Histological Spectrum of Hepatitis-Virus Associated Glomerulonephritis

Introduction

Hepatitis virus-associated glomerulonephritis (HVGN) is a recognized extrahepatic manifestation of Hepatitis B (HBV) and C viral (HCV) infections.^1-3 Pathogenesis of HVGN includes immune complex-mediated injury, direct cytotoxic viral effects, and/or due to cryoglobulins.⁴ Glomerular disease can occur in 2.6–4.3% of patients with chronic HBV infection.^5-8 The histological spectrum of HBV-associated GN commonly includes membranous nephropathy (MN), membranoproliferative glomerulonephriti (MPGN), IgA nephropathy (IgAN), and focal segmental glomerulosclerosis (FSGS).⁹ In HCV-associated GN, the glomerular pathology is commonly MPGN with or without Type 2 cryoglobulinemia, followed by MN, FSGS, and mesangioproliferative GN.³ Studies on hepatitis virus-associated renal disease are limited. The time between the detection of hepatitis via serology and the development of HVGN needs to be documented.

The factors for the renal prognosis of HVGN are not well-documented. There are no reports on HVGN from Southeast Asia and the Indian subcontinent. We report the prevalence and histological spectrum of HVGN in a tertiary care center in North India.

Materials and Methods

The study included renal biopsies of patients with serological evidence of HBV and/or HCV infection over nine years (2014 to 2022) received in the Department of Pathology, SGPGIMS. Patients with HBsAg, HBV DNA, HCV antibody, and HCV RNA positive serology were included.

Clinical data and viral serology, namely HBsAg, HBeAg, HBe-Ab, HBV DNA, and HCV RNA levels were retrieved. Laboratory parameters, including autoimmune workup, urinalysis, seruigm biochemistry, and serum complement levels were noted from the records. The estimated glomerular filtration rate (eGFR) was calculated using the chronic kidney disease epidemiology collaboration (CKD-EPI) creatinine equation. Oxford classification of IgA nephropathy criteria of tubular atrophy/interstitial fibrosis was used to assess the degree of tubulointerstitial injury.

All the biopsies were processed for light microscopy, immunofluorescence (IF), and electron microscopy using the standard protocol and stained with hematoxylin and eosin stain (H&E).

Sections of formalin-fixed paraffin-embedded (FFPE) renal tissue were analyzed by IHC using prediluted HbsAg (A10F1, Master Diagnostica, Spain). The staining was performed according to the manufacturer’s instructions on a BenchMark-XT slide stainer (Ventana Medical Systems, Inc.). Liver biopsy tissue from an HBsAg-positive patient was used as a positive control. The Institute Ethics Committee approved the study (2024-151-IP-EXP-59) and patient consent was obtained.

Results

A total of 5179 native kidney biopsies were received between January 2014 and December 2022 in the department of pathology, SGPGIMS, Lucknow. HbsAg was positive in 49 patients; HBV DNA levels ranged from 20 to 406 million/IU. HCV antibody was positive in 10 patients, HCV RNA levels ranged from 1.64 to 3319 million/IU. The prevalence of HVGN was 1.1%, mostly due to HBV-related GN (0.95%), which constituted 83% of all HVGN.

Table 1 shows the demographic and clinical features of HBV- and HCV-related GN. Transmission electron microscopy and evaluation were available for 21 biopsies. The findings were incorporated while analyzing the biopsies.

HBV-related renal disease

Of 49 patients, 12 were known to be HBsAg positive, and 37 were detected during workup for renal disease. In the former group, the duration from HBV infection to onset of renal disease was 8-168 (median-24) months [Table 1]; five patients (41%) were on antiviral therapy, and three were on remission. They also had a significantly (p=0.028) higher (33%) incidence of chronic liver disease (CLD) as compared to the newly diagnosed (6.8%).

Nephrotic-range proteinuria was observed in 27 patients (55%). Liver function tests were deranged in 18 (37%), of which 4 (8%) previously had HBV infection. One patient, positive for HBsAg, was co-infected with HIV. No significant clinical and histological features were associated with the presence of HBeAg.

IgAN (n=13; 26.5%) was the most common histological diagnosis [Figure 1a-b], followed by membranous nephropathy (n=8; 16.3%) [Figure 1c-e]. The other histological patterns included diffuse global glomerulosclerosis (DGGS) (n=8; 16.3%), minimal change disease (n=3; 6.1%), FSGS (n=3, 6.1%), amyloidosis (n=3; 6.1%), pauci-immune crescentic GN (n=3; 6.1%), and immune-complex mediated MPGN (n=2; 4%,). Tubulointerstitial pathology (acute tubular injury and tubulointerstitial nephritis) was noted in six (12%) biopsies. Histology for patients co-infected with HIV showed IgAN. Histology of five patients with CLD showed DGGS (n=2), MN (n=1), IgAN (n=1), and MPGN (n=1).

Close

Figure 1:

IgA nephropathy: (a) Glomerulus shows diffuse mesangial hypercellularity (H&E,400x); (b) 3+ glomerular mesangial immune deposit (IF,400x). Membranous nephropathy (MN): (c) Glomerulus is enlarged with thickened and stiff capillary loops (PAS,400x); (d) Glomerulus showing subepithelial argyrophilic spikes (PASM, 400x); (e) 3+ fine granular subepithelial immune deposits (IF, 400x). Cryoglobulinemic glomerulonephritis: (f) Glomerulus shows fuchsinophilic subendothelial deposits and intra-capillary hyaline thrombi (MT,400X). (g) Positive control of Hepatitis B-virus (HBV) surface antigen IHC in a case of HBV positive liver biopsy (IHC,400x); (h) A case of MN negative for immunohistochemistry (IHC,400x).

Export to PPT

Most biopsies of IgAN showed segmental sclerosis (S1) (9/13) and mesangial hypercellularity (M1) (7/13). Only two biopsies showed endocapillary hypercellularity (E1), and one showed crescents (C1); significant tubular atrophy and interstitial fibrosis (T1-2 lesions) were present in six. In MN, one biopsy was PLA2R IHC positive, and three had raised serum anti-PLA2R levels. Two patients were not tested for serum PLA2R. IF did not show light chain restriction in all three amyloids and serum electrophoresis was negative for monoclonal proteins. Two patients had a history of long-standing tuberculosis, suggesting associated Serum Amyloid A (SAA).

IHC was performed in 40/49 biopsies, and none were positive for HBsAg expression [Figure 1f-g].

Discussion

The prevalences of HBV and HCV infection in India are 0.87-21.4% and 0.19- 53.7%, respectively.¹⁰ The data on hepatitis virus-associated renal disease from India and Southeast Asia are limited. We studied the prevalence and clinicopathological characteristics of HBV and HCV-associated renal disease over nine years in a tertiary care Institute in North India.

We observed a 1.1% prevalance of HVGN, primarily due to HBV-related GN (0.95%), which is much lower than other Asian countries, like China, where it has been reported as 2.6-4.3%.^5-8

The pathogenesis of hepatitis-associated GN involves the deposition of in-situ or circulating immune complexes, virus-induced immunological effector mechanisms, or direct cytotoxic glomerular or tubular injury.¹¹ The size, charge, and molecular weights of hepatitis antigens determine their pathogenicity and location of immune deposits.¹² Formed HBeAg antigen-antibody immune complexes are cationic and small and tend to deposit in the subepithelial region. In contrast, HBsAg or HBcAg-antibody complexes are anionic and large and restricted to the mesangial and subendothelial regions.² Renal tubular epithelial cells have been known to express HBsAg and HBcAg.¹³ Studies show that serum from patients of chronic hepatitis B promotes apoptosis of renal tubular cells by upregulating the Fas-signaling pathway.¹⁴

In our study, IgAN and concomitant MPGN and IgAN were the most common renal manifestations of HBV and HCV-associated renal diseases, respectively. The higher prevalence of IgAN could be related to liver dysfunction in these patients. In contrast, Kong et al.¹⁵ and Chen et al.¹⁶ reported MN as the most common GN. However, Zhang et al.¹⁷ reported mesangioproliferative GN as the most common type. HBV infection was identified as an independent risk factor for CKD in rural China.¹⁸

Kong et al., studied 500 renal biopsies and detected HBsAg and HbcAg by IHC in only three (0.6%) and six (1.2%) of them, respectively.¹⁵ IHC of all biopsies in our study were negative for HBsAg, despite a positive control in each staining batch. This may be because we performed IHC on FFPE tissue, while many of the previous studies demonstrated immune staining on frozen tissue.^15,17 Similar results were observed by Nikolopoulou A et al. when they evaluated the staining in six renal biopsies.¹⁹ Zhang Y et al. detected HBsAg in all ten biopsies with acute post-infectious HBV GN.²⁰ Chen et al. and Zhang et al. demonstrated HBsAg in 76.2% and 41.3% of renal biopsies, respectively.^16,17 Table 2 shows the summary of various studies evaluating IHC for HBsAg in renal biopsies of positive patients. Prospective studies utilizing more sensitive methods on fresh renal tissue in known HBV/HCV patients are required to understand the pathogenesis of HV-associated renal disease.

On follow-up, 15% of our patients developed end-stage renal disease, requiring renal transplant. There are studies on the effect of HBV infection in renal transplant recipients. HBsAg-positive kidney transplant recipients face a 2.5- and 1.5-fold higher mortality risk and risk of allograft loss, respectively.²¹ There was 80% graft survival after 6 months of transplantation in HCV-positive renal transplant recipients compared to 91% in HCV-negative patients (p < 0.001).²² Mo et al. found significant differences in survival between positive and negative HBsAg patients and those receiving antiviral therapy (p = 0.020). The study concluded that lifelong antiviral therapy is essential, and patients with high preoperative HBV titers should be closely monitored for reactivation.²³

The European Association for the Study of the Liver (EASL) guidelines suggest that combined liver and kidney transplantation should be considered in cirrhotic patients with renal disease.²⁴ Direct-acting antiviral therapy has significantly improved outcomes in HCV-positive renal transplant recipients.²⁵ Therefore, studies are needed to determine if patients with hepatitis C can undergo kidney transplants with acceptable long-term outcomes.

In conclusion, we present our study of HBV and HCV-related renal disease, with a focus on HBV. IgAN was the most common GN associated with HBV infection, while MPGN, both immune-complex mediated and C3GN, and IgAN were most common in HCV-related GN. We suggest routine screening of all patients with renal disease for hepatitis infection.