Our study is a retrospective cohort study which included subjects with biopsy-proven renal non-AA amyloidosis. All native kidney biopsies obtained between January 2005 and December 2019 were retrospectively reviewed. We identified 92 subjects with renal non-AA amyloidosis. Of 92 subjects, 18 patients were excluded: 14 were referred from other hospitals and 4 had inadequate data. Demographic and laboratory findings at the time of renal biopsy, treatment regimens and outcome data were collected from electronic medical reports.
All kidney biopsy specimens were obtained by ultrasound guided needle-biopsy, embedded in paraffin and cut into 4–6 µm sections. The sections were stained with hematoxylin and eosin, periodic acid–Schiff, Masson’s trichrome, Jones methamine silver and Congo red and evaluated by light microscopy (LM). For immunohistochemical (IHC) evaluation, paraffin-embedded tissue sections were stained with antibodies to amyloid A, fibrinogen alpha, transthyretin, kappa, lambda and lysosome by a fully automated device (Benchmark XT, Ventana Medical Systems, Tucson, USA). Cryosections were stained with polyclonal FITC-conjugated antibodies to kappa, lambda, IgG, IgA, IgM, C1q, C3c and fibrinogen (1/20 dilution, DAKO, Glostrup, Denmark) for immunoflourescence microscopy (IF).
Congo red stain was performed for detection of amyloid deposits and assessed by LM. Amyloid was identified as eosinophilic deposits which demonstrated apple-green birefringence under polarized light. AL amyloidosis was determined by IF, IHC and laboratory findings. Unclassified (UC) amyloidosis was defined when IHC and IF revealed equivocal or inconclusive results together with laboratory finding (Figure). All biopsies were evaluated according to the renal amyloidosis scoring and grading system previously described by Sen and Sarsik (6). Pattern and quantity of amyloid deposition in glomeruli, interstitium, vessels and tubulointerstitial changes were scored (Supp Data). By addition of all scores of glomerular, vascular and tubulointerstitial involvement, a renal amyloid prognostic score (RAPS) was obtained.
All patients with AL amyloidosis underwent bone marrow aspiration and biopsy with Jamshidi needles from iliac crest. After fixation with Hollant, bone marrow biopsy specimen were stained with haematoxylin and eosin, congo red and periodic acid–Schiff reagent. Immunohistochemical analysis done with Ventana automatic machine for kappa, lambda and immunoglobulins. Plasma cell monoclonality was defined as a κ/λ ratio of more than 3 (κ clone) or less than 1 (λ clone). Plasma cell percentages were assessed on immunoperoxidase-stained sections for κ and λ immunoglobulins and Wright-Giemsa–stained aspirate smears. Amyloid deposits were stained Congo red and detected by apple-green birefringence under polarized light. Multiple myeloma was diagnosed according to International Myeloma Working Group criteria (13).
Estimated glomerular filtration rate (eGFR) was calculated by using CKD-EPI Eq. (14). End stage kidney disease (ESKD) was defined as requirement of renal replacement therapy (RRT).
Primary outcomes were kidney and patient survival. Follow-up period was defined as duration from kidney biopsy to last visit or death. Renal survival was calculated from date of biopsy to the initiation of RRT. Patients who died without requiring RRT were censored for analysis of renal survival (death-censored renal survival).
Categorical variables were presented as percentage and compared with the chi-square test. Normality of distribution were assessed using the Kolmogorov-Smirnov test. Continuous variables were presented as mean ± standard deviation or median and interquartile ranges. Continuous variables were compared with one-way ANOVA or Mann-Whitney U test according to distribution of normality. Ordinary regression analysis was performed for estimating of relationship between age, gender, baseline eGFR, proteinuria and RAPS grade. Kaplan-Meier analysis was used for evaluation of death-censored renal survival and patient survival. A p value < 0.05 was statistically significant. All statistics were performed with Statistical Package of Social Science (SPSS) software version 14.0 for Windows.