Anat Embryol (2001) 203:147173

Springer-Verlag 2001

R E V I E W A RT I C L E

Felix Eckstein Maximilian Reiser Karl-Hans Englmeier Reinhard Putz

In vivo morphometry and functional analysis of human articular cartilage with quantitative magnetic resonance imaging from image to data, from data to theory
Accepted: 6 December

Abstract Analyses of form-function relationships and disease processes in human articular cartilage necessitate in vivo assessment of cartilage morphology and deformational behavior. MR imaging and advanced digital post-processing techniques have opened novel possibilities for quantitative analysis of cartilage morphology, structure, and function in health and disease. This article reviews work on three-dimensional post-processing of MR image data of articular cartilage, summarizing studies on the accuracy and precision of quantitative analyses in human joints. It presents normative values on cartilage volume, thickness, and joint surface areas in the human knee, and describes the correlation between different joints and joint surfaces as well as their association with gender, body dimensions, and age. The article summarizes ongoing work on functional adaptation of articular cartilage to mechanical loading, analyses of in situ cartilage deformation in intact joints in vivo and in vitro, and the quantitative evaluation of cartilage tissue loss in osteoarthritis. We describe evolving techniques for assessment of the structural/biochemical composition of articular cartilage, and discuss future perspectives of quantitative cartilage imaging in the context of joint mechanics, mechano-adaptation, epidemiology, and osteoarthritis research. Specifically, we show that fat-suppressed gradient echo sequences permit valid analysis of cartilage morphology, both in healthy and severely osF. Eckstein () R. Putz Muskuloskeletal Research Group, Anatomische Anstalt, Ludwig-Maximilians-Universitt Mnchen, Pettenkoferstrasse 11, 80336 Mnchen, Germany e-mail: eckstein@anat.med.uni-muenchen.de Tel.: +49-89-5160 4847, Fax: +49-89-5160 4802 M. Reiser Institut fr Klinische Radiologie, Klinikum der Ludwig-Maximilian Universitt Mnchen, Grohadern, 81377 Mnchen, Germany K.-H. Englmeier Institut fr Medizinische Informatik und Systemforschung, GSF-Forschungszentrum fr Umwelt und Gesundheit Neuherberg, 85764 Oberschleiheim, Germany

teoarthritic joints, as well as highly reproducible measurements (CV%=1 to 3% in the knee, and 2 to 10% in the ankle). Relatively small differences in cartilage morphology exist between both limbs of the same person (~5%), but large differences between individuals (CV% ~20%). Men display only slightly thicker cartilage then women (~10%), but significantly larger joint surface areas (~25%), even when accounting for differences in body weight and height. Weight and height represent relatively poor predictors of cartilage thickness (r2 <15%), but muscle cross section areas display more promising correlations (r2 >40%). The level of physical exercise (sportive activity) does not account for interindividual differences in cartilage thickness. The thickness appears to decrease slightly in the elderly in particular in women, even in the absence of osteoarthritic cartilage lesions. Strenuous physical exercises (e.g., knee bends) cause a 6% patellar cartilage deformation in young individuals, but significantly less deformation in elderly men and women (<3%). The time required for full recovery after exercise (fluid flow back into the matrix) is relatively long (~90 min). Static in situ compression of femoropatellar cartilage with 150% body weight produces large deformations after 4 h (~30% volume change), but only very little deformation during the first minutes of loading. Quantitative analyses of magnetization transfer and proton density hold promise for biochemical evaluation of articular cartilage, and are shown to be related to the deformational behavior of the cartilage. Application of these techniques to larger cohorts of patients in epidemiological and clinical studies will establish the role of quantitative cartilage imaging not only in basic research on form-function relationships of articular cartilage, but also in clinical research and management of osteoarthritis. Key words Morphometry Articular cartilage Magnetic resonance imaging Osteoarthritis

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Background, technique development, and validation

Articular cartilage a complex buffer with unique properties Synovial joints allow for locomotion, for orientation of the body relative to its surroundings, and for interaction with the environment. However, their function is not only to permit movement between segments of the body, but to reduce bending stresses and to guarantee a mechanically efficient distribution of tissue in the skeleton (Pauwels 1965, 1980). Muscles display smaller lever arms than forces generated by the mass of the body or from outside; joints therefore encounter forces of several times the body weight during normal activity. Measurements with telemetric hip endoprostheses (Bergmann et al. 1993) have confirmed that during gait forces of approximately 300% body weight are generated in the hip, and these can rise up to 800% for exceptional activities. Articular cartilage is the tissue that is capable of transferring these loads from one skeletal element to the other, and of providing almost frictionless gliding of the joint bodies during loading. Cartilage displays unique morphological and mechanical properties, and it has so far not been possible to produce an artificial material with comparable load-bearing capacity. The tissue has no metabolic functions except for the maintenance of its mechanical properties; these properties are not directly determined by cells, but by the interstitial matrix. The matrix consists of approximately 70% fluid and 30% structural elements, the proteoglycans (PGs) and collagens being the main components. Triple helical type II collagen molecules condense into fibrils to give cartilage its tensile strength (Akizuki et al. 1986). These represent heterotypic alloys with collagen XI in their center (Blaschke et al. 2000). Collagen XI and several PGs (e.g., fibromodulin) have been suggested to control the process of fibril formation, leading to fibrils with a relatively uniform diameter of 20 nm. Collagen IX represents a surface coating of collagen II, likely providing bridges between fibrils and with other matrix components (Olsen 1997). Collagen II molecules are cross-linked through pyridinoline. A number of other cartilage components (COMP=cartilage oligomeric matrix protein, Decorin, and others) have been suggested to link collagen and the PGs (Rosenberg et al. 1998). The latter, and in particular aggrecan with its highly charged chondroitin-sulfate and keratan-sulfate side chains, have the capacity to bind a vast amount of cations and water molecules, and produce an endoosmotic swelling pressure that puts the collagen matrix under substantial tensile stress (Maroudas 1976). The matrix composition varies substantially throughout the depth of the tissue and between different joints (Buckwalter and Mankin 1997a; Huch et al. 1997; Mow and Ratcliffe 1997). In mature cartilage, this biochemical heterogeneity is accompanied by structural variation, collagen fibrils forming a complex three-dimensional network (Jeffery et al. 1991; Buckwalter and Mankin

1997a). These fibrils display a preferential tangential orientation in the surface (tangential) layer, a radial orientation in the deeper (radial) zone, and a relatively random orientation inbetween (transitional zone). In the radial zone, tightly packed collagenous elements with a longitudinal as well as helical arangement form tubular structures with an internal diameter of 12 m (Ap Gwynn et al. 2000). The lumen of the tubules is lined by circumferential 10-nm fibres, spaced at 5070 nm intervals. The tangential zone is composed of a spongy arrangement of collagen fibrils, containing bunches of small (<1 m) tangentially orientated tubules (Ap Gwynn et al. 2000). This zone is rich in a mucin-like glycoprotein, known as superficial zone protein (Flannery et al. 1999) or lubricin (Jay et al. 2000). This protein provides cartilage with a friction coefficient that is almost zero, i.e., lower than that of ice gliding on ice (Mow and Ratcliffe 1997; Jay et al. 2000). Mechanically, cartilage does not behave like an elastic tissue, the relation between load and deformation (stress and strain) being non-linear and time-dependent. Although there is a linear relationship between the stress and strain in the instantaneous response to compression and in equilibrium (with very different ratios, however), this is not the case for the deformational state in between these two idealized situations (Fig. 1). The linear biphasic theory regards the tissue as a composite of a porous "solid" matrix on the one hand and a fluid phase on the other (Mow et al. 1984, 1993; Mow and Ratcliffe 1997). This theory has been very successful in describing the fundamental mechanical behavior of cartilage. Loading of the cartilage leads to instantaneous hydrostatic pressurization in the tissue, with relatively little deformation (Fig. 1). It has been shown in theory and experiments that the applied load is supported initially by the fluid phase, shielding the solid collagen-PG matrix from excessive strain (Ateshian et al. 1994; Ateshian and Wang 1995; Wu et al. 1996; Soltz and Ateshian 1998, 2000; Herberhold et al. 1999). This explains why cartilage can tolerate high loading peaks without suffering damage. When loading continues, the fluid starts to stream through the porous matrix, which, however, displays relatively low permeability (Fig. 1). Step by step, the load is then transferred from the fluid to the solid phase. Finally, the hydrostatic pressure reaches zero, deformation ceases (equilibrium), and the total load is supported by the solid matrix (Fig. 1). Since loading events are generally of short duration under physiological conditions, and since hydrostatic pressurization is maintained over relatively long periods of time (Ateshian et al. 1994; Wu et al. 1996), the matrix appears to encounter relatively little strain during normal joint loading. It must, however, be kept in mind that cartilage displays very complex properties, such as tension-compression nonlinearity (Akizuki et al. 1986), anisotropy, spatial inhomogeneity, etc. (Mow and Ratcliffe 1997), which make it difficult to comprehensively describe its mechanical behavior. This has lead to extensions of the linear biphasic theory (e.g., Ateshian et al. 1997; Donzelli et al. 1999), but these the-

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Fig. 1 Simplified model demonstrating the biphasic properties and load partitioning of the cartilage between the fluid and solid phase during loading. The solid phase (proteogycan-collagen matrix) is symbolized by an elastic spring within a tube, the interstitial fluid phase by circles within the tube. During loading, fluid can only escape through the small pore at the top of the tube: Instantaneous response of the cartilage There is no fluid flow and no solid matrix stress, the entire load is supported by hydrostatic pressurization of the interstitial fluid (red circles). Intermediate phase Fluid streams through the porous matrix (symbolized by a hole at the top of the tube); the load is shared between the fluid and solid phase. Equilibrium (steady state) Fluid flow ceases, the hydrostatic pressure is zero, the entire load is supported by the solid phase (symbolized by the red spring within the tube)

ories still represent considerable simplifications of the unique mechanical properties of articular cartilage. Cartilage failure the irreversible pathway Osteoarthritis (OA) is the most prevalent chronic disease in the elderly, affecting more than 50% of those 65 years and older (Peyron 1986; Felson 1988; 1990; Felson et al. 1995). It causes pain and functional deficits, with substantial effects on the quality of life (Guccione et al. 1994). Joint disease has resulted in costs of more than $ 65 billion per year in the US in 1992 (Yelin and Callaghan 1995), and the direct and indirect costs have been currently estimated to approach 1% of the gross national product of the US (Yelin 1998). With the expected aging of the population, this socio-economic burden will increase considerably over the next decades. The magnitude of the problem is highlighted by the official declaration of the years 2000 to 2010 as the Bone and Joint

Decade (http://www.bonejointdecade.org/). This initiative receives support by the WHO, the UN, and other institutions. Its goal is to raise awareness of the growing burden of musculoskeletal disorders on society, and to advance understanding through research on prevention and treatment. Although there is certain evidence that genetic (Cicuttini and Spector 1996; 1997) and mechanical (Radin et al. 1991; Sharma et al. 2000) factors play a pivotal role in the evolution of OA, yet it is still unclear which specific genes or factors are involved in the initiation and progression of cartilage damage. It is also not understood why some joints are more often affected than others, or why cartilage degenerates relatively early in life in some individuals, whereas in others it maintains normal morphology and function over eighty years and more. Most likely, OA is a multifactorial disease and constitutes a common final pathway (Radin et al. 1991) rather than one specific process. Nevertheless, some common features have been identified: after an initial insult, the cell metabolism and matrix turnover is stimulated in an attempt to maintain matrix integrity. If this process fails, the matrix undergoes intrinsic biochemical alterations (loss of PGs and rise in water content). Later structural changes occur (cleavage of collagens) that are associated with a release of degradation products (biochemical markers). The enzymes involved in collagen and PG degradation are currently identified, as well as ways to pharmacologically block associated processes (e.g., Mitchell et al. 1996; Lark et al. 1997; Arner et al. 1998, 1999; Ishigura et al. 1999; Dahlberg et al. 2000). Continuous matrix degradation leads to a deterioration of mechanical properties, initiating a vicious circle in which

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(normal) mechanical loading can lead to elevated mechanical strain and further insult to the matrix. This is particularly problematic when the surface layer is affected. The surface zone has been shown to regulate the fluid flow between the cartilage and the joint space, and to guarantee adequate hydrostatic pressurization during loading (Setton et al. 1993). If its permeability increases, a higher proportion of the load must be supported by the matrix; progressive insult then leads to macro-morphological changes of the cartilage (fissures, small lesions, or uniform cartilage thinning). At this stage, there is a net loss of tissue (volume) until the cartilage is eroded. Although it is unclear at which stage the subchondral bone starts to show associated changes, conventional radiography displays visible alterations (e.g., cyst formation or sclerosis) only in the later stages of this process. Articular cartilage is known to display only limited capacity for regeneration and repair, and the quality of repair tissue is clearly inferior to that of normal cartilage. Conventional approaches for treating OA range from conservative measures (administration of anti-inflammatory and analgesic drugs, physiotherapy, braces) to surgical intervention (debridement, anterograde drilling of subchondral bone, correction of malalignment) and eventually joint replacement (Buckwalter and Mankin 1997b). Currently, there is no curative treatment for the disease, and available drugs have failed to demonstrate "structure-modifying" activity. Although OA is not inevitably progressive, it must be considered an irreversible pathway. There is presently only limited potential, therefore, to alter its natural course. However, novel therapeutic strategies are emerging with potential for structure modification; these include inhibitors of metalloproteinases and their upstream regulators, transplants (homologous, autologous, and cell seeding), perichondral grafting, microfracturing of subchondral bone, use of artificial matrices, growth factors, gene therapy, and others (Buckwalter et al. 1997b). These approaches have stimulated hope for more effective treatment of OA. A major reason for the current lack of knowledge regarding form-function relationships of normal and osteoarthritic human articular cartilage is that analyses of its morphology and mechanical properties have been essentially confined to post-mortem specimens. With the advent of non-invasive in vivo techniques for evaluating cartilage, it will, however, become feasible to collect systematic data at different stages of life and with detailed individual histories. It will become possible to follow individuals longitudinally while systematically changing the (mechanical) environment, to screen groups that are at high risk of sustaining joint disorders (e.g., patients with joint instability), and to evaluate the efficacy of novel therapeutic compounds. Since clinical symptoms correlate poorly with tissue loss in OA, objective measures are needed to determine disease stage and rate of progression as well as the appropriate time point and type of therapeutic intervention.

Conventional methods of cartilage imaging fishing in the dark The methods currently available for evaluating cartilage include arthroscopy, biochemical markers, radiography, and magnetic resonance imaging (MRI). Diagnostic arthroscopy is considered a gold standard for evaluating surface alterations, but it has the disadvantage that it is invasive. The examination provides, however, only little information on cartilage properties or its alterations through the depth. Biochemical markers cannot give specific information on certain joints when being obtained from the serum or urine. Joint aspiration, on the other hand, causes pain and suffers from limited compliance. Conventional radiography can provide indirect information about cartilage destruction from signs such as joint space narrowing, subchondral sklerosis or cysts, or the presence of osteophytes. However, it is a two-dimensional technique that is sensitve to artifacts resulting from malpositioning (Buckland-Wright et al. 1995a) or meniscal subluxation (Gale et al. 1999). Several authors have reported little agreement between standard radiography and the actual cartilage thickness and/or state of the articular surface in arthroscopy (Fife et al. 1991; Brandt et al. 1991; Karvonen et al. 1994). For reproducible measurements of the joint space width, radiographs must be obtained in standing semiflexed position, preferably with fluroscopic control (Buckland-Wright et al. 1995a, 1999). This technique has been shown to produce relatively accurate data in the medial (but not in the lateral) femorotibial compartment of the knee of OA patients (Buckland-Wright et al. 1995b). However, radiography is unable to differentiate between femoral and tibial cartilage loss, is less sensitive to focal than to general cartilage lesions, and cannot demonstrate the pattern of cartilage destruction throughout the joint surface. Due to these limitations, measurements of the joint space width in radiographs are not ideal for reliably evaluating cartilage thickness and surface alterations. Magnetic resonance imaging the protons make the difference MRI uses a strong magnetic field and high frequency radiowaves (rather than ionizing radiation) for obtaining sectional images. The technique has so far been shown to have no adverse effects on health. Other important advantages are its multiplanar capabilities and its superior soft tissue contrast (Peterfy and Genant 1996; Stbler et al. 2000; Peterfy 2000). In MRI, the tissue contrast can be substantially modulated by choosing different types of pulse sequences, and by changing the specific parameters of these sequences (repetition time, echo time, flip angle, etc.). Hence, a variety of specific sequences can be selected for optimal delineation of specific tissues, or even for specific aspects of these tissues. However, cartilage presents an imaging challenge due to its short transverse relaxation time (T2) and to various sources of artifact, in particular at the bone interface.

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Several MR sequences have been advocated for qualitatively evaluating cartilage and joints in general (Recht and Resnick 1994, 1998; Peterfy and Genant 1996; Stbler et al. 2000; Peterfy 2000). For diagnostic purposes, proton density weighted and T2-weighted spinecho sequences with fat-suppression have been recommended as well as fat-suppressed gradient echo images. These allow to detect thinning of the cartilage, focal lesions, signal alterations, and osteochondral defects (Bredella et al. 1999; Stbler et al. 2000). Qualitative scoring systems have been developed for clinical evaluation of the disease status in synovial joints (Peterfy and Genant 1996; Peterfy 2000), but these depend on the experience of the reader and may therefore be less reliable in longitudinal studies. In particular, qualitative scores are only of limited usefulness in a scientific context when addressing fundamental questions concerning the morphology and functional adaptation of cartilage tissue statistically. For these reasons, there has been increasing interest in deriving quantitative parameters from MR image data. For the analysis of cartilage macro-morphology (volume, thickness, surface areas) the bone cartilage interface and the articular surface need to be delineated accurately. In particular, the spatial resolution must be sufficient to permit quantitative measurements throughout its thickness. For these reasons, a high-resolution pulse sequence is required that visualizes cartilage with high contrast to its surrounding tissues. Some investigators have used two different pulse sequences and digital subtraction techniques to improve contrast (Robson et al. 1995; Mnsterer et al. 1996). However, today it is widely accepted that T1-weighted gradient echo sequences with spectral fat suppression are best suited for this purpose (Recht et al. 1993). These sequences produce images in which the cartilage appears bright (hyperintense) compared to all other tissues, and in which the chemical shift artifact at the bone cartilage interface is eliminated. Spectral fat-suppression techniques require high field systems (>0.5 T). Fat-suppression is achieved by applying a prepulse, preventing the fat-bound protons from creating a signal during the subsequent data acquisition (Recht et al. 1993). However, sequences with selective excitation of only the water-bound protons have been introduced recently, in which fat-signal elimination can be obtained with much shorter acquisition times (Hardy et al. 1998; Hyhlik Drr et al. 2000; Graichen et al. 2000a; Glaser et al. 2000). This technique permits to acquire a data set of the human knee joint (about 60 slices of 1.5-mm thickness) at an in-plane resolution of 0.30.3 mm3 (about 16 million voxels) in less than 10 min. (Fig. 2a). In this context one must be aware that in MRI an improvement in resolution by a factor of 2 (in all 3 dimensions) requires a prolongation of the image time by a factor of 64 (n6), if an identical contrastto-noise ratio is to be achieved. Water-excitation sequences therefore also have the advantage that the spatial resolution can be improved without requiring excessive imaging time. In the elbow and foot, for instance, it

has become possible to obtain image data at a resolution of 10.250.25 mm3 with acquisition times that are tolerable in vivo (<20 min). Another potentially valuable technique for quantitative cartilage imaging is driven equilibrium Fourier transform (DEFT; Hargreaves et al. 1999). This method enhances signal intensity without waiting for full T1 recovery and provides a good combination of bright cartilage and high contrast with surrounding tissue. The accuracy and precision of quantitative assessment of cartilage morphology, however, remain to be established. Three-dimensional post-processing from image to data General considerations Based on sectional images alone, the comparison between individuals or the longitudinal study of tissue alterations are unreliable, because corresponding section locations and orientations cannot be reproduced. Therefore, three-dimensional (3D) digital post-processing techniques are advantageous, since they can provide objective measurements, independent of the specific section location and orientation. Moreover, quantitative evaluation should always include the entire joint surface and should not rely on single images. The image-processing steps required for these analyses include the segmentation of the cartilage, 3D analysis (including 3D reconstruction and 3D thickness computations), and potentially 3D registration (matching). Segmentation Segmentation is the process by which appropriate image points (voxels) are assigned to a specific anatomic structure, such as a cartilage plate (Fig. 2a). MRI provides insufficient contrast for fully automated segmentation of articular cartilage based on the gray value distribution alone. Volume-growing algorithms (Eckstein et al. 1996; Piplani et al. 1996) are sensitive to irregularities at the cartilage surface and often fail in regions where contrast is low. We have therefore developed a B-spline Snake (deformable contour) algorithm that relies on the interaction of image forces (gray value gradients), model forces (stiffness of a parameterized B-spline curve), and coupling forces (segmentation of previous sections). This approach can accelerate the interactive segmentation process and increases consistency between observers (Stammberger et al. 1999a). Other groups have employed different algorithms, such as active shape models (Solloway et al. 1997), edge detection (Robson et al. 1995; Kshirsagar et al. 1998), fitting of B-spline curves to ma-nually digitized points (Cohen et al. 1999), immersion based watershed segmentation (Ghosh et al. 2000), and live-wire algorithm (Steines et al. 2000). None of these approaches have, however, succeeded in fully automated segmentation of articular

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Fig. 2ad Methods for quantitative 3D analysis of cartilage morphology from MR imaging. a Sagittal MR image of human knee obtained with a fat-suppressed (water-excitation) gradient echo sequence; segmentation of femoral cartilage with B-spline Snake algorithm (Stammberger et al. 1999a). b 3D volume reconstruction of the femoral cartilage. c Analysis of joint surface area by a triangulation technique (Eckstein et al. 2001a). d Computation of 3D thickness distribution, independent of section orientation (Stammberger et al. 1999b)

3D analysis the cartilage volume and beyond The initial approach of quantifying articular cartilage has been to determine its volume (Pilch et al. 1994; Peterfy et al. 1994; Eckstein et al. 1994). This is achieved by numerical integration (multiplication of the voxel number assigned to a cartilage plate with the voxel size; Fig. 2b). The measurement of cartilage volume has the advantage that it can be used as a direct parameter of cartilage growth, adaptation, and tissue loss in OA. However, it lacks focal, region-specific information on the quantitative distribution of the tissue within a joint surface, and it is determined by both the cartilage thickness as well as the size of the joint surface. In longitudinal studies (in which the joint surface can be assumed to be constant) changes in volume directly reflect changes in thickness, but for some applications it is necessary to clearly separate these two parameters.

cartilage, and post-processing for an entire human knee joint is generally in the range of several hours. Further improvements may be achieved by more sophisticated model-based approaches. These improvements are important for implementation of the technique in the (routine) clinical assessment of OA patients.

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The size of the joint surface area can be determined by triangulation (Eckstein et al. 2001a; Fig. 2c). Since the angle between the MR images and the articular surface varies throughout the joint surface and from acquisition to acquisition, the computation of cartilage thickness must take into account the local out-of-plane deviations of the distance vectors. This is achieved by computing normal vectors of the bone-cartilage interface (Cohen et al. 1999) or minimal distances between the articular surface and the interface (Lsch et al. 1997). Both implementations are mathematically identical, but can lead to slightly different results when being applied to discrete voxel objects (Lsch et al. 1997). Stammberger et al. (1999b) developed a computational method based on 3D Euclidean distance transformation that avoids the explicit calculation of surface normals. It provides robust results around cartilage edges and lesions and can visualize the cartilage thickness distribution throughout cartilage plates (Fig. 2d). The thickness values are determined from approximately 1,000 computations per square centimeter. Comparison of thickness computations from sectional images with different spatial orientation have produced consistent results, showing that the algorithms works independent of the specific section orientation (Eckstein et al. 2000a). 3D registration (matching) In order to depict local/regional cartilage thickness changes directly, Stammberger et al. (2000) developed a 3D matching algorithm, in which the bone-cartilage interfaces of two data sets are registered. First, a principal axis decomposition is used to align the surfaces. In a second step, corresponding image points are identified by elastically deforming the surfaces based on local geometric similarity measures (Euclidean distance and orientation compatibility). The method can be used, for instance, to depict the spatial pattern of cartilage deformation during compression (Fig. 3). Kshirsagar et al. (1998) developed a rigid 3D registration algorithm and observed a higher precision of localized cartilage volume versus total cartilage volume measurements (2.0% vs. 3.8%). These methods are particularly useful in longitudinal studies that aim to monitor local changes of thickness, for instance in epidemiological studies on disease progression, or in animal models of OA. Validation and precision can we really trust it? Knee joint Most quantitative MRI analyses of human cartilage have been performed in the knee, because it displays the largest cartilage thickness and is the site of earliest cartilage degeneration. Numerous sources of artifacts can occur in MRI (partial volume-, chemical shift-, susceptibility-, truncation-, motion-artifacts, etc.), requiring thorough
Fig. 3ac Analysis of changes in regional cartilage thickness during compression with the 3D matching algorithm (Stammberger et al. 2000). a Patellar cartilage thickness before compression. b Patellar cartilage thickness after compression with 150% body weight for 4 h (color legend for a and b: dark blue 00.6 mm, light blue 0.61.2 mm, then 0.6 mm intervals up to gray >4.2 mm). c Direct visualization of local thickness changes by 3D matching (color legend for c: black no deformation, violet 00.4 mm, then 0.4 mm intervals up to dark blue >2.4 mm)

154 Fig. 4 Determination of cartilage morphology in severe osteoarthritis (prior to knee joint arthroplasty): a reconstruction showing incomplete cartilage cover (green) on the proximal tibia (red); b Correlation between in vivo cartilage volume measurements with MRI and water displacement of surgically removed tissue after arthroplasty (adapted from Burgkart et al. 2000)

validation of quantitative measurements. Since these artifacts do not only depend on the equipment used, but on the specific composition of the tissue under investigation, validation studies must be made directly on the biological object of interest. Knee-joint cartilage volume measurements (obtained with T1-weighted, fat-suppressed gradient echo sequences) have been shown to deviate not more than 5 to 10% on average from water displacement of surgically retrieved tissue (Peterfy et al. 1994; Dupuy et al. 1996; Piplani et al. 1996; Cicuttini et al. 1999); anatomical sectioning (Sittek et al. 1996; Eckstein et al. 1996), and CT arthrography (Eckstein et al. 1998a, 2000a). We have re-

cently compared in vivo (pre-operative) MRI data of tibial cartilage (obtained with a water excitation sequence) to volume displacement of surgically removed cartilage tissue of the resected tibial plateau after total knee replacement (Burgkart et al. 2000). Accurate volume measurements were obtained in these patients with moderate and severe OA, with only a slight (12%) underestimation vs. surgically removed tissue and a correlation coefficient of r=0.98 between both methods (Fig. 4). Regional distribution patterns of cartilage thickness were also found to be consistent with those derived from sectioning (Sittek et al. 1996, Eckstein et al. 1996, Kladny et al. 1996), A mode ultrasound (Eckstein et al.

155 Table 1 Precision errors [CV% (and standard deviation) of repeated measurements] obtained for cartilage volume measurements with different sequences, resolutions, and section orientations. The CV% (and SD) are given here as the root-mean-square (RMS) average (Gler et al. 1995), whereas in some of the original papers cited these have been given as the mean CV% or SD. Patella Short-term precision healthy volunteers Tieschky et al. 1997 (tra, fs, 20.6, n=7, 6 rep) Eckstein et al. 1998b (sag, fs, 20.31, n=8, 6 rep) Eckstein et al. 2000c (tra, we, 1.50.31, n=12, 4 rep) Heudorfer et al. 2000 (sag, we, 1.50.31 n=8, 4 rep) Hyhlik-Drr et al. 2000 (cor, we, 1.20.31, n=8, 6 rep) Long-term precision Burgkart et al. 2001 (sag, we, 1.50.31, n=8, 3 rep) Patients with severe OA Burgkart et al. 2000 (cor, we, 1.20.31, n=8, 24 rep) Femur (tra transverse section orientation, sag sagittal, cor coronal; fs spectral fat suppression with prepulse, we selective water excitation; the resolution is given in mm, first giving the section thickness, and then the in-plane resolution; n number of individuals examined, rep number of scan repetitions with repositioning) Medial Tibia Lateral Tibia

1.6% 1.5% 1.0% 2.3%

2.1% 2.8%

3.2% 2.5% 2.3% (48 mm3)

3.8% 2.9% 2.6% (66 mm3)

3.9%

2.4%

3.6%

3.5%

5.5% (56 mm3)

3.8% (59 mm3)

1997), and stereophotogrammetry (Cohen et al. 1999). By exchanging frequency- and phase-encoding directions, it has been excluded that inhomogeneities of the magnetic field (introduced by susceptibility artefact) lead to relevant geometric distortion of the images (Eckstein et al. 2000a). Glaser et al. (2000) compared knee-joint cartilage measurements with a frequency selective fat saturation (prepulse) vs. water excitation technique and found deviations of only 1 to 4%, with slightly higher values for the water excitation sequence. These were attributed to a shorter echo time (TE) and higher signal intensity of the deeper cartilage layers. The in vivo precision (reproducibility) has been studied in healthy volunteers and patients, by repeating measurements after joint repositioning and reshimming of the magnet. These studies permit to estimate how reliably volunteers and patients can be discriminated in cross-sectional studies, and how reliably changes can be detected longitudinally. Generally, the precision error is expressed as a standard deviation (SD) of repeated measurements or as a coefficient of variation (CV%=SD divided by the mean). As a rule, the minimal interval of change that can be detected with 95% confidence in a single individual is 2.8 times the precision error, and the minimal detectable difference in a group of 10 individuals is in the range of the precision error (Cummings and Black 1986). The reproducibility of cartilage volume and thickness measurements was found to be relatively high (Peterfy et al. 1994; Pilch et al. 1994; Marshall et al. 1995; Dupuy et al. 1996; Eckstein et al. 1996; Kshirsagar et al. 1998; Cicuttini et al. 1999); Table 1 summarizing

the findings of our group for different protocols, resolutions, and section orientations. The highest precision (around 1%) was obtained for the patella (transverse section orientation; water excitation sequence). In other knee-joint surfaces the precision error was 24%, this being substanitally lower than the interindividual variation. The long-term precision error (imaging sessions several weeks apart) was somewhat higher than that obtained for short-term conditions (Burgkart et al. 2001), but the differences were relatively small. This suggests that factors such as scanner drift and changes in imaging conditions (temperature, for instance) are not a critical problem in quantitative cartilage imaging. In patients with severe OA (prior to knee arthroplasty), the CV% was higher (due to lower absolute mean values), but the SDs of repeated measurements were found to be similar to those in healthy volunteers (Burgkart et al. 2000). In view of an estimated tissue loss of more than 1000 mm3 in tibial cartilage volume before knee arthroplasty (Hyhlik-Drr et al. 2000; Burgkart et al. 2000), these data suggest that reliable staging and monitoring of OA progression can be performed with quantitative MRI. Computations of the mean cartilage thickness and joint surface areas was shown to yield similar precision errors to those of the cartilage volume, whereas the reproducibility of the maximal cartilage thickness value was found to be somewhat lower (Stammberger et al. 1999b; Hyhlik-Drr et al. 2000; Heudorfer et al. 2000). With digital image analysis it could be demonstrated that regional distribution patterns of articular cartilage throughout joint surfaces also display satisfactory preci-

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sion (Eckstein et al. 1996; Tieschky et al. 1997), the matching algorithm by Stammberger et al. (2000) being currently employed to determine the precision of local/regional (in contrast to global) thickness changes in the human knee-joint cartilage. Other joints with thin cartilage layers Since other joints of the human body display a significantly smaller cartilage thickness than the knee (Adam et al. 1998), spatial resolution is a critical issue in quantitative imaging of these articulations (Link et al. 1998). Peterfy et al. (1995) reported a satisfactory accuracy and precision of volume measurements in the metacarpophalangeal articulation. In elbow joint specimens (average cartilage thickness around 1 mm), we found good agreement of cartilage measurements with a water excitation sequence (resolution 10.250.25 mm3, interpolated to an in-plane resolution of 0.1250.125 mm2), in relation to CT arthrography and A-mode ultrasound (Graichen et al. 2000a). When determining the in vivo precision of this imaging protocol in joints of the hind foot (Fig. 5a) with an average cartilage thickness 0.5 to 0.9 mm), we observed precision errors between 2.1 and 10.9% for single joint surfaces, but smaller errors (<3%) for cumulative measures of several joints (Al-Ali et al. 2001). These results demonstrate that quantitative cartilage imaging is not confined to the knee, but can also be employed for other joints. In the hip and shoulder, however, circular extremity coils cannot be used, and total body coils do not provide adequate resolution. Advances will therefore depend on the design of more efficient surface coils, to permit measurements in all major joints of the human body.

Application to the study of cartilage morphology and function

Determinants of cartilage morphology what makes us so different? Having shown that MRI can be used for in vivo cartilage morphometry, our efforts have focussed on identifying the determinants of the individual cartilage volume, thickness, and surface areas. This question is not only interesting from an anatomical point of view, but is also relevant in the context of retrospectively estimating tissue loss in patients, when symptoms start to occur.

Fig. 5 Quantitative cartilage imaging in the hind foot: a sagittal MR image (10.250.25 mm3) of the human hind foot; b correlation of mean cartilage thickness (mm) in the talocrural joint with the mean cartilage thickness in the talotarsal (subtalar and talocacaneonavicular) joint and the knee joint (c), respectively. Data from 16 young healthy volunteers (8 men, 8 women)

157 Table 2 Interindividual variability (CV%) of knee joint cartilage volume, mean cartilage thickness, and size of the surface areas in 95 healthy individuals (49 men, 46 women; age 20 to 30 years), and side (left-right) differences in 15 individuals (8 men, 7 women; age 2356 years). (Diff absolute difference; Th thickness; med medial; lat lateral) Men (n=49) Patella Volume Mean Th Surface area Femur Volume Mean Th Surface area Med. tibia Volume Mean Th Surface area Lat. tibia Volume Mean Th Surface area Total knee Volume Mean Th Surface area 19% 16% 15% 19% 18% 13% 24% 19% 16% 23% 20% 14% 15% 13% 12% Women All Side Diff (n=46) (n=95) (n=15) 21% 16% 12% 16% 13% 12% 23% 19% 15% 21% 15% 15% 14% 10% 10% 23% 17% 18% 23% 17% 17% 29% 20% 20% 29% 19% 20% 20% 13% 16% 6.35.4% 3.93.5% 3.62.8% 6.04.2% 4.94.2% 4.33.1% 10.16.7% 6.04.5% 7.25.3% 7.35.5% 6.03.9% 6.74.2% 5.03.7% 3.83.1% 3.61.9%

Inter-individual variability and side differences An initial analysis on a small sample of mixed gender (n=20) reported a high degree of inter-individual variability of the cartilage volume, and relatively low associations with the body weight and height (Eckstein et al. 1998c). A subsequent study in 27 adult men (Eckstein et al. 2001a) confirmed that a high degree of variability is also present within one gender (CV% ~20%), and that the variability of cartilage volume is determined both by considerable variation in cartilage thickness and joint surface areas (CV% ~15%, respectively). A high interindividual variability has also been described in the elbow (Springer et al. 1998) and in the joints of the hind foot (Al-Ali et al. 2001). Recently, our data base on knee-joint cartilage has been extended to 95 young healthy volunteers (49 men, 46 women; age 20 to 30 years), the high interindividual variability having been confirmed in this larger sample (Table 2, Fig. 6). When evaluating side differences in the knee (Mller et al. 2000), no significant differences were found between the left and right. The absolute deviations were substantially lower (~5%) than the inter-individual variability (Table 2). Gender differences As seen from the normative data in young healthy volunteers (Fig. 6), men and women display significant differences in cartilage morphology. Cicuttini et al. (1999) reported gender differences of cartilage volume in patients with knee pain (but no other apparent disorders). These differences were significant for the femur (60%) and the patella (47%), but not for the tibia (49%), and remained

Fig. 6 Normative data and gender differences on cartilage morphology in 95 young healthy volunteers (49 men, 46 women), aged 20 to 30 years: a cartilage volume, b cartilage thickness, c cartilage surface areas. Levels of significance of gender differences are given in Table 3

significant after adjusting for age, body weight, height, and femoral (condylar) bone volume. Faber et al. (2001) analyzed healthy men and women with a normal weight (body mass index <25), which had been physically inactive throughout life (no sportive activity for more than 1 h per week). They reported a similar (although statistically significant) gender difference for tibial cartilage volume (+43% in the medial and +47% in the lateral condyle; P< 0.01), but smaller differences in the patella

158 Table 3 Gender differences on knee joint cartilage volume, mean cartilage thickness, and size of the surface areas in (a) physically inactive men and women (n=18); (b) young (2030 years) healthy individuals (n=95); (c) men and women matched for body weight (n=26); (d) men and women matched for body height (n=30). ( From Faber et al. (2001); BW body weight, BH body height; Physically inactive men and women (n=9 vs. 9) Patella Volume Mean Th Surface area Volume Mean Th Surface area Volume Mean Th Surface area Volume Mean Th Surface area Volume Mean Th Surface area +20% +9% +23% ** +27% * +5% +21% ** +47% ** +13% +33% ** +43% ** +9% +33% ** +29% ** +8% +23% ** Th thickness; Med medial; Lat lateral; body weight=607 kg in women, and 769 kg in men; body height=1696 cm in women, and 1816 cm in men; # body weight=66.73.4 kg in women, and 66.61.8 kg in men; + body height=1753 cm in women, and 1753 cm in men) Men and women matched for BW # (n=13 vs. 13) +12% +2% +18% *** +16% +3% +19% *** +29% ** +9% +23% ** +28% ** +9% +22% *** +18% ** +5% +20% *** Men and women matched for BH + (n=15 vs. 15) +14% +5% +11% ** +14% +8% +14% ** +25% * +13% +17% * +32% *** +12% +22% *** +18% *** +7% +15% ***

Young healthy men and women (n=49 vs. 46) +27% *** +7% * +25% *** +38% *** +13% *** +26% *** +36% *** +12% ** +28% *** +41% *** +10% * +31% *** +32% *** +10% *** +26% ***

Femur

Med. tibia

Lat. tibia

Total knee

Table 4 Correlation of cartilage volume, thickness and joint surface areas between the knee, the talocrural and talotarsal joints, and the muscle cross-scetional areas of the thigh and calf, respectively (8 men, 8 women; age 22 to 27 years). (Th thickness; taloTalo-crural joint Knee Volume Mean Th Surface area Volume Mean Th Surface area Volume Mean Th Surface area 0.52* 0.19 0.80***

tarsal joint subtalar + talocalcaneonavicular joint; MCA muscle cross-sectional area; * taken at 50% between the greater trochanter and the lateral joint space of the knee; + taken at the maximum circumference) MCA thigh* 0.83*** 0.77*** 0.78 0.79*** 0.63** 0.76*** 0.79*** 0.48 0.77*** MCA calf+ 0.70** 0.62* 0.79*** 0.82*** 0.61* 0.82*** 0.75*** 0.53* 0.72**

Talo-tarsal joint 0.50 0.36 0.71** 0.92*** 0.62** 0.80***

Talocrural joint

Subtalar+ TCN joint

(+20%; P< 0.01) and femur (+27%; P< 0.01) than Cicuttini et al. (1999). It was shown that the gender differences in cartilage volume originate primarily from differences in the joint surface area size (total knee +23%; P< 0.01), but to a lesser extent from differences in cartilage thickness (+ 8%; difference not statistically significant). In a larger sample (n=95) gender differences of cartilage thickness become significant, but are still considerably smaller than those of joint surface areas (Table 3). When matching men and women with identical body weight or height, the joint surfaces are still significantly larger in men. Cartilage thickness values show a trend to be larger in the men, but the differences do not reach statistical significance (Table 3).

Correlations between joints and joint surfaces Quantitative parameters displayed low correlations amongst the cartilage plates of the knee (patella, femur, medial, and lateral tibia), the coefficients (r) ranging from 0.16 to 0.70 for volume, 0.24 to 0.62 for surface areas, and 0.31 to 0.78 for thickness (Eckstein et al. 2001a). The knee-joint cartilage tissue thus appears to be distributed relatively randomly onto the various cartilage plates, and high or low values in one surface are not necessarily associated with high or low values in another. The patellar cartilage, for instance, can take up between 11 and 27% of the knee volume. Within the hind foot (talocrural joint, subtalar joint, and talocalcaneonavicular joint), the correlation among different cartilage plates tended to be higher than within the knee. The correlation between the talocrural joint and talotarsal joint was r=0.92 for cartilage volume, 0.80 for

159 Table 5 Correlation of knee joint cartilage volume, thickness, and surface areas with the body weight, body height, tibial head diameter, and muscle cross-sectional areas of the thigh. (Th thickness; cross-sect cross-sectional) Body weight All (n=95) Volume Mean Th Surface area Volume Mean Th Surface area Volume Mean Th Surface area 0.62*** 0.38*** 0.64*** 0.30* 0.24 0.25 0.22 0.04 0.26 Body height 0.70*** 0.36*** 0.78*** 0.41** 0.19 0.48*** 0.45** 0.05 0.68*** Tibial head diameter 0.60*** 0.24* 0.85*** 0.03 0.18 0.71*** 0.51*** 0.19 0.65*** Muscle cross-sect area (thigh) 0.76*** (n=35) 0.65*** 0.69*** 0.40 (n=19) 0.35 0.25 0.63** (n=16) 0.61* 0.49

Men (n=49)

Women (n=46)

the surface areas, and 0.62 for the mean thickness (Table 4, Fig. 5). When comparing these joints with the knee in the same individuals, there was a positive correlation for the size of the joint surface areas (range r=+0.24 to +0.84), but not for the cartilage thickness (range r=-0.63 to +0.47) (Table 4, Fig. 5). Cartilage thickness values in different joints in the body appear thus not to be determined by identical factors. Scale effects and correlation with anthropometric variables A relatively weak correlation was observed between body weight or height and the cartilage volume of the knee or elbow. However, a higher association was found with the local bone size (diameter of the tibial head and the humeral condyles, respectively; Eckstein et al. 1998c; Springer et al. 1998). In a sample of men (Eckstein et al. 2001a), the correlations between cartilage volume and body weight/height were also found to be low (r=0.06 to 0.27/0.28 to 0.45 for the various cartilage plates, whereas the size of the bones was shown to display higher correlation coefficients with the cartilage volume (r=0.39 to 0.67). Interestingly, however, the size of the joint surface area and the mean cartilage thickness were not significantly associated (r=0.02 to 0.34), showing that individuals with larger joints do not necessarily display thicker cartilage (Eckstein et al. 2001a). Although Karvonen et al. (1994) reported a significant relationship between the tibial plateau area and cartilage thickness in the knee (as determined from local measurements in single MR images), our current data demonstrates that this association is relatively weak. Table 5 shows the correlation of knee-joint cartilage parameters with the body weight, the body height, and the medio-lateral diameter of the tibial head in 95 healthy volunteers. Body weight and height were significantly associated with cartilage volume and surface areas, and to a lesser extent with the cartilage thickness, when pooling data from men and women. However, the correlation coefficients were considerably lower when analyzing men and women separately (Table 5). The tibial head diameter (determined from the MRI scans) was not bet-

ter in predicting cartilage volume than body weight or height, this being in contrast to our earlier report in a smaller sample (Eckstein et al. 1998c). The correlation with the muscle cross-sectional areas of the thigh, however, showed higher correlation coefficients than body weight or height, in particular for the mean cartilage thickness (Table 5). This suggests that muscle forces may play an important role in the ontogenesis and/or maintenance of individual cartilage morphology. When comparing quantitative cartilage parameters in the hind foot and knee joint with the muscle cross sectional areas of the thigh and calf, respectively, the muscles at the thigh did not display a significantly higher correlation with cartilage volume in the knee than with the hind foot and vice versa (Table 4). The substantial heterogeneity of cartilage thickness between the knee and hindfoot (see above) can thus not be explained by local deviations in muscle force. It thus remains to be established which specific factors are responsible for the heterogeneity of cartilage thickness in different joints and different skeletal regions of the same individual. Functional adaptation of articular cartilage the lazy tissue? Mechanical stimulation Mechanical stimuli have been shown to be very potent regulators of muscle and bone tissue turnover. During immobilization and weightlessness (space flight), there are almost immediate signs of atrophy, whereas intense physical training leads to substantial increases in tissue mass. The biosynthetic activity of chondrocytes is known to be affected by mechanical stimuli, both in cell culture and in explants of cartilage tissue (Sah et al. 1989; Urban 1994). There is evidence that static loading suppresses the production of matrix components, whereas dynamic loading increases PG and collagen synthesis. Spatially varying patterns of cartilage deformation have been associated with spatial variation in metabolic activity (Kim et al. 1995; Buschmann et al. 1996; Wong et al. 1997; Quinn et al. 1998). Animal studies have suggested that cartilage thickness decreases after immobilization,

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Fig. 7 Knee joint cartilage thickness in triathletes vs. physically inactive individuals (PIV). No statistically significant differences are observed between both groups (adapted from Mhlbauer et al. 2000)

but investigations with increased levels of exercise have produced inconclusive and partly contradictory results (Helminen et al. 1992; Newton et al. 1997). Carter and Wong (1988) and Carter et al. (1991) proposed that hydrostatic pressure prevents the enchondral ossification process to progress to the joint surface, and that the magnitude of the pressure determines the thickness of the cartilage layer in joint development. It has therefore been assumed that the inter-individual variability in cartilage thickness is explained by inter-individual differences in the mechanical loading history (Carter et al. 1991). To test this hypothesis, we examined triathletes, who had been training for at least 10 h per week over the last three years and had been physically active also throughout childhood and adolescence. These were compared with individuals who had been never physically active (<1 h sport per week throughout life), had no job involving any physical activity, and had a normal weight (body mass index <25). Surprisingly, male thriathletes did not display significant differences in cartilage thickness compared with physically inactive volunteers (Fig. 7). These findings suggest that (opposite to muscle and bone) substantial increases in mechanical stimulation do not increase the thickness of the cartilage, and that the natural variation in cartilage thickness is not explained by different levels of physical exercise (Mhlbauer et al. 2000). Interestingly, however, we found that triathletes display larger knee joint surface areas (+ 9%/P< 0.01 in men; + 7%/P=0.08 in women; Eckstein et al. 2001b). These data suggest that the mechanism to reduce stress may be by an increase in the bearing surface rather than by an increase in cartilage thickness. The reason for this may be that beyond a certain thickness the nutritive situation of the cartilage becomes critical (Mow and Ratcliffe 1997; Buckwalter and Mankin 1997a), and/or that the stress distribution (load partitioning) within the cartilage becomes unfavorable (Ateshian et al. 1994; Wu et al. 1996). With thicker cartilage, there is more space for the interstitial fluid to es-

Fig. 8 Comparison of patellar cartilage properties after 6 weeks of immobilization vs. the contralateral limb. The findings indicate differences in cartilage morphology (atrophy) and ultrastructural/biochemical changes: mean cartilage thickness; magnetization transfer coefficient (MTC; potential association with macromolecules, specifically with collagen); proton density (potential association with interstitial water content). The error bars show the standard deviation of two repeated measurements

cape laterally from the site of contact, the mechanism of hydrostatic pressurization (Fig. 1) being impaired. With larger contact areas, in contrast, pressurization is enhanced. By demonstrating that the adaptational processes are directed towards improving hyrostatic pressurization rather than elastic damping properties of the cartilage, our observations lend indirect support to the concept of hydrostatic pressurization being the biologically relevant mechanism of load transmission through cartilage. These findings may thus reveal a general principle of joint development, also in phylogenesis, large animals displaying much larger joint surfaces than man, but not necessarily thicker cartilage. Immobilization We recently had the opportunity to examine an individual who had been immobilized for 6 weeks (no weight bearing and limitation of the knee flexion angle to 60) after arthroscopic removal of a joint body and suture of a small meniscal tear). We observed a 36% reduction in quadriceps cross-sectional area (relative to the contralateral side), and an 8% reduction in bone density (DXA) in the left leg (24% in the proximal tibia). The patellar cartilage displayed an 11% lower thickness in the immobilized limb (Fig. 8), whereas the medial and lateral tibial cartilage did not display differences compared with the contralateral knee (0.7 and 0.8%, respectively). These preliminary results suggest that cartilage atrophy may occur when the mechanical stimulus falls short of a certain threshold. It will have to be clarified whether these changes are reversible when loading returns to normal levels, and whether they affect the susceptibility to develop OA. These findings have implications for the

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clinical management of the postoperative period in bone and joint surgery and may potentially require active countermeasures, to avoid cartilage atrophy during immobilization. Aging It has been controversial whether cartilage thinning occurs during the normal aging process (possibly as a result of reduction in mechanical loading) or whether this only affects people with OA. In cadaver studies, Meachim (1971) found no significant decrease of cartilage thickness in the shoulder with age. In the patella, they (Meachim et al. 1977) described thinning of articular cartilage, in particular in women above the age of 50, but these changes were attributed to OA and not to matrix shrinkage. Karvonen et al. (1994) concluded (from local measurements in MR images) that age accounted for a significant linear decrease in cartilage thickness both in the absence and presence of OA. However this was confined to the site of frequent femorotibial contact of the lateral and medial femoral condyle, but did not apply for the patella, tibia, or posterior aspects of the femoral condyles. We recently investigated 15 elderly men and 15 women (50 to 75 years) with no history of knee pain, trauma, surgery (Hudelmaier et al. 2000a). These were compared with volunteers aged less than 30 years. Approx. 25% of the elderly volunteers displayed obvious cartilage lesions, despite being asymptomatic. In those without lesions, we found no significant differences in patellar cartilage thickness between elderly and young men (6%), but a significantly lower thickness (12%; P< 0.05) in elderly women. This indicates that there may exist gender-specific differences in cartilage thinning during aging, which may be due to a higher reduction of muscle force in elderly women compared with elderly men. Analysis of in situ cartilage deformation squeezing the sponge Although the mechanical properties of articular cartilage have been extensively investigated in vitro (both in tissue samples and in indentation Mow et al. 1984, 1993; Mow and Ratcliffe 1997), there have been no reliable data on the amount of in situ cartilage deformation occurring in the intact joint under in vivo loading conditions. These data cannot be extrapolated from in vitro studies, because the magnitude of the joint loads occurring during normal (in particular dynamic) exercise are unknown. The same applies for the effect of the boundary conditions (e.g., non-linear contact conditions between two incongruous joint surfaces, presence of the synovial fluid). Data on the in vivo deformation of articular cartilage are, however, required for estimating the strain (and stress) to which the cartilage is subjected under normal conditions (e.g., as input data for experiments in cell bi-

Fig. 9 MR-based analysis of patellar cartilage deformation (volume change) and recovery in vivo (adpated from Eckstein et al. 1999): a 6 sets (I to VI) of 50 knee bends at 15 min intervals. Data acquisition 37 min and 812 min after exercise; b Cartilage recovery after 100 knee bends for the same individuals shown in a. The cartilage requires >90 min to fully recover after the exercise

ology), and to characterize the mechanical target environment for tissue engineered cartilage. In vivo deformation We have therefore optimized fat-suppressed imaging protocols for fast acquisition of the total patellar cartilage in less than 4 min (Tieschky et al. 1997; Table 1). With this sequence we have quantified the cartilage volume of healthy volunteers after 1 h of physical rest, who were then asked to perform 50 knee bends outside the MR scanner. They were then repositioned in the magnet, and data sets were acquired at 37 min after exercise. We observed deformation of the cartilage in all volunteers (average 6%), the interstitial fluid being displaced from the matrix (Eckstein et al. 1998d). It has been further

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demonstrated that multiple sets of 50 knee bends (at 15 min intervals) do not lead to an increase in cartilage deformation (Fig. 9a), and that the time required for the cartilage volume to attain pre-exercise levels is as long as 90 min (Fig. 9b), due to a relatively low fluid flux through the surface (approximately 0.027 m/s; Eckstein et al. 1999). An interesting finding was that the individual rate (not magnitude) of cartilage recovery was highly correlated (r=0.97) with the level of deformation after the knee bends, both parameters varying considerably between individuals (deformation 2.4% to 8.6%). During the recovery phase of the experiment, no external loading was applied, and the individuals remained relaxed in the scanner throughout the entire 90 min period. The rate of recovery was thus exclusively determined by the properties of the cartilage. The high correlation with the deformation after knee bends therefore suggests that the latter may be determined by inter-individual differences in mechanical cartilage properties. This idea is intriguing, since it involves the possibility to determine functional properties of articular cartilage in vivo. As it is still unclear why some individuals develop OA early in life and others do not. It may therefore be interesting to examine whether functional properties of the cartilage are related to the individual susceptibility of developing OA. Comparing static (90 squatting for 20 s) vs. dynamic loading (30 deep knee bends to 120), differences were found both in the amount (Fig. 10) and in the pattern (Fig. 11) of cartilage deformation throughout the joint surface (Eckstein et al. 2000b). Normal level walking (5 min) was observed to cause only little deformation, whereas running (200 m at level + 54 stairs up and down within approximately 4 min) caused volume changes similar to those seen in squatting (Lemberger et al. 2000) (Fig. 10). The deformation after running was highly correlated (r >0.80) with that after squatting and knee bends, respectively. Waterton et al. (2000) investigated healthy volunteers early in the morning and after a day of mainly standing activity. They reported no change in overall femoral cartilage volume and thickness, but thinning of the cartilage in the femoropatellar and femorotibial contact zones. They also observed an increase in thickness in those areas not subjected to loading during standing and hypothesized that this results from negative intra-articular pressure during joint extension by the quadriceps. However, it has been suggested alternatively that interstitial fluid is displaced from load-bearing to non-load-bearing areas within the cartilage. In elderly individuals without symptoms (50 to 75 years), we found a considerably smaller degree of patellar cartilage deformation than in younger individuals (Hudelmaier et al. 2000b; Fig. 12). This finding was unexpected, since previous in vitro experiments have suggested that cartilage becomes more compliant with aging (Armstrong and Mow 1982). The findings suggest that under physiological condtions cartilage tissue and chondrocytes do not encounter higher, but lower strains in el-

Fig. 10 Magnitude of patellar cartilage deformation (volume change) after different types of exercises: level walking (5 min), running (200 m at level + 54 stairs up and down within approximately 4 min), squatting (90 for 20 s), and 30 knee bends

derly persons. Since the metabolic function of chondrocytes has been to be enhanced by dynamic loading (Sah et al. 1989; Urban 1994), cartilage thinning with aging may be due to a decrease in mechanical stimulation of cartilage cells. A potential explanation for the reduced deformation may be that elderly individuals develop different motor strategies (Papa and Cappozzo 2000) and thus subject their knee joints to smaller loads during knee bending. It has, however, also been demonstrated that certain collagen cross-links increase with aging (pentosidine), both in animals and in humans (Takahashi et al. 1995; Brama et al. 1999). These authors suggested that this may render the cartilage matrix stiffer than in the young. More standardized in vivo loading protocols may be used in the future to clarify the question whether functional properties of the cartilage change with aging. Also, milder forms of cartilage compression may be used clinically to evaluate functional properties in the course of joint disease, and to study the effects of different types of therapy on cartilage quality. In vitro deformation Cartilage deformation during (rather than shortly after) loading cannot be investigated in vivo, since it is impossible to apply relevant loads to the joint of a living person within the MR scanner and to keep the joint in a constant position relative to the coil. To overcome this limitation, we have constructed a non-metallic compression apparatus that is capable of generating loads of up to 1500 N and fits into the extremity coil of a clinical MRI scanner. With this apparatus, the time-dependent deformation of femoropatellar cartilage has been studied in situ in intact cadaveric knee joints, the joint capsule being fully intact (Herberhold et al. 1999). The apparatus

163 Fig. 11 Pattern of patellar cartilage deformation after squatting (a) and deep knee bends (b), as derived by 3D registration of pre- and post exercise thickness reconstructions (Stammberger et al. 2000). Posterior view of the right patellar articular surface (superior aspect at the top and lateral facet on the right). Red areas display regions of high deformation. These are confined to small areas after squatting (a), but are more widely distributed after knee bends (b) (adapted from Eckstein et al. 2000b)

permits the study of cartilage deformation with faster 2D sequences and of obtaining images at 1 min. intervals. Patellar and femoral compression were studied for 4 h under continuous static loading with 150% body weight. A maximal thickness reduction of 5715% was observed for patellar cartilage (Fig. 3) and a volume change of >30%, suggesting that more than 50% of the interstitial fluid were displaced from the matrix (Fig. 13). However, only very little deformation occurred during the first few minutes of loading (3% after 1 min and 11% after 8 min). Two important conclusions were reached from this study: (1) Under in situ conditions the patellar cartilage requires several (>4) h to reach equilibrium under physiologically realistic load magnitudes. This suggests that the state in which the entire load is borne by the solid matrix (Fig. 1) is likely never reached during in vivo

loading. (2) During the first minutes of loading, cartilage deforms relatively little, showing that there is little time for the interstitial fluid to escape initially (Fig. 1). The mechanism of load transmission in articular cartilage is thus not by elastic deformation, but by hydrostatic compression. In combination with biphasic finite element analysis (Donzelli and Spilker 1998), these experiments can be used to calculate the load-partitioning between the fluid phase (hydrostatic pressurization) and the proteoglycan-collagen-matrix, the stresses in the latter eventually causing tissue failure. Other investigators have constructed compression devices for subjecting cartilage probes to uniaxial compression and have observed cartilage properties with high field MR systems (Rubenstein et al. 1996; Kaufman et al. 1999). These experiments have shown that MR relax-

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Fig. 12 Patellar cartilage deformation in elderly (5075 years) asymptomatic individuals (men and women) compared with young (2030 years) healthy volunteers. Elderly men and women display a significantly smaller degree of deformation than younger individuals. No significant differences were observed between genders

ation parameters and signal intensity gradients change during compression in a depth-dependent fashion, affecting the layering of the cartilage in MR images. Kaufman et al. (1999) observed a rise in permeability and decrease in stiffness upon trypsin digestion of the interstitial matrix, with changes in T1 (but not in T2) during compression of degraded cartilage. Performing compression experiments under different spatial orientations, Grnder and Kanowski (1998) have shown that changes of cartilage appearance (layering) depend on the orientation relative to the static magnetic field. Regatte et al. (1999) reported sodium relaxation parameters to be sensitive to PG depletion both in compressed and in uncompressed cartilage, whereas proton relaxation parameters were only sensitive in compressed cartilage. These studies underline that MRI cannot only be used to derive geometric data on cartilage tissue, but also structural and biochemical properties. Structural analysis toward non-invasive histology Clinically, local changes in signal intensity are used for diagnostic purposes (Peterfy and Genant 1996; Peterfy 2000), but similar to the evaluation of cartilage thickness quantitative 3D techniques will be necessary to provide objective and reproducible data on cartilage quality in vivo. A layering of articular cartilage has been observed in many MRI sequences (Lehner et al. 1989; Modl et al. 1991; Rubenstein et al. 1993, 1996; Xia 1998; 2000; Grnder et al. 1998; Kim et al. 1999; Goodwin et al. 2000), and there is great interest in relating the signal intensity gradients observed in specific sequences to the structural variation of the cartilage (histo-

Fig. 13 In vitro compression of patellar cartilage within a nonmetallic loading apparatus over a period of 4 h. The plot shows the cartilage thickness changes of a central transverse image throughout the patella (medial to lateral) as a function of time. A volume change of >30% was recorded, suggesting that more than 50% of the interstitial fluid has been displaced from the matrix during compression (Herberhold et al. 1999)

logic zones) and its biochemical composition. However, some of these layers have been shown to represent artifacts (Erickson et al. 1996; Frank et al. 1997). Due to the anisotropic arrangement of collagen fibrils and the magic angle effect, cartilage appearance and layering is dependent on the relative orientation to the static magnetic field (Xia 1998, 2000; Grnder et al. 1998; Kim et al. 1999; Goodwin et al. 2000). Collagen fiber orientation appear to have a dominant influence on the appearance of layers in hyaline cartilage in long-TR MR imaging through T2 effects; this influence is likely caused by restriction of water mobility and the resulting magic angle effect (Goodwin et al. 2000). Macromolecules and interstitial water content Paul et al. (1993) was one of the first to hypothesize that signal intensity variations across the cartilage (in a spin

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echo sequence) were correlated with the spatial distribution of the PGs. More recent work indicates that gadolinium-diethylene triamine pentaacetic acid (Gd-DTPA) enhanced MR imaging can be potentially used to quantify the PG content of normal and degenerated articular cartilage in vivo (Bashir et al. 1996, 1997, 1999; Allen et al. 1999; Trattnig et al. 1999). Gd-DTPA is a negatively charged contrast agent that distributes inversely to the concentration of glycosaminoglycans in cartilage. Experiments have also been performed with a positively charged nitroxide contrast agents (Bacic et al. 1997; Lattanzio et al. 2000), but these are not yet approved for in vivo usage. Other investigators have propagated T1rho relaxation (Duvvuri et al. 1997) and sodium imaging for measuring PGs (Insko et al. 1997, 1999; Reddy et al. 1998; Regatte et al. 1999), but these techniques require specific hardware, not yet available with clinical MRI scanners. Wolff et al. (1991) employed the magnetization transfer (MT) to evaluate cartilage macromolecules. This technique uses a low-power, off-resonance radio-frequency field, resulting in a decreased signal intensity in regions with tight magnetic coupling between fluid and macromolecules. Some authors suggested that the MT is almost exclusively caused by collagen (Kim et al. 1993; Aoki et al. 1996; Seo et al. 1996), whereas others have found contribution from the PGs (Gray et al. 1995; Wachsmuth et al. 1997). Clinically, it may be feasible to gain specific information on macromolecules (collagen) by obtaining MR images with and without an MT pre-pulse (Vahlensieck et al. 1994, 1998; Peterfy and Genant 1996). Attempts to evaluate the interstitial water content in cartilage have been based on the measurement of the transverse relaxation time (T2; Dardzinski et al. 1997, Frank et al. 1999, Lsse et al. 2000). Alternatively, Selby et al. (1995) developed a protocol for determining the proton density (PD) from gradient echo sequences with variable echo times and flip angles, finding good agreement with direct measurements of the weight fraction of interstitial water. These approaches may permit to diagnose ultrastructural and biochemical alterations of the cartilage earlier, and to initiate preventive measures or treatment before macro-morphological changes occur. Towards quantitative parameters We have therefore developed computational techniques by which the segmentation of the cartilage boundaries is first performed on a T1-weighted, fat-suppressed MR sequence. This topographic information is then transferred to data-sets obtained with pulse-sequences that carry specific information on the structural and biochemical composition of the cartilage. The signal intensity is then analyzed throughout the entire cartilage plate (Hohe et al. 2000a). In case of motion artifacts between acquisitions, the image data can be registered with a 3D leastsquares fit algorithm. Hohe et al. (2000a) reported significant differences in MT and PD between patellar and tibial cartilage. Using

the compression apparatus described above, Faber et al. (1999) found the MT to increase and the PD to decrease during compression of femoropatellar cartilage, these changes being consistent with an increase in macromolecular density during compression, and a decrease in interstitial water content. Mhlbauer et al. (2001) found the reproducibility of in vivo analyses of MT and PD in volunteers to be only moderate and not yet satisfactory. However, they were able to detect a reproducible increase in patellar PD and MT after immobilization (Fig. 8), indicating that intrinsic changes of the cartilage during non-use can be measured with MRI. When averaging data of several acquisition, they reported a positive correlation (r=0.60) of the PD with the deformational behavior after knee bends. This is in agreement with findings in cartilage explants that an increase in interstitial water content is associated with a reduction in mechanical stiffness (Armstrong and Mow 1982). These observations indicate that MR-derived parameters of cartilage properties are associated with the mechanical function of the tissue, setting the path to non-invasive histology and evaluation of functional properties. As biochemical and structural parameters are known to vary topographically by layer and region throughout cartilage, Hohe et al. (2000b) developed a computational technique for analyzing the signal intensity characteristics throughout the depth of the tissue in a layer-specific manner. This technique can be used to quantify MR signal of any given sequence (or combination of sequences) throughout entire cartilage plates, independent of the specific section orientation (Fig. 14). Perspectives and directions what's for the future? Joint mechanics Quantitative morphometry of articular cartilage in the living can be used as an input parameter for computer models of diarthrodial joints. These permit to simulate the load transmission in joints (Blankevoort et al. 1991; Heegaard et al. 1995; Merz et al. 1997), and the adaptational processes in the tissue (van Rietbergen et al. 1992; Eckstein et al. 2000c). These models can be important for the pre-operative planing of surgical interventions that aim to reduce the mechanical stress acting on articular surfaces, and for optimizing the effect of operations on joint mechanics. There is growing evidence for a key role for joint laxity in the natural history of OA (Sharma et al. 1999a,b). Open MR imaging systems now permit to apply external loads during imaging (Graichen et al. 1999), testing the effect of normal or pathologic neuromuscular control on skeletal motion patterns (Graichen et al. 2000b) or joint stability (Graichen et al. 2000c). These techniques have been recenty applied to study patients with shoulder instability (von Eisenhart et al. 2000) and cruciate ligament deficiency of the knee (Bringmann et al. 2000). Cohen et al. (1998) determined the in situ joint contact areas from MR imaging data. They obtained image

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167 Fig. 15 Quantitative analysis of surface curvature from MR image data: patellar cartilage. The method determines the distribution of maximal and minimal principal curvatures as well as the mean curvature within the cartilage surface (green convex areas; red concave areas)

data of femoral and patellar joint surface morphology, superimposed these to data for different joint position (45 flexion), and used a proximity criterion to compute femoropatellar contact zones. The cross-registration of high-resolution morphological data on articular cartilage with either open MR imaging (including the application of external forces) or kinematic/gait analysis (Alexander et al. 2000) may thus be developed into efficient tools for assessing joint mechanics during normal neuromuscular control, and to analyze the specific mechanical factors that initiate or promote cartilage degeneration. As the contact mechanics and contact stresses of joints are determined by the congruity/ incongruity of
v

the articular surfaces (Eckstein et al. 2000c), methods have been developed for analyzing the surface curvatures of diathrodial joints from stereophogrammetric data of cadaver joint surfaces (Ateshian 1993; Ateshian and Soslowsky 1997). Similar methods are currently used by us to determine the spatial distribution of maximal, minimal, and mean principal curvatures throughout articular surfaces from in vivo MRI data (Fig. 15). This is achieved by fitting B-spline surfaces to the segmented cartilage contours and performing Gaussian curvature analysis (Hohe et al. 2001). Quantitative data on surface geometry and incongruity are potentially valuable as risk factors for predicting the initiation and progression of osteoarthritis. Mechano-adaptation of articular cartilage Our current data suggest that no adaptation of the cartilage thickness occurs to increased mechanical stimulation, and that the normal variability in cartilage thickness cannot be explained by differences in loading history. These data will have to be confirmed in athletes that perform different types of activities (e.g., sprinters and

Fig. 14ae Layer-specific analysis of the regional cartilage MR signal intensity. a Transverse section through patellar cartilage, indicating the regions and layers for which the signal intensity is computed. Graphs be show the distribution of magnetization transfer (MT) throughout the articular surface (intensity projectiong) of patellar cartilage: a) averaging throughout the entire depth of the tissue. b Surface layer (superficial 33%). c Middle 33%. d Deep 33%. The technique can be employed to derive and visualize biochemical/structural properties of the cartilage in various depths throughout the entire joint surface (Color legend: <41% red, 4144% orange, then intervals of 3%..., blue >59%)

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weight lifters), which involve more intense (although less frequent) loading, and larger increases in speed and maximal muscle force (rather than endurance). Since reliable measurements can now also be obtained in the ankle joint, it is possible to study patients with Van Nees rotationplasty (Gottsauner et al. 1991; Steenhoff et al. 1993). This operation is used to maintain knee joint function (using the rotated ankle joint) in cases of above knee amputation. This intervention can be used to investigate whether ankle joint cartilage undergoes morphological changes when being subjected to loads that normally occur in the knee, either increases in its thickness (adaptation) or damage (degeneration). As our preliminary observations indicate that tissue atrophy may occur during immobilization, this issue will have to be studied in further detail, since it has important implications for clinical management of the postoperative period in bone and joint surgery. The observation of cartilage atrophy must be verified longitudinally, for instance in astronauts, bed-rest studies, or patients subject to immobilization. It will be important to clarify which minimal mechanical stimulus is required to maintain cartilage thickness, and whether remobilization (after prolonged periods of non-use) can restore the original cartilage thickness or not. This is of particular importance, since animal studies have indicated that morphological, biochemical, and biomechanical cartilage alterations cannot be fully reversed after immobilization (Kiviranta et al. 1994; Setton et al. 1997; Haapala et al. 1999, 2000). Osteoarthritis (clinical mangagement, epidemiology, drug efficacy studies) Quantitative cartilage imaging with MR has great potential to become a powerful tool in the reserach and clinical management of OA. The relatively large variability of normal cartilage morphology represents a current obstacle in reliably estimating tissue loss retrospectively in cross-sectional studies. However, Burgkart et al. (2000) have shown that t-scores (deviations of patients vs. healthy young volunteers, expressed in standard deviations) can be enhanced when normalizing the cartilage volume to the original (now partly denuded) joint surface area (Fig. 4b). These MR-based parameters will have to be compared directly with radiographic evaluation of the joint, to demonstrate whether quantitative MRI is more sensitive than current methodology. The most important step will be to longitudinally determine the annual rate of change of cartilage volume in different cartilage plates in OA patients, in order to allow for reliable sample size calculations. The 3D matching algorithm by Stammberger et al. (2000) may prove particularly useful in this context, since it permits to analyze the regional pattern of cartilage loss throughout the joint surfaces. In epidemiological studies, quantitative cartilage imaging can now be readily employed to determine the risk factors of both the onset and progression of OA in larger

cohorts. Since more specific information can be gained on the spatial and temporal pattern of knee joint cartilage loss, these studies may be able to draw a more detailed picture of the natural history of OA, and to objectively differentiate subsets of patients at different stages of disease, and with different degrees of OA progression. Eventually, the method may prove powerful in testing the effect of new therapeutic compounds in clinical trials. Because of the immense costs involved in developing new agents, efficient clinical trials are very critical in drug development. The methodology now available may help to identify the most promising compounds early. It can also be potentially applied to animal models of OA, in which drug effects can be demonstrated on a much smaller time scale (e.g., Beckmann et al. 1995; Gahunia et al. 1995; Watson et al. 1996; 1997; Bacic et al. 1997). Methods for in vivo analysis of cartilage biochemistry and structure are still awaiting validation, but these will be important in detecting cartilage changes at the earliest possible time point.

Conclusions
In vivo assessment of human articular cartilage morphology and deformation has set the path for achieving more detailed knowledge on form-function relationships in synovial joints in health and disease. Dedicated magnetic resonance imaging sequences and 3D digital postprocessing techniques now permit to determine cartilage morphology with high accuracy and precision. Important insights have been gained on cartilage deformation in situ, and theories are beginning to emerge to explain variations in normal cartilage morphology and its functional adaptation to mechanical stimuli. Epidemiologic studies may allow to identify the specific factors involved in disease initiation and progression. Noninvasive analysis of cartilage quantity and quality may also become a valuable tool in the clinical management of OA, including the selection of the most effective therapeutic compounds in clinical trials.
Acknowledgments We would like to express our thanks to Rainer Burgkart, Sonja Faber, Christian Glaser, Heiko Graichen, Jan Hohe, Martin Hudelmaier, Roland Mhlbauer, and Tobias Stammberger for their contributions to this article, and to all students involved in these studies. This work has been supported by the Deutsche Forschungsgemeinschaft.

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