从细胞水平研究龟鹿二仙含药血清在治疗骨质疏松中的作用

从细胞水平研究龟鹿二仙含药血清在治疗骨质疏松中的作用.(1)用酶消化法获得新生大鼠的成骨细胞.碱性磷酸酶(ALP)染色进行鉴定,通过对成骨细胞增殖、胰岛素样生长因子(IGF-1)分泌的检测来考察成骨细胞功能.(2)用1,25(OH)2D3诱导骨髓单核细胞获得破骨细胞,抗酒石酸酸性磷酸酶(TRAP)染色鉴定破骨细胞.电镜扫描观察破骨细胞形成的骨凹陷情况;同时检测TRAP( )细胞数和破骨细胞TRAP活性.(3)用血清药理学方法制备龟鹿二仙含药血清,分别加入不同剂量组10%含药血清进行干预,观察该中药血清对上述成骨细胞和破骨细胞功能指标的影响.龟鹿二仙高剂量组血清可促进成骨细胞增殖,增加IGF-1的分泌量.而中剂量组血清可明显抑制骨髓细胞向破骨细胞的转化,抑制骨凹陷形成,降低细胞内TRAP的活性.本研究表明龟鹿二仙具有显著的抗骨质疏松作用.     

GM-CSF和其诱导的破骨细胞对成骨细胞的影响

目的：探讨GM-CSF和其诱导的破骨细胞对成骨细胞生长的影响。方法：采用分组对照、细胞计数等方法进行研究。结果：GM-CSF和其诱导的破骨细胞可以促进成骨细胞生长。结论：GM-CSF和其诱导的破骨细胞促进成骨细胞增殖、分化，可能是通过破骨细胞内GM-CSF介导的信号通路而发挥作用。     

破骨细胞在骨组织工程中的意义及其研究策略

破骨细胞是骨吸收细胞,在骨塑造和重塑中发挥重要作用。目前在破骨细胞对成骨细胞调节方面的影响了解很少。最近的研究表明在骨发育过程中破骨细胞的缺乏会导致骨基质排列紊乱、矿化减少、成骨细胞行为异常。破骨细胞在骨组织工程的引入将会解决骨组织工程面临的许多问题。破骨细胞前体在组织工程化骨上生成破骨细胞是一条生理、实际的破骨细胞引入途径。     

白细胞介素33介导的骨免疫北大核心

背景:骨免疫学是近年来骨代谢疾病机制研究的热点。白细胞介素33是骨免疫学中的一个重要的新型细胞因子,在类风湿关节炎、骨质疏松等多种骨代谢疾病进展中发挥重要作用,其传导通路可能成为临床治疗的潜在靶点。目的:综述白细胞介素33在骨代谢过程中的骨免疫调节作用及其研究进展。方法:检索PubMed、Web of Science、中国知网及万方数据库从建库至2022年6月收录的白细胞介素33和骨代谢及骨免疫学研究相关文献,最终纳入67篇文献进行总结。结果与结论:①白细胞介素33介导骨免疫对骨代谢具有调节作用。②白细胞介素33对骨代谢的调节可通过降低硬化蛋白的表达,进而激活成骨细胞Wnt/β-catenin通路,该途径通过多种机制增加骨量,包括干细胞更新、刺激成骨细胞前复制、诱导成骨细胞生成、抑制成骨细胞和骨细胞凋亡。白细胞介素33可通过刺激Bcl6、MafB、Irf-8等抗破骨细胞基因的表达,下调破骨细胞活性T细胞核因子c1的表达,进而抑制破骨细胞的形成,并且可诱导凋亡分子的表达,促进破骨细胞凋亡。③白细胞介素33可下调2型天然淋巴细胞RANKL的表达,促进白细胞介素13、白细胞介素4和粒细胞-巨噬细胞集落刺激因子的产生来抑制破骨细胞的分化。白细胞介素33可诱导调节性T细胞向炎症部位募集,通过抑制Th1、CD8+T细胞和M1巨噬细胞,以支持成骨前体细胞的分化,抑制破骨细胞共刺激分子的表达,导致破骨细胞的分化和功能受到抑制。白细胞介素33可激活幼稚T细胞向Th2型表型分化,产生Th2型细胞因子,可诱导破骨细胞在局部的募集,造成局部骨吸收增加,软骨退化。④白细胞介素33可诱导内皮细胞增殖、迁移和形态分化,促进血管形成,对缺血性疾病有一定的保护作用。⑤白细胞介素33对骨代谢的调节具有双向性,造成骨代谢紊乱的病因多样,并且体内不同部位微环境又不相同,控制白细胞介素33的调节作用是治疗骨代谢相关疾病的一个新的方向。     

PTH对破骨细胞骨吸收功能的影响及成骨细胞介导作用

采用分离、培养兔破骨细胞和成骨细胞的方法，体外研究甲状旁腺激素（ＰＴＨ）对破骨细胞骨吸收功能的影响，以及成骨细胞和破骨细胞之间的相互作用。结果表明，ＰＴＨ对破骨细胞的骨吸收功能无直接影响，但在成骨细胞参与下，ＰＴＨ对破骨细胞性骨吸收有明显的促进作用。说明成骨细胞在ＰＴＨ调节破骨细胞功能活动中有着重要的介导作用。     

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.     

骨细胞的力学感受器

骨细胞是骨骼中最丰富和寿命最长的细胞,是骨重建的调节器。骨细胞在内分泌调节和钙磷酸盐代谢中发挥重要作用,也是力学刺激的主要响应者,感知力学刺激以直接或间接的方式对刺激做出反应。骨细胞中的力学转导是一个复杂而精细的调节过程,涉及细胞与其周围环境、相邻细胞以及细胞内部不同功能的力学感受器之间的相互作用。目前已知的骨细胞主要力学感受器包括初级纤毛、piezo离子通道、整合素、细胞外基质以及基于连接蛋白的细胞间连接。这些力学感受器在骨细胞中发挥着至关重要的作用,它们能够感知并转导力学信号,进而调节骨稳态。本文对5种力学感受器进行系统的介绍,以期为理解骨细胞如何响应力学刺激和维持骨组织稳态提供新的视角和认识。     

成骨细胞参与白细胞介素—1调节破骨细胞功能的实验研究

建立了成骨细胞和破骨细胞共同培养体系，探讨了白细胞介素－１（ＩＬ１）促进骨吸收的作用机理。研究发现，破骨细胞＋ＩＬ１组的骨吸收陷窝数目和面积，与单纯破骨细胞组比较，差异无显著性意义。破骨细胞＋成骨细胞＋ＩＬ１组，骨吸收陷窝数目和面积均显著增加，与破骨细胞＋ＩＬ１组及单纯破骨细胞对照组比较，差异均有显著性意义。提示ＩＬ１对破骨细胞缺乏直接作用，而是在成骨细胞介导下，发挥调节破骨细胞的骨吸收作用。     

免疫活性细胞对骨代谢的调控作用

1 骨免疫学的概念 骨形成和骨吸收之间的平衡调节着骨内环境稳定，这包括有成骨细胞（Osteoblasts，OB）和破骨细胞（Osteoclasts，OC）间相互的协调作用。OB是骨形成细胞，可分泌骨基质分子；而来源于造血前体细胞的OC则吸收骨基质。     

Recent progress in studies on osteocytes--osteocytes and mechanical stress

Although osteocytes are of the most abundant cells in bone, our knowledge about the role of osteocytes in bone metabolism is still poor compared with that about osteoblasts and osteoclasts, both being on the surface of bone. Osteocytes are terminally differentiated bone-forming cells. During bone formation, some of the osteoblasts lining the surface of bone are incorporated into the newly formed osteoid matrix and become osteocytes, while the other osteoblasts remain on the surface as lining cells. During this transition from osteoblasts to osteocytes, the cells lose numerous osteoblastic phenotypes and acquire osteocytic characteristics such as high expression of osteocalcin and particularly their specific morphology. Osteocytes are connected with each other in bone and with osteoblasts on the bone surface through canaliculi, forming cellular networks; and gap-junctions present at the contact sites mediate their intercellular communication. Several roles of osteocytes in bone have been proposed so far. Of them, based on the morphological characteristics of osteocytes, sensation of mechanical stress loaded onto bone is suspected to be one of their functions. One of the mechanical stresses on bone is fluid shear stress. Between the osteocyte's plasma membrane and the bone matrix is the periosteocytic space. This space exists both in the lacunae and in the canaliculi, and it is filled with extracellular fluid (ECF). Application of mechanical stress to bone locally deforms the tissue. This periodical deformation subsequently causes an increase in the flow of ECF in the periosteocytic space, resulting in shear stress on the surface of the osteocytes. Experimental studies demonstrated that bone cells were equivalently or more sensitive to the fluid shear stress than epithelial cells. Osteocytic cells cultured enhanced expression of prostaglandin (PG) G/H synthase-2 (COX-2) mRNA in response to shear stress. PGE2 is a potent regulator of proliferation and function of osteoblasts and osteoclasts. Therefore, a metabolic response by osteoblasts and osteoclasts lining the bone surface may be caused by PGE2 produced by osteocytes in response to shear stress when the prostanoid reaches the surface through the canaliculi. In conclusion, osteocytes play an important role in sensing extracellular mechanical stress, and the mechanical signals mediated by osteocytes may regulate the overall metabolism of cells in bone tissue.     

Some osteocytes released from their lacunae are embedded again in the bone and not engulfed by osteoclasts during bone remodeling

It is generally accepted that osteocytes derive from osteoblasts that have secreted the bone around themselves. Osteocytes are cells embedded in the lacunae in the bone, and they are characteristically in contact with other cells by many slender cytoplasmic processes in canaliculi. During bone remodeling, many osteocytes in the bone are released from their lacunae by osteoclasts; however it remains unclear what happens to these released osteocytes. The cortical bone of the rat mandibular body was used in this study. Mandibles were fixed, decalcified, and then embedded in Epon 812. Specimens were sectioned in the frontal direction into serial 0.5 microm-thick semithin or 0.1 microm-thick ultrathin sections, and then examined by light or transmission electron microscopy. Cells that fitted in the osteocytic lacunae with canaliculi extending to the bone were identified as osteocytes in this study. Among many osteocytes released by osteoclasts in cutting cones, there were osteocytes half-released from their lacunae. These cells fitted in their lacunae with canaliculi extending to the bone and showed developed cell organelles in the cytoplasm. In closing cones, many osteocytes were situated in the bone away from cement lines; however, there were half-embedded osteocytes in the bone formed on cement lines. These cells fitted in their lacunae with canaliculi extending to the bone formed below cement lines and showed developed cell organelles in the cytoplasm. These results show that half-embedded osteocytes in closing cones derive from half-released osteocytes in cutting cones. Osteocytes encircled by osteoclasts were sometimes observed on one section, but serial sections showed that these osteocytes fitted in their remaining lacunae in the bone on other sections. This shows that not all osteocytes released from their lacunae are engulfed by osteoclasts. Consequently, the present results suggests that some osteocytes released from their lacunae are embedded again in the bone and not engulfed by osteoclasts during bone remodeling.     

Interaction of dichloromethylene diphosphonate and vitamin D on bone of thyroparathyroidectomized rats.

Dichloromethylene diphosphonate (Cl2MDP) antagonized the action of vitamin D on bone in thyroparathyroidectomized rats by reducing the metabolic activity of osteoblasts and osteocytes and decreasing the number of osteoclasts. Ultrastructurally, osteoblasts in Cl2MDP-treated rats were interpreted to be less active in bone matrix synthesis. Osteocytes in Cl2MDP-treated rats were interpreted ultrastructurally to be inactive; there was no evidence of bone resorption when compared to osteocytes in rats given vitamin D alone. Abnormal osmiophilic densities in the pericellular bone matrix of rats given vitamin D alone were not present in rats given vitamin D and Cl2MDP. The ultrastructure of osteoclasts was unaltered by Cl2MDT. These cellular changes were associated with a decrease in serum calcium and increase in bone ash and magnesium concentration in rats given high levels (10 mg/kg) of Cl2MDP. Bone adenosine triphosphatase and alkaline phosphatase activities were not affected by Cl2MDP. These results suggest that Cl2MDP may limit the hypercalcemia of hypervitaminosis D by directly inhibiting bone cells in addition to its physicochemical action.     

Histological identification of osteocytes in the allegedly acellular bone of the sea breams Acanthopagrus australis,Pagrus auratus and Rhabdosargus sarba (Sparidae,Perciformes, Teleostei)

The bone of advanced teleost fishes such as those of the family Sparidae is said to lack osteocytes or to be acellular. Acellularity has been determined by apparent lack of osteocyte lacunae. This study questions the validity of this criterion. Scanning electron and light microscopy of paraffin and resin sections were used to show that the sides of sea bream mandibles consist of laminar parallel-fibred bone that we call tubular bone, because it contains tubules, and localised regions of Sharpey fibre bone. Osteocytes lie along the walls of tubules that also contain collagen fibril bundles (T-fibres), or in the lumens of tubules that do not contain T-fibres. We show that the osteocytes are derived from osteoblasts. The T-fibre system is different from other fibre systems that have been described. The tubules enclose wide T-fibres (lenticular in cross-section, maximum width about 8 μm) that taper at their ends and continue as thin T-fibres (round in crosssection, about 2 μm wide). The T-fibres originate in the periosteum. In mature tubular bone, spaces of increasing size develop around the osteocytes. Osteocytes are released from the bone matrix and become postosteocytes or bone-lining cells. Secondary bone lines the largest spaces. In Sharpey fibre bone, small osteocytes in small lacunae (about 2 μm wide) are found in columns parallel to the Sharpey fibres. Large osteocytes are found in large round spaces and are much larger than comparable osteocytes in lacunae in the bone of the salmon Salmo salar. We conclude that an absence of visible or conventional osteocyte lacunae does not mean that the cells themselves are absent. There are cells and two types of collagen fibre bundle in the tubules. The cells are osteocytes derived from osteoblasts, and these osteocytes apparently resorb bone with the result that large amounts of bone are destroyed. “Acellular” tubular and Sharpey fibre bone are types of cellular bone that differ from each other and from conventional cellular bone.     

The ultrastructural effect of estrogens on bone cells in thyroparathyroidectomized rats.

To determine the direct effect of estrogen on bone cells, female rats were thyroparathyroidectomized and given either 200 microng of estradiol cypionate or placebo treatment for 4 to 30 days. After 8 days, an osteosclerosis of the tibial metaphysis developed in rats treated with estrogens. Osteoblasts in estrogen-treated rats were interpreted ultrastructurally to be less active in bone formation than controls. Osteocytes in estrogen-treated rats were inactive compared to osteocytes in control rats which were continuing the process of bone formation. Osteoclasts were decreased in numbers in rats treated with estrogen; however, the electron microscopic characteristics of osteoclasts did not differ from controls. These cellular changes were associated with a severe hypocalcemia in both estrogen-treated and control rats. Serum phosphorus and urinary hydroxyproline were lowered significantly by the administration of estrogen. It is concluded that metaphyseal osteosclerosis develops in estrogen-treated rats independent of parathyroid hormone and calcitonin. The osteoclerosis is due to an effect of estrogens to reduce osteoclast-numbers and a lesser inhibitory effect on the bone-forming activity of osteoblasts and osteocytes. (Am J Pathol 87:311-322, 1977).     

Osteocyte: the impresario in the electrical stimulation for bone fracture healing

Delayed healing and nonunions of bone fracture are critical problems in orthopedic surgery. Electrical stimulation has been used as a therapeutic method for enhancing bone healing for a long time. Despite unanimous clinical success, the underlying mechanism concerning bone tissue in response to electrical stimulation remains poorly understood. In the meantime, emerging evidences suggest that osteocytes, with their unique location and morphologies, play an important role in regulating the behaviors of other bone cells, including osteoblasts, osteoclasts and their progenitor cells. In this paper, we hypothesize that osteocytes are the sensory cells for the electrical stimulation, and they orchestrate the whole process of new bone formation and remodeling in the electrotherapy for bone fracture. The postulated electrosensory transduction pathway might be a coupling effect of osteoblasts and osteoclasts, which is regulated by the biochemical signals expressed from osteocytes after sensing the membrane potential changes. It is believed that better understanding of this mechanism would facilitate optimizing the electrotherapy for bone disorders and assist in solving these clinical problems.     

In vivo leptin expression in cartilage and bone cells of growing rats and adult humans

The present investigation was carried out to analyse, immunohistochemically, in vivo leptin expression in cartilage and bone cells, the latter restricted to the elements of the osteogenic system (stromal cells, osteoblasts, osteocytes, bone lining cells). Observations were performed on the first lumbar vertebra, tibia and femur of four rats and on the humerus, femur and acromion of four patients. Histological sections of paraffin-embedded bone samples were immunostained using antibody to leptin. The results showed that, in growing rat bone, leptin is expressed in chondrocytes and stromal cells, but not in osteoblasts; bone lining cells were not found in the microscopic fields examined. In adult human bone, leptin is expressed in chondrocytes, stromal cells and bone lining cells; osteoblasts were not found in the microscopic fields examined. Osteocytes were found to be leptin positive only occasionally and focally in both rat and human bone. The in vivo findings reported show, for the first time, that leptin appears to be expressed only in the cells of the osteogenic lineage (stromal cells, bone lining cells, osteocytes) that, with respect to osteoblasts, are permanent and inactive, i.e. in those cells that according to our terminology constitute the bone basic cellular system (BBCS). Because the BBCS seems to be primarily involved in sensing and integrating mechanical strains and biochemical factors and then in triggering and driving bone formation and/or bone resorption, it appears that leptin seems to be mainly involved in modulating the initial phases of bone modelling and remodelling processes.     

Histological identification of osteocytes in the allegedly acellular bone of the sea breams Acanthopagrus australis, Pagrus auratus and Rhabdosargus sarba (Sparidae, Perciformes, Teleostei)

The bone of advanced teleost fishes such as those of the family Sparidae is said to lack osteocytes or to be acellular. Acellularity has been determined by apparent lack of osteocyte lacunae. This study questions the validity of this criterion. Scanning electron and light microscopy of paraffin and resin sections were used to show that the sides of sea bream mandibles consist of laminar parallel-fibred bone that we call tubular bone, because it contains tubules, and localised regions of Sharpey fibre bone. Osteocytes lie along the walls of tubules that also contain collagen fibril bundles (T-fibres), or in the lumens of tubules that do not contain T-fibres. We show that the osteocytes are derived from osteoblasts. The T-fibre system is different from other fibre systems that have been described. The tubules enclose wide T-fibres (lenticular in cross-section, maximum width about 8 m) that taper at their ends and continue as thin T-fibres (round in crosssection, about 2 m wide). The T-fibres originate in the periosteum. In mature tubular bone, spaces of increasing size develop around the osteocytes. Osteocytes are released from the bone matrix and become postosteocytes or bone-lining cells. Secondary bone lines the largest spaces. In Sharpey fibre bone, small osteocytes in small lacunae (about 2 m wide) are found in columns parallel to the Sharpey fibres. Large osteocytes are found in large round spaces and are much larger than comparable osteocytes in lacunae in the bone of the salmon Salmo salar. We conclude that an absence of visible or conventional osteocyte lacunae does not mean that the cells themselves are absent. There are cells and two types of collagen fibre bundle in the tubules. The cells are osteocytes derived from osteoblasts, and these osteocytes apparently resorb bone with the result that large amounts of bone are destroyed. Acellular tubular and Sharpey fibre bone are types of cellular bone that differ from each other and from conventional cellular bone.     

Ultrastructure of bone-tissue cells

The ultrastructure of bone cells is delineated by investigation of bone tissue in young mice. The characteristic cytoplasmicstic pattern of osteoblasts, osteocytes and osteoclasts is described. The morphologic findings are discussed with respect to the function of these cells.     

The osteopetrotic rabbit: Skeletal cytology and ultrastructure

The lethal, autosomal recessive osteopetrotic mutation in the rabbit, osteosclerosis (os/os), has recently been made available for experimental investigation. We have examined the cytology and ultrastructure of skeletal cells in mutants and report abnormalities in osteoblasts, osteocytes, and osteoclasts. Mutant osteoclasts lack a well-defined ruffled border and show few morphological signs of bone resorption. Osteoblasts in mutants form bone in neonatal life but show signs of degeneration by 2 weeks after birth. Mutant osteoblasts and osteocytes contain large, electron-dense cytoplasmic inclusions. External surfaces of mutant long bones show no evidence of bone resorption by scanning electron microscopy, and fibrosis of intertrabecular spaces is a prominent feature in mutants. These data, considered with recent evidence that the functions of osteoblasts and osteoclasts are interrelated, suggest that reduced bone resorption, a characteristic feature of osteopetrosis, may be related to osteoblast incompetence in this mutation.