Membranoproliferative glomerulonephritis (MPGN) is a pattern of glomerular injury. MPGN is characterized by mesangial expansion and hypercellularity, endocapillary hypercellularity, and double contour formation along the capillary walls. MPGN can result from 2 main causes: 1) deposition of Ig/immune-complexes (IC) and 2) deposition of complement factors. Based on the etiology of MPGN, Drs. Sethi and Fervenza proposed a new histologic classification of MPGN into Ig/IC-mediated MPGN and complement-mediated MPGN. In this recording, Dr. Sethi demonstrates the use of the new classification with case studies.
Presenters and Credentials:
Sanjeev Sethi, M.D., Ph.D., Professor of Laboratory Medicine and Pathology and Consultant in the Division of Anatomic Pathology at Mayo Clinic in Rochester, Minnesota.
Our speaker for this program is Dr. Sanjeev Sethi, Professor in the Department of Laboratory Medicine and Pathology, as well as a consultant in Anatomic Pathology at Mayo Clinic in Rochester, Minnesota. Dr. Sethi has recorded 2 Hot Topics on the topic of membranoproliferative glomerulonephritis or MPGN. In part 2, Dr. Sethi discusses additional cases of MPGN and provides an algorithm for diagnosis and differentiation of the MPGN types, including C3 glomerulopathy. Thank you, Dr. Sethi, for presenting with us today.Welcome to Mayo Medical Laboratory’s Hot Topics. These presentations provide short discussion of current topics and may be helpful to you in your practice.
Okay, so let’s start from here. So patient 1, patient 2 had immune-complex mediated membranoproliferative glomerulonephritis or MPGN. Patient 1 was an infection. Patient 2 had a dysproteinemia, and I can give you many examples of MPGN from autoimmune diseases as well. So, those are the immune-complex mediated MPGNs. But what about the complement-mediated MPGNs, or what’s called C3 glomerulopathy? And that was patient 3 and patient 4.
So this is a sort of a cartoon that I drew up to explain these 2 disease entities. And, if you look on that first phase, you get injury, basically, from immunoglobulins. That activates the complement system, typically, the classical pathway of complement, and you get all these deposits. That’s the immunoglobulin, the blue stuff. And the yellow stuff is all the complement that’s deposited. Once they are deposited along the glomerular capillary wall, so that’s the subendothelial region, that’s the GBM, and then these are the podocytes, okay? So everything is in circulation and gets deposited along the capillary walls. Once it gets deposited over here, regardless whether this is coming from an infection, regardless whether this is coming from an autoimmune disease, regardless whether it’s coming from our monoclonal gammopathy, or what’s called a paraprotein or a dysproteinemia, this drives inflammation. So once you have this stuff here, first the neutrophils, then the monocytes, they come in to clean this mess up. Once they are trying to clean this mess up, they release their proteases; that’s the little p that I have in there. These proteases are enzymes that are present in the granules within the neutrophils and the monocytes, and what this does is it damages these capillary walls, makes big holes over there. And as a result, the red blood cells leak out; you can see it over here. That’s how these patients get hematuria. That’s how they get proteinuria. That’s how they get renal failure. And this is what the proliferative phase is, okay?
What the glomerulus does is, it’s maybe in my own way of thinking, it is trying to fool these neutrophils and monocytes into not seeing these immunoglobulins and complement because that’s what they’re coming after. So it’s kind of hiding these bad elements, and what it does is, it makes a new basement membrane over here. And, as a result, now you have this extremely thick capillary wall. Here is the old basement membrane. Here is the new basement membrane, and this is what’s the double contour or the tram-track appearance. But remember, you’re seeing immunoglobulin and you’re seeing complements. So as pathologists over here, the immunofluorescence is extremely important, and it tells me what’s entrapped there. But looking at the material that’s entrapped in these thick capillary walls, I can tell you what the underlying etiology is. Is this an infection, is this autoimmune disease, or is this a paraprotein, or a monoclonal gammopathy? Just by looking at this material based on the immunofluorescence studies, I am able to give you an underlying etiology.
Here is the other part of the story. You don’t really need immunoglobulins. So if you have overactivation of the complement system one way or the other, the complement by themselves can also activate, can also cause injury, and cause neutrophils and monocytes to come in and, then again, cause similar pattern of injury or red blood cells leaking in. Eventually you get these thick capillary walls and you get these double contours. But remember, the key finding here is the almost complete absence of immunoglobulins. The complement factors that are deposited on the capillary walls themselves are capable of injuring the capillary walls. They are capable of calling in the neutrophils and monocytes and causing the ensuing damage, and then again, you get this thick GBM. But when I look in the biopsy in these patients, I find almost no immunoglobulins or very little immunoglobulins and, mostly it’s completely loaded up with just complement factors. So, we call that complement-mediated MPGN or C3 glomerulopathy.
And those were, indeed, our 2 patients. Patient number 3 and patient number 4 had only complement in them. And the term that’s commonly used for this complement-mediated glomerulonephritis is C3 glomerulopathy, and it basically encompasses the 2 disease entities. One is dense-deposit disease. This is when the deposits are intramembranous. When you get dense intramembranous deposits, we call it dense-deposit disease. On the other hand, when the deposits are not intramembranous or dense and there are subepithelial, subendothelial, and mesangial all over the place, we call it C3 glomerulonephritis. C3 glomerulonephritis, in the past, was one of the commonly misdiagnosed as MPGN type I and type III. Many of these cases now are being relabeled as C3 glomerulonephritis.
So what’s the underlying etiology? I wanted to show you a few more examples of C3 glomerulonephritis or C3 glomerulopathy. So while MPGN pattern of injury–that lobular accentuation, the double contours, the endocapillary proliferation–is the most common presentation on the biopsy, not every patient has that. This shows you the classic MPGN look, you can see the double contours. This one also shows you the MPGN-like picture. So the first 2 patients show you the MPGN pattern of injury. This one actually shows you what’s called the segmental sclerosis or the focal sclerosis, and you can see there is segmental adhesion of the capillary tufts to the Bowman’s capsule up there and up here. Here again, MPGN. This, on the other hand, shows you a mesangial proliferative lesion. It’s not so much of an MPGN; it’s more in the mesangium. And then finally, you have crescents. So, bottom line is, although the MPGN pattern is common, other patterns are also seen. So a mesangial proliferative, or a segmental sclerosis, all represent a glomerulonephritis, may all be present. So MPGN, by and large, is not the only pattern of injury that you see in patients with C3 glomerulopathy.
IF- Bright C3, Negative Ig
So just to show you, these 5 patients that I just showed you, how we make the diagnosis. The IF only shows you the C3; all immunoglobulins are negative, so it fits the story of C3 glomerulopathy.
EM: Mesangial, Subendothial and Subepithelial Deposits
The EM shows you the subendothelial deposits here. In this case, there’s a subepithelial. So in the past, you might have called this type III. Same thing here, you would have called this type III, but these are just C3 glomerulopathies. And this last patient has this dense deposit. So it’s really dense-deposit disease. So patients 1, 2, 3, and 4 are C3 glomerulonephritis, because they don’t have those dense deposits. They just have these scattered mesangial and capillary wall deposits. Regardless of whether that’s subepithelial and subendothelial, these are all C3 glomerulonephritis. And this patient with dense deposit is dense-deposit disease. So, indeed, these are all cases of C3 glomerulopathy 1, 2, 3, 4; you could subcategorize it as C3 glomerulonephritis, and patient 5 would be dense-deposit disease.
Kidney Biopsy Diagnosis
Like I said, MPGN pattern is the most important or the most common, and then you can get all of the other patterns as well. The diagnostic criteria is the presence of complement in the mesangium and along capillary walls, but minimal or no staining for immunoglobulins.
Criteria: C3 Glomerulonephritis
In the setting of C3 glomerulonephritis, you’ll get these large subendothelial and mesangial deposits. If you have intramembranous deposits, you would call it dense-deposit disease.
Clinical Features and Laboratory Evaluation: C3 Glomerulonephritis
Now, in the setting of dense-deposit disease, you can have a bimodal, patient may be young or be elderly, but in general, you can see where the sex distribution is wide. And you’ll see over here, young children from the age of 8 to 73, and older patients from the age of 73 can get it; a mean of 42. Both males and females are affected. Most patients will present with a rise in creatinine, or meaning they have renal failure. Most patients have hypertension. Most patients will have hematuria and proteinuria. Sometimes, the proteinuria can be fairly severe. We have seen patients with even 15 g of protein a day. Because the complement system is activated, C3 levels are low. C4 are often normal because this indicates activation of the alternative pathway, whereas C4 is a part of the classical pathway, so the C4 levels often are normal.
Classical Pathway- Alternative Pathway
So, this is an important slide. So let’s try and figure out what’s going on in patients with C3 glomerulopathy. And to understand that, we need to know the very basics of the complement system or at least the very basics of the alternative pathway of complement system that is involved in C3 glomerulopathy. What happens is in circulation, C3 is being degraded at a very low level; it’s sort of a hydrolysis, autohydrolysis, to form small amounts of this component called C3b. C3b is almost immediately degraded into its degradation products to 2 important complement-regulating proteins. And one is factor I and it needs factor H, so factor I with a cofactor, factor H, that’s CFH. C3b is almost instantaneously degraded into its small degradation products. If this does not happen, some amounts of C3 may escape and then combine back with the C3. And what this forms is a sort of an intermediate enzyme called the C3 convertase. The C3 convertase is a very active enzyme. What it does instead is now it goes back and breaks up even more C3 to C3b. The C3 convertase then breaks up C3 into more C3b. Once you get more C3b, it can combine with more C3 to form more C3 convertase. So basically, this becomes sort of an amplification loop. So the key regulation really lies over here. That means the C3 that forms C3b, most of the C3b is then degraded very rapidly so that the C3b doesn’t combine with C3 to form this C3 convertase. So that’s one major regulation over there. The other regulation is once C3 convertase forms, it is further broken down, also fairly rapidly, by the presence of factor H. And then there are certain regulating factors on the endothelial cells themselves that can regulate the breakdown of C3 convertase. But keep in mind that one of the major role-players over here is factor H, which helps break down the C3b into C3b degradation products and then also breaks down the C3 convertase. The C3 convertase, once it is formed, is very active, breaks down more C3, forms more C3b, and there’s a whole amplification loop that takes place.
The C3 convertase, on the other hand, then proceeds down this particular aspect is the alternative pathway. Once it goes down this is the common or the terminal pathway, it then forms a C5 convertase. The C5 convertase then breaks down C5 into C5b, which combines with C6, 7, 8, and 9 and forms a terminal complex. The terminal complex is what’s the active part of the alternative pathway. It can sit on bacteria and punch holes through their cell membrane. It can sit on dead cells and punch holes. It can sit on cancer cells. So it’s basically the end-product of the alternative pathway activation. And as you can see, the alternative pathway is fairly well controlled over here because you don’t want this going haywire. You don’t want a lot of this particular thing being formed, the sMAC being formed, so that it keeps destroying cells in a haywire manner.
AP Workup in 5 Cases of MPGN
So patient 1, when we tested this particular patient had an autoantibody to factor H. Patient 2 had an autoantibody to the C3 convertase. Patient 3 actually had a mutation in factor H. Patient 4, although didn’t have any mutations but had polymorphisms to factor H; and patient 5, the one that had dense-deposit disease, had polymorphisms to factor H and also had an autoantibody at very high titer to the C3 convertase.
The Alternative Pathway of Complement
So let’s look at this pathway one more time. So here’s the alternative pathway. It forms C3 in small amounts, forms C3b. The C3b, like I said in the presence of factor I and factor H, is then broken down into these degradation products. If C3b escapes, it forms the C3 convertase. The C3 convertase then forms a C5 convertase, breaks the C5 into C5b, which combines with these C6, 7, 8, 9, to form this terminal complex, and so it’s called the soluble MAC or the membrane attack complex. Our patient 1 had antibodies to C3 convertase and that’s called C3NeF or C3 nephritic factor. What that does is it stabilizes the C3 convertase; remember I said the C3 convertase is a very active enzyme; it breaks down C3 into C3b. Now, the C3 convertase has a very short half-life. It’s broken down almost instantaneously. When you have autoantibodies to C3 convertase or the C3Nef, now the C3 convertase becomes very stable. And as a result, it can go on for a much longer time. That means it keeps breaking C3 into C3b, and then the cycle goes on. The other scenario is patient has an autoantibody. So, patient 2 had an autoantibody to the C3 convertase, so C3NeF. Patient 1 actually had an autoantibody to factor H. So there you go. That’s patient 2. That’s patient 1. If you make an autoantibody to factor H, now factor H can no longer do its job. That means it can’t really break down C3b into its degradation products, number 1. Number 2, Factor H is also required to break down C3 convertase, and no longer can the C3 convertase be broken down, so it sort of goes unchecked, and you get more C3 into C3b. Patient 3 had a mutation in factor H, and patients 4 and 5 had polymorphisms.
DDD vs C3GN Pathophysiology
So anything wrong with the factor H, that means you have a mutation or you have these alleles in factor H that predisposes you to poor function of factor H can also result in overactivity of the C3 convertase. It can also result in overactivity of C3 just breaking down into C3b, and now you cannot make these degradation products. Sometimes, you can have a mutation in factor I. Again, if you have a mutation in factor I, the C3b cannot be degraded, and you get lots of C3b accumulating. Finally, rarely, you can also have a mutation in C3. If you have a mutation in C3, the mutation is at such a site that the factor H and the factor I can’t act on it. So you have a mutation in C3 itself, it will lead to overactivity. So these are just some ways that the alternative pathway can become abnormal or can get overactive. It gets more complex, but these are the more common abnormalities of the alternative pathway that result in overactivity.
So what’s the difference between dense-deposit disease and C3 glomerulonephritis? So why does one patient, for example, over here who has a factor H mutation, get C3 glomerulonephritis, and then another patient that has a factor I mutation get dense-deposit disease? We don’t know for sure, okay, at this point, but the speculation is that when the C3 convertase is active, overactive, and you also have overactivity of the C5 convertase, and when both C3 and C5 convertases are overactive, we think that these patients end up with C3 glomerulonephritis.
On the other hand, when the C3 convertase is more active than the C5 convertase, we think that these patients might end up with dense-deposit disease. So this is just speculative at this point, and that’s what I’m trying to show you over here that in dense-deposit disease, there is overactivity of the C3 convertase compared to the C5. In C3GN, the thinking is both C3 convertase and C5 convertase are both equally active. So when you have both equally active, you get more of these larger products, the sMAC being deposited, and that causes these large deposits. On the other hand, in dense-deposit disease, it’s more C3 convertase; that means you get these small breakdown products of C3 being accumulated and they form the dense-deposit disease.
Acquired and Genetic Abnormalities Associtaed with Complement-Mediated MPGN (C3 Glomerulapathy)
So the take-home message in C3 glomerulopathy is these are abnormalities of the alternative pathway of complement. So, very simply put, how can these abnormalities come across? One is these are inherited or they could be acquired. So, what’s acquired? Acquired basically means making antibodies, making antibodies to complement-regulating proteins such as factor H, factor I, factor B, or you can make antibodies to the C3 convertase itself, that’s the C3 nephritic factor. So that’s the first group. So these are the acquired. On the other hand, the second one would be the inherited or the genetic abnormalities, and these are mutations in all the complement-regulating proteins or the allele variants. Rarely, you can get a mutation in a complement factor itself like C3, and bottom line is, all of these results in abnormalities or overactivity of the C3 convertase, and that results in a complement-mediated MPGN or the so-called C3 glomerulopathy.
Pathophysiology of MPGN: Keeping it Simple
So for my final slide, keeping MPGN simple. If you see immune complexes on a biopsy, you think of an immune complex-mediated MPGN. Immune complexes typically can come from 1 of 3 areas. So the patient needs to be worked up for infections, particularly chronic infections. Number 2: patient needs to be worked up for autoimmune diseases, lupus or whatever, arthritis, Sjögren’s disease, Hashimoto’s, you name it, or the patient needs to be worked up for a monoclonal gammopathy. These are the 3 major groups of disease that result in immune complex MPGN. And you can clearly see that the treatment for infections vs. autoimmune disease vs. monoclonal gammopathy is completely different. On the other hand, MPGN that does not show you much immunoglobulins, but shows you a lot of complement deposits, typically means this is a complement-mediated MPGN. We can subclassify it into C3 glomerulonephritis and dense-deposit disease based on EM. The exact significant of subclassifying, as yet, is not known. And this results from alternative pathway abnormalities and as you can clearly see, some patients may have an autoantibody, some patients may have a mutation, and obviously, the treatment for autoimmune antibody is likely to be different from those having an underlying mutation in the alternative pathway of complement proteins. Is there an idiopathic subgroup? Probably, 5% to 10% of these patients with MPGN might not fit into either of these particular groups.
These are some of the key references of C3 glomerulopathy and MPGN.