Urinary methotrexate dosage in rheumatoid arthritis, in patients treated for at least 6 months: a potential marker of adherence.

OBJECTIVES: Non-adherence to rheumatoid arthritis (RA) treatments must be identified. A methotrexate (MTX) urinary dosage (METU) was recently developed. The aim of our study was to assess adherence to MTX in RA using METU in real-life conditions and to compare it with indirect adherence measurement technics. METHODS: We performed a cross-sectional study at Reims University Hospital. We included over 18-year-old patients with RA treated by MTX for more than 6 months. Patients were invited to complete demographic, clinical and psychological questionnaires and adherence measurement technics (Compliance Questionnaire of Rheumatology (CQR) and Medication Possession Ratio (MPR)). A urinary sample was collected to measure MTX and information about tolerance was evaluated through Methotrexate Intolerance Severity Score. RESULTS: 84 patients were included, 26 using oral MTX, 58 subcutaneous (SC) MTX. Among them, 73% were female, mean age was 61.5 years, MTX mean dose was 15 mg/week and 61.9% were treated by biological DMARDs (Disease Modifying Antirheumatic Drugs). 77 patients (91.7%) were adherent to treatment according to METU, whereas MPR and CQR reported less adherence (69.5% and 61.9%, respectively). MPR and METU were not significantly different in SC MTX users (p=0.059). Non-adherent patients had a higher number of tender joints and C reactive protein value (p<0.05). CONCLUSION: This is the first largest study evaluating MTX adherence in patients with RA using a urinary dosage. We identified that indirect adherence measurements did not reflect real-life adherence. It would be appreciable to realise METU, in a new study, in patients with RA with unexplained response to treatment, to consider it before escalating therapeutic strategy.

Serum and urine lipidomic profiles identify biomarkers diagnostic for seropositive and seronegative rheumatoid arthritis.

Objective: Seronegative rheumatoid arthritis (RA) is defined as RA without circulating autoantibodies such as rheumatoid factor and anti-citrullinated protein antibodies; thus, early diagnosis of seronegative RA can be challenging. Here, we aimed to identify diagnostic biomarkers for seronegative RA by performing lipidomic analyses of sera and urine samples from patients with RA. Methods: We performed untargeted lipidomic analysis of sera and urine samples from 111 RA patients, 45 osteoarthritis (OA) patients, and 25 healthy controls (HC). These samples were divided into a discovery cohort (n = 97) and a validation cohort (n = 84). Serum samples from 20 patients with systemic lupus erythematosus (SLE) were also used for validation. Results: The serum lipidome profile of RA was distinguishable from that of OA and HC. We identified a panel of ten serum lipids and three urine lipids in the discovery cohort that showed the most significant differences. These were deemed potential lipid biomarker candidates for RA. The serum lipid panel was tested using a validation cohort; the results revealed an accuracy of 79%, a sensitivity of 71%, and a specificity of 86%. Both seropositive and seronegative RA patients were differentiated from patients with OA, SLE, and HC. Three urinary lipids showing differential expression between RA from HC were identified with an accuracy of 84%, but they failed to differentiate RA from OA. There were five lipid pathways that differed between seronegative and seropositive RA. Conclusion: Here, we identified a panel of ten serum lipids as potential biomarkers that can differentiate RA from OA and SLE, regardless of seropositivity. In addition, three urinary lipids had diagnostic utility for differentiating RA from HC.

Relationship Between Inflammation and Metabolism in Patients With Newly Presenting Rheumatoid Arthritis.

Background: Systemic inflammation in rheumatoid arthritis (RA) is associated with metabolic changes. We used nuclear magnetic resonance (NMR) spectroscopy-based metabolomics to assess the relationship between an objective measure of systemic inflammation [C-reactive protein (CRP)] and both the serum and urinary metabolome in patients with newly presenting RA. Methods: Serum (n=126) and urine (n=83) samples were collected at initial presentation from disease modifying anti-rheumatic drug naïve RA patients for metabolomic profile assessment using 1-dimensional 1H-NMR spectroscopy. Metabolomics data were analysed using partial least square regression (PLS-R) and orthogonal projections to latent structure discriminant analysis (OPLS-DA) with cross validation. Results: Using PLS-R analysis, a relationship between the level of inflammation, as assessed by CRP, and the serum (p=0.001) and urinary (p<0.001) metabolome was detectable. Likewise, following categorisation of CRP into tertiles, patients in the lowest CRP tertile and the highest CRP tertile were statistically discriminated using OPLS-DA analysis of both serum (p=0.033) and urinary (p<0.001) metabolome. The most highly weighted metabolites for these models included glucose, amino acids, lactate, and citrate. These findings suggest increased glycolysis, perturbation in the citrate cycle, oxidative stress, protein catabolism and increased urea cycle activity are key characteristics of newly presenting RA patients with elevated CRP. Conclusions: This study consolidates our understanding of a previously identified relationship between serum metabolite profile and inflammation and provides novel evidence that there is a relationship between urinary metabolite profile and inflammation as measured by CRP. Identification of these metabolic perturbations provides insights into the pathogenesis of RA and may help in the identification of therapeutic targets.

Rheumatoid arthritis in low-level toluene-exposed workers based on nationwide medical surveillance data in Korea.

BACKGROUND: There is growing evidence that exposure to organic solvents can play a role in the etiology of rheumatoid arthritis (RA). This prospective cohort study aimed to investigate the association between RA and toluene exposure. METHODS: The study cohort consisted of Korea Occupational Safety and Health Agency data from male workers exposed to toluene who had undergone a toluene-associated special medical examination at least once between January 1, 2000 and December 31, 2004 (n = 148,870). The morbidity from RA based on hospital admission records was estimated from 2000 to 2005 using National Health Insurance Claim Data. The standardized admission ratio (SAR) for RA was calculated with reference to the general population. Levels of urinary hippuric acid (HA), a metabolite of toluene, were measured and used for exposure assessment. RESULTS: Toluene-exposed workers were at an elevated risk of seropositive rheumatoid arthritis (ICD-10 code M05) with an SAR of 2.38 (95% confidence interval [CI]: 1.14-4.37) and other rheumatoid arthritis (M06) with an SAR of 1.22 (95% CI: 0.91-1.59). When data were stratified according to the duration of toluene exposure and by tertiles of urinary HA level, no significant difference was apparent. CONCLUSION: SARs of the toluene-exposed workers are higher than that of the general reference population, indicating that exposure to toluene may contribute to an increased risk of RA. Further studies of toluene-exposed workers with longer follow-up are needed.

A wide-targeted urinary and serum metabolomics strategy reveals the effective substance of the Wu-tou decoction.

As an important Chinese medicine decoction, Wu-tou decoction has been used to treat rheumatic arthritis for more than a thousand years. We previously reported that the Wu-tou decoction could change the urinary and serum metabolites in adjuvant-induced arthritis rats significantly. The purpose of this research was to confirm the potential biomarkers obtained by previous non-targeted metabolomics study through quantitative analysis by liqui chromatography with tandem mass spectrometry, in the meantime, to further study the effective material basis of Wu-tou decoction. Firstly, the important compounds in the tryptophan metabolism pathway, the arginine and proline metabolism pathway, the amino acid metabolism pathway, the tricarboxylic acid cycle, the vitamin B6 metabolism pathway, and the phenylalanine metabolism pathway, which were identified as potential biomarkers in previous study, were selected for quantitative analysis. Then the linearity, limit of detection, limit of quantification, selectivity, accuracy, precision, stability, recovery, and matrix effect of the quantitative method were examined. Finally, ten and eighteen metabolites were quantitatively analyzed in the serum and urine, respectively. The results showed that seven out of ten serum potential biomarkers and ten out of eighteen urine potential biomarkers were confirmed as real biomarkers. This research provides a powerful reference for the study on effective material basis of Wu-tou decoction.

Metabolomic approach to the exploration of biomarkers associated with disease activity in rheumatoid arthritis.

We aimed to investigate metabolites associated with the 28-joint disease activity score based on erythrocyte sedimentation rate (DAS28-ESR) in patients with rheumatoid arthritis (RA) using capillary electrophoresis quadrupole time-of-flight mass spectrometry. Plasma and urine samples were collected from 32 patients with active RA (DAS28-ESR≥3.2) and 17 with inactive RA (DAS28-ESR<3.2). We found 15 metabolites in plasma and 20 metabolites in urine which showed a significant but weak positive or negative correlation with DAS28-ESR. When metabolites between active and inactive patients were compared, 9 metabolites in plasma and 15 in urine were found to be significantly different. Consequently, we selected 11 metabolites in plasma and urine as biomarker candidates which significantly correlated positively or negatively with DAS28-ESR, and significantly differed between active and inactive patients. When a multiple logistic regression model was built to discriminate active and inactive cohorts, three variables-histidine and guanidoacetic acid from plasma and hypotaurine from urine-generated a high area under the receiver operating characteristic (ROC) curve value (AUC = 0.8934). Thus, this metabolomics approach appeared to be useful for investigating biomarkers of RA. Combination of plasma and urine analysis may lead to more precise and reliable understanding of the disease condition. We also considered the pathophysiological significance of the found biomarker candidates.

Innovative HPTLC method for simultaneous determination of ternary mixture of certain DMARDs in real samples of rheumatoid arthritis patients: an application of quality by design approach.

A uniquely developed high performance thin-layer chromatographic (HPTLC) method coupled with UV detection was applied using quality by design approach (QbD) for simultaneous determination of methotrexate (MTX), sulfasalazine (SSZ) and hydroxychloroquine (HCQ) in serum and urine samples of rheumatoid arthritis patients. MTX, SSZ with HCQ are the most common disease-modifying antirheumatic drugs (DMARDs) combination used for the treatment of rheumatoid arthritis. This ternary mixture with montelukast (MK) added as internal standard, were separated by using a mixture of ethyl acetate: methanol: 25% ammonia, (8: 2: 3, v/v/v) as a mobile phase system. The separation was achieved on HPTLC precoated silica gel plate 60 F254 and the detection was carried out at 306 nm for MTX and at 340 nm for both SSZ and HCQ. The design was planned to obtain the most optimum retardation factors (Rf) with best resolution. The Rf values for MTX, SSZ, MK and HCQ were of 0.31 ±â€¯0.03, 0.62 ±â€¯0.02, 0.71 ±â€¯0.02 and 0.83 ±â€¯0.03; respectively. The interactive response optimizer achieved the most favorable conditions with acceptable composite desirability of 0.9703. Linear relationship with good correlation coefficients (r = 0.9990-0.9994) were also obtained with detection and quantification limits of 13.94-260.64 and 46.84-1810.01(ng/ml); respectively. The suggested method was established in accordance with the guidelines of Food and Drug Administration (FDA). The established QbD-HPTLC method achieved simple, sensitive and selective quantification of the studied drugs in serum and urine samples in the presence of their metabolites with no interferences. This method can be extended effectively for therapeutic drug monitoring and pharmacokinetics studies of these drugs.

NEPHROPATHY IN EARLY RHEUMATOID ARTHRITIS PATIENTS: DOES A SIGNIFICANT RISK EXIST?

Objective of the study layed in assessment of the pathophysiological relation between cell-mediated immunity (tumor necrosis factor-alpha (TNF-α) inflammatory cytokine) activation and renal dysfunction in the patients with early rheumatoid arthritis. We analyzed the data from 35 early rheumatoid arthritis (RA) patients of average age of 50,71±2,25 years (ranged 18-76 years, 80% of women) with 9,21±0,43 months mean duration of the disease by the time of the study initiation. Urine and blood tests were performed to verify the main indicators of kidney function and inflammation cytokines significant interaction. All signs of renal dysfunction at the baseline in the patients with early RA were associated with glomerular filtration rate decrease and excretion of urine protein increase. Dynamics of albuminuria, according to the analysis of variance for one-factor scheme, were significantly determined by the state of disease activity, reflecting the severity of joint damage. High urine ß-2-microglobulin level was significantly associated with the expression rate of main inflammatory cytokines as per binary regression analysis. The obtained dependence showed the dynamics of expression of tubular disorders in early RA with a progressive deterioration which did associate with the levels of TNF-α expression, and variety of the urine miсroglobulin rates in the interval 200-350 µg/L. Reliable correlation (r=0.51, p<0.05) between beta-2-microglobulinuria and TNF-α levels was clearly shown, revealing the relationship described by the formula MGU=- 81+937×log10 (TNF-α) as per regression analysis. The severity of tubular damage in early RA is associated with TNF-α expression, especially in the patients with TNF-α above 250 pg/mL, when microalbuminuria rates were significantly higher (p=0.00043). We identified robust data that in the early RA patients with high TNF-α, the number of reported cases of microalbuminuria was significantly higher than in those with low levels.

Urinary Orosomucoid - 2 and Soluble CD14 as Potential Biomarkers for Assessment of Disease Activity in Rheumatoid Arthritis.

Rheumatoid arthritis (RA) is the most common inflammatory joint disease leading to severe disability and premature mortality. Current blood biomarkers for assessing RA activity are invasive and are not highly sensitive or specific to changes in disease activity. Therefore, there is a need for new biomarkers that can accurately indicate disease activity. The present study evaluated the use of urinary orosomucoid (ORM) - 2 and soluble CD14 (sCD14) as non-invasive biomarkers for precise assessment of disease activity of RA, in order to improve treatment outcomes in RA patients. The study included 36 female patients with RA, were divided into three groups of mild, moderate and severe disease activity according to disease activity score 28 (DAS 28) based on erythrocyte sedimentation rate (ESR) and were compared to control group. Urinary levels of ORM-2 and sCD14 were measured by ELISA. All patients showed significant increase in urinary ORM-2 and sCD14 levels in comparison to controls. There were significant positive correlations of urinary ORM-2 and sCD14 levels with DAS28score and also with the conventional blood biomarkers (C- reactive protein (CRP) and ESR). The receiver operating characteristic curve analysis revealed that both urinary ORM-2 and sCD14 have higher predictive value for disease activity than CRP and ESR. In conclusion, Urinary ORM-2 and sCD14 levels were increased in patients with RA and were correlated with the disease activity. Thus, the urinary biomarkers might be able to replace blood measures for RA activity as they could provide non-invasive and precise assessment of disease activity in RA patients.

Quantitative assessment of betamethasone dual-acting formulation in urine of patients with rheumatoid arthritis and ankylosing spondylitis after single-dose intramuscular administration and its application to long-term pharmacokinetic study.

Quantitative evaluation and assessment of pharmacokinetic parameters of Diprospan® (suspension for injection 7mg/mL (2mg+5mg/mL) of betamethasone) were performed in urine samples taken from patients with rheumatoid arthritis or ankylosing spondylitis for 28days after systemic intramuscular administration in routine clinical practice in an open-comparative prospective cohort study. The maximum betamethasone concentration was reached at day 4 of the follow-up; in some cases, ß-phase of elimination of the drug was appeared at day 14 or at day 21 of the follow-up. The deferred ß-phase elimination was likely a consequence of the physiological characteristics of the patients or of the influence of non-steroidal agents. The half-life of betamethasone was 8.5days. The elimination rate constant was 2.49h-1; the mean clearance was 4.72L/d. The recommended frequency of the drug administration to its complete elimination was estimated up to 48days. Mann-Whitney test showed no significant differences in pharmacokinetic characteristics between male and female subjects. The prolonged elimination phase was observed in patients with deviations in their body mass index, continual treatment by diclofenac and nimesulide or, possibly, after consuming an alcohol. The study was recorded in Clinical Trials open source with identifier NCT03119454.

Association between urinary sodium and potassium excretion and blood pressure and inflammation in patients with rheumatoid arthritis.

Hypertension is highly prevalent in patients with rheumatoid arthritis (RA). In other populations, high sodium (Na+) and low potassium (K+) intake are associated with an increased risk of hypertension, and in animal models, a high salt intake exacerbated arthritis. Patients with RA have many comorbidities associated with salt sensitivity, but their salt intake and its relationship to blood pressure and inflammation is unknown. Using the Kawasaki formula, Na+ and K+ urinary excretion (reflecting intake) was estimated in 166 patients with RA and 92 controls, frequency matched for age, sex, and race. Inflammatory markers and disease activity were measured in RA patients. We tested the associations between blood pressure and Na+ and K+ excretion. Estimated 24-h Na+ excretion was similarly high in both RA (median [IQR] 5.1 g, [3.9-6.6 g]) and controls (4.9 g, [4.0-6.5 g]), p = 0.9, despite higher rates of hypertension in RA (54 vs. 39%, p = 0.03). The Na+:K+ excretion ratio was significantly higher in RA (2.0 [1.6-2.4]) vs. 1.7 [1.5-2.1]), p = 0.02] compared to controls. In RA, a lower K+ excretion was inversely correlated with diastolic blood pressure (adjusted ß = - 1.79, p = 0.04). There was no significant association between Na+ or K+ excretion and inflammatory markers. Despite a similar Na+ excretion, patients with RA had higher rates of hypertension than controls, a finding compatible with increased salt sensitivity. Patients with RA had a lower Na+:K+ excretion ratio than controls, and lower K+ excretion was associated with higher diastolic blood pressure in RA.

[Effects of heat-reinforcing acupuncture on urine metabolites in rheumatoid arthritis rabbits].

OBJECTIVE: To compare the effects of different acupuncture methods on urine metabolites in rheumatoid arthritis (RA) rabbits, and to explore the specificity mechanism of heat-reinforcing acupuncture for RA. METHODS: A total of 40 clean purple-blue rabbits were randomly allocated to a normal group, a model group, a mild reinforcing-reducing needling (MRRN) group, a twirling-reinforcing needling (TRN) group and a heat-reinforcing needling (HRN) group, 8 rabbits in each one. Except the normal group, the rabbits in the remaining groups were treated with ovalbumin and freezing to establish RA model. The rabbits in the MRRN group, TRN group and HRN group were treated with MRRN, TRN and HRN at "Zusanli" (ST 36), respectively, 30 min per treatment, once a day for seven days. After treatment, 24-h urine was collected. The rabbits were sacrificed to collect synovial tissues of knee to perform morphology observation; the liquid chromatography quadrupole time-of-flight mass spectrometry (LC-Q/TOF-MS) was applied to measure urine metabolites. All the data were analyzed by principal component analysis (PCA) and partial least squares discriminant analysis (PLS-DA). RESULTS: Compared with the normal group, the leucine-related metabolites, as main urine metabolites, were decreased in the model group (P<0.05), while the purine-related metabolites and tryptophane-related metabolites were increased (P<0.05). Compared with the model group, the leucine-related metabolites, as main urine metabolites, were increased in the three needling groups after treatment (P<0.05), while the tryptophan-related metabolites andpurine-related metabolites were decreased (P<0.05), moreover, the leucine-related metabolites in the HRN group were obviously higher than those in the MRRN group and TRN gruop (P<0.05). CONCLUSIONS: MRRN, TRN and HRN can regulate the pathway of leucine metabolism (energy metabolism), purine metabolism (oxidative damage) and tryptophane metabolism (immune regulation) for RA, The specificity of HRN for RA focuses on regulation of leucine metabolism (energy metabolism).

Increased Frequency of Rheumatoid Arthritis and Allergic Rhinitis among Pesticide Sprayers and Associations with Pesticide Use.

Objective: The aim of this study was to identify diseases linked with the pesticide sprayer occupation and explore possible associations with exposure history data. Methods: Α cross sectional study was conducted among pesticide sprayers (n = 80) and the general population (n = 90) in Thessaly (Greece). Medical history, demographic characteristics and detailed exposure history were recorded by conducting personal interviews. Lifetime exposure indicators were calculated for several pesticide chemical subclasses. Moreover, organophosphate metabolite levels were quantified in urine samples of all participants by using gas chromatography -mass spectrometry (GC-MS). Multinomial analysis was used to determine associations between occupational pesticide exposure and diseases or disorders. Results: In the pesticide sprayers group, significantly higher frequencies for rheumatoid arthritis (RA) and allergic rhinitis were observed compared with the control group (p = 0.002 and p = 0.024 respectively). Within the pesticide sprayers group, high lifetime pesticide exposure was associated with increased risk for reporting RA (OR: 43.07 95% CI: 3.09-600.67) and allergic rhinitis (OR: 9.72 95% CI: 2.31-40.89), compared with low pesticide exposure. Exposure to organophsphate, guanidine and quinone pesticides were associated with RA while organophosphates, pyrethroids and paraquat were associated with allergic rhinitis. Despite the higher levels of certain pesticide metabolites observed among participants with rheumatoid arthritis, the differences were not statistically significant. One metabolite (diethylthiophosphate) was found to be significantly increased in allergic rhinitis cases (p = 0.037). Conclusions: The results from the current study suggest a possible association of occupational pesticide exposure with RA and allergic rhinitis that should be further investigated.

Cross-Reactivity of Chloroquine and Hydroxychloroquine With DRI Amphetamine Immunoassay.

BACKGROUND: Chloroquine and hydroxychloroquine are medical drugs used to treat the chemoprophylaxis of malaria and a second-line anti-inflammatory drug. METHODS: We performed a study of cross-reactivity of chloroquine and hydroxychloroquine in the DRI Amphetamine Assay inspired by a case report of a self-ingestion of chloroquine after a family dispute, that involved the following: (1) an in vitro study with control samples of healthy subjects, (2) an in vivo study with samples of patients with rheumatoid arthritis, and (3) an evaluation of the cross-reactivity of chloroquine and hydroxychloroquine in 3 additional immunoassays. RESULTS: In the case report, the Amphetamine DRI assay resulted positive both at 1000 ng/mL cutoff (1507 and 1137 ng/mL) and at 500 ng/mL cutoff (1178 and 642 ng/mL). Chloroquine urine levels were 103,900 and 100,900 ng/mL at 5 and 9 hours after ingestion. The results with control samples showed a positive cross-reactivity of chloroquine in the DRI Amphetamine Assay (approximately 0.74% and 0.89% at cutoff of 1000 and 500 ng/mL, respectively). Hydroxychloroquine did not cross-react with the DRI Amphetamine Assay up to 1,000,000 ng/mL. In patients treated with chloroquine or hydroxychloroquine, DRI Amphetamine did not produce false-positive results. The comparative assay study showed a positive cross-reactivity of chloroquine in the Emit II Plus Amphetamines Assay with control samples. CONCLUSIONS: Chloroquine can cause false-positive results in the DRI Amphetamine Assay when it is present at high concentrations. Hydroxychloroquine did not produce false-positive results neither in the DRI Amphetamine Assay nor in the others immunoassays evaluated.

A non-target urinary and serum metabolomics strategy reveals therapeutical mechanism of Radix Astragali on adjuvant-induced arthritis rats.

Radix Astragali has been used traditionally in China to treat rheumatoid arthritis (RA) in formulas. In this paper, we conducted a holistic evaluation of Radix Astragali acted on adjuvant-induced arthritis (AIA) rats by urinary and serum metabolomic studies. Histological results and hind paw swelling were used to assess the joint damage, while the levels of IL-1ß, TNF-α, SOD and MDA in serum were used to assess inflammation injury and oxidative stress. Metabolomic study and multivariate statistical analyses were used to investigate the differences between different groups. After processing with multivariate statistical analysis, 13 and 21 potential biomarkers were respectively found in urine and serum when Radix Astragali treatment group compared with model group. The main metabolism pathways in which Radix Astragali affected on AIA rats were tryptophan metabolism, phenylalanine metabolism, citrate cycle metabolism, fatty acid metabolism, vitamin B6 metabolism and so on. The present study demonstrates that urinary and serum metabolomics method could be a potentially powerful tool to understand the holistic therapeutic effect and the mechanisms of herb medicines.

Urinary haptoglobin, alpha-1 anti-chymotrypsin and retinol binding protein identified by proteomics as potential biomarkers for lupus nephritis.

The study was aimed at identification by proteomics and validation by enzyme-linked immunosorbent assay (ELISA) of potential urinary biomarkers for lupus nephritis. Study subjects comprised 88 systemic lupus erythematosus (SLE) patients and 60 controls (rheumatoid arthritis, diabetes mellitus and healthy individuals). Based on the SLE disease activity index (SLEDAI), patients were classified as active renal (AR), active non-renal (ANR) or inactive disease (ID). Urinary proteins from a group of patients with AR or ID were resolved by two-dimensional gel electrophoresis and identified by matrix-assisted laser desorption ionization-time of flight-mass spectrometry (MALDI-TOF-MS/MS). The selected biomarkers were validated by ELISA using samples from all patients and controls. AR patients were followed-up for 12 months after start of therapy. Three urinary proteins, alpha-1 anti-chymotrypsin (ACT), haptoglobin (HAP) and retinol binding protein (RBP), were detected in patients with AR and not ID. Upon validation, ACT levels were higher in AR patients than the other groups (P < 0·001) and showed good correlation with renal SLEDAI (r = 0·577, P < 0·001) as well as SLEDAI (r = 0·461, P < 0·001). Similarly, HAP levels were > 10-fold higher in AR than other groups (P < 0·001) and correlated well with renal SLEDAI (r = 0·594, P < 0·001) and SLEDAI (r = 0·371, P < 0·01). RBP levels were also higher in AR patients than in other groups (P < 0·05), except diabetes, and showed moderate correlation with renal SLEDAI (r = 0·284, P < 0·008) and SLEDAI (r = 0·316, P < 0·003). Upon follow-up with treatment, levels of all three proteins declined at 6 and 12 months (P < 0·01). Multiple logistic regression identified ACT as the best marker to differentiate AR from ANR. Urinary HAP, ACT and RBP are potential biomarkers for lupus nephritis activity.

Urinary proteomics can define distinct diagnostic inflammatory arthritis subgroups.

Current diagnostic tests applied to inflammatory arthritis lack the necessary specificity to appropriately categorise patients. There is a need for novel approaches to classify patients with these conditions. Herein we explored whether urinary proteomic biomarkers specific for different forms of arthritis (rheumatoid arthritis (RA), psoriatic arthritis (PsA), osteoarthritis (OA)) or chronic inflammatory conditions (inflammatory bowel disease (IBD)) can be identified. Fifty subjects per group with RA, PsA, OA or IBD and 50 healthy controls were included in the study. Two-thirds of these populations were randomly selected to serve as a training set, while the remaining one-third was reserved for validation. Sequential comparison of one group to the other four enabled identification of multiple urinary peptides significantly associated with discrete pathological conditions. Classifiers for the five groups were developed and subsequently tested blind in the validation test set. Upon unblinding, the classifiers demonstrated excellent performance, with an area under the curve between 0.90 and 0.97 per group. Identification of the peptide markers pointed to dysregulation of collagen synthesis and inflammation, but also novel inflammatory markers. We conclude that urinary peptide signatures can reliably differentiate between chronic arthropathies and inflammatory conditions with discrete pathogenesis.

Urinary interleukin-6 as a predictor of radiographic progression in rheumatoid arthritis: A 3-year evaluation.

Previously, we demonstrated that the urine proteome signature of patients with rheumatoid arthritis (RA) reflects inflammation-related cellular processes. Here, we measured interleukin (IL)-6, IL-8, and chemokine ligand 2 (CCL2) concentrations in the urine of RA patients and prospectively investigated their role in predicting RA activity and prognosis. One hundred seventy-three RA patients and 62 non-RA controls were recruited. Urinary IL-6, CCL2, and IL-8 levels were elevated in RA patients and correlated well with disease activity. Urinary IL-6 level at presentation was an independent risk factor of radiographic progression at 1 and 3 years. High urinary IL-6 level increased the risk ratio of radiographic progression by 2.9-fold, which was comparable to high serum CRP. Moreover, combination of urinary IL-6 and serum CRP measures synergistically increased the predictability of radiographic progression. In a subgroup with normal ESR, patients with the highest tertile of urinary IL-6 were at 6.4-fold greater risk of radiographic progression. Conclusively, high urinary IL-6 level at presentation is an independent risk factor for radiographic progression of RA, reflecting disease activity. Urinary IL-6 in combination with serum CRP may be a useful parameter for estimating RA prognosis.

Urine metabolome profiling of immune-mediated inflammatory diseases.

BACKGROUND: Immune-mediated inflammatory diseases (IMIDs) are a group of complex and prevalent diseases where disease diagnostic and activity monitoring is highly challenging. The determination of the metabolite profiles of biological samples is becoming a powerful approach to identify new biomarkers of clinical utility. In order to identify new metabolite biomarkers of diagnosis and disease activity, we have performed the first large-scale profiling of the urine metabolome of the six most prevalent IMIDs: rheumatoid arthritis, psoriatic arthritis, psoriasis, systemic lupus erythematosus, Crohn's disease, and ulcerative colitis. METHODS: Using nuclear magnetic resonance, we analyzed the urine metabolome in a discovery cohort of 1210 patients and 100 controls. Within each IMID, two patient subgroups were recruited representing extreme disease activity (very high vs. very low). Metabolite association analysis with disease diagnosis and disease activity was performed using multivariate linear regression in order to control for the effects of clinical, epidemiological, or technical variability. After multiple test correction, the most significant metabolite biomarkers were validated in an independent cohort of 1200 patients and 200 controls. RESULTS: In the discovery cohort, we identified 28 significant associations between urine metabolite levels and disease diagnosis and three significant metabolite associations with disease activity (P FDR < 0.05). Using the validation cohort, we validated 26 of the diagnostic associations and all three metabolite associations with disease activity (P FDR < 0.05). Combining all diagnostic biomarkers using multivariate classifiers we obtained a good disease prediction accuracy in all IMIDs and particularly high in inflammatory bowel diseases. Several of the associated metabolites were found to be commonly altered in multiple IMIDs, some of which can be considered as hub biomarkers. The analysis of the metabolic reactions connecting the IMID-associated metabolites showed an over-representation of citric acid cycle, phenylalanine, and glycine-serine metabolism pathways. CONCLUSIONS: This study shows that urine is a source of biomarkers of clinical utility in IMIDs. We have found that IMIDs show similar metabolic changes, particularly between clinically similar diseases and we have found, for the first time, the presence of hub metabolites. These findings represent an important step in the development of more efficient and less invasive diagnostic and disease monitoring methods in IMIDs.