Causes and consequences of proteinuria after renal transplantation

INTRODUCTION

The presence of proteinuria is a frequent finding after renal transplantation, and affects 30-45% of patients per year 1 . Traditionally, the literature on proteinuria after transplantation discussed the differential diagnosis of high-level proteinuria and its relation to patient graft survival 2 . In the last five years we have learned that proteinuria, at all levels, is an important biological marker that identifies grafts and patients with poor prognosis. These studies suggest two questions that we will try to answer in this review: What are the causes of low and high level proteinuria after transplantation? And why is the relationship between proteinuria and the reduction in survival of both the graft and the patient?

PREVALENCE

The prevalence of proteinuria varies between 15 and 45% in different studies, and this variation is mainly due to differences in the level of proteinuria used to define the value considered as abnormal 1 and at the time proteinuria has been determined . We believe that it is important to diagnose proteinuria during the first months after transplantation, which allows us to identify grafts and patients at high risk. For this purpose, we showed in Figure 1 the prevalence of proteinuria at one year of transplantation in live donor and corpse kidney recipients. As can be observed, there are no significant differences in the prevalence or level of proteinuria between these two groups.

Data on the prevalence of albuminuria after transplantation are scarcer. In studies conducted at our service, we demonstrated that albuminuria is common and affects most patients with proteinuria, even of low level. For example, more than 80% of patients with proteinuria between 150 and 500 mg / day and 100% of patients with higher levels of proteinuria have albuminuria 3 . Among patients without proteinuria, approximately 15% have albuminuria above 30 mg / day 3 .

CAUSES OF PROTEINURIA

Proteinuria in renal transplantation may be due to multiple causes (Table 1). Patients with high proteinuria (> 1,500 mg / day) frequently have glomerulopathy in the graft (in 80% of cases) 3 . However, in patients with lower levels, establishing the cause of proteinuria can be difficult. We will briefly consider the different causes of proteinuria.

Residual Proteinuria

The presence of proteinuria from the native kidneys may complicate the interpretation of proteinuria detected after transplantation. This is common in patients who receive an early renal transplant or shortly after initiating dialysis and, therefore, with a significant renal function and residual diuresis 4 . Results of two studies provide us with practical guidelines to aid in the interpretation of proteinuria in these patients. First, these studies demonstrate that pretransplantation proteinuria, even when it is of nephrotic range, decreases abruptly during the first weeks after receiving a normal transplant renal transplant 4,5 . This decrease is probably due to the decrease in blood flow that occurs in the native kidneys after a transplant, if the graft has a good function 5 . This latter point is important because, in our experience, in recipients with poor initial graft function the blood flow in the native kidneys is maintained and 'native' proteinuria persists. A second study helps us in the interpretation of proteinuria 4 . First, in patients with normal-graft graft, the presence of proteinuria greater than 3,000 mg / day at three weeks after transplantation should not be attributed to the native kidneys but indicates the presence of glomerular disease in the graft (probably a recurrence of one Glomerulopathy). Second, proteinuria greater than 1.  500 mg / day per year of the transplant and / or an increase in proteinuria from the third week to the year post-transplant in more than 500 mg / day indicate a pathology in the graft. Third, the native kidneys may have low levels of proteinuria (less than 500 mg / day) even one year after transplantation, although proteinuria is expected to decrease with time.

Glomerular graft diseases

In a previous study we assessed protocol biopsies in patients with proteinuria year after transplantation 3 . Only 9% of these patients had glomerulopathy. However, among patients with proteinuria levels of more than 1,500 mg / day, 80% had evidence of glomerular disease. Other studies support these results 2 . We must consider three types of glomerulopathy in the graft: recurrent disease, de novo disease and transplant glomerulopathy.

Table 2 summarizes the prevalence and consequences for graft recurrence of the most common glomerulopathies. It emphasizes that in most studies, the diagnosis of recurrence has been based on the presence of proteinuria. Segmental and focal glomerulosclerosis (GSF) is a disease with a high recurrence risk, affecting approximately 30% of patients. Previous studies have identified subgroups of patients with GSF that have a much higher risk and include: patients diagnosed before age 18, patients with rapid progression (less than three years) of disease 6  and, in particular, patients with History of recurrences in previous transplants. Approximately 50% of patients with GSF recurrence lose the graft. Membranous nephropathy (NM) is also associated with a high risk of recurrence. Protocol biopsy studies demonstrated that histologic recurrence of NM occurs generally during the first months after transplantation in 40% of cases and that, initially, histologic changes do not cause proteinuria 7 . IgA nephropathy frequently occurs (> 50%) after transplantation, although histological changes are usually mild and consist of the presence of IgA deposits detected by immunofluorescence and small mesangial deposits detected by electron microscopy, but without proliferation of mesangial cells or clinical manifestations. It is rare that recurrent IgA nephropathy causes loss of graft, Although we have observed rare cases with an aggressive clinical behavior accompanied by a high level of proteinuria. Mesangiocapillar (or membranoproliferative, GNMP) glomerulopathy has acquired great interest lately due to new data that allow us to distinguish several subtypes of this disease 8,9 . From the point of view of recurrence, distinguishing between these types of GNMP has important clinical consequences. For example, in cases of GNMP associated with monoclonal proteins, the recurrence of the disease can occur rapidly and has an aggressive clinical presentation 10 . Patients with type I MPAG associated with hypocomplementemia also have a high risk of recurrence 10,11 . In general,

Protocol biopsy studies revealed that, in many cases, the recurrence of glomerulopathy is not associated with proteinuria 7,10 . This is of interest for a number of reasons: Firstly, the histological diagnosis by protocol biopsies gives us for the first time information on when the recurrence occurs and also, for the first time, gives us information about the histological changes that occur during the phases Initials of these diseases. Secondly, it is reasonable to think that the treatment of these recurrences in their initial stages will be more effective than in more advanced stages. Third, we must remember that low-level proteinuria may be the first manifestation of a recurrence of native glomerulopathy, Which may have negative implications for long-term grafting. Therefore, it is important to investigate patients with low-level proteinuria and follow them closely, through periodic determinations and performing a biopsy if a progressive increase in proteinuria is demonstrated.

In some cases, the graft develops glomerulopathy that is not recurrent but has the histological features of native kidney glomerulopathy. These glomerulopathies are generally diagnosed later than recurrent and may result in graft loss 13 . The most common histological types include GSF 14 , NM and mesangiocapillar. They are infrequent diseases of poorly defined etiology.

Transplant glomerulopathy (GT) has been recognized for many years as a disease that is generally diagnosed several years after transplantation and can cause high-level proteinuria, even of nephrotic range 15,16 . In recent years we have learned that this disease, in most cases, is due to damage of the capillaries produced by anti-HLA class II antibodies 17 . The use of protocol biopsies and the ability to effectively measure these antibodies have allowed us to recognize that, in many patients, WG is developed during the first months after transplantation and that its clinical presentation frequently consists of progressive loss of function Graft with severe hypertension and low level proteinuria.

In more recent studies (presented at the American Transplant Congress, ATC, 2011), we have shown that proteinuria is an early marker of damage to the glomerular capillaries by anti-HLA II antibodies, before the histological manifestations of GT were visible . The presence of proteinuria among all patients with anti-HLA II antibodies identifies a subgroup of patients with a high incidence of glomerulitis and a high risk of developing GT in the future. Studies performed with an electron microscope showed that damage to the endothelial cells of the glomerular capillary precedes the histological changes that we consider to be WG diagnoses 18 .

Proteinuria associated with tubular and interstitial damage

Published studies many years ago proteinuria attributed to "chronic transplant rejection ' 15 . In most of these cases, the biopsy showed GT changes. Biopsy studies in patients with proteinuria 3 demonstrated that there is a subgroup of patients with chronic transplant nephropathy (interstitial fibrosis and tubular atrophy 19 ), without glomerular disease, but with low levels of proteinuria. Frequently, these patients have albuminuria 3 and, therefore , they may have occult glomerular disease. At the same time, the proximal tubule reabsorbs large amounts of albumin 20 , making it possible that the proteinuria in these patients is due to tubular damage.

Proteinuria secondary to the use of m-TOR inhibitor

The use of sirolimus or everolimus has been associated with the development of proteinuria in renal transplantation in numerous studies 1,21,22 . Initially, this observation occurred in patients with chronic graft nephropathy after switching from calcineurin inhibitors (ICN) to sirolimus with the intention of preserving renal function. For this reason it was suggested that proteinuria associated with sirolimus was the result of the hemodynamic effect secondary to CNI. At this time, the evidence indicates that m-TOR inhibitors have direct effects on protein filtration in the glomerulus and specifically on the podocyte. Letavernier, et al. 23 demonstrated that sirolimus affects the synthesis of VEGF, which is essential for podocyte and endothelial cell survival, As well as for intracellular Akt signaling, which is critical for differentiation, adhesion and survival of epithelial cells. More recent studies have shown a decrease in the expression of constituent proteins of the diaphragmatic cleft, an essential structure for the selective ultrafiltration of the glomerulus 24,25 .

Clinical studies indicate that the presence of proteinuria in a patient treated with sirolimus may have negative consequences for the graft. In studies of conversion of ICN to sirolimus it was found that specifically in patients with proteinuria, conversion to sirolimus had negative effects on graft survival 26 . In isolated cases 27 the development of GSF has been detected in grafts treated with sirolimus. In particular, in patients with incipient glomerulopathies (mainly recurrent IgA), m-TOR inhibitor drugs may produce a significant increase in proteinuria and impairment of renal function. Conversely, There is no clear evidence that sirolimus impairs grafting in patients who maintain a low and stable level of proteinuria (<500 mg / day). However, these patients should be followed closely, measuring proteinuria periodically and, if this increase progressively, the suspension of the m-TOR inhibitor should be considered.

Proteinuria related to donor and recipient factors

Interestingly, among all patients with proteinuria at one year of transplantation, 51% have a clear cause; Include glomerulopathy on biopsy, sirolimus or anti-HLA class II antibodies. However, 49% of patients with proteinuria do not present any of these factors. This observation led us to explore other factors that may be related to proteinuria (table 3) (abstract presented in ATC, 2011) 1 .

It is interesting to consider the relationship between proteinuria and donor and recipient demographic factors 3 . Proteinuria is more common and more abundant in transplants from smaller or lesser-functioning donors (major donors, female donors, relatively lower donors), and in larger recipient transplants (male recipients with a higher body mass index [ BMI]). These data suggest that the difference in size / function between the donor and the recipient determines, in part, the risk of proteinuria. Physiologically, it is possible that this difference in size causes glomerular hyperfiltration, which may lead to proteinuria and progressive deterioration of renal function 28,29 . A possible example of this model is shown in figure 2, In which it is emphasized that the percentage of patients with proteinuria increases with the age of the donor. At the same time, in each donor group defined by age, the risk of proteinuria increases progressively the higher the weight of the recipient.

RELATIONSHIP BETWEEN PROTEINURIA AND INJERENCE SURVIVAL

Interest in proteinuria after transplantation is basically due to its relationship with graft survival. In general, when the level of proteinuria increases, graft survival decreases (figure 3). In this figure, the risk of graft loss progressively increases with the level of proteinuria, and this risk is noticeable even in patients who are considered to have a low proteinuria level (<500 mg / day). In previous studies 3, we calculated that, compared to grafts without proteinuria, the risk of graft loss with proteinuria levels between 150 and 500 mg / day increased to 2.45 times, to 6.07 times in patients with proteinuria between 501 and 1,500 mg / day and to 14.3 times with proteinuria levels above 1,500 mg / day.

What is the relationship between the level of proteinuria and the survival of the graft? Our proposal is that proteinuria is an indicator of a series of aggressions that can affect the graft and that these are mainly, and not proteinuria, that cause the loss of the transplant. For example, it is very likely that glomerulopathy is the leading cause of graft loss in patients with high levels of proteinuria due to glomerular disease. It is also possible that in these patients the presence of high levels of proteinuria can produce interstitial damage to the graft 30 . It is much less clear why grafts with low levels of proteinuria also have a compromised survival 31,32 and it is in this case that, again, We have to consider the cause of proteinuria (Table 1 and Table 3). Low levels of proteinuria may indicate: 1) the first stages of recurrence of glomerulopathy; 2) the effect of anti-HLA class II antibodies, or 3) the existence of a significant disproportion between graft function and recipient needs. In each of these cases proteinuria is related to a reduction in graft survival, but the mechanism of graft damage is different. There are two forms of low-level proteinuria, which are probably not associated with an unfavorable prognosis: residual proteinuria and proteinuria induced by m-TOR inhibitors. Even in these cases, We have to take precautions and follow the patient closely to confirm that it does not increase during follow-up. If this occurs, we can rule out the possibility that proteinuria is residual 4 and we must look for other causes.

To understand the relationship between proteinuria and graft survival we must also consider that the presence of proteinuria is associated with other characteristics of the donor, recipient and graft 3 . For example, proteinuria is associated with decreased renal function, and although it is important to consider this relationship, statistically the relationship between proteinuria and graft survival is independent of graft function 3 .

RELATIONSHIP BETWEEN PROTEINURIA AND PATIENT SURVIVAL

Proteinuria in renal transplantation is associated with a reduction in patient survival. Several studies already published have shown this relationship 33-35 . Specifically, other studies showed that patients with proteinuria have a risk of increased cardiovascular 2.45 times as compared to patients without proteinuria 36 . In more recent studies in our group (abstract presented in ATC, 2011) we confirmed the relationship between a progressive increase in proteinuria and a decrease in patient survival and we began a study of the factors that may explain or contribute to this relationship Figure 4). This figure highlights that even low levels of proteinuria, less than 500 mg / day,

We must consider three types of factors to try to explain the relationship between proteinuria and patient survival: first, it is possible that the factors that cause proteinuria also increase the risk of the patient. An examination of these factors (Table 3) does not suggest that this is a plausible explanation and, in fact, in preliminary studies a relationship between factors related to proteinuria and patient survival has not been found. Second, it is possible that patients with proteinuria may acquire or have other factors that may be related to patient survival 3 . We have recently expanded these studies (ATC, 2011) and, indeed, Proteinuria is related to other biochemical parameters (elevated lipids and decreased albumin and hemoglobin), blood pressure and decreased graft function. All these variables are related to patient survival. Thirdly, it is well known that, in the general population, microalbuminuria is related to a higher mortality from all causes, including cardiovascular causes 37,38 . This relationship is generally attributed to an association between albuminuria / proteinuria and alterations in endothelial function and / or inflammation. Recently, in a retrospective study it was observed that microalbuminuria is related to the survival of renal transplant patients 39 . In this study, what has been surprising is that albuminuria was related not only to an increase in cardiovascular risk but also to an increase in the risk of cancer mortality. Albuminuria is very common after transplantation, so it is important to confirm these relationships and to study factors that may explain the mechanisms underlying the relationship between proteinuria and renal transplant survivorship.

TREATMENT

Treatment of a patient with proteinuria should include three aspects: 1) specific treatment of the cause of proteinuria; 2) reduction of proteinuria using non-specific treatments, and 3) treatment of cardiovascular risk. Treatment of proteinuria, particularly of a low level, is often limited to control of blood pressure or the use of angiotensin converting enzyme (ACE) inhibitors, which may reduce the concentration of protein in the urine. Although these maneuvers are probably useful, in our opinion it is a mistake not to try to investigate the possible cause of proteinuria. Establishing such a cause not only can give us prognostic information about the graft,

Specific Treatments

In this review we can not discuss in detail all the therapies we can use to treat the multiple causes of proteinuria in transplantation (Table 1). In general, the treatment of recurrent glomerulopathies follows the same guidelines used in the treatment of these diseases in the native kidney. However, when we treat these diseases in the transplanted patient we must consider several particular factors:

1. The use of antirejection drugs may modify the behavior of certain glomerulopathies in the graft. For example, if we consider the low prevalence of recurrence of diseases such as lupus erythematosus. In addition, antirejection drugs may modify the effects of other drugs. For example, anti-CD20 antibodies are known to cause B lymphocyte depletion over a much longer period in transplant patients than in other patients 40,41 . In our experience, cyclophosphamide may be used in the treatment of aggressive acute graft glomerulopathies, but its use should be avoided at the same time as azathioprine or mycophenolate, given the high risk of leukopenia.

2. Patients who received a transplant obviously had glomerulopathy in the native kidney. Therefore, it is possible that in transplant patients these diseases may be especially aggressive. This, of course, is not applicable to all patients and, for example, the recurrence of IgA nephropathy is generally mild. However, other studies have suggested that the aggressiveness of the disease in the native kidney is reproduced in the graft, for example in the case of GSF 42 . We also detected a low spontaneous remission of NM in the graft: among 34 patients diagnosed of recurrent NM by biopsies per protocol and with minimal clinical manifestations, 33 have presented a progressive increase of proteinuria and, in those cases with follow-up biopsies, A histological worsening of nephropathy. These considerations have implications for deciding when to treat the patient with recurrence.

3. In the graft, glomerulopathies can be diagnosed in the initial stages because in these patients proteinuria is measured periodically 43 . In our service, the use of biopsies by protocol allows us to diagnose these diseases frequently when there are no clinical manifestations. Perhaps for this reason certain treatments are more effective in grafting than in the native kidney. For example, plasmapheresis may be effective in the treatment of certain cases of GSF 44 and GNMP 10.45 . The early diagnosis of these diseases can theoretically improve their response to treatment, as we have seen, in the case of NM 40 .

In patients with proteinuria, it is useful to measure the level of anti-HLA antibodies for several reasons: 1) most patients with transplant glomerulopathy have or have had anti-HLA class II antibodies, so their presence is diagnostic; 2) the existence of anti-HLA II antibodies and proteinuria is an early index of capillary damage by antibodies and is associated with reduced graft survival; 3) the level of anti-HLA class II antibodies is related to graft survival and this relationship is independent of other factors such as biopsy markers (mainly the presence of C4d in capillaries) and biochemical markers (proteinuria and renal function) 46 ; 4) the appearance of new anti-HLA antibodies indicates that immunosuppression is not effective, either because the patient is not taking his medication regularly, or because the doses indicated are not sufficient; 5) in patients who develop anti-HLA antibodies after transplantation it is sometimes possible to suppress the level of antibodies with higher doses of immunosuppressants and our recommendation is to increase the dose of mycophenolate and / or tacrolimus for a limited period of approximately six months, Periodically measuring serum antibody levels.

In patients receiving m-TOR inhibitors it is essential to monitor proteinuria periodically and if this increases significantly we recommend stopping the drug. Sirolimus can dramatically increase proteinuria due to glomerulopathy, even when it is mild. In our experience, discontinuation of this medication results in an improvement in proteinuria over a period of a few months. In patients with a high level of proteinuria, sirolimus can produce acute renal failure 47 . Finally, it is important to mention that in patients with proteinuria a change from ICN to m-TOR inhibitors can be harmful 26 . 

As discussed earlier, we have detected that approximately 50% of patients with transplant proteinuria may be related to donor or recipient factors indicative of a significant discrepancy between the limited functional capacity of the graft to respond to the recipient's demands. Possibly, in these cases proteinuria is due to glomerular hypertension and glomerular hyperfiltration and, if this is the case, the inhibition of the renin-angiotensin system (RAS) should be considered as a specific treatment for the cause of proteinuria 48 .

Non-specific treatment of proteinuria in renal transplantation

Based on the results of studies of proteinuria control in chronic kidney disease, it seems reasonable to recommend the following non-specific measures in patients undergoing transplantation: 1) blood pressure control (systolic blood pressure less than 130 mmHg); 2)  use of ACE inhibitors and / or angiotensin II receptor antagonists (AIIRAs) at the highest tolerated dose, even if there is no hypertension; 3) lipid profile control, preferably with statins; 4) maintain a proper BMI; 5) protein restriction in the diet, and 6) smoking cessation 49 .

First, we must insist that, in patients with transplanted kidney problems, these measures reduce the progression of kidney failure, but in general, do not prevent their appearance 50 . Therefore, we must consider these non-specific measures as a treatment to be added to the specific treatment. Second, although inhibition of RAS reduces transplant proteinuria, there is insufficient evidence to improve graft survival. Thirdly, it is important to emphasize that the use of ACE inhibitors or ARBs in patients undergoing transplantation can have important negative consequences; Therefore, special precautions should be taken, including the use of low doses and monitoring of creatinine,

A systematic review 51 of 21 randomized studies on the use of ACE inhibitors and / or AIIRAs and including 1,549 transplant patients concluded that these drugs produce a clinically significant reduction of proteinuria but also hematocrit and hematocrit. Glomerular filtration rate. In this study, no significant differences were observed in blood pressure control or graft survival. However, the studies included in this review have limitations due to the small sample size, the reduced follow-up time and the start of treatment at different times after transplantation. A prospective, randomized, Controlled and multicenter study evaluated the effect of an ARI II (candesartan) on graft survival and cardiovascular morbidity and mortality in 502 renal transplant patients 52 . This study was earlier than expected because of a lower-than-expected rate of events but it was observed that patients treated with candesartan had a significant decrease in proteinuria and a better control of blood pressure compared to patients receiving placebo . Two observational and retrospective studies attempted to elucidate the effect of inhibition of RAS on graft survival, but with conflicting results. In the first of them 53 , A significant beneficial effect on the survival of both the graft and the patient was found to be independent of the presence of proteinuria. In contrast, the second of these studies 54 concluded that inhibition of RAS has no beneficial effects either on graft survival or on patient survival, even in subgroups of patients at high cardiovascular risk. In summary, with the data available today it would be premature to recommend treatment with ACEI and / or ARI II in order to improve the survival of the graft or the patient in the renal transplant, since this effect is still unknown. The second of these studies 54 concluded that the inhibition of RAS has no beneficial effects either on graft survival or on patient survival, even in subgroups of patients with high cardiovascular risk. In summary, with the data available today it would be premature to recommend treatment with ACE inhibitors and / or ARAII in order to improve the survival of the graft or the patient in the renal transplant, since this effect is still unknown. The second of these studies 54 concluded that the inhibition of RAS has no beneficial effects either on graft survival or on patient survival, even in subgroups of patients with high cardiovascular risk. In summary, with the data available today it would be premature to recommend treatment with ACEI and / or ARI II in order to improve the survival of the graft or the patient in the renal transplant, since this effect is still unknown.

Cardioprotection
The presence of proteinuria, even low - level, primarily identifies patients with reduced survival, but not only for increased cardiovascular risk 34,36 . The mechanism that causes this increased risk is not currently known and we do not have enough data (as we have seen in the previous section) to conclude whether the reduction of proteinuria or the use of RAS inhibitors improves the prognosis of these patients. However, it is reasonable to intensify cardioprotective measures in patients with proteinuria; These measures should include: blood pressure monitoring (less than 130 mmHg); Use of statins by maintaining the LDL cholesterol level below 100 mg / dl (less than 80 mg / dl in high-risk patients 55 ); Strict glycemic control in patients with diabetes (measure debatable on the basis of recent studies 56 ); Avoid tobacco use; Use of aspirin, and use of beta-blockers. Among these measures, we have only evidence, based on prospective, controlled and randomized studies, that the use of statins is effective in transplant patients 57 .

KEY CONCEPTS

1. Proteinuria after transplantation is common and may be due to multiple causes.

2. Proteinuria, even when low level (<500 mg / day), is a sensitive indicator of reduced graft survival.

3. The reduction in the survival of the graft with proteinuria is probably due to the same problem that causes proteinuria. Therefore, it is critical to investigate the cause of proteinuria after transplantation.

4. Proteinuria, even when low level (<500 mg / day), is also associated with a reduction in patient survival mainly due to an increase in cardiovascular risk.

5. The treatment of proteinuria should consider three aspects: the cause, the reduction with non-specific measures and the reduction of the cardiovascular risk associated with it.