The alarmins high mobility group box protein 1 and S100A8/A9 display different inflammatory profiles after acute knee injury.

OBJECTIVE: To compare the concentrations of high mobility group box 1 protein (HMGB1) and S100A8/A9 in synovial fluid between patients with knee injuries and osteoarthritis (OA), and knee healthy subjects. To investigate associations of alarmin levels with different joint injuries and with biomarkers of inflammation, Wnt signaling, complement system, bone and cartilage degradation. METHODS: HMGB1 and S100A8/A9 were measured in synovial fluid by immunoassays in patients with knee injuries, with OA and from knee healthy subjects, and were related to time from injury and with biomarkers obtained from previous studies. Hierarchical cluster and enrichment analyses of biomarkers associated to HMGB1 and S100A8/A9 were performed. RESULTS: The synovial fluid HMGB1 and S100A8/A9 concentrations were increased early after knee injury; S100A8/A9 levels were negatively associated to time after injury and was lower in the old compared to recent injury group, while HMGB1 was not associated to time after injury. The S100A8/A9 levels were also increased in OA. The initial inflammatory response was similar between the alarmins, and HMGB1 and S100A8/A9 shared 9 out of 20 enriched pathways. The alarmins displayed distinct response profiles, HMGB1 being associated to cartilage biomarkers while S100A8/A9 was associated to proinflammatory cytokines. CONCLUSIONS: HMGB1 and S100A8/A9 are increased as an immediate response to knee trauma. While they share many features in inflammatory and immunoregulatory mechanisms, S100A8/A9 and HMGB1 are associated to different downstream responses, which may have impact on the OA progression after acute knee injuries.

E8002 Reduces Adhesion Formation and Improves Joint Mobility in a Rat Model of Knee Arthrofibrosis.

Knee arthrofibrosis is a common complication of knee surgery, caused by excessive scar tissue, which results in functional disability. However, no curative treatment has been established. E8002 is an anti-adhesion material that contains L-ascorbic acid, an antioxidant. We aimed to evaluate the efficacy of E8002 for the prevention of knee arthrofibrosis in a rat model, comprising injury to the surface of the femur and quadriceps muscle 1 cm proximal to the patella. Sixteen male, 8-week-old Sprague Dawley rats were studied: in the Adhesion group, haemorrhagic injury was induced to the quadriceps and bone, and in the E8002 group, an adhesion-preventing film was implanted between the quadriceps and femur after injury. Six weeks following injury, the restriction of knee flexion owing to fibrotic scarring had not worsened in the E8002 group but had worsened in the Adhesion group. The area of fibrotic scarring was smaller in the E8002 group than in the Adhesion group (p < 0.05). In addition, the numbers of fibroblasts (p < 0.05) and myofibroblasts (p < 0.01) in the fibrotic scar were lower in the E8002 group. Thus, E8002 reduces myofibroblast proliferation and fibrotic scar formation and improves the range of motion of the joint in a model of knee injury.

Anti-Inflammatory Therapeutic Approaches to Prevent or Delay Post-Traumatic Osteoarthritis (PTOA) of the Knee Joint with a Focus on Sustained Delivery Approaches.

Inflammation plays a central role in the pathogenesis of knee PTOA after knee trauma. While a comprehensive therapy capable of preventing or delaying post-traumatic osteoarthritis (PTOA) progression after knee joint injury does not yet clinically exist, current literature suggests that certain aspects of early post-traumatic pathology of the knee joint may be prevented or delayed by anti-inflammatory therapeutic interventions. We discuss multifaceted therapeutic approaches that may be capable of effectively reducing the continuous cycle of inflammation and concomitant processes that lead to cartilage degradation as well as those that can simultaneously promote intrinsic repair processes. Within this context, we focus on early disease prevention, the optimal timeframe of treatment and possible long-lasting sustained delivery local modes of treatments that could prevent knee joint-associated PTOA symptoms. Specifically, we identify anti-inflammatory candidates that are not only anti-inflammatory but also anti-degenerative, anti-apoptotic and pro-regenerative.

Morphological Changes in Articular Cartilage and Free-Radical Lipid Peroxidation in Rats with Posttraumatic Osteoarthrosis.

Using the rat model of posttraumatic osteoarthrosis of the knee joint induced by surgical transection of their anterior cruciate ligaments, we showed that irreversible loss of hyaluronan by the extracellular matrix of the joint cartilage tissue against the background of oxidative stress accompanied by accumulation of intermediate LPO products in blood serum and formation of thiol system incompetence was one of the key patterns of dystrophic degeneration of the cartilage tissue. Considerable metabolic shifts were associated with structural modification of the articular hyaline cartilage: its thinning and a decrease of chondrocyte density and their abnormal spatial distribution in the matrix with predominance of solitary isolated cells with signs of karyopyknosis and karyolysis.

Metabolism of Subchondral Bone Tissue in Posttraumatic Osteoarthrosis in Rats.

In rats with modeled posttraumatic knee osteoarthrosis, negative changes in subchondral bone metabolism were revealed: a tendency to an increase in osteocalcin concentration, a decrease in sclerostin and osteoprotegerin levels, and a significant increase in FGF-23 concentration accompanied by a slight elevation of inorganic phosphorous and significant increase in total calcium levels in comparison with the corresponding parameters in intact controls. These findings demonstrate crucial importance of structural integrity of the subchondral bone, because its protection improves the results of reconstructive therapy for local cartilage defects.

Lubricin in experimental and naturally occurring osteoarthritis: a systematic review.

OBJECTIVE: Lubricin is increasingly being evaluated as an outcome measure in studies investigating post-traumatic and naturally occurring osteoarthritis. However, there are discrepancies in results, making it unclear as to whether lubricin is increased, decreased or unchanged in osteoarthritis. The purpose of this study was to review all papers that measured lubricin in joint injury or osteoarthritis in order to draw conclusions about lubricin regulation in joint disease. DESIGN: A systematic search of the Pubmed, Web of Knowledge, and EBSCOhost databases for papers was performed. Inclusion criteria were in vivo studies that measured lubricin in humans or animals with joint injury, that investigated lubricin supplementation in osteoarthritic joints, or that described the phenotype of a lubricin knock-out model. A methodological assessment was performed. RESULTS: Sixty-two studies were included, of which thirty-eight measured endogenous lubricin in joint injury or osteoarthritis. Nineteen papers found an increase or no change in lubricin and nineteen reported a decrease. Papers that reported a decrease in lubricin were cited four times more often than those that reported an increase. Fifteen papers described lubricin supplementation, and all reported a beneficial effect. Eleven papers described lubricin knock-out models. CONCLUSIONS: The human literature reveals similar distributions of papers reporting increased lubricin as compared to decreased lubricin in osteoarthritis. The animal literature is dominated by reports of decreased lubricin in the rat anterior cruciate ligament transection model, whereas studies in large animal models report increased lubricin. Intra-articular lubricin supplementation may be beneficial regardless of whether lubricin increases or decreases in OA.

Explant models for meniscus metabolism, injury, repair, and healing.

Purpose/Aim: Knee meniscus is a wedge-shaped fibrocartilaginous tissue, playing important roles in maintaining joint stability and function. Injuries to the meniscus, particularly with the avascular inner third zone, hardly heal and frequently progress into structural breakdown, followed by the initiation of osteoarthritis. As the importance of meniscus in joint function and diseases is being recognized, the field of meniscus research is growing. Not only development, biology, and metabolism but also injury, repair, and healing of meniscus are being actively investigated. As meniscus functions as an integrated unit of a knee joint, in vivo models with various species have been the predominant method for studying meniscus pathophysiology and for testing healing/regeneration strategies. However, in vivo models for meniscus studies suffer from low reproducibility and high cost. To complement the limitations of in vivo animal models, several types of meniscus explants have been applied as highly controlled, standardized in vitro models to investigate meniscus metabolism, pathophysiology, and repair or regeneration process. This review summarizes and compares the existing meniscus explant models. We also discuss the advantages and disadvantages of each explant model.Conclusion: Despite few outstanding challenges, meniscus explant models have potential to serve as an effective tool for investigations of meniscus metabolism, injury, repair and healing.

Microarray analysis of cartilage: comparison between damaged and non-weight-bearing healthy cartilage.

AIM: A limited healing response to focal cartilage lesions is frequently encountered in the clinical cartilage pathology. This study compares the gene expression patterns of damaged and undamaged regions of cartilage obtained from the same patient with focal cartilage lesions. The aim of this study is to provide new genes and proteins, which may be a potential future target of research. METHODS: During the autologous chondrocyte implantation (MACI) surgery, cartilage tissues (healthy non-weight bearing and Damaged-lesion side) were obtained from 10 patients with knee focal cartilage lesions. The degeneration status of the cartilage was characterized according to ICRS criteria. Whole genome microarray gene expression profiling was performed and some of the differentially regulated genes were validated with RT-PCR. RESULTS: Damaged and undamaged non-weight bearing cartilage showed distinct gene expression profiles. Genes involved in cell signaling, matrix degradation, hypoxia, and the inflammatory response showed significant up- or down-regulation. In the focal lesions, expression of genes such as HIF1α, TIMP-2, EID1, EID2, NCOA3, NBR1, SP100, and HSP90AA1 was significantly higher compared to healthy non-weight bearing cartilage from the same joint, whereas TIMP-4 was lower. CONCLUSION: The genes examined in this study differ distinctly between focal cartilage (ICRS 3-4) lesions and undamaged sites of the same joint. We believe that the data set forth in this study may be used for clinical purposes and be a guide in the development of new biological approaches for therapy.

The effects of age on the severity of joint damage and intra-articular inflammation following a simulated medial meniscus injury in 3, 6, and 9 month old male rats.

Purpose: Aging is a known risk factor for osteoarthritis (OA). Several transgenic rodent models have been used to investigate the effects of accelerated or delayed aging in articular joints. However, age-effects on the progression of post-traumatic OA are less frequently evaluated. The objective of this study is to evaluate how animal age affects the severity of intra-articular inflammation and joint damage in the rat medial collateral ligament plus medial meniscus transection (MCLT+MMT) model of knee OA.Methods: Forty-eight, male Lewis rats were aged to 3, 6, or 9 months old. At each age, eight rats received either an MCLT+MMT surgery or a skin-incision. At 2 months post-surgery, intra-articular evidence of CTXII, IL1ß, IL6, TNFα, and IFNγ was evaluated using a multiplex magnetic capture technique, and histological evidence of OA was assessed via a quantitative histological scoring technique.Results: Elevated levels of CTXII and IL6 were found in MCLT+MMT knees relative to skin-incision and contralateral controls; however, animal age did not affect the severity of joint inflammation. Conversely, histological investigation of cartilage damage showed larger cartilage lesion areas, greater width of affected cartilage, and more evidence of hypertrophic cartilage damage in MCLT+MMT knees with age.Conclusions: These data indicate the severity of cartilage damage subsequent to MCLT+MMT surgery is related to the rat's age at the time of injury. However, despite greater levels of cartilage damage, the level of intra-articular inflammation was not necessarily affected in 3, 6, and 9 month old male rats.

Characterization of Synovial Cytokine Patterns in Bucket-Handle and Posterior Horn Meniscal Tears.

The specific etiology of meniscal tears, including the mechanism of lesion, location, and orientation, is considered for its contribution to subsequent joint cytokine responsiveness, healing outcomes, and by extension, appropriate lesion-specific surgical remediation. Meniscal repair is desirable to reduce the probability of development of posttraumatic osteoarthritis (PTOA) which is strongly influenced by the coordinate generation of pro- and anti-inflammatory cytokines by the injured cartilage. We now present biochemical data on variation in cytokine levels arising from two particular meniscal tears: bucket-handle (BH) and posterior horn (PH) isolated meniscal tears. We selected these two groups due to the different clinical presentations. We measured the concentrations of TNF-α, IL-1ß, IL-6, IL-8, and IL-10 in knee synovial fluid of 45 patients with isolated meniscal lesions (BH tear, n = 12; PH tear, n = 33). TNF-α levels were significantly (p < 0.05) greater in the BH group compared with the PH group, whereas IL-1ß levels were significantly greater (p < 0.05) in the PH group compared with the BH group. Both BH and PH groups were consistent in presenting a positive correlation between concentrations of IL-6 and IL-1ß. A fundamental difference in IL-10 responsiveness between the two groups was noted; specifically, levels of IL-10 were positively correlated with IL-6 in the BH group, whereas in the PH group, levels of IL-10 were positively correlated with IL-1ß. Collectively, our data suggest a possible influence of the meniscal tear pattern to the articular cytokine responsiveness. This differential expression of inflammatory cytokines may influence the risk of developing PTOA in the long term.

IGF-1 and BMP-7 synergistically stimulate articular cartilage repairing in the rabbit knees by improving chondrogenic differentiation of bone-marrow mesenchymal stem cells.

Knee injury is known as a frequently occurred damage related to sports, which may affect the function of cartilage. This study aims to explore whether Insulin-like growth factor 1 (IGF-1) and bone morphogenetic protein-7 (BMP-7)-modified bone-marrow mesenchymal stem cells (BMSCs) affect the repair of cartilage damage found in the knee. Primarily, BMSCs were treated with a series of pEGFP-C1, IGF-1, and BMP-7, followed by determination of IGF-1 and BMP-7 expression. A rabbit cartilage defect model was also established. Afterfward, cell morphology, viability, cartilage damage repair effect, and expression of collagen I and collagen II at the 6th and the 12th week were measured. BMSCs treated with pEGFP-C1/IGF-1, pEGFP-C1/BMP-7, and pEGFP-C1/BMP-7-IGF-1 exhibited elevated expression of BMP-7 and IGF-1. Besides, BMSCs in the P10 generation displayed decreased cell proliferation. Moreover, BMSCs treated with IGF-1, BMP-7, and IGF-1-BMP-7 showed reduced histological score and collagen I expression while elevated collagen II expression, as well as better repair effect, especially in those treated with IGF-1-BMP-7. Collectively, these results demonstrated a synergistic effect of IGF-1 and BMP-7 on the BMSC chondrogenic differentiation on the articular cartilage damage repair in the rabbit knees, highlighting its therapeutic potential for the treatment of articular cartilage damage.

CCL22 is a biomarker of cartilage injury and plays a functional role in chondrocyte apoptosis.

BACKGROUND: Osteoarthritis (OA) is one of the leading causes of disability worldwide. Previous history of knee injury is a significant risk factor for OA. It has been established that low-level chronic inflammation plays a pivotal role in the onset and pathogenesis of OA. The primary aim of this research was to determine if a history of knee joint injury is associated with systemic inflammation. A secondary aim was to determine if systemic inflammation is related to knee pain and joint structure. METHODS: Differences in serum cytokine association networks, knee joint structural changes (MRI), and self-reported pain (i.e., Knee Injury and Osteoarthritis Outcome Score Pain subscale, KOOSPAIN and Intermittent and Constant Osteoarthritis Pain score, ICOAP) between individuals who had sustained a youth (aged 15-26 years) sport-related knee injury 3-10 years previously and age- and sex-matched controls were examined. Proteins of interest were also examined in an OA rat model. RESULTS: Cytokine association networks were found to differ significantly between study groups, yet no significant associations were found between networks and KOOSPAIN or MRI-defined OA. A group of cytokines (MCP1/CCL2, CCL22 and TNFα) were differentially associated with other cytokines between study groups. In a pre-clinical rat OA model, serum CCL22 levels were associated with pain (r = 0.255, p = 0.045) and structural changes to the cartilage. CCL22 expression was also observed in human OA cartilage and furthermore, CCL22 induced apoptosis of isolated human chondrocytes. DISCUSSION: These results suggest that CCL22 may be an early factor in the onset/pathogenic process of cartilage degeneration and/or related to pain OA.

Cell origin, volume and arrangement are drivers of articular cartilage formation, morphogenesis and response to injury in mouse limbs.

Limb synovial joints are composed of distinct tissues, but it is unclear which progenitors produce those tissues and how articular cartilage acquires its functional postnatal organization characterized by chondrocyte columns, zone-specific cell volumes and anisotropic matrix. Using novel Gdf5CreERT2 (Gdf5-CE), Prg4-CE and Dkk3-CE mice mated to R26-Confetti or single-color reporters, we found that knee joint progenitors produced small non-migratory progenies and distinct local tissues over prenatal and postnatal time. Stereological imaging and quantification indicated that the columns present in juvenile-adult tibial articular cartilage consisted of non-daughter, partially overlapping lineage cells, likely reflecting cell rearrangement and stacking. Zone-specific increases in cell volume were major drivers of tissue thickening, while cell proliferation or death played minor roles. Second harmonic generation with 2-photon microscopy showed that the collagen matrix went from being isotropic and scattered at young stages to being anisotropic and aligned along the cell stacks in adults. Progenitor tracing at prenatal or juvenile stages showed that joint injury provoked a massive and rapid increase in synovial Prg4+ and CD44+/P75+ cells some of which filling the injury site, while neighboring chondrocytes appeared unresponsive. Our data indicate that local cell populations produce distinct joint tissues and that articular cartilage growth and zonal organization are mainly brought about by cell volume expansion and topographical cell rearrangement. Synovial Prg4+ lineage progenitors are exquisitely responsive to acute injury and may represent pioneers in joint tissue repair.

Definition of a Critical Size Osteochondral Knee Defect and its Negative Effect on the Surrounding Articular Cartilage in the Rat.

BACKGROUND: Joint trauma is predisposing to the incidence of osteoarthritis (OA) of the knee. There is a limited knowledge on the impact of posttraumatic osteochondral defects on the whole joint. This study was designed to define a critical size osteochondral defect in the knee of rats and to investigate a possible association between osteochondral defects and degeneration of the surrounding joint surface. METHODS: Cylindrical osteochondral defects of different sizes were created in the knee joint of rats. The natural course of these lesions was studied by macroscopic observation, histology, and immunohistochemistry. Gene expression of the articular cartilage surrounding the defects in vivo and of articular chondrocytes cultured in vitro in IL1ß and fibroblast growth factor 2 (FGF2) supplemented media was evaluated by quantitative polymerase chain reaction (qPCR). RESULTS: In defects of 0.9 mm diameter, spontaneous joint surface healing was observed but also upward advancing of the subchondral bone plate at 16 weeks. Larger 1.4 mm diameter defects were critical size, not resulting in successful healing at any time point. Importantly, the articular cartilage surrounding the defects expressed FGF2 and IL1ß, but not ACAN and Col2. Chondrocytes cultured in IL1ß and FGF2 supplemented media lost the natural fibroblast growth factor receptors - FGFr1/FGFr3 balance and showed decreased viability. CONCLUSIONS: A critical size osteochondral defect was defined as 1.4 mm in diameter in rat. Subchondral bone plate advancement occured rapidly. The articular cartilage surrounding osteochondral defects showed catabolic activity with expression of IL1ß, FGF2 and a disturbed FGFr1/FGFr3 balance, potentially initiating a process of early osteoarthritic disease.

Biomarker Changes in Anterior Cruciate Ligament-Deficient Knees Compared With Healthy Controls.

PURPOSE: To establish how synovial fluid biomarker concentrations change in patients after anterior cruciate ligament (ACL) tears, with and without associated cartilage injury, with comparisons made to healthy controls. METHODS: Patients were prospectively enrolled between January 2013 and December 2014. Inclusion criteria included any patient undergoing knee arthroscopy. Patients with a confirmed ACL tear were allocated to either the ACL tear with cartilage injury group or the ACL tear without cartilage injury group based on intraoperative assessment. Patients who underwent an arthroscopic procedure with no injury history or symptoms in their contralateral knee were asked to provide samples to serve as healthy controls. These subjects may or may not have been the same ones with noted ACL pathology. The concentrations of 20 biomarkers were determined using a multiplex magnetic bead immunoassay. Biomarker concentrations were then compared between the 3 study groups (ACL tears with and without cartilage injury, and uninjured contralateral knees) using an analysis of variance test with pairwise comparisons. The minimal clinically important difference was calculated based on the standard error of measurement. RESULTS: The study included synovial fluid samples from 134 knees: 34 ACL tears without cartilage injury (mean age 34.0 years), 28 ACL tears with cartilage injury (mean age 36.3 years), and 72 healthy controls (mean age 41.1 years). Analysis of variance testing showed significant differences among groups for matrix metalloproteinase-3 (F = 81.8; P < .001), tissue inhibitor of metalloproteinase (TIMP)-1 (F = 7.9; P ≤ .001), TIMP-2 (F = 4.5; P = .015); fibroblast growth factor-2 (F = 4.9; P = .011), interleukin-6 (F = 8.2; P = .001), and macrophage inflammatory protein-1 beta (F = 7.3; P = .001). Pairwise comparisons showed no significant differences between ACL tears with, and without cartilage injury, but did show that both groups of ACL tears had significantly higher concentrations of (first P value = ACL tears with and then ACL tears without cartilage injury): matrix metalloproteinase-3 (P < .001; P < .001), TIMP-1 (P < .001; P = .011), interleukin-6 (P = .009; P = .038), and macrophage inflammatory protein-1 beta (P = .003; P = .045) compared with contralateral controls. ACL tears without associated cartilage damage had significantly lower concentrations of TIMP-2 (P = .011) and fibroblast growth factor-2 (P = .014) compared with controls. All biomarker concentration differences that reached statistical significance were also larger than calculated minimal clinically important differences. CONCLUSIONS: The current study identified 6 pro- and anti-inflammatory synovial fluid biomarkers whose concentrations after ACL injury were significantly different compared with uninjured controls. No significant differences in synovial fluid biomarker concentrations were seen between ACL injured knees with and without associated cartilage damage. LEVEL OF EVIDENCE: Level III, retrospective comparative study of prospectively gathered data.

Molecular changes indicative of cartilage degeneration and osteoarthritis development in patients with anterior cruciate ligament injury.

BACKGROUND: Anterior cruciate ligament (ACL) tear is considered a risk factor for osteoarthritis development. The purpose of our study was to investigate the expression levels of the apoptotic enzyme caspase 3, pro-inflammatory cytokines interleukin-1ß (IL-1ß) and interleukin-6 (IL-6) and degrading enzyme matrix metalloproteinase 13 (MMP-13), all indicative of cartilage degeneration and osteoarthritis development in patients' chondrocytes after ACL rupture. METHODS: We investigated the correlation between grade of cartilage degradation and time from injury or patients' age. IL-1ß, IL-6 and MMP-13 mRNA expression levels were investigated in normal (n = 4) and chondrocytes from patients with ACL rupture (n = 33) using real-time polymerase chain reaction (PCR). Moreover, MMP-13 and caspase-3 protein expression levels were evaluated by western blot analysis. Trend analysis and correlation coefficient were performed to derive the relations between gene expression (MMP13, IL-6, IL-1ß) and grading of cartilage defects and between gene expression (MMP13, IL-6, IL-1ß) and patients' age, respectively. RESULTS: Correlations were established between grade of cartilage degradation and time from injury. MMP-13, IL-6, IL-1ß and caspase 3 expression levels were significantly upregulated in chondrocytes from ACL-deficient knee compared to normal. Among the patients with ACL-deficient knees, a significant upregulation of MMP-13 was observed in patients with ACL-rupture > 18 months from the time of injury to arthroscopy compared to patients with ACL-injury up to 18 months, whereas IL-6 and IL-1ß expression was higher in chondrocytes from patients with more than 10 months ACL injury compared to those that underwent surgery within the first 10 months after injury. Νο association was observed between IL-1ß, IL-6 and MMP-13 expression levels and cartilage defects or patients' age. CONCLUSION: Our results showed that increased levels of apoptotic, inflammatory and catabolic factors in chondrocytes are associated with time from injury and could contribute to cartilage degradation and osteoarthritis development after ACL rupture.

Correlation of Synovial Fluid Biomarkers With Cartilage Pathology and Associated Outcomes in Knee Arthroscopy.

PURPOSE: To correlate the intraoperative concentrations of 20 synovial fluid biomarkers with preoperative symptoms, intraoperative findings, and postoperative outcomes in patients undergoing knee arthroscopy, with comparisons made to samples obtained from asymptomatic knees. METHODS: Synovial fluid samples were obtained from 81 patients undergoing knee arthroscopy meeting the inclusion criteria, which included 70 samples from operative knees and 32 samples from contralateral knees. Preoperatively, baseline data obtained from clinical questionnaires including a visual analog scale (VAS) score, the Lysholm score, and the Knee Injury and Osteoarthritis Outcome Score-Physical Function Short Form were recorded. Synovial fluid was collected from both the operative knee and asymptomatic contralateral knee. Synovial fluid was stored with a protease inhibitor at -80°C until analysis. Intraoperative findings, procedures performed, and International Cartilage Repair Society (ICRS) cartilage status scores in all operative knees were documented. The concentrations of the following 20 biomarkers were measured using a multiplex magnetic bead immunoassay: matrix metalloproteinase (MMP) 3; MMP-13; tissue inhibitor of metalloproteinase (TIMP) 1; TIMP-2; TIMP-3; TIMP-4; fibroblast growth factor 2; eotaxin; interferon γ; interleukin (IL) 10; platelet-derived growth factor BB; IL-1 receptor antagonist; IL-1ß; IL-6; monocyte chemotactic protein 1 (MCP-1); macrophage inflammatory protein 1α; macrophage inflammatory protein 1ß; RANTES (regulated upon activation, normal T cell expressed and secreted); tumor necrosis factor α; and vascular endothelial growth factor. Clinical outcome scores were obtained in 83% of patients at a mean of 17 months' follow-up postoperatively. Analysis of variance and Pearson correlation analysis were performed to determine statistical significance between preoperative data, intraoperative findings, postoperative outcomes, and synovial fluid biomarker concentrations compared with asymptomatic contralateral knees. RESULTS: Analysis was performed on 70 operative and 32 contralateral samples. There were strong positive correlations between ICRS score and age, symptom duration, VAS score, and Knee Injury and Osteoarthritis Outcome Score-Physical Function Short Form. A strong positive correlation was found between MCP-1 and IL-6 concentrations, intraoperative ICRS score, and continued pain at the time of final follow-up. MCP-1 and IL-6 were the strongest predictors of severe cartilage lesions, whereas IL-1 receptor antagonist was inversely related. MMP-3 levels were consistently elevated in all operative samples and directly correlated to increased preoperative VAS scores. RANTES, vascular endothelial growth factor, and platelet-derived growth factor BB were the strongest predictors of postoperative improvement at final follow-up regardless of injury and cartilage status. CONCLUSIONS: Synovial fluid biomarkers have the capacity to reflect the intra-articular environment before surgery and potentially predict postoperative clinical outcomes. Recognition of key molecular players may yield future therapeutic targets, and large clinical trials exploring these discoveries are anticipated. LEVEL OF EVIDENCE: Level III, therapeutic case-control study.

Type II collagen C2C epitope in human synovial fluid and serum after knee injury--associations with molecular and structural markers of injury.

PURPOSE: Investigate in a cross-sectional study time-dependent changes of synovial fluid type II collagen epitope C2C concentrations after knee injury and correlate to other joint injury biomarkers. METHODS: Synovial fluid samples were aspirated between 0 days and 7 years after injury (n = 235). Serum was collected from 71 of the knee injured patients. Synovial fluid from 8 knee-healthy subjects was used as reference. C2C was quantified by immunoassay and structural injury was determined from magnetic resonance images (MRI) of the injured knee acquired 1-38 days after injury (n = 98). Additional joint injury biomarker results were from earlier investigations of the same samples. RESULTS: Synovial fluid C2C concentrations were higher in injured knees than in knees of reference subjects from 1 day up to 7 years after injury. C2C concentrations in synovial fluid and serum were correlated (r = 0.403, P < 0.001). In synovial fluid from subjects early after injury (0-33 days), C2C concentrations were correlated with cross-linked C-telopeptide of type II collagen (r = 0.444, P = 0.003), ARGS-aggrecan (r = 0.337, P < 0.001), osteocalcin (r = 0.345, P < 0.001), osteopontin (r = 0.371, P < 0.001) and IL-8 (r = -0.385, P < 0.001), but not with structural joint injury as visualized on MRI. CONCLUSION: The increased levels of synovial fluid C2C after injury, together with the associations seen with several other injury-related biomarkers, suggest that an acute knee injury is associated with an immediate and sustained local degradation of type II collagen.

Interleukin-6 and tumor necrosis factor alpha in synovial fluid are associated with progression of radiographic knee osteoarthritis in subjects with previous meniscectomy.

OBJECTIVE: To explore potential associations between proinflammatory cytokines in synovial fluid and progression of osteoarthritis (OA) in meniscectomized subjects. DESIGN: We studied 132 subjects on average 18 years after meniscectomy, with a second examination 4-10 years later. We measured concentrations of interleukin (IL)-6, -8 and tumor necrosis factor (TNF)-α by multiplex immunoassay, graded radiographic features of tibiofemoral and patellofemoral OA according to the Osteoarthritis Research Society International (OARSI) atlas, scored patient-reported outcomes using the Knee Injury and Osteoarthritis Outcome Score (KOOS), and used logistic regression (adjusted for age, gender, body mass index, and time between examinations) for assessment of associations. RESULTS: Higher first examination concentrations of IL-6 and TNF-α were associated with increased risk for subsequent osteophyte progression (odds ratios (OR); 95% confidence intervals 1.05; 1.00-1.09 and 1.35; 1.03-1.75). Higher second examination concentrations of TNF-α were associated with having progressed in loss of joint space (OR 1.70; 1.15-2.52) or having worsened in the activity of daily living subscale of KOOS (OR 1.50; 1.07-2.09) in the preceding years. Subjects with increasing concentrations of IL-6 or TNF-α between examinations were five times more likely to have progressed in joint space narrowing between the same examinations, as compared to those with stable or decreasing concentrations (OR 5.17; 1.54-17.32 and 5.01; 1.32-18.92). CONCLUSIONS: In subjects with previous meniscectomy, higher or over time increasing synovial fluid levels of IL-6 and TNF-α seems to be associated with increased risk for progression of radiographic OA.

Alterations in subchondral bone plate, trabecular bone and articular cartilage properties of rabbit femoral condyles at 4 weeks after anterior cruciate ligament transection.

OBJECTIVE: To quantify early osteoarthritic-like changes in the structure and volume of subchondral bone plate and trabecular bone and properties of articular cartilage in a rabbit model of osteoarthritis (OA) induced by anterior cruciate ligament transection (ACLT). METHODS: Left knee joints from eight skeletally mature New Zealand white rabbits underwent ACLT surgery, while the contralateral (CTRL) right knee joints were left unoperated. Femoral condyles were harvested 4 weeks after ACLT. Micro-computed tomography imaging was applied to evaluate the structural properties of subchondral bone plate and trabecular bone. Additionally, biomechanical properties, structure and composition of articular cartilage were assessed. RESULTS: As a result of ACLT, significant thinning of the subchondral bone plate (P < 0.05) was accompanied by significantly reduced trabecular bone volume fraction and trabecular thickness in the medial femoral condyle compartment (P < 0.05), while no changes were observed in the lateral compartment. In both lateral and medial femoral condyles, the equilibrium modulus and superficial zone proteoglycan (PG) content were significantly lower in ACLT than CTRL joint cartilage (P < 0.05). Significant alterations in the collagen orientation angle extended substantially deeper into cartilage from the ACLT joints in the lateral femoral condyle relative to the medial condyle compartment (P < 0.05). CONCLUSIONS: In this model of early OA, significant changes in volume and microstructure of subchondral bone plate and trabecular bone were detected only in the femoral medial condyle, while alterations in articular cartilage properties were more severe in the lateral compartment. The former finding may be associated with reduced joint loading in the medial compartment due to ACLT, while the latter finding reflects early osteoarthritic changes in the lateral compartment.