软骨下矿化组织在骨性关节炎进展中的作用及其机制

[目的]综述软骨下矿化组织在骨性关节炎(osteoarthritis,OA)中的改变,探讨软骨下矿化组织在OA发病机制中的作用,展望可能的研究方向。[方法]广泛查阅近年来有关软骨下骨和钙化软骨在OA中病理变化的文献,并对其正常结构和功能、生物力学性能、微观结构的变化等总结分析。[结果]软骨下矿化组织的改变与OA的发生发展密切相关。[结论]进一步加强对软骨下矿化组织的了解和认识,将对OA的研究起到积极的作用。     

关节软骨钙化层相关研究进展

关节软骨钙化层(calcified cartilage zone, CCZ)是软骨组织中一层重要的界面结构,对骨软骨组织生理功能的发挥有着重要意义。相关基础研究及临床研究表明钙化层结构有利于骨关节炎的防治以及软骨组织的再生修复,本文对钙化层的理化性质,及在防治骨关节炎和软骨修复方面的研究进展作一综述。     

膝关节原发性骨关节炎软骨和软骨下骨病理改变的定量研究

目的观察膝关节原发性骨关节炎(osteoarthritis,OA)胫骨平台软骨和软骨下骨病理改变特点,对比内、外侧平台软骨和软骨下骨结构参数,探讨钙化层和软骨下骨在OA发病机制中的作用。方法取2009年10月-2011年5月行人工全膝关节置换术治疗的30例30膝原发性OA患者自愿捐赠的新鲜胫骨平台标本进行实验。其中男11例,女19例;年龄55～78岁,平均65.1岁。病程10～25年,平均16.6年;患膝内翻畸形1～23°,平均9.3°。大体观察胫骨平台后在内、外侧中央负重区取材,常规制备脱钙石蜡切片,行HE和番红O/固绿染色,观察关节软骨退变特点,参照Mankin评分标准评分并分期;观察钙化层及软骨下骨病理改变。应用Image Pro Plus 6.0图像分析软件测量软骨和软骨下骨结构参数,包括软骨全层(total articular cartilage,TAC)厚度、钙化层(articular calcified cartilage,ACC)厚度、ACC/TAC比值、软骨下骨板(subchondral bone plate,SCP)厚度以及骨小梁体积分数(trabecular bone volume,BV/TV)。结果大体观察内侧平台软骨退变较外侧严重,内侧平台软骨Mankin评分为(12.4±1.1)分,显著高于外侧平台的(8.3±1.6)分(t=12.173,P=0.000)。根据Mankin评分结果在60个标本中,14个为OA早期,可见软骨浅表层裂隙、潮线复制和软骨下骨增厚;19个为OA中期,可见软骨深层裂隙、多发软骨下骨吸收陷窝和明显增厚的软骨下骨;27个为OA晚期,可见软骨全层缺失、软骨内化骨和"象牙化"软骨下骨。软骨和软骨下骨结构参数测定示:内侧平台TAC厚度显著低于外侧平台,ACC/TAC比值、BV/TV及SCP厚度显著高于外侧平台,差异均有统计学意义(P<0.05)。内、外侧平台ACC厚度比较,差异无统计学意义(P>0.05)。结论钙化层和软骨下骨可能在OA发生与进展中发挥了重要作用。     

骨关节炎中关节软骨和周围骨的变化

骨关节炎(OA)是最常见的关节炎,同时也是造成中老年人伤残的重要原因。曾经一度认为关节软骨的改变是骨关节炎主要的病变,但现在普遍认为,所有关节结构包括软骨钙化区、软骨下骨、骨小梁、关节囊等都会受到影响。然而骨与软骨所起到的作用、之间的关系如何还不明确。本文就骨关节炎中关节软骨和周围骨的变化作一综述。     

软骨钙素

软骨钙素是存在于软骨基质中的一种钙结合蛋白。它主要分布在生长板软骨肥大细胞层和关节软骨的深层至潮线。其组成和结构与Ⅱ型胶原羧基端前肽相同。软骨钙素在软骨生理、病理钙化中发挥重要作用。     

正常膝关节软骨钙化层形态结构研究

目的探索关节软骨钙化层的结构形态及其与软骨非钙化层和软骨下骨之间的界面连接方式。方法组织库获取自愿捐献的人体正常股骨髁新鲜标本20个,男10个,女10个;年龄17～45岁。常规制备石蜡横、纵切片,番红O／固绿和冯库萨染色观察钙化层形态结构;扫描电镜观察软骨各层之间的界面连接方式;连续切片结合建模技术建立骨软骨三维模型。结果关节骨软骨复合组织番红O／固绿染色结果示软骨红染,软骨下骨蓝染,钙化层位于潮线与黏合线之间;冯库萨染色结果示钙化层黑染,结构及边界清晰,上界面以波浪状潮线结构与非钙化层紧密连接,下界面以凹凸不平的梳齿状结构与软骨下骨相互锚合：关节骨软骨剥离面及断面扫描电镜示钙化层与非钙化层以沟壑镶嵌方式相互嵌合;关节骨软骨复合组织纵切面观察,钙化层与软骨下骨间的黏合线呈凹凸不平的梳齿状结构;骨软骨三维模型观察结果与关节软骨自然剥离横断面扫描电镜观察结果基本一致。结论钙化层是关节软骨的重要结构,它通过特有界面连接方式将软骨牢牢固定在软骨下骨上。     

实验性骨关节炎中关节软骨钙化层厚度测定及其意义

作者借助髋关节骨关节炎（ＯＡ）实验动物模型、观察到：（１）关节软骨钙化层在负重区分布最厚，非负重区最薄。（２）ＯＡ严重度和钙化层厚度呈正相关，和软骨非钙化软骨厚度呈负相关。（３）ＯＡ病理发展过程中具有非钙化软骨层变薄，钙化软骨层增厚，潮线前移等特征。该结果为分析ＯＡ病因及病理机制提供直接依据。     

软骨下骨在骨关节炎进展中的作用

骨关节炎(OA)以关节软骨进行性退变和软骨下骨反应性增生为主要病理特征.关节软骨损害是OA最明显变化,OA研究长期以来主要集中在关节软骨,但软骨下骨作为与之毗邻的组织,在OA发病中的作用不容忽视,认清软骨下骨在OA中的变化和作用,有助于更全面和深刻地认识OA发病机制,为治疗提供新思路.该文就软骨下骨在OA中的病理变化及OA进展中的作用作一综述.     

生物力学在关节软骨修复中的作用

活动关节软骨是无血管的透明软骨,损伤后修复困难。传统的修复方式以手术为主,但修复的软骨组织常常无法满足透明软骨的结构条件。软骨组织工程是修复关节软骨的又一途径,在过去几十年,研究者们除了关注"细胞、支架、生长因子"3要素,也开始关注力学条件对构建组织工程软骨的作用。活动关节有复杂的力学性能,关节软骨、软骨基质和其中的细胞都受到不同强度、频率和不同方向的力学刺激,从而影响其功能和结构。在构建组织工程软骨的过程中,添加了力学刺激对软骨细胞的功能、间充质细胞的分化都有重要作用。何种力学条件最有利于构建具有类似天然透明软骨结构和功能的组织工程软骨是该研究领域的热点。     

骨关节炎相关危险因素研究

骨关节炎(osteoarthritis,OA)以关节软骨变性为特征,累及滑膜、关节囊、软骨下骨、韧带以及关节周围的肌肉和肌腱,是一种全关节病。它是由多种因素综合作用而发生的一种复杂病理过程。主要表现为关节疼痛、僵硬、肿胀、活动受限,影像学上表现为骨赘形成、软骨下骨囊性变、软骨下骨硬化以及关节间隙变窄。据估计,65岁以上人群中OA的发病率男性为60%,女性为70%[1]。本文就此相关内容作一综述。1关节软骨的结构和组成关节软骨功能上和结构上可分为4个区带:浅层带、中层带(或过渡带)、深层带和钙化软骨带。在深层带和钙化软骨带之间有潮水标分…     

保留钙化层结构的猪股骨滑车全厚软骨缺损模型建立

目的建立保留钙化层结构的猪股骨滑车全厚软骨缺损模型,为观察组织工程软骨在保留钙化层的膝关节软骨缺损模型中的修复效果提供良好的实验研究平台。方法选取6月龄清洁级贵州小香猪9只,体重40～50 kg,用标准的软骨缺损制作套件在其右后肢股骨滑车切迹旁制备直径6 mm、深0.2～0.5 mm、不伤及钙化层结构的圆柱形全厚软骨缺损模型。造模4周后行3.0T MRI观察,取材后进行大体、体视显微镜观察及固绿-番红O、阿利新蓝、天狼星红组织学染色观察缺损处软骨修复情况。结果造模后实验动物均存活,术后切口无感染,无髌骨脱位;术后即可下地行走并部分负重,1周后均能自由活动,无跛行。造模后4周,MRI检查可见滑车处有明显连续信号中断,异常信号深及软骨下骨,缺损周边深层未见明显信号异常。标本大体观察示缺损底部有少量填充物、出血点,与周围正常软骨界限清楚。体式显微镜观察示钙化层基本完整,缺损局部软骨下骨板有塌陷。普通显微镜下,固绿-番红O及阿利新蓝染色示缺损处无软骨细胞及染料着色;偏光显微镜下,天狼星红染色示缺损底部被连续、强折光性的纤维组织少量填充。结论通过该造模方法制作的不伤及钙化层结构的猪股骨滑车全厚软骨缺损模型,可用于骨关节炎早期软骨病变修复的研究及猪软骨钙化层结构作用研究的动物模型。     

The ultrastructure and biomechanical significance of the tidemark of articular cartilage.

Thirty specimens of human articular cartilage obtained at surgery were examined by scanning electron microscopy to determine the ultrastructure of the tidemark, the junction of the non-calcified and calcified portions of mature articular cartilage. Three distinct variations of the collagen framework of the tidemark were observed: (1) A band of randomly oriented compacted fibrils that appeared to be continuous with those of the non-calcified and calcified zones. (2) A band of flattened fibrils paralleling the undulating surface of the calcified cartilage. (3) A band of perpendicularly oriented fibrils having a distinct continuous transition between the non-calcified and calcified zones, the amount of calcified material applied about the fibrils rapidly increasing as the fibrils entered the calcified zone. The tidemark may serve to provide a tethering mechanism for the relatively flexible and perpendicularly oriented collagen fibrils of the deepest portion of the non-calcified articular cartilage and may prevent them from being sheared at their point of anchorage to the calcified zone. The undulating pattern of the tidemark affords a strong geometric pattern in providing resistance to the shearing action of articulation. Small gaps present in the tidemark may provide pathways for the passage of nutrients into the deep non-calcified zone of articular cartilage from the subchondral bone.     

Effects of hydrocortisone on the vertebral cartilage plate in mice. A light and electron microscopic study

Together with other side effects, the clinical use of steroid appears to produce disorders in the spinal column, especially in young patients. However, morphologic details of steroid induced changes in the spinal column are little known. In this study, mice were treated with daily intramuscular injections of hydrocortisone at doses of 1, 5, and 10 mg/kg body weight for 1, 2, or 4 weeks after 2 weeks of age, and the cartilage plates of the lumbar vertebrae were examined, compared with controls, by light and electron microscopy. Cartilage plates consist of an outer zone, abutting the nucleus pulposus and an inner zone oriented toward the vertebral body. The outer zone is divided into a superficial layer and a deep calcified layer. With steroid therapy, chondrocytes in the inner zone and the superficial layer of the outer zone became degenerative or necrotic. In mice treated with 5 and 10 mg hydrocortisone/kg body weight, ossification appeared earlier in the deep calcified layer than in control animals. After 2 weeks of treatment, the cartilage tissue of the inner zone in mice treated with 10 mg hydrocortisone contained ossification gaps, columnar bone tissue, connecting the bony vertebral body, and the deep calcified layer of the outer zone. The thickness of each layer was measured by light microscopy. By hydrocortisone treatment, the whole of the cartilage plate decreased in thickness, the two cartilage layers of the inner zone and the superficial layer of the outer zone became thinner, and the calcified or ossified layer of the outer zone became widened.(ABSTRACT TRUNCATED AT 250 WORDS)     

Early Effects of Efferent Duct Ligation on the Rat Rete Testis

Early effects of efferent duct ligation (EDL) were studied in adult rats using histological serial sectioning and electron microscopy.
From 30 min to 3 h after EDL the rete cavities dilated and there was accumulation of sperm and cellular debris. The lumen of the transitional zone, the junction of the seminiferous tubules with the tubuli recti, opened and the Sertoli cell bodies straightened up. The direction of sperm tails in the lumen of the seminiferous tubules near the transitional zone changed suggesting a backflow from the rete towards the seminiferous tubules. These changes progressively increased until 12 h. The distensibility of various parts of the rete was unequal. Likewise, the susceptibility of the junctional zone to pressure effects was related on its structure and position. Early changes in the rete testis and the transitional zone epithelium were fully reversible and seemed to be caused by mechanical influence of flow direction only. No evidence of a valve function of the transitional zone was obtained.     

Ultrastructural Modifications of the Extracellular Matrix Upon Calcification of Growth Plate Cartilage as Revealed by Quick-Freeze Deep Etching Technique

The ultrastructural changes in cartilage matrix that occur during calcification have been examined in chick epiphyseal growth plate cartilage prepared by quick-frozen, deep-etched, and rotary shadowed replicas. The extracellular cartilage matrix contains a reticular network closely associated with an extensive network of collagen. The components of the reticular network, including thick and thin filaments, are attached directly to the cell membrane, matrix vesicle membrane, and collagen fibrils. This network, which interconnects the matrix vesicles and collagen, fills the extracellular matrix. The dimensions of the reticular network seem to remain almost constant in size from the reserve and proliferative zones to the calcifying zone. The collagen fibrils seem to consist of subfibrillar structures that branch and anastomose. In optimally quick-frozen, deep-etched, prepared collagen, a cross-banding pattern was exposed. Globular structures stud the collagen fibrils, which gradually diminish in number from the reserve zone down to the calcifying zone. The matrix vesicles, when fractured, showed a granular appearance. In most cases, the fracture plane passed through the bilayer of the matrix vesicle membrane. The true surface of the matrix vesicle membrane, therefore, was exposed after deep etching. At the calcifying zone, crystal deposition had occurred in needle-like and/or plate-like form within the membrane-bound matrix vesicles. The reticular network was still intact in the vicinity of the calcified matrix, but in the intercrystalline space, neither the reticular structure nor the globular structure was detectable. Within the calcified matrix, both reticular and granular structures had disappeared from the interfibrillar space of the collagen fibrils. Received: 19 November 1996 / Accepted: 15 October 1997     

Localization of type X collagen in canine growth plate and adult canine articular cartilage

Type X collagen was extracted from ends of canine growth plates by pepsin digestion after 4 M guanidine hydrochloride extraction, purified by stepwise salt precipitation (2.0 M NaCl in 0.5 M acetic acid), and chromatographed on a Bio-Gel A1.5 M column in 1.0 M CaCl2. Without reduction on sodium dodecyl sulfate (SDS) polyacrylamide gels, the preparation yielded a single, high-molecular-weight (mol wt) band; after reduction, a single band of relative mol wt 5.0 x 10(4) was found. Polyclonal sera were raised against the purified collagen and used in the immunolocalization of canine type X collagen. As expected, indirect immunoperoxidase (IP) or indirect immunofluorescent staining with the polyclonal sera demonstrated that most of the immunoreactivity was localized in the zone of provisional calcification of the growth plate and in cartilage remnants in the metaphyseal region of the physis. A progressive decrease in staining toward the diaphysis of the fetal canine long bone was apparent as the trabecular structures were remodeled to bone. Unexpectedly, type X collagen was also detected in the zone of calcified, mature articular cartilage. It was concentrated in the pericellular matrix of the chondrocytes, appeared at or just above the tidemark, and was expressed immediately before mineralization. Identification of type X collagen in both the canine growth plate and the zone of calcified articular cartilage suggests that cells in the deep layer of cartilage and in the zone of calcified cartilage in the adult animal retain some characteristics of a growth plate and may be involved in regulation of mineralization at this critical interface. The expression of growth plate-like properties would allow the deep chondrocytes of mature articular cartilage to play a role in remodeling of the joint with age and in the pathogenesis of osteoarthritis.     

Elastic modulus of calcified cartilage is an order of magnitude less than that of subchondral bone

The elastic moduli of calcified cartilage and subchondral bone tissues were measured experimentally with use of a three-point bending test. Specimens were obtained from a bovine patella and the distal end of a bovine femur, from two different animals. Fifteen specimens were tested as “pure” subchondral bone beams, and 15 were tested as composite calcified cartilage/subchondral bone beams. A least-squares optimization scheme was used to obtain modulus values from the composite beams. The elastic modulus for subchondral bone calculated from the “pure” subchondral bone beams was 2.3 ± 1.5 GPa (3.9 ± 1.5 GPa for specimens from the femur and 1.6 ± 0.7 GPa for specimens from the patella). The composite beam optimization resulted in a modulus for subchondral bone of 5.7 ± 1.9 GPa and a modulus for calcified cartilage of 0.32 ± 0.25 GPa. The modulus for the calcified cartilage was more than an order of magnitude lower than the modulus of the underlying subchondral bone. This supports the idea that the zone of calcified cartilage forms a transitional zone of intermediate stiffness between the articular cartilage and the subchondral bone.     

Ultrastructural cytochemistry of complex carbohydrates in osteoblasts, osteoid, and bone matrix

Glycosaminoglycans (GAGs) and glycoproteins are essential components for osteogenesis. We have examined rat osteoblasts, osteoid, transitional zone, and fully calcified bone matrix, utilizing Spicer's high-iron diaminethiocarbohydrazide-silver protein (HID-TCH-SP) method for sulfated glycoconjugates and Thiéry's periodate-TCH-SP (PA-TCH-SP) method for vicinal glycol-containing glycoconjugates. HID-TCH-SP stained cytoplasmic granules of osteoblasts. Stain deposits in the extracellular matrix were observed in decreasing amounts in osteoid, the transitional zone, and fully calcified bone matrix. Enzyme digestion with testicular hyaluronidase removed most HID-TCH-SP stain deposits. PA-TCH-SP staining was observed with increasing intensity in rough endoplasmic reticulum, Golgi saccules, and cytoplasmic granules. Collagen fibrils in osteoid were weakly stained with PA-TCH-SP, and their staining appeared even weaker in fully calcified bone matrix. In contrast, collagen fibrils in calcified cartilage stained intensely with the PA-TCH-SP method. Focal circular profiles (0.1-0.5 mumol in diameter), which lacked collagen fibrils but reacted moderately with PA-TCH-SP, were frequently seen in the transitional zone and fully calcified bone matrix, but were only occasionally present in osteoid. The presence of testicular hyaluronidase-resistant GAG and acid phosphatase in these focal areas suggests that they represent sites of GAG degradation. The eventual loss of HID-TCH-SP staining in the bone matrix suggests that removal of sulfated glycoconjugates may be a requisite for expansion of initial calcification sites and/or complete calcification.