力学载荷对破骨细胞凋亡的影响及其调节机制

破骨细胞是参与骨代谢的基本功能细胞之一.破骨细胞在骨重建过程中主要承担旧骨组织的破坏和吸收,因此,破骨细胞凋亡的微小变化都可能会改变骨重建的进程.调节破骨细胞凋亡的因素有很多,如雌激素、二磷酸盐等生物化学因素,但力学载荷对于破骨细胞生物学活性影响的研究相对较少.综述了力学载荷对破骨细胞生物学活性的影响以及细胞凋亡与破骨细胞凋亡的调节.     

影响破骨细胞功能的主要因素

骨是一种不断重塑的活性组织。骨重塑是由骨吸收和新骨形成两个过程协调完成。对骨吸收起主要作用的是多核—破骨细胞。破骨细胞是一种多核巨细胞，每个细胞具有2—100个核不等，直径可达100μm，位于哈佛系统内的骨内膜表面和骨膜下表面。尽管许多调节破骨细胞生成及骨吸收的因素仍     

护骨素的研究进展

护骨素是近年发现的肿瘤坏死因子家族的新成员具有犯制破骨细胞分化及骨吸收活性的一种分泌型糖蛋白。护骨素与其护骨素配基之间竞争性结合机制是调控破骨细胞分化、增殖、凋亡及发挥重作用过程中最重要的信号通路,是雌激素调节破骨细胞生成和抗骨吸收的作用途径之一。对其研究将促进因破骨细胞活性增强导致骨吸收、骨破坏为特点的疾病的病因、预防和治疗等研究的深入。     

破骨细胞的生物学新观

骨是一种不断进行重建的有活力组织。骨重建 (remodeling)是一个偶联过程 ,骨吸收紧随新骨形成。负责骨吸收的细胞主要是多核破骨细胞 ,关于调节破骨细胞的形成及溶骨作用的因子还有许多问题未找到答案 ,但最近在理解破骨细胞的细胞生物学和分子生物学及骨髓微环境在调节破骨细胞的形成及溶骨作用方面已获得了重大进展。1　破骨细胞形态学破骨细胞 osteoclast)是一种大的多核巨细胞 ,含 2～ 1 0 0个核 ,通常为 1 0～ 2 0个 ,直径可达 1 0 0 μm,数量极少 ,通常仅 2～ 3个 /μm3,但在骨转换活跃的部位 ,如成长中骨的干骺端 ,其数量增加。破骨…     

破骨细胞骨吸收中肿瘤坏死因子α可提高空泡型质子泵表达与活性

背景:空泡型的ATP酶(V-ATPase)在破骨细胞的骨吸收功能中起重要作用,肿瘤坏死因子α对破骨细胞中的V-ATPase表达与活性的影响尚不明确。目的:通过V-ATPase的表达与酶的活性变化,探讨肿瘤坏死因子α促进破骨细胞骨吸收功能的机制。方法:体外诱导培养破骨细胞,分别给予不同质量浓度的肿瘤坏死因子α干预(5,10,30μg/L)。采用荧光定量PCR及Western Blot检测肿瘤坏死因子α对破骨细胞V-ATPase表达的影响;根据吸光度值计算出V-ATPase的相对活性;倒置显微镜下观察骨吸收陷窝形成情况,采用Image J软件分析骨吸收面积。结果与结论:破骨细胞经过48 h的肿瘤坏死因子α干预以后,V-ATPase的表达与活性均有显著的增加,并且提高肿瘤坏死因子α的干预浓度均可增强此效应。破骨细胞经肿瘤坏死因子α干预后,培养板的骨吸收面积明显增加,这种作用随着肿瘤坏死因子α干预浓度的增加而增强。由此推测肿瘤坏死因子α作为一个重要的炎症递质参与病理性骨质吸收的过程,除促进破骨细胞形成以外,其可能的机制是肿瘤坏死因子α通过增加V-ATPase的表达,并提高V-ATPase活性,从而增强破骨细胞的骨质吸收活动。     

破骨细胞融合分子机制的研究进展

破骨细胞是体内介导骨吸收的主要细胞。破骨细胞融合过程包括:识别、半融合结构及融合孔形成,最后融合孔扩张、细胞内容物混合。破骨细胞融合与破骨细胞的大小及其骨吸收能力密切相关,对破骨细胞融合分子机制的研究可为骨吸收亢进性疾病的治疗寻求新的突破口。本文就国内外破骨细胞融合过程及其分子机制的最新研究进展进行综述。     

白细胞介素1β对破骨细胞空泡型质子泵活性与胞内酸化的影响

目的:通过体外实验研究白细胞介素1β(IL-1β)对破骨细胞空泡型质子泵与细胞内pH值的影响,以探讨IL-1β促进破骨细胞骨吸收的机制。方法:骨髓单核巨噬细胞经过诱导成为破骨细胞后,予以0.1、0.3和0.5μg/L的IL-1β干预48 h,检测破骨细胞空泡型质子泵mRNA和蛋白的表达以及IL-1β对破骨细胞胞内酸化的影响,观察空泡型质子泵活性的改变与破骨细胞骨吸收功能的变化。结果:研究结果提示破骨细胞经IL-1β干预48 h后,空泡型质子泵的表达与活性均显著增加,并且细胞内pH值下降。破骨细胞与含有IL-1β的细胞培养液共同孵育后,其骨吸收能力增强,培养液中IL-1β的浓度越高,该作用越趋明显。结论:IL-1β可能是通过上调空泡型质子泵的表达与酶的活性,促进氢离子的产生和(或)转运,引发破骨细胞骨吸收活动增加,最终导致骨质破坏,由此参与骨关节炎症性疾病的发生。     

破骨细胞骨吸收的分子机制

破骨细胞性骨吸收是在骨的微环境内进行的复杂分子生物学反应过程,涉及到众多蛋白质和调控因子的参与。有关破骨细胞的活化,骨基质的吸收,骨吸收的调控等方面,现有的数据还不够充分。本文综述了金属基质蛋白酶(M atrix m eta lloprote inases,MM P s)在破骨细胞移行和骨基质吸收方面的重要作用,以及破骨细胞分化因子(R eceptor activator of NF-κB-ligand,RANKL)和护骨素(O steoprotegerin,OPG)在骨吸收调控网络中的地位。     

实验性家兔骨折愈合过程中破骨细胞的形态学测量

为了探讨中成药生骨再造散对骨折愈合中骨吸收与破骨细胞形态学特征的影响及二者的关系,将30只青紫蓝家兔建成双侧桡骨3mm骨缺损的骨折模型,随机分为生骨再造散组(A组)、仙灵骨葆组(B组)及对照组(C组),每组各10只,于术后14d和31d每组各处死动物5只取材,进行骨吸收与破骨细胞形态学计量检测和相关性分析。结果:术后A组骨吸收较对照组强,破骨细胞形态学也发生了一定变化,差异有统计学意义(P<0.05);B组对破骨细胞形态也有一定影响,但对骨吸收影响不大。相关分析表明:骨吸收主要与破骨细胞数目相关,其次还与破骨细胞形状因子、面积、光密度等相关。实验结果提示,生骨再造散在术后14d和31d均可促进骨吸收,可能是通过早期提高破骨细胞数量,后期则增强其功能实现的,对骨折愈合有一定促进作用。     

应用破骨细胞-颅骨联合培养体系研究先天性成骨不全的骨再建过程

目的　采用先天性成骨不全(OI)小鼠,oim/oim为动物模型,应用破骨细胞-颅骨联合培养体系研究OB和OC两种细胞在OI骨再建过程中的功能改变和相互作用。 方法　实验采用小鼠颅骨（CAL）组织培养,实验设两组：WTCAL-WTOC组：联合培养对照组颅（WTCAL） 与对照破骨细胞（WTOC）;OICAL-OIOC组：联合培养OI颅骨（OICAL）与OI破骨细胞（OIOC）。以免疫组化染色方法　-TRAP识别破骨细胞,ALP免疫组化染色方法识别成骨细胞。破骨细胞骨吸收活性为骨吸收陷窝占颅骨表面百分比。单位OC吸收面积为总骨吸收陷窝除以破骨细胞数。 结果　于7d,OICAL-OIOC组破骨细胞数低于WTCAL-WTOC组;OICAL-OIOC组的OC/OB比例低WTCAL-WTOC组;OICAL-OIOC组单位破骨细胞吸收能力高于WTCAL-WTOC组。 结论　OI的小鼠模型骨再建中骨量丢失一方面由于其成骨细胞功能异常,另一方面也可能因为其破骨细胞的代偿性功能活跃。     

Receptor Activator for Nuclear Factor kappa B Ligand (RANKL) as an osteoimmune key regulator in bone physiology and pathology

The strength and integrity of the human skeleton depends on a delicate equilibrium between bone resorption and bone formation. Bone resorption is an elementary cellular activity in the modelling of the skeleton during growth and development. Later in life a most important physiological process in the skeleton is bone remodelling, which is locally initiated by resorption. During remodelling bone resorption is coupled to new bone formation that ensures renewal of bone with only minor local and temporary bone loss. Cells responsible for bone resorption and subsequent bone formation are the osteoclasts and osteoblasts, respectively. The osteoclast is derived from the pluripotent hematopoietic stem cell, which gives rise to a myeloid stem cell that can further differentiate into megakaryocytes, granulocytes, monocytes/macrophages and osteoclasts. The respective bone resorbing and forming actions of osteoclasts and osteoblasts are finely coupled, so that bone mass remains remarkably stable in a healthy adult. Imbalance between osteoclast and osteoblast activities can arise from a wide variety of hormonal changes or perturbations of inflammatory and growth factors resulting in postmenopausal osteoporosis, Paget's disease, lytic bone metastases, or rheumatoid arthritis, leading to increased bone resorption and crippling bone damage. In view of the critical role of osteoclasts in diverse pathology, there has been immense effort aimed at understanding the biology of this unique cell. The present review is focused on the current knowledge of the mechanisms that regulate the functional links between bone turnover and the immune system helping us to understand the main factors that lead to bone loss observed in osteoporosis, cancer and in rheumatoid arthritis. The aim of this review paper is to consider the key molecular interactions involved in the formation of osteoclast cells in normal and pathological conditions.     

Stimulation of osteoclast formation by inflammatory synovial fluid

Peri-articular bone resorption is a feature of arthritis due to crystal deposition and rheumatoid disease. Under these conditions, the synovial fluid contains numerous inflammatory cells that produce cytokines and growth factors which promote osteoclast formation. The aim of this study was to determine whether inflammatory synovial fluid stimulates the formation of osteoclasts. Synovial fluid from rheumatoid arthritis (RA), pyrophosphate arthropathy (PPA) and osteoarthritis (OA) patients was added to cultures (n=8) of human peripheral blood mononuclear cells (PBMCs) in the presence and absence of macrophage colony-stimulating factor (M-CSF) and the receptor activator of NF-κB ligand (RANKL). Osteoclast formation was assessed by the formation of cells positive for tartrate-resistant acid phosphatase (TRAP) and vitronectin receptor (VNR) and the extent of lacunar resorption. The addition of 10% OA, RA and PPA synovial fluid to PBMC cultures resulted in the formation of numerous multinucleated or mononuclear TRAP⁺ and VNR⁺ cells which were capable of lacunar resorption. In contrast to PBMC cultures incubated with OA synovial fluid, there was marked stimulation of osteoclast formation and resorption in cultures containing inflammatory RA and PPA synovial fluid which contained high levels of tumour necrosis factor alpha, a factor which is known to stimulate RANKL-induced osteoclast formation.     

Osteoclastogenesis and arthritis

There is emerging interest for osteoclasts as key players in the erosive and inflammatory events leading to joint destruction in chronic arthritis. In fact, chronic inflammatory joint diseases such as psoriatic arthritis and rheumatoid arthritis are often characterized by destruction of juxta-articular bone and erosions due to the elevated activity of osteoclasts, which are involved in bone resorption. The main step in inflammatory bone erosion is an imbalance between bone resorption and bone formation: osteoclast formation is enhanced by proinflammatory cytokines such as TNF-α, IL-1β, and IL-17 and is not balanced by increased activity of bone-forming osteoblasts. T-cells, stromal cells, and synoviocytes enhance osteoclast formation via expression of RANKL and, under pathologic conditions, of proinflammatory cytokines. In rheumatoid arthritis, accumulation of osteoclasts in synovial tissues and their activation associated with osteoclastogenic cytokines and chemokines at cartilage erosion sites suggest that they could be usefully selected as therapeutic target. In particular, in consideration of the primary role of RANKL and TNF-α in osteoclastogenesis, the control of the production of RANKL and the inhibition of TNF-α represent important strategies for reducing bone damage in this disease.     

IL-23 promotes osteoclast formation by up-regulation of receptor activator of NF-kappaB (RANK) expression in myeloid precursor cells

Inflammation-mediated bone loss is a major feature of various bone diseases including rheumatoid arthritis, osteoarthritis and advanced periodontitis. Enhanced osteoclast development or activity at the inflammation site results in bone resorption. IL-23 is a heterodimeric cytokine belonging to the IL-6/IL-12 family that has been implicated in the pathogenesis of rheumatoid arthritis and demonstrated to play a role in osteoclastogenesis via stimulation of IL-17 production. In this study we investigated whether IL-23 contributes to the regulation of osteoclast differentiation independent of the IL-17 pathway. We show that IL-23 dose-dependently up-regulates receptor activator of NF-kappaB expression in primary murine bone marrow macrophages and RAW264.7 cells and thereby promotes commitment of myeloid precursor cells to receptor activator of NF-kappaB ligand-mediated osteoclastic differentiation. However, IL-23 by itself is insufficient to induce osteoclastogenesis. Increased osteoclastic differentiation of cells was associated with enhanced cathepsin K expression and dentine resorption indicating enhanced formation of functional osteoclasts. IL-17 was not detectable in culture supernatants and when added to cultures, did not promote differentiation of RAW264.7 cells. These results demonstrate that IL-23 can act directly on myeloid precursor cells in addition to indirectly stimulating receptor activator of NF-kappaB ligand production in osteoblasts and explains its potency in driving osteoclast development in inflammation-mediated bone pathology.     

Rheumatoid arthritis synovial macrophage-osteoclast differentiation is osteoprotegerin ligand-dependent

Osteoprotegerin ligand (OPGL) is a newly discovered molecule which is essential for osteoclast differentiation. Both OPGL and its soluble decoy receptor, osteoprotegerin (OPG), which inhibits osteoclast formation, are known to be produced by osteoblasts and inflammatory cells found in the rheumatoid arthritis (RA) synovium. In this study, RA synovial macrophages were incubated in the presence or absence of OPGL, macrophage-colony stimulating factor (M-CSF), and dexamethasone for various time points. The results indicated that osteoclast formation from RA synovial macrophages is OPGL-dependent and that OPGL and M-CSF are the only humoral factors required for RA synovial macrophage-osteoclast differentiation. OPG was found to inhibit osteoclast formation by RA synovial macrophages in a dose-dependent manner. This study has shown that macrophages isolated from the synovium of RA patients are capable of differentiating into osteoclastic bone-resorbing cells; this process is OPGL- and M-CSF-dependent and is modulated by corticosteroids. Cellular (T and B cells, dendritic cells) and humoral factors in RA synovium and bone may influence osteoclast formation and bone resorption by controlling OPGL/OPG production.     

Review: Immune cells and mediators of inflammatory arthritis

Cytokine expression in the inflamed synovial membrane of patients with rheumatoid arthritis and other forms of chronic inflammatory arthritis and other forms of chronic inflammatory arthritis leads to formation of osteoclasts. These cells are primarily involved in the resorption of mineralized cartilage and bone and thus participate in joint damage in the course of chronic arthritides. Osteoclastogenesis in the synovial membrane is driven by cytokines such as RANKL, MCSF but also classical proinflammatory mediators such as TNF, IL-1 and IL-6. Inhibition of osteoclast formation has proven as an effective approach to inhibit structural damage in experimental arthritis and preliminary data suggest that such approaches are also effective in human RA.     

Review: Immune cells and mediators of inflammatory arthritis

Cytokine expression in the inflamed synovial membrane of patients with rheumatoid arthritis and other forms of chronic inflammatory arthritis and other forms of chronic inflammatory arthritis leads to formation of osteoclasts. These cells are primarily involved in the resorption of mineralized cartilage and bone and thus participate in joint damage in the course of chronic arthritides. Osteoclastogenesis in the synovial membrane is driven by cytokines such as RANKL, MCSF but also classical proinflammatory mediators such as TNF, IL-1 and IL-6. Inhibition of osteoclast formation has proven as an effective approach to inhibit structural damage in experimental arthritis and preliminary data suggest that such approaches are also effective in human RA.     

Repair of local bone erosions and reversal of systemic bone loss upon therapy with anti-tumor necrosis factor in combination with osteoprotegerin or parathyroid hormone in tumor necrosis factor-mediated arthritis

Local bone erosion and systemic bone loss are hallmarks of rheumatoid arthritis and cause progressive disability. Tumor necrosis factor (TNF) is a key mediator of arthritis and acts catabolically on bone by stimulating bone resorption and inhibiting bone formation. We hypothesized that the concerted action of anti-TNF, which reduces inflammation and parathyroid hormone (PTH), which stimulates bone formation, or osteoprotegerin (OPG), which blocks bone resorption and could lead to repair of local bone erosions and reversal of systemic bone loss. To test this, human TNF-transgenic mice with established erosive arthritis and systemic bone loss were treated with PTH, OPG, and anti-TNF, alone or in combination. Local bone erosions almost fully regressed, on combined treatment with anti-TNF and PTH and/or OPG, suggesting repair of inflammatory skeletal lesions. In contrast, OPG and anti-TNF alone led to arrest of bone erosions but did not achieve repair. Treatment with PTH alone had no influence on the progression of bone erosions. Local bone erosions all showed signs of new bone formation such as the presence of osteoblasts, osteoid formation, and mineralization. Furthermore, systemic bone loss was completely reversed on combined treatment and this effect was mediated by osteoblast stimulation and osteoclast blockade. In summary, we conclude that local joint destruction and systemic inflammatory bone loss because of TNF can regress and that repair requires a combined approach by reducing inflammation, blocking bone resorption, or stimulating bone formation.     

Synovial fluid macrophages are capable of osteoclast formation and resorption

To determine whether synovial fluid (SF) macrophages isolated from the SF of osteoarthritis (OA), rheumatoid arthritis (RA) and pyrophosphate arthropathy (PPA) joints are capable of osteoclast formation, and to investigate the cellular and humoral factors required for this to occur, SF macrophages (CD14+) were isolated from the knee joint SF from patients with OA, RA and PPA and cultured for up to 14 days with macrophage-colony stimulating factor (M-CSF) and soluble receptor activator for nuclear factor-kappaB ligand (RANKL) or tumour-necrosis factor-alpha (TNFalpha) and interleukin-1alpha (IL-1alpha). Osteoclast differentiation was assessed by expression of tartrate-resistant acid phosphatase (TRAP) and vitronectin receptor (VNR), F-actin ring formation and lacunar resorption. Osteoclast formation and lacunar resorption was seen in RANKL-treated cultures of SF macrophages isolated from OA, RA and PPA joints with the largest amount of resorption noted in RA and PPA SF macrophage cultures. In TNFalpha/IL-1alpha-treated RA and PPA SF macrophage cultures, osteoclasts capable of lacunar resorption were also formed. Lacunar resorption was more extensive in RANKL than TNFalpha/IL-1alpha-treated cultures. These findings indicate that SF macrophages are capable of differentiating into mature osteoclasts capable of lacunar resorption. M-CSF in combination with RANKL or TNFalpha/IL-1alpha was required for osteoclast formation. As inflammatory synovial fluids contain an increase in the number of macrophages and an increase in the amounts of RANKL, TNFalpha and IL-1alpha, these findings suggest that one means whereby bone erosions may form in rheumatoid or crystal arthritis is by differentiation of synovial fluid macrophages into osteoclasts.     

Roles of Wnt signals in bone resorption during physiological and pathological states

Osteoclasts, multinucleated giant cells, are responsible for bone resorption in physiological and pathological conditions such as osteoporosis and rheumatoid arthritis. Osteoclasts develop from the monocyte/macrophage lineage under the strict control of bone-forming osteoblasts. Osteoblast-lineage cells express two cytokines essential for osteoclast differentiation, colony-stimulating factor-1, and receptor activator of nuclear factor κB ligand (RANKL) and also express osteoprotegerin, a soluble decoy receptor for RANKL. The signaling molecule Wnt has been shown to be important for the differentiation of osteoblasts through β-catenin-dependent canonical and β-catenin-independent noncanonical pathways. Recent studies have established that Wnt-mediated signals are also crucial for bone resorption in both physiological and pathological conditions. In this review, we introduce recent advances in roles of Wnt signaling in bone formation and bone resorption.