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Written by : Gerren Hobby, MD
We will be discussing complement-mediated glomerulonephritis in this blog post. This has been an exciting topic to watch over the past several years as we have come to a better understanding of the role of complement in glomerulonephritis. The role of complement in ANCA vasculitis as well as IgA nephropathy has been studied in recent years, but some of the most striking examples of complement involvement are in membranoproliferative glomerulonephritis (MPGN). This has led to a change from the old classification of MPGN, which relied on histological appearance only, to a sharper focus on the pathophysiological driver of disease – the alternative complement pathway. This blog is part one of two on the subject. Another blog post will discuss the treatment of complement-mediated glomerular disorders in detail. For this blog, we will discuss the classification and diagnosis of MPGN, as well as how this new classification clarifies the diagnosis for patients right now and informs future studies.
Let's start with the history of membranoproliferative glomerulonephritis classification. The old system split MPGN into three types based on histological findings on light microscopy and the location of deposits on electron microscopy.
MPGN Type I: mesangial and subendothelial deposits
MPGN Type II: mesangial and intramembranous deposits
MPGN Type III: mesangial, subendothelial, subepithelial, and/or intramembranous deposits
This classification was found to be inadequate for multiple reasons. Overlap between the types of MPGN could occur, but the main issue is that the classification only indicated the presence of a histological appearance and made no attempt at suggesting an underlying driver of the disease. Because of this, once an MPGN pattern was seen on a kidney biopsy, the clinician was tasked with an evaluation of potential secondary causes such as HCV, HBV, HIV, collagen vascular disorders, malignancy and other infections. If this workup was negative, then the patient was deemed to have “idiopathic MPGN.” This classification left us with only vague ideas of the true driver of the disease process. This meant that patients with MPGN actually comprised a cohort of patients with a large number of mechanisms of disease. This heterogeneity of disease drivers, combined with its rarity, made it difficult to study effectively in clinical trials. Subsequently, we were left with the fact that 30-50% of patients progressed to ESKD within 10 years of diagnosis.
As more became known about the complement system in glomerular disease, peculiarities with the old classification arose. There were reports of cases with type I MPGN morphology but with low C3 and normal C4 levels, which suggested complement-mediated disease. Additionally, other case reports of MPGN were found to have intense C3 staining, but no immunoglobulin (Ig) staining, again suggesting alternative complement pathway activation. Lastly, descriptions of genetically mediated forms of MPGN due to complement gene mutations added to our understanding of the alternative pathway in these glomerular disorders. In sum, a need for a new taxonomy of the disease that gave us actionable diagnoses that could be addressed through treatments tailored to the specific immunologic defect was evident.
Other glomerular diseases have made this shift. Goodpasture’s syndrome, for example, moved from eponym to the term anti-GBM disease after the target epitope was discovered. ANCA vasculitis has also seen a change in naming, and we are currently observing the refinement of the categorization of membranous nephropathy due to the prolific rise in culprit antigens.
In 2010, a new classification for MPGN came about that was based on the increased appreciation of the alternative complement pathway in membranoproliferative glomerulonephritis. The new classification reclassified MPGN subtypes into immune complex membranoproliferative glomerulonephritis (IC-MPGN) and complement 3 glomerulopathy (C3G). There is a further subdivision of complement 3 glomerulopathy into C3 glomerulonephritis (C3GN) and dense deposit disease (DDD). In this classification, dense deposit disease essentially replaced MPGN type II. MPGN type I and III now fall into C3 glomerulonephritis or immune complex glomerulonephritis.
With a new classification based on the mechanism of disease, we can shift to a discussion of pathogenesis. We are finding that many glomerular disorders ultimately involve complement in their pathogenesis despite the inciting event. In anti-GBM, a conformational change in the NC1 domain of type IV collagen creates a neoepitope to which autoantibodies form. Downstream of this, T and B cells are ultimately involved as well as the complement system. In ANCA vasculitis, autoantibodies prime neutrophils to create vessel damage, but an increased appreciation for the alternative pathway has gained traction. So what about C3G and IC-MPGN? In these disease states, the inciting event differs by disease process, thereby forming the basis for the initial categorization. In C3G, the pathogenesis begins with the alternative complement pathway rather than an upstream event. Since the alternative complement pathway is constitutively active, loss of regulation causes overactivation and glomerular damage. With IC-MPGN, the pathogenesis begins with the classical pathway but is now known to involve the alternative pathway as well. To better understand how these diseases occur, let’s dive into the alternative complement pathway.
The Alternative Complement Pathway
The complement system is composed of more than 30 soluble and membrane-bound proteins, which make up the classical, lectin-binding, and alternative pathways. These pathways have a seemingly over-complex naming system, but this is due to the fact that many of the proteins are zymogens that need to be cleaved to become active. After realizing that, the taxonomy makes more sense as it denotes the smaller cleavage fragment “a” and the larger fragment “b” (i.e. C3a and C3b, respectively). All three pathways converge on C3 convertase, and the end result is the formation of the membrane attack complex (MAC).
To begin, let’s discuss the classical pathway, which is initiated when the C1qrs complex binds the Fc region of IgG or IgM.The bound C1qrs complex cleaves the zymogens C4 and C2 to form the C4bC2b C3 convertase. The lectin-binding pathway is a bit different as it is initiated when hexamers of mannose-binding lectins bind bacterial carbohydrate motifs and associate with MBL-associated serine proteases to cleave C4 and C2 to again form the C4bC2b C3 convertase.
In contrast, the initiation of the alternative pathway is distinct due to the fact that activation is continuous, spontaneous, and begins with C3. Each day, 1% of total C3 undergoes hydrolysis at a thioester bond to spontaneously activate the alternative pathway. This process is called “tick over,” and it allows C3 to recruit factor D, which cleaves factor B, a zymogen, to form it into the active serine esterase Bb (a.k.a. C3 convertase) that cleaves C3 to C3a and C3b. Next, C3b associates with Bb to form C3bBb, a C3 convertase that then amplifies the entire process.
Due to the constitutive nature of the alternative complement pathway activation, regulation is key to keeping the system in check. Some of the regulatory proteins include:
Factor H: a serum protein that promotes the decay of C3 and C5 convertases; acts as a cofactor for factor I to inactivate C3b
Factor I: inactivates C3b
Membrane cofactor protein (MCP): a surface protein that is a cofactor for serum factor I, which cleaves and inactivates C3b
Complement receptor 1 (CR1): accelerates decay of C3 and C5 convertases; also a cofactor for serum factor I
Decay accelerating factor (DAF): accelerates decays of C3 and C5 convertases
This tight regulation of the alternative pathway can be dysregulated through several mechanisms. The first and most common way are autoantibodies that stabilize C3 convertase, thereby delaying its decay. These autoantibodies are referred to as C3 nephritic factors (C3NeFs). Why is this important? In contrast to C3 which lasts a relatively long time, the C3 convertase C3bBb only has a half-life of 90 seconds due to the action of multiple regulatory proteins. When this regulation is compromised, complement-mediated disorders occur. This loss of regulator activity in the alternative pathway can happen due to:
Autoantibodies to factor H that prevent its action on C3b
Mutations in complement factor H or I that prevent it from inactivating C3b
C3 nephritic factor which stabilizes C3 convertase by preventing it’s degradation by factor H
C5 nephritic factor which stabilizes C5 convertase
MCP mutations that disrupt it from functioning as a cofactor to factor I. This leads to higher C3b levels
CFHR mutations which allow these proteins to bind to C3b as factor H would, but lack regulatory function
Monoclonal immunoglobulins that inhibit factor H
Pathogenesis of C3G and IC-MPGN:
In C3G, genetic variants in alternative pathway genes or autoantibodies, such as C3 nephritic factor, lead to the overactivation of the alternative pathway. IC-MPGN on the other hand, is initiated by activation of the classical pathway. In many patients, IC-MPGN is secondary to infections, autoimmune conditions, or a monoclonal gammopathy. Despite the fact that the classical pathway is the initial pathogenic event in IC-MPGN, there is a large amount of evidence suggesting excessive alternative pathway activation in its pathogenesis. For instance, the pattern of low C3 levels along with normal C4 is seen to a similar degree in IC-MPGN as in C3G. Additionally, there are similar levels of genetic variants and autoantibodies in IC-MPGN as in C3G. Taken together, although the classical pathway initiates the complement system in IC-MPGN and there are a variety of secondary causes, a strong role for the alternative pathway in its pathogenesis is coming into clear view, specifically for cases of idiopathic IC-MPGN. This increasing recognition of the role of the alternative pathway in IC-MPGN has important diagnostic and therapeutic implications.
Clinical Presentation and Diagnosis:
Patients with C3G or IC-MPGN present with proteinuria with or without hematuria with wide variation in the degree of proteinuria and kidney function. Dense deposit disease primarily occurs in the pediatric population, whereas C3GN and IC-MPGN tend to occur later in life. Although a low C3 and normal C4 suggest alternative pathway activation, a normal C3 level can be seen in over a third of cases of C3G and IC-MPGN. Conversely, a pattern of both low C3 and C4 can be seen in IC-MPGN secondary to hepatitis C.
On kidney biopsy, once an MPGN pattern is seen, the new classification system utilizes immunofluorescence to help narrow down the driver of disease. As mentioned above, the first step is immunofluorescent evaluation for C3 and immunoglobulin staining. This step splits MPGN into two major groups.
Complement 3 glomerulonephritis (C3G): characterized by predominant C3 immunofluorescence that is at least two orders of magnitude higher than other immune reactants. Ig staining can be present but must be less prominent than C3 staining. C3G is then subdivided into:
C3 glomerulonephritis (C3GN): has a “cloudy” appearance of C3 immunofluorescence due to granular mesangial and segmental subendothelial deposits
Dense deposit disease (DDD): has “ribbon-like” C3 deposits along the GBM and lamina densa which have recently been shown to be apolipoprotein E (ApoE) rich. Immunohistochemistry for Apo3 may be useful in the future for the diagnosis of DDD. More importantly, though, it may play a role in the terminal complement cascade and this informs future research in pathogenesis and therapeutics.
Immune complex membranoproliferative glomerulonephritis (IC-MPGN): immunofluorescence of both Ig and C3
From a pathogenesis standpoint, a histological finding of C3G suggests alternative complement pathway overactivation. This necessitates a search for the underlying driver of alternative pathway overactivation via genetic testing as well as testing for autoantibodies like C3 nephritic factor. Genetic variants in alternative pathway components (Table 1) can be seen in up to 40% of patients with C3GN and 33% of those with DDD. Evaluation for C3 nephritic factor is useful as it is seen in 45% of patients with C3GN and 86% of patients with DDD.
A histological finding of IC-MPGN means that the next step is a search for secondary causes of IC-MPGN. Sometimes the kidney biopsy helps narrow the list of possibilities. Light chain restriction, for example, indicates a monoclonal gammopathy. A biopsy showing immunofluorescence with IgM, C3, kappa/lambda, +/- IgG and no C1q suggests HCV infection. Lastly, a full-house pattern with IgA, IgM, IgG, kappa/lambda, C1q, C3, and C4 suggests MPGN secondary to autoimmune diseases such as lupus, Sjogren’s disease, or rheumatoid arthritis. Past these findings that suggest a particular disease, a dizzying array of chronic infections exist that are associated with an MPGN pattern, including HCV, HBV, endocarditis, other chronic bacterial infections, and schistosomiasis. A thorough history, as well as testing for secondary disorders, is a vital component to combine with a biopsy for a proper evaluation of the patient. Once secondary causes are excluded, the patient is deemed to have idiopathic disease, which suggests a large role for alternative pathway involvement.
Our understanding and classification of MPGN has evolved and is likely to continue to do so in the future. The pathogenesis of this collection of disease processes is complex and not yet fully understood. A large number of disease mechanisms converge to create the pattern of MPGN on kidney biopsy. A recent KI Reports paper highlights the complexity in the evaluation of a patient with MPGN and a wide range of presentations of the disease with seven illustrative cases.
Prognosis:
C3G and IC-MPGN are rare diseases which makes it difficult to precisely define the prognosis. That being said, the rate of progression is high with low figures of 30-35% of patients progressing to ESKD within 10 years of diagnosis. Higher For DDD, specifically, 50% of children reach ESKD within 10 years of diagnosis. As such, many of these patients ultimately receive a kidney transplant. Unfortunately, patients with DDD have more than a 65% rate of recurrent disease after kidney transplant. A recent KI Reports article examined the outcome of C3G after kidney transplants in a Swiss cohort of 41 patients with C3G and IC-MPGN. In a mean follow-up of 4.7 years, seven patients had disease recurrence. Those with disease recurrence experienced a 28% rate of graft loss. A systemic analysis of C3GN and DDD by other researchers showed a rate of graft loss of 53% for DDD, even after treatment with eculizumab and a 22-70% rate of graft loss for C3GN, depending on treatment.
Conclusion:
Over the past several decades, our understanding of MPGN has evolved considerably. What was once merely viewed as a pattern of disease on kidney biopsy has gained clarity based on the involvement of the alternative pathway in its pathogenesis. This is important for several reasons. Firstly, we obtain biopsies to find out the cause of a patient's disease. The new classification helps us narrow the list of possibilities to a smaller number of options. Secondly, any time we study a disease process, a homogenous cohort of patients helps us define the natural history of the disease, pathogenesis, and potential treatments. A more precise definition of the glomerular disease helps with this. Lastly, there has been significant progress in the development of alternative complement pathway inhibitors, which gives us hope for improving outcomes. In total, we now have a more precise definition of MPGN which will help us study the disease better and develop treatment protocols. It has been exciting to watch this greater understanding of pathophysiology and associated evolving treatments that will lead to improved patient outcomes in the coming years.
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