#4635 CIRCULATING NEPHRIN AUTOANTIBODIES IN PATIENTS WITH POST-TRANSPLANT RECURRENCE OF PRIMARY FOCAL SEGMENTAL GLOMERULOSCLEROSIS
Authors: Hattori Motoshi , Shirai Yoko , Kanda Shoichiro , Ando Taro , Ishizuka Kiyonobu , Hashimoto Taeko , Miura Kenichiro 
Published: June 2023

Abstract

Background and Aims
Post-transplant recurrence of primary focal segmental glomerulosclerosis (FSGS) is a major challenge in the field of nephrology and kidney transplantation. Primary FSGS is thought to be caused by circulating factors (CFs) that injure podocytes, but these factors remain unknown despite intensive efforts to identify them. Recently, one post-transplant recurrent FSGS patient with pretransplant nephrin autoantibodies was reported (Watts AJB et al, JASN 33:238, 2022). Furthermore, we reported very early post-transplant recurrence changes of podocytes and possible roles of circulating nephrin autoantibodies in a patient with FSGS recurrence (Hattori M et al, AJT 22:2478, 2022). To confirm these preliminary results, we examined the autoantibodies to nephrin and the punctate deposition of IgG colocalized with nephrin using stocked plasma samples and grafted kidney biopsy specimens of the patients with FSGS recurrence.

Method
This study was approved by the local Ethics Committee of Tokyo Women's Medical University (approval no. 2021-0184). The clinical and research activities reported here are consistent with the Principles of the Declaration of Istanbul as outlined in the “Declaration of Istanbul on Organ Trafficking and Transplant Tourism”. From 1989 to 2022, a total of 78 patients with FSGS underwent kidney transplantation in our institution, and 16 patients experienced post-transplant recurrence of FSGS. Since the stoked plasma samples were available in 14 out of 16 patients, these 14 patients were included in this study. All 14 patients showed immediate post-transplant recurrence and had no pathogenic variants in FSGS-related genes including NPHS1. Circulating anti-nephrin autoantibodies were measured using the ELISA methods based on the Watts's report with one modification, that is, recombinant extracellular domain of human nephrin was obtained commercially (R&D systems, USA). To validate the ELISA system, circulating anti-nephrin autoantibodies were also measured in 9 genetic FSGS patients, 13 membranous nephropathy (MN) patients, 4 lupus nephritis (LN) patients, and 13 healthy controls. Dual immunofluorescence (IF) staining of nephrin and IgG and image acquisition using the two-dimensional structured illumination microscopy mode with a Nikon microscope were performed as previously reported (Hattori M et al, AJT 22:2478, 2022).

Results
The cut-off level of anti-nephrin antibody calculated from the results of a total 30 cases including 13 MN patients, 4 LN patients, and 13 healthy controls using Frey's formula (J Immunol Method, 1998) was 172.2 U/mL All 14 recurrent patients enrolled in this study had higher titter than the cut-off level and the mean ± SD of the anti-nephrin antibody level in these patients was 939.9 ± 261.8 U/mL. In contrast, the mean anti-nephrin antibody level in patients with genetic FSGS was 98.6 ± 77.3 U/mL, which was comparable with healthy controls. The IF staining of graft biopsy specimens showed punctate depositions of IgG colocalized with nephrin in all 14 recurrent patients.

Conclusion
The autoantibodies to nephrin in plasma samples and the punctate deposition of IgG colocalized with nephrin in grafted kidney biopsy specimens were observed in all 14 patients with post-transplant recurrent FSGS. Further studies in larger numbers of patients as well as in patients of other ethnicities are clearly required to confirm this surprising result; however, circulating nephrin autoantibodies could be a possible candidate for CFs in the pathogenesis of recurrent FSGS.