Mechanical behavior of intact and low-grade degenerated cartilage.

Young's modulus, elastic and plastic deformation, mechanical hardness and load at failure were determined for low-grade degenerated hyaline cartilage in a porcine model. Osteochondral plugs from the medial condyle of 30 female pigs were used. Cartilage defects were classified using the International Cartilage Repair Society (ICRS) protocol. Mechanical hardness was measured using a Shore A testing device. Total stiffness and plastic deformation was evaluated in the range 50-200 N using a 5-mm indenter. The load at failure was then determined. ICRS grade I specimens showed significantly lower stiffness than grade 0 specimens. ICRS grade 0 specimen showed no significant plastic deformation within the load range 25-100 N. In degenerated cartilage, plastic deformation started at a significantly lower load (50 N). The Young's modulus at 25 N in ICRS grade 0 specimens (18.8 MPa) was significantly higher than in grade I (11.1 MPa) or grade II (10.5 MPa) specimens. Intact cartilage showed significantly higher tension at failure and mechanical Shore A hardness. Young's modulus and tension at failure showed strong correlation. Cartilage degeneration is associated with a significant loss of elasticity and mechanical stress resistance. Shore hardness measurement is an adequate method for rapid biomechanical evaluation of cartilage specimens.     

水的饱和辛醇溶液水分标准物质的研制

生物燃料的国家标准规定了产品的技术指标和相应的检测方法。水的饱和辛醇溶液的水分标准物质,用于生物燃料水分测量时仪器的校准和方法的验证,能够保障测量结果的准确可靠和等效一致。该标准物质采用卡尔·费休库仑法、卡尔·费休容量法和定量核磁共振等三种不同原理的方法定值。通过方法研究和改进,实现了库仑法和容量法的一致;通过引入新的核磁共振方法,提高了结果的准确性。最终标准物质的水分量值为4.76％,扩展不确定度为0.09％。     

A computational algorithm to simulate disorganization of collagen network in injured articular cartilage

Cartilage defects are a known risk factor for osteoarthritis. Estimation of structural changes in these defects could help us to identify high risk defects and thus to identify patients that are susceptible for the onset and progression of osteoarthritis. Here, we present an algorithm combined with computational modeling to simulate the disorganization of collagen fibril network in injured cartilage. Several potential triggers for collagen disorganization were tested in the algorithm following the assumption that disorganization is dependent on the mechanical stimulus of the tissue. We found that tensile tissue stimulus alone was unable to preserve collagen architecture in intact cartilage as collagen network reoriented throughout the cartilage thickness. However, when collagen reorientation was based on both tensile tissue stimulus and tensile collagen fibril strains or stresses, the collagen network architecture was preserved in intact cartilage. Using the same approach, substantial collagen reorientation was predicted locally near the cartilage defect and particularly at the cartilage–bone interface. The developed algorithm was able to predict similar structural findings reported in the literature that are associated with experimentally observed remodeling in articular cartilage. The proposed algorithm, if further validated, could help to predict structural changes in articular cartilage following post-traumatic injury potentially advancing to impaired cartilage function.     

Uranyl tris nitrato as a luminescent probe for trace water detection in acetonitrile

Uranyl tris nitrato i.e. [UO₂(NO₃)₃]^– was formed by adding tetramethylammonium nitrate to uranyl nitrate in acetonitrile medium. The luminescence features of this complex in acetonitrile are very sensitive to water content, which could lead to the use of it as a luminescent probe for water present in acetonitrile. The luminescence intensity ratio of 507 to 467 nm peak of uranyl tris nitrato showed a linear response in the range 0–5% (v/v) water content in acetonitrile. The present method was applied for three synthetic samples of acetonitrile for water detection and the results obtained were compared using Karl Fischer titration. There was a good agreement in the values obtained by both the methods.     

Viability of diced, crushed cartilage grafts and the effects of Surgicel (oxidized regenerated cellulose) on cartilage grafts

The viability of cartilage grafts has been well documented; however, controversy still exists about the viability of crushed cartilage. Recently, there has been a tendency to use diced cartilage grafts wrapped with oxidized regenerated cellulose (Surgicel) sheets for improving dorsal contour in rhinoplasty. The viability of diced cartilage grafts and the effect of Surgicel on cartilage grafts are not well known. In this study, we used ear cartilage from 18 New Zealand rabbits. Cartilage grafts were transplanted to surgically created subcutaneous pockets on the back of the rabbits on both the left and right sides. There were three groups: (1) intact cartilage grafts, (2) crushed cartilage grafts, and (3) diced cartilage grafts. The grafts that were transplanted to the right side were wrapped with Surgicel. Cartilage grafts in all groups were viable. In grafts that were wrapped with Surgicel, a marked increase in the collagen content was investigated. Grafts that were wrapped with Surgicel demonstrated no evidence of proliferation, whereas the bare cartilage grafts demonstrated significant amounts of proliferation.     

On the Determination of Water Content in Biomass Processing

Processing of lignocellulosic materials to fuels such as methane and bioethanol may involve several processing steps including pretreatment, saccharification, fermentation, and anaerobic digestion. The amounts of substrate used in these processes are usually based on dry matter content, and the processes themselves typically lead to a change in dry matter content. Thus, it is of great importance to be able to measure dry matter accurately. Dry matter content is commonly determined by measuring loss of water during oven drying. We have used Karl Fischer (KF) titration to measure the water content in a wide range of biomass fractions and have compared these data to results obtained by oven drying. This revealed considerable differences for all tested materials. For lignocellulosic materials, oven drying tends to overestimate dry matter content for untreated material. On the other hand, oven drying generally underestimates dry matter content in pretreated materials due to loss of organic volatiles. These differences have major consequences for the calculation of mass balances and yields in bioprocessing. The KF method gives more accurate water determination than oven drying due to the unique selectivity of the analysis. The method is suitable for the analysis of lignocellulosic biomasses and is particularly useful for determination of water content in pretreated materials, where oven drying usually underestimates the dry matter content due to loss of volatiles.     

New selective method for quantification of organosilanol groups in silicone pre-elastomers

The silicone elastomers used for drug delivery are normally reinforced by fumed silica, which contains a high density of silanol groups. These inorganic silanol groups have to be deactivated in order to avoid stiffening the uncured pre-elastomer, also called creep hardening. One commonly used way of achieving this deactivation is to mix the material with low molecular mass organosilanols at an elevated temperature. It is important to be able to quantify the nonbonded organosilanols remaining in the material after manufacture. Traditional testing does not distinguish between inorganic silanols and organosilanols. A new selective method for the quantification of organosilanol groups in silicone pre-elastomers has therefore been developed. This method is based on derivatization of the silanol groups with a mixture of dimethylphenylchlorosilane and tetramethyldiphenylsilazane, so that the silanol groups are replaced with a dimethylphenyl group. The derivatized organosilanols are then determined by liquid chromatography using a size exclusion column and a UV detector. No interference was found from other groups normally present in medical grade pre-elastomers, such as vinyls, hydrides, and inorganic bonded silanol on silica or water. The results agreed well with the nonselective Karl Fischer titration for some short chain silanols.     

骨碎补结合组织工程软骨促进软骨再生的实验研究

目的:评估骨碎补结合组织工程软骨治疗对实验兔软骨缺陷模型软骨再生的疗效。方法:将h IGF-1基因转染MSCs,并与脱细胞真皮基质(ADM)构建组织工程软骨。24只新西兰白兔随机分为A、B、C、D四组,A、C组进行自体软骨移植,B、D组进行改建的细胞-ADM移植。C、D组用40%骨碎补汤喂养4周,150 m L/d。第12周处死实验动物,分离缺损关节软骨部位,蜡块包埋染色,通过总体形态评价软骨再生组织。采用组织学评分评估再生软骨质量。采用甲苯胺蓝染色评价缺损部位产生软骨糖胺聚糖的情况。结果:与B组比较,C组和D组的新生软骨覆盖度、新骨髓的颜色、缺损边缘和表面粗糙度均显著提高(P0.05);再生软骨的组织学评分软骨表面评分显著改善(P0.05)。C组与D组具有比其他组更好的基质、细胞分布和表面指数。并且有较厚的透明样软骨组织,具有正常的糖胺聚糖产生。表明该治疗方法可以通过再生透明样软骨且没有不良事件来减少软骨缺陷。结论:工程软骨结合骨碎补治疗可显著改善兔膝关节软骨缺损修复的质量,为临床治疗软骨病变提供重要理论依据。     

Quantitative ultrasound can assess the regeneration process of tissue-engineered cartilage using a complex between adherent bone marrow cells and a three-dimensional scaffold

Articular cartilage (hyaline cartilage) defects resulting from traumatic injury or degenerative joint disease do not repair themselves spontaneously. Therefore, such defects may require novel regenerative strategies to restore biologically and biomechanically functional tissue. Recently, tissue engineering using a complex of cells and scaffold has emerged as a new approach for repairing cartilage defects and restoring cartilage function. With the advent of this new technology, accurate methods for evaluating articular cartilage have become important. In particular, in vivo evaluation is essential for determining the best treatment. However, without a biopsy, which causes damage, articular cartilage cannot be accurately evaluated in a clinical context. We have developed a novel system for evaluating articular cartilage, in which the acoustic properties of the cartilage are measured by introducing an ultrasonic probe during arthroscopy of the knee joint. The purpose of the current study was to determine the efficacy of this ultrasound system for evaluating tissue-engineered cartilage in an experimental model involving implantation of a cell/scaffold complex into rabbit knee joint defects. Ultrasonic echoes from the articular cartilage were converted into a wavelet map by wavelet transformation. On the wavelet map, the percentage maximum magnitude (the maximum magnitude of the measurement area of the operated knee divided by that of the intact cartilage of the opposite, nonoperated knee; %MM) was used as a quantitative index of cartilage regeneration. Using this index, the tissue-engineered cartilage was examined to elucidate the relations between ultrasonic analysis and biochemical and histological analyses. The %MM increased over the time course of the implant and all the hyaline-like cartilage samples from the histological findings had a high %MM. Correlations were observed between the %MM and the semiquantitative histologic grading scale scores from the histological findings. In the biochemical findings, the chondroitin sulfate content increased over the time course of the implant, whereas the hydroxyproline content remained constant. The chondroitin sulfate content showed a similarity to the results of the %MM values. Ultrasonic measurements were found to predict the regeneration process of the tissue-engineered cartilage as a minimally invasive method. Therefore, ultrasonic evaluation using a wavelet map can support the evaluation of tissue-engineered cartilage using cell/scaffold complexes.     

The acute effect of bipolar radiofrequency energy thermal chondroplasty on intrinsic biomechanical properties and thickness of chondromalacic human articular cartilage

Radio frequency energy (RFE) thermal chondroplasty has been a widely-utilized method of cartilage debridement in the past. Little is known regarding its effect on tissue mechanics. This study investigated the acute biomechanical effects of bipolar RFE treatment on human chondromalacic cartilage. Articular cartilage specimens were extracted (n?=?50) from femoral condyle samples of patients undergoing total knee arthroplasty. Chondromalacia was graded with the Outerbridge classification system. Tissue thicknesses were measured using a needle punch test. Specimens underwent pretreatment load-relaxation testing using a spherical indenter. Bipolar RFE treatment was applied for 45?s and the indentation protocol was repeated. Structural properties were derived from the force-time data. Mechanical properties were derived using a fibril-reinforced biphasic cartilage model. Statistics were performed using repeated measures ANOVA. Cartilage thickness decreased after RFE treatment from a mean of 2.61?mm to 2.20?mm in Grade II, II-III, and III specimens (P?相似文献   

Regional differences of tibial and femoral cartilage in the chondrocyte gene expression,immunhistochemistry and composite in different stages of osteoarthritis

The function of articular cartilage as an avascular tissue is mainly served by collagen type II and proteoglycan molecules. Within this matrix homeostasis between production and breakdown of the matrix is exceptionally sensitive.The current study was conducted to identify regional differences in specific alterations in cartilage composition during the osteoarthritic process of the human knee joint. Therefor the changes in the expression of the key molecules of the extracellular matrix were measured in dependence of the anatomical side (femoral vs tibial) and associated with immunohistochemistry and quantitative measurement.60 serial osteochondral femoral condyle and the tibial plateau samples of patients undergoing implantation of total knee endoprosthesis of areas showing mild (Group A, macroscopically ICRS grade 1b) respectively advanced (Group B, macroscopically ICRS grade 3a/3b) (30 each) osteoarthritis according to the histological-histochemical grading system (HHGS) were compared with 20 healthy biopsies with immunohistochemistry and histology. We quantified our results on the gene expression of collagen type I and II and aggrecan with the help of real-time (RT)-PCR. Proteoglycan content was measured colorometrically.In group A slightly increased colour intensity was found for collagen II in deeper layers, suggesting a persisting but initially still intact repair process. But especially on the medial tibia plateau the initial Col II increase in gene expression is followed by a decrease leading to the lowest over all Col II expression on the medial plateau, here especially in the central part. There in late stage diseases the collagen type I expression was also more pronounced. Markedly decreased safranin O staining intensity was observed in the radial zone and less reduced intensity in the transitional zone with loss of zonal anatomy in 40% of the specimens in group A and all specimens in group B. Correlation between colorometrically analysed proteoglycan GAG content and aggrecan Real Time PCR is mainly weak.Tibial and femoral cartilage in contrast to patellar cartilage both are preferential exposed to compressive stresses, but presence of menisci affects the load distribution at the tibial side, which creates varying conditions for the different cartilage surfaces in the knee.As directly measured Poissońs ratio in tibial cartilage is higher but Youn?s modulus is lower than in femoral cartilage, different resulting feedback amplification loops interact with proceeding cartilage damage. The initial loss of aggrecan may support Matrix metalloproteinases (Mmps) in the access to the collagen network and the considerably differing mechanical properties at both joint surfaces result in varying increased synthesis and release of matrix degrading enzymes.The present study has identified a selection of events which reflect the response of cartilage structure and composite, chondrocytes itself and their productivity to changes in mechanical stress depending on the anatomical site.     

The measurement of negative charge content in cartilage using a colloid titration technique.

A colloid titration technique has been used to determine the sulfate and carboxylate content of various glycosaminoglycans and has been validated by comparing the results with data obtained using well-established techniques. The method has been applied to the measurement of the negative charge content of cartilage slices at various depths from the articular surface and to the determination of sulfate and carboxylate contents in bovine nasal septa. Titrations of nasal septa were performed on milled cartilage, on cartilage digested with papain and on proteoglycans purified by cesium chloride gradient centrifugation of guanidinium chloride extracts. The sulfate content was similar for all three preparations (0.5 mu eq per milligram dry cartilage). However, the carboxylate content determined on milled cartilage was 40% higher than that obtained for cartilage digested with papain or for purified proteoglycans; this implies the possible contribution of carboxyl groups from structural glycoproteins present in the extracellular matrix. The carboxylate content determined on purified proteoglycans was in excellent agreement with values calculated from chemical analyses.     

Structural changes of hip osteoarthritis using magnetic resonance imaging

Introduction

Few data are available concerning structural changes at the hip observed by magnetic resonance imaging (MRI) in people with or without hip osteoarthritis (OA). The aim of this study was to compare cartilage volume and the presence of cartilage defects and bone marrow lesions (BMLs) in participants with and without diagnosed hip OA.

Methods

Femoral head cartilage volume was measured by MRI for 141 community-based persons with no diagnosed hip OA, and 19 with diagnosed hip OA. Cartilage defects and BMLs were regionally scored at the femoral head and acetabulum.

Results

Compared with those without diagnosed hip OA, people with diagnosed hip OA had less femoral head cartilage volume (1763 mm³ versus 3343 mm³; p <0.001) and more prevalent cartilage defects and BMLs (all p ≤0.05) at all sites other than the central inferomedial region of the femoral head. In those with no diagnosed hip OA, cartilage defects in the anterior and central superolateral region of the femoral head were associated with reduced femoral head cartilage volume (all p ≤0.02). Central superolateral BMLs at all sites were associated with reduced femoral head cartilage volume (all p ≤0.003), with a similar trend occurring when BMLs were located in the anterior region of the hip (all p ≤0.08).

Conclusions

Compared with community-based adults with no diagnosed hip OA, people with diagnosed hip OA have less femoral head cartilage volume and a higher prevalence of cartilage defects and BMLs. For people with no diagnosed hip OA, femoral head cartilage volume was reduced where cartilage defects and/or BMLs were present in the anterior and central superolateral regions of the hip joint. Cartilage defects and BMLs present in the anterior and central superolateral regions may represent early structural damage in the pathogenesis of hip OA.     

The restoration of full-thickness cartilage defects with mesenchymal stem cells (MSCs) loaded and cross-linked bilayer collagen scaffolds on rabbit model

Cartilage has a limited self-repair capability and the repair of large cartilage defects remains a challenge in clinic. This study aimed to investigate the effect of mesenchymal stem cells (MSCs) loaded three-dimensional bilayer collagen scaffold for cartilage repair. Cross-linked three-dimensional bilayer collagen scaffolds seeded with or without MSCs were implanted into large cartilage defects (4 mm in diameter; 3 mm in depth) in rabbit knees. The untreated cartilage defects served as control. The tissue response was evaluated at 6 and 12 weeks after implantation by general histology and semi-quantitative histological grading systems. In addition, the repaired tissues were evaluated by mechanical test at 12 weeks after implantation. The MSCs-loaded collagen scaffold group showed the most hyaline cartilage, highest histological scores and compressive modulus. Moreover, it showed a good integration with the subchondral bone and adjacent cartilage. The structure of the novel bilayer collagen scaffolds provided architectural support for the differentiation of MSCs and demonstrated successful induction of in vivo chondrogenesis. These findings suggested that MSCs-loaded bilayer collagen scaffold could be an appealing candidate to be used for cartilage regeneration.     

On the influence of mechanical conditions in osteochondral defect healing

Despite the introduction of new surgical techniques, the treatment of cartilage defects remains challenging. Delay or complete failure of cartilage healing is associated with problems in biological regeneration. The influence of mechanical conditions on this process, however, remains unevaluated. Osteochondral defects were generated on the left femoral condyle in 18 Yucatan minipigs. After 4, 6 and 12 weeks the defect filling, trabecular orientation and bone density were compared to the intact contralateral side. The mechanical straining during this period was then analyzed using an adaptive finite element technique. Histologically, the osteochondral defects showed bone resorption at the base and bone formation from the circumference. At 12 weeks, the macroscopically healed specimens showed fibrous cartilage formation, a minimally organized trabecular structure and increased trabecular volume fraction compared to the controls (p < 0.002). The amount of cancellous, cartilagineous, and fibrous tissue and the defect size as measured in histomorphometric analysis for the three time points (4, 6 and 12 weeks) was comparable in magnitude to that predicted by finite element analysis. The simulated osteochondral healing process was not fully capable of re-establishing a hyaline-like cartilage layer. The correlation between simulation and histology allows identification of mechanical factors that appear to have a larger impact on the healing of osteochondral defects than previously considered.     

Effect of acetabular labral tears,repair and resection on hip cartilage strain: A 7 T MR study

BackgroundTears of the acetabular labrum are frequently present in patients with groin pain. While it is clear that the labrum contributes to the surface area articulating with the femoral head, it is not clear whether labral repair yields different load distribution in the hip compared to labral resection.PurposeDetermine whether labral repair reduces cartilage strain more effectively than labral resection.MethodsSix human cadaveric hips (mean age 37 years) were loaded in a simulated single-leg stance within the bore of a 7 T MR scanner. After cartilage had reached a steady-state thickness distribution, a scan of the cartilage was acquired with a voxel size of 0.1×0.1×0.3 mm. This method was repeated for each of six specimens when the labrum was intact, after a surgically simulated labral tear, after an arthroscopic labral repair and after labral resection. Cartilage thickness and strain in an anterosuperior region of interest were measured from the MR scans. A paired t-test was used to compare mean and maximum cartilage strain when the labrum was intact vs. torn, torn vs. repaired and repaired vs. resected. Three-dimensional patterns of cartilage strain distribution were qualitatively compared for the different labral conditions.ResultsFor the number of specimens tested we found no change in mean and maximum cartilage strain, and little obvious change in the pattern of cartilage strain distribution after a simulated labral tear. Labral repair caused a 2% decrease in mean cartilage strain compared to a torn labrum (p=0.014). Labral resection caused a 4% and 6% increase in mean and maximum cartilage strain, respectively, compared to labral repair (p=0.02), and the cartilage strain distribution was elevated throughout the region of interest.ConclusionBased on our ex vivo findings of increased cartilage strain after labral resection when compared to labral repair, we have demonstrated the associated consequences to the mechanical environment of the cartilage following surgical treatment of the labrum.     

Measurement of the strongly held water of lysozyme by drying

A method for the measurement of water that is strongly held by lysozyme is described. This water is slowly removed by vaccum drying of lyophilized and can be titrated with the Karl Fischer reagent. Drying curves were obtained by mechanical pumping (moderate vacuum) and diffusion pumping (high vacuum) at 20°, 10°, 0°, ?10° and ?20°C. About 23 water molecules per lysozyme molecule are at least moderately held by the protein. These water molecules fall into several classes. Three to four of them are quite strongly held and may correspond to the three buried water molecules observed in the x-ray analysis of lysozyme structure. The presence of tri-N-acetylglycosamine in the lysozyme active cleft has no effect on drying at 0°C. The method shows promise of being generally applicable to the measurement of small amounts of water which are strongly held by biological structures.     

Hydrogen bonding between sugar and protein is responsible for inhibition of dehydration-induced protein unfolding.

The nature of the interaction responsible for the inhibition of protein unfolding and subsequent damage by sugars during dehydration is unclear. The relationship between sample moisture content measured by coulometric Karl Fischer titration and the apparent moisture content predicted by the area of the protein side chain carboxylate band at approximately 1580 cm-1 in infrared spectra of dried protein-sugar samples was examined. For samples in which a high level of native protein structure was retained in the dried solid, the apparent moisture content predicted by the carboxylate band area was greater than the actual moisture content, indicating that protection results from direct sugar-protein hydrogen bonding and not entrapment of water at the protein surface. Further, we show that the degree of structural protection conferred by sucrose and trehalose apparent in second derivative, amide I infrared spectra, correlates with the extent of hydrogen bonding between sugar and protein. The failure of dextran to inhibit dehydration-induced lysozyme unfolding is shown to result from the inability of the polymer to hydrogen bond adequately to the protein. Therefore, formation of an amorphous phase alone is not sufficient to maintain protein structure during dehydration. Glucose hydrogen bonds to a high degree with dried lysozyme, but is incapable of inhibiting lyophilization-induced protein unfolding in the absence of an effective cryoprotectant. However, the addition of polyethylene glycol, which is known to protect proteins during freezing, but not drying, to glucose protected lysozyme structure during lyophilization. Together, these results show that hydrogen bonding between carbohydrate and protein is necessary to prevent dehydration-induced protein damage. However, hydrogen bonding alone is not sufficient to protect proteins during lyophilization in the absence of adequate freezing protection.     

微载体技术在软骨损伤修复中的应用与展望北大核心CSCD

软骨内部无血管结构、细胞外基质含量高的特点,使软骨组织的自我恢复能力很差。在临床治疗中,轻度的软骨缺损通常采用物理治疗或药物治疗方式,严重者需进行手术治疗。近年来,软骨组织工程技术为治疗软骨缺损提供了新的思路,与传统的手术治疗方式相比,结合软骨组织工程技术进行治疗具有创口小、恢复佳的优点。将微载体技术融入组织工程支架的设计中,可以利用微载体直径小、能够负载多种生长因子的特点,进一步扩展支架功能、促进软骨组织再生。文中首先对微载体技术进行介绍,对近年来微载体的主要制备方式和创新内容进行了概括总结,作为后续介绍的基础内容。然后对应用于软骨修复中的微载体进行了材料和功能上的划分,介绍了不同材料、不同功能微载体的属性特征和在软骨修复方面的具体应用,最后结合该领域发展历程对其今后发展趋势及方向进行展望,并基于笔者团队关于骨软骨一体化层状支架的研究,提出了通过微载体优化层状支架性能的思路,有望制备出更贴合天然软骨结构特征的仿生支架。     

The effects of proteolytic enzymes on the mechanical properties of adult human articular cartilage.

The effects of the lysosomal proteinase cathepsin D on the mechanical properties of adult human articular cartilage were examined in detail in 7 joints within the age range 21 to 72 years. The results of a preliminary study on the effects of the lysosomal proteinase cathepsin B1 and clostridial collagenase on the mechanical properties of cartilage are also presented. Cartilage which had been incubated with either cathepsin D or cathepsin B1 showed increased deformation in uniaxial compression perpendicular to the articular surface. The enzyme-treated cartilage also showed decreased tensile stiffness at low values of stress. This effect was more pronounced in specimens from the deeper zone of cartilage than in specimens from the superficial zone. It was also more pronounced in specimens which were aligned perpendicular to the predominant alignment of the collagen fibres in the superficial zone than in specimens which were parallel to the collagen fibres. At higher stresses the tensile stiffness of the treated cartilage was not significantly different from that of the untreated tissue. The tensile fracture stress of the cartilage was also not significantly reduced by the action of cathepsin D. In contrast to the effects observed with the cathepsins, the preliminary results obtained by incubating cartilage for 24 h with clostridial collagenase showed that both the tensile stiffness and the fracture stress were considerably lower than the corresponding values for the untreated tissue. Biochemical analysis of the incubation media, and the specimens, revealed that a large proportion of the proteoglycans was released from the cartilage by each of the three enzymes. The proportion of the total collagen which was released from the cartilage was different for each enzyme: cathepsin D released between 0 and 1.5 per cent, cathepsin B1 released between 2.3 and 4.3 per cent and collagenase released between 5.3 and 27.8 per cent of the collagen after 24 h.