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The terminal complement complex (TCC) has been described as a potential driver in the pathogenesis of posttraumatic osteoarthritis (PTOA). However, sublytic TCC deposition might also play a crucial role in bone development and regeneration. Therefore, we elucidated the effects of TCC on joint-related tissues using a rabbit PTOA model. In brief, a C6-deficient rabbit breed was characterized on genetic, protein, and functional levels. Anterior cruciate ligament transection (ACLT) was performed in C6-deficient (C6−/−) and C6-sufficient (C6+/−) rabbits. After eight weeks, the progression of PTOA was determined histologically. Moreover, the structure of the subchondral bone was evaluated by µCT analysis. C6 deficiency could be attributed to a homozygous 3.6 kb deletion within the C6 gene and subsequent loss of the C5b binding site. Serum from C6−/− animals revealed no hemolytic activity. After ACLT surgery, joints of C6−/− rabbits exhibited significantly lower OA scores, including reduced cartilage damage, hypocellularity, cluster formation, and osteophyte number, as well as lower chondrocyte apoptosis rates and synovial prostaglandin E2 levels. Moreover, ACLT surgery significantly decreased the trabecular number in the subchondral bone of C6−/− rabbits. Overall, the absence of TCC protected from injury-induced OA progression but had minor effects on the micro-structure of the subchondral bone. [ABSTRACT FROM AUTHOR]

Background: Cryotherapy is routinely administered after sports injuries of synovial joints. Although positive clinical effects on periarticular swelling and pain have been described, the effects on the cell biological activities of cartilage and synovial cells remain largely unknown so far. Hypothesis: Local hypothermia alleviates synovial reactions and prevents chondrocyte death as well as cartilage destructive processes after blunt cartilage trauma. Study Design: Controlled laboratory study. Methods: Human cartilage explants were impacted by a drop-tower apparatus (0.59 J) and cultured at 24 hours or 7 days in different temperature conditions (2 hours [short term], 16 hours [medium term], or throughout [long term] at 27°C; afterwards or throughout at 37°C). Besides, isolated human fibroblast-like synoviocytes (FLS) were stimulated with traumatized cartilage conditioned medium and cultured as mentioned above up to 4 days. The effects of hypothermia were evaluated by cell viability, gene expression, type II collagen synthesis and cleavage, as well as the release of matrix metalloproteinase (MMP)–2, MMP-13, and interleukin 6 (IL-6). Results: Seven days after trauma, hypothermic treatment throughout improved cell viability (short term: 10.1% [P = .016]; medium term: 6% [P = .0362]; long term: 12.5% [P = .0039]). Short-term hypothermia attenuated the expression of catabolic MMP-13 (mRNA: –2.2-fold [P = .0119]; protein: –2-fold [P = .0238]). Whereas type II collagen synthesis (1.7-fold [P = .0227]) was increased after medium-term hypothermia, MMP-13 expression (mRNA: –30.8-fold [P = .0025]; protein: –10.3-fold [P < .0001]) and subsequent cleavage of type II collagen (–1.1-fold [P = .0489]) were inhibited. Long-term hypothermia further suppressed MMP release (pro–MMP-2: –3-fold [P = .0222]; active MMP-2: −5.2-fold [P = .0183]; MMP-13: −56-fold [P < .0001]) and type II collagen breakdown (–1.6-fold [P = .0036]). Four days after FLS stimulation, hypothermia significantly suppressed the gene expression of matrix-destructive enzymes after medium-term (MMP-3: –4.1-fold [P = .0211]) and long-term exposure (a disintegrin and metalloproteinase with thrombospondin motifs 4 [ADAMTS4]: –4.3-fold [P = .0045]; MMP-3: –25.8-fold [P = .014]; MMP-13: –122-fold [P = .0444]) and attenuated IL-6 expression by trend. Conclusion: After blunt cartilage trauma, initial hypothermia for only 2 hours and/or 16 hours induced significant cell-protective and chondroprotective effects and promoted the anabolic activity of chondrocytes, while the expression of matrix-destructive enzymes by stimulated FLS was attenuated by prolonged hypothermia. Clinical Relevance: The findings of this preliminary ex vivo investigation indicate that optimized cryotherapy management after cartilage trauma might prevent matrix-degenerative processes associated with the pathogenesis of posttraumatic osteoarthritis. [ABSTRACT FROM AUTHOR]

Cartilage injury can trigger crucial pathomechanisms, including excessive cell death and expression of matrix-destructive enzymes, which contribute to the progression of a post-traumatic osteoarthritis ( PTOA). With the intent to create a novel treatment strategy for alleviating trauma-induced cartilage damage, we complemented a promising antioxidative approach based on cell and chondroprotective N-acetyl cysteine ( NAC) by chondroanabolic stimulation. Overall, three potential pro-anabolic growth factors - IGF-1, BMP7 and FGF18 - were tested comparatively with and without NAC in an ex vivo human cartilage trauma-model. For that purpose, full-thickness cartilage explants were subjected to a defined impact (0.59 J) and subsequently treated with the substances. Efficacy of the therapeutic approaches was evaluated by cell viability, as well as various catabolic and anabolic biomarkers, representing the present matrix turnover. Although monotherapy with NAC, FGF18 or BMP7 significantly prevented trauma-induced cell dead and breakdown of type II collagen, combination of NAC and one of the growth factors did not yield significant benefit as compared to NAC alone. IGF-1, which possessed only moderate cell protective and no chondroprotective qualities after cartilage trauma, even reduced NAC-mediated cell and chondroprotection. Despite significant promotion of type II collagen expression by IGF-1 and BMP7, addition of NAC completely suppressed this chondroanabolic effect. All in all, NAC and BMP7 emerged as best combination. As our findings indicate limited benefits of the simultaneous multidirectional therapy, a sequential application might circumvent adverse interferences, such as suppression of type II collagen biosynthesis, which was found to be reversed 7 days after NAC withdrawal. [ABSTRACT FROM AUTHOR]

Background: Sport injuries of the knee often lead to posttraumatic arthritis. In addition to direct damage of the cartilage, trauma-associated intra-articular bleeding may cause hemarthrosis. Both blood exposure and trauma are known to induce cell death and inflammation and to enhance proteoglycan release in cartilage. Hypothesis: Blood exposure increases chondrocyte death as well as inflammatory and degenerative processes in traumatized cartilage. Study Design: Controlled laboratory study. Methods: Human macroscopically intact osteoarthritic (OA) cartilage explants were impacted by a drop-tower system (0.59 J) and cultivated with or without 10% blood. Interactive effects were studied concerning cell survival, gene expression, and the release of mediators over 24 hours and 96 hours. To evaluate the effects of trauma and hemarthrosis in vivo, a newly established blunt cartilage trauma model in the rabbit was used. Treatment of the knee joints of mature New Zealand White rabbits consisted of the following groups: control (C), arthrotomy (A), arthrotomy with cartilage trauma (AT; 1.0 J), and arthrotomy with cartilage trauma and blood injection (ATH). After 1 and 12 weeks, inflammatory mediators in the synovial fluid and histological changes of the cartilage were determined, and immunohistological staining was performed. Results: The in vitro studies revealed a significant additional or synergistic effect of blood exposure on trauma-induced chondrocyte death, interleukin (IL)–1β and prostaglandin-E2 (PGE2) release, and matrix metalloproteinase (MMP)/pro-MMP level. Singular arthrotomy in vivo induced a temporary inflammation. Histologically, cartilage trauma caused significant OA changes that were not aggravated by an additional hemarthrosis. Trauma led to a persistent deposition of terminal complement complex (TCC), being enhanced by hemarthrosis. However, trauma-induced formation of osteophytes and arthrotomy-induced elevation of tumor necrosis factor–α release were reduced by hemarthrosis. Conclusion: While blood exposure clearly aggravated trauma-induced OA processes in the in vitro model, a singular blood injection revealed heterogeneous effects in vivo, enhancing TCC deposition but reducing trauma-induced osteophyte formation while the histological score of traumatized cartilage was not further impaired. Clinical Relevance: The results of this study indicate that a singular, limited bleeding event might not exacerbate early trauma-induced cartilage degeneration in joint injuries. An early removal of intra-articular blood may not prevent the final resulting cartilage damage. [ABSTRACT FROM AUTHOR]

Intermediate filament networks are part of the cytoskeleton and protect cellular integrity during large deformations. In cells from mesenchymal lineage the cytoskeleton is centrally involved in signal transduction, thereby influencing differentiation. We study the ultrastructure of IF networks in three human mesenchymal cell types, namely undifferentiated mesenchymal stem cells, chondrocytes, and osteoblasts. In order to capture the high morphological variability of IF networks we apply techniques from image analysis to extract the network graph from 2D scanning electron microscopy (SEM) images in a fully automatic way, which allows for a high-throughput analysis of SEM data. The extracted network graphs are analyzed by techniques from spatial statistics to detect differences in network morphology between different cell types and infer strategies of network remodeling used by the cells to adapt their mechanical properties during migration and differentiation. [ABSTRACT FROM AUTHOR]

Background: Clinically oriented and easy to handle animal models are urgently needed to test pharmacologic treatment of cartilage trauma to reduce the resulting tissue damage by chondrocyte apoptosis and induction of matrix-degrading enzymes. Aim: To develop a biomechanically defined cartilage trauma model. Material and Methods: We constructed a novel trauma device that allows biomechanically defined force application to the load-bearing region of the medial and lateral femoral condyles in adult rabbits. The fixation to the femur was specially designed to avoid uncontrolled influx of blood into the joint. The device was tested on the articular femoral surface of cadaveric rabbits. Results: At a lower energy (1.0 J), the tests showed that superficial and partially deep fissuring, partial necrosis of the chondrocytes, and early proteoglycan loss occurred at the region of impact. Subchondral fractures could be excluded by micro CT. At higher energy (≥1.4 J), we observed more pronounced deep fissuring and in some cases complete shearing of the articular cartilage from the subchondral bone. Conclusion: Our model represents an easy to use method to create a biomechanically defined cartilage trauma and offers some advantages with respect to handling under aseptic surgical conditions and prevention of uncontrolled intra-articular bleeding from the bone marrow compartment for pharmacologic studies. [ABSTRACT FROM AUTHOR]

The aim of this study was to determine if extracorporeal shock wave therapy (ESWT) in vivo affects the structural integrity of articular cartilage. A single bout of ESWT (1500 shock waves of 0.5 mJ/mm2) was applied to femoral heads of 18 adult Sprague-Dawley rats. Two sham-treated animals served as controls. Cartilage of each femoral head was harvested at 1, 4, or 10 weeks after ESWT ( n = 6 per treatment group) and scored on safranin-O-stained sections. Expression of tenascin-C and chitinase 3-like protein 1 (Chi3L1) was analyzed by immunohistochemistry. Quantitative real-time polymerase chain reaction (PCR) was used to examine collagen (II)α1 (COL2A1) expression and chondrocyte morphology was investigated by transmission electron microscopy no changes in Mankin scores were observed after ESWT. Positive immunostaining for tenascin-C and Chi3L1 was found up to 10 weeks after ESWT in experimental but not in control cartilage. COL2A1 mRNA was increased in samples 1 and 4 weeks after ESWT. Alterations found on the ultrastructural level showed expansion of the rough-surfaced endoplasmatic reticulum, detachment of the cell membrane and necrotic chondrocytes. Extracorporeal shock waves caused alterations of hyaline cartilage on a molecular and ultrastructural level that were distinctly different from control. Similar changes were described before in the very early phase of osteoarthritis (OA). High-energy ESWT might therefore cause degenerative changes in hyaline cartilage as they are found in initial OA. © 2010 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 28:1050-1056, 2010 [ABSTRACT FROM AUTHOR]