Uncoupling of collagen II metabolism in newly diagnosed, untreated rheumatoid arthritis is linked to inflammation and antibodies against cyclic citrullinated peptides.

OBJECTIVE: To investigate the relationship between markers of collagen II synthesis and degradation with disease activity measures, autoantibodies, and radiographic outcomes in a 4-year protocol on patients with early rheumatoid arthritis (RA) who are naïve to disease-modifying antirheumatic drugs. METHODS: One hundred sixty patients with newly diagnosed, untreated RA entered the Cyclosporine, Methotrexate, Steroid in RA (CIMESTRA) trial. Disease activity and radiograph status were measured at baseline and 4 years. The N-terminal propeptide of collagen IIA (PIIANP) and the cross-linked C-telopeptide of collagen II (CTX-II) were quantified at baseline by ELISA. PIIANP was also assayed at 2 and 4 years. Anticyclic citrullinated peptide (anti-CCP) was recorded at baseline. An uncoupling index for cartilage collagen metabolism was calculated from PIIANP and CTX-II measurements. RESULTS: PIIANP was low at diagnosis and 4 years on (p < 0.001), irrespective of treatment and disease activity. PIIANP was lowest in anti-CCP positive patients (p = 0.006), and there was a negative correlation between PIIANP and anti-CCP titers (rho = -0.25, p 0.002). CTX-II was increased (p < 0.001) and correlated positively with disease activity and radiographic progression, but not with anti-CCP (p = 0.93). The uncoupling index was not superior to CTX-II in predicting radiographic changes. CONCLUSION: These results suggest that cartilage collagen formation and degradation are unbalanced when RA is diagnosed. The different associations of collagen II anabolism (PIIANP) and collagen II degradation (CTX-II) with anti-CCP, synovitis, and radiographic progression indicate that at this early stage of RA, cartilage collagen degradation is mainly driven by synovitis, while anti-CCP antibodies may interfere with cartilage regeneration by inhibiting collagen IIA formation. Trial registration j.nr NCT00209859.

Structures of strontium diformate and strontium fumarate. A synchrotron powder diffraction study.

The crystal structures of strontium diformate in space groups P2(1)2(1)2(1) (alpha form, 295 K), P4(1)2(1)2 (beta form, 334 and 540 K) and I4(1)/amd (delta form, 605 K), and strontium fumarate in space groups Fddd (beta form, 105 K) and I4(1)/amd (alpha form, 293 K) have been determined from synchrotron X-ray powder diffraction data. Except for the alpha-strontium diformate, all the structures are based on a diamond-like Sr-ion arrangement, as in strontium acetylene dicarboxylate. The formate ions are disordered in the delta phase owing to steric hindrance. The fumarate ions are disordered over four (alpha) or two (beta) symmetry-equivalent orientations. alpha-Strontium fumarate crystallizes with a unique 90 degree carboxylate dihedral angle, and is stable up to 773 K.

Application of inductively coupled plasma-mass spectrometry (ICP-MS) and quality assurance to study the incorporation of strontium into bone, bone marrow, and teeth of dogs after one month of treatment with strontium malonate.

The strontium content of serum, bone, marrow, and teeth was determined by inductively-coupled plasma mass spectrometry (ICP-MS). Significant correlations were obtained after the data were subjected to quality assurance (QA) performed according to validated procedures. After four weeks of treatment with strontium malonate, strontium levels increased from 76 +/- 9 microg g(-1) in placebo-treated dogs to levels of 7.2 +/- 1.7 mg g(-1), 9.5 +/- 2.7 mg g(-1), and 9.8 +/- 2.7 mg g(-1) in groups treated with 300, 1000, and 3000 mg kg(-1) day(-1), respectively. Strontium induced a highly significant increase in the bone formation marker, bone-specific alkaline phosphatase (BSAP), and an excellent correlation was found with the bone-strontium content. In females, the placebo-treated group showed a decrease in BSAP of 53%, whereas the three strontium malonate-treated groups showed an increase of 60, 276, and 278% for the groups treated with 300, 1000, and 3000 mg kg(-1) day(-1), respectively. For males the corresponding values were -44%, +142%, +194%, and +247% increases in BSAP in the placebo, 300, 1000, and 3000 mg kg(-1) day(-1) groups respectively.

Posttranslational Protein Modifications in Type 1 Diabetes - Genetic Studies with PCMT1, the Repair Enzyme Protein Isoaspartate Methyltransferase (PIMT) Encoding Gene.

BACKGROUND: Posttranslational protein modifications have been implicated in the development of autoimmunity. Protein L-isoaspartate (D-aspartate) O-methyltransferase (PIMT) repairs modified proteins and is encoded by PCMT1, located in a region linked to type 1 diabetes (T1D), namely IDDM5. AIM: To evaluate the association between genetic variations in the PCMT1 gene and T1D. METHODS: Firstly, PCMT1 was sequenced in 26 patients with T1D (linked to IDDM5) and 10 control subjects. The variations found in PCMT1 were then tested (alone and interacting with a functional polymorphism in SUMO4 and with HLA) for association with T1D in 253 families (using transmission disequilibrium test). In a third step, the association of the functional variation in PCMT1 (rs4816) with T1D was analyzed in 778 T1D patients and 749 controls (using chi-square test). In vitro promoter activity was assessed by transfecting INS-1E cells with PCMT1 promoter constructs and a reporter gene, with or without cytokine stimulation. RESULTS: Four polymorphisms in complete linkage disequilibrium were identified in PCMT1 (5' to the gene (rs11155676), exon 5 (rs4816) and exon 8 (rs7818 and rs4552)). In the whole cohort of 253 families, the allele associated with increased PIMT enzyme activity (rs4816, allele A) was less frequently transmitted to the affected than to the non-affected offspring (46% vs. 53%, p = 0.099). This finding was even more evident in the subset of families where the proband had high-risk SUMO4 (p = 0.069) or low-risk HLA (p = 0.086). Surprisingly, in the case-control study with 778 cases and 749 controls, an inverse trend was found (40.36% of patients and 36.98% of controls had the allele, p = 0.055). PCMT1 promoter activity increased with cytokine stimulation, but no differences were detected between the constructs adjacent to rs11155676. CONCLUSION: PCMT1 was virtually associated with T1D in groups defined by other risk genes (SUMO4 and HLA). A general association in a not further defined sample of T1D patients was not evident. Verification in a larger population is needed.

Biomarkers predict radiographic progression in early rheumatoid arthritis and perform well compared with traditional markers.

OBJECTIVE: To evaluate the performance of biochemical and traditional markers in predicting radiographic progression in rheumatoid arthritis (RA). METHODS: One hundred thirty-two patients with early RA were treated with nonbiologic therapies for 2 years and studied longitudinally. Genomic DNA was analyzed for presence of the shared epitope. Levels of matrix metalloproteinases (matrix metalloproteinase 1 [MMP-1], MMP-13, and MMP-3), tissue inhibitor of metalloproteinases 1 (TIMP-1), and cartilage oligomeric matrix protein (COMP) were assessed in serially obtained serum samples. The presence of pyridinoline (Pyr), deoxypyridinoline, glycosylated Pyr (Glc-Gal-Pyr), and C-telopeptide of type II collagen (CTX-II) was assessed in urine samples. Radiographs obtained at entry and at 2 years were evaluated using the modified Larsen score. RESULTS: Baseline and 2-year radiographs were available from 118 patients. Larsen scores worsened during the 2 years in 50 patients, while 68 patients had no radiographic progression. Levels of a variety of biochemical markers, i.e., MMP-3, CTX-II, COMP, TIMP-1, Pyr, and Glc-Gal-Pyr, correlated significantly with radiographic progression at entry and longitudinally as assessed by area under the curve (AUC). By multivariate analysis, a model including MMP-3 and CTX-II was identified as providing the best prediction of radiographic progression at entry (predictive accuracy by receiver operating characteristic [ROC] AUC = 0.76 [95% confidence interval 0.66-0.85]), while a combination of MMP-3, CTX-II, and swollen joint count formed the best longitudinal AUC model (predictive accuracy by ROC AUC = 0.81 [95% confidence interval 0.73-0.89]). Patient-reported measures (Health Assessment Questionnaire, pain scores) were of limited use. In a subset of 50 patients who were treated with methotrexate (MTX) during the followup period, median serum MMP-3 levels decreased after the initiation of MTX therapy (P = 0.0003). CONCLUSION: These results indicate that biochemical markers are useful predictors of radiographic progression in RA and that serum MMP-3 levels decrease significantly with MTX therapy. Multivariate models that include MMP-3 and CTX-II perform better than existing traditional markers in predicting radiographic outcome in RA.

Strontium D-glutamate hexahydrate and strontium di(hydrogen L-glutamate) pentahydrate.

[Sr(C5H7NO4)].6H2O, (I), and [Sr(C5H8NO4)2].5H2O, (II), both crystallize with similar strontium-glutamate-water layers. In (I), the neutral layers are connected through hydrogen bonds by water molecules, while in (II), the positively charged layers are connected through hydrogen bonds and electrostatic interactions by interleaving layers of hydrogen glutamate anions and water molecules.

New serum biochemical markers (Coll 2-1 and Coll 2-1 NO2) for studying oxidative-related type II collagen network degradation in patients with osteoarthritis and rheumatoid arthritis.

OBJECTIVE: Protein nitration is a prominent feature of inflammatory processes in the joint. We have developed immunoassays specific for a peptide of the alpha-helical region of type II collagen 108HRGYPGLDG116 (Coll 2-1) and its nitrated form 108HRGY(NO2)PGLDG116 (Coll 2-1 NO2) in biological fluids. DESIGN: Coll 2-1 and Coll 2-1 NO2 peptides were injected into rabbits. Two antisera (D3 and D37) were selected for their specificity and affinity and used to develop specific immunoassays. Coll 2-1 and Coll 2-1 NO2 were measured in sera of 242 healthy subjects (N), 67 patients with primary knee osteoarthritis (OA) and 19 patients with rheumatoid arthritis (RA). RESULTS: In healthy subjects, Coll 2-1 and Coll 2-1 NO2 concentrations were 125.13+/-3.71 nM and 0.16+/-0.08 nM, respectively. In OA and RA, Coll 2-1 and Coll 2-1 NO2 serum levels were found to be significantly increased compared to controls of the same range of age (Coll 2-1: OA: 200.80+/-8.98 nM, RA: 172.30+/-19.05 nM, normal: 126.60+/-6.70 nM and Coll 2-1 NO2: OA: 0.26+/-0.02, RA: 0.38+/-0.05, normal: 0.12+/-0.01 nM). Coll 2-1 NO2 levels were significantly more elevated in RA than in OA patients (P<0.05). As a consequence, the ratio Coll 2-1 NO2/Coll 2-1 was 1.6 times higher in RA than in OA subjects. No relationship was found between the radiological OA severity and the levels of Coll 2-1 and Coll 2-1 NO2 in serum. Coll 2-1 NO2, but not Coll 2-1, was correlated with C-reactive protein in the sera of OA and RA patients. CONCLUSIONS: 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 effect on cartilage of different forms of application of postmenopausal estrogen therapy: comparison of oral and transdermal therapy.

The effect on urine C-telopeptides of type II collagen (uCTX-II) of oral and transdermal estradiol treatment was compared using samples from two randomized, double-blind, placebo-controlled trials. A total of 171 healthy, Danish postmenopausal women, 45-65 years of age completed the 2-year study periods. The uCTX-II marker assessed cartilage degradation, and this response was compared with the effect on urine C-telopeptides of type I collagen (uCTX-I), considered a specific marker of bone resorption. Doses in the oral estradiol treatment groups (continuous combined therapy) were 1 mg 17-beta-estradiol+1 mg drosperinone or 1 mg 17-beta-estradiol+2 mg drosperinone or 1 mg 17-beta-estradiol+3 mg drosperinone or placebo. Doses in the transdermal estradiol treatment groups (continuous combined therapy) were 45 microg 17-beta-estradiol+30 levonorgestrel or 45 microg 17-beta-estradiol+40 microg levonorgestrel or placebo. The effect of oral and transdermal estradiol therapy on cartilage degradation was reflected as a decrease of 19-30% in uCTX-II (P=0.02 and P=0.003 vs. placebo) after 1 year of treatment. uCTX-I decreased 70% (P<0.0001 vs. placebo) reflecting a pronounced effect on bone resorption that was consistent with a 2-year increase in spine and hip BMD of 7-8% and 4-6%, respectively. The results indicate that different regimens of postmenopausal HRT both have an effect on cartilage and bone thus protecting against osteoporosis and osteoarthritis (OA). However, long-term clinical trials are needed to further investigate this issue.

Characterization of aged osteocalcin fragments derived from bone resorption.

INTRODUCTION: Osteocalcin (OC) is a small bone matrix protein exclusively found in mineralized tissue. OC measured in serum or plasma provides an index of bone formation. In the present study a sensitive inhibition ELISA was established that could quantify fragments derived from the OC Mid-region in human urine. METHODS: The ELISA was based on a monoclonal antibody directed against residues 21-29 of human OC (Mid-OC Urine ELISA). OC fragments were isolated from human urine by immunoaffinity chromatography. OC fragments were purified further by reversed phase high performance chromatography for characterization by N-terminal sequencing and mass-spectrometry. OC fragments were assayed in bone cell culture supernatants and in serum and urine from patients undergoing anti-resorptive bisphosphonate therapy using the Mid-OC urine ELISA. RESULTS AND CONCLUSION: It was demonstrated that the release of OC fragments was highly correlated with osteoclast-mediated pit formation (r2= 0.89) and with an established marker of bone resorption (CTX; r2=0.91). Mid-OC values were decreased markedly after 3 and 10 days of anti-resorptive bisphosphonate treatment further indicating that the marker reflects bone resorption. The molecular characterization revealed that most of these molecules were less than 15 amino acids in length and many contained modified aspartyl residues (D-aspartyl and isoaspartyl) characteristic of aged proteins. The presence of such modifications shows that these molecules have resided in the bone matrix for an extended period and thus they cannot be derived directly from bone formation. In conclusion, these findings demonstrate that OC-fragments are released during osteoclastic bone resorption and that the quantification of specific age-modified OC fragments can provide an index of bone resorption.

Suppression of elevated cartilage turnover in postmenopausal women and in ovariectomized rats by estrogen and a selective estrogen-receptor modulator (SERM).

OBJECTIVE: Several observational studies indicate that estrogen deficiency increases the incidence of osteoarthritis in postmenopausal women. To validate this observation, we investigated the effects of ovariectomy (OVX) on cartilage erosion in rats using histology and an established bio-assay of cartilage-specific collagen type II degradation products (CTX-II). Furthermore, we investigated whether estrogen and levormeloxifene, a selective estrogen-receptor modulator (SERM), can prevent the OVX-induced changes in cartilage degradation. The clinical relevance was assessed in postmenopausal women by measuring the changes in CTX-II during 12-month treatment with levormeloxifene versus placebo. DESIGN: Sixty 6-month-old rats were divided in five groups. One group was subjected to sham and the others to OVX, followed by treatment with vehicle alone, estradiol or 0.2 mg/kg/day or 5 mg/kg/day of levormeloxifene. The rats were treated for 9 weeks with biweekly blood and urine sampling for measurement of bone resorption and cartilage turnover. After study termination, hind knees were removed for histological analysis of erosions. The effect of levormeloxifene in post-menopausal women was assessed by measuring CTX-II in samples from 301 women who were participating in a phase II study of this SERM. RESULTS: OVX rats showed significant increases in the urinary excretion of CTX-II. After 9 weeks this was manifested as increased surface erosion of knee articular cartilage compared with sham-operated rats. Treatment with estrogen or levormeloxifene prevented the OVX-induced changes. There was a significant correlation between the 4-week changes in CTX-II and cartilage erosion at week 9 (r = 0.64, P < 0.001). In postmenopausal women treated with levormeloxifene, the urinary excretion of CTX-II was decreased by approximately 50% and restored CTX-II levels to the premenopausal range. CONCLUSIONS: This study is the first to demonstrate that a SERM suppresses cartilage degradation in both rodents and humans, suggesting potential therapeutical benefits in the prevention of destructive joint diseases such as osteoarthritis.

Hormone replacement therapy, calcium and vitamin D3 versus calcium and vitamin D3 alone decreases markers of cartilage and bone metabolism in rheumatoid arthritis: a randomized controlled trial [ISRCTN46523456].

This study aimed to evaluate the effects of hormone replacement therapy (HRT), known to prevent osteoporosis and fractures, on markers of bone and cartilage metabolism. Furthermore, we assessed whether changes in these markers corresponded to alterations in bone mineral density and radiographic joint destructions in postmenopausal women with rheumatoid arthritis. Eighty-eight women were randomized to receive HRT, calcium, and vitamin D3, or calcium and vitamin D3 alone, for 2 years. Bone turnover was studied by analyzing serum levels of C-terminal telopeptide fragments of type I collagen (CTX-I), C-terminal telopeptide of type I collagen (ICTP), bone sialoprotein, and C-terminal propeptide of type I procollagen (PICP) and cartilage turnover by urinary levels of collagen type II C-telopeptide degradation fragments (CTX-II) and cartilage oligomeric matrix protein (COMP) in serum. Treatment with HRT resulted in decrease in CTX-I (P < 0.001), ICTP (P < 0.001), PICP (P < 0.05), COMP (P < 0.01), and CTX-II (P < 0.05) at 2 years. Reductions in CTX-I, ICTP, and PICP were associated with improved bone mineral density. Of the markers tested, CTX-I reflected bone turnover most sensitively; it was reduced by 53 +/- 6% in the patients receiving HRT. Baseline ICTP (P < 0.001), CTX-II (P < 0.01), and COMP (P < 0.05) correlated with the Larsen score. We suggest that biochemical markers of bone and cartilage turnover may provide a useful tool for assessing novel treatment modalities in arthritis, concerning both joint protection and prevention of osteoporosis.

Post-translational modifications of proteins: implications for aging, antigen recognition, and autoimmunity.

Proteins are complex organic molecules susceptible to numerous post-translational modifications occurring spontaneously during aging or as a consequence of physiologic or pathologic processes. Antigenicity and interactions of proteins with components of the immune system may be profoundly affected by post-translational modifications. Thus, modified self-antigens may be absent (not-tolerated) during early T-cell selection and trigger reactions by the immune system as they arise later in life. In turn, this may play a role in the initiation and pathogenesis of autoimmune diseases. This Review article presents an overview of protein modifications that have been shown to affect antigenicity and presentation of protein antigens in autoimmune diseases. The relevance of these observations is discussed, and the implications for future prophylactic and therapeutic interventions are outlined.

Cartilage destruction in collagen induced arthritis assessed with a new biochemical marker for collagen type II C-telopeptide fragments.

OBJECTIVE: To assess the ability of a marker of collagen type II degradation (CTX-II) to quantify cartilage turnover in vitro in cartilage explants and in vivo in rats with collagen induced arthritis (CIA). METHODS: Bovine articular cartilage explants were cultured in the presence of interleukin 1a, oncostatin M, and plasminogen to induce cartilage degradation. CTX-II, CTX-I (C-telopeptide fragment of collagen type I), glycosaminoglycan, and hydroxyproline contents in culture supernatants were measured. CIA was induced in 12-week-old female Lewis rats by immunization with bovine type II collagen. The incidence and severity of arthritis were monitored by measuring paw swelling, and urinary levels of CTX-II and CTX-I were determined. The knee joints of rats were histopathologically examined after sacrifice. Results. CTX-II but not CTX-I levels correlated well with collagen degradation in bovine articular cartilage in vitro quantified by hydroxyproline release. Urinary CTX-II levels as well as paw volume of CIA rats were significantly higher than normal rats on Days 21, 28, and 42 and were apparently correlated with cartilage destruction, assessed histopathologically. Urinary CTX-I level began to increase on Day 21, but only on Day 42 was it significantly different between CIA and normal rats. The elevation in CTX-I level appeared to occur later than that of CTX-II, in accord with the more delayed onset of bone erosion in the CIA model of rheumatoid arthritis. CONCLUSION: Urinary CTX-II may be a useful marker for evaluation of dynamics of cartilage destruction in CIA rats.

An immunoassay for measuring fragments of newly synthesized collagen type I produced during metastatic invasion of bone.

Degradation products of collagen type I can be measured by CrossLaps (CTX) immunoassays, providing an index of bone resorption. The CTX epitope EKAHDGGR comprises a DG-motif susceptible to post-translational modifications. In newly synthesized collagen this motif is in the native form denoted alphaCTX, but converts to an isomerized form (betaCTX) during aging of bone. Furthermore, the lysine residue (K) within the CTX epitope participates in inter-molecular cross-links in mature bone. The present paper describes an assay, ALPHA CTX ELISA for measurement of cross-linked alphaCTX molecules (alpha-alphaCTX) in urine. The ALPHA CTX ELISA demonstrated a high specificity and technical precision for measuring such fragments. The assay was evaluated in a cross-sectional study, comparing the urinary excretion of the marker in 100 breast cancer patients with bone metastases (BC+) and 15 breast cancer patients without metastases to bone (BC-) as well as 31 age-matched healthy post-menopausal women (PM). For comparison alpha, beta, and beta-betaCTX was also measured using commercially available immunoassays. In BC+ urinary alpha-alphaCTX increased significantly compared to BC- and PM with p-values of 0.005 and <0.0001, respectively. In contrast, the age-modified form beta-betaCTX, representing the degradation of old bone, was less increased. Z-score analysis was used to compare the ability of the CTX markers to discriminate between BC+ and PM. The alpha-alphaCTX marker was found to provide a far better discrimination (Z=7.5) than beta-betaCTX (Z=3.6). In conclusion, measurement of alpha-alphaCTX fragments may provide a clinically relevant assessment of bone resorption related to bone metastases.

Ovariectomized rats as a model of postmenopausal osteoarthritis: validation and application.

We aimed to assess the effect of ovariectomy on cartilage turnover and degradation, to evaluate whether ovariectomized (OVX) rats could form an experimental model of postmenopausal osteoarthritis. The effect of ovariectomy on cartilage was studied using two cohorts of female Sprague-Dawley rats, aged 5 and 7 months. In a third cohort, the effect of exogenous estrogen and a selective estrogen receptor modulator was analyzed. Knee joints were assessed by histological analysis of the articular cartilage after 9 weeks. Cartilage turnover was measured in urine by an immunoassay specific for collagen type II degradation products (CTX-II), and bone resorption was quantified in serum using an assay for bone collagen type I fragments (CTX-I). Surface erosion in the cartilage of the knee was more severe in OVX rats than in sham-operated animals, particularly in the 7-month-old cohort (P = 0.008). Ovariectomy also significant increased CTX-I and CTX-II. Both the absolute levels of CTX-II and the relative changes from baseline seen at week 4 correlated strongly with the severity of cartilage surface erosion at termination (r = 0.74, P < 0.01). Both estrogen and the selective estrogen receptor modulator inhibited the ovariectomy-induced acceleration of cartilage and bone turnover and significantly suppressed cartilage degradation and erosion seen in vehicle-treated OVX rats. The study indicates that estrogen deficiency accelerates cartilage turnover and increases cartilage surface erosion. OVX rats provide a useful experimental model for the evaluation of the chondroprotective effects of estrogens and estrogen-like substances and the model may be an in vivo representation of osteoarthritis in postmenopausal women.

Endothelin 1 promotes osteoarthritic cartilage degradation via matrix metalloprotease 1 and matrix metalloprotease 13 induction.

OBJECTIVE: Degradation of the collagenous extracellular matrix by metalloproteases (MMPs) plays an important role in the pathogenesis of osteoarthritis (OA). Recently, it was suggested that endothelin 1 (ET-1), a potent vasoconstrictor, may be involved in MMP regulation. This study investigated the role of ET-1 in OA cartilage degradation. METHODS: We explored ET-1 expression and synthesis in normal and OA cartilage and synovial membrane by reverse transcription-polymerase chain reaction and immunohistochemistry. MMP-1 and MMP-13 gene expression and protein synthesis were investigated using Northern blotting and enzyme-linked immunosorbent assays. Additionally, ET-1-induced collagenase activity, type II collagen metabolites, and tissue inhibitor of metalloproteases 1 (TIMP-1) protein were evaluated. RESULTS: We found expression and synthesis of ET-1, in situ, in both normal and OA cartilage and synovial membrane. We demonstrated that ET-1 induced gene expression and protein synthesis of both MMP-1 and MMP-13. These enzymes were produced in OA chondrocyte cultures, and the production increased in a dose-dependent manner in the presence of ET-1. In OA cartilage, ET-1 also induced type II collagen-derived neoepitopes concomitantly with an increase in collagenase activity and a decrease in TIMP-1 protein. CONCLUSION: Our results provide strong evidence of the catabolic role of ET-1 in OA cartilage via MMP-1 and MMP-13 up-regulation. As well, ET-1 increased the net MMP/TIMP balance and secondarily increased collagen degradation. Hence, ET-1 becomes an attractive factor to target in the conception of new therapeutic approaches for OA and other diseases in which MMP-13 and MMP-1 actions are crucial in tissue alteration.

Breast cancer patients with bone metastases are characterised by increased levels of nonisomerised type I collagen fragments.

BACKGROUND: Fragments of collagen type I containing the epitope AHDGGR (CTX) are generated during bone resorption. The aspartyl-glycine (DG) site within CTX is synthesised in the L-aspartyl peptide (alphaL) form, but converts to the age-modified forms L-isoaspartyl peptide (betaL) and D-aspartyl peptide (alphaD) over time. The purpose of the present study was to test the ability of the various CTX forms to identify breast cancer patients with bone metastases and to investigate whether such patients had an altered CTX excretion pattern. METHODS: In this cross-sectional study we compared CTX excretion in healthy premenopausal and postmenopausal women with CTX levels in patients with breast cancer. The breast cancer cohort comprised eight hypercalcemic patients with bone metastases (HC+), 100 normocalcemic patients with bone metastases (NC+) and 15 normocalcemic patients without bone metastases (NC-). RESULTS: In HC+ patients and NC+ patients, the excretion of alphaL CTX was highly increased compared with NC- patients (P < 0.01), with Z scores of 3.4 and 2.0, respectively. The excretion of the age-modified forms (betaL and alphaD CTX) was less increased in HC+ patients and in NC+ patients as compared with NC- patients, with Z scores of 2.2 and 1.0, respectively, for betaL CTX, and of 1.6 and 0.8, respectively, for alphaD CTX. CONCLUSION: Assays for the various isoforms of CTX have different sensitivities to identify patients affected by bone metastases. The alphaL CTX isoform reflecting resorption of young bone appeared to provide the best differentiation of patients affected by breast cancer-induced bone metastases. In conclusion, patients affected by metastatic bone disease present an altered excretion pattern of CTX isoforms.

Association of baseline levels of markers of bone and cartilage degradation with long-term progression of joint damage in patients with early rheumatoid arthritis: the COBRA study.

OBJECTIVE: The known risk factors for radiologic progression in rheumatoid arthritis (RA) are not optimally discriminative in patients with early disease who do not have evidence of radiologic damage. We sought to determine whether urinary C-terminal crosslinking telopeptide of type I (CTX-I) and type II (CTX-II) collagen (markers of bone and cartilage destruction, respectively) are associated with long-term radiologic progression in patients with early RA. METHODS: This was a prospective study of 110 patients with early RA who were participating in the COBRA (Combinatietherapie Bij Reumatoïde Artritis) clinical trial and followup study, a randomized controlled trial comparing the efficacy of oral pulse prednisolone, methotrexate, plus sulfasalazine with sulfasalazine alone. We investigated the relationship between baseline levels of urinary CTX-I and CTX-II and the mean annual progression of joint destruction over a median of 4 years, as measured by changes in the modified Sharp score (average of 2 independent readers). RESULTS: In multivariate logistic regression analysis, baseline urinary CTX-I and CTX-II levels in the highest tertile were the strongest predictors of radiologic progression (Sharp score increase >2 units/year; odds ratio 7.9 and 11.2, respectively), independently of treatment group, erythrocyte sedimentation rate (ESR), Disease Activity Score in 28 joints, rheumatoid factor (RF), and baseline joint damage (Sharp score). The likelihood ratios for a positive test were 3.8 and 8.0 for CTX-I and CTX-II, respectively, which compared favorably with the likelihood ratios for the ESR (3.0), baseline joint damage (1.6), and RF (1.8). When patients were grouped according to the presence (Sharp score >/=4, n = 49) and absence (Sharp score <4, n = 61) of joint damage at baseline, CTX-I and CTX-II levels were predictive only in those without baseline joint damage (odds ratio 14.9 and 25.7, respectively). CONCLUSION: High baseline levels of urinary CTX-I and CTX-II independently predict an increased risk of radiologic progression over 4 years in patients with early RA, especially those without radiologic joint damage. Urinary CTX-I and CTX-II may be useful for identifying individual RA patients at high risk of progression very early in the disease, before erosions can be detected radiographically. Such patients may be in special need of treatments that inhibit bone and cartilage degradation.