A case of membranous nephropathy during asymptomatic syphilis

Abstract

We experienced membrane nephropathy associated with syphilis infection. A 51-year-old woman had an increase in body weight with leg edema. Blood and urine examination showed low serum protein and heavy proteinuria. She was diagnosed with nephrotic syndrome. The light microscope showed normal glomeruli with no bubbling appearance or spike lesions. Immunofluorescence analysis on the renal biopsy sample showed granular deposition of IgG, C3, C1q, κ and λ on capillary walls. The electron microscope showed a hump-like subepithelial and mesangial electron-dense deposit. Her pathologic diagnosis was Ehrenreich-Churg stage I membrane nephropathy. Negative staining of IgG4 on glomerular walls indicates secondary membrane nephropathy. The immunology test indicated active syphilis infection despite the lack of any skin lesions or constitutional symptoms related to syphilis. NDNF, which was recently identified as an antigen of syphilis-associated MN, was positive on capillary walls. The antibiotic treatment led to rapid, complete remission without using immunosuppressive treatment.

Introduction

Syphilis is an infectious disease caused by Treponema pallidum. The primary infection presents as a localized skin ulcer lesion. Many patients do not notice since an ulcer caused by syphilis is painless. The patients who do not get initial treatment properly can show various clinical symptoms, including fever, adenopathy, and rash. Nephrotic syndrome is a rare complication after syphilis infection [1]. The renal pathology can present as minimal change disease, membranoproliferative glomerulonephritis (MPGN), or acute interstitial nephritis; however, it mostly presents as membranous nephropathy (MN). The number of syphilis-associated MN cases would be increasing due to the increase in the number of patients with syphilis in worldwide [2].

Membranous nephropathy (MN) is a glomerular disease that can cause nephrotic syndrome. MN is classified into idiopathic or secondary MN. Secondary MN is associated with various diseases, including infection, autoimmune diseases, tumors, and drug use [3]. Secondary MN shares many pathological features with idiopathic MN; however, there are several distinct patterns. IgG subclass staining is useful to differentiate idiopathic from secondary MN. IgG4 is dominantly deposited in idiopathic MN, while IgG1 and IgG3 are dominant in secondary MN [4]. This discrimination from idiopathic MN is important, as treatment for secondary MN is the elimination of the antigens that cause nephrotic syndrome. Otherwise, immunosuppressive therapy, including steroids, is often required for remission.

The exploration for the antigen for membrane nephropathy has progressed. PLA2R (M-type phospholipase A2 receptor) was identified as a major antigen in idiopathic MN [5]. PLA2R is reported to account for more than half of idiopathic MN [5]. The immunofluorescent staining of PLA2R using renal biopsy samples is now routinely performed in many laboratories [6]. Recent mass spectrometry analysis has successfully identified the remaining antigens of membrane nephropathy [1], including THSD7A, NELL1, EXT1/EXT2, NCAM1, PCDH7, SEMA3B, CNTN1, FAT1, and PCSK6 [6]. Neuron-derived neurotrophic factor (NDNF) was also newly discovered by using mass spectrometry as a novel antigenic target in syphilis-associated MN [7]. Of note, in the study, NDNF was detected in all cases of syphilis infection, suggesting NDNF was a specific antigen. All cases were stage I MN, and the dominant IgG subclass was IgG1 [7]. The renal deposition of NDNF has been shown on glomerular walls, but the pathological meaning remains unclear. Serologic diagnosis without renal biopsy has been suggested for several target antigens, including NDNF, but the method is not established at the level of commercial use. A summary of case series of NDNF-positive syphilis-associated MN showed prompt remission only using antibiotic treatment. Most cases present a skin rash before nephrotic syndrome.

Here, we report a case of secondary MN followed by asymptomatic syphilis infection. Serological tests for syphilis infection should be considered when encountering secondary MN, even if patients do not have any rash or symptoms related to syphilis infection.

Case report

A 51-year-old woman was aware of rapidly progressive leg edema with body weight increase and presented to our hospital. Laboratory tests showed low levels of serum protein and albumin with heavy proteinuria, consistent with nephrotic syndrome. She was admitted to our hospital for examination. She has an allergy to some fruits. She occasionally drinks alcohol, but never smokes. She has a history of Salpingectomy 3 months before admission. She has not taken regular medication or any supplements. Family history was unremarkable. On admission, her height was 157 cm, and her body weight was 55 kg, which was a 4 kg increase over 1 week. She had a decrease in her appetite. Body temperature was 36.7 °C. Blood pressure was 136/94 mmHg, and heart rate was 79 bpm. The abdomen was flat and soft with no tenderness. Leg edema was prominent. No rash and lymphadenopathy were noted. Chest radiography and Abdominal CT showed bilateral pleural effusion. Kidney size was normal (Fig. 1).

Fig. 1.

Chest radiography and abdominal CT images

Serum biochemical analyses showed a decrease in total protein (TP; 5.5 g/dL) and albumin (Alb; 1.5 g/dL). The lipid profile showed an increase in total cholesterol (T-Chol; 251 mg/dL) and low-density lipoprotein (LDL-C; 182 mg/dL). Kidney function was not impaired. Blood urea nitrogen (BUN) was 10.0 mg/dL, and serum creatinine (Cr) was 0.56 mg/dL. The levels of serum electrolytes were mostly normal. Sodium (Na) was 139 mmol/L, potassium (K) was 5.0 mmol/L, chloride (Cl) was 107 mmol/L, calcium (Ca) was 7.7 mg/dL, and phosphate (P) was 4.6 mg/d. C-reactive protein (CRP) was 0.47 mg/dL. Urine analysis showed occult blood (OB) was 3+, and urine protein (UP) was 4+. Urinary sediment analysis showed that the urine red blood cell (RBC) count was more than 100/high-power field (HPF). No dysmorphic red blood cells were observed. Urine white blood cell (WBC) was 5–9/HPF. Spot urinary protein excretion was 15.83 g/gCr. The selectivity index (S.I.) of the ratio of transferrin clearance to immunoglobulin G (IgG) clearance was calculated as 0.14, indicating high selectivity. Urine protein levels were consistently high in the repeated urine examination after admission, but hematuria and leukocyturia were diminished.

An immunological test was performed. Serum complement analysis was normal. C3 was 146 mg/dL (normal range: 73–138 mg/dL). C4 was 15 mg/dL (normal range: 11–31 mg/ dL), and CH50 was 40.9 U/mL (normal range: 31.6–57.6 U/mL). The levels of serum immunoglobulin were as follows: IgG was 1803 mg/dL, IgA was 346 mg/dL, IgM was 192 mg/dL, and IgE was 165 mg/dL. Monoclonal protein was not detected. Testing for infectious diseases was investigated. Antibodies for hepatitis B virus surface antigen (HBs-Ab) and hepatitis C virus (HCV-Ab) were negative. Human Immunodeficiency Virus (HIV) Antigen–Antibody Combination Assay (HIV-AgAb) was negative. The levels of rapid plasma reagin (RPR) were 512 (normal range 0–1), and Treponema pallidum antibody (TPAb) were 1154.9 T.U. (normal range 0–9.9), indicating active syphilis infection. Full data is shown in Table 1. Of note, results of RPR and TPAb were both negative when she underwent salpingectomy performed 3 months before noticing nephrotic syndrome, which suggests she got infected during these 3 months. The patient completely denied any recent sexual activity, and she did not undergo any blood transfusion during the salpingectomy.

Table 1.

Characteristic of blood test

Serum			Blood			Coagulation
CRP	0.47	mg/dl	WBC	7100	/µ	PT	106.4	%
TP	5.5	g/dl	RBC	482×104	/µ	PT-INR	0.96
Alb	1.5	g/dl	Hb	11.7	g/dL	APTT	46.4	Sec
AST	16	U/L	Ht	38.2	%	Fib	557	mg/dl
ALT	11	U/L	MCV	79.3	fL	D-dimer	2.7	µg/dl
LDH	163	U/L	Plt	37.8×104	/L
CK	138	U/L				Urine
Na	139	mmol/L	Immunology			Gravity	1.047
K	5.0	mmol/L	ANA	(–)		pH	6.5
Cl	107	mmol/L	IgG	1803	mg/dL	UP	(4 +)
Ca	7.7	mg/dl	IgA	346	mg/dL	OB	(3 +)
P	4.6	mg/dl	IgM	192	mg/dL	RBC	100	/HPF
UA	3.8	mg/dL	C3	146	mg/dL	WBC	5–9	/HPF
BUN	10.0	mg/dL	C4	15	mg/dL	UPCR	15.8	g/gCr
Cre	0.56	mg/dL	CH50	40.9	IU/mL	β2-MG	480	µg/L
eGFR	88.0	mL/min/1.73 m2	PR-3 ANCA	(–)
Glu	82	mg/dL	MPO-ANCA	(–)		Other
HbA1c	5.0	%	HBsAg	(–)		S.I	0.14
T-Chol	251	mg/dL	HBsAb	(–)
TG	153	mg/dL	HCV Ab	(–)
LDL-C	182	mg/dL	TPHA	( +)
HDL-C	43	mg/dL	RPR	( +)
TSH	1.68	mIU/L	HBsAg	(–)
Free T3	2.21	pg/ml	HBsAb	(–)
Free T4	0.68	Ng/dL

UPCR: Urinary protein creatinine ratio. S.I. selectivity index; less than 0.2 indicates high selectivity

A kidney biopsy was performed for further evaluation. The light microscope of the renal biopsy sample showed mild interstitial fibrosis, tubular atrophy, and arteriolar hyalinosis. We found 17 normal glomeruli. No global sclerosis, segmental sclerosis, adhesion, or crescent lesion was observed in the glomeruli. Thickness of GBM, spikes, or bubbling appearance on the glomerular basement membrane was not observed (Fig. 2). Immunofluorescence (IF) analysis showed bright granular deposits of IgG, C3, C1q, κ, and λ in the glomerular capillary wall (Fig. 3). IgA, IgM, and C4 deposition was weakly positive. Subclass staining for IgG showed deposit of IgG1, IgG2, and IgG3, but not IgG4, in the glomerular capillary wall (Fig. 4), indicating secondary MN. Immunostaining for phospholipase A2 receptor (PLA2R) was negative. Electron microscopy (EM) showed severe foot process effacement and electron-dense deposits (EDD) on the subepithelial and mesangial lesion. The subepithelial EDD deposition appeared to be a hump (Fig. 5). The diagnosis was Ehrenreich–Churg stage I MN.

Fig. 2.

Light microscope image analysis on renal biopsy sample. Upper panel; (left) Periodic Acid-Schiff (PAS) stain, (right) Masson’s trichrome stain. Bar; 50 µm. Lower panel; Periodic Acid-Methenamine Silver (PAM) stain. Right panel shows enlarged images of the red box in the left panel. There was no GBM thickening, spike lesion, and bubbling appearance

Fig. 3.

Immunofluorescence staining. Scale bar, 50 μm. IgG, C3, C1q, κ, λwas positive on the glomerular capillary walls. As for the C4 staining, mouse monoclonal anti-C4d Ab (clone 10–11, BioRad, Cat#22,228,004) was used

Fig. 4.

Immunofluorescence staining for IgG subclass staining. Bright deposition was detected in IgG1, IgG2, and IgG3. The following antibodies were used for IgG subclass staining; IgG1(Cat # MA1-83,240, Invitrogen, clone 2C11), IgG2 (Cat# I5635, Sigma-Aldrich, clone HP-6014), IgG3(Cat# I7260, Sigma-Aldrich, clone HP-6050), IgG4(Cat# A-10651, Invitrogen, clone HP-6025)

Fig. 5.

Electron microscope image. Electron dense deposit (EDD) was detected in the subepithelial and mesangial lesion. Scale bar, 5 μm. White arrows indicate EDD in the subepithelial and mesangial lesion, enlarged from the white box

Recent case reports [4] suggest that NDNF can be an antigen for syphilis-associated MN. We performed the IF and IHC examination for NDNF using renal biopsy samples fixed with 10% formalin. After the deparaffinization procedure, anti-retrieval was performed using EDTA solution. Polyclonal Rabbit anti-Human NDNF Antibody (#LS-C168140) was incubated at room temperature for 60 min, followed by secondary Ab-conjugated Alexa555 or HRP. In the IF study, NDNF signal was brightly positive along with the glomerular wall in the renal biopsy sample of the patient, whereas the signal of NDNF was not detected in the normal kidney sample obtained from the transplant patient. (Fig. 6). We started to treat with Amoxicillin (1500 mg/day) right after the syphilis infection was confirmed. She did not show any signs of Jarisch-Herxheimer reaction, which is a constitutional symptom often observed right after antibiotic treatment. Her proteinuria was diminished rapidly without using immunosuppression therapy, and reached remission (UPCR<0.2 g/gCr) in 4 weeks after antibiotic treatment (Fig. 7).

Fig. 6.

NDNF staining. (Upper panel; IF, Lower panel; IHC). The NDNF was stained along with glomerular membrane. Scale bar, 20 μm. Control; normal kidney sample obtained from the transplant patient

Fig. 7.

Clinical course

Discussion

We report a case with membranous nephropathy followed by syphilis infection. There are several case reports of MN associated with syphilis infection [8]. Consistent with previous reports, the pathological findings show an early stage of MN and the hump-like deposition of EDD in the subepithelial and mesangial lesion. However, the clinical course was unique in our case. In the previous cases, typical rash or vaginal discharge preceded nephrotic syndrome [8]. In contrast, our case did not show typical clinical symptoms related to syphilis. We speculated the infectious timing was within 3 months before the development of nephrotic syndrome based on the negative results of syphilis markers in the salpingectomy. The absence of a Jarisch-Herxheimer reaction may support the possibility of a relatively small burden of syphilis. These findings indicate early latent infection of syphilis. We would like to highlight that it may be possible that nephrotic syndrome precedes the development of other manifestations of syphilis. The positive result of STS and TPAb suggests active infection of syphilis; however, the patient denied any sexual activity despite our closed question. Syphilis can be transmitted by the blood supply, but there was no history of recent blood infusion. Overall, we could not identify the exact infectious route of syphilis. Our case suggests the importance of screening for serum syphilis markers when encountered in nephrotic syndrome, even if no apparent symptoms related to syphilis are present.

NDNF was recently discovered by using mass spectrometry in the recent study [4]. Validated cohort in the study identified NDNF in all patients with syphilis MN, suggesting NDNF is a specific antigen in syphilis MN. A caution from the study is that NDNF can be detected in patients with HIV or lung cancer [4]. Our patient was not positive for HIV markers, and the lung CT did not show any suspicious shadow. Our case is consistent with clinicopathological features reported by Haas, including full-house pattern on the immunofluorescent method with bright signal of IgG, C3 and C1q, dominant staining for IgG1, IgG2 and IgG3 subtypes, subepithelial hump-like lesion, early stage of MN, and rapid complete remission after antibiotics treatment [4]. Although the cases reported by Haas were mostly young men, our case was a woman. A case study nicely showed the presence of NDNF in the serum from syphilis MN with Western blot [9]. In the future, it may be possible to detect NDNF using a serum sample with an ELISA assay [10, 11]. The pathogenesis of NDNF remained unknown, but it is believed to be involved in forming immune deposits in glomerular walls [12]. A case of renal biopsy after remission of nephrotic syndrome still showed IgG deposition, questioning the pathogenesis of immune deposition in NDNF-positive syphilis MN [11].

The infectious case of Syphilis has been increasing over the years, giving a pandemic caution in worldwide [2]. Therefore, cases of MN secondary to syphilis infection would be expected to be increasing in the future. We suggest that a screening test for syphilis infection is essential in patients with nephrotic syndrome, even if there are no symptoms related to syphilis.

Declarations

Conflict of interest

All authors declare no conflicts of interest. All authors declare that they have no conflict of interest.

Research involving human participants and/or animals

This article does not contain any studies with human participants or animals performed by any of the authors.

Informed consent

Informed consent was obtained from the patient.