Interleukin-10 protects against blood-induced joint damage.

Despite prophylactic treatment, haemophilia patients suffer from spontaneous joint bleeds, which lead to severe joint damage. Also after joint trauma, an intra-articular haemorrhage can add to joint damage over time. This study evaluated interleukin 10 (IL-10) in the search for possible interventions to prevent or limit the damaging effects of joint bleeds. Human articular cartilage tissue explants were cultured in the presence or absence of 50% v/v blood (or its cellular components) for 4 d (the expected blood load in vivo after a joint haemorrhage), followed by a recovery period of 12 d. Pharmacological dosages of IL-10 reached during treatment (1 or 10 ng/ml) were added. Additionally, cartilage and synovial tissue obtained from joints with end-stage haemophilic arthropathy (HA) were cultured in the presence of IL-10 (10 ng/ml). IL-10 protected cartilage from the damaging effects of blood exposure, measured by its effects on proteoglycan turnover. In addition, IL-10 beneficially influenced cartilage from patients with HA and reduced the production of the inflammatory cytokines IL-1beta and tumour necrosis factor-alpha by haemophilic synovial tissue. Taken together, although effects were obtained in vitro, IL-10 protects against blood-induced joint damage and might be further evaluated as candidate in treatment of tissue damaging effects of joint haemorrhages.

Initiation of degenerative joint damage by experimental bleeding combined with loading of the joint: a possible mechanism of hemophilic arthropathy.

OBJECTIVE: To investigate the effect of a limited number of experimental joint bleedings, combined with loading of the affected joint, on the development of progressive degenerative joint damage. METHODS: The right knee of 8 mature beagle dogs was injected with freshly collected autologous blood 3 times per week for 4 weeks, to mimic a limited number of joint hemorrhages occurring over a short period. To ensure loading of the experimental joint, the contralateral control knee of the animals was fixed to the trunk 4 hours per day, 3 days per week. Ten weeks after the last injection, cartilage tissue and synovium were collected from both knees to analyze features of joint degeneration. Cartilage was prepared for analysis of proteoglycan turnover (synthesis, retention, release, and content) and histologic features. Synovium was prepared for histologic analysis. RESULTS: The rate of proteoglycan synthesis was significantly increased, characteristic of degenerative cartilage damage as seen in osteoarthritis. Release of newly formed proteoglycans (as a measure of retention) and total loss of proteoglycans from the cartilage matrix were increased. Cartilage matrix integrity was adversely altered, as shown by histologic damage. Histologic analysis also revealed signs of synovial inflammation. These effects were not observed 10 weeks after the experimental bleedings in joints that did not undergo forced loading. CONCLUSION: Experimental joint bleedings when combined with loading of the affected joint resulted in features of progressive degenerative joint damage, whereas similar joint hemorrhages without joint loading did not. This might reflect a possible mechanism of joint damage in hemophilia.

Blood-induced joint damage: longterm effects in vitro and in vivo.

OBJECTIVE: We previously showed that 4-day in vitro exposure of human cartilage to blood, as well as a single experimental joint bleeding in dogs, resulted in a disturbed cartilage matrix turnover lasting at least 2 weeks. We now evaluate the longterm outcome of the adverse in vitro and in vivo effects of blood on cartilage matrix turnover. METHODS: Human and canine articular cartilage tissue was cultured in the presence of homologous whole blood during 4 days. The in vitro cartilage matrix turnover was analyzed directly after blood exposure or following culture for additional periods of 2, 5, and 10 weeks in the absence of blood. The in vivo longterm effects were determined by injecting autologous blood into the right knee of 12 Beagle dogs. Six dogs were killed shortly after blood injections; the 6 remaining dogs were killed 10 weeks later. Cartilage matrix turnover and the cartilage destructive properties of the synovial tissue were analyzed. RESULTS: Short term (4 days) in vitro exposure of human or canine cartilage to whole blood inhibited proteoglycan synthesis by more than 98% (day 4), an inhibition which lasted until week 10 (70 and 75% inhibition, respectively). Also the in vivo short term exposure of cartilage to blood induced the adverse changes in cartilage proteoglycan turnover seen shortly after exposure. However, in vivo 10 weeks after the last injection, normalization of cartilage matrix turnover was observed. Synovial inflammation was absent and no destructive activity was found. CONCLUSION: These data show a discrepancy between the in vitro and in vivo longterm effects of blood on cartilage. A possible explanation for the in vivo recovery after experimental joint bleeding in dogs could be that the observed changes in cartilage only predispose to acute damage but that additional (e.g., mechanical) factors are needed to induce permanent joint damage.

Immature articular cartilage is more susceptible to blood-induced damage than mature articular cartilage: an in vivo animal study.

OBJECTIVE: Cartilage of young but skeletally mature dogs is more susceptible to blood-induced damage than that of old dogs. The aim of the present study was to investigate whether cartilage of skeletally immature individuals is even more adversely affected by exposure to blood than that of mature individuals, as suggested by clinical practice experience with humans. METHODS: Right knees of 3 groups of 6 beagle dogs (skeletally immature, young mature, and old animals) were injected with autologous blood on days 0 and 2. On day 4, cartilage matrix proteoglycan turnover (content, synthesis, and release), synovial inflammation, and cartilage-destructive properties of the synovial tissue were determined and compared with those of the left uninjected control knees. RESULTS: Subsequent to intraarticular bleeding, cartilage proteoglycan content decreased in an age-dependent manner, with the largest decrease occurring in cartilage of immature animals. Proteoglycan synthesis per cell also decreased in an age-dependent manner, with the largest decrease occurring in the immature animals. Cartilage proteoglycan release increased in all 3 groups, but the decrease was not age dependent. Interestingly, immature animals showed a large increase in cartilage DNA content upon exposure to blood, whereas mature animals did not. Histologic analysis showed a mild synovitis in animals of all ages, but catabolic inflammatory activity was found only in immature animals. CONCLUSION: Joints of skeletally immature dogs appeared to be more susceptible than joints of mature dogs to the adverse effects of a joint hemorrhage. These data suggest that for humans, specifically young children are at risk for joint damage after a joint hemorrhage.

Short-term exposure of cartilage to blood results in chondrocyte apoptosis.

Studies have shown that joint bleeding leads to cartilage degradation independent of concurrent synovitis. We hypothesized that the blood-induced cartilage damage is because of increased chondrocyte apoptosis after short-term exposure of whole blood or isolated mononuclear cells plus red blood cells to cartilage. Human cartilage tissue samples were co-cultured for 4 days with whole blood (50% v/v) or with mononuclear cells plus red blood cells (50% v/v equivalents). Cartilage matrix proteoglycan synthesis ((35)SO(4)(2-) incorporation) was determined after 4 days as well as at day 16 (after a 12-day recovery period in the absence of any additions). To test the involvement of apoptosis a specific caspase-3 inhibitor (acDEVDcho, 0 to 500 micro mol/L) as well as a pan-caspase inhibitor (zVADfmk, 0 to 500 micro mol/L) were added. Chondrocyte apoptosis was evaluated by immunohistochemical staining of single-strand DNA and by terminal dUTP nick-end labeling. Cartilage co-cultured with whole blood as well as mononuclear cells plus red blood cells induced a long-term inhibition of proteoglycan synthesis (74% and 78% inhibition on day 16, respectively). Immunohistochemistry showed a threefold increase in apoptotic chondrocytes in cultures with 50% whole blood as well as with mononuclear cells plus red blood cells. Both the specific caspase-3 inhibitor and the pan-caspase inhibitor partially restored proteoglycan synthesis in the cartilage after blood exposure. This effect was accompanied by a decrease in the number of apoptotic chondrocytes. These data suggest that a single joint hemorrhage (a 4-day exposure of cartilage to 50% v/v blood) results in induction of chondrocyte apoptosis, responsible for the observed inability of the chondrocytes to restore the proteoglycan synthesis during recovery from a short-term exposure to blood. This reduced restoration could eventually lead to cartilage degeneration and ultimately joint destruction.