A 56-year-old Japanese woman was referred because of severe lower-limb edema and general fatigue. She had been healthy prior to admission with no history of renal disease. One year before admission, laboratory data showed no abnormalities, with a serum creatinine level of 0.68 mg/dL, serum total protein level 7.0 g/mg, low-density lipoprotein (LDL)-cholesterol level 110 mg/dL, and no urinary protein excretion. An examination showed a body height of 152 cm, weight 60 kg (gained about 6 kg in the month after onset), blood pressure (BP) 132/84 mmHg, and heart rate 104 beats/min. Chest and abdominal X-ray showed no abnormalities. The laboratory data revealed severe proteinuria (6.8 g/day), hypoproteinemia (serum total protein 4.2 g/dL, albumin 1.8 g/dL), normal renal function [serum creatinine level 0.66 mg/dL, estimated glomerular filtration ratio (eGFR) 72.2 mL/min/1.73 m2], and dyslipidemia [LDL-cholesterol level 300 mg/dL, triglyceride (TG) level 220 mg/dL, high-density lipoprotein (HDL)-cholesterol level 56 mg/dL]. Serology was negative for anti-nuclear, anti-glomerular basement membrane, myeloperoxidase anti-neutrophil cytoplasmic (MPO-ANCA), proteinase 3-ANCA, hepatitis C antibodies, and hepatitis B antigen. Serum complement factors and IgG, IgM, IgA levels were within normal limits. Thus, she had no findings suggestive of infection such as hepatitis B and C viruses, malignancy, collagen diseases including systemic lupus erythematosus and rheumatic arthritis, or drug-induced secondary MN. She was therefore diagnosed with IMN.
Despite being treated with conventional diuretics, including a loop diuretic and thiazide (on admission, furosemide 40 mg/day and trichlormethiazide 2 mg/day were administered, and the furosemide dose was increased to 80 mg/day on the 3rd day), her lower-limb edema and urinary volume remained unchanged. On the 5th day, renal biopsy was performed. Histopathology showed typical stage II MN including diffuse thickening of the basement membrane and a large number of electron-condensing substances on the subepithelium. An immunofluorescence examination showed granular IgG and C3 deposition on the capillary loops. IgG subclasses were also stained. IgG1, and IgG3 were negative in the glomeruli, but IgG2 and IgG4 were strongly positive (Fig. 1). AQP-2 immunohistochemistry was performed via the following method (polymer technique): The kidney tissues were fixed in 20% neutral buffered formalin and embedded in paraffin, and sections 2-3 mum in thickness were prepared. The details of the method have been reported previously. Briefly, the simple statin MAX-PO (MULTI) (NICHIREI BIOSCIENCES INC., Tokyo, Japan) Polymer reagent instillation was performed as follows: incubation at room temperature for 30 minutes, and then washing with TBS for 5 minutes, 3 times. The samples then underwent color development with adjusted DAB substrate solution. The reagent on the slide was washed for a few minutes with purified water and then immersed in purified water. The specimens were finally stained with Hematoxylin and Eosin staining as a contrast agent and then washed for 2 minutes, the color enhanced, and the specimens dehydrated, penetrated, and mounted. The specimens were observed using light microscopy. The primary antibody was anti-AQP-2 antibody (NB110-74682, 1:200, Novus Biologicals, Littleton, Colorado, USA). Case 2 was also stained under the same conditions. AQP-2 immunostaining in the kidney was strongly positive in the collecting duct cells (Fig. 2A). The urinary AQP-2 levels were 3.20 ng/mgCre and urine osmolality 510 mOsm/kgH2O. After the renal biopsy, tolvaptan 15 mg/day was initiated, and the furosemide dose was reduced to 40 mg/day and trichlormethiazide discontinued due to a marked increase of urine volume. Three days later, the urine volume increased markedly to 3,800 mL/day (Fig. 3, blue line), and tolvaptan was reduced to 7.5 mg/day and then discontinued on the 14th day along with furosemide. The urinary AQP-2 level and urine osmolality also decreased to 0.88 ng/mgCre and 188 mOsm/kgH2O, respectively, on the 14th day. The patient's urine volume was maintained at approximately 2,000 mL/day by the 56th day. The urinary protein excretion decreased in a serial manner without the administration of an immunosuppressant such as prednisolone (PSL) and had reduced to approximately 0.5 g/day by the 56th day (Fig. 3, blue bar).
The changes in the serum sodium level are shown in Fig. 4. Serum BNP levels were reduced from 250 to 44 pg/mL after tolvaptan treatment on the 14th day. The cardiac function measured by echocardiography remained unchanged within normal limits throughout the treatment period [baseline: left ventricular end-diastolic dimension (LVDd) 48 mm, left ventricular ejection fraction (LVEF) 66%, E/e' 6.6; 14th day: LVDd 47 mm, LVEF 68%, E/e' 6.5].