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The exostosin 1 and 2 (EXT1/2) complex was recently identified as a potential antigen associated with systemic autoimmunity in PLA2R-negative membranous nephropathy patients. There are currently no reports of circulating anti-EXT1/2 antibodies and no clinical studies of EXT1/2-positive membranous nephropathy in the Chinese population.
Immunohistochemical staining for PLA2R, THSD7A and NELL-1 was performed in 1442 consecutive patients with biopsy-proven MN from January 2011 to December 2019 at Beijing Anzhen Hospital. A total of 201 MN patients were PLA2R-, THSD7A- and NELL-1-negative and enrolled in the study. Patient medical records were reviewed for clinical data, including demographics, laboratory tests, treatment, and follow-up. The glomerular expression of EXT1 and EXT2 was screened. The anti-EXT1 and anti-EXT2 antibodies were also detected in the sera of patients with EXT1/2–positive membranous nephropathy by western blot, ELISA and immunoprecipitation separately. Clinical and pathologic features were comparable between PLA2R-, THSD7A- and NELL-1-negative patients with EXT1/2 or without EXT1/2 positive.
Among these 201 patients, 19 of 83 (22.9%) with lupus membranous nephropathy (class 5 and class 5+3/4), and 13 of 118 (11.0%) with nonlupus membranous nephropathy exhibited positive EXT1/2 staining. Thirteen patients with EXT1/2-positive non-LMN had no definitive secondary cause, but 7 patients (53.8%, 7/13) were seropositive for ANA. Baseline anti–double-stranded DNA antibody and serum antinuclear antibody positivity were independent predictors of glomerular EXT1/2-positive expression in patients with lupus membranous nephropathy (pure class 5, class 5 + 3/4) and nonlupus membranous nephropathy, respectively. In 32 patients with EXT1/2-positive MN, 22 patients with available serum samples collected at the time of biopsy were tested for anti-EXT1 and anti-EXT2 antibodies. Sixteen patients were positive for antibodies. According to the molecular weight of positive bands, we speculated that 5 patients were positive for both anti-EXT1 and anti-EXT2 antibodies, 11 patients were positive only for anti-EXT1 antibodies, and no patients were positive only for anti-EXT2 antibodies. There were no significantly differences between the characteristics of 22 EXT1/2-positive MN patients with or without antibody seropositivity. Two patients with EXT1/2-positive membranous nephropathy underwent repeated renal biopsies. When the pathological type changed to lupus nephritis (LN) or the LN worsened clinically, the renal tissue staining intensity of EXT1/2 increased. Renal EXT1/2 staining positivity occurred earlier than the diagnosis of lupus.
This is the first study to detect circulating anti-EXT1/2 antibodies, and the functional significance should be further studied. The renal EXT1/2 staining positive was earlier than diagnosis of lupus. whether the level of serum EXT1/2 antibody can be used as a predictor for disease progression and prognosis needs to be further evaluated in larger sample studies.