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Biomarkers of acute kidney injury
Clinical Queries: Nephrology 0101 (2012) 13–17
Contents lists available at ScienceDirect
Clinical Queries: Nephrology January–March 2012, Vol. 1/Issue 1
j o u r n a l h o m e p a g e : h t t p : / / w w w. e l s e v i e r. c o m / l o c a t e / c q n
ISSN No.: 2211-9477
Biomarkers of acute kidney injury R.K. Sharma*,† *Head of the Department of Nephrology, †Director, Sanjay Gandhi Postgraduate Institute of Medical Sciences, Lucknow – 226014, India.
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Article history: Received 2 November 2011 Accepted 4 November 2011
Keywords: Acute kidney injury Biomarker Creatinine Cystatin C Interleukin-18 Neutrophil gelatinase-associated lipocalin Renal function
a b s t r a c t
Acute kidney injury (AKI) represents a common serious clinical problem. Incidence of AKI is increasing rapidly to epidemic proportions. Development of AKI, especially in intensive care settings, is associated with increased morbidity, mortality, and hospitalization costs. Currently available diagnostic tools are mostly insensitive for early diagnosis. However, prompt diagnosis and risk stratification are necessary for guiding therapy and preventing progression of disease. Finding an early, reliable, suitable, easily reproducible, economical, and accurate biomarker for AKI is a top research priority. In recent years, many urinary and serum proteins have been investigated as possible early markers of AKI and some of them have shown great promise. This topic reviews some of the emerging biomarkers of AKI.
Introduction For the past 30 years, there have been no major improvements in the mortality rate of hospitalized patients with severe acute kidney injury (AKI), despite advances in supportive care. One key reason is because a change in the serum creatinine (SCr), which has been the standard metric for detection and progression of AKI, is not sufficiently sensitive for an early diagnosis of AKI. The absence of a more sensitive biomarker has impaired progress in this field and has had a detrimental effect on the design and outcome of AKI clinical trials. Recently, several proteins emerged as sensitive and specific biomarkers with a capacity to be used in the detection of early kidney injury and grading of injury severity. Acute kidney injury Despite more than half a century of investigation, AKI remains a major healthcare issue in medicine today. Reported to occur in 1–32% of all hospital admissions and 10–90% of intensive care unit admissions, the wide variation reflects different criteria used to define AKI. However, independent of definition, a diagnosis of AKI is consis tently associated with an increase in both short- and long-term morbidity and mortality. Even the mildest forms of AKI are independently
*Corresponding author. E-mail address: [email protected] ISSN: 2211-9477 Copyright © 2012. Reed Elsevier India Pvt. Ltd. All rights reserved. doi: 10.1016/S2211-9477(11)70003-6
Copyright © 2012, Reed Elsevier India Pvt. Ltd. All rights reserved.
associated with increased early as well as long-term mortality, the risk increasing as severity of renal injury increases.1,2 Furthermore, the incidence of AKI is increasing. Based on a large administrative database study of hospital admissions between 1992 and 2001, Xue et al estimated an 11% increase per year in the incidence of AKI.3 However, of even greater concern is the failure to develop effective interventions to prevent or treat AKI, meaning that the current management remains directed toward supportive therapy while awaiting recovery of renal function.
The urgent need for biomarkers Traditional blood (creatinine and blood urea nitrogen [BUN]) and urinary markers of kidney injury (urinary casts and fractional excretion of sodium) do not allow for early detection of AKI.4 The change in SCr does not discriminate the time and type of renal insult nor the site and extent of glomerular or tubular injury. Previous animal studies clearly demonstrated that treatment should be initiated well before the rise of SCr and very early after the insult.5–9 Therefore, the absence of sensitive and specific biomarkers for the early detection of AKI impairs progress in the diagnosis and treatment of patients with AKI and has a detrimental effect on the design and, possibly, the outcomes of clinical trials. Many potential therapeutic agents, such as atrial natriuretic peptide, insulin-like growth factor-1, and endothelin-receptor antagonist,10–13 have been tried, but have had little success. The lack of reliable biomarkers for early injury detection leads to a delay in the introduction of treatment until well into the course of renal disease. At present, there is no single intervention
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or sequence of clinical interventions that will significantly improve renal function after the onset of acute tubular injury or necrosis. Dialysis remains the only Food and Drug Administration (FDA)approved treatment option for established AKI. Acknowledging the inherent deficiencies of SCr to diagnose AKI, the American Society of Nephrology in 2005 designated identification, characterization, and development of new AKI biomarkers as a key research area for the next 5 years.
Characteristics of an ideal biomarker for acute kidney injury An ideal biomarker would identify patients at highest risk for AKI in a timely manner, thus allowing early and potentially effective intervention. Characteristics of the ideal AKI biomarker have been described and include early identification of injury, stratification according to injury severity, etiologic specificity for the injury, and providing valuable prognostic information. However, the wide spectrum of pathophysiology leading to AKI makes it unlikely that any single biomarker will achieve all these aims. Several promising biomarkers of AKI have been identified, both in urine and plasma, and are currently the subject of ongoing studies defining their clinical utility. However, the translational process from bench to bedside is complicated. Interpretation of novel biomarkers to detect minor but significant renal injury undetected by SCr proved difficult. Although insensitive and slow to respond, creatinine remains the only marker validated against clinically relevant outcomes. Any potential replacement must therefore demonstrate the ability to identify clinically meaningful injury and be useful in guiding suitable interventions or other management decisions. Although the molecular pathways mediating renal injury are increasingly understood, with potential to quantify individual components of these pathways in the laboratory,14 the focus of this clinical commentary is on biomarkers that reflect renal injury, which is frequently the result of multiple contemporaneous mechanisms in clinical practice. Besides establishing the early diagnosis, biomarkers are needed for several other purposes in AKI (summarized in Table 1). Thus, biomarkers are needed for: (1) pinpointing the location of primary injury (proximal tubule, distal tubule, interstitium, or vasculature); (2) determining the duration of kidney failure (AKI, chronic kidney disease, or ‘acute on chronic’ kidney disease); (3) discerning AKI subtypes (pre-renal, intrinsic renal, or post-renal); (4) identifying AKI etiologies (ischemia, toxins, sepsis, or a combination); (5) differentiating AKI from other forms of acute kidney disease (urinary tract infection, glomerulonephritis, or interstitial nephritis); (6) risk stratification and prognostication (duration and severity of AKI, need for renal replacement therapy, length of hospital stay, and mortality); (7) defining the course of AKI; and (8) monitoring the response to AKI interventions. Biomarkers are also desperately needed for use as surrogate endpoints in clinical trials evaluating potential therapeutics for AKI.
Biomarkers of acute kidney injury Changes in excretion of specific markers in the urine have been proposed as reflecting injury to specific regions of the nephron. Table 1 Areas of need for biomarkers in acute kidney injury. Biomarkers are needed to determine: 1. Location of injury 2. Duration of AKI 3. AKI subtypes 4. AKI etiologies 5. Differentiate from other forms of acute kidney disease 6. Risk stratification and prognostication 7. Defining course of AKI 8. Monitoring interventions AKI: acute kidney injury.
For example, high molecular weight proteinuria is proposed as an indication of glomerular injury;15–18 low molecular weight proteinuria,19,20 brush border antigens21–23 urinary enzymes,24–33 and other urinary proteins34–42 are proposed to indicate damage to the proximal convoluted tubule. As a result, insults to specific areas of the nephron, as might occur with a particular nephrotoxin, may be associated with different urinary excretion patterns of these markers. However, attempts to use these markers in screening patients for renal injury and identifying the site of kidney injury have been disappointing; this is due to a lack of sufficient validation, the lack of standardized assays, instability in urine, and the changes in pattern specificity of urinary marker excretion with advancing renal dysfunction. Recently, several protein biomarkers emerged through the application of functional genomics and proteomics to human and animal models of AKI, and proved to be promising biomarkers as non-invasive indicators of AKI.
Neutrophil gelatinase-associated lipocalin Deoxyribonucleic acid microarray techniques searching for candidate biomarkers of AKI found neutrophil gelatinase-associated lipo calin (NGAL) as one of the maximally-induced genes in a murine model of renal ischemia-reperfusion injury. A 25-KDa glycoprotein covalently bound to gelatinase, its resistance to proteolysis further enhanced potential suitability as a clinical biomarker. It is synthesized and secreted by tubular epithelial cells of the proximal and distal segment. It is freely filtered by the glomerulus, undergoing rapid clearance by the proximal tubule via receptor binding and endocytosis. In healthy kidneys, it is barely detectable in either plasma or urine. However, in the setting of acute tubular injury, NGAL undergoes rapid and profound up-regulation with large increases in both urine and plasma. Distinct from traditional markers of function such as creatinine, this rapid response enables NGAL to potentially identify injured kidney much earlier than was previously possible. The endogenous role of NGAL remains unclear. It seems to be involved with iron transportation to and from the proximal tubular epithelial cells, and animal studies demonstrate a renoprotective effect of exogenously administered NGAL in the setting of acute ischemic injury. The NGAL demonstrated a near-perfect performance for identifying AKI after pediatric cardiac surgery with an area under the receiver operator characteristic curve (AUCROC) of 0.99 and 1.0 at 2 and 4 hours, respectively, after cardiopulmonary bypass (CPB), respectively.43 Subsequent studies of both urinary and plasma NGAL in pediatric cardiac surgery support this, further demonstrating it to be an independent predictor of AKI severity and duration as well as hospital length of stay. However, results in adult cardiac surgery have been mixed, with different studies reporting widely varying diagnostic characteristics for NGAL. The AUCROC for early diagnosis of AKI by urinary NGAL has varied from 0.61 at 18 hours post-CPB44 to 0.96 at 2 hours post-CPB.45 Similarly, performance of plasma NGAL for the diagnosis of AKI has varied from an AUCROC of 0.54 within 6 hours of CPB46 to 0.87 at 24 hours post-CPB.47 Although specificity was consistently 70–80% in these studies, sensitivity has varied greatly, ranging from 40% to 90%. The reason for such widely discrepant findings is unclear. Studies reporting poorer diagnostic performance of NGAL have typically included patients with a wide spectrum of baseline renal function, and it is unknown whether this impacts the diagnostic performance of NGAL. The additional comorbidities typical of an adult cardiac surgical population may also introduce potential confounding variables, thus increasing the etiologic heterogeneity of AKI in this population. Furthermore, it is uncertain whether age itself modifies NGAL synthesis in AKI. Recently, urinary NGAL at hospital admission in multitrauma patients provided excellent early identification of AKI developing during the subsequent 5 days (AUCROC 0.98) with both sensitivity and specificity > 90%.48 Also, NGAL has been demonstrated to identify delayed graft function within hours of kidney transplantation,49 and ongoing studies are exploring its utility for early identification of renal injury in liver transplantation.
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The biologic rationale for NGAL as an early AKI biomarker is strong. However, a number of key issues, including the wide variability in reported diagnostic performance, require clarification before adoption into clinical practice. Although a rapid point-of-care test is available, experience is limited and its reliability remains to be confirmed with most studies to date using a research-based enzyme-linked immunosorbent assay (ELISA) technique for NGAL quantification.
Kidney injury molecule-1 Kidney injury molecule-1 (KIM-1) is a type 1 transmembrane glycoprotein containing a novel 6-cysteine immunoglobulin-like domain plus a threonine/serine and proline-rich domain characteristic of mucin-like O-glycosylated proteins. It is undetectable in normal kidney tissue or urine, but is expressed at high levels in dedifferentiated proximal tubule epithelial cells from human and rodent kidneys after ischemic or toxic injury, and in renal cell carcinoma.50–53 It is also known as hepatitis A virus cellular receptor 1 and T-cell immunoglobulin and mucin-domain-containing molecule 1.54–56 Also, KIM-1 has been implicated as a urinary biomarker of AKI in pediatric and adult patients. In a small cross-sectional study, Han et al demonstrated that soluble forms of cleaved KIM-1 can be detected in the urine of patients with established AKI; elevated urinary KIM-1 levels were found within 12 hours after an initial ischemic insult and prior to the appearance of granular casts in the urine.51 Urinary KIM-1 was also evaluated after pediatric cardiac surgery in a case control study,57 using samples from the same cohort in the NGAL study previously described. Urinary KIM-1 had an AUCROC of 0.57 at 2 hours, 0.83 at 12 hours, and 0.78 at 24 hours. High urinary KIM-1 expression was also associated with adverse clinical outcomes in patients with established AKI. In a prospective study, a cohort of 201 patients with established AKI, Liangos et al demonstrated that urinary KIM-1 and N-acetyl-b-D-glucosaminidase (NAG) were significantly associated (AUCROC of 0.78) with the clinical composite endpoints of death or renal replacement therapy.58 Kidney injury molecule-1 is particularly interesting given its ‘kidney dominance’ of expression. In a follow-up study involving a larger number of adult patients, the temporal expression patterns and the utility of urinary KIM-1 for the early detection of AKI are being studied.
Interleukin-18 Interleukin-18 (IL-18) is a proinflammatory cytokine that is known to be induced and cleaved in the proximal tubule, and subsequently can be easily detected in the urine following ischemic AKI in animal models. In an initial cross-sectional study, urine IL-18 levels mea sured by a commercially available ELISA were markedly elevated in patients with established AKI, but not in subjects with urinary tract infection, chronic kidney disease, nephrotic syndrome or pre-renal azotemia. In a subsequent study of patients who developed AKI 2–3 days after cardiac surgery, urinary IL-18 levels peaked at 12 hours post-surgery, and predicted AKI with an AUCROC of 0.75.59 Urine IL-18 is also considered an early biomarker of AKI in the intensive care setting, being able to predict this complication about 2 days prior to the rise in SCr. Early elevated urine IL-18 levels correlated with the severity of AKI as well as mortality. Thus, IL-18 may also represent a promising biomarker for AKI. Interleukin-18 is more specific to ischemic AKI, and levels are largely unaffected by nephrotoxins, chronic kidney disease or urinary tract infections. However, IL-18 measurements may be influenced by a number of co-existing variables, since renal IL-18 mRNA levels are known to be induced in other disease states such as endotoxemia, immunological injury and cisplatin toxicity. Furthermore, plasma IL-18 levels are known to be increased in various systemic inflammatory states, and the relationships between plasma and urine IL-18 remain unexplored.
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Cystatin C A small cysteine proteinase inhibitor, cystatin-C, has many features of an ideal glomerular filtration marker. Synthesized and released into plasma by all nucleated cells at a constant rate, its small size (13 KDa) and positive charge at physiologic pH makes it freely filtered at the glomerulus. It is neither secreted nor reabsorbed by renal tubules but undergoes almost complete catabolism by proximal tubular cells, and thus little, if any, appears in the urine. With a half-life of about 2 hours, serum cystatin-C reflects glomerular filtration rate better than creatinine. Although it is generally considered less subject to the non-renal variables that impact creatinine, recent studies suggest that cystatin-C levels may in fact be affected by various anthropometric measures as well as inflammatory processes, use of corticosteroids, and changes in thyroid function, thereby potentially confounding perioperative interpretation.60 Although increasingly reported as an endpoint in clinical studies, the diagnostic and prognostic characteristics of cystatin-C for AKI remain incompletely defined. In a mixed critical care population, cystatin-C enabled a diagnosis of AKI 1.5 days earlier than plasma creatinine, with moderate ability to predict dialysis requirement.61 However, a similar repeat study found that cystatin-C identified AKI no earlier than creatinine and did not predict mortality.62 Results in adult cardiac surgery are similarly mixed, with one study reporting good discrimination for AKI within 6 hours of surgery (AUCROC 0.83, sensitivity 77%, specificity 86%),63 whereas a further study of similar size found serum cystatin-C no better than chance for identifying AKI.64 However, the latter study did find that urinary cystatin-C identified AKI within 6 hours of surgery (AUCROC 0.72, sensitivity 94%, specificity 40%), suggesting that tubular injury may impair the usual catabolism of cystatin-C, leading to its appearance in urine. Several studies report a rise in cystatin-C within 8 hours of exposure to radiocontrast agents but without adequate description of the diagnostic characteristics for contrast-induced nephropathy. Although a commercial platform for rapid and reliable cystatin-C measurement is available, the total number of patients studied remains small and with inconsistencies in results that are not well understood.
N-Acetyl-b-D-Glucosaminidase Although several enzymes present in tubular epithelial cells have been investigated as potential markers of renal injury, NAG remains the most extensively investigated to date. A lysosomal enzyme, it is abundantly present in proximal tubular epithelial cells and its relatively large size (130 KDa) prevents glomerular filtration with the result that urinary NAG represents enzyme leakage from proximal tubular cells into the tubular lumen. It is stable in urine across a range of pH and temperature, and it is easily quantified by commercially available colorimetric or spectrophotometric methods. Although a large number of studies have profiled the release of NAG across a diverse range of clinicopathologic conditions, each purporting to demonstrate subtle proximal renal tubular damage, few studies address either the diagnostic capability for AKI or a direct link with clinical outcomes. The NAG was used extensively through the 1990s by anesthesiologists investigating the potential nephrotoxicity of compound A, a degradation product of sevoflurane. Despite several studies indicating increases in urinary NAG with sevoflurane exposure, as well as other potential markers of renal injury, such as albumin, microglobulins, glutathione-S-transferase, and glucose, a link with histopathologic changes, increasing SCr, or adverse clinical outcomes was unable to be established, thus leaving the interpretation of these transient perioperative changes unclear. However, in a cross-sectional study, urinary NAG provided excellent discrimination of patients with established AKI from a control group including both normal individuals and patients with urinary tract infection (AUCROC 0.97)65 and in a hospital population referred for nephrology consultation for acute renal failure, increasing urinary NAG was associated with a 3.0-, 3.7-, and 9.1-fold higher odds for the composite outcome of in-hospital mortality or dialysis with each increasing
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quartile relative to the first, a relationship maintained despite adjusting for multiple comorbidities.66 The NAG levels increased within 6 hours of pediatric cardiac surgery, remaining elevated through 48 hours. Although higher in patients who developed AKI, diagnostic utility was modest 12 hours post-CPB (AUCROC 0.69) and despite sensitivity > 80%, specificity remained poor across a range of potential threshold values. In contrast, urinary NAG levels were not consistently different between patients with or without AKI after cardiac surgery in adults, and a single study in liver transplant recipients failed to show a difference in post-operative urinary NAG levels between patients with or without AKI.67
Finally, larger prospective studies are necessary to validate the temporal expression pattern of various biomarkers for early detection of AKI, to determine how to combine multiple biomarkers for early detection of AKI, and to discover how the temporal course relates to the onset, severity, and outcome of AKI. More information must be gathered regarding the accuracy of measurement in the presence of interfering substances and the stability of the various biomarkers in routine clinical storage conditions, including a number of freeze–thaw cycles.
Liver fatty acid binding protein
There is a need for a new standard definition of AKI that is not based on a change in SCr. The new definition of AKI should be based on multiple biomarkers and clinical parameters, and capable of detecting initial renal injury within minutes to hours. Any potential AKI biomarker should undergo several clinical validation processes for both assay performance and diagnostic utility. Furthermore, several requirements must be met in order for AKI biomarkers to be useful in daily clinical practice. The AKI biomarkers must:
Liver fatty acid binding protein (LFABP) is a 14 KDa protein normally expressed in the proximal convoluted and straight tubules of the kidney, and up-regulated in animal models of AKI. An ELISA for this analyte is commercially available; it was first used to demonstrate that urinary L-FABP levels were significantly increased prior to the increase in SCr in those patients who developed AKI post-contrast dye. In a recent prospective study of children undergoing cardiac surgery, urine LFABP increased at 4 hours post-bypass, with an AUC of 0.810 for a cut-off value of 486 ng/mg creatinine.68 Thus, L-FABP also appears to represent a promising AKI bio marker. However, urinary L-FABP measurements may also be influenced by a number of confounding variables. Several studies have documented increased urinary L-FABP levels in patients with nondiabetic chronic kidney disease, early diabetic nephropathy, idiopathic focal glomerulo sclerosis and polycystic kidney disease. Additionally, LFABP is also abundantly expressed in the liver, and urinary L-FABP may be influenced by serum LFABP levels.
Limitations of biomarkers for early detection of acute kidney injury in clinical situation Additional studies are necessary before biomarkers may be used in routine clinical practice. Currently, these promising biomarkers have been tested in only small studies and limited clinical situations. None of these biomarkers have been systematically evaluated in the various clinical settings of AKI. Furthermore, the studies have been insufficiently powered to establish a cut-off value that is predictive of AKI. In addition, there are very limited data available, at present, about temporal expression patterns for various biomarkers including NGAL, KIM-1, IL-18, and Cys-C in the various clinical settings of AKI from the onset of renal insult. The heterogeneity of AKI suggests that more than one marker may be necessary to obtain sufficient sensitivity and specificity for AKI screening. An analysis of multiple biomarkers may optimize early detection of AKI, as in the case of prostate cancer detection. Analyses of multiple markers may also provide insight into the pathophysiology and sites of nephron injury, since different markers may be chosen to reflect injury to different nephron segments. Nevertheless, it would be naïve to call the result positive if one or more biomarkers are positive, since such an approach would be inefficient and increase sensitivity at the expense of specificity, or vice versa. A linear combination that would maximize the AUCROC for detection of AKI may be useful, but this approach remains to be validated by future studies.57 Currently, there is no standard procedure to combine the multiple biomarkers for clinical use. Another major challenge is the development of a rapid assay for validated biomarkers at the bedside or in a clinical laboratory for the detection of AKI. Current quantification methods include ELISA, Luminex®-based assay, and nephelometry, which are not fast enough for rapid assay. Recently, Biosite Inc. launched a prospective multicenter study evaluating NGAL in early and evolving stages of AKI. The study is using the Triage® NGAL test, which supposedly can accurately measure NGAL concentrations in blood and provide results within 15 minutes.
Conclusion
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Allow for early detection of renal injury. Identify the severity of AKI. Provide a rationale for risk stratification in clinical studies, including the identification of patients at risk for AKI. Guide timing of therapy. Reflect improvement and worsening of the kidney injury. Be amenable to quick and reliable measurement at the bedside or clinical laboratory.
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16. Bazzi C, Petrini C, Rizza V, Arrigo G, D’Amico G. A modern approach to selectivity of proteinuria and tubulointerstitial damage in nephrotic syndrome. Kidney Int 2000;58:1732–41. 17. Mackinnon B, Shakerdi L, Deighan CJ, et al. Urinary transferrin, high molecular weight proteinuria and the progression of renal disease. Clin Nephrol 2003; 59:252–8. 18. Corso A, Zappasodi P, Pascutto C, et al. Urinary proteins in multiple myeloma: correlation with clinical parameters and diagnostic implications. Ann Hematol 2003;82:487–91. 19. Tolkoff-Rubin NE, Rubin RH, Bonventre JV. Noninvasive renal diagnostic studies. Clin Lab Med 1988;8:507–26. 20. Bazzi C, Petrini C, Rizza V, et al. Urinary excretion of IgG and alpha(1)-microglobulin predicts clinical course better than extent of proteinuria in membranous nephropathy. Am J Kidney Dis 2001;38:240–8. 21. Taniguchi N, Tanaka M, Kishihara C, et al. Determination of carbonic anhydrase C and beta 2-microglobulin by radioimmunoassay in urine of heavy-metalexposed subjects and patients with renal tubular acidosis. Environ Res 1979; 20:154–61. 22. Zager RA, Johannes GA, Sharma HM. Quantitating the severity of proximal tubular brush border injury by a simple direct binding radioimmunoassay. Am J Kidney Dis 1982;1:353–8. 23. Mutti A, Lucertini S, Valcavi P, et al. Urinary excretion of brushborder antigen revealed by monoclonal antibody: early indicator of toxic nephropathy. Lancet 1985;26:914–7. 24. Gibey R, Dupond JL, Alber D, Leconte des Floris R, Henry JC. Predictive value of urinary N-acetyl-beta-D-glucosaminidase (NAG), alanine-aminopeptidase (AAP) and beta-2-microglobulin (beta 2M) in evaluating nephrotoxicity of gentamicin. Clin Chim Acta 1981;116:25–34. 25. Stonard MD, Gore CW, Oliver GJ, Smith IK. Urinary enzymes and protein patterns as indicators of injury to different regions of the kidney. Fundam Appl Toxicol 1987;9:339–51. 26. Kohli MM, Ganguly NK, Naur S, Sharma VK. Urinary excretion of renal brush border membrane enzymes in leprosy patients: effect of multidrug therapy. Experientia 1996;52:127–30. 27. Donaldio C, Puccini R, Lucchesi A, Giordani R, Rizzo G. Urinary excretion of proteins and tubular enzymes in renal transplant patients. Ren Fail 1998;20:707–15. 28. Nouwen EJ, De Broe ME. Human intestinal versus tissue-nonspecific alkaline phosphatase as complementary urinary markers for the proximal tubule. Kidney Int 1994;47:S43–51. 29. Olbricht CJ, Steinker M, Auch-Schwelk W, et al. Effect of cyclosporin on kidney proteolytic enzymes in men and rats. Nephrol Dial Transplant 1994;9:22–6. 30. Girolami JP, Bascands JL, Pecher C, et al. Renal kallikrein excretion as a distal nephrotoxicity marker during cadmium exposure in rats. Toxicology 1989;55:117–29. 31. Usuda K, Kono K, Dote T, et al. Urinary biomarkers monitoring for experimental fluoride nephrotoxicity. Arch Toxicol 1998;72:104–9. 32. Branten AJ, Mulder TP, Peters WH, Assmann KJ, Wetzels JF. Urinary excretion of glutathione S transferases alpha and pi in patients with proteinuria: reflection of the site of tubular injury. Nephron 2000;85:120–6. 33. Boldt J, Brenner T, Lang J, Kumle B, Isgro F. Kidney-specific proteins in elderly patients undergoing cardiac surgery with cardiopulmonary bypass. Anesth Analg 2003;97:1582–9. 34. Lynn KL, Marshall RD. Excretion of Tamm-Horsfall glycoprotein in renal disease. Clin Nephrol 1984;22:253–7. 35. Torffvit O, Jorgensen PE, Kamper AL, et al. Urinary excretion of Tamm-Horsfall protein and epidermal growth factor in chronic nephropathy. Nephron 1998; 79:167–72. 36. Taman M, Liu Y, Tolbert E, Dworkin LD. Increase urinary hepatocyte growth factor excretion in human acute renal failure. Clin Nephrol 1997;48:241–5. 37. Sugimura K, Goto T, Tsuchida K, et al. Production and activation of hepatocyte growth factor in acute renal failure. Ren Fail 2001;23:597–603. 38. du Cheyron D, Daubin C, Poggioli J, et al. Urinary measurement of Na+/H+ exchanger isoform 3 (NHE3) protein as new marker of tubule injury in critically ill patients with ARF. Am J Kidney Dis 2003;42:497–506. 39. Muramatsu Y, Tsujie M, Kohda Y, et al. Early detection of cysteine rich protein 61 (CYR61, CCN1) in urine following renal ischemic reperfusion injury. Kidney Int 2002;62:1601–10. 40. Kwon O, Molitoris BA, Pescovitz M, Kelly KJ. Urinary actin, interleukin-6, and interleukin-8 may predict sustained ARF after ischemic injury in renal allografts. Am J Kidney Dis 2003;41:1074–87. 41. Mariano F, Guida G, Donati D, et al. Production of platelet-activating factor in patients with sepsis-associated acute renal failure. Nephrol Dial Transplant 1999;14:1150–7. 42. Zhou H, Pisitkun T, Aponte A, et al. Exosomal fetuin-A identified by proteomics: a novel urinary biomarker for detecting acute kidney injury. Kidney Int 2006; 70:1847–57.
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43. Mishra J, Dent C, Tarabishi R, et al. Neutrophil gelatinase-associated lipocalin (NGAL) as a biomarker for acute renal injury after cardiac surgery. Lancet 2005; 365:1231–8. 44. Wagener G, Gubitosa G, Wang S, et al. Urinary neutrophil gelatinase-associated lipocalin and acute kidney injury after cardiac surgery. Am J Kidney Dis 2008;52: 425–33. 45. Tuladhar SM, Puntmann VO, Soni M, et al. Rapid detection of acute kidney injury by plasma and urinary neutrophil gelatinase-associated lipocalin after cardio pulmonary bypass. J Cardiovasc Pharmacol 2009;53:261–6. 46. Koyner JL, Bennett MR, Worcester EM, et al. Urinary cystatin C as an early biomarker of acute kidney injury following adult cardiothoracic surgery. Kidney Int 2008;74:1059–69. 47. Haase-Fielitz A, Bellomo R, Devarajan P, et al. Novel and conventional serum biomarkers predicting acute kidney injury in adult cardiac surgery—a prospective cohort study. Crit Care Med 2009;37:553–60. 48. Makris K, Markou N, Evodia E, et al. Urinary neutrophil gelatinase-associated lipocalin (NGAL) as an early marker of acute kidney injury in critically ill multiple trauma patients. Clin Chem Lab Med 2009;47:79–82. 49. Lebkowska U, Malyszko J, Lebkowska A, et al. Neutrophil gelatinase-associated lipocalin and cystatin C could predict renal outcome in patients undergoing kidney allograft transplantation: a prospective study. Transplant Proc 2009;41:154–7. 50. Ichimura T, Bonventre JV, Bailly V, et al. Kidney Injury Molecule-1 (KIM-1), a putative epithelial cell adhesion molecule containing a novel immunoglobulin domain, is up-regulated in renal cells after injury. J Biol Chem 1998;273: 4135–42. 51. Han WK, Bailly V, Abichandani R, Thadhani R, Bonventre JV. Kidney Injury Molecule-1 (KIM-1): a novel biomarker for human renal proximal tubule injury. Kidney Int 2002;62:237–44. 52. Ichimura T, Hung CC, Yang SA, Stevens JL, Bonventre JV. Kidney injury molecule-1: a tissue and urinary biomarker for nephrotoxicant-induced renal injury. Am J Physiol. Renal Physiol 2004;286:F552–63. 53. Han WK, Alinani A, Wu CL, et al. Human kidney injury molecule-1 is a tissue and urinary tumor marker of renal cell carcinoma. J Am Soc Nephrol 2005;16: 1126–34. 54. Feigelstock D, Thompson P, Mattoo P, Kaplan GG. Polymorphisms of the hepatitis A virus cellular receptor 1 in African green monkey kidney cells result in antigenic variants that do not react with protective monoclonal antibody 190/4. J Virol 1998;72:6218–22. 55. Feigelstock D, Thompson P, Mattoo P, Zhang Y, Kaplan GG. The human homolog of HAVcr-1 codes for a hepatitis A virus cellular receptor. J Virol 1998;72: 6621–8. 56. Kuchroo VK, Umetsu DT, DeKruyff RH, Freeman GJ. The TIM gene family: emerging roles in immunity and disease. Nat Rev Immunol 2003;3:454–62. 57. Han WK, Waikar SS, Johnson A, et al. Urinary biomarkers for detection of acute kidney injury. Kidney Int 2008;73:863–9. 58. Liangos O, Perianayagam MC, Vaidya VS, et al. Urinary N-acetyl-beta-(D)glucosaminidase activity and kidney injury molecule-1 level are associated with adverse outcomes in acute renal failure. J Am Soc Nephrol 2007;18:904–12. 59. Parikh CR, Mishra J, Thiessen-Philbrook H, et al. Urinary IL-18 is an early predictive biomarker of acute kidney injury after cardiac surgery. Kidney Int 2006;70: 199–203. 60. Knight EL, Verhave JC, Spiegelman D, et al. Factors influencing serum cystatin C levels other than renal function and the impact on renal function measurement. Kidney Int 2004;65:1416–21. 61. Herget-Rosenthal S, Marggraf G, Husing J, et al. Early detection of acute renal failure by serum cystatin C. Kidney Int 2004;66:1115–22. 62. Ahlstrom A, Tallgren M, Peltonen S, Pettila V. Evolution and predictive power of serum cystatin C in acute renal failure. Clin Nephrol 2004;62:344–50. 63. Haase-Fielitz A, Bellomo R, Devarajan P, et al. Novel and conventional serum bio markers predicting acute kidney injury in adult cardiac surgery—a prospective cohort study. Crit Care Med 2009;37:553–60. 64. Koyner JL, Bennett MR, Worcester EM, et al. Urinary cystatin C as anearly biomarker of acute kidney injury following adult cardiothoracic surgery. Kidney Int 2008;74:1059–69. 65. Han WK, Waikar SS, Johnson A, et al. Urinary biomarkers in the early diagnosis of acute kidney injury. Kidney Int 2008;73:863–9. 66. Liangos O, Perianayagam MC, Vaidya VS, et al. Urinary N-acetyl-beta-(D)glucosaminidase activity and kidney injury molecule-1 level are associated with adverse outcomes in acute renal failure. J Am Soc Nephrol 2007;8:904–12. 67. Hei Z, Li X, Shen N, Pang H, Zhou S, Guan J. Prognostic values of serum cystatin C and beta2 microglobulin, urinary beta2 microglobulin and N-acetyl-betaD-glucosaminidase in early acute renal failure after liver transplantation. Chin Med J 2008;121:1251–6. 68. Portilla D, Dent C, Sugaya T, et al. Liver fatty acid-binding protein as a biomarker of acute kidney injury after cardiac surgery. Kidney Int 2008;73:465–72.
Contents lists available at ScienceDirect
Clinical Queries: Nephrology January–March 2012, Vol. 1/Issue 1
j o u r n a l h o m e p a g e : h t t p : / / w w w. e l s e v i e r. c o m / l o c a t e / c q n
ISSN No.: 2211-9477
Biomarkers of acute kidney injury R.K. Sharma*,† *Head of the Department of Nephrology, †Director, Sanjay Gandhi Postgraduate Institute of Medical Sciences, Lucknow – 226014, India.
a r t i c l e
i
n f o
Article history: Received 2 November 2011 Accepted 4 November 2011
Keywords: Acute kidney injury Biomarker Creatinine Cystatin C Interleukin-18 Neutrophil gelatinase-associated lipocalin Renal function
a b s t r a c t
Acute kidney injury (AKI) represents a common serious clinical problem. Incidence of AKI is increasing rapidly to epidemic proportions. Development of AKI, especially in intensive care settings, is associated with increased morbidity, mortality, and hospitalization costs. Currently available diagnostic tools are mostly insensitive for early diagnosis. However, prompt diagnosis and risk stratification are necessary for guiding therapy and preventing progression of disease. Finding an early, reliable, suitable, easily reproducible, economical, and accurate biomarker for AKI is a top research priority. In recent years, many urinary and serum proteins have been investigated as possible early markers of AKI and some of them have shown great promise. This topic reviews some of the emerging biomarkers of AKI.
Introduction For the past 30 years, there have been no major improvements in the mortality rate of hospitalized patients with severe acute kidney injury (AKI), despite advances in supportive care. One key reason is because a change in the serum creatinine (SCr), which has been the standard metric for detection and progression of AKI, is not sufficiently sensitive for an early diagnosis of AKI. The absence of a more sensitive biomarker has impaired progress in this field and has had a detrimental effect on the design and outcome of AKI clinical trials. Recently, several proteins emerged as sensitive and specific biomarkers with a capacity to be used in the detection of early kidney injury and grading of injury severity. Acute kidney injury Despite more than half a century of investigation, AKI remains a major healthcare issue in medicine today. Reported to occur in 1–32% of all hospital admissions and 10–90% of intensive care unit admissions, the wide variation reflects different criteria used to define AKI. However, independent of definition, a diagnosis of AKI is consis tently associated with an increase in both short- and long-term morbidity and mortality. Even the mildest forms of AKI are independently
*Corresponding author. E-mail address: [email protected] ISSN: 2211-9477 Copyright © 2012. Reed Elsevier India Pvt. Ltd. All rights reserved. doi: 10.1016/S2211-9477(11)70003-6
Copyright © 2012, Reed Elsevier India Pvt. Ltd. All rights reserved.
associated with increased early as well as long-term mortality, the risk increasing as severity of renal injury increases.1,2 Furthermore, the incidence of AKI is increasing. Based on a large administrative database study of hospital admissions between 1992 and 2001, Xue et al estimated an 11% increase per year in the incidence of AKI.3 However, of even greater concern is the failure to develop effective interventions to prevent or treat AKI, meaning that the current management remains directed toward supportive therapy while awaiting recovery of renal function.
The urgent need for biomarkers Traditional blood (creatinine and blood urea nitrogen [BUN]) and urinary markers of kidney injury (urinary casts and fractional excretion of sodium) do not allow for early detection of AKI.4 The change in SCr does not discriminate the time and type of renal insult nor the site and extent of glomerular or tubular injury. Previous animal studies clearly demonstrated that treatment should be initiated well before the rise of SCr and very early after the insult.5–9 Therefore, the absence of sensitive and specific biomarkers for the early detection of AKI impairs progress in the diagnosis and treatment of patients with AKI and has a detrimental effect on the design and, possibly, the outcomes of clinical trials. Many potential therapeutic agents, such as atrial natriuretic peptide, insulin-like growth factor-1, and endothelin-receptor antagonist,10–13 have been tried, but have had little success. The lack of reliable biomarkers for early injury detection leads to a delay in the introduction of treatment until well into the course of renal disease. At present, there is no single intervention
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or sequence of clinical interventions that will significantly improve renal function after the onset of acute tubular injury or necrosis. Dialysis remains the only Food and Drug Administration (FDA)approved treatment option for established AKI. Acknowledging the inherent deficiencies of SCr to diagnose AKI, the American Society of Nephrology in 2005 designated identification, characterization, and development of new AKI biomarkers as a key research area for the next 5 years.
Characteristics of an ideal biomarker for acute kidney injury An ideal biomarker would identify patients at highest risk for AKI in a timely manner, thus allowing early and potentially effective intervention. Characteristics of the ideal AKI biomarker have been described and include early identification of injury, stratification according to injury severity, etiologic specificity for the injury, and providing valuable prognostic information. However, the wide spectrum of pathophysiology leading to AKI makes it unlikely that any single biomarker will achieve all these aims. Several promising biomarkers of AKI have been identified, both in urine and plasma, and are currently the subject of ongoing studies defining their clinical utility. However, the translational process from bench to bedside is complicated. Interpretation of novel biomarkers to detect minor but significant renal injury undetected by SCr proved difficult. Although insensitive and slow to respond, creatinine remains the only marker validated against clinically relevant outcomes. Any potential replacement must therefore demonstrate the ability to identify clinically meaningful injury and be useful in guiding suitable interventions or other management decisions. Although the molecular pathways mediating renal injury are increasingly understood, with potential to quantify individual components of these pathways in the laboratory,14 the focus of this clinical commentary is on biomarkers that reflect renal injury, which is frequently the result of multiple contemporaneous mechanisms in clinical practice. Besides establishing the early diagnosis, biomarkers are needed for several other purposes in AKI (summarized in Table 1). Thus, biomarkers are needed for: (1) pinpointing the location of primary injury (proximal tubule, distal tubule, interstitium, or vasculature); (2) determining the duration of kidney failure (AKI, chronic kidney disease, or ‘acute on chronic’ kidney disease); (3) discerning AKI subtypes (pre-renal, intrinsic renal, or post-renal); (4) identifying AKI etiologies (ischemia, toxins, sepsis, or a combination); (5) differentiating AKI from other forms of acute kidney disease (urinary tract infection, glomerulonephritis, or interstitial nephritis); (6) risk stratification and prognostication (duration and severity of AKI, need for renal replacement therapy, length of hospital stay, and mortality); (7) defining the course of AKI; and (8) monitoring the response to AKI interventions. Biomarkers are also desperately needed for use as surrogate endpoints in clinical trials evaluating potential therapeutics for AKI.
Biomarkers of acute kidney injury Changes in excretion of specific markers in the urine have been proposed as reflecting injury to specific regions of the nephron. Table 1 Areas of need for biomarkers in acute kidney injury. Biomarkers are needed to determine: 1. Location of injury 2. Duration of AKI 3. AKI subtypes 4. AKI etiologies 5. Differentiate from other forms of acute kidney disease 6. Risk stratification and prognostication 7. Defining course of AKI 8. Monitoring interventions AKI: acute kidney injury.
For example, high molecular weight proteinuria is proposed as an indication of glomerular injury;15–18 low molecular weight proteinuria,19,20 brush border antigens21–23 urinary enzymes,24–33 and other urinary proteins34–42 are proposed to indicate damage to the proximal convoluted tubule. As a result, insults to specific areas of the nephron, as might occur with a particular nephrotoxin, may be associated with different urinary excretion patterns of these markers. However, attempts to use these markers in screening patients for renal injury and identifying the site of kidney injury have been disappointing; this is due to a lack of sufficient validation, the lack of standardized assays, instability in urine, and the changes in pattern specificity of urinary marker excretion with advancing renal dysfunction. Recently, several protein biomarkers emerged through the application of functional genomics and proteomics to human and animal models of AKI, and proved to be promising biomarkers as non-invasive indicators of AKI.
Neutrophil gelatinase-associated lipocalin Deoxyribonucleic acid microarray techniques searching for candidate biomarkers of AKI found neutrophil gelatinase-associated lipo calin (NGAL) as one of the maximally-induced genes in a murine model of renal ischemia-reperfusion injury. A 25-KDa glycoprotein covalently bound to gelatinase, its resistance to proteolysis further enhanced potential suitability as a clinical biomarker. It is synthesized and secreted by tubular epithelial cells of the proximal and distal segment. It is freely filtered by the glomerulus, undergoing rapid clearance by the proximal tubule via receptor binding and endocytosis. In healthy kidneys, it is barely detectable in either plasma or urine. However, in the setting of acute tubular injury, NGAL undergoes rapid and profound up-regulation with large increases in both urine and plasma. Distinct from traditional markers of function such as creatinine, this rapid response enables NGAL to potentially identify injured kidney much earlier than was previously possible. The endogenous role of NGAL remains unclear. It seems to be involved with iron transportation to and from the proximal tubular epithelial cells, and animal studies demonstrate a renoprotective effect of exogenously administered NGAL in the setting of acute ischemic injury. The NGAL demonstrated a near-perfect performance for identifying AKI after pediatric cardiac surgery with an area under the receiver operator characteristic curve (AUCROC) of 0.99 and 1.0 at 2 and 4 hours, respectively, after cardiopulmonary bypass (CPB), respectively.43 Subsequent studies of both urinary and plasma NGAL in pediatric cardiac surgery support this, further demonstrating it to be an independent predictor of AKI severity and duration as well as hospital length of stay. However, results in adult cardiac surgery have been mixed, with different studies reporting widely varying diagnostic characteristics for NGAL. The AUCROC for early diagnosis of AKI by urinary NGAL has varied from 0.61 at 18 hours post-CPB44 to 0.96 at 2 hours post-CPB.45 Similarly, performance of plasma NGAL for the diagnosis of AKI has varied from an AUCROC of 0.54 within 6 hours of CPB46 to 0.87 at 24 hours post-CPB.47 Although specificity was consistently 70–80% in these studies, sensitivity has varied greatly, ranging from 40% to 90%. The reason for such widely discrepant findings is unclear. Studies reporting poorer diagnostic performance of NGAL have typically included patients with a wide spectrum of baseline renal function, and it is unknown whether this impacts the diagnostic performance of NGAL. The additional comorbidities typical of an adult cardiac surgical population may also introduce potential confounding variables, thus increasing the etiologic heterogeneity of AKI in this population. Furthermore, it is uncertain whether age itself modifies NGAL synthesis in AKI. Recently, urinary NGAL at hospital admission in multitrauma patients provided excellent early identification of AKI developing during the subsequent 5 days (AUCROC 0.98) with both sensitivity and specificity > 90%.48 Also, NGAL has been demonstrated to identify delayed graft function within hours of kidney transplantation,49 and ongoing studies are exploring its utility for early identification of renal injury in liver transplantation.
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The biologic rationale for NGAL as an early AKI biomarker is strong. However, a number of key issues, including the wide variability in reported diagnostic performance, require clarification before adoption into clinical practice. Although a rapid point-of-care test is available, experience is limited and its reliability remains to be confirmed with most studies to date using a research-based enzyme-linked immunosorbent assay (ELISA) technique for NGAL quantification.
Kidney injury molecule-1 Kidney injury molecule-1 (KIM-1) is a type 1 transmembrane glycoprotein containing a novel 6-cysteine immunoglobulin-like domain plus a threonine/serine and proline-rich domain characteristic of mucin-like O-glycosylated proteins. It is undetectable in normal kidney tissue or urine, but is expressed at high levels in dedifferentiated proximal tubule epithelial cells from human and rodent kidneys after ischemic or toxic injury, and in renal cell carcinoma.50–53 It is also known as hepatitis A virus cellular receptor 1 and T-cell immunoglobulin and mucin-domain-containing molecule 1.54–56 Also, KIM-1 has been implicated as a urinary biomarker of AKI in pediatric and adult patients. In a small cross-sectional study, Han et al demonstrated that soluble forms of cleaved KIM-1 can be detected in the urine of patients with established AKI; elevated urinary KIM-1 levels were found within 12 hours after an initial ischemic insult and prior to the appearance of granular casts in the urine.51 Urinary KIM-1 was also evaluated after pediatric cardiac surgery in a case control study,57 using samples from the same cohort in the NGAL study previously described. Urinary KIM-1 had an AUCROC of 0.57 at 2 hours, 0.83 at 12 hours, and 0.78 at 24 hours. High urinary KIM-1 expression was also associated with adverse clinical outcomes in patients with established AKI. In a prospective study, a cohort of 201 patients with established AKI, Liangos et al demonstrated that urinary KIM-1 and N-acetyl-b-D-glucosaminidase (NAG) were significantly associated (AUCROC of 0.78) with the clinical composite endpoints of death or renal replacement therapy.58 Kidney injury molecule-1 is particularly interesting given its ‘kidney dominance’ of expression. In a follow-up study involving a larger number of adult patients, the temporal expression patterns and the utility of urinary KIM-1 for the early detection of AKI are being studied.
Interleukin-18 Interleukin-18 (IL-18) is a proinflammatory cytokine that is known to be induced and cleaved in the proximal tubule, and subsequently can be easily detected in the urine following ischemic AKI in animal models. In an initial cross-sectional study, urine IL-18 levels mea sured by a commercially available ELISA were markedly elevated in patients with established AKI, but not in subjects with urinary tract infection, chronic kidney disease, nephrotic syndrome or pre-renal azotemia. In a subsequent study of patients who developed AKI 2–3 days after cardiac surgery, urinary IL-18 levels peaked at 12 hours post-surgery, and predicted AKI with an AUCROC of 0.75.59 Urine IL-18 is also considered an early biomarker of AKI in the intensive care setting, being able to predict this complication about 2 days prior to the rise in SCr. Early elevated urine IL-18 levels correlated with the severity of AKI as well as mortality. Thus, IL-18 may also represent a promising biomarker for AKI. Interleukin-18 is more specific to ischemic AKI, and levels are largely unaffected by nephrotoxins, chronic kidney disease or urinary tract infections. However, IL-18 measurements may be influenced by a number of co-existing variables, since renal IL-18 mRNA levels are known to be induced in other disease states such as endotoxemia, immunological injury and cisplatin toxicity. Furthermore, plasma IL-18 levels are known to be increased in various systemic inflammatory states, and the relationships between plasma and urine IL-18 remain unexplored.
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Cystatin C A small cysteine proteinase inhibitor, cystatin-C, has many features of an ideal glomerular filtration marker. Synthesized and released into plasma by all nucleated cells at a constant rate, its small size (13 KDa) and positive charge at physiologic pH makes it freely filtered at the glomerulus. It is neither secreted nor reabsorbed by renal tubules but undergoes almost complete catabolism by proximal tubular cells, and thus little, if any, appears in the urine. With a half-life of about 2 hours, serum cystatin-C reflects glomerular filtration rate better than creatinine. Although it is generally considered less subject to the non-renal variables that impact creatinine, recent studies suggest that cystatin-C levels may in fact be affected by various anthropometric measures as well as inflammatory processes, use of corticosteroids, and changes in thyroid function, thereby potentially confounding perioperative interpretation.60 Although increasingly reported as an endpoint in clinical studies, the diagnostic and prognostic characteristics of cystatin-C for AKI remain incompletely defined. In a mixed critical care population, cystatin-C enabled a diagnosis of AKI 1.5 days earlier than plasma creatinine, with moderate ability to predict dialysis requirement.61 However, a similar repeat study found that cystatin-C identified AKI no earlier than creatinine and did not predict mortality.62 Results in adult cardiac surgery are similarly mixed, with one study reporting good discrimination for AKI within 6 hours of surgery (AUCROC 0.83, sensitivity 77%, specificity 86%),63 whereas a further study of similar size found serum cystatin-C no better than chance for identifying AKI.64 However, the latter study did find that urinary cystatin-C identified AKI within 6 hours of surgery (AUCROC 0.72, sensitivity 94%, specificity 40%), suggesting that tubular injury may impair the usual catabolism of cystatin-C, leading to its appearance in urine. Several studies report a rise in cystatin-C within 8 hours of exposure to radiocontrast agents but without adequate description of the diagnostic characteristics for contrast-induced nephropathy. Although a commercial platform for rapid and reliable cystatin-C measurement is available, the total number of patients studied remains small and with inconsistencies in results that are not well understood.
N-Acetyl-b-D-Glucosaminidase Although several enzymes present in tubular epithelial cells have been investigated as potential markers of renal injury, NAG remains the most extensively investigated to date. A lysosomal enzyme, it is abundantly present in proximal tubular epithelial cells and its relatively large size (130 KDa) prevents glomerular filtration with the result that urinary NAG represents enzyme leakage from proximal tubular cells into the tubular lumen. It is stable in urine across a range of pH and temperature, and it is easily quantified by commercially available colorimetric or spectrophotometric methods. Although a large number of studies have profiled the release of NAG across a diverse range of clinicopathologic conditions, each purporting to demonstrate subtle proximal renal tubular damage, few studies address either the diagnostic capability for AKI or a direct link with clinical outcomes. The NAG was used extensively through the 1990s by anesthesiologists investigating the potential nephrotoxicity of compound A, a degradation product of sevoflurane. Despite several studies indicating increases in urinary NAG with sevoflurane exposure, as well as other potential markers of renal injury, such as albumin, microglobulins, glutathione-S-transferase, and glucose, a link with histopathologic changes, increasing SCr, or adverse clinical outcomes was unable to be established, thus leaving the interpretation of these transient perioperative changes unclear. However, in a cross-sectional study, urinary NAG provided excellent discrimination of patients with established AKI from a control group including both normal individuals and patients with urinary tract infection (AUCROC 0.97)65 and in a hospital population referred for nephrology consultation for acute renal failure, increasing urinary NAG was associated with a 3.0-, 3.7-, and 9.1-fold higher odds for the composite outcome of in-hospital mortality or dialysis with each increasing
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quartile relative to the first, a relationship maintained despite adjusting for multiple comorbidities.66 The NAG levels increased within 6 hours of pediatric cardiac surgery, remaining elevated through 48 hours. Although higher in patients who developed AKI, diagnostic utility was modest 12 hours post-CPB (AUCROC 0.69) and despite sensitivity > 80%, specificity remained poor across a range of potential threshold values. In contrast, urinary NAG levels were not consistently different between patients with or without AKI after cardiac surgery in adults, and a single study in liver transplant recipients failed to show a difference in post-operative urinary NAG levels between patients with or without AKI.67
Finally, larger prospective studies are necessary to validate the temporal expression pattern of various biomarkers for early detection of AKI, to determine how to combine multiple biomarkers for early detection of AKI, and to discover how the temporal course relates to the onset, severity, and outcome of AKI. More information must be gathered regarding the accuracy of measurement in the presence of interfering substances and the stability of the various biomarkers in routine clinical storage conditions, including a number of freeze–thaw cycles.
Liver fatty acid binding protein
There is a need for a new standard definition of AKI that is not based on a change in SCr. The new definition of AKI should be based on multiple biomarkers and clinical parameters, and capable of detecting initial renal injury within minutes to hours. Any potential AKI biomarker should undergo several clinical validation processes for both assay performance and diagnostic utility. Furthermore, several requirements must be met in order for AKI biomarkers to be useful in daily clinical practice. The AKI biomarkers must:
Liver fatty acid binding protein (LFABP) is a 14 KDa protein normally expressed in the proximal convoluted and straight tubules of the kidney, and up-regulated in animal models of AKI. An ELISA for this analyte is commercially available; it was first used to demonstrate that urinary L-FABP levels were significantly increased prior to the increase in SCr in those patients who developed AKI post-contrast dye. In a recent prospective study of children undergoing cardiac surgery, urine LFABP increased at 4 hours post-bypass, with an AUC of 0.810 for a cut-off value of 486 ng/mg creatinine.68 Thus, L-FABP also appears to represent a promising AKI bio marker. However, urinary L-FABP measurements may also be influenced by a number of confounding variables. Several studies have documented increased urinary L-FABP levels in patients with nondiabetic chronic kidney disease, early diabetic nephropathy, idiopathic focal glomerulo sclerosis and polycystic kidney disease. Additionally, LFABP is also abundantly expressed in the liver, and urinary L-FABP may be influenced by serum LFABP levels.
Limitations of biomarkers for early detection of acute kidney injury in clinical situation Additional studies are necessary before biomarkers may be used in routine clinical practice. Currently, these promising biomarkers have been tested in only small studies and limited clinical situations. None of these biomarkers have been systematically evaluated in the various clinical settings of AKI. Furthermore, the studies have been insufficiently powered to establish a cut-off value that is predictive of AKI. In addition, there are very limited data available, at present, about temporal expression patterns for various biomarkers including NGAL, KIM-1, IL-18, and Cys-C in the various clinical settings of AKI from the onset of renal insult. The heterogeneity of AKI suggests that more than one marker may be necessary to obtain sufficient sensitivity and specificity for AKI screening. An analysis of multiple biomarkers may optimize early detection of AKI, as in the case of prostate cancer detection. Analyses of multiple markers may also provide insight into the pathophysiology and sites of nephron injury, since different markers may be chosen to reflect injury to different nephron segments. Nevertheless, it would be naïve to call the result positive if one or more biomarkers are positive, since such an approach would be inefficient and increase sensitivity at the expense of specificity, or vice versa. A linear combination that would maximize the AUCROC for detection of AKI may be useful, but this approach remains to be validated by future studies.57 Currently, there is no standard procedure to combine the multiple biomarkers for clinical use. Another major challenge is the development of a rapid assay for validated biomarkers at the bedside or in a clinical laboratory for the detection of AKI. Current quantification methods include ELISA, Luminex®-based assay, and nephelometry, which are not fast enough for rapid assay. Recently, Biosite Inc. launched a prospective multicenter study evaluating NGAL in early and evolving stages of AKI. The study is using the Triage® NGAL test, which supposedly can accurately measure NGAL concentrations in blood and provide results within 15 minutes.
Conclusion
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Allow for early detection of renal injury. Identify the severity of AKI. Provide a rationale for risk stratification in clinical studies, including the identification of patients at risk for AKI. Guide timing of therapy. Reflect improvement and worsening of the kidney injury. Be amenable to quick and reliable measurement at the bedside or clinical laboratory.
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