Management of patient with Lupus Nephritis (LN) continues to remain a challenge for the treating physicians because of considerable morbidity and even mortality. The search of biomarkers in serum and urine is a focus of researchers to unravel new targets for therapy. In the present study, the utility of NMR-based serum metabolomics has been evaluated for the first time in discriminating LN patients from non-nephritis lupus patients (SLE) and further to get new insights into the underlying disease processes for better clinical management. Metabolic profiling of sera obtained from 22 SLE patients, 40 LN patients and 30 healthy controls (HC) were performed using high resolution 1D 1H-CPMG and diffusion edited NMR spectra to identify the potential molecular biomarkers. Using multivariate analysis, we could distinguish SLE and LN patients from HC and LN from SLE patients. Compared to SLE patients, the LN patients had increased serum levels of lipid metabolites (including LDL/VLDL lipoproteins), creatinine and decreased levels of acetate. Our results revealed that metabolic markers especially lipids and acetate derived from NMR spectroscopy has high sensitivity and specificity to distinguish LN among SLE patients and has the potential to be a useful adjunctive tool in diagnosis and clinical management of LN.

Introduction. Th17, γδT, NK, and NKT cells in peripheral blood and serum IL-17 and IL-23 in Takayasu arteritis (TA) were measured and correlated with disease activity. Methods. Th17 (anti-CD3APC, CD4PECy7, and IL-17PE), NKT, NK (anti-CD3APC, CD56FITC), and γδT (anti-CD3FITC and γδTCRAPC) cells were enumerated by flow cytometry in peripheral blood of 30 patients with TA (ACR1990 criteria) and 20 healthy controls, serum IL-17 and IL-23 measured by ELISA. Relation with disease activity (NIH criteria, ITAS2010) was analyzed (using nonparametric tests, median with interquartile range). Results. Mean age of patients was 33.47 ± 11.78 years (25 females); mean symptom duration was 7.1 ± 5.3 years. 13 were not on immunosuppressants; 12 were active (ITAS2010 ≥ 4). The percentage of Th17 cells was significantly expanded in TA (patients 2.1 (1.5–3.2) versus controls 0.75 (0.32–1.2); p < 0.0001) with no differences in other cell populations. Serum IL-17 and IL-23 (pg/mL) in patients (6.2 (4.6–8.5) and 15 (14.9–26.5), resp.) were significantly higher (p < 0.001) than controls (3.9 (3.9–7.3) and undetectable median value, resp.). Subgroup analysis revealed no correlation of Th17 cells, serum IL-17, and IL-23 with disease activity or medications, nor any significant difference before and after medication. Conclusions. There is significant expansion of Th17 cells and elevated serum IL-17 and IL-23 levels in TA patients compared to healthy controls.

B cell activating factor (BAFF) and a proliferation-inducing ligand (APRIL) help in B cell activation, maintenance and plasma cell survival. B cell infiltration has been demonstrated in kidneys of patients with lupus nephritis (LN). Serum levels of BAFF and APRIL have shown inconsistent relationships with lupus disease activity. We evaluated urinary levels of BAFF and APRIL as biomarker for LN. Thirty-six patients with proliferative lupus nephritis (AN), 10 with active lupus without nephritis (AL) and 15 healthy controls (HC) were studied. APRIL and BAFF levels were measured in both serum and urine using enzyme-linked immunosorbent assay (ELISA). Urine levels were normalized for urinary creatinine excretion. Urine levels were correlated with conventional disease activity markers and histology. Levels were reassessed in 20 AN patients at 6 months after treatment with cyclophosphamide. Urinary APRIL (uAPRIL) and BAFF (uBAFF) levels were raised significantly in AN. uAPRIL, but not uBAFF, correlated moderately with renal Systemic Lupus Erythematosus Disease Activity Index (SLEDAI) in AN (r = 0·36, P < 0·05). On receiver operator curve (ROC) analysis, uBAFF and uAPRIL showed an area under the curve (AUC) of 0·825 and 0·781, respectively, in differentiating between nephritis and non-nephritis, which performed better than low C3, C4 and raised anti-dsDNA antibodies. There was no correlation of serum levels with uBAFF (r = 0·187, P = 0·261) and uAPRIL (r = 0·114, P = 0·494). uAPRIL levels reduced after treatment (mean 125 pg/mg to 36 pg/mg, P < 0·05). uBAFF levels reduced in 16 responders while two of four non-responders had increase in levels. Thus, uBAFF and uAPRIL are potential biomarkers of proliferative lupus nephritis.

Reactive arthritis (ReA) is a member of seronegative spondyloarthropathy (SSA), which involves an acute/subacute onset of asymmetrical lower limb joint inflammation weeks after a genitourinary/gastrointestinal infection. The diagnosis is clinical because it is difficult to culture the microbes from synovial fluid. Arthritis patients with a similar clinical picture but lapsed history of an immediate preceding infection that do not fulfill the diagnostic criteria of other members of SSA, such as ankylosing spondylitis, psoriatic arthritis, and arthritis associated with inflammatory bowel disease, are labeled as peripheral undifferentiated spondyloarthropathy (uSpA). Both ReA and uSpA patients show a strong association with class I major histocompatibility complex allele, HLA-B27, and a clear association with an infectious trigger; however, the disease mechanism is far from clear. Because the clinical picture is largely dominated by rheumatoid-arthritis (RA)-like features including elevated levels of inflammatory markers (such as ESR, CRP, etc.), these overlapping symptoms often confound the clinical diagnosis and represent a clinical dilemma, making treatment choice more generalized. Therefore, there is a compelling need to identify biomarkers that can support the diagnosis of ReA/uSpA. In the present study, we performed NMR-based serum metabolomics analysis and demonstrated that ReA/uSpA patients are clearly distinguishable from controls and further that these patients can also be distinguished from the RA patients based on the metabolic profiles, with high sensitivity and specificity. The discriminatory metabolites were further subjected to area under receiver operating characteristic curve analysis, which led to the identification of four metabolic entities (i.e., valine, leucine, arginine/lysine, and phenylalanine) that could differentiate ReA/uSpA from RA.

Takayasu arteritis (TA) is a large vessel vasculitis of unknown pathogenesis. Assessment of disease activity is a challenge, and there is an unmet need for relevant biomarker(s). In our previous study, NMR based serum metabolomics had revealed distinctive metabolic signatures in TA patients compared with age/sex matched healthy controls and systemic lupus erythematosus (SLE). In this study we investigate whether the metabolites correlate with disease activity. Patients with TA fulfilling American College of Rheumatology (ACR) criteria were enrolled, and disease activity was assessed using Indian Takayasu Clinical Activity Score using acute phase reactant-erythrocyte sedimentation rate [ITAS-A (ESR)]. Sera were analyzed using 800 MHz NMR spectrometer to identify metabolites [based on partial least squares discriminant analysis (PLS-DA) VIP (variable importance in projection) score > 1.0 and permutation test, p-value <0.01]. 45 active and 53 inactive TA patients with median age 27 [(IQR) 22-35 years] and 27 [(IQR) 23-37 years], female to male ratio 3.5:1 and 4.9:1, and median duration of illness 5 [(IQR) 2-9 years] and 3 [(IQR) 1-6 years], respectively, were enrolled. The key metabolites with highest discriminatory potential in active TA (ITAS-A ≥ 4) were glutamate and N-acetyl glycoprotein (NAG), both elevated, with area under the curve 0.775 and 0.769 ( p-value <0.001). On follow up assessment, metabolic spectra started to differ with change in disease activity. This large cohort of patients revealed metabolic profiles discriminating between clinically active and inactive TA patients. It suggests glutamate and NAG have strong potential as biomarkers for disease activity in TA and may serve as a guide to therapy. We are now working to further validate these results in longitudinal studies.