In situ measurement of transport between subchondral bone and articular cartilage

Subchondral bone and articular cartilage play complementary roles in load bearing of the joints. Although the biomechanical coupling between subchondral bone and articular cartilage is well established, it remains unclear whether direct biochemical communication exists between them. Previously, the calcified cartilage between these two compartments was generally believed to be impermeable to transport of solutes and gases. However, recent studies found that small molecules could penetrate into the calcified cartilage from the subchondral bone. To quantify the real‐time solute transport across the calcified cartilage, we developed a novel imaging method based on fluorescence loss induced by photobleaching (FLIP). Diffusivity of sodium fluorescein (376 Da) was quantified to be 0.07 ± 0.03 and 0.26 ± 0.22 µm²/s between subchondral bone and calcified cartilage and within the calcified cartilage in the murine distal femur, respectively. Electron microscopy revealed that calcified cartilage matrix contained nonmineralized regions (～22% volume fraction) that are either large patches (53 ± 18 nm) among the mineral deposits or numerous small regions (4.5 ± 0.8 nm) within the mineral deposits, which may serve as transport pathways. These results suggest that there exists a possible direct signaling between subchondral bone and articular cartilage, and they form a functional unit with both mechanical and biochemical interactions, which may play a role in the maintenance and degeneration of the joint. © 2009 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 27:1347–1352, 2009     

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.     

Elevated cross-talk between subchondral bone and cartilage in osteoarthritic joints

Osteoarthritis (OA) is a degenerative joint disease and one of the leading causes of disability in the United States and across the world. As a disease of the whole joint, OA exhibits a complicated etiology with risk factors including, but not limited to, ageing, altered joint loading, and injury. Subchondral bone is hypothesized to be involved in OA development. However, direct evidence supporting this is lacking. We previously detected measurable transport of solute across the mineralized calcified cartilage in normal joints, suggesting a potential cross-talk between subchondral bone and cartilage. Whether this cross-talk exists in OA has not been established yet. Using two models that induced OA by either ageing or surgery (destabilization of medial meniscus, DMM), we tested the hypothesis that increased cross-talk occurs in OA. We quantified the diffusivity of sodium fluorescein (mol. wt. 376Da), a marker of small-sized signaling molecules, within calcified joint matrix using our newly developed fluorescence loss induced by photobleaching (FLIP) method. Tracer diffusivity was found to be 0.30±0.17 and 0.33±0.20μm(2)/s within the calcified cartilage and 0.12±0.04 and 0.07±0.03μm(2)/s across the osteochondral interface in the aged (20-24-month-old, n=4) and DMM OA joints (5-month-old, n=5), respectively, which were comparable to the control values for the contralateral non-operated joints in the DMM mice (0.48±0.13 and 0.12±0.06μm(2)/s). Although we did not detect significant changes in tissue matrix permeability in OA joints, we found i) an increased number of vessels invading the calcified cartilage (and sometimes approaching the tidemark) in the aged (+100%) and DMM (+50%) joints relative to the normal age controls; and ii) a 60% thinning of the subchondral bone and calcified cartilage layers in the aged joints (with no significant changes detected in the DMM joints). These results suggested that the capacity for cross-talk between subchondral bone and articular cartilage could be elevated in OA. Further studies are needed to identify the direction of the cross-talk, the signaling molecules involved, and to test whether subchondral bone initiates OA development and could serve as a pharmaceutical target for OA treatment. This article is part of a Special Issue entitled "Osteoarthritis".     

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。     

Variations in articular calcified cartilage by site and exercise in the 18-month-old equine distal metacarpal condyle

OBJECTIVES: To interrelate articular calcified cartilage thickness, mineralisation density, tidemark count and tidemark linear accretion rate by site in the equine third metacarpal distal condyle. To determine the effects of exercise during early life on articular calcified cartilage. METHOD: Six of 12 pasture-raised Thoroughbred horses were exercised from 10 days old. Calcein labels were given 19 and 8 days prior to euthanasia at 18 months old. Osteochondral specimens were cut from the distal third metacarpal condyle and imaged using confocal scanning light microscopy (CSLM) and quantitative backscattered electron scanning electron microscopy (qBSE). Articular calcified cartilage thickness and total thickness mineralisation density were measured on montaged qBSE image sets, and inter-label mineralisation density, tidemark count and linear accretion rate measured on registered CSLM-qBSE image pairs. RESULTS: Calcified cartilage thickness, mineralisation density, tidemark count and linear accretion rate varied significantly between sites. Regions with thinner calcified cartilage had greater linear accretion rates, hence rapid chondroclastic resorption. Mineralisation density was positively correlated with linear accretion rate. Fewer multiple tidemarks were counted in regions with greater linear accretion rates. Lag time between the tidemark and cement line was estimated (180 days; in the range of 0-648 days). Exercise had little effect on measured parameters. CONCLUSION: The major determinant of articular calcified cartilage thickness is the rate of chondroclastic resorption, not tidemark linear accretion rate. Our evidence supports coupled, mechanosensitive regulation of chondroclastic resorption and linear accretion rate in articular calcified cartilage. Exercising pasture-reared foals causes little additional adaptation in distal third metacarpal articular calcified cartilage.     

The cartilage-bone interface

In the knee joint, the purpose of the cartilage-bone interface is to maintain structural integrity of the osteochondral unit during walking, kneeling, pivoting, and jumping--during which tensile, compressive, and shear forces are transmitted from the viscoelastic articular cartilage layer to the much stiffer mineralized end of the long bone. Mature articular cartilage is integrated with subchondral bone through a approximately 20 to approximately 250 microm thick layer of calcified cartilage. Inside the calcified cartilage layer, perpendicular chondrocyte-derived collagen type II fibers become structurally cemented to collagen type I osteoid deposited by osteoblasts. The mature mineralization front is delineated by a thin approximately 5 microm undulating tidemark structure that forms at the base of articular cartilage. Growth plate cartilage is anchored to epiphyseal bone, sometimes via a thin layer of calcified cartilage and tidemark, while the hypertrophic edge does not form a tidemark and undergoes continual vascular invasion and endochondral ossification (EO) until skeletal maturity upon which the growth plates are fully resorbed and replaced by bone. In this review, the formation of the cartilage-bone interface during skeletal development and cartilage repair, and its structure and composition are presented. Animal models and human anatomical studies show that the tidemark is a dynamic structure that forms within a purely collagen type II-positive and collagen type I-negative hyaline cartilage matrix. Cartilage repair strategies that elicit fibrocartilage, a mixture of collagen type I and type II, are predicted to show little tidemark/calcified cartilage regeneration and to develop a less stable repair tissue-bone interface. The tidemark can be regenerated through a bone marrow-driven growth process of EO near the articular surface.     

The structure of the human subchondral plate

To study the anatomy of subarticular bone and cartilage, fresh specimens of cartilage on bone from the human shoulder, hip and knee were treated with bleach or papain, or were fixed and decalcified. All were compared using scanning electron microscopy. Papain digestion selectively removed cartilage to the tidemark. The tidemark contour was highly variable; irregularities were indirectly related to degenerative lesions and were most prominent in peripheral non-weight-bearing areas of joints with central fibrillation. Decalcification exposed the interface between the bone and calcified cartilage. Collagen fibrils in articular cartilage did not interdigitate with those of bone. The subchondral bone was appositional, avascular, smooth and very thin in most areas of human joints. Perforations through subchondral bone or calcified cartilage were rare. Bleach maceration destroyed important details.     

Osteoarthrotic changes after acute transarticular load. An animal model.

The canine patellofemoral joint was subjected to a standardized transarticular load of 2170 newtons for two milliseconds, and the gross and histological changes were examined at two, twelve, and twenty-four weeks after injury. Initially, the load creates fractures in the zone of calcified cartilage, with minimum damage to the articular cartilage surface. Surface fissures were visible in all patellae only after staining with India ink. Histologically, these surface clefts extended into the transitional or superficial radial zone, and they did not communicate with the subchondral bone except in two patellae. However, there were reproducible clefts in the region of the subchondral bone and the zone of calcified cartilage in all patellae. Six months after loading, there was a loss of safranin-O staining above the deep clefts, and there was new-bone formation in the subchondral region and fibrillation of the cartilaginous surface. Thus, the initial changes had progressed to osteoarthrotic-like conditions at six months. In this animal model, the joint is not invaded and the changes that result from loading are reproducible. The injury to the joint creates superficial disruption of the cartilage and subchondral changes that lead to arthritic-like degeneration of the cartilage within six months.     

The normal human chondro-osseous junctional region: evidence for contact of uncalcified cartilage with subchondral bone and marrow spaces

Background  

The chondro-osseous junctional region of diarthrodial joints is peculiarly complex and may be considered to consist of the deepest layer of non-calcified cartilage, the tidemark, the layer of calcified cartilage, a thin cement line (between the calcified cartilage and the subchondral bone) and the subchondral bone. A detailed knowledge of the structure, function and pathophysiology of the normal chondro-osseous junction is essential for an understanding of the pathogenesis of osteoarthrosis.     

Role of endochondral ossification of articular cartilage and functional adaptation of the subchondral plate in the development of fatigue microcracking of joints

The mechanisms that regulate functional adaptation of the articular ends of long bones are poorly understood. However, endochondral ossification of articular cartilage and modeling/remodeling of the subchondral plate and epiphyseal trabeculae are important components of the adaptive response. We performed a histologic study of the distal end of the third metacarpal/metatarsal bone of Thoroughbreds after bones were bulk-stained in basic fuchsin and calcified sections were prepared. The Thoroughbred racehorse is a model of an extreme athlete which experiences particularly high cyclic strains in distal limb bones. The following variables were quantified: microcrack boundary density in calcified cartilage (N.Cr/B.Bd); blood vessel boundary density in calcified cartilage (N.Ve/B.Bd); calcified cartilage width (Cl.Cg.Wi); duplication of the tidemark; and bone volume fraction of the subchondral plate (B.Ar/T.Ar). Measurements were made in five joint regions (lateral condyle and condylar groove; sagittal ridge; medial condylar and condylar groove). N.Cr/B.Bd was site-specific and was increased in the condylar groove region; this is the joint region from which parasagittal articular fatigue (condylar) fractures are typically propagated. Formation of resorption spaces in the subchondral plate was co-localized with microcracking. N.Ve/B.Bd was also site-specific. In the sagittal ridge region, N.Ve/B.Bd was increased, Cl.Cg.Wi was decreased, and B.Ar/T.Ar was decreased, when compared with the other joint regions. Multiple tidemarks were seen in all joint regions. Cumulative athletic activity was associated with a significant decrease in B.Ar/T.Ar in the condylar groove regions. N.Cr/B.Bd was positively correlated with B.Ar/T.Ar (P < 0.05, r(s) = 0.29) and N.Ve/B.Bd was negatively correlated with B.Ar/T.Ar (P < 0.005, r2 = 0.14) and Cl.Cg.Wi (P < 0.05, r2 = 0.07). We conclude that endochondral ossification of articular cartilage and modeling/remodeling of the subchondral plate promote initiation and propagation of site-specific fatigue microcracking of the joint surface, respectively, in this model. Microcracking of articular calcified cartilage likely represents mechanical failure of the joint surface. Propagation of microcracks into the subchondral plate is a critical factor in the pathogenesis of articular condylar fatigue (stress) fracture. Functional adaptation of the joint likely protects hyaline cartilage from injury in the short-term but may promote joint degeneration and osteoarthritis with ongoing athleticism.     

The effect of varus stress on the moving rabbit knee joint

Unicompartmental osteoarthritis was produced by applying varus stress to moving rabbit knee joints. Degenerative changes were confined to the medial tibial and the medial femoral articular surfaces. Within the range of varus stress used, duration appears to be more important than magnitude of varus stress in determining the severity of cartilage damage. The calcified zone remained histologically unchanged despite advanced changes in the noncalcified zone superficial to the tidemark. Intrachondral degenerative cysts were frequently found in the basilar layers of the noncalcified cartilage adjacent to the tidemark where shear stresses were likely to be highest and diffusible nutrients least available. Highly cellular cartilaginous tissue was noted in the subchondral marrow spaces in the specimens with advanced cartilage degeneration. These areas appeared to be continuous with the overlying degenerated cartilage through gaps in the calcified cartilage. Subchondral bone did not show remarkable trabecular thickening despite advanced degenerative changes in the articular cartilage.     

Interaction between zonal populations of articular chondrocytes suppresses chondrocyte mineralization and this process is mediated by PTHrP

OBJECTIVE: Articular cartilage is separated from subchondral bone by the tidemark and a calcified cartilage zone. Advancement of the calcified region and tidemark duplication are both hallmarks of osteoarthritis (OA). Currently the mechanisms controlling post-natal articular cartilage mineralization are poorly understood. The objective of this study is to test the hypothesis that cellular communication between different cartilage layers regulates articular chondrocyte mineralization. DESIGN: Co-culture models were established to evaluate the interaction of chondrocytes derived from the surface, middle and deep zones of articular cartilage. The cultures were stimulated with triiodothyronine (T3) to promote chondrocyte hypertrophy. The effects of zonal chondrocyte interactions on chondrocyte mineralization were examined over time. RESULTS: Co-culture of deep zone chondrocytes (DZCs) with surface zone chondrocytes (SZCs) suppressed the T3-induced increase in alkaline phosphatase (ALP) activity and related mineralization. Moreover, SZC-DZC co-culture was associated with a significantly higher parathyroid hormone-related peptide (PTHrP) expression when compared to controls. When PTHrP(1-40) was added to the DZC-only culture, it suppressed DZC ALP activity similar to the inhibition observed in co-culture with SZC. In addition, treatment with PTHrP reversed the effect of T3 stimulation on the expression of hypertrophic markers (Indian hedgehog, ALP, matrix metalloproteinases-13, Type X collagen) in the DZC cultures. Moreover, blocking the action of PTHrP significantly increased ALP activity in SZC+DZC co-culture. CONCLUSION: Our findings demonstrate the role of zonal chondrocyte interactions in regulating cell mineralization and provide a plausible mechanism for the post-natal regulation of articular cartilage matrix organization. These findings also have significant implications in understanding the pathology of articular cartilage as well as devising strategies for functional cartilage repair.     

Effects of weight bearing on the tidemark and osteochondral junction of articular cartilage: Histomorphometric analyses of 7 normal femoral heads

To study the effect of weight bearing on the tidemark and osteochondral junction, we compared the morphology of these two boundaries in weight-bearing and less weight-bearing regions of normal human femoral heads. We measured the irregularities of the boundaries in the two regions using an X-Y digitizer connected to a computer in histological whole sections of femoral heads in 7 subjects without joint diseases.

The irregularity of the tidemark was small, showing no difference between the two regions. However, the irregularity of the osteochondral junction in the weight-bearing region was greater than in the less weight-bearing region, which was confirmed by three-dimensional reconstructed images. Our findings suggest that mechanical stresses greatly influence the morphology of the osteochondral junction compared to the influence of such stresses on the tidemark, and that the marked irregularity of the osteochondral junction in the weight-bearing region is a reactive phenomenon against mechanical fragility due to simple contact between calcified cartilage and subchondral bone without fibrous connections.     

Effects of weight bearing on the tidemark and osteochondral junction of articular cartilage: histomorphometric analyses of 7 normal femoral heads

To study the effect of weight bearing on the tidemark and osteochondral junction, we compared the morphology of these two boundaries in weight-bearing and less weight-bearing regions of normal human femoral heads. We measured the irregularities of the boundaries in the two regions using an X-Y digitizer connected to a computer in histological whole sections of femoral heads in 7 subjects without joint diseases. The irregularity of the tidemark was small, showing no difference between the two regions. However, the irregularity of the osteochondral junction in the weight-bearing region was greater than in the less weight-bearing region, which was confirmed by three-dimensional reconstructed images. Our findings suggest that mechanical stresses greatly influence the morphology of the osteochondral junction compared to the influence of such stresses on the tidemark, and that the marked irregularity of the osteochondral junction in the weight-bearing region is a reactive phenomenon against mechanical fragility due to simple contact between calcified cartilage and subchondral bone without fibrous connections.     

Lead accumulation in tidemark of articular cartilage

OBJECTIVE: Determination of the spatial distribution of the toxic element lead (Pb) and other trace elements in normal articular cartilage and subchondral bone from adult humans with no history of work-related exposure to Pb. METHODS: Four macroscopically normal femoral heads and three patellas were harvested from randomly selected forensic autopsies. All subjects died of acute illnesses, had no history of work-related exposure to Pb and had no metabolic bone disease. The elemental distribution of lead (Pb) together with zinc (Zn), strontium (Sr) and calcium (Ca) in the chondral and subchondral region was detected using high resolution synchrotron radiation induced micro X-ray fluorescence (SR mu-XRF) analysis. SR mu-XRF line scans in conventional and SR mu-XRF area scans in confocal geometry were correlated to backscattered electron (BE) images visualizing the mineralized tissue. RESULTS: In all samples, we found a highly specific accumulation of Pb in the tidemark, the transition zone between calcified and non-calcified articular cartilage. Pb fluorescence intensities in the tidemark, which is thought to be a metabolically active mineralization front, were 13-fold higher when compared to subchondral bone. Pb intensities in the subchondral region were strongly correlated with Zn, but were distinctly different from Ca and Sr. CONCLUSIONS: The finding of the highly specific accumulation of lead in the tidemark of human articular cartilage is novel. However at this point, the exact mechanisms of the local Pb accumulation as well as its clinical implications are unknown.     

Subfracture insult to the human cadaver patellofemoral joint produces occult injury

The current criterion used by the automotive industry for injury to the lower extremity is based on visible bone fracture. Studies suggest, however, that chronic joint degeneration may occur after subfracture impact loads on the knee. We hypothesized that subfracture loading of the patellofemoral joint could result in previously undocumented microtrauma in areas of high contact pressure. In the current study, seven patellofemoral joints from human cadavers were subjected to impact with successively greater energy until visible fracture was noted. Transverse and comminuted fractures of the patella were noted at 6.7 kN of load. Approximately 45% of the impact energy then was delivered to the contralateral joint. Subfracture loads of 5.2 kN resulted in no gross bone fracture in five of seven specimens. Histological examination of the patellae revealed occult trauma in four of the seven specimens in the subfracture group; trauma consisted of a horizontal split fracture in the subchondral bone, at the tidemark, or at the interface of calcified cartilage and subchondral bone. The trauma appeared predominantly on the lateral facet, adjacent to or directly beneath preexisting fibrillation of the articular surface. Surface fibrillation was noted in histological sections of control patellae (not subjected to impact loading), but occult damages were not observed. Although the mechanism of this occult trauma is unknown, similar damage has been shown to occur from direct shear loading. As these microcracks can potentiate a disease process in the joint, this study may suggest that the current criterion for injury, based on bone fracture alone, is not sufficiently conservative.     

Cyclical articular joint loading leads to cartilage thinning and osteopontin production in a novel in vivo rabbit model of repetitive finger flexion

OBJECTIVE: An in vivo rabbit model of repetitive joint flexion and loading was used to characterize the morphological effects of cyclical loading on articular cartilage. DESIGN: The forepaw digits of eight anesthetized New Zealand White adult female rabbits were repetitively flexed at 1 Hz with a mean peak digit load of 0.42 N for 2 h per day for 60 cumulative hours. Metacarpophalangeal joints were collected from loaded and contra-lateral control limbs, fixed, decalcified, embedded, and thin-sectioned. Serial sections were stained for histology or for immunohistochemistry. Morphometric data including the mean thicknesses of the uncalcified cartilage and of the calcified cartilage were collected from digital photomicrographs of safranin O-stained sections. The number of cells stained with anti-osteopontin antibody was counted. RESULTS: We observed a decrease in uncalcified cartilage mean thickness with no significant change in calcified cartilage thickness. We also observed a significant increase in the number of cells positive for osteopontin (OPN) in the uncalcified cartilage. These changes occurred without overt cartilage surface degeneration. CONCLUSIONS: Cyclical loading leads to changes at the tissue and cellular levels in articular cartilage. These changes are suggestive of tidemark advancement and may indicate a reactivation of cartilage mineralization steps analogous to endochondral ossification. This novel in vivo rabbit model of repetitive flexion and loading can be used to investigate the effects of cyclical loading on articular joints.     

Changes induced by chronic in vivo load alteration in the tibiofemoral joint of mature rabbits

We investigated the relationship between the magnitude and duration of chronic compressive load alteration and the development and progression of degenerative changes in the rabbit tibiofemoral joint. Varus loading devices were attached to the hind limb of mature NZW rabbits. Altered compressive loads of 0%, 50%, and 80% body weight (BW) were applied to the tibiofemoral joint for 12 h per day for 12 and 24 weeks (n = 4 animals/group). Compartment‐specific assessment of the tibial plateau included histological assessments (articular cartilage, calcified cartilage, and subchondral bone thicknesses, degeneration score, and articular cartilage cellularity) and biomechanical measures (aggregate modulus, permeability, Poisson's ratio). Analyses of variance techniques were used to examine the relationship between each outcome measure with load magnitude and duration as independent variables in the model. Degenerative changes developed in the medial compartment with increased magnitude of compressive loading and included fibrillation, increased degeneration score, and reduced cellularity of the articular cartilage. Increased calcified cartilage thickness was observed in both the medial and lateral compartments following exposure to altered loading of 80% BW for 24 weeks. This work demonstrates that in vivo chronic compressive load alteration to the tibiofemoral joint can initiate progressive macroscopic and histological‐based degenerative changes analogous to the early changes occurring in OA. © 2012 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 30:1413–1422, 2012     

Regenerating hyaline cartilage in articular defects of old chickens using implants of embryonal chick chondrocytes embedded in a new natural delivery substance

Summary Partial and full thickness defects were created mechanically in articular cartilage and subchondral bone of the tibiotarsal joint condyles of 3-year-old chickens. The wounds were then repaired using embryonal chick chondrocytes embedded in a new biocompatible, hyaluronic acid-based delivery substance. Controls were similarly operated on but received either no treatment or implants of the delivery substance only. Animals were killed from 1 week to 6 months postoperatively. Sections from the two groups were examined and compared macroscopically, histologically, and histochemically. Results of 6-month follow-up showed that only the defects of the experimental chickens were completely filled with reparative hyaline cartilage tissue, with no signs of inflammation or immunologic rejection. Initially the entire defect cavity, whether partial thickness or full thickness up to the deep regions in the subchondral bone, was filled with cartilaginous reparative tissue. Relatively rapid maturation occurred under the tidemark; chondrocytes hypertrophied, were invaded with vascular elements and ossified. In the superficial areas, the reparative tissue remained cartilaginous and matured as typical hyaline cartilage tissue. These results indicate that aged chicken cartilage and its accompanying thin and spongy osteoporotic bone offer a favorable host environment for embryonal cell implants.     

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。