Ectopic mineralization of articular cartilage in the bullfrog Rana catesbeiana and its possible involvement in bone closure

Mineralization of the articular cartilage is a pathological condition associated with age and certain joint diseases in humans and other mammals. In this work, we describe a physiological process of articular cartilage mineralization in bullfrogs. Articular cartilage of the proximal and distal ends of the femur and of the proximal end of the tibia-fibula was studied in animals of different ages. Mineralization of the articular cartilage was detected in animals at 1 month post-transformation. This mineralization, which appeared before the hypertrophic cartilage showed any calcium deposition, began at a restricted site in the lateral expansion of the cartilage and then progressed to other areas of the epiphyseal cartilage. Mineralized structures were identified by von Kossa's staining and by in vivo incorporation of calcein green. Element analysis showed that calcium crystals consisted of poorly crystalline hydroxyapatite. Mineralized matrix was initially spherical structures that generally coalesced after a certain size to occupy larger areas of the cartilage. Alkaline phosphatase activity was detected at the plasma membrane of nearby chondrocytes and in extracellular matrix. Apoptosis was detected by the TUNEL (TDT-mediated dUTP-biotin nick end-labeling) reaction in some articular chondrocytes from mineralized areas. The area occupied by calcium crystals increased significantly in older animals, especially in areas under compression. Ultrastructural analyses showed clusters of needle-like crystals in the extracellular matrix around the chondrocytes and large blocks of mineralized matrix. In 4-year-old animals, some lamellar bone (containing bone marrow) occurred in the same area as articular cartilage mineralization. These results show that the articular cartilage of R. catesbeiana undergoes precocious and progressive mineralization that is apparently stimulated by compressive forces. We suggest that this mineralization is involved in the closure of bone extremities, since mineralization appears to precede the formation of a rudimentary secondary center of ossification in older animals.

Comparative study by computed radiography, histology, and scanning electron microscopy of the articular cartilage of normal goats and in chronic infection with caprine arthritis-encephalitis virus

In the northeast of Brazil, caprine arthritis-encephalitis (CAE) is one of the key reasons for herd productivity decreasing that result in considerable economic losses. A comparative study was carried out using computed radiography (CR), histological analysis (HA), and scanning electronic microscopy (SEM) of the joints of CAE infected and normal goats. Humerus head surface of positive animals presented reduced joint space, increased bone density, and signs of degenerative joint disease (DJD). The carpal joint presented no morphological alterations in CR in any of the animals studied. Tarsus joint was the most affected, characterized by severe DJD, absence of joint space, increased periarticular soft tissue density, edema, and bone sclerosis. Histological analysis showed chronic tissue lesions, complete loss of the surface zone, absence of proteoglycans in the transition and radial zones and destruction of the cartilage surface in the CAE positive animals. Analysis by SEM showed ulcerated lesions with irregular and folded patterns on the joint surface that distinguished the limits between areas of normal and affected cartilage. The morphological study of the joints of normal and CAE positive goats deepened understanding of the alteration in the tissue bioarchitecture of the most affected joints. The SEM finding sustained previous histological reports, similar to those found for rheumatoid arthritis, suggesting that the goat infected with CAE can be considered as a potential model for research in this area.

Treatment of knee articular cartilage defects: surgical strategies, results and analysis of factors determining the clinical outcome

Articular cartilage lesions, with their inherent limited healing potential, are hard to treat and remain a challenging problem for orthopedic surgeons. Despite the development of several treatment strategies, the real potential of each procedure in terms of clinical benefit and effects on the joint degeneration processes is not clear. Aim of this PhD project was to evaluate the results, both in terms of clinical and imaging improvement, of new promising procedures developed to address the challenging cartilage pathology. Several studies have been followed in parallel and completed over the 3-year PhD, and are reported in detail in the following pages. In particular, the studies have been focused on the evaluation of the treatment indications of a scaffold based autologous chondrocyte implantation procedure, documenting its results for the classic indication of focal traumatic lesions, as well as its use for the treatment of more challenging patients, older, with degenerative lesions, or even as salvage procedure for more advanced stages of articular degeneration. The second field of study involved the analysis of the results obtained treating lesions of the articular surface with a new biomimetic osteochondral scaffold, which showed promise for the treatment of defects where the entire osteochondral unit is involved. Finally, a new minimally invasive procedure based on the use of growth factors derived from autologous platelets has been explored, showing results and underlining indicatios for the treatment of cartilage lesions and different stages of joint degeneration. These studies shed some light on the potential of the evaluated procedures, underlining good results as well as limits, they give some indications on the most appropriate candidates for their application, and document the current knowledge on cartilage treatment procedures suggesting the limitations that need to be addressed by future studies to improve the management of cartilage lesions.

T2 star relaxation times for assessment of articular cartilage at 3 T: a feasibility study

T2 mapping techniques use the relaxation constant as an indirect marker of cartilage structure, and the relaxation constant has also been shown to be a sensitive parameter for cartilage evaluation. As a possible additional robust biomarker, T2* relaxation time is a potential, clinically feasible parameter for the biochemical evaluation of articular cartilage.

The structure of articular cartilage

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Evaluation of articular cartilage in patients with femoroacetabular impingement (FAI) using T2* mapping at different time points at 3.0 Tesla MRI: a feasibility study

To define the feasibility of utilizing T2* mapping for assessment of early cartilage degeneration prior to surgery in patients with symptomatic femoroacetabular impingement (FAI), we compared cartilage of the hip joint in patients with FAI and healthy volunteers using T2* mapping at 3.0 Tesla over time.

The structural architecture of adult mammalian articular cartilage evolves by a synchronized process of tissue resorption and neoformation during postnatal development

OBJECTIVE: During postnatal development, mammalian articular cartilage acts as a surface growth plate for the underlying epiphyseal bone. Concomitantly, it undergoes a fundamental process of structural reorganization from an immature isotropic to a mature (adult) anisotropic architecture. However, the mechanism underlying this structural transformation is unknown. It could involve either an internal remodelling process, or complete resorption followed by tissue neoformation. The aim of this study was to establish which of these two alternative tissue reorganization mechanisms is physiologically operative. We also wished to pinpoint the articular cartilage source of the stem cells for clonal expansion and the zonal location of the chondrocyte pool with high proliferative activity. METHODS: The New Zealand white rabbit served as our animal model. The analysis was confined to the high-weight-bearing (central) areas of the medial and lateral femoral condyles. After birth, the articular cartilage layer was evaluated morphologically at monthly intervals from the first to the eighth postnatal month, when this species attains skeletal maturity. The overall height of the articular cartilage layer at each juncture was measured. The growth performance of the articular cartilage layer was assessed by calcein labelling, which permitted an estimation of the daily growth rate of the epiphyseal bone and its monthly length-gain. The slowly proliferating stem-cell pool was identified immunohistochemically (after labelling with bromodeoxyuridine), and the rapidly proliferating chondrocyte population by autoradiography (after labelling with (3)H-thymidine). RESULTS: The growth activity of the articular cartilage layer was highest 1 month after birth. It declined precipitously between the first and third months, and ceased between the third and fourth months, when the animal enters puberty. The structural maturation of the articular cartilage layer followed a corresponding temporal trend. During the first 3 months, when the articular cartilage layer is undergoing structural reorganization, the net length-gain in the epiphyseal bone exceeded the height of the articular cartilage layer. This finding indicates that the postnatal reorganization of articular cartilage from an immature isotropic to a mature anisotropic structure is not achieved by a process of internal remodelling, but by the resorption and neoformation of all zones except the most superficial (stem-cell) one. The superficial zone was found to consist of slowly dividing stem cells with bidirectional mitotic activity. In the horizontal direction, this zone furnishes new stem cells that replenish the pool and effect a lateral expansion of the articular cartilage layer. In the vertical direction, the superficial zone supplies the rapidly dividing, transit-amplifying daughter-cell pool that feeds the transitional and upper radial zones during the postnatal growth phase of the articular cartilage layer. CONCLUSIONS: During postnatal development, mammalian articular cartilage fulfils a dual function, viz., it acts not only as an articulating layer but also as a surface growth plate. In the lapine model, this growth activity ceases at puberty (3-4 months of age), whereas that of the true (metaphyseal) growth plate continues until the time of skeletal maturity (8 months). Hence, the two structures are regulated independently. The structural maturation of the articular cartilage layer coincides temporally with the cessation of its growth activity - for the radial expansion and remodelling of the epiphyseal bone - and with sexual maturation. That articular cartilage is physiologically reorganized by a process of tissue resorption and neoformation, rather than by one of internal remodelling, has important implications for the functional engineering and repair of articular cartilage tissue.

T2 mapping of hip articular cartilage in healthy volunteers at 3T: a study of topographic variation

PURPOSE: To perform baseline T(2) mapping of the hips of healthy volunteers, focusing on topographic variation, because no detailed study has involved hips. T(2) mapping is a quantitative magnetic resonance imaging (MRI) technique that evaluates cartilage matrix components. MATERIALS AND METHODS: Hips of 12 healthy adults (six men and six women; mean age = 29.5 +/- 4.9 years) were studied with a 3.0-Tesla MRI system. T(2) measurement in the oblique-coronal plane used a multi-spin-echo (MSE) sequence. Femoral cartilage was divided into 12 radial sections; acetabular cartilage was divided into six radial sections, and each section was divided into two layers representing the superficial and deep halves of the cartilage. T(2) of these sections and layers were measured. RESULTS: Femoral cartilage T(2) was the shortest (-20 degrees to 20 degrees and -10 degrees to 10 degrees , superficial and deep layers), with an increase near the magic angle (54.7 degrees ). Acetabular cartilage T(2) in both layers was shorter in the periphery than the other parts, especially at 20 degrees to 30 degrees . There were no significant differences in T(2) between right and left hips or between men and women. CONCLUSION: Topographic variation exists in hip cartilage T(2) in young, healthy adults. These findings should be taken into account when T(2) mapping is applied to patients with degenerative cartilage. J. Magn. Reson. Imaging 2007;26:165-171. (c) 2007 Wiley-Liss, Inc.

Effect of multislice acquisition on T1 and T2 measurements of articular cartilage at 3T

PURPOSE: The aim of this study was to investigate the effect of magnetization transfer on multislice T(1) and T(2) measurements of articular cartilage. MATERIALS AND METHODS: A set of phantoms with different concentrations of collagen and contrast agent (Gd-DTPA(2-)) were used for the in vitro study. A total of 20 healthy knees were used for the in vivo study. T(1) and T(2) measurements were performed using fast-spin-echo inversion-recovery (FSE-IR) sequence and multi-spin-echo (MSE) sequence, respectively, in both in vitro and in vivo studies. We investigated the difference in T(1) and T(2) values between that measured by single-slice acquisition and that measured by multislice acquisition. RESULTS: Regarding T(1) measurement, a large drop of T(1) in all slices and also a large interslice variation in T(1) were observed when multislice acquisition was used. Regarding T(2) measurement, a substantial drop of T(2) in all slices was observed; however, there was no apparent interslice variation when multislice acquisition was used. CONCLUSION: This study demonstrated that the adaptation of multislice acquisition technique for T(1) measurement using FSE-IR methodology is difficult and its use for clinical evaluation is problematic. In contrast, multislice acquisition for T(2) measurement using MSE was clinically applicable if inaccuracies caused by multislice acquisition were taken into account. J. Magn. Reson. Imaging 2007;26:109-117. (c) 2007 Wiley-Liss, Inc.

In vivo biochemical 7.0 Tesla magnetic resonance: preliminary results of dGEMRIC, zonal T2, and T2* mapping of articular cartilage

INTRODUCTION: Ultra-high-field whole-body systems (7.0 T) have a high potential for future human in vivo magnetic resonance imaging (MRI). In musculoskeletal MRI, biochemical imaging of articular cartilage may benefit, in particular. Delayed gadolinium-enhanced MRI of cartilage (dGEMRIC) and T2 mapping have shown potential at 3.0 T. Although dGEMRIC, allows the determination of the glycosaminoglycan content of articular cartilage, T2 mapping is a promising tool for the evaluation of water and collagen content. In addition, the evaluation of zonal variation, based on tissue anisotropy, provides an indicator of the nature of cartilage ie, hyaline or hyaline-like articular cartilage.Thus, the aim of our study was to show the feasibility of in vivo dGEMRIC, and T2 and T2* relaxation measurements, at 7.0 T MRI; and to evaluate the potential of T2 and T2* measurements in an initial patient study after matrix-associated autologous chondrocyte transplantation (MACT) in the knee. MATERIALS AND METHODS: MRI was performed on a whole-body 7.0 T MR scanner using a dedicated circular polarization knee coil. The protocol consisted of an inversion recovery sequence for dGEMRIC, a multiecho spin-echo sequence for standard T2 mapping, a gradient-echo sequence for T2* mapping and a morphologic PD SPACE sequence. Twelve healthy volunteers (mean age, 26.7 +/- 3.4 years) and 4 patients (mean age, 38.0 +/- 14.0 years) were enrolled 29.5 +/- 15.1 months after MACT. For dGEMRIC, 5 healthy volunteers (mean age, 32.4 +/- 11.2 years) were included. T1 maps were calculated using a nonlinear, 2-parameter, least squares fit analysis. Using a region-of-interest analysis, mean cartilage relaxation rate was determined as T1 (0) for precontrast measurements and T1 (Gd) for postcontrast gadopentate dimeglumine [Gd-DTPA(2-)] measurements. T2 and T2* maps were obtained using a pixelwise, monoexponential, non-negative least squares fit analysis; region-of-interest analysis was carried out for deep and superficial cartilage aspects. Statistical evaluation was performed by analyses of variance. RESULTS: Mean T1 (dGEMRIC) values for healthy volunteers showed slightly different results for femoral [T1 (0): 1259 +/- 277 ms; T1 (Gd): 683 +/- 141 ms] compared with tibial cartilage [T1 (0): 1093 +/- 281 ms; T1 (Gd): 769 +/- 150 ms]. Global mean T2 relaxation for healthy volunteers showed comparable results for femoral (T2: 56.3 +/- 15.2 ms; T2*: 19.7 +/- 6.4 ms) and patellar (T2: 54.6 +/- 13.0 ms; T2*: 19.6 +/- 5.2 ms) cartilage, but lower values for tibial cartilage (T2: 43.6 +/- 8.5 ms; T2*: 16.6 +/- 5.6 ms). All healthy cartilage sites showed a significant increase from deep to superficial cartilage (P < 0.001). Within healthy cartilage sites in MACT patients, adequate values could be found for T2 (56.6 +/- 13.2 ms) and T2* (18.6 +/- 5.3 ms), which also showed a significant stratification. Within cartilage repair tissue, global mean values showed no difference, with 55.9 +/- 4.9 ms for T2 and 16.2 +/- 6.3 ms for T2*. However, zonal assessment showed only a slight and not significant increase from deep to superficial cartilage (T2: P = 0.174; T2*: P = 0.150). CONCLUSION: In vivo T1 dGEMRIC assessment in healthy cartilage, and T2 and T2* mapping in healthy and reparative articular cartilage, seems to be possible at 7.0 T MRI. For T2 and T2*, zonal variation of articular cartilage could also be evaluated at 7.0 T. This zonal assessment of deep and superficial cartilage aspects shows promising results for the differentiation of healthy and affected articular cartilage. In future studies, optimized protocol selection, and sophisticated coil technology, together with increased signal at ultra-high-field MRI, may lead to advanced biochemical cartilage imaging.