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. Author manuscript; available in PMC: 2020 Jul 30.
Published in final edited form as: N Engl J Med. 2020 Jan 30;382(5):416–426. doi: 10.1056/NEJMoa1911481

Soluble Urokinase Receptor and Acute Kidney Injury

Salim S Hayek 1,#, David E Leaf 1,#, Ayman Samman Tahhan 1,#, Mohamad Raad 1, Shreyak Sharma 1, Sushrut S Waikar 1, Sanja Sever 1, Alex Camacho 1, Xuexiang Wang 1, Ranadheer R Dande 1, Nasrien E Ibrahim 1, Rebecca M Baron 1, Mehmet M Altintas 1, Changli Wei 1, David Sheikh-Hamad 1, Jenny S-C Pan 1, Michael W Holliday Jr 1, James L Januzzi 1, Steven D Weisbord 1, Arshed A Quyyumi 1, Jochen Reiser 1
PMCID: PMC7065830  NIHMSID: NIHMS1565140  PMID: 31995687

Abstract

BACKGROUND

Acute kidney injury is common, with a major effect on morbidity and health care utilization. Soluble urokinase plasminogen activator receptor (suPAR) is a signaling glycoprotein thought to be involved in the pathogenesis of kidney disease. We investigated whether a high level of suPAR predisposed patients to acute kidney injury in multiple clinical contexts, and we used experimental models to identify mechanisms by which suPAR acts and to assess it as a therapeutic target.

METHODS

We measured plasma levels of suPAR preprocedurally in patients who underwent coronary angiography and patients who underwent cardiac surgery and at the time of admission to the intensive care unit in critically ill patients. We assessed the risk of acute kidney injury at 7 days as the primary outcome and acute kidney injury or death at 90 days as a secondary outcome, according to quartile of suPAR level. In experimental studies, we used a monoclonal antibody to urokinase plasminogen activator receptor (uPAR) as a therapeutic strategy to attenuate acute kidney injury in transgenic mice receiving contrast material. We also assessed cellular bioenergetics and generation of reactive oxygen species in human kidney proximal tubular (HK-2) cells that were exposed to recombinant suPAR.

RESULTS

The suPAR level was assessed in 3827 patients who were undergoing coronary angiography, 250 who were undergoing cardiac surgery, and 692 who were critically ill. Acute kidney injury developed in 318 patients (8%) who had undergone coronary angiography. The highest suPAR quartile (vs. the lowest) had an adjusted odds ratio of 2.66 (95% confidence interval [CI], 1.77 to 3.99) for acute kidney injury and 2.29 (95% CI, 1.71 to 3.06) for acute kidney injury or death at 90 days. Findings were similar in the surgical and critically ill cohorts. The suPAR-overexpressing mice that were given contrast material had greater functional and histologic evidence of acute kidney injury than wild-type mice. The suPAR-treated HK-2 cells showed heightened energetic demand and mitochondrial superoxide generation. Pretreatment with a uPAR monoclonal antibody attenuated kidney injury in suPAR-overexpressing mice and normalized bioenergetic changes in HK-2 cells.

CONCLUSIONS

High suPAR levels were associated with acute kidney injury in various clinical and experimental contexts. (Funded by the National Institutes of Health and others.)

The incidence of acute kidney injury is increasing globally. Acute kidney injury occurs in 2 to 5% of hospitalized adults and has a major effect on morbidity and health care utilization.1–4 The largest burden of acute kidney injury occurs in critically ill patients and in persons with cardiovascular disease, who are at increased risk for both acute kidney injury and chronic kidney disease owing to their older age and multiple coexisting conditions, as well as their greater likelihood of undergoing procedures that may directly affect the kidneys, such coronary angiography or cardiac surgery.4–6 Despite recent gains in our understanding of the causes and underlying mechanisms of acute kidney injury, few therapeutic or preventive options exist.7 Thus, uncovering new therapeutic targets for the prevention of acute kidney injury is of importance.

Inflammation and oxidative stress are central components of the pathogenesis of acute kidney injury, implicating multiple subtypes of immune cells.8,9 Evidence of a pathway linking the bone marrow to kidney injury has emerged, involving soluble urokinase plasminogen activator receptor (suPAR)7,10–17 — the circulating form of a glycosylphosphatidylinositol–anchored three-domain membrane protein. This receptor is normally expressed at very low levels on a variety of cells, including endothelial cells, podocytes, and, with induced expression, immunologically active cells such as monocytes and lymphocytes.11,16,18 Levels of suPAR are strongly predictive of progressive decline in kidney function.17,19–23 Long-term exposure to elevated suPAR levels directly affects the kidneys by means of pathologic activation of αvβ3 integrin expressed in podocytes, resulting in proteinuria.7,12,16,24 Whether suPAR has an effect on kidney tubular cells — the cells most affected in acute kidney injury — is unclear.

We investigated whether a high level of suPAR was associated with acute kidney injury in patients undergoing coronary angiography and sought to replicate the findings in two other clinical contexts in which patients are at high risk for acute kidney injury: cardiac surgery and critical illness. We then used experimental models to determine whether the overexpression of suPAR led to worsening of renal function and assessed the potential for prevention of acute kidney injury by means of pharmacologic inhibition of suPAR.

METHODS

ACUTE KIDNEY INJURY AND SUPAR

We evaluated the association between suPAR levels and postprocedural acute kidney injury in two prospective cohorts of patients undergoing coronary angiography for suspected coronary artery disease: the Emory Cardiovascular Biobank (EmCAB) and the Catheter Sampled Blood Archive in Cardiovascular Diseases (CASABLANCA). To determine whether suPAR was associated with acute kidney injury unrelated to the use of contrast material we sought to replicate our findings in patients at high risk for acute kidney injury who were undergoing cardiac surgery and in critically ill patients who had been admitted to the intensive care unit (ICU).25–27

CORONARY ANGIOGRAPHY COHORTS

EmCAB and CASABLANCA are prospective observational cohorts consisting of adult patients (≥18 years of age) undergoing coronary angiography for suspected ischemic heart disease.25,26 EmCAB enrolled patients at three Emory Healthcare sites in Atlanta between 2003 and 2015, and CASABLANCA enrolled patients at Massachusetts General Hospital in Boston between 2008 and 2011. EmCAB excluded patients with congenital heart disease, severe anemia, a recent blood transfusion, myocarditis, or a history of active inflammatory disease or cancer. The only exclusion criterion in CASABLANCA was an unwillingness to participate.

Patients without end-stage kidney disease who had a serum creatinine–based measurement of kidney function at baseline and at least one measurement obtained within 7 days after angiography were included in this analysis. We measured suPAR in blood samples that were obtained at the time of the procedure before the injection of contrast material. Both studies were approved by the institutional review board at the respective institutions.

The authors had full access to the data and take responsibility for the completeness and accuracy of the data and for the integrity of the analysis. All the participants provided written informed consent at the time of enrollment. Details of the cardiac surgery cohort and the ICU cohort are provided in Section I in the Supplementary Appendix, available with the full text of this article at NEJM.org.

MEASUREMENT OF KIDNEY FUNCTION AND DEFINITION OF ACUTE KIDNEY INJURY

Measurements of serum creatinine at enrollment and all subsequent values obtained during the index hospitalization were obtained from electronic medical records. The estimated glomerular filtration rate (eGFR) was calculated with the use of the Chronic Kidney Disease Epidemiology Collaboration equation.28 Acute kidney injury was defined according to the Kidney Disease: Improving Global Outcomes (KDIGO) Work Group criteria as an absolute increase in the creatinine level of at least 0.3 mg per deciliter (30 μmol per liter) within the first 48 hours after the procedure or ICU admission, a relative increase of at least 50% in the creatinine level within the first 7 days after the procedure or ICU admission, or use of dialysis.29 The creatinine measurement obtained immediately before the procedure or on admission to the ICU was used as the baseline value for all analyses.

SAMPLE COLLECTION AND MEASUREMENT OF SUPAR

Blood samples were obtained as described, and plasma was stored at −80°C in EDTA-coated tubes. Experienced technicians who were unaware of the clinical data measured suPAR in plasma using a commercially available enzyme-linked immunosorbent assay (ViroGates). The lower limit of detection was 100 pg per milliliter. The interassay coefficient of variation, which was determined with the use of blinded replicate samples obtained from study patients, was 10.9%. We and others have found that suPAR levels are stable in stored plasma and serum samples and that levels are reproducible in samples that have been stored for more than 5 years at −80°C despite exposure to multiple freeze–thaw cycles.14,30

ANIMAL MODEL OF ACUTE KIDNEY INJURY

We used C57BL/6J wild-type mice and transgenic mice overexpressing suPAR in an animal model of contrast-induced nephropathy to study the effect of suPAR on kidney function.12,17 The median suPAR level in the suPAR-transgenic mice at 10 weeks of age was 210 ng per milliliter (interquartile range, 160 to 256). We injected iohexol intraperitoneally in suPAR-transgenic mice (20 mice, 9 of which were male) and in wild-type controls (16 mice, 8 of which were male) following published protocols.31 The mice were also randomly assigned to receive an intraperitoneal injection of either urokinase plasminogen activator receptor (uPAR)–blocking monoclonal antibody or the same concentration of IgG isotype. Serum creatinine and kidney histologic tests were used to assess the severity of acute kidney injury. Experimental details are provided in the Supplementary Appendix.14,16,32,33 The study was approved by the Institutional Animal Care and Use Committee of Rush University in Chicago.

EFFECT OF SUPAR ON KIDNEY TUBULAR CELL BIOENERGETICS

We quantified the generation of reactive oxygen species and cellular bioenergetics of human kidney proximal tubular (HK-2) cells that were exposed to recombinant suPAR at a concentration of 10 ng per milliliter, with or without anti-uPAR antibody, using the MitoSOX (Invitrogen) and Seahorse Extracellular Flux Analyzer (Agilent), respectively. Details of the experiments are provided in the Supplementary Appendix.34

STATISTICAL ANALYSIS

Continuous variables are presented as means (±SD) or as medians (with interquartile ranges) for normally and nonnormally distributed data, respectively. Categorical variables are presented as percentages. To compare patients across quartiles of suPAR level, we used analysis of variance or the Kruskal–Wallis test for continuous variables and chi-square tests for categorical variables.

We used logistic regression to characterize the association between suPAR levels and acute kidney injury at 7 days as the primary outcome and acute kidney injury or death at 90 days as a secondary outcome. We assessed suPAR levels both as a continuous variable (natural log–transformed) and as quartiles, with the lowest quartile serving as the reference group. We adjusted for covariates using three models. In all the cohorts, model 1 was unadjusted. In the coronary angiography cohort, model 2 was adjusted for age, sex, race, smoking history, diabetes mellitus, congestive heart failure, hypertension, and cohort (EmCAB or CASABLANCA); model 3 incorporated the aforementioned variables in addition to acute myocardial infarction, revascularization, volume of contrast material, and baseline eGFR. In the cardiac surgery cohort, model 2 was adjusted for age, sex, race, diabetes mellitus, congestive heart failure, and preoperative eGFR; model 3 incorporated the aforementioned variables in addition to urgent procedure and cardiopulmonary-bypass time of more than 120 minutes. In the ICU cohort of critically ill patients, model 2 was adjusted for age, sex, race, baseline eGFR, diabetes mellitus, congestive heart failure, chronic lung disease, and chronic liver disease; model 3 was further adjusted for vasopressors received during the first 24 hours of ICU admission, mechanical ventilation during the first 24 hours of ICU admission, and the hemoglobin level and white-cell count at ICU admission.

To investigate the possibility of effect modification attributed to differences in baseline characteristics, we computed odds ratios for the association between suPAR levels and acute kidney injury in relevant subgroups and performed tests of interaction. Finally, we calculated the area under the curve (AUC) to assess the incremental value of adding suPAR to the Simplified Integer Risk Score for Calculating the Risk of Acute Kidney Injury, a validated clinical score derived from the National Cardiovascular Data Registry (NCDR) and used to predict the risk of contrast-induced nephropathy; the score includes age, preprocedural eGFR, history of stroke, history of heart failure, history of percutaneous coronary intervention, acute coronary syndrome on presentation, diabetes, chronic lung disease, hypertension, cardiac arrest, anemia, heart failure at presentation, balloon-pump use, and cardiogenic shock.35

For the experiments in animals, we used a two-way analysis of variance and post hoc tests (least significant differences) to compare creatinine levels and kidney injury scores between suPAR-transgenic mice and wild-type mice and between the mice that received IgG isotype and those that received uPAR monoclonal antibody. Two-tailed P values of 0.05 or less were considered to indicate statistical significance. All the analyses were performed with the use of SPSS software, version 24 (IBM).

RESULTS

BASELINE CHARACTERISTICS OF PATIENTS AND DETERMINANTS OF ACUTE KIDNEY INJURY

The study included 3827 patients undergoing coronary angiography: 2752 from the EmCAB cohort, and 1075 from the CASABLANCA cohort. Summary statistics for each of these cohorts are reported in Table S1 in the Supplementary Appendix.

Postprocedural acute kidney injury developed in 318 patients (8%). The mean increase in the creatinine level was 0.44±0.54 mg per deciliter (39±50 μmol per liter) among patients with acute kidney injury, as compared with 0.01±0.22 mg per deciliter (1±20 μmol per liter) among those without acute kidney injury. The majority of cases of acute kidney injury within 7 days after angiography were mild (98% of the cases were of KDIGO stage 1), with 28 patients having KDIGO stage 2 acute kidney injury and 3 patients having KDIGO stage 3 acute kidney injury. Patients with acute kidney injury after coronary angiography were more likely than those without acute kidney injury to be older, to have diabetes mellitus, to have a history of heart failure, to have a higher suPAR level, to have a lower baseline eGFR, to have received a lower volume of contrast material, and to have undergone percutaneous coronary intervention at the time of angiography (Table 1). In multivariable analysis, only diabetes mellitus, history of heart failure, lower eGFR, and higher suPAR levels were independently associated with acute kidney injury.

Table 1.

Demographic and Clinical Characteristics of Patients Who Underwent Coronary Angiography, Stratified According to Incidence of Postprocedural Acute Kidney Injury.*

Characteristic No Acute Kidney Injury (N = 3509) Acute Kidney Injury (N = 318) P Value
Age — yr 66±12 68±12 <0.001
Male sex — no. (%) 2413 (69) 224 (70) 0.54
Black race — no. (%)† 467 (13) 37 (12) 0.40
Body-mass index‡ 29±6 30±7 0.26
Smoking — no. (%) 2314 (66) 196 (62) 0.12
Type 2 diabetes mellitus — no. (%) 1206 (34) 139 (44) 0.001
Hypertension — no. (%) 2783 (79) 261 (82) 0.24
History of myocardial infarction — no. (%) 959 (27) 87 (27) 0.99
History of heart failure — no. (%) 1147 (33) 135 (42) <0.001
Estimated glomerular filtration rate§
 Mean — ml/min/1.73 m2 of body-surface area 71±22 62±22 <0.001
 <60 ml/min/1.73 m2 — no. (%) 1098 (31) 157 (49) <0.001
Median suPAR level (IQR) — pg/ml 3162 (2451–4115) 3937 (2935–5070) <0.001
Percutaneous coronary intervention — no. (%) 1905 (54) 143 (45) 0.001
Acute myocardial infarction — no. (%) 449 (13) 47 (15) 0.10
Median volume of contrast material (IQR) — ml 157 (95–230) 136 (77–210) 0.002
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*

Plus–minus values are means ±SD. IQR denotes interquartile range, and suPAR soluble urokinase plasminogen activator receptor.

†

Race was reported by the patient.

‡

The body-mass index is the weight in kilograms divided by the square of the height in meters.

§

The estimated glomerular filtration rate was calculated with the use of the Chronic Kidney Disease Epidemiology Collaboration equation.

ASSOCIATION OF SUPAR AND ACUTE KIDNEY INJURY

The characteristics of the patients, stratified according to quartiles of suPAR level, are shown in Table S2. After coronary angiography, the incidence of acute kidney injury was 14% in the highest suPAR quartile (≥4184 pg per milliliter) and 4% in the lowest quartile (<2475 pg per milliliter), which yielded an unadjusted odds of acute kidney injury that was 3.8 times as high in the highest quartile as in the lowest quartile (Fig. 1A). The association between suPAR level and postprocedural acute kidney injury persisted despite adjustment for clinical characteristics (model 2), including the volume of contrast material and baseline kidney function (model 3; adjusted odds ratio, 2.66; 95% confidence interval [CI], 1.77 to 3.99). The results were consistent when we examined the suPAR level as a continuous variable (per natural log) (adjusted odds ratio, 2.10; 95% CI, 1.54 to 2.87). The suPAR level was also strongly associated with the combined outcome of acute kidney injury or death from any cause at 90 days (adjusted odds ratio, 2.29; 95% CI, 1.71 to 3.06) (Table S3). In subgroup and sensitivity analyses, the odds ratios (per natural log of suPAR) for acute kidney injury remained consistent across relevant subgroups, including each cohort separately (Fig. 1B).

Figure 1 (facing page). Risk of Acute Kidney Injury after Coronary Angiography.

Figure 1 (facing page).

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Panel A shows the odds ratios and 95% confidence intervals (CIs; I bars) for acute kidney injury according to quartiles of soluble urokinase plasminogen activator receptor (suPAR) level before the procedure. Model 1 was unadjusted; model 2 was adjusted for age, sex, race, smoking history, diabetes mellitus, congestive heart failure, hypertension, and cohort (EmCAB [Emory Cardiovascular Biobank] or CASABLANCA [Catheter Sampled Blood Archive in Cardiovascular Diseases]); and model 3 incorporated the aforementioned variables in addition to acute myocardial infarction, revascularization, volume of contrast material, and baseline kidney function (estimated glomerular filtration rate). Quartile 1 was the reference group (1.00) in all models, with a suPAR level of less than 2475 pg per milliliter. The suPAR levels in quartiles 2, 3, and 4 were 2475 to 3198 pg per milliliter, 3199 to 4183 pg per milliliter, and 4184 pg per milliliter or more, respectively. Panel B shows the odds ratios for acute kidney injury per 1 unit natural log of suPAR according to subgroup in the unadjusted analysis (model 1). Stage 3 chronic kidney disease was defined as an estimated glomerular filtration rate of less than 60 ml per minute per 1.73 m2 of body-surface area.

Last, we examined the incremental value of adding suPAR to the NCDR Simplified Integer Risk Score in predicting the risk of acute kidney injury after coronary angiography.35 The AUC for the NCDR risk score was 0.579 (95% CI, 0.560 to 0.597). The addition of suPAR to the NCDR score modestly improved the AUC to 0.628 (95% CI, 0.610 to 0.647), with a change in the AUC of 0.050 (95% CI, 0.013 to 0.087).

CARDIAC SURGERY AND ICU COHORTS

The demographic and clinical characteristics of the patients in the surgical and ICU cohorts are listed in Tables S4 through S7. Among 250 patients who underwent cardiac surgery, the incidence of acute kidney injury was 40% in the highest suPAR quartile (≥5100 pg per milliliter) and 16% in the lowest quartile (<2860 pg per milliliter). Among 692 critically ill patients admitted to the ICU, the incidence of acute kidney injury was 30% (53 patients) in the highest suPAR quartile (≥9440 pg per milliliter) and 9% (15 patients) in the lowest quartile (<5150 pg per milliliter).

Among patients in the surgical cohort, acute kidney injury developed postoperatively in 67 (27%); of those, 14 (6%) had severe (stage 2 or 3) acute kidney injury, and 8 (3%) underwent dialysis. In both the surgical and ICU cohorts, the risk of acute kidney injury increased steadily with increasing suPAR quartiles, with an increase of 3.5 to 4 times in the risk of acute kidney injury in the highest suPAR quartile as compared with the lowest suPAR quartile. The association between suPAR and acute kidney injury was only minimally attenuated in multivariable analyses and did not differ between subgroups. (Details are provided in Figs. S1 through S3.)

SUPAR OVEREXPRESSION AND WORSENING ACUTE KIDNEY INJURY IN EXPERIMENTAL MODELS

Before the injection of iohexol, baseline kidney function and histologic findings were similar in the wild-type mice and the suPAR-transgenic mice at 10 weeks of age (Fig. 2A, 2D, and 2G), despite higher suPAR levels in the transgenic mice than in the wild-type mice (210±56 ng per milliliter vs. 2±1 ng per milliliter). At 24 hours after the injection of contrast material, both wild-type mice and suPAR-transgenic mice had an increase in the serum creatinine level. However, suPAR-transgenic mice had significantly higher creatinine levels and more severe histopathological features of acute kidney injury than their wild-type counterparts that had received the IgG isotype (Fig. 2E and 2H). One mouse in the suPAR-transgenic group that received IgG died unexpectedly, so data are shown for 19 mice.

Figure 2 (facing page). Acute Kidney Injury in Wild-Type and Transgenic Mice before and after Treatment with Anti-uPAR Monoclonal Antibody.

Figure 2 (facing page).

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Panels A through F show representative kidney histologic findings, on high-power view, with the use of periodic acid–Schiff stain in samples obtained from wild-type mice and suPAR-transgenic mice at baseline (Panels A and D) and 48 hours after the administration of iohexol (Panels B, C, E, and F) stratified according to treatment (IgG isotype, in Panels B and E; or urokinase plasminogen activator receptor [uPAR] monoclonal antibody, in Panels C and F). Wild-type mice and suPAR-transgenic mice had largely normal kidney morphologic features at baseline. At 48 hours after iohexol administration, tubular vacuolization could be seen in all wild-type and suPAR-transgenic mice (arrows). The suPAR-transgenic mice that received IgG (Panel E) had more severe renal injuries than mice in any other studied groups. The suPAR-transgenic mice that received the uPAR monoclonal antibody (Panel F) had significantly less severe tubular vacuolization than their counterparts that received the IgG isotype (Panel E). Panel G shows serum creatinine levels measured before and after the administration of contrast material. To convert the values for creatinine to micromoles per liter, multiply by 88.4. As compared with baseline, the serum creatinine level at 24 hours after iohexol injection was increased in all examined groups. The suPAR-transgenic mice that received the IgG isotype had much higher creatinine levels than mice in any other groups. There was no significant between-group difference at baseline. Panel H shows a semiquantitative scoring system that accounts for glomerular and tubular changes associated with acute kidney injury; kidney-injury scores range from 0 to 12, with higher scores indicating more severe kidney injury. (One mouse in the suPAR-transgenic group that received IgG died unexpectedly, so data are shown for 19 mice.) The analyses in Panels G and H were conducted with two-way analysis of variance. In Panels G and H, bars represent means, and I bars ±1 SD; circles or squares indicate values in individual mice.

ATTENUATION OF ACUTE KIDNEY INJURY WITH ANTI-UPAR ANTIBODY

Mice that were pretreated with a uPAR monoclonal antibody had lower creatinine levels at 24 hours than their counterparts that received the IgG isotype (Fig. 2G). When comparing the renal histopathological findings in the wild-type mice and suPAR-transgenic mice, we found that both groups had largely normal histologic features at baseline (Fig. 2A and 2D). At 48 hours after the administration of iohexol, all the mice had histologic features typical of contrast-induced acute kidney injury, including tubular vacuolization, tubular necrosis, and casts (Fig. 2B, 2C, 2E, and 2F). The suPAR-transgenic mice that were pretreated with a uPAR monoclonal antibody had milder histopathological features of acute kidney injury and lower kidney injury scores than the mice that received the IgG isotype (Fig. 2E and 2F).

EFFECT OF SUPAR ON BIOENERGETIC PROFILE AND OXIDATIVE STRESS OF HK-2 CELLS AND PODOCYTES

HK-2 cells that were exposed to suPAR had significantly higher energetic demand under baseline conditions, with increased mitochondrial basal respiration and ATP production, and higher maximum rate of respiration and spare respiratory capacity than cells exposed to media alone. The suPAR-treated cells also had a higher rate of nonmitochondrial oxygen consumption, indicating an active involvement of other cellular oxygen–consuming reactions in addition to that catalyzed by the mitochondrial cytochrome c oxidase. The oxygen-consuming rates that were attributed to proton leak across the mitochondrial membrane did not differ between suPAR-exposed and nonexposed cells, which indicated that mitochondria were not damaged by suPAR and that the mitochondrial integrity was maintained after suPAR treatment. Superoxide generation was increased by a factor of 2 in the presence of suPAR, an effect that was completely abrogated by co-exposure to uPAR antibody. These effects were attenuated when uPAR antibody was coadministered with suPAR. These effects were not seen in podocytes that were exposed to suPAR. (Details are provided in Figs. S4 and S5.)

DISCUSSION

This study showed that suPAR was associated with subsequent acute kidney injury in several cohorts (4769 patients who were exposed to intra-arterial contrast material for coronary angiography, who underwent cardiac surgery, or who were critically ill). Concurrently, we obtained experimental evidence that suPAR may be directly involved in the pathogenesis of acute kidney injury by sensitizing kidney proximal tubules to injury through modulation of cellular bioenergetics and increased oxidative stress. Inhibiting suPAR with the use of a monoclonal antibody attenuated the effect of iohexol on kidney function in mice overexpressing suPAR and abrogated bioenergetic changes in HK-2 cells exposed to suPAR.

There has been little progress in the overall risk stratification, prevention, and treatment of acute kidney injury. Therapies such as intravenous saline hydration, acetylcysteine, and sodium bicarbonate have had little success.27,36 Biomarkers that are currently under study, such as cystatin C, neutrophil gelatinase–associated lipocalin, and kidney injury molecule 1, are early markers of acute kidney injury whose levels increase only after renal injury has occurred.37,38 We found that preprocedural suPAR levels were predictive of acute kidney injury in both low-risk and high-risk cohorts and across subgroups, independent of relevant clinical characteristics, including baseline kidney function. In addition, suPAR modestly improved risk discrimination when this variable was added to the NCDR Simplified Integer Risk Score for acute kidney injury. These findings are in line with one previous smaller study involving 107 patients who underwent cardiac surgery.39 Improved assessment of the preprocedural risk of acute kidney injury would allow for more informed decision making and would help to identify a subgroup of patients who would benefit from an intervention to minimize procedural acute kidney injury, potentially in the form of anti-suPAR therapies.

The wide spectrum of clinical contexts in which suPAR levels are associated with incident acute kidney injury suggests that the underlying mechanism is not dependent on the type of inciting event. On the basis of our animal models, we speculate that there may be a synergistic effect between suPAR — which acts as a metabolic sensitizer and increases the workload of tubular cells — and various injuries such as ischemia, cytotoxic effects, and oxidative stress (e.g., induced by cardiac surgery or the use of iodinated radioactive contrast material).40,41 In response to high suPAR levels, proximal tubular cells, but not podocytes, showed an increase in mitochondrial respiration. Extramitochondrial oxygen consumption was increased in both types of cells by suPAR, but to a greater degree in tubular cells than in podocytes — a finding that suggests activation of extramitochondrial enzymatic oxidation.

Before the injection of contrast material, we found no histopathological or biochemical measures of renal dysfunction in 10-week-old mice overexpressing suPAR. After the injection of contrast material, the suPAR-transgenic mice had significantly more severe acute kidney injury than the wild-type mice. The effect of the administration of contrast material on kidney injury in suPAR-transgenic mice was attenuated with a monoclonal antibody to uPAR, which suggests that chronically elevated suPAR levels sensitized the kidney to acute injury (although suPAR concentrations were >50 times as high as median levels in humans) and that this sensitizing effect of suPAR could be reversed pharmacologically. These conclusions are in line with a recent report showing that targeting the urokinase receptor in a rat model of diabetic kidney disease resulted in improvement in kidney function.42 The mechanisms of suPAR in kidney dysfunction have focused on its source of production and its role in binding and activating podocyte αvβ3 integrins.15,17 Other reports have suggested that suPAR also affects proximal tubules and drives kidney fibrosis in an integrin-dependent manner.14,16,17,43 Although it will be helpful for studies to be expanded into other models of acute kidney injury, it is plausible that prolonged suPAR exposure affects podocytes and tubular cells by means of different mechanisms, a hypothesis in line with the different bioenergetic profiles seen in HK-2 cells and podocytes in response to suPAR.

Our study has several strengths. We found consistent results across several well-characterized cohorts in three clinical contexts and a total of 4769 participants, allowing for subgroup analyses and adjustment for confounders. The accompanying experimental findings have compelling translational and therapeutic implications. Our study also has several important limitations. One is the retrospective nature of the cohort studies and risk of selection bias. However, the collection of blood samples and the end points were prespecified in all the cohorts included in the study, and the incidence of acute kidney injury in these cohorts was consistent with rates reported in the NCDR registry.35 We could not compare suPAR to other biomarkers such as kidney injury molecule 1 and neutrophil gelatinase–associated lipocalin in the present study, because these were not systematically measured in all cohorts.

In conclusion, high suPAR levels were associated with incident acute kidney injury in several patient cohorts. The experimental models used here suggest that suPAR may be a pathogenic factor in acute kidney injury.

Supplementary Material

Supplement1

Acknowledgments

Supported by grants (1R61HL138657-02, 1P30DK111024-03S1, 5R01HL095479-08, 3RF1AG051633-01S2, 5R01AG042127-06, 2P01HL086773-08, U54AG062334-01, 1R01HL141205-01, 5P01HL101398-02, 1P20HL113451-01, 5P01HL086773-09, 1RF1AG051633-01, R01NS064162-01, R01HL89650-01, HL095479-01, 1DP3DK094346-01, and 2P01HL086773, to Dr. Quyyumi; R01HL089650-01 and R01DK101350, to Drs. Sever, Wei, and Reiser; K23DK106448, to Dr. Leaf; and R01HL142093, to Dr. Baron) from the National Institutes of Health, by a grant (15SFCRN23910003, to Dr. Quyyumi) from the American Heart Association, by an American Society of Nephrology Foundation for Kidney Research Carl W. Gottschalk Research Scholar Grant (to Dr. Leaf), by the Hutter Family Professorship (to Dr. Januzzi), by a grant (W81XWH1810667, to Dr. Baron) from the Department of Defense, by a Merit Award (BX002006, to Dr. Sheikh-Hamad) and a Career Development Award (BX002912, to Dr. Pan) from the Department of Veterans Affairs, and by the American Society of Nephrology Foundation for Kidney Research George B. Rathmann Research Fellowship Award, as part of the Ben J. Lipps Research Fellowship Program (to Dr. Holliday).

Disclosure forms provided by the authors are available with the full text of this article at NEJM.org.

We thank the members of the Emory Cardiovascular Biobank team, specifically Yi-An Ko for database maintenance and statistical support, Mosaab Awad and Ayman Alkhoder for serving as coordinators, the staff of the Emory Clinical Cardiovascular Research Institute, and the staff of the Atlanta Clinical and Translational Science Institute for the recruitment of participants, compilation of data, and preparation of samples; Beata Samelko and Jing Li for technical help with mouse tissue and enzyme-linked immunosorbent assay for soluble urokinase plasminogen activator receptor; the members of the Brigham and Women’s Hospital Registry of Critical Illness (Mayra Pinilla, Sam Ash, Paul Dieffenbach, Laura Fredenburgh, and Anthony Massaro); and Myles Wolf (Duke University) and Monnie Wasse (Rush University) for critical reading of an earlier version of the manuscript.

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