Post-transplant membranous glomerulonephritis as a manifestation of chronic antibody-mediated rejection Hyeon Joo Jeong, Beom Jin Lim, Myoung Soo Kima, Yu Seun Kima, Soon Il Kima Departments of Pathology and aSurgery, Yonsei University College of Medicine, Seoul, Korea Introduction Membranous glomerulonephritis (MGN) may recur or develop de novo after renal transplantation, of which de novo form is more common than recurrent form. Although immune mechanisms are suspected, the pathogenesis of post-transplant MGN is uncertain yet. Recently, it has been known that peritubular capillary (PTC) C4d expression is associated with de novo post-transplant MGN. In this study, we tried to further explore the relationship between post-transplant MGN and antibody-mediated immunity by C4d immunohisto-chemistry and electron microscopy. Table 1. Histologic features of post-transplant MGN. Post-transplant MGN Post-transplant IgAN p-value Global sclerosis (%) 20.6±23.4 24.8±21.2 0.352 Segmental sclerosis (%) 5.1±16.2 7.2±10.9 0.078 Interstitial inflammation 0.021 Absent or mild 16 31 Moderate or severe 1 Tubulitis 0.389 17 44 3 Interstitial fibrosis 0.473 13 38 4 9 Tubular atrophy 0.616 36 11 Arterial hyalinosis 0.140 10 7 Arterial fibrointimal thickening 0.536 45 2 Mesangial proliferation 0.605 Absent or focal 15 42 diffuse 5 C4d deposit in PTCs 0.023 Negative 22 Positive(diffuse and focal) 6 Materials and Methods 1. Subjects Of 2,619 patients received kidney transplantation at Severance Hospital between 1988 and 2008, post-transplant MGN was diagnosed in 20 cases. A total of 18 cases (16 males and 2 females) with sufficient paraffin-embedded tissue were selected. Two of them were recurrent forms, 7 de novo forms and the other 9 unknown. Forty-seven cases of post-transplant IgA nephropathy (IgAN) were retrieved as a control group. 2. Tissue processing and microscopic examination Biopsy samples were examined by light microscopy, immuno-fluorescence and electron microscopy according to a routine protocol. Immunohistochemistry for C4d was performed using paraffin-embedded tissues. 3. Assessment of renal allograft morphology The degree of histologic injuries was scored according to the Banff 97 criteria. C4d staining was interpreted as diffuse positive when more than 50% of cortical PTCs were linearly stained and focal positive when the staining was less than 50%. Positive staining of a few capillaries was interpreted negative. Multilayering of PTC basement membrane was evaluated by electron microscopy. At least five fields were taken at 1:20,000 magnification. The degree of layering was categorized as low- (3-4 layers) and high-grade (≥5 layers). 4. Statistical analysis A comparison of histologic injury between post-transplant MGN and IgAN was performed using Mann-Whitney U-test and Pearson’s Chi-square method. All parameters are scored according to Banff 97 criteria. Figure 1.Histologic features of post-transplant MGN in glomerulus (A) and tubulointerstitium (B). Granular IgG deposits along the capillary loops were observed in all cases (C). C4d deposits in PTCs were demonstrated by immunohistochemistry (D). The electron dense deposits were observed in subepithelial spaces in all cases (E) and multilayering of PTC basement membrane was observed in some cases (F) C E F A D B Results 1. Patient demography Mean age of recipients at transplantation was 38.2 yrs. Fifty percent of them received grafts from the opposite gender and 33% from the living related donors. Number of HLA-DR mismatch was 1 in 41.7% and 2 in 8.3% of unrelated donors. The biopsy indication was proteinuria (>1g/24hr) and increased serum creatinine levels. No protocol biopsy was performed. Median time to diagnosis was 4.8 yrs after transplantation. 3. PTC basement membrane multilayering Multilayering of PTC basement membrane was observed in 10 cases. The grade of multilayering was high in 3 cases and low in 7 cases. 2. Renal allograft histology and C4d deposit in PTCs (Figure 1) Stages of MGN were I in 5 cases, II in 9 cases, III in 3 cases and IV in 1 case. Acute rejection was present in 3 cases and chronic rejection in 4 cases. On immunofluorescence, granular IgG deposits were observed in all samples. C4d positivity in PTCs was observed in 9 cases (diffuse in 8 cases, and focal in 1 case). Two were associated with acute rejection and 3 were associated with chronic rejection. The C4d positivity increased with stages of MGN. Histologic features were compared to those of post-transplant IgAN (Table 1). Conclusion The histologic features of post-transplant MGN were not different from those of post-transplant IgAN except the degree of interstitial inflammation. A higher frequency of PTC C4d positivity and basement membrane multilayering suggests an involvement of chronic antibody-mediated injury in the evolution of post-transplant MGN.