit biochemical markers of bone formation in vivo, but it is unclear to what extent this is a consequence of osteoclast inhibition or a direct inhibitory effect on cells of the osteoblast lineage. In order to investigate this issue, we studied the effects of various bisphosphonates on osteoblast growth and differentiation in vitro. The aminobisphosphonates pamidronate and alendronate inhibited osteoblast growth, caused osteoblast apoptosis, and inhibited protein prenylation in osteoblasts in a dose-dependent manner over the concentration range 20-100 microM. Further studies showed that alendronate in a dose of 0.1 mg/kg inhibited protein prenylation in calvarial osteoblasts in vivo, indicating that alendronate can be taken up by osteoblasts in sufficient amounts to inhibit protein prenylation at clinically relevant doses. Pamidronate and alendronate inhibited bone nodule formation at concentrations 10-fold lower than those required to inhibit osteoblast growth. These effects were not observed with non-nitrogen-containing bisphosphonates or with other inhibitors of protein prenylation and were only partially reversed by cotreatment with a fourfold molar excess of ss-glycerol phosphate. We conclude that aminobisphosphonates cause osteoblast apoptosis in vitro at micromolar concentrations and inhibit osteoblast differentiation at nanomolar concentrations by mechanisms that are independent of effects on protein prenylation and may be due in part to inhibition of mineralization. While these results need to be interpreted with caution because of uncertainty about the concentrations of bisphosphonates that osteoblasts are exposed to in vivo, our studies clearly demonstrate that bisphosphonates exert strong inhibitory effects on cells of the osteoblast lineage at similar concentrations to those that cause osteoclast inhibition. This raises the possibility that inhibition of bone formation by bisphosphonates may be due in part to a direct inhibitory effect on cells of the osteoblast lineage.     

Immunoreactive serum calcitonin and skeletal histology in chronic renal failure

Serum calcitonin and serum parathyroid hormone (PTH) were measured in 50 patients undergoing regular haemodialysis for end-stage chronic renal failure, and an analysis of osteoclast and osteoblast activities was made in bone biopsies obtained by iliac crest trephine. Osteoclast and osteoblast activities were studied in a multivariate analysis in relation to factors which might reasonably be thought to influence activity, namely serum calcitonin, serum PTH, serum calcium, serum inorganic phosphate, and bone aluminium. Only serum PTH correlated strongly with osteoclast activity (p = 0.0047). Serum PTH correlated also with osteoblast activity (p = 0.0024). Serum inorganic phosphate correlated negatively with osteoblast activity (p = 0.0082). Serum calcitonin did not correlate with osteoclast or osteoblast activities but did correlate strongly with bone aluminium in a multivariate analysis (p = 0.0078). Bone aluminium did not correlate independently with osteoclast or osteoblast activities. This study affirms the implied powerful role of PTH in influencing osteoclast and osteoblast activities in end-stage chronic renal failure.     

Alendronate affects long bone length and growth plate morphology in the oim mouse model for Osteogenesis Imperfecta

Alendronate, a bisphosphonate drug, has shown promise in reducing remodeling and bone loss in postmenopausal osteoporosis. Alendronate acts directly on the osteoclast, inhibiting its resorption capability. This inhibition of osteoclast activity has led to the use of bisphosphonates in the treatment of the osteogenesis imperfecta condition. Treatment of osteogenesis imperfecta with bisphosphonates enhances bone strength, but the consequences on linear bone growth are not well defined. Using the oim mouse model for type III osteogenesis imperfecta, two doses of alendronate, low (0.125 mg/kg/wk) and high (2.5 mg/kg/wk) were administered weekly via intraperitoneal injection starting at 4 weeks of age and ending at 12 weeks of age to assess the effects of alendronate on humerus and ulna length. The higher dose of alendronate reduced humerus and ulna length in the oim/wt and wt/wt genotypes for both sexes (P < 0.05). The oim/oim humerus and ulna were not significantly affected by the higher dose of alendronate in females, but reduced bone length in males (P < 0.0085). Proximal humerus growth plate area was affected by both genotype and alendronate dose and growth plate diameter was increased at the chondro-osseous junction by both alendronate doses (P < 0.011). Genotype and alendronate dose affected growth plate height. The oim/oim genotype displayed taller growth plates. The high dosage of alendronate increased overall growth plate height, particularly within the hypertrophic zone, which suggests a failure of vascular invasion-induced apoptosis in the hypertrophic cells. In conclusion, these results indicate that high doses of alendronate (>2.5 mg/kg/wk) inhibit long bone length in mice through alteration of the growth plate and possibly reduced resorption at the chondro-osseous junction.     

Bone origin of the serum complex of calcium, phosphate, fetuin, and matrix Gla protein: biochemical evidence for the cancellous bone-remodeling compartment.

We previously described the discovery of a fetuin-matrix Gla protein (MGP)-mineral complex in the serum of rats treated with the bone-active bisphosphonate etidronate and showed that the appearance of this complex in serum correlates with the inhibition of bone mineralization by etidronate. In this study we show that the inhibition of bone resorption by treatment with the hormone calcitonin, the cytokine osteoprotegerin, or the drug alendronate, completely inhibits the generation of the fetuin-mineral complex in response to etidronate injection. These observations can be explained best by the bone-remodeling compartment (BRC), a cancellous bone compartment in which the concentrations of calcium and phosphate are determined directly by the combined actions of the osteoclast and the osteoblast. When bone mineralization is acutely inhibited by etidronate, the BRC model predicts that the continuing action of osteoclasts will cause a sharp rise in the concentrations of calcium and phosphate in the aqueous solution of the BRC with the consequent spontaneous formation of calcium phosphate crystal nuclei in which growth then would be arrested by formation of a complex with fetuin. When the inhibition of bone resorption by calcitonin, osteoprotegerin, or alendronate is combined with the acute inhibition of bone mineralization with etidronate, the BRC model correctly predicts that there will no longer be a sharp rise in calcium and phosphate, and, therefore, there will no longer be the formation of the fetuin-mineral complex. The vascular nature of the BRC is supported by the observations that the fetuin component of the fetuin-mineral complex is derived from plasma fetuin and that the fetuin mineral complex appears in plasma within minutes of the inhibition of bone mineralization with etidronate.     

Comparative Study on Osteoconductivity by Synthetic Octacalcium Phosphate and Sintered Hydroxyapatite in Rabbit Bone Marrow

Octacalcium phosphate (OCP) is thought to be a precursor of the mineral crystals in biological apatite. Synthetic OCP has been shown to be converted into an apatite structure when implanted in murine calvarial bone, to enhance bone regeneration more than synthetic hydroxyapatite (HA), and to degrade faster than biodegradable β-tricalcium phosphate. This study was designed to investigate whether OCP implantation enhances the formation and resorption of new bone (remodeling) concomitant with OCP degradation when implanted intramedullary in a rabbit femur for 12 weeks, compared to sintered HA ceramic. Histological and histomorphometric analyses using undecalcified specimens showed that the area of bone apposition was significantly higher on OCP than on HA between 2 and 3 weeks, whereas it subsequently became smaller on OCP than on HA. The area attacked by multinucleated giant cells, including tartrate-resistant acid phosphatase (TRAP)-positive cells, was significantly higher for OCP than for HA at 8 weeks. Radiography revealed resorption of OCP but not of HA. The results disclose some osteoconductive characteristics of synthetic OCP in the bone marrow space: (1) enhancement of bone regeneration at the initial bone apposition stage and (2) stimulation of resorption of the newly formed bone coupled with OCP biodegradation mediated by TRAP-positive osteoclast-like cells. These results suggest that synthetic OCP would be a more useful bone substitute than HA in implant applications where rapid bone formation and concomitant implant resorption are important considerations.     

Differential growth factor control of bone formation through osteoprogenitor differentiation

The osteogenic factors bone morphogenetic protein (BMP-7), platelet-derived growth factor (PDGF)-BB, and fibroblast growth factor (FGF-2) regulate the recruitment of osteoprogenitor cells and their proliferation and differentiation into mature osteoblasts. However, their mechanisms of action on osteoprogenitor cell growth, differentiation, and bone mineralization remain unclear. Here, we tested the hypothesis that these osteogenic agents were capable of regulating osteoblast differentiation and bone formation in vitro. Normal human bone marrow stromal (HBMS) cells were treated with BMP-7 (40 ng ml(-1)), PDGF-BB (20 ng ml(-1)), FGF-2 (20 ng ml(-1)), or FGF-2 plus BMP-7 for 28 days in a serum-containing medium with 10 mM beta-glycerophosphate and 50 microg ml(-1) ascorbic acid. BMP-7 stimulated a morphological change to cuboidal-shaped cells, increased alkaline phosphatase (ALKP) activity, bone sialoprotein (BSP) gene expression, and alizarin red S positive nodule formation. Hydroxyapatite (HA) crystal deposition in the nodules was demonstrated by Fourier transform infrared (FTIR) spectroscopy only in BMP-7- and dexamethasone (DEX)-treated cells. DEX-treated cells appeared elongated and fibroblast-like compared to BMP-7-treated cells. FGF-2 did not stimulate ALKP, and cell morphology was dystrophic. PDGF-BB had little or no effect on ALKP activity and biomineralization. Alizarin Red S staining of cells and calcium assay indicated that BMP-7, DEX, and FGF-2 enhanced calcium mineral deposition, but FTIR spectroscopic analysis demonstrated no formation of HA similar to human bone in control, PDGF-BB-, and FGF-2-treated samples. Thus, FGF-2 stimulated amorphous octacalcium phosphate mineral deposition that failed to mature into HA. Interestingly, FGF-2 abrogated BMP-7-induced ALKP activity and HA formation. Results demonstrate that BMP-7 was competent as a sole factor in the differentiation of human bone marrow stromal cells to bone-forming osteoblasts confirmed by FTIR examination of mineralized matrix. Other growth factors, PDGF, and FGF-2 were incompetent as sole factors, and FGF-2 inhibited BMP-7-stimulated osteoblast differentiation.     

Liver X receptors orchestrate osteoblast/osteoclast crosstalk and counteract pathologic bone loss

Osteoporosis is characterized by enhanced differentiation of bone‐resorbing osteoclasts, resulting in a rapid loss of functional trabecular bone. Bone‐forming osteoblasts and osteoblast‐derived osteocytes perform a key role in the regulation of osteoclast development by providing both the pro‐osteoclastogenic cytokine receptor activator of NF‐κB ligand (RANKL) and its natural decoy receptor osteoprotegerin (OPG). By regulating the RANKL/OPG ratio, osteoblasts hence determine the rate of both osteoclast differentiation and bone turnover. Here, we describe a novel role for liver X receptors (LXRs) during the crosstalk of bone‐forming osteoblasts and bone‐resorbing osteoclasts. By using a system of osteoblast/osteoclast cocultures, we identify LXRs as regulator of RANKL expression and the RANKL/OPG ratio in osteoblasts. Activation of LXRs drastically reduced the RANKL/OPG ratio and interfered with osteoblast‐mediated osteoclast differentiation in vitro. During an ovariectomy (OVX)‐induced model of postmenopausal osteoporosis, the application of an LXR agonist shifted the RANKL/OPG ratio in vivo, ameliorated the enhanced osteoclast differentiation, and provided complete protection from OVX‐induced bone loss. These results reveal an unexpected involvement of LXRs in the regulation of bone turnover and highlight a potential role for LXRs as novel targets in the treatment of osteoporosis and related diseases. © 2012 American Society for Bone and Mineral Research.     

负载BMP-2掺锶磷酸钙复合材料对成骨细胞增殖及功能的影响

目的探索负载BMP-2掺锶磷酸钙复合材料对成骨细胞增殖及功能的影响。方法获取SD大鼠成骨细胞,随机分为空白对照组(Con组)、磷酸钙组(CPC组)和复合材料组(BSCPC组);培养基中分别添加安慰剂、磷酸钙和负载BMP-2掺锶磷酸钙共培养一段时间,通过CCK-8法检测成骨细胞的增殖情况,碱性磷酸酶(alkalinephosphatase,ALP)及茜素红染色观察细胞的功能状态,蛋白电泳观察骨保护素(osteoprotegerin,OPG)、核因子κB受体活化因子(receptor activator of nuclear factor-KB,RANK)及核因子κB受体活化因子配体(ligand of receptor activator of nuclear factor-κB,RANKL)蛋白的表达情况。结果共培养1、3、5和7 d,和Con组比较,CPC组和BSCPC组的成骨细胞数目明显增加(P0.05),且以BSCPC组成骨细胞数目最多(P0.05);共培养14 d及21 d,和Con组比较,CPC组和BSCPC组的成骨细胞的矿化能力及ALP活性明显增加(P0.05),且以BSCPC组细胞钙化能力最强及ALP活性最高(P0.05);共培养7 d,和Con组比较,CPC组和BSCPC组的成骨细胞的OPG表达明显增加,而RANK及RANKL蛋白表达明显降低(P0.05),且以BSCPC组的成骨细胞蛋白OPG、RANK及RANKL蛋白表达量改善最为显著(P0.05)。结论负载BMP-2掺锶磷酸钙复合材料促进成骨细胞增殖分化和改善细胞活性和功能。     

Proton concentrations can be a major contributor to the modification of osteoclast and osteoblast differentiation,working independently of extracellular bicarbonate ions

We established a system to separately analyze the role of protons and bicarbonate ions in vitro in which the pH of the medium was controlled by HEPES at various concentrations of sodium bicarbonate (NaHCO₃) in the absence of carbon dioxide (CO₂). Using this system, we demonstrated that acidosis promoted osteoclast formation independently of extracellular NaHCO₃ in a short-term culture. Protons and bicarbonate ions acted on osteoclast differentiation with opposite effects, the former positively and the latter negatively. The HEPES-based system maintained pH in the absence of extracellular NaHCO₃ without CO₂. Therefore, we could demonstrate that osteoblast differentiation was promoted at higher pH in a long-term culture system without NaHCO₃ in which ALP activity and nodule mineralization were enhanced. This finding indicates that protons negatively control osteoblast differentiation independently of extracellular bicarbonate ions. However, the difference in the concentration of NaHCO₃ did not have any influence on nodule mineralization. The opposite effects of protons, the promotion of osteoclast formation and the inhibition of osteoblast differentiation, were suppressed in the presence of 5 mM N-acetyl cysteine, a reagent activating the scavenging of reactive oxygen species (ROS), implying that ROS act on both systems, the promotion of large osteoclast formation and the deterioration of osteoblast formation under acidosis.     

Microtubule assembly affects bone mass by regulating both osteoblast and osteoclast functions: Stathmin deficiency produces an osteopenic phenotype in mice

Cytoskeleton microtubules regulate various cell signaling pathways that are involved in bone cell function. We recently reported that inhibition of microtubule assembly by microtubule‐targeting drugs stimulates osteoblast differentiation and bone formation. To further elucidate the role of microtubules in bone homeostasis, we characterized the skeletal phenotype of mice null for stathmin, an endogenous protein that inhibits microtubule assembly. In vivo micro–computed tomography (µCT) and histology revealed that stathmin deficiency results in a significant reduction of bone mass in adult mice concurrent with decreased osteoblast and increased osteoclast numbers in bone tissues. Phenotypic analyses of primary calvarial cells and bone marrow cells showed that stathmin deficiency inhibited osteoblast differentiation and induced osteoclast formation. In vitro overexpression studies showed that increased stathmin levels enhanced osteogenic differentiation of preosteoblast MC3T3‐E1 cells and mouse bone marrow–derived cells and attenuated osteoclast formation from osteoclast precursor Raw264.7 cells and bone marrow cells. Results of immunofluorescent studies indicated that overexpression of stathmin disrupted radial microtubule filaments, whereas deficiency of stathmin stabilized the microtubule network structure in these bone cells. In addition, microtubule‐targeting drugs that inhibit microtubule assembly and induce osteoblast differentiation lost these effects in the absence of stathmin. Collectively, these results suggest that stathmin, which alters microtubule dynamics, plays an essential role in maintenance of postnatal bone mass by regulating both osteoblast and osteoclast functions in bone. © 2011 American Society for Bone and Mineral Research     

Unique roles of phosphorus in endochondral bone formation and osteocyte maturation

The mechanisms by which inorganic phosphate (P_i) homeostasis controls bone biology are poorly understood. Here we used Dmp1 null mice, a hypophosphatemic rickets/osteomalacia model, combined with a metatarsal organ culture and an application of neutralizing fibroblast growth factor 23 (FGF‐23) antibodies to gain insight into the roles of P_i in bone biology. We showed (1) that abnormal bone remodeling in Dmp1 null mice is due to reduced osteoclast number, which is secondary to a reduced ratio of RANKL/OPG expressed by osteoclast supporting cells and (2) that osteoblast extracellular matrix mineralization, growth plate maturation, secondary ossification center formation, and osteoblast differentiation are phosphate‐dependent. Finally, a working hypothesis is proposed to explain how phosphate and DMP1 control osteocyte maturation. © 2011 American Society for Bone and Mineral Research.     

Emodin Regulates Bone Remodeling by Inhibiting Osteoclastogenesis and Stimulating Osteoblast Formation

Bone remodeling, a physiological process in which new bone is formed by osteoblasts and the preexisting bone matrix is resorbed by osteoclasts, is vital for the maintenance of healthy bone tissue in adult humans. Imbalances in this process can cause various pathological conditions, including osteoporosis. Emodin, a naturally occurring anthraquinone derivative found in Asian herbal medicines, has numerous beneficial pharmacologic effects, including anticancer and antidiabetic activities. However, the effect of emodin on the regulation of osteoblast and osteoclast activity has not yet been investigated. We show here that emodin is a potential target for osteoporosis therapeutics, as treatment with this agent enhances osteoblast differentiation and bone growth and suppresses osteoclast differentiation and bone resorption. In this study, emodin suppressed receptor activator of nuclear factor‐κB (NF‐κB) ligand (RANKL)‐induced osteoclast differentiation of bone marrow macrophages (BMMs) and the bone‐resorbing activity of mature osteoclasts by inhibiting RANKL‐induced NF‐κB, c‐Fos, and NFATc1 expression. Emodin also increased ALP, Alizarin Red‐mineralization activity, and the expression of osteoblastogenic gene markers, such as Runx2, osteocalcin (OCN), and ALP in mouse calvarial primary osteoblasts, as well as activated the p38‐Runx2 pathway, which enhanced osteoblast differentiation. Moreover, mice treated with emodin showed marked attenuation of lipopolysaccharide (LPS)‐induced bone erosion and increased bone‐forming activity in a mouse calvarial bone formation model based on micro‐computed tomography and histologic analysis of femurs. Our findings reveal a novel function for emodin in bone remodeling, and highlight its potential for use as a therapeutic agent in the treatment of osteoporosis that promotes bone anabolic activity and inhibits osteoclast differentiation. © 2014 American Society for Bone and Mineral Research.     

Mst2 Controls Bone Homeostasis by Regulating Osteoclast and Osteoblast Differentiation

Mammalian sterile 20‐like kinase 2 (Mst2) plays a central role in the Hippo pathway, controlling cell proliferation, differentiation, and apoptosis during development. However, the roles of Mst2 in osteoclast and osteoblast development are largely unknown. Here, we demonstrate that mice deficient in Mst2 exhibit osteoporotic phenotypes with increased numbers of osteoclasts and decreased numbers of osteoblasts as shown by micro–computed tomography (µCT) and histomorphometric analyses. Osteoclast precursors lacking Mst2 exhibit increased osteoclastogenesis and Nfatc1, Acp5, and Oscar expression in response to receptor activator of NF‐κB ligand (RANKL) exposure. Conversely, Mst2 overexpression in osteoclast precursors leads to the inhibition of RANKL‐induced osteoclast differentiation. Osteoblast precursors deficient in Mst2 exhibit attenuated osteoblast differentiation and function by downregulating the expression of Runx2, Alpl, Ibsp, and Bglap. Conversely, ectopic expression of Mst2 in osteoblast precursors increases osteoblastogenesis. Finally, we demonstrate that the NF‐κB pathway is activated by Mst2 deficiency during osteoclast and osteoblast development. Our findings suggest that Mst2 is involved in bone homeostasis, functioning as a reciprocal regulator of osteoclast and osteoblast differentiation through the NF‐κB pathway. © 2015 American Society for Bone and Mineral Research.     

Prednisolone induces osteoporosis-like phenotype in regenerating zebrafish scales

Summary

We demonstrate that glucocorticoids induce an osteoporotic phenotype in regenerating scales of zebrafish. Exposure to prednisolone results in altered mineral content, enhanced matrix breakdown, and an osteoporotic gene-expression profile in osteoblasts and osteoclasts. This highlights that the zebrafish scale provides a powerful tool for preclinical osteoporosis research.

Introduction

This study aims to evaluate whether glucocorticoid (prednisolone) treatment of zebrafish induces an osteoporotic phenotype in regenerating scales. Scales, a readily accessible dermal bone tissue, may provide a tool to study direct osteogenesis and its disturbance by glucocorticoids.

Methods

In adult zebrafish, treated with 25 μM prednisolone phosphate via the water, scales were removed and allowed to regenerate. During regeneration scale morphology and the molar calcium/phosphorus ratio in scales were assessed and osteoblast and osteoclast activities were monitored by time profiling of cell-specific genes; mineralization was visualized by Von Kossa staining, osteoclast activity by tartrate-resistant acid phosphatase histochemistry.

Results

Prednisolone (compared to controls) enhances osteoclast activity and matrix resorption and slows down the build up of the calcium/phosphorus molar ratio indicative of altered crystal maturation. Prednisolone treatment further impedes regeneration through a shift in the time profiles of osteoblast and osteoclast genes that commensurates with an osteoporosis-like imbalance in bone formation.

Conclusions

A glucocorticoid-induced osteoporosis phenotype as seen in mammals was induced in regenerating scalar bone of zebrafish treated with prednisolone. An unsurpassed convenience and low cost then make the zebrafish scale a superior model for preclinical studies in osteoporosis research.     

恒定磁场对骨组织中细胞影响的研究

目的阐明恒定磁场对成骨细胞和破骨细胞的生长和功能的影响和对骨髓基质细胞向成骨细胞方向分化的作用，从而在细胞水平较全面的解释磁场治疗骨质疏松的机理。方法利用体外原代培养的成骨细胞、破骨细胞和骨髓基质细胞，分别外加0．38T、0．48T恒定磁场处理后，观察恒定磁场对这三种细胞生长、分化和功能的影响。结果恒定磁场处理可促进骨髓基质细胞向成骨细胞方向分化，促进成骨细胞的增殖、分化及功能的表达，并且抑制破骨细胞的生长、分化和功能。结论恒定磁场促进骨组织的成骨作用，抑制骨分解，是其治疗骨质疏松良好效果的部分机制。     

Roles of macrophage-colony stimulating factor and osteoclast differentiation factor in osteoclastogenesis

We have isolated osteoclast precursors (OCPs) from cocultures of mouse calvarial cells and bone marrow cells without adding any osteotropic factors. OCPs expressed Mac-1, Mac-2, and Gr-1 antigens but not osteoclast markers such as tartrate-resistant acid phosphatase (TRAP) and calcitonin receptors, and they differentiated into TRAP-positive cells within 48 h on a fixed calvarial cell layer pretreated with osteotropic factors such as 1α,25-dihydroxyvitamin D₃. In the present study, we investigated the regulatory mechanisms of OCP formation from hemopoietic cells and TRAP-positive cell formation from OCPs. Calvarial osteoblasts obtained from macrophage-colony stimulating factor (M-CSF) -deficient op/op mice failed to support OCP formation or the differentiation of OCPs into TRAP-positive cells. Both OCP formation and TRAP-positive cell formation supported by osteoblasts were completely inhibited by osteoclastogenesis inhibitory factor (OCIF, also called OPG), which is a decoy receptor of osteoclast differentiation factor (ODF; also called TRANCE, RANKL, and OPGL). When bone marrow cells were cultured for 4 days with soluble ODF (sODF/sRANKL) together with M-CSF, OCPs were formed even in the absence of osteoblasts. When OCPs were treated with sODF/sRANKL and M-CSF in the absence of osteoblasts, they differentiated into TRAP-positive cells within 48 h even in the presence of hydroxyurea. Northern blotting analysis revealed that osteoblasts constitutively expressed a certain level of ODF/RANKL mRNA. These results indicated that M-CSF and sODF/sRANKL produced by osteoblasts are two essential factors for both OCP formation and TRAP-positive osteoclast formation. Received: July 17, 1999 / Accepted: Nov. 12, 1999     

Reduced bone formation and relatively increased bone resorption in absorptive hypercalciuria

Absorptive hypercalciuria (AH), a common stone-forming condition characterized biochemically by intestinal hyperabsorption of calcium and hypercalciuria may be associated with bone loss. In AH type I (AH-1), hypercalciuria persists despite restriction in dietary calcium intake. We therefore hypothesized that the skeleton may contribute to the hypercalciuria in this subgroup of patients. Histomorphometric analysis of iliac crest biopsies were performed on nine stone-formers with AH-1 and on nine matched normal subjects. After stabilization on a stone-prevention diet, calcium homeostasis in the stone formers was then evaluated on inpatient constant metabolic diet before and after short-term blockade of bone resorption by alendronate (10 mg daily, 17 days total). Compared with controls, the stone-formers had lower indices of bone formation (osteoblast surface/bone surface 1.8+/-2.1 vs 3.0+/-1.5%, P=0.04; wall thickness 35.8+/-6.9 vs 47.2+/-7.6%, P=0.001) and relatively higher bone resorption (osteoclast surface/bone surface 0.4+/-0.2 vs 0.2+/-0.2%, P=0.05). In the stone-formers, a short-term course of alendronate treatment corrected fasting urinary calcium (0.14+/-0.06 to 0.06+/-0.04 mg Ca/mg Cr, P=0.001) and marginally reduced 24-h urinary calcium by 48 mg/day (P=0.06). Increased intestinal calcium absorption and hypercalciuria persisted, but estimated calcium balance improved (P=0.007). Our results suggest that the hypercalciuria of AH-1 originates primarily from intestinal hyperabsorption of calcium, but bone resorption in excess of bone formation may contribute.     

阿仑膦酸修饰的水凝胶负载柚皮素骨靶向系统构建及其增强成骨分化作用

目的 中药黄酮类单体存在水溶性较差、生物利用度低等特点，低效率全身给药的方式限制其在治疗骨疾患的应用，本研究旨在改善黄酮类单体治疗骨质疏松局限现状。方法 本研究制备了阿仑膦酸修饰的水凝胶负载中药骨碎补活性黄酮单体柚皮素的凝胶载药系统，通过活体荧光技术验证其骨靶向性能，并进一步采用PCR、WB等技术探究了该凝胶载药系统的促进成骨分化效用。结果 制备的凝胶载药系统促进成骨且稳定性好，释药部位集中于大鼠骨组织，干预成骨细胞MC3T3-E1细胞后，其成骨因子及相关蛋白表达明显增强。结论 阿仑膦酸修饰的水凝负载柚皮素可实现骨靶向大鼠骨组织，且能有效促进成骨分化，从而可潜在实现治疗骨质疏松。