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Summary Objective To perform a molecular mechanism-based investigation of the chondroprotective potential of hylan G-F 20. Method The effects of hylan G-F 20 on IL-1β-induced glycosaminoglycan (GAG) depletion and matrix metalloproteinase (MMP) expression in bovine and human cartilage explants were evaluated. Three weekly intra-articular hylan G-F 20 or control injections were administered 4 weeks post-operatively to rabbits with surgically induced osteoarthritis (OA). Cartilage histopathologic scores and osteophyte size were evaluated at 1, 4, and 8 weeks post-injections. Histomorphometry and immunostaining were used to quantify cartilage area and type II collagen (Col II) intensity, and real-time polymerase chain reaction (PCR) was used to examine the mRNA levels of Col2A1 , MMP-13 , - 16 and IL-1β at 1 week. Results Hylan G-F 20 retained GAG in IL-1β-exposed bovine and human cartilage explants and abrogated IL-1β-mediated increases in MMP-1, -3, and -13 in human explant culture. Hylan G-F 20‒treated OA joints had significantly better cartilage integrity at 1 and 4 weeks post-treatment and significantly smaller osteophytes at 4 weeks compared with control. Col2A1 mRNA increased with hylan G-F 20 treatment, which correlated with a trend toward increased Col II immunostaining. MMP - 13 and - 16 mRNAs increased in OA cartilage, but were not significantly altered by hylan G-F 20. IL-1β mRNA was undetectable in cartilage and unaltered in the synovium. Conclusions Hylan G-F 20 improved cartilage integrity and decreased osteophyte formation in the rabbit model of OA. Our results suggest that hylan G-F 20 may stimulate cartilage repair by increasing Col II, and inhibit IL-1β-mediated matrix degradation by decreasing MMPs.

Summary Objective To compare the effects of glucosamine (GlcN), curcumin, and diacerein in immortalized human C-28/I2 chondrocytes at the cellular and the gene expression level. This study aimed to provide insights into the proposed beneficial effects of these agents and to assess the applicability of the C-28/I2 cell line as a model for the evaluation of chondroprotective action. Methods Interleukin-1beta (IL-1β)-stimulated C-28/I2 cells were cultured in the presence of GlcN, curcumin, and diacerein prior to the evaluation of parameters such as viability, morphology and proliferation. The impact of GlcN, curcumin, and diacerein on gene expression was determined using quantitative real-time RT-PCR (qPCR). Results At the transcriptional level, 5mM GlcN and 50μM diacerein increased the expression of cartilage-specific genes such as aggrecan (AGC) and collagen type II (COL2), while reducing collagen type I (COL1) mRNA levels. Moreover, the IL-1β-mediated shift in gene expression pattern was antagonized by GlcN and diacerein. These effects were associated with a significant reduction in cellular proliferation and the development of chondrocyte-specific cell morphology. In contrast, curcumin was not effective at lower concentrations but even damaged the cells at higher amounts. Conclusions Both GlcN and diacerein promoted a differentiated chondrocytic phenotype of immortalized human C-28/I2 chondrocytes by altering proliferation, morphology, and COL2/COL1 mRNA ratios. Moreover, both agents antagonized inhibitory effects of IL-1β by enhancing AGC and COL2 as well as by reducing COL1 mRNA levels.

Summary Objective Articular cartilage may experience iatrogenic injury during routine orthopaedic/arthroscopic procedures. This could cause chondrocyte death, leading to cartilage degeneration and posttraumatic osteoarthritis. In an in vitro cartilage injury model, chondrocyte death was reduced by increasing the osmolarity of normal saline (NS), the most commonly-used irrigation solution. Here, we studied the effect of hyperosmolar saline (HS) on chondrocyte viability and cartilage repair in an in vivo injury model. Design Cartilage injury was induced by a single scalpel cut along the patellar groove of 8 week old rats in the absence of irrigation or with either NS (300 mOsm) or HS (600 mOsm). The percentage of cell death (PCD) within the injured area was assessed using confocal microscopy. Repair from injury was evaluated by histology/immunostaining, and inflammatory response by histology, cytokine array analysis and ELISA (enzyme-linked immunosorbent assay). Results The PCD in saline-irrigated joints was increased compared to non-irrigated (NI) joints [PCD = 20.8% (95%CI; 14.5, 27.1); PCD = 9.14% (95%CI; 6.3, 11.9); P = 0.0017]. However, hyperosmotic saline reduced chondrocyte death compared to NS (PCD = 10.4% (95%CI; 8.5, 12.3) P = 0.0024). Repair score, type II collagen and aggrecan levels, and injury width, were significantly improved with hyperosmotic compared to NS. Mild synovitis and similar changes in serum cytokine profile occurred in all operated joints irrespective of experimental group. Conclusions Hyperosmotic saline significantly reduced the chondrocyte death associated with scalpel-induced injury and enhanced cartilage repair. This irrigation solution might be useful as a simple chondroprotective strategy and may also reduce unintentional cartilage injury during articular reconstructive surgery and promote integrative cartilage repair, thereby reducing the risk of posttraumatic osteoarthritis.

Rationale The aim of the treatment of osteoarthritis (OA) is to improve symptoms and function; ideally, treatment would also prevent the structural progression of the disease. The targets of structure-modifying OA drugs (STMOADs) can be one of the various lesions of OA, including cartilage destruction, osteophytes, bone sclerosis, cysts, and bone attrition. However, prevention of cartilage destruction (chondroprotection) is currently accepted as the most promising target of STMOADs. The possibility of preventing progression of cartilage damage has been demonstrated in various animal models of OA. Chondroprotection is now being investigated in human OA. Several trials are in progress, and two positive studies have already been reported 1,2 . The goal of these initial trials was mainly to demonstrate that a reduction of the natural progression of OA joint space narrowing is possible in humans. Today, chondroprotection is no longer considered a myth, and the questions now are whether STMOADs provide a clinical benefit for the patient and how to evaluate that benefit. These new and relevant questions are difficult to answer. Can we expect a clinical benefit from STMOADs? Drugs evaluated for a structure-modifying effect may or may not also have symptomatic effects in the short term. STMOADs with a symptomatic effect can be expected to lead to clinical improvement in the OA patient in the long term. Evaluation of the clinical benefit of STMOADs could be possible with the tools presently used for the assessment of symptom-modifying drugs in OA. The clinical benefit that can be expected from a drug without known symptomatic effects but with a structuremodifying effect and how to measure that clinical benefit are as yet unknown. An effective STMOAD is not expected to induce a repair of the structural lesions of OA, but it is expected to stop or decrease their progression. Such a structural effect in the long term might result in symptom improvement or might only prevent a progression of symptoms. It also might have no clinical effect, since symptoms and lesions are poorly correlated in OA patients. The time required to demonstrate the clinical benefit of a STMOAD is unknown. It is possible that there may be a long delay (several years) between the beginning of STMOAD efficacy and the realization of any clinical benefit. The extent of clinical benefit could be related to the degree of effectiveness of the STMOAD. If this were true, the delay in clinical benefit could be very long for drugs able to only partially reduce the structural progression of the disease. Finally, the clinical benefit of a STMOAD might be missed by our usual tools and could require new methods of evaluation.

Summary Objective Cell-based tissue engineering strategies are currently in clinical use and continue to be developed at a rapid pace for the repair of cartilage defects. Regardless of the repair methodology, chondrocytes within newly regenerated cartilage remain susceptible to the abnormal inflammatory and mechanical environments that underlie osteoarthritic disease, likely compromising the implant's integration, function, and longevity. The present study investigates the use of parathyroid hormone-related peptide (PTHrP) overexpression for chondroprotection. Design Bovine articular chondrocytes were transfected with human PTHrP (hPTHrP) constructs (1-141 or 1-173) and subjected to injurious cyclic tensile strain (CTS; 0.5Hz and 16% elongation) for 48h. mRNA expression of matrix remodeling, inflammatory signaling, hypertrophic, and apoptotic genes were examined with real-time reverse transcription polymerase chain reaction. Nitric oxide (NO) and prostaglandin E 2 (PGE 2 ) production were measured using the Griess assay and enzyme immunoassay (EIA), respectively. Results CTS-induced an arthritic phenotype in articular chondrocytes as indicated by increased gene expression of collagenases and aggrecanases and increased production of NO and PGE 2 . Additionally, CTS increased collagen type X (Col10a1) mRNA expression, whereas overexpression of either hPTHrP isoform inhibited CTS-induced Col10a1 gene expression. However, hPTHrP 1-141 augmented CTS-induced NO and PGE 2 production, and neither hPTHrP isoform had any significant effect on apoptotic genes. Conclusions Our results suggest that chondrocytes overexpressing PTHrP resist mechanical strain-induced hypertrophic-like changes. Therapeutic PTHrP gene transfer may be considered for chondroprotection applications in newly regenerated cartilage.

SummaryObjectiveTo evaluate the impact of forced joint exercise following acute knee injury on lubricin metabolism and its relationship to cartilage degeneration and to assess chondroprotection of a single-dose purified human lubricin injection in exercised injured joints.MethodsAnterior cruciate ligament transection (ACLT) was performed in rats with six experimental groups; 3-week post-ACLT, 3-week post-ACLT + exercise, 5-week post-ACLT, 5-week post-ACLT + exercise, and 5-week post-ACLT + exercise treated with intra-articular phosphate buffered saline (PBS) or lubricin. Joint exercise was achieved using a rotating cylinder at a speed of 6 rpm for 30 min daily, 5 days a week starting 1 week following surgery. Cartilage lubricin expression in injured joints was determined. Histological analyses included Safranin O/Fast Green, activated caspase-3, and lubricin mRNA in-situ hybridization. Assessment of cartilage damage was performed by osteoarthritis research society international (OARSI) modified Mankin scoring and urinary CTXII (uCTXII) levels.ResultsAt 3 weeks, lubricin expression in exercised ACLT joints was significantly (P