The determination of both Coll 2-1 and Coll 2-1 NO2 in serum of arthritic patients seems to be a promising useful tool for the detection of oxidative-related cartilage degradation episode. Further, these markers could be helpful for monitoring the effects of anti-inflammatory or antioxidant drugs on cartilage degradation.

The in-vitro effects of avocado and soybean unsaponifiable residues on neutral metalloproteinase activity, cytokines and prostaglandin E2 (PGE2) production by human articular chondrocytes were investigated. Avocado and soybean unsaponifiable residues were mixed in three ratios: 1:2 (A1S2), 2:1 (A2S1) or 1:1 (A2S2). Freshly isolated human chondrocytes were cultured for 72 h in the absence or presence of interleukin-1beta, (IL-1beta) (17 ng/ml), with or without unsaponifiable residue mixtures at a concentration of 10 microg/ml. A/S unsaponifiable residues were also tested separately at concentrations of 3.3, 6.6 and 10 microg/ml. All A/S unsaponifiable mixtures reduced the spontaneous production of stromelysin, interleukin-6 (IL-6), interleukin-8 (IL-8) and prostaglandin E2 (PGE2) by chrondrocytes. At concentrations of 3.3 and 6.6 microg/ml, A/S residues, tested separately, were potent inhibitors of the production of IL-8 and PGE2. Nevertheless, only avocado residue inhibited IL-6 production at these concentrations. A/S unsaponifiable mixtures had a more pronounced inhibitory effect on cytokine production than avocado or soybean residues added alone. As anticipated, IL-1beta induced a marked release of collagenase, stromelysin, IL-6, IL-8 and PGE2. A/S unsaponifiable mixtures partially reversed the IL-1 effects on chrondrocytes. These findings suggest a potential role for A/S unsaponifiable extracts in mitigating the deleterious effects of IL-1beta: on cartilage.

Based on previous studies showing that strontium ranelate (S12911) modulates bone loss in osteoporosis, it could be hypothesized that this drug also is effective on cartilage degradation in osteoarthritis (OA). This was investigated in vitro on normal and OA human chondrocytes treated or not treated with interleukin-1␤ (IL-1␤). This model mimics, in vitro, the imbalance between chondroformation and chondroresorption processes observed in vivo in OA cartilage. Chondrocytes were isolated from cartilage by enzymatic digestion and cultured for 24 -72 h with 10 ؊4 -10 ؊3 M strontium ranelate, 10 ؊3 M calcium ranelate, or 2 ⅐ 10 ؊3 M SrCl 2 with or without IL-1␤ or insulin-like growth factor I (IGF-I). Stromelysin activity and stromelysin quantitation were assayed by spectrofluorometry and enzyme amplified sensitivity immunoassay (EASIA), respectively. Proteoglycans (PG) were quantified using a radioimmunoassay. Newly synthesized glycosaminoglycans (GAGs) were quantified by labeled sulfate (Na 2 35 SO 4 ) incorporation. This method allowed the PG size after exclusion chromatography to be determined. Strontium ranelate, calcium ranelate, and SrCl 2 did not modify stromelysin synthesis even in the presence of IL-1␤. Calcium ranelate induced stromelysin activation whereas strontium compounds were ineffective. Strontium ranelate and SrCl 2 both strongly stimulated PG production suggesting an ionic effect of strontium independent of the organic moiety.

The normalisation of Coll2-1 levels 3 months after surgery indicates that Coll2-1 is a disease-specific marker that is sensitive to the structural changes occurring in a single joint. Furthermore, the immunohistochemical findings are consistent with the concept that the major source of serum Coll2-1 is the damaged articular cartilage. Finally, serum MPO levels decreased after joint replacement indicating that neutrophil activation occurs in OA joints, even in the late stage of the disease.

Cytokines are potent regulators of the chondrocyte functions. Some of them are produced by chondrocytes and interact to regulate cartilage metabolism. In this study, we investigated the production of interleukin-1 beta (IL-1 beta), IL-6, IL-8 and leukemia inhibitory factor (LIF) by human chondrocytes and examined the modulation of their secretion by exogenous cytokines. Human articular chondrocytes were isolated from their extracellular matrix by a triple successive enzymatic digestion of the cartilage. Subsequently, chondrocytes were stimulated by increased amounts of human recombinant cytokines [IL-1 beta, tumour necrosis factor alpha (TNF alpha), IL-8, LIF, IL-6]. IL-1 beta, IL-6, IL-8 and LIF were assayed into culture media and inside cell extracts by specific enzyme amplified sensitivity immunoassays (EASIAs). Under these experimental conditions, we have identified various interactions between cytokines. IL-beta and TNF alpha highly stimulated IL-6, LIF and IL-8 productions. IL-6 decreased IL-8 synthesis and increased LIF production. IL-8 slightly enhanced IL-6 production. Finally, LIF stimulated IL-1 beta, IL-6 and IL-8 productions. Using neutralizing antibodies against IL-1, we demonstrated that the effects of LIF were secondary to the stimulation by LIF of IL-1 beta production by the chondrocytes. In conclusion, chondrocytes secrete a variety of immunocompetent cytokines including IL-1 beta, IL-6, IL-8 and LIF that can interact to regulate chondrocytes metabolism. These results also define new biological activities of LIF and IL-6, and raise questions concerning their role in the pathogenesis of joint diseases.

At baseline, Coll 2-1 and Coll 2-1 NO(2) urinary levels were indicative of the clinical activity of knee OA and the increase of these peptides over 1 year was predictive of the radiological progression of knee OA.

Objective. Compared with wild-type (WT) mice, biglycan/fibromodulin double-deficient mice develop severe knee osteoarthritis. We undertook this study to compare type II collagen catabolism in the 2 genotypes and to compare the usefulness of 3 biomarkers of collagen degradation (C2C [also known as Col2-3/4C long mono ] as well as the peptide Coll2-1 and its nitrated form, Coll2-1NO 2 ) for evaluating collagen catabolism in vivo.Methods. In 15 WT mice and 15 biglycan/ fibromodulin double-deficient mice, we determined serum levels of C2C at ages 66 and 141 days, and we determined serum levels of Coll2-1 and Coll2-1NO 2 at ages 49, 81, 95, and 141 days. Expression of the biomarkers in knee sections was examined using immunohistochemistry.Results. The mean concentrations of C2C and Coll2-1 were higher in biglycan/fibromodulin doubledeficient mice at all time points. For C2C and Coll2-1, the ratio of the serum concentration in biglycan/ fibromodulin double-deficient mice to that in WT mice (the double-deficient:WT ratio) was constant over time and was ϳ1.63 and ϳ1.15, respectively. In contrast, the double-deficient:WT ratio for Coll2-1NO 2 varied and, depending on age, was >1 or <1. No significant correlation was found between the expression of the different biomarkers, except for a weak, negative correlation between Coll2-1NO 2 and C2C. In both genotypes, antibodies to each biomarker labeled some fibroblasts in the tendons and menisci as well as chondrocytes above the tidemark in articular cartilage. Growth plates were unstained. For each biomarker, extracellular staining was limited to fibrocartilage areas in the tendons and menisci in all mice and was limited to some focal lesions of the cartilage in biglycan/fibromodulin doubledeficient mice.Conclusion. The different double-deficient:WT ratios observed with C2C, Coll2-1, and Coll2-1NO 2 in the absence of any correlation between the expression of the 3 biomarkers indicate that these biomarkers give complementary, rather than redundant, information about in vivo type II collagen catabolism.Osteoarthritis (OA) is one of the leading causes of pain and disability in the elderly. Its high prevalence and its moderate-to-severe impact on daily life pose a significant public health problem (1). Despite intensive research, a cure remains elusive for this disease with important socioeconomic consequences. The intrinsic difficulty in finding a cure is compounded by the low sensitivity of diagnostic and monitoring tools. For example, the destruction of articular cartilage is an important feature of OA, and radiographic change in joint space width (JSW) is widely recognized as the gold standard end point for measuring destruction of articular cartilage and OA progression. However, this end point is an indirect measure of cartilage integrity, since cartilage is invisible on radiographs and its integrity must be inferred from the spacing between bones. Furthermore, change in JSW has a low signal-to-noise ratio; annual changes are only 0.1-0.2 mm in OA patients (2), close to the p...

These findings suggest that IFN gamma and IL-1 synergistically stimulate the production of IL-6, IL-1ra, NO and PGE(2)and inhibit PG synthesis. By contrast, IL-1 beta and IFN gamma have opposite effects on IL-8, IL-10 and stromelysin productions. These effects are not reversed by L-NMMA, suggesting that NO is not the principal mediator involved in responses of chondrocytes to IFN gamma.