Investigation of Glycosaminoglycans in Urine and Their Alteration in Patients with Juvenile Idiopathic Arthritis.

(1) Background: In this study, we evaluated the modulation of urine glycosaminoglycans (GAGs), which resulted from etanercept (ETA) therapy in patients with juvenile idiopathic arthritis (JIA) in whom methotrexate therapy failed to improve their clinical condition. (2) Methods: The sulfated GAGs (sGAGs, by complexation with blue 1,9-dimethylmethylene), including chondroitin-dermatan sulfate (CS/DS) and heparan sulfate (HS), as well as non-sulfated hyaluronic acid (HA, using the immunoenzymatic method), were determined in the blood of 89 children, i.e., 30 healthy children and 59 patients with JIA both before and during two years of ETA treatment. (3) Results: We confirmed the remodeling of the urinary glycan profile of JIA patients. The decrease in the excretion of sGAGs (p < 0.05), resulting from a decrease in the concentration of the dominant fraction in the urine, i.e., CS/DS (p < 0.05), not compensated by an increase in the concentration of HS (p < 0.000005) and HA (p < 0.0005) in the urine of patients with the active disease, was found. The applied biological therapy, leading to clinical improvement in patients, at the same time, did not contribute to normalization of the concentration of sGAGs (p < 0.01) in the urine of patients, as well as CS/DS (p < 0.05) in the urine of sick girls, while it promoted equalization of HS and HA concentrations. These results indicate an inhibition of the destruction of connective tissue structures but do not indicate their complete regeneration. (4) Conclusions: The metabolisms of glycans during JIA, reflected in their urine profile, depend on the patient's sex and the severity of the inflammatory process. The remodeling pattern of urinary glycans observed in patients with JIA indicates the different roles of individual types of GAGs in the pathogenesis of osteoarticular disorders in sick children. Furthermore, the lack of normalization of urinary GAG levels in treated patients suggests the need for continued therapy and continuous monitoring of its effectiveness, which will contribute to the complete regeneration of the ECM components of the connective tissue and thus protect the patient against possible disability.

Plasma and Urine Levels of Glycosaminoglycans in Patients with Systemic Sclerosis and Their Relationship to Selected Interleukins and Marker of Early Kidney Injury.

Systemic sclerosis (SSc) is a chronic connective tissue disease characterized by immune system dysfunction, vasculopathy, and progressive fibrosis of the skin and internal organs, resulting from excessive accumulation of extracellular matrix (ECM) elements, including collagen and proteoglycans (PGs). An uncontrolled PG proliferation, caused by disturbances in their metabolism in tissues, is most likely reflected in the quantitative changes of their components, i.e., glycosaminoglycans (GAGs), in body fluids. Therefore, the aim of this study was to quantify the different types of GAGs in the blood and urine of systemic sclerosis patients. Chondroitin/dermatan sulfates (CS/DS) and heparan sulfates/heparin (HS/H) were quantified by hexuronic acid assay and electrophoretic fractionation, while hyaluronic acid (HA) and keratan sulfates were evaluated using ELISA tests. In turn, individual urinary GAGs were determined using the Blyscan™ Sulfated Glycosaminoglycan Assay Kit. Our results showed that the plasma concentrations of CS/DS, HS/H, HA, and KS in systemic sclerosis patients were significantly higher compared with those in healthy subjects. In the case of urine measurements, we have found that in SSc patients, CS/DC concentrations were significantly higher, while HA concentrations were significantly lower compared with the values observed in the urine of healthy subjects. Importantly, the found by us correlations between plasma keratan sulfate levels and both the duration of the disease and the severity of skin lesions, as expressed by the Rodnan scale, seems to suggest this GAG as a potential marker in assessing disease progression and activity. In addition, a level of urinary excretion of all types of GAGs due to their high positive correlation with uACR, may be a valuable complementary test in the diagnosis of early renal dysfunction in the course of SSc.

GAAGs, COMP, and YKL-40 as Potential Markers of Cartilage Turnover in Blood of Children with Juvenile Idiopathic Arthritis Treated with Etanercept-Relationship with ADAMTS4, ADAMTS5, and PDGF-BB.

We quantified galactosaminoglycans (GAAGs), oligomeric cartilage matrix protein (COMP), and human cartilage glycoprotein 39 (YKL-40) in blood obtained from juvenile idiopathic arthritis (JIA) before and during 2-year treatment with etanercept (ETA), as potential biomarkers of cartilage extracellular matrix (ECM) dysfunction and indicators of efficacy of biologic therapy. We also evaluated the relationship of the mentioned markers with the factors that regulate their metabolism, disintegrin and thrombospondin motif metalloproteinases 4 (ADAMTS4), ADAMTS5, and platelet-derived growth factor BB (PDGF-BB). METHODS: We studied 38 children diagnosed with JIA and 45 healthy children. We quantified GAAGs by assessing the concentration of unsaturated disaccharide units formed by digestion of isolated glycosaminoglycans with chondroitinase ABC, while COMP, YKL-40, and PDGF-BB were quantified using immunoenzymatic methods. RESULTS: Compared to the control group, GAAGs and COMP levels were significantly lower, while YKL-40 levels were higher in the blood of patients with aggressive JIA, qualified for ETA treatment. ETA therapy leading to clinical improvement simultaneously promoted normalization of COMP and YKL-40 levels, but not GAAGs. After 24 months of taking ETA, glycan levels were still significantly lower, relative to controls. GAAGs, COMP, and YKL-40 levels were significantly influenced by ADAMTS4, ADAMTS5, and PDGF-BB levels both before and during ETA treatment. CONCLUSIONS: The dynamics of changes in marker concentrations during treatment seem to indicate that measurement of COMP and YKL-40 levels can be used to assess the chondroprotective biological efficacy of therapy. In contrast, changes in GAAGs concentrations reflect systemic extracellular matrix transformations in the course of JIA.

Plasma Glycosaminoglycans in Children with Juvenile Idiopathic Arthritis Being Treated with Etanercept as Potential Biomarkers of Joint Dysfunction.

We assessed the effect of two-year etanercept (ETA) therapy on the metabolism of the cartilage extracellular matrix (ECM) in patients with juvenile idiopathic arthritis (JIA). METHODS: We performed a quantitative evaluation of glycosaminoglycans (GAGs) (performed by the multistage extraction and purification method) in blood obtained from patients before and during 24 months of ETA treatment, as potential biomarker of joint dysfunction and indicators of biological effectiveness of therapy. Since the metabolism of GAGs is related to the activity of proteolytic enzymes and prooxidant-antioxidant factors, we decided to evaluate the relationship between GAGs and the levels of metalloproteinases (MMP), i.e., MMP-1 and MMP-3 (using immunoenzymatic methods), as well as the total antioxidative status (TAS) (using the colorimetric method) in blood of the JIA patients. RESULTS: When compared to the controls, GAGs and TAS concentrations were significantly lower in patients with an aggressive course of JIA qualified for ETA treatment. MMP-1 and MMP-3 levels were significantly higher versus control values. An anti-cytokine therapy leading to clinical improvement does not lead to the normalization of any of the assessed parameters. GAGs concentration is significantly related to MMP-1, MMP-3, TAS, TOS, and CRP levels. CONCLUSION: The results of the present study indicate the necessity of constant monitoring of the dynamics of destructive processes of articular cartilage in children with JIA. We suggest that GAGs may be a useful biomarker to assess the clinical status of the extracellular matrix of joints.

The Effects of TNF-α Inhibition on the Metabolism of Cartilage: Relationship between KS, HA, HAPLN1 and ADAMTS4, ADAMTS5, TOS and TGF-ß1 Plasma Concentrations in Patients with Juvenile Idiopathic Arthritis.

We assessed the effect of 24-month anti-tumor necrosis factor alpha (TNF-α) treatment on the remodeling of the cartilage extracellular matrix (ECM) in patients with juvenile idiopathic arthritis (JIA). METHODS: Quantitative evaluation of keratan sulfate (KS), hyaluronic acid (HA), hyaluronan and proteoglycan link protein 1 (HAPLN1), as potential biomarkers of joint dysfunction, and the levels of a disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS) 4 and 5, total oxidative status (TOS) and transforming growth factor (TGF-ß1) was performed (using immunoenzymatic methods) in blood obtained from patients before and after 24 months of etanercept (ETA) treatment. RESULTS: When compared to the controls, KS, HA and HAPLN1 levels were significantly higher in patients with an aggressive course of JIA qualified for ETA treatment. An anti-cytokine therapy leading to clinical improvement promotes the normalization only of the HA level. Proteolytic and pro-oxidative factors, present in high concentrations in patients before the treatment, correlated with HAPLN1, but not with KS and HA levels. In these patients, negative correlations were found between the levels of TGF-ß1 and KS, HA and HAPLN1. CONCLUSION: The anti-TNF-α therapy used in patients with JIA has a beneficial effect on ECM cartilage metabolism, but it does not completely regenerate it. The changes in the plasma HA level during the anti-cytokine therapy suggest its potential diagnostic utility in monitoring of disease activity and may be used to assess the efficacy of ETA treatment.

Urinary sulphated glycosaminoglycans excretion in obese patients with type 2 diabetes mellitus treated with metformin.

OBJECTIVE: The pattern of urinary excretion of total sulphated glycosaminoglycans (GAGs) and their particular types: chondroitin sulphate/dermatan sulphate (CS/DS) and heparan sulphate (HS) was analysed in obese patients with type 2 diabetes mellitus (T2DM) treated with metformin in monotherapy for the period of six months. METHODS: The urinary sulphated glycosaminoglycans were quantitated using standardised dye (1.9-dimethylmethylene blue)-binding method and normalised to creatinine level. RESULTS: Urinary total GAGs, CS/DS and HS levels were significantly higher in untreated diabetic patients in comparison to healthy subjects. Moreover, it was observed that urinary total GAGs, CS/DS and HS levels in diabetic patients after six-month metformin therapy were significantly decreased versus pre-treatment situation. CONCLUSIONS: The obtained results suggest that the six-month treatment with metformin in obese patients with T2DM has a regulating influence on the systemic changes in proteoglycans/glycosaminoglycans, resulting in a decrease in the urinary excretion of total GAGs, CS/DS and HS.

Concerted Actions by PIICP, CTXII, and TNF-α in Patients with Juvenile Idiopathic Arthritis.

Joint destruction in juvenile idiopathic arthritis (JIA), initiated in the early, preclinical stage of the disease, is diagnosed on the basis of clinical evaluation and radiographic imaging. The determination of circulating cartilage-matrix turnover markers can facilitate the diagnosis and application of better and earlier treatment strategies for JIA. We have shown that 96 JIA patients have elevated levels of procollagen II C-terminal propeptide (PIICP), reflecting the extent of joint cartilage biosynthesis, and C-telopeptide of type II collagen (CTXII), a biomarker of the resorption of this tissue. Patients who did not respond to treatment had particularly high levels of these markers. JIA treatment resulted in the normalization of these markers in remissive patients, but not in those with active JIA. We showed correlations between examined variables and inflammatory process indicators, i.e., C-reactive protein (CRP), erythrocyte sedimentation rate (ESR), and tumor necrosis factor-α (TNF-α). The TNF-α of patients responding to treatment correlated with PIICP, especially in the patients before treatment (r = 0.898, p < 0.001). Significant changes in serum PIICP during JIA therapy suggest its potential diagnostic utility in the monitoring of disease activity and the possibility of its use in assessing treatment towards remission. Understanding changes in type II collagen metabolism over the course of the discussed arthritis may allow the implementation of both new diagnostic tools and new therapeutic strategies in children with JIA.

Plasma Glycosaminoglycan Profiles in Systemic Sclerosis: Associations with MMP-3, MMP-10, TIMP-1, TIMP-2, and TGF-Beta.

The aim of the study was to determine whether plasma levels of total glycosaminoglycans (GAGs), matrix metalloproteinases (MMPs) (MMP-3, MMP-10), and their tissue inhibitors (TIMPs) (TIMP-1, TIMP-2) as well as transforming growth factor ß (TGF-ß) differ in the patients with systemic sclerosis (SSc) in relation to the healthy subjects. Plasma samples were obtained from 106 people (64 patients with SSc and 42 healthy individuals) and measured for MMP-3, MMP-10, TIMP-1, TIMP-2, and TGF-ß levels using ELISA methods. GAGs isolated from plasma samples were quantified using a hexuronic acid assay. The plasma levels of total GAGs, TIMP-1, TIMP-2, and TGF-ß were significantly higher, while MMP-3 was significantly decreased in SSc patients compared to the controls. We have revealed a significant correlation between plasma GAGs and TGF-ß (r = -0.47) and TIMP-2 (r = 0.38), respectively. The results of this study revealed that remodeling of the extracellular matrix, reflected by quantitative changes in plasma glycosaminoglycans, occurs during systemic sclerosis. Thus, the alterations in GAG metabolism connected with SSc may lead to systemic changes in the properties of the connective tissue extracellular matrix.

Significant Remodeling Affects the Circulating Glycosaminoglycan Profile in Adult Patients with both Severe and Mild Forms of Acute Pancreatitis.

Acute pancreatitis (AP) manifests itself either as a mild, self-limiting inflammation or a severe, systemic inflammatory process that is associated with various complications and a high mortality rate. It is unknown whether these two forms of the disease can differ in the profile of circulating glycosaminoglycans, which are molecules with huge biological reactivity due to a high density of negative electric charge. Plasma glycosaminoglycans were characterized/quantified in 23 healthy controls, 32 patients with mild AP, and 15 individuals with severe disease using electrophoresis with enzymatic identification (chondroitin sulfate and heparan sulfate) or an ELISA-based test (hyaluronan). Moreover, the correlations between the glycosaminoglycan levels and clinical parameters were evaluated. Both forms of AP showed similar remodeling of the plasma profile of the sulfated glycosaminoglycans. In contrast, only in the patients with mild AP was the level of circulating hyaluronan significantly decreased as compared to the healthy controls. Both forms of AP are associated with systemic changes in the metabolism of glycosaminoglycans. However, the alterations in hyaluronan metabolism may contribute to the disease evolution. The circulating hyaluronan may have some clinical value to predict the severity of AP and to evaluate the clinical status of patients with severe AP.

Association of Circulating COMP and YKL-40 as Markers of Metabolic Changes of Cartilage with Adipocytokines in Juvenile Idiopathic Arthritis.

The aim of this study was to evaluate the association of circulating cartilage oligomeric matrix protein (COMP) and human cartilage glycoprotein-39 (YKL-40) as markers of metabolic changes of cartilage, with leptin, adiponectin, and resistin in juvenile idiopathic arthritis (JIA) patients before and after treatment. A significant decrease of COMP and an increase of YKL-4 were found in blood of untreated patients. JIA treatment leading to clinical improvement resulted in normalization of COMP levels only. Concentrations of both markers in treated patients, while showing no clinical improvement, differed from those in controls and patients with remission. The leptin level decreased (p < 0.05) in untreated patients; however, concentrations of adiponectin and resistin increased (p < 0.05) as compared to controls. JIA treatment resulted in normalization of adipocytokine levels in remissive patients but not those with active JIA. Untreated patients showed a correlation between COMP and leptin, adiponectin, and body mass index (BMI) and between YKL-40 and leptin, adiponectin, BMI, C-reactive protein (CRP), and erythrocyte sedimentation rate (ESR). In inactive JIA, a correlation between YKL-40 and leptin was shown. Treated patients with an active JIA demonstrated a correlation between COMP and adiponectin and between YKL-40 and leptin, adiponectin, BMI, CRP, and ESR. The results of this work indicate that leptin and adiponectin but not resistin may be involved in the development and progression of joint dysfunction in JIA. Additionally, we suggest that YKL-40 may be a useful biomarker of disease activity and may be used to assess treatment towards remission, as compared to COMP.

Circulating C1q/TNF-Related Protein 3, Omentin-1 and NGAL in Obese Patients with Type 2 Diabetes During Insulin Therapy.

The aim of the study was to quantify the plasma concentration of omentin-1, neutrophil gelatinase-associated lipocalin (NGAL), and complement C1q tumor necrosis factor-related protein-3 (CTRP3) in obese patients with type 2 diabetes, before introducing insulin therapy, in relation to the plasma expression profiles of these regulatory molecules in the same patients after a 6-month insulin mixture therapy and in obese controls. Elevated plasma NGAL concentrations were found in type 2 diabetic patients as compared with subjects with metabolically healthy obesity. In turn, a 6-month insulin mixture therapy has shown a marked increase in the plasma concentration of omentine-1 and a significant decrease in plasma CTRP3 concentration in obese patients with type 2 diabetes, in relation to the values found in these patients before the implementation of insulin therapy. Insulin mixture therapy has also proved to be an important factor modifying the plasma profile of NGAL, increasing the concentration of this bioactive molecule in the plasma of patients with type 2 diabetes, after 6 months of its use, in relation to the concentration before treatment. The significant changes in the plasma profile of omentin-1, NGAL and CTRP3 during insulin therapy suggest their potential diagnostic utility in monitoring metabolic changes associated with the introduction of insulin treatment in type 2 diabetic patients.

Adiponectin, Leptin, and Leptin Receptor in Obese Patients with Type 2 Diabetes Treated with Insulin Detemir.

The aim of the present study is to quantitatively assess the expression of selected regulatory molecules, such as leptin, leptin receptor, and adiponectin in the blood of obese patients with type 2 diabetes both before treatment and after six months of pharmacological therapy with the long-lasting insulin analogue, insulin detemir. A significant decrease in the analysed regulatory molecules, i.e., leptin receptor and adiponectin, was found in blood plasma of the patients with untreated type 2 diabetes. These changes were accompanied by an increase in plasma leptin concentrations. Insulin treatment resulted in the normalization of plasma leptin receptor and adiponectin concentrations. The circulating leptin level did not change following anti-diabetic therapy with insulin detemir. Gender was a significant factor modifying the circulating level of all the analysed regulatory active compounds. Bioinformatic analysis was performed using Matlab with the Signal Processing Toolbox. The conducted discriminant analysis revealed that the leptin receptor, Δw(19), and adiponectin, Δw(21), were the parameters undergoing the most significant quantitative changes during the six-month therapy with insulin detemir. The conducted examinations indicated the contribution of adipocytokines-the biologically-active mediators of systemic metabolism, such as leptin and adiponectin in the pathomechanism of disorders being the basis for obesity which leads to development of insulin resistance, which, in turn, results in the occurrence of type 2 diabetes.

Laboratory Indicators of Aggrecan Turnover in Juvenile Idiopathic Arthritis.

OBJECTIVES: Evaluation of chondroitin sulfate (CS), as an early marker of aggrecan degradation, and chondroitin sulfate 846 epitope (CS846), as a biomarker of CS synthesis, is an attempt at answering the question whether the therapy used in juvenile idiopathic arthritis (JIA) patients contributes to the normalization of biochemical changes in aggrecan. METHODS AND RESULTS: Serum levels of CS and CS846 as well as catalase (CT), superoxide dismutase (SOD), and glutathione peroxidase (GPx) activities in erythrocyte were assessed in patients before and after treatment. In the course of JIA, aggrecan metabolism is disturbed, which is reflected by a decrease (p < 0.001) in CS serum level and an increase (p < 0.05) in CS846 concentration. Furthermore, increased (p < 0.001) activities of CT, SOD, and GPx in untreated JIA patients were recorded. The anti-inflammatory treatment resulted in the normalization of CS846 level and SOD and GPx activities. In untreated patients, we have revealed a significant correlation between serum CS and CS846, CT, CRP, ESR, MMP-3, and ADAMTS-4, respectively, as well as between CS846 and CT, GPx, CRP, ESR, and TGF-ß1, respectively. CONCLUSION: The observed changes of CS and CS846 in JIA patients indicate a further need of the therapy continuation aimed at protecting a patient from a possible disability.

Relationship between adiponectin, leptin, IGF-1 and total lipid peroxides plasma concentrations in patients with systemic sclerosis: possible role in disease development.

AIM: The relationship between adiponectin, leptin, insulin-like growth factor-1 (IGF-1) and total lipid peroxide (TLP) concentrations, and its possible role in the development of diffuse cutaneous systemic sclerosis (dcSSc), were evaluated in this study. METHODS, RESULTS: Plasma adipokines and IGF-1 levels were determined using the enzyme-linked immunosorbent assay method, whereas TLP levels were determined using a photometric test, in 36 dcSSc patients and 40 healthy controls matched by age, sex and body mass index (BMI). Plasma levels of adipokines were significantly lowered, while TLP and IGF-1 were increased in dcSSc patients compared to controls. Adiponectin correlated significantly with leptin (r = 0.44), TLP (r = -0.54), CRP (r = -0.47), erythrocyte sedimentation rate (ESR) (r = -0.40) and duration of disease (r = -0.44). A significant relationship was found between leptinemia and IGF-1 (r = -0.40), TLP (r = 0.44), duration of disease (r = -0.38) and BMI (r = 0.65). TLP correlated with IGF-1 (r = -0.43), C-reactive protein (r = 0.47), ESR (r = 0.49) and duration of disease (r = 0.46), while IGF-1 correlated with ESR (r = -0.40). CONCLUSIONS: Adipose tissue may play a complex role in the development of dcSSc, affecting both the metabolic state of the organism, as well as free radical-induced connective tissue degradation. Although, leptin seems to exert a pro-oxidative effect and both adiponectin and IGF-1 appear to prevent free radical damage, confirmation of the above effects requires further research.

Plasma and urinary glycosaminoglycans in the course of juvenile idiopathic arthritis.

OBJECTIVES: The aim of the study was to perform analyses of plasma and urinary glycosaminoglycan isolated from juvenile idiopathic arthritis (JIA). METHODS, RESULTS: Chondroitin/dermatan sulfate (CS/DS), heparan sulfate/heparin (HS/H) and hyaluronic acid (HA) were evaluated in samples obtained from JIA patients before and after treatment. Electrophoretic analysis of GAGs identified the presence of CS, DS and HS/H in plasma of healthy subjects and JIA patients. CS were the predominant plasma GAGs constituent in all investigated subject. The plasma CS level in untreated patients was significantly decreased. Therapy resulted in an increase in this glycan level. However, plasma CS concentration still remained higher than in controls. Increased levels of DS and HA in untreated JIA patients were recorded. Anti-inflammatory treatment led to normalization of these parameters concentrations. Plasma and urinary concentrations of HS/H were similar in all groups of individuals. Urinary CS/DS and HA were decreased only in untreated patients. CONCLUSIONS: The data presented indicate that changes in plasma and urinary glycosaminoglycan occur in the course of JIA. There are probably the expression of both local articular cartilage matrix and systemic changes in connective tissue remodeling.

Circulating keratan sulfate as a marker of metabolic changes of cartilage proteoglycan in juvenile idiopathic arthritis; influence of growth factors as well as proteolytic and prooxidative agents on aggrecan alterations.

BACKGROUND: The aim of this study was to evaluate the plasma keratan sulfate (KS) level as a potential marker of joint damage in children with juvenile idiopathic arthritis (JIA). The influence of growth factors as well as proteolytic and prooxidative agents on aggrecan alterations were evaluated in this study. METHODS: Plasma levels of KS, transforming growth factor ß1 (TGF-ß1), platelet-derived growth factor BB (PDGF-BB), a disintegrin and metalloproteinase with thrombospondin motifs 4 and 5 (ADAMTS-4 and ADAMTS-5), and thiol groups (TG) were quantified in samples obtained from 30 healthy subjects and 30 patients with JIA before and after treatment. RESULTS: Increased (p<0.01) plasma KS was observed in JIA patients before treatment. Therapy resulted in a decrease in KS level. However, plasma KS level remained higher (p<0.05) than in controls. Increased levels of TGF-ß1 (p<0.01) and PDGF-BB (p<0.05) in untreated JIA patients were recorded. Clinical improvement was accompanied by significant decrease in TGF-ß1 and PDGF-BB, compared with a pretreatment condition and a control group. The concentrations of proteinases were characterized by different trends of alterations. When the ADAMTS-4 level increased (p<0.01) in the blood of untreated patients, the concentration of ADAMTS-5 was found to be reduced (p<0.0001), compared with controls. JIA treatment resulted in the normalization of ADAMTS-4 level. Plasma TG concentration was decreased only in untreated patients (p<0.05). We have revealed a significant correlation between plasma KS level and ADAMTS-4, TGF-ß1, TG, C-reactive protein, and erythrocyte sedimentation rate levels. CONCLUSIONS: Plasma KS level in JIA patients, reflecting the aggrecan structure, indicates that treatment that modifies inflammation simultaneously does not contribute to total regeneration of articular matrix components and signalizes the need for further treatment.

Influence of proteolytic-antiproteolytic enzymes and prooxidative-antioxidative factors on proteoglycan alterations in children with juvenile idiopathic arthritis.

OBJECTIVES: The influence of proteolytic-antiproteolytic enzymes and prooxidative-anti-oxidative factors on proteoglycan alterations in children with juvenile idiopathic arthritis (JIA) was evaluated in this study. DESIGN, METHODS, RESULTS: Plasma and urinary glycosaminoglycans (GAGs), as well as plasma levels of matrix metalloproteinases (MMPs) (MMP-3, MMP-10), tissue inhibitors of metalloproteinases (TIMPs) (TIMP-1, TIMP-2), total oxidative status (TOS) and total antioxidative status (TAS), were quantified in samples obtained from 30 healthy subjects and 30 JIA patients before and after treatment. Significantly decreased plasma and urinary concentration of GAGs in JIA patients before treatment was observed. Therapy resulted in an increase in the concentration of the above listed parameters. However, the plasma GAG level still remained significantly lower compared to that in controls. Increased levels of MMP-3 and TIMP-1 in both JIA patient groups were recorded. The plasma MMP-10 and TIMP-2 concentrations in untreated patients were significantly decreased. Anti-inflammatory treatment led to normalization of these parameter concentrations. Significant increase of TOS but decrease of TAS was found in the blood of untreated patients. The treatment resulted only in the normalization of TOS concentration. We have revealed a significant correlation between plasma GAGs and: MMP-3 (r=0.54), TOS (r=0.64) and urinary GAGs (r=0.55), respectively. CONCLUSIONS: Proteoglycan/glycosaminoglycan alterations in JIA patients, which are stimulated by MMP-3 and reactive oxygen species (ROS), indicate rather systemic disturbance of extracellular matrix metabolism, and not merely local changes which occur in articular structures. Given the destructive potential of ROS and MMPs and their hyperexpression in JIA, inhibition of these compounds should bring a substantial clinical benefit.

Antioxidant activity and structural modifications of serum chondroitin sulfate in Graves' disease.

OBJECTIVES: Disturbances in the metabolism of proteoglycans/glycosaminoglycans related to free radical overproduction can be detected in Graves' ophthalmopathy. However, it is not evident whether these disorders appear before the clinical manifestation of ophthalmopathy and contribute to the development of nonthyroidal symptoms. They might also reflect the body's defence against the effects of an autoimmune attack, or they merely result from these disorders. Our study conducted in Graves' patients, free of ocular changes at the time of recruitment, is an attempt at explaining these ambiguities. We decided to investigate the relation between the changes in extracellular matrix components, namely the total glycosaminoglycans (GAGs) and chondroitin sulfates (CS), as well as parameters characterising oxidative stress, i.e. the total antioxidant status (TAS), and the thiol group (TG) concentration. DESIGN, METHODS, RESULTS: GAGs, TAS and TG were quantified in serum samples obtained from 30 healthy subjects and 30 Graves' patients before and after treatment. An increased serum concentration of total GAGs and CS in Graves' patients before and after treatment was observed. Additionally, increased sulfation of CS in both Graves' patient groups was recorded. The serum TG and TAS concentrations in untreated patients were significantly decreased. Subsequent euthyreosis led to the normalisation of TG concentration. We have revealed a significant correlation between changes of extracellular matrix components, especially chondroitin-4-sulfate, thyroid status parameters, and parameters characterising oxidative stress. CONCLUSIONS: The structural modifications of serum CS in Graves' patients clinically free of ocular complications are probably the expression of systemic connective tissue remodelling. Simultaneously, these modifications may participate in a defence response to free radical damage underlying this pathology.

Age- and gender-related alteration in plasma advanced oxidation protein products (AOPP) and glycosaminoglycan (GAG) concentrations in physiological ageing.

BACKGROUND: The authors studied the role of increased oxidative stress in the development of oxidative protein damage and extracellular matrix (ECM) components in ageing. The age- and gender-associated disturbances in connective tissue metabolism were evaluated by the plasma chondroitin sulphated glycosaminoglycans (CS-GAG) and non-sulphated GAG-hyaluronan (HA) measurements. Plasma concentration of advanced oxidation protein products (AOPP) was analysed in order to assess oxidative protein damage and evaluate the possible deleterious role of oxidative phenomenon on tissue proteoglycans' metabolism during the physiological ageing process. METHODS: Sulphated and non-sulphated GAGs as well as AOPP were quantified in plasma samples from 177 healthy volunteers. RESULTS: A linear age-related decline of plasma CS-GAG level was found in this study (r=-0.46; p<0.05). In contrast, HA concentrations rise gradually with age (r=0.44; p<0.05) in plasma samples. For both ECM components, the observed differences were not gender-specific. A strong age-dependent relationship has been shown in regard to AOPP. AOPP levels significantly increased with age (r=0.63; p<0.05), equally strongly in both men (r=0.69; p<0.05) and women (r=0.57; p<0.05) during physiological ageing. A significant correlation was found between the concentrations of AOPP and both CS-GAG (r=-0.31; p<0.05) and HA (r=0.33; p<0.05). CONCLUSIONS: Proceeding with age changes in the ECM are reflected by CS-GAG and HA plasma levels. Strong correlations between AOPP and ECM components indicate that oxidative stress targets protein and non-protein components of the connective tissue matrix during human ageing.

Age- and gender-dependent changes in connective tissue remodeling: physiological differences in circulating MMP-3, MMP-10, TIMP-1 and TIMP-2 level.

BACKGROUND: the mechanisms which cause age-dependent remodeling of connective tissue are still not fully understood. Matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs) constitute an important proteolytic pathway affecting physiological matrix remodeling. OBJECTIVE: the way in which changes in the extracellular matrix metabolism during the ageing process influence the level of circulating MMP-3 and MMP-10, as well as their tissue inhibitors TIMP-1 and TIMP-2, in a healthy population was investigated in this study. METHODS: blood samples were taken from 81 healthy individuals aged 6-62 years and measured for MMP-3, MMP-10, TIMP-1 and TIMP-2 levels using enzyme-linked immunosorbent assays. Polyacrylamide gel electrophoresis followed by Western immunoblotting allowed for the detection of pro- and active forms of both MMPs. RESULTS: Serum MMP-3 and TIMP-1 values were positively correlated with age (r = 0.44, p = 0.00001 and r = 0.28, p = 0.012, respectively). A contrary tendency was found for MMP-10 and TIMP-2 serum levels. A strong age-related decrease in MMP-10 (-0.53; p = 0.000) and TIMP-2 (-0.52; p = 0.000) was noticed in our study. Gender was a significant factor modifying MMP/TIMP potential, except for the MMP-10 level. CONCLUSIONS: the data presented indicate that changes in MMP/TIMP balance occur in physiological ageing. Moreover, these findings highlight the necessity of utilizing age- and sex-matched values for analysis of MMPs and TIMPs in the pathological conditions.