Urinary HMGB1 levels are associated with CD4+ T-cells in urine in patients with ANCA-associated vasculitis and active nephritis

Abstract

Background High mobility group box 1 (HMGB1) is a nuclear non-histone protein that acts as an alarmin when released in the extra-cellular medium by necrotic or activated cells. High serum HMGB1 levels have been found in patients with antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV) and active nephritis as well as in patients with granulomatosis with polyangiitis (GPA) and predominant granulomatous manifestations. High serum and urinary HMGB1 levels have also been found in patients with systemic lupus erythematosus (SLE) and active nephritis. Objectives To evaluate serum and urinary levels of HMGB1, CD4+ T-cells and effector memory T-cells (CD4+ T EM )in peripheral blood (PB) and urine from patients with AAV and active nephritis. Methods 24 AAV patients with active nephritis and 10 healthy controls (HC) were evaluated in a cross-sectional study. Clinical diagnosis of GPA or microscopic polyangiitis (MPA) was made according to the European Medicines Agency algorithm whereas patients with isolated renal involvement, ANCA positivity and/or biopsy-proven pauci immune necrotizing glomerulonephritis were classified as renal limited vasculitis (RLV). Serum and urinary levels of HMGB1 were measured by Western blot while CD4+ T-cells and CD4+ T EM cells (CD4+CD45+RO+CCR7-) in urine were analyzed by flow cytometry. Disease activity was evaluated according to the Birmingham Vasculitis Activity Score (BVAS). Results Median urinary intensity of HMGB1 was significantly higher in AAV patients when compared to HC [5.15 (IQR 3.52-9.25) vs. 2.58 (1.88-4.24); P =0.006] whereas no difference was found among different AAV subsets regarding median urinary intensity of HMGB1 [GPA: 4.1 (2.6-16.4) vs. MPA: 5.1 (4.4-5.6) vs. RLV 5.7 (1.4-13.6); P =0.951] and HMGB1/creatinine ratio [GPA: 0.93 (0.29-3.91) vs. MPA: 1.29 (0.90-1.89) vs. RLV 0.54 (0.14-1.28); P =0.186]. In urine of AAV patients, median CD4+ T-cells was 125.00 (58.93-429.28) cells/ml and CD4+ T EM cells was 93.24 (43.21-343.37) cells/ml. Urine levels of HMGB1 were positively correlated with CD4+ T-cell counts in urine (rho: 0.454; P =0.014) and with CD4+ T EM cells in urine (rho: 0.378; P =0.049) while a tendency for a negative correlation between urinary HMGB1 and CD4+ T-cells in PB (rho: -0.307; P =0.083). No correlation could be found between urine levels of HMGB1 and 24 hour proteinuria (rho: 0.065; P =0.390), creatinine clearance (rho: -0.049; P =0.832), BVAS (rho: -0.203; P =0.171) and serum HMGB1 (rho: -0.046; P =0.419). Conclusions Urine HMGB1 levels are increased in AAV patients with active nephritis when compared to HC and are associated with CD4+ T-cells in urine suggesting that urine HMGB1 is a biomarker for renal activity in AAV.