氟化物对骨形成中成骨细胞的影响及机制

氟化物作为促骨形成药物之一 ,最早被推荐应用于骨质疏松症的临床治疗 ,目前仍是唯一可供临床使用的有效的促骨形成的药物。近年来由于成骨细胞 (osteoblast OB)培养技术及基因工程技术的发展 ,对氟化物刺激骨形成作用有了更进一步的认识 ,有助于我们对氟中毒时氟骨症机制的探讨。1　氟化物对成骨细胞的直接作用　　氟是一种已知可影响骨形成的非激素因子。具有双相调节作用。长期小剂量氟可促进骨形成 ;大剂量可引起骨质疏松或骨硬化。在人体和动物实验研究中 ,较多学者报道氟中毒时骨量增多是由于 OB数增多 ,活性增加 ,生命周期延长 [1 …     

前列腺素E2促进老年大鼠成骨细胞骨形成及其机制研究

目的　探讨前列腺素E2 (PGE2 )对老年雄性大鼠胫骨上段松质骨的作用。　方法　取2 0月龄Wistar雄性大鼠 ,分别皮下注射PGE2 (3mg·kg 1·d 1) 10d和 30d。用体内双荧光标记法、不脱钙组织切片 ,以骨组织形态计量学方法观察胫骨上段松质骨骨小梁和骨髓腔的动态和静态参数变化。　结果　PGE2 作用 10d骨小梁表面成骨细胞周长OB/BS〔(12 3± 7 6 ) %〕和类骨质周长OS/BS〔(2 0 4± 7 2 ) %〕明显高于对照组〔(1 6± 0 7) %和 (4 3± 1 7) % ,P <0 0 1〕 ,骨小梁表面和髓腔出现多层排列的前成骨细胞OPC ,形成多细胞成分而钙化不全的新生骨小梁WB ;PGE2 作用 30d矿化骨形成率BFR/BV〔(815 4± 137 9) %·年 1〕和骨量BV/TV〔(4 2 1± 12 6 ) %〕明显高于对照组〔(15 4 9± 14 6 5 ) %·年 1和 (13 5± 3 2 ) % ,P <0 0 1〕 ,骨髓脂肪组织面积F/TV由 (18 2± 5 6 ) %减少到 (11 4± 3 6 ) % (P <0 0 5 )。　结论　PGE2 在短期内有刺激老年大鼠松质骨成骨细胞骨形成 ,增加骨量的作用。     

Osterix：与成骨细胞分化和骨形成有关的转录因子

Osterix(Osx)是一种新发现的与成骨细胞分化和骨形成有关的转录因子，只在发育的骨组织中特异性表达。Osx基因剔除的Osx(-／-)小鼠完全丧失骨形成能力。Osx(-／-)小鼠／胚胎间叶细胞仍能正常表达Runx2，Osx可能位于成骨细胞分化路径中Runx2的下游，前成骨细胞分化为成熟的成骨细胞需要Osx的存在。同时Osx可能是转录因子Sox9和软骨细胞的一种负向调控子，可阻止骨／软骨祖细胞向软骨细胞的分化。     

雌二醇对成骨细胞护骨素基因表达的影响

目的 研究在MG－63细胞和成人成骨细胞培养的不同阶段,雌激素对护骨素（OPG）基因mRNA表达的影响,探讨雌激素对成骨细胞的作用机制。方法 在细胞培养的不同时间,测定细胞内碱性磷酸酶（ALP）活性、骨钙素（OC）含量和进行Van Gieson胶原染色,以确定细胞培养的增殖、成熟、矿化,于成熟阶段用17β－雌二醇（E2）对培养细胞进行处理,用半定量RT－PCR技术测定雌二醇对成骨细胞OPG基因mRNA表达的影响。结果 ALP活性、骨钙素含量及胶原染色均表明,细胞培养汇片后12天达到成熟。正常成人髂骨分离的成骨细胞（HOB）及成骨肉瘤细胞株MG－63均能表达OPG,且表达量与培养的时程有关。培养12天时,两各OPG基因mRNA表达均达最高峰（P＜0．01）。10^-8mol/L的E2能使MG－63细胞OPG基因mRNA的表达至基础值的10倍（P＜0．001）,HOB的OPG基因mRNA表达增加至基础值的7－8倍（P＜0．01）。结论 OPG在成骨细胞表达。雌激素使成骨细胞OPG基因mRNA表达的增加是雌激素缺乏导致骨质疏松的重要机制之一。     

辛伐他汀对成骨细胞成骨能力的影响

目的　研究辛伐他汀促进成骨作用的机制及量效关系。方法　取成人的髂骨松质骨 ,采用胶原 胰蛋白酶消化 ,获得松质骨中的成骨细胞 ,进行纯化和培养。在此基础上添加不同浓度的辛伐他汀 ( 1× 1 0 - 1 1 、1× 1 0 - 9、1× 1 0 - 7、1× 1 0 - 6、1× 1 0 - 5、5× 1 0 - 5 mol/L)并进行培养。用生化法测定细胞碱性磷酸酶 (ALP)活性 ,用放射免疫法测定细胞培养液中骨钙素 (OC)和细胞间质含钙量。结果　辛伐他汀对成骨细胞ALP活性和骨钙素分泌的影响均呈正相关 ,即各浓度辛伐他汀对ALP活性、骨钙素的分泌和成骨细胞的成骨能力均有刺激作用 ,并与辛伐他汀剂量呈正相关。结论　辛伐他汀能增加成骨细胞的ALP活性和骨钙素的产生 ,提高成骨细胞的成骨能力。     

氟对成骨细胞的影响

氟是一种已知可影响骨形成的非激素因子,对骨形成具有双相调节作用;长期小剂量氟可促进骨形成,大剂量可引起骨质疏松或骨硬化。成骨细胞是骨发生和骨形成的重要细胞,具有合成、分泌组成骨基质的胶原和糖蛋白的作用,并通过钙化基质形成骨组织,另外成骨细胞在维持机体内环境的稳定、生理机制调节和骨代谢性疾病中发挥重要作用。     

Osterix:与成骨细胞分化和骨形成有关的转录因子

Osterix(Osx)是一种新发现的与成骨细胞分化和骨形成有关的转录因子,只在发育的骨组织中特异性表达。Osx基因剔除的Osx(-/-)小鼠完全丧失骨形成能力。Osx(-/-)小鼠/胚胎间叶细胞仍能正常表达Runx2,Osx可能位于成骨细胞分化路径中Runx2的下游,前成骨细胞分化为成熟的成骨细胞需要Osx的存在。同时Osx可能是转录因子Sox9和软骨细胞的一种负向调控子,可阻止骨/软骨祖细胞向软骨细胞的分化。     

成骨细胞与破骨细胞的相互作用对骨重塑的调节

骨骼是一个动态活性组织,它通过持续的重塑来维持其矿化平衡及自身的结构完整。在骨重塑的过程中,协调成骨细胞,骨细胞和破骨细胞之间的活性,能保持骨重塑过程的动态耦联平衡,其中成骨细胞（骨形成功能）和破骨细胞（骨吸收功能）在骨重塑过程中起关键作用。成骨细胞和破骨细胞之间的相互调节在骨重塑过程实现骨形成和骨吸收平衡的基础。两组细胞实现细胞间相互作用主要有三种方式：直接接触,分泌旁分泌因子及细胞与骨基质作用,成骨细胞和破骨细胞之间3种相互作用方式对骨重塑过程起重要调节作用。     

结缔组织生长因子与骨形成蛋白-9在骨代谢中的研究进展

<正>结缔组织生长因子(CTGF)是一种细胞外基质分泌蛋白,生理状态下,在心、脑、肝、肾等组织器官中呈基础量表达,病理状态下,如在某些增生性或纤维性疾病中,CTGF过度表达与疾病的发生发展紧密相关。CTGF调节细胞功能多样化,参与体内多种病理生理过程,如胚胎发育、雌性生殖系统、恶性肿瘤、纤维化疾病等。CTGF是骨形态发生蛋白(BMPs)信号通路的靶基因,可作用于BMPs信号通路〔1〕,而BMP-9是已知BMPs中     

1,25(OH)2D3对体外培养成骨细胞骨保护素mRNA表达的影响

维生素Ｄ体内活性形式 1,2 5 (ＯＨ) 2 Ｄ3具有促进骨吸收及骨形成作用。其骨吸收作用通过成骨细胞介导 ,然而机制尚未完全清楚。骨保护素 (ｏｓｔｅｏｐｒｏｔｅｇｅｒｉｎ ,ＯＰＧ)是近年分离出的一种因子 ,由成骨细胞合成和分泌 ,在破骨细胞的分化形成中起信号传导作用〔1〕。我们于 2 0 0 0年 6～ 12月实验观察 1,2 5 (ＯＨ) 2 Ｄ3对体外培养乳鼠成骨细胞ＯＰＧ基因表达的影响 ,以探讨其对骨吸收功能的调节机制。　　一、材料与方法　　 1.细胞培养 :取新生 2 4ｈＳＤ大鼠颅骨 ,Ⅰ型胶原酶分阶段消化收集细胞 ,5 %ＣＯ2 、3 7℃培养 ,每…     

Effects of BMP-2 on osteoblastic cells and on skeletal growth and bone formation in unloaded rats.

A previous study showed that skeletal unloading induced by hindlimb suspension for 14 days in rats reduces osteoblastic cell proliferation, inhibits skeletal growth and bone formation and induces metaphyseal bone loss. This study investigated the effect of recombinant human bone morphogenetic protein-2 (rhBMP-2) in this model. In vitro analysis showed that rhBMP-2 (25-100 ng/ml, 48-96 h) increased alkaline phosphatase activity, an early marker of osteoblast differentiation, in rat neonatal calvaria cells and adult marrow stromal cells, showing that rhBMP-2 induced the differentiation of osteoblast precursor cells in vitro. In contrast, rhBMP-2 did not increase rat calvaria or marrow stromal cell proliferation. Biochemical and histomorphometric analysis showed that systemic infusion with rhBMP-2 (2 microg/kg/day) in unloaded rats had no significant effect on serum osteocalcin levels and on histomorphometric indices of bone formation. Accordingly, rhBMP-2 infusion did not prevent the decreased skeletal growth, trabecular bone bone volume and bone mineral content induced by unloading. The present data indicate that, although rhBMP-2 stimulates osteoblastic cell differentiation, rhBMP-2 infusion is not effective in increasing bone formation and in preventing trabecular bone loss induced by unloading in rats.     

葡萄糖对大鼠骨髓基质干细胞向成骨细胞分化的影响

目的 体外观察不同葡萄糖浓度在促骨髓基质干细胞向成骨细胞分化过程中对细胞因子碱性磷酸酶、骨形成蛋白-2、转化生长因子β1表达的影响,并探讨其对骨代谢的作用机制.方法 无菌条件下从2周龄SD大鼠长骨骨髓中分离获取骨髓基质干细胞,采用全骨髓贴壁培养法对骨髓基质干细胞进行纯化、传代扩增,随后在不同糖浓度(5.5 mmol/L、25.0 mmol/L)干预下向成骨细胞诱导分化培养21 d,进行茜素红染色观察矿化结节并测定成骨细胞的标记物碱性磷酸酶、骨形成蛋白-2及转化生长因子β1表达,比较各组中成骨细胞分化的情况.组间比较采用单因素方差分析法.结果 25.0 mmol/L糖浓度组与5.5 mmol/L糖浓度组比较,钙结节形成比例分别为(45.3±0.7)%、(68.3±0.8)%,差异具有统计学意义(P＜0.05),碱性磷酸酶(0.350±0.020、0.563±0.043)、骨形成蛋白-2[(590±27)、(744±41)μg/L]及转化生长因子β1[(875±40)、(1188±52)μg/L]活性比较,差异具有统计学意义(均P＜0.05).结论 在高糖条件下,骨髓基质干细胞向成骨细胞的分化减弱,这可能是糖尿病性骨质疏松的重要机制之一.     

Diabetes causes decreased osteoclastogenesis, reduced bone formation, and enhanced apoptosis of osteoblastic cells in bacteria stimulated bone loss

The most common cause of inflammatory bone loss is periodontal disease. After bacterial insult, inflammation induces bone resorption, which is followed by new reparative bone formation. Because diabetics have a higher incidence and more severe periodontitis, we examined mechanisms by which diabetes alters the response of bone to bacterial challenge. This was accomplished with db/db mice, which naturally develop type 2 diabetes. After inoculation of bacteria osteoclastogenesis and bone resorption was measured. Both parameters were decreased in the diabetic group. Diabetes also suppressed reparative bone formation measured histologically and by the expression of osteocalcin. The impact of diabetes on new bone formation coincided with the effect of diabetes on apoptosis of bone-lining cells. Within 5 d of bacterial challenge, apoptosis declined in the wild-type animals yet remained significantly higher in the diabetic group. Thus, diabetes may cause a net loss of bone because the suppression of bone formation is greater than the suppression of bone resorption. The uncoupling of bone formation and resorption may be due in part to prolonged apoptosis of bone lining cells.     

高血糖状态对骨形成和骨吸收作用及骨密度的影响

目的 探讨不同血糖水平对骨形成、骨吸收和骨密度的影响.方法 对糖尿病组(DM)148例,葡萄糖耐量低减组(IGT)30例,空腹血糖受损组(IFG)30例,健康对照组(Control)50例,分别测定血浆葡萄糖(BG)、糖化血红蛋白(HbA1c)、血清骨钙素(BGP)、尿脱氧吡啶啉(DPD)/肌酐(Cr)、腰椎(L2、L3、L4)和髋部(股骨颈、Ward三角、大转子)骨密度(BMD),并对上述指标进行T检验和方差分析.结果 四组间方差分析显示BMD在Ward三角、L4差异有统计学意义(P<0.05).血清BGP水平在四组呈递增趋势,尿DPD/Cr水平在四组呈递减趋势,方差分析显示尿DPD/Cr水平差异有显著统计学意义(P<0.001).尿DPD/Cr水平随HbA1c水平递增逐渐增高.结论 糖尿病的前期阶段(IGT),骨吸收已呈现高于同龄正常人水平,糖尿病患者骨形成低于同龄正常人而骨吸收高于同龄正常人,骨平衡呈现负平衡,骨量逐渐丢失;IGT开始骨密度已较正常人低,良好的血糖控制对减少骨吸收和延缓骨量下降具有保护作用.应在糖调节受损的早期阶段对骨质疏松进行干预治疗.     

Effects of tumor necrosis factor on bone formation in vitro

Tumor necrosis factor (TNF) was studied for its effects on bone formation in cultured rat calvariae. TNF alpha at 100-100,000 U/ml stimulated [3H]thymidine incorporation into DNA, an effect that appeared after 24 h of treatment and lasted 96 h. Transient (24-h) treatment with TNF alpha increased [3H]proline incorporation into type I collagen 24-72 h after the factor was removed; this effect was DNA synthesis dependent and blocked by hydroxyurea. Transient treatment with TNF alpha also increased alkaline phosphatase activity. In contrast, continuous treatment with TNF alpha for 48-96 h caused a marked inhibition on [3H]proline incorporation into type I collagen and alkaline phosphatase activity. TNF alpha caused a small increase in collagen degradation. Lymphotoxin had similar effects to those of TNF alpha. In conclusion, TNF alpha stimulates calvarial DNA synthesis which causes an increased number of collagen-synthesizing cells, but TNF alpha has a direct inhibitory effect on osteoblastic function.     

Effects of etidronate-mediated suppression of bone remodeling on aluminum-induced de novo bone formation.

The mechanism by which aluminum chloride stimulates de novo bone formation is unknown. To evaluate the role of bone remodeling and mature osteoblastic function in aluminum-induced neoosteogenesis, we compared the osteogenic effects of aluminum in normal beagles to those in animals with low turnover osteomalacia induced by treatment with etidronate [1-hydroxyethane-1,1-diphosphoric acid (HEBP)]. As assessed by quantitative bone histomorphology, beagles treated with HEBP developed low turnover osteomalacia characterized by a 78% reduction in osteoblast number, a 5.5-fold increase in osteoid volume, complete absence of active mineralization, and diminished resorption surfaces compared to untreated controls. The iv administration of aluminum chloride to normal dogs generated new trabecular structures in the marrow cavity consistent with induction of de novo bone formation. This response consisted of increased trabecular bone volume and number, accumulation of woven osteoid, and increased number of bone-forming cells. The concomitant administration of HEBP failed to prevent induction of de novo bone formation by aluminum. Instead, the neoosteogenic process was superimposed on low turnover osteomalacia in HEBP-treated dogs. Serum aluminum concentrations were increased 2-fold, whereas bone aluminum accumulation was reduced by 58% in HEBP- and aluminum-treated dogs compared to that in aluminum-treated controls. These findings indicate that aluminum stimulation of neoosteogenesis in beagles is independent of mature osteoblast function, normal bone remodeling, and total bone aluminum accumulation. Rather, aluminum-induced de novo bone formation appears to result from stimulation of mesenchymal precursors to form a primitive type of bone which is distinct from coupled bone formation.     

Runx2 is a target of mechanical unloading to alter osteoblastic activity and bone formation in vivo

Molecular mechanisms underlying unloading-induced reduction of bone formation have not yet been fully understood. In vitro, Runx2 has been suggested to be involved in mechanical signaling in osteoblasts. However, the roles of Runx2 in vivo during the bone response to mechanical stimuli have not yet been known. The purpose of this paper was to examine the roles of Runx2 in unloading-induced bone loss in vivo. Tail suspension was conducted for 2 wk using 9- to 11-wk-old Runx2 heterozygous knockout mice (Runx2(+/-)) and wild-type (Wt) littermates. Bones were subjected to two-dimensional micro-x-ray computed tomography, bone histomorphometry and RT-PCR analyses. Loss of half Runx2 gene dosage-exacerbated unloading-induced bone loss in trabecular and cortical envelopes. Unloading-induced reduction in mineral apposition rate and bone formation rate in cortical bone as well as trabecular bone was exacerbated in Runx2(+/-) mice, compared with Wt mice. Bone resorption parameters were not significantly affected by unloading or Runx2(+/-) genotype. Basal Runx2 and osterix mRNA levels in bone were reduced by 50% in Wt, whereas unloading in Runx2(+/-) mice did not further alter Runx2 and osterix mRNA levels. In contrast, osteocalcin mRNA levels were reduced by unloading, regardless of Runx2 gene dosage. These data demonstrated that full Runx2 gene dosage is required for maintaining normal function of osteoblasts in mechanical unloading or nonphysiological condition. Finally, we propose Runx2 as a critical target gene in unloading to alter osteoblastic activity and bone formation in vivo.     

Effects of hypoxia on granulocytic-monocytic progenitors in rats. Role of bone marrow stroma

Hemorrhagic shock leads to hypoxia and is associated with bone marrow (BM) failure. Hemorrhagic shock is also a predisposing factor in immune dysregulation. Since the BM is the major organ of immune cells in the adult, its failure following hemorrhagic shock may explain the increased susceptibility to infection. The in vitro evidence indicates that hypoxia mediates altered functions in BM stroma. Since similar hematopoietic alterations are reported in hypoxia and hemorrhagic shock, hypoxia alone could be a representative model to study BM responses during hemorrhagic shock. In this study, we use an animal model to dissect the hematopoietic effects of hypoxia. We subjected rats to hypoxia, and at days 1 and 5 post-hypoxia we determined the numbers of granulocytic-monocytic progenitors (CFU-GM) in the BM. We found significant increase (P < 0.05) in CFU-GM at day 1 and a downward trend by day 5. Enhanced BM cellularity could not explain the increase in CFU-GM by day 1. BM stromal cells mediated most of the stimulatory effects by hypoxia. CFU-GM was inversely proportional to bioactive TGF-beta and directly proportional to IL-1. Compared to normoxic rats, IL-6 production was suppressed in BM cells from hypoxic rats. The results show that hypoxia alone initiate a stimulatory response in CFU-GM progenitors. These effects are at least partially mediated through the BM stroma. In the absence of a second insult, CFU-GM reverts to baseline. The data also suggest that hypoxia mediates complex responses that include cytokine production. These results add to the current understanding of hematopoietic responses by hypoxia and adds to the mechanisms of immune dysfunctions following hemorrhagic shock.     

Effects of transforming growth factor-beta on osteoblastic osteosarcoma cells

Transforming growth factor-beta (TGF beta), a polypeptide that controls growth and differentiation in many cell types and has recently been found in abundant amounts in bone, was examined for its effects on cells with the osteoblast phenotype using the clonal osteoblastic osteosarcoma cell line ROS 17/2.8. TGF beta increased alkaline phosphatase (AP) activity and the rate of collagen synthesis per cell. Cell proliferation was inhibited, and the morphological appearance of the cells was markedly changed. All effects were observed at concentrations as low as 0.1 ng/ml TGF beta. Increases in AP activity were detectable after 24 h and increased progressively with time. TGF beta increased AP activity under serum-free conditions and during thymidine-induced inhibition of DNA synthesis. The increase in AP activity mediated by TGF beta could be completely inhibited with actinomycin D and cycloheximide. 1,25-Dihydroxyvitamin D3 at 10(-7) M slightly increased AP activity in ROS 17/2.8 cells, but strongly inhibited AP activity when the cells were pretreated with TGF beta. The data suggest that TGF beta stimulates expression of the osteoblastic phenotype in ROS 17/2.8 cells and that TGF beta may be an important regulator of local bone remodeling.