Susan Chubinskaya

Post‐traumatic osteoarthritis: Improved understanding and opportunities for early intervention

Even with current treatments of acute joint injuries, more than 40% of people who suffer significant ligament or meniscus tears, or articular surface injuries, will develop osteoarthritis (OA). Correspondingly, 12% or more of all patients with lower extremity OA have a history of joint injury. Recent research suggests that acute joint damage that occurs at the time of an injury initiates a sequence of events that can lead to progressive articular surface damage. New molecular interventions, combined with evolving surgical methods, aim to minimize or prevent progressive tissue damage triggered by joint injury. Seizing the potential for progress in the treatment of joint injuries to forestall OA will depend on advances in (1) quantitative methods of assessing the injury severity, including both structural damage and biologic responses, (2) understanding of the pathogenesis of post‐traumatic OA, taking into account potential interactions among the different tissues and the role of post‐traumatic incongruity and instability, and (3) application of engineering and molecular research to develop new methods of treating injured joints. This paper highlights recent advances in understanding of the structural damage and the acute biological response following joint injury, and it identifies important directions for future research.

OP-1/BMP-7 in cartilage repair

Three years ago we published a book chapter on the role of bone morphogenetic proteins (BMPs) in cartilage repair. Since that time our understanding of the function of osteogenic protein-1 (OP-1) or BMP-7 in cartilage homeostasis and repair has substantially improved and therefore we decided to devote a current review solely to this BMP. Here we summarise the information accumulated on OP-1 from in vitro and ex vivo studies with cartilage cells and tissues as well as from in vivo studies of cartilage repair in various animal models. The primary focus is on articular chondrocytes and cartilage, but data will also be presented on nonarticular cartilage, particularly from the intervertebral disc. The data show that OP-1 is a unique growth factor which, unlike other members of the same BMP family, exhibits in addition to its strong pro-anabolic activity very prominent anti-catabolic properties. Animal studies have demonstrated that OP-1 has the ability to repair cartilage in vivo in various models of articular cartilage degradation, including focal osteochondral and chondral defects and osteoarthritis, as well as models of degeneration in intervertebral disc cartilage. Together our findings indicate a significant promise for OP-1 as therapeutic in cartilage repair.RésuméIl y a trois ans a été publié le chapitre d’un livre sur le rôle des BMP dans la réparation cartilagineuse. Depuis cette époque, notre compréhension du mécanisme d’action de l’OP-1 ou BMP-7 dans la régulation cartilagineuse et sa réparation ont été améliorées. Nous avons décidé de passer en revue les différents travaux portant sur cette BMP. Nous avons résumé les informations accumulées sur l’OP-1, les études in-vitro et ex-vivo sur les cellules cartilagineuses et sur les tissus à partir de travaux sur la réparation cartilagineuse chez différents modèles d’animaux. Ces travaux ont été essentiellement centrés sur les chondrocytes du cartilage articulaire mais l’on peut également trouver des données sur le cartilage non articulaire particulièrement au niveau du disque inter vertébral. Ces données montrent que l’OP-1 est le seul facteur de croissance. Dans la famille des autres BMP les activités pro-anaboliques sont plus importantes que les propriétés anti anaboliques. Les études animales démontrent que l’OP-1 permet une réparation cartilagineuse parmi les différents modèles de lésions de ce cartilage, outre les lésions cartilagineuses, les lésions d’arthrose et les lésions de dégénérescence des disques intervertébraux. Tout cela nous fait entrevoir une sérieuse avancée thérapeutique grâce à l’OP-1 dans les champs des réparations cartilagineuses.

Human Articular Chondrocytes Express Osteogenic Protein-1

This study demonstrates for the first time that human articular chondrocytes express osteogenic protein-1 (OP-1). OP-1 was originally purified from bone matrix and was shown to induce cartilage and bone formation. Both OP-1 protein and message were present in human normal and osteoarthritic (OA) cartilages. OP-1 mRNA was upregulated in OA cartilage compared with normal adult tissues. However, the level of mature OP-1 protein in the same OA tissues was downregulated, whereas the pro-OP-1 remained high. Moreover, these two forms of OP-1 were localized in an inverted manner. Mature OP-1 was primarily detected in the superficial layer, whereas the pro-form was mostly in the deep layer of cartilage. The presence of pro- and mature OP-1 in extracts of normal and OA cartilages was confirmed by Western blotting. These findings imply that articular chondrocytes continue to express and synthesize OP-1 throughout adulthood. The observed patterns of the distribution of pro- and mature OP-1 also suggest differences in the processing of this molecule by normal and OA chondrocytes and by the cells in the superficial and deep layers. Distinct distribution of OP-1 and its potential activation in deep zones and regions of cloning in OA cartilages may provide clues to the potential involvement of endogenous OP-1 in repair mechanisms.

Regulation of osteogenic proteins by chondrocytes.

The purpose of this review is to summarize the current scientific knowledge of bone morphogenetic proteins (BMPs) in adult articular cartilage. We specifically focus on adult cartilage, since one of the major potential applications of the members of the BMP family may be a repair of adult tissue after trauma and/or disease. After reviewing cartilage physiology and BMPs, we analyze the data on the role of recombinant BMPs as anabolic agents in tissue formation and restoration in different in vitro and in vivo models following with the endogenous expression of BMPs and factors that regulate their expression. We also discuss recent transgenic modifications of BMP genes and subsequent effect on cartilage matrix synthesis. We found that the most studied BMPs in adult articular cartilage are BMP-7 and BMP-2 as well as transforming growth factor-beta (TGF-beta). There are a number of contradicting reports for some of these growth factors, since different models, animals, doses, time points, culture conditions and devices were used. However, regardless of the experimental conditions, only BMP-7 or osteogenic protein-1 (OP-1) exhibits the most convincing effects. It is the only BMP studied thus far in adult cartilage that demonstrates strong anabolic activity in vitro and in vivo with and without serum. OP-1 stimulates the synthesis of the majority of cartilage extracellular matrix proteins in adult articular chondrocytes derived from different species and of different age. OP-1 counteracts the degenerative effect of numerous catabolic mediators; it is also expressed in adult human, bovine, rabbit and goat articular cartilage. This review reveals the importance of the exploration of the BMPs in the cartilage field and highlights their significance for clinical applications in the treatment of cartilage-related diseases.

Long-Term Effects of Bupivacaine on Cartilage in a Rabbit Shoulder Model

Background Previous investigations have reported on the chondrotoxicity of bupivacaine in short-term in vivo and in vitro models. This study was designed to provide additional information on the long-term effects of bupivacaine infusion on articular cartilage in an established rabbit shoulder model. Hypothesis Infusion of bupivacaine into the rabbit shoulder will have long-term deleterious effects on articular cartilage. Study Design Controlled laboratory study. Methods Thirty-six rabbits were randomized into 3 groups and were infused over 48 hours with saline (S), bupivacaine alone (B), or bupivacaine with epinephrine (B+E) into the glenohumeral joint. Animals were sacrificed after 3 months, and tissue samples were analyzed with live/dead cell assay, proteoglycan (PG) synthesis and content assays, and conventional histological evaluation. Results No macroscopic or radiographic changes were detected in the infused shoulders. Sulfate uptake of infused shoulders relative to controls was elevated to 112% ± 39% (S), 166% ± 67% (B), and 210% ± 127% (B+E). Statistical analysis of PG content demonstrated significantly increased levels in bupivacaine groups compared with saline. There were no significant differences among groups in cell count, percentage of living cells, or histological grade. Conclusions No permanent impairment of cartilage function was detected 3 months after intra-articular infusion of bupivacaine. Cartilage metabolism was increased, indicating a possible reparative response. This suggests that, at least in the model used, articular cartilage has the ability to recover from the chondrotoxic effects of bupivacaine infusion. Before extrapolating these results to human cartilage, other factors including underlying cartilage injury or disease, decreased chondrocyte density, and increased bupivacaine dosing need to be taken into account. Clinical Relevance Bupivacaine toxicity has recently been implicated in the development of chondrolysis after arthroscopic shoulder procedures, but these findings suggest that additional noxious stimuli might be required before permanent damage ensues.

Age-related changes in cartilage endogenous osteogenic protein-1 (OP-1).

Articular cartilage has a poor reparative capacity. This feature is exacerbated with aging and during degenerative joint conditions, contributing to loss of motion and impairment of quality of life. This study focused on osteogenic protein-1 (OP-1) and its ability to serve as a repair-stimulating factor in articular cartilage. The purpose of this work was to develop a quantitative method for the assessment of the content of OP-1 protein in extracts from human articular cartilage and to investigate the changes in OP-1 mRNA expression and protein levels with aging of normal adult cartilage. Full thickness cartilage was dissected from femoral condyles of donors with no history of joint disease within 24 h of death. Levels of OP-1 mRNA expression were measured by a semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) method; concentration of OP-1 protein was detected by new sandwich enzyme-linked immunosorbent assay (ELISA); qualitative changes in OP-1 forms were evaluated by Western blots with various anti-OP-1 antibodies. The sensitivity of the ELISA method allowed the detection of picogram quantities of OP-1 in cartilage extracts. We found that (1) concentration of OP-1 in normal cartilage is within the range of biological activity of OP-1 in vitro; and (2) during aging of human adult, articular cartilage, levels of OP-1 protein and message are dramatically reduced (more than 4-fold; p<0.02). The major qualitative changes affected primarily mature OP-1. The results of the current study suggest the possibility that OP-1 may be critical for chondrocytes to maintain their normal homeostasis and could also serve as a repair factor during joint disease or aging.

Restoration of articular cartilage.

➤ Novel (i.e., quantitative and semiquantitative) cartilage imaging techniques can evaluate cartilage composition to augment information obtained from traditional magnetic resonance imaging sequences that detail morphology.➤ A well-defined role for drugs leading to chondroprotection has not yet been determined.➤ Shortcomings of bone marrow stimulation include limited production of hyaline repair tissue, unpredictable repair cartilage volume, and a negative impact on later cellular transplantation if required.➤ The role of biological augments, such as cellular concentrates or platelet-rich plasma, remains undefined. When their use is reported in the literature, it is important that their process of production and characterization be detailed.➤ Rehabilitation programs, incorporating controlled exercise and progressive partial weight-bearing, are an important part of cartilage repair surgery and should be detailed in reports on operative techniques applied.➤ Malalignment, meniscal injury, and ligament deficiency should be corrected in a staged or concomitant fashion to reduce the overall likelihood of mechanical failure in cartilage repair surgery.

BMP-7 protects against progression of cartilage degeneration after impact injury

In vivo studies were used to characterize a model of cartilage injury leading to osteoarthritis progression in the medial femorotibial joint of sheep. In three subsequent studies, bilateral impact injuries were created and one joint received intraarticular injections of 340 µg of rhBMP‐7 protein in a collagen particle carrier while the contralateral knee received the vehicle alone. Sheep were allocated to three groups that received intraarticular injections on day 0 (group A), 21 (group B), or 90 (group C) after experimental knee injury. In each group the, joints were evaluated for signs of osteoarthritis progression 90 days after the last treatment using India ink stained area, OARSI histological scoring, cartilage sGAG content, immunostaining for apoptosis (TUNEL), caspase‐3, collagen degradation (Col 2 3/4C short collagen epitope), and the endogenous (pro‐) form of BMP‐7 protein. Knee joints that received rhBMP‐7 immediately after injury had small focal lesions at the injury site that did not progress into the surrounding cartilage. Joints that received BMP‐7 3 weeks after injury were improved and had limited progression compared to controls, but joints that received the protein 12 weeks after injury had no statistically significant improvement. These studies suggest that BMP‐7 may be chondroprotective after traumatic injury in patients if it is administered within 3 to 4 weeks of the index injury. The mechanism of protection after sublethal injury appeared to be an increased survival of chondrocytes that are able to participate in the repair process.

Gene Expression by Human Articular Chondrocytes Cultured in Alginate Beads

Culture of articular chondrocytes in alginate beads offers several advantages over culture in monolayer; cells retain their phenotype for 8 months or longer. Earlier studies of chondrocytes cultured in alginate concentrated on collagen and proteoglycan synthesis. However, gene expression by in situ hybridization (ISH) has not been investigated. The purposes of the present study on human chondrocytes were (a) to modify the ISH procedure for the alginate beads to examine the mRNA expression of α1 (II) procollagen, aggrecan, and two matrix metalloproteinases (MMP-3 and MMP-8) thought to be involved in cartilage matrix degradation, and (b) to compare expression in cultured chondrocytes with that in chondrocytes of intact human cartilage. The modifications made for ISH include the presence of CaCl2 and BaCl2 in the fixation and washing steps and exclusion of cetyl pyridinium chloride. By ISH we show that aggrecan, MMP-3, and MMP-8 are continuously expressed during 8 months of culture. The α1 (II) procollagen gene is expressed only during the first 2 months of culture and after 3 months its expression is undetectable, which is consistent with its absence in adult articular cartilage. By Western blotting, Type II collagen protein had been synthesized and deposited in both the cell-associated and further-removed matrix compartments at 7 and 14 days of culture. These data indicate that chondrocytes cultured in alginate beads could be preserved for immunohistochemistry and ISH and that culture of human chondrocytes in alginate beads may serve as a good model for studying cartilage-specific phenotype as well as factors that influence cartilage matrix turnover.

Osteogenic protein-1 (osteogenic protein-1/bone morphogenetic protein-7) inhibits degeneration and pain-related behavior induced by chronically compressed nucleus pulposus in the rat.

Study Design. To study the therapeutic efficacy of intradiscal injection of osteogenic protein-1 (OP-1) to reduce degeneration and associated discogenic pain. Objective. To evaluate if intradiscal injection of OP-1 can reverse disc degeneration and reduce hyperalgesia, a pain-related behavior. Summary of Background Data. We showed that induction of hyperalgesia was higher in rats exposed to compressed nucleus pulposus (NP). It has been reported that intradiscal injection of OP-1 stimulates synthesis of proteoglycans and collagen in normal intervertebral discs. Methods. Rats were divided into several groups. In the sham group, the rings of an Ilizarov-type apparatus were only applied to the tail without compression. In the compressed NP group, the apparatus was used to apply chronically compression to the tail. Four weeks after surgery, the NP group was subdivided into 3 groups: saline-treated and OP-1-treated, which was divided into 2 groups (i.e., the continuous compression OP-1 [COP-1] group, in which compression was continuously applied to the tail for 4 weeks after OP-1 treatment and the release compression OP-1 [ROP-1] group, in which compression was released at treatment. Either physiologic saline or OP-1 was injected into the instrumented NP. The treated NP was harvested and applied to the left lumbar nerve roots 4 weeks after injection. Hyperalgesia was measured up to 3 weeks after surgery. The degree of disc degeneration and the appearance of the extracellular matrix in the intervertebral discs were evaluated by histology. Results. Mechanical hyperalgesia was observed in the sham and saline groups, but not in the OP-1 treated group. In the saline group, NP cells became spindle-shaped. In the OP-1 group, the NP cells became swollen with vacuolated cytoplasm, and the content of the extracellular matrix was markedly increased. Conclusion. OP-1 injection into degenerative intervertebral disc resulted in the enhancement of the extracellular matrix and the inhibition of pain-related behavior.